Stem Cell Technology represents a major breakthrough in anti-aging and regenerative skin care, by protecting, strengthening, and replenishing our own human skin cells. Where Peptides stimulate different functions acting as messengers to skin cells, stem cell technology improves the life of the core of the cell. Working in synergy with peptides, they enhance the effectiveness of peptides and other active ingredients.

Antiaging effects - The stem cells in our skin have a limited life expectancy due to DNA damage, aging and oxidative stress. As our own skin stem cells age, they become more difficult to repair and replenish. Protection of our stem cells becomes more and more beneficial as our skin ages, and with the advent of stem cells, we are now able to delay the natural aging process even further than before.

Expected benefits of stem cells technology for regenerative skin care:

Stem Cell Replenishing Serum Featuring a potent concentration of apple and edelweiss plant stem cells, state-of-the-art peptides, and other cutting edge ingredients, the Stem Cell Replenishing Serum is thoroughly formulated to produce age defying results, restoring the youthful look and vitality to aging skin.

Stem Cell Moisturizing Cream Also featuring a healthy concentration of apple and edelweiss plant stem cells, peptides, and numerous botanical extracts, the Stem Cell Moisturizing Cream is formulated to produce age defying results while also helping to maintain healthy and youthful looking skin as a daily moisturizer.

Our Stem Cell Applications:

LPAR Stem Cell Products contain a wide variety of stem cells with healthy and potent concentrations in order to deliver the results skin care consumers strive for. The first stem cell ingredient discovered and produced is a liposomal preparation based on the stem cells of a rare Swiss apple. The revolutionary active ingredient, Malus Domestica by PhytoCellTec is based on a high tech plant cell culture technology. It has been proven to protect the longevity of skin stem cells and provide significant anti-wrinkle effects. Since the discovery and the worldwide success of Apple Stem Cells introduction to the cosmetic and skin care marketplace, other new and exciting stem cell ingredients have been discovered to provide extraordinary results for all skin types.

We were proud to be the first skin care line to offer the ground-breaking combination of Apple and Edelweiss stem cells, and are dedicated to formulating the best new and existing stem cell ingredients into our product line as the technology continues to develop.

To inquire about purchasing LPAR Stem Cell products. visit our Retail Locator page.

Featuring a luxurious and potent blend of three major botanical stem cells (Apple, Gardenia Jasminoides, Echinacea Angustifolia) two state-of-the-art peptides (Nutripeptides, Matrixyl synthe6), and numerous botanical extracts and minerals, the Stem Cell Nourishing Mask is thoroughly formulated to nourish, firm, and energize mature skin. Total Stem Cell Concentration: 5.5% - Total Peptide Concentration: 9.0%

Directions: Using fingertips, apply on clean, dry skin twice weekly. Avoid the eye area. The mask can be left on the skin for prolonged periods (during the day or overnight). Allow at least 10-15 minutes for the mask to penetrate the skin before rinsing with water or applying additional product For external use only.

Ingredients: Water (Aqua), Glycerin, Glyceryl Acrylate/Acrylic Acid Copolymer, Hydrolyzed Rice Protein (Nutripeptides), Sodium Hyaluronate, Hydroxypropyl Cyclodextrin, Palmitoyl Tripeptide-38 (Matrixyl synthe6), Biosaccharide Gum-1, Olea Europaea (Olive) Fruit Oil, Gardenia Jasminoides Meristem Cell Culture, Xanthan Gum, Malus Domestica Fruit Cell Culture, Lecithin, Porphyridium Polysaccharide, Echinacea Angustifolia Meristem Cell Culture, Carbomer, Triethanolamine, Mentha Pipertita (Peppermint) Extract, Camellia Sinensis (Green Tea) Leaf Extract, Palmaria Palmata (Dulce) Extract, Chamomilla Recutita (Matricaria) Flower Extract, Phenoxyethanol, Caprylyl Glycol, Ethylhexylglycerin, Hexylene Glycol, Copper PCA, Zinc PCA, Dipotassium Glycyrrhizate, Olea Europaea (Olive) Fruit Extract, Aloe Barbadensis Leaf Juice Powder, Fragrance (Parfum)

Featuring a plant and fruit stem cell enhanced blend of three major stem cells (Apple, Edelweiss, Alpine Rose), state-of-the-art peptides (Eyeseryl, Nutripeptides), the Stem Cell Eye Therapy is an advanced eye formula designed to nourish, firm, and increase skin elasticity and skin smoothness around the eye area. Total Stem Cell Concentration: 6.75% - Total Peptide Concentration: 11.0%

Directions: Using fingertips, apply product around both eyes on clean, dry skin once or twice daily before applying a moisturizer or night cream. For external use only.

Ingredients: Water, Acetyl Tetrapeptide-5 (Eyeseryl), Sodium Hyaluronate, Hydrolyzed Rice Protein (Nutripeptides), Glycerin, Leontopodium Alpinum Meristem Cell Culture (Edelweiss Stem Cells), Xanthan Gum, Malus Domestica Fruit Cell Culture (Apple Stem Cells), Lecithin, Porphyridium Polysaccharide, Camellia Sinensis (Green Tea) Leaf Extract, Cucumis Sativus (Cucumber) Fruit Extract, Phenoxyethanol, Caprylyl Glycol, Ethylhexylglycerin, Hexylene Glycol, Carbomer, Triethanolamine, Rhododendron Ferrugineum Leaf Cell Culture Extract (Alpine Rose Stem Cells) Isomalt, Sodium Benzoate, Lactic Acid, Sodium Polystyrene Sulfonate, Allantoin, Copper PCA, Aloe Barbadensis Leaf Juice Powder

Plant stem cells represent a major breakthrough in skin care, launching the beginning of a new system of treating the skin...by protecting and replenishing the building blocks of what makes up our own skin: Stem Cells. Rather than working around the natural aging process of our skin stem cells, we now have the technology available to improve the life of our skins most important and central component.

Featuring a potent combination of apple, edelweiss, and grape stem cells, state-of-the-art peptides, and other cutting edge ingredients, the Stem Cell Replenishing Serum is thoroughly formulated to produce age defying results, restoring the youthful look and vitality to aging skin.

Directions: Apply with fingertips on clean, dry skin once or twice daily. Avoid the eye area by approximately 1 cm. Suitable for mature skin types. For external use only.

Ingredients: Water (Aqua), Glycerin, Dipeptide Diaminobutyroyl Benzylamide Diacetate, Acetyl Octapeptide-3, Malus Domestica Fruit Cell Culture (Apple Stem Cells), Hydrolyzed Ceratonia Siliqua Seed Extract, Palmitoyl Tripeptide-5, PEG-8 Dimethicone, Saccharide Isomerate, Imperata Cylindrica (Root) Extract, Polysorbate 20, Leontopodium Alpinum Meristem Cell Culture (Edelweiss Stem Cells), Leucojum Aestivum Bulb Extract, Triethanolamine, Carbomer, Xanthan Gum, Vitis Vinifera Fruit Cell Extract (Grape Stem Cells), Isomalt, Sodium Benzoate, Lecithin, Disodium EDTA, Allantoin, Aloe Barbadensis Leaf Juice Powder, Phenoxyethanol, Caprylyl Glycol, Ethylhexylglycerin, Hexylene Glycol, PEG-8-Carbomer, Fragrance (Parfum)

Plant stem cells represent a major breakthrough in skin care, launching the beginning of a new system of treating the skin...by protecting and replenishing the building blocks of what makes up our own skin: Stem Cells. Rather than working around the natural aging process of our skin stem cells, we now have the technology available to improve the life of our skins most important and central component.

Featuring a healthy concentration and a diverse group of stem cells (apple, edelweiss, grape), peptides, and numerous botanical extracts, the Stem Cell Moisturizing Cream is formulated to produce age-defying results, while also helping to maintain healthy and youthful looking skin as a daily moisturizer.

Directions: For mature skin and/or skin conditioning, apply onto clean, dry skin with fingertips once daily. Avoid the eye. For external use only.

Ingredient Highlights: Plant/Fruit Stem Cells 4% - Malus Domestica (Apple Stem Cells) - Leontopodium Alpinum Cell Culture Extract (Edelweiss Stem Cells) - Vitis Vinifera Fruit Cell Extract (Grape Stem Cells)

Ingredients: Water (Aqua), Glycerin, Isopropyl Myristate, Caprylic/Capric Triglyceride, Cetearyl Olivate, Sorbitan Olivate, Sorbitol, Saccharide Isomerate, Sodium Hyaluronate, Leucojum Aestivum Bulb Extract, Malus Domestica Fruit Cell Extract (Apple Stem Cells), Leontopodium Alpinum Meristem Cell Culture (Edelweiss Stem Cells), Vitis Vinifera Fruit Cell Extract (Grape Stem Cells), Crambe Abyssinica Seed Oil, Dimethicone, Cetyl Alcohol, Imperata Cylindrica (Root) Extract, Acetyl Octapeptide-3 (SNAP-8), Dipeptide Diaminobutyroyl Benzylamide Diacetate(SYN-AKE), Palmitoyl Tripeptide-3 (SYN-COL), Hydrolyzed Ceratonia Siliqua Seed Extract, Aloe Barbadensis Leaf Juice Powder, Olea Europaea (Olive) Leaf Extract, Glyceryl Stearate, Xantham Gum, Cetyl Palmitate, Sorbitan Palmitate, Bisabolol, Tocopheryl Acetate, Fragrance, Phenoxyethanol, Caprylyl Glycol, Ethylhexyglycerin, Hexylene Glycol, PEG-8, Carbomer, Lecithin, Isomalt, Sodium Benzoate, Disodium EDTA

[ pH: 5.00 ]

Featuring high concentrations of Vitamin C (Tetrahexyldecyl Ascorbate), Orange Stem Cells, and Peptides, this is a multi-beneficial cream with state-of-the-art actives formulated to deliver significant and lasting results.

Tetrahexyldecyl Ascorbate is a stable, oil soluble form of Vitamin C that penetrates deeper into the skin than traditional ascorbic acid based Vitamin C. It's a proven skin lightener, a powerful Anti-Oxidant, DNA protector, and increases collagen synthesis more effectively than ascorbic acid. Orange Stem Cells work to increase elasticity and skin resistance to the dermis, which increase firmness and diminish wrinkles while also working synergistically with peptides to further increase skin elasticity and collagen support.

How to Use: Smooth a pearl sized drop onto the face once daily (morning or evening). Avoid the eye area while applying. Follow with Solar Protection if used during the day.

Ingredients: Water (Aqua), Tetrahexyldecyl Ascorbate (Vitamin C Ester), Glycerin, Hexyl Laurate, Caprylic/Capric Triglyceride, Butylene Glycol, Sorbitol, Stearic Acid, Glyceryl Stearate, PEG-100 Stearate, Cetyl Alcohol, Sorbitan Stearate, Polysorbate 60, Acetyl Hexapeptide-8, Sodium Hyaluronate, Squalane, Dimethicone, PPG-12/SMDI Copolymer, Citrus Aurantium Dulcis Callus Culture Extract (Orange Stem Cells), Tocopheryl Acetate, Cetearyl Ethylhexanoate, Linoleic Acid, Glycine Soja (Soybean) Sterols, Phospholipids, Di-PPG-2 Myreth-10 Adipate, Retinol, Polysorbate 20, Hydrolyzed Glycosaminoglycans, Alcohol, Ectoin, Lecithin, Cyclotetrapeptide-24 Aminocyclohexane Carboxylate, Glucosamine HCl, Algae Extract, Yeast Extract, Urea, Micrococcus Lysate, Plankton Extract, Arabidopsis Thaliana Extract, Magnesium Aluminum Silicate, Xanthan Gum, Phenoxyethanol, Caprylyl Glycol, Ethylhexylglycerin, Hexylene Glycol, Disodium EDTA, Citrus Aurantium Dulcis (Orange) Peel Oil

[ pH: 4.7 ]

The Vitamin C Stem Cell Mask combines a potent blend of Vitamin C Ester (Tetrahexyldecyl Ascorbate), highly concentrated plant and fruit stem cells (Argan, Sea Fennel), and Aldenine, a unique peptide that acts as a cellular detoxifier and a collagen III booster.

Directions: Apply on clean, dry skin. Avoid the eye area. The mask may be left on the skin (i.e. during the day or overnight), or it may be rinsed off with lukewarm water after 10 - 15 minutes. Suitable for mature skin types.

Ingredients: Water (Aqua), Tetrahexyldecyl Ascorbate, Kaolin, Glycerin, Glyceryl Stearate, Sorbitan Olivate, Cetearyl Olivate, Cetyl Palmitate, Sorbitol, Sorbitan Palmitate, Stearic Acid, Caprylic/Capric Triglyceride, Cyclopentasiloxane, Cyclhexasiloxane, Carthamus Tinctorius (Safflower) Seed Oil, Punica Granatum Extract, Butylene Glycol, Ananas Sativus (Pineapple) Fruit Extract, Carica Papaya Fruit Extract, Hydrolyzed Wheat Protein, Hydrolyzed Soy Protein, Tripeptide-1, Argania Spinosa (Argan Stem Cells) Sprout Cell Extract, Crithmum Maritimum (Sea Fennel Stem Cells) Callus Culture Filtrate, Oligopeptide-68, Sodium Oleate, Phenoxyethanol, Caprylyl Glycol, Ethylhexylglycerin, Hexylene Glycol, Polyacrylamide, C13-14 Isoparaffin, Laureth-7, Isomalt, Hydrogenated Lecithin, Lecithin, Sodium Benzoate, Allantoin, Citrus Aurantium Dulcis (Orange) Peel Oil, Magnesium Aluminum Silicate, Xanthan Gum, Disodium EDTA

[ pH: 6.00 ]

Originally designed to prepare and increase the skins receptiveness to our Professional Peptide Peel, the Premier Peptide Serum has gone on to become our most powerful anti-wrinkle product for year-round home care due to its high concentration and diversity of peptides. Composed of a total concentration of 65% peptides, the Premier Peptide Serum is a state of the art facial serum expertly formulated to reduce the signs of aging, energizing mature skin.

The Intensive Clarifying Peptide Cream is a unique and high potency moisturizing cream formulated with an abundance of natural skin lighteners, peptides, and botanical extracts that combine to clarify and firm mature skin, while effectively minimizing fine lines and wrinkles.

The Collagen Peptide Complex builds off of our original Collagen Copper Activating Complex, and includes an advanced formulation of peptides, including Syn-Coll, a small but powerful peptide that stimulates collagen synthesis at a cellular level, helping to compensate for any collagen deficit in the skin.

Boasting a remarkable collection of natural and innovative ingredients from exotic plants and enhanced peptides, the neck firming cream has been designed & tested to firm and energize mature skin, while providing increased smoothness and elasticity to the often neglected neck area.

Providing sufficient hydration is the most essential way to keep our skin healthy and youthful. While many of our products assist in hydrating the skin, hydration is the main focus of the Nano-Peptide B5 Complex, acting as the foundation for your home care regimen. Fortified with Sodium Hyaluronate (30%) and Pantothenic acid, it provides an especially deep and complete hydration. Because of the presence of peptides, it also assists in tightening and firming the skin while allowing for maximum absorption and effectiveness.

Designed for mature skin, this sophisticated moisturizer promotes cell renewal, stimulating the dermis layer of the skin with a high potency blend of peptides (Argireline, Matrixyl, & Biopeptide-CLTM) and botanical extracts that make it a particularly refined and effective moisturizing cream for age management.

The A&M Eye Recovery Therapy is an advanced age management treatment, applying the most tried and true peptides and delivery systems; Argireline & Matrixyl, to the highly wrinkle prone and fragile eye area, providing diminished wrinkle depth, and increased firmness and elasticity. The peptide Eyeliss is added to further enhance this treatment by counteracting skin slackening, puffiness, and decreasing irritation.

The A&M Facial Recovery Therapy is an advanced age-management treatment that blends the most tried and true peptides and delivery systems; Argireline & Matrixyl. Stimulating the deeper layers of the skin, the A&M Facial Recovery Therapy provides diminished wrinkle depth, as well as an increase in skin elasticity and firmness.

Originally designed to prepare and increase the skins receptiveness to our Professional Peptide Peel, the Premier Peptide Serum has gone on to become our most powerful anti-wrinkle product for year-round home care due to its high concentration and diversity of peptides. Composed of a total concentration of 65% peptides, the Premier Peptide Serum is a state of the art facial serum expertly formulated to reduce the signs of aging, energizing mature skin.

Directions: For mature skin types; apply at least three weeks before beginning the Lucrece Professional Peptide Peel treatment, and use twice a day leading up to the Peel. For year round application, apply once per day after the Collagen Peptide Complex. Avoid the eye area by at least 1 cm during application.

Peptides: SYN-AKE: A small peptide (Dipeptide Diaminobutyroyl Benzylamide Diacetate) that mimics the activity of Waglerin 1, a polypeptide that is found in the venom of the Temple Viper, Tropidolaemus wagleri. Clinical trials have shown SYN-AKE is capable of reducing wrinkle depth by inhibiting muscle contractions. SNAP-8: An anti-wrinkle (Acetyl Octapeptide-3) elongation of the famous Hexapeptide Argireline. The study of the basic biochemical mechanisms of anti-wrinkle activity led to the revolutionary Hexapeptide which has taken the cosmetic world by storm. ARGIRELINE: (Acetyl Hexapeptide-8) MATRIXYL: (Palmitoyl Pentapeptide-4) REGU-AGE: (Hydrolyzed Rice Bran Protein - Oxido Reductases - Soybean Protein) BIOPEPTIDE CL: (Palmitoyl Oligopeptide) RIGIN: (Palmitoyl Tetrapeptide-7) EYELISS: (Dipeptide-2 & Palmitoyl Tetrapeptide-7) INYLINE: (Acetyl Hexapeptide 30)

Other Ingredients: Water, Sodium Hyaluronate, Spiraea Ulmaria Flower Extract & Centella Asiatica Extract & Echinacea Purpurea Extract, Phenoxyethanol & Benzyl Alcohol & Potassium Sorbate & Tocopherol, Meadowsweet, Hydrocotyl Extract, Leucojum Aestivum Bulb Extract, Amino Acids, Diazolidinyl Urea, Imperata Cylindrica Extract, SMDI Copolymer, Hydroxyethylcellulose

[ pH: 5.00 ]

This unique and high potency moisturizing cream is formulated with an abundance of natural skin lighteners, peptides, and botanical extracts that combine to help clarify and energize mature skin.

Directions: Smooth a pearl size drop onto the face, gently massaging in with fingertips once per day (morning), avoiding the eye area. Follow with solar protection if applicable.

Skin Lightening Agents: Mulberry Bark, Saxifrage Extract, Grape Extract, Scutellaria Root Extracts, Vitamin C Ester (Tetrahexyldecyl Ascorbate), Emblica Fruit Extract, Licorice Root Extract.

Ingredients: Water (Aqua), Saxifrage Extract & Grape Extract & Butylene Glycol & Water & Mulberry Bark Extract & Scutellaria Root Extract, Prunus Amygdalus Dulcis (Sweet Almond) Oil, Caprylic/Capric Triglycerides, Sesamum Indicum (Sesame) Seed Oil, Cetearyl Olivate & Sorbitan Olivate, Glycerin, Palmitoyl Pentapeptide-4 (Matrixyl), Tetrahexyldecyl Ascorbate (C-Ester), Glyceryl Stearate & PEG 100 Stearate, Stearic Acid, Theobroma Cocao (Cocoa) Seed Butter, PPG-12/SMDI Copolymer, Butyrospermum Parkii (Shea) Butter, Tocopheryl Acetate (Vitamin E), Phyllanthus Emblica Fruit Extract, Palmitoyl Tripeptide-5 (Syn-Coll), Triethanolamine, Phenoxyethanol, Mangifera Indica (Mango) Seed Butter, Darutoside, Tricholoma Matsutake Singer (Mushroom) Extract, Imperata Cylindrica (Root) Extract, Fragrance (Parfum), Glucosamine HCL & Algae Extract & Yeast Extract & Urea, Retinyl Palmitate (Vitamin A), Centella Asiatica Extract & Echinacea Purpurea Extract, Xanthan Gum, Arctostaphylos Uva Ursi Leaf Extract, Glycyrrhiza Glabra Root Extract, Magnesium Aluminum Silicate, Disodium EDTA

[ pH: 5.75 ]

Specializing in firming the skin, the Collagen Peptide Complex builds off of our original Collagen Copper Activating Complex, and adds a combination of (5) major peptides, helping to keep the skin looking its youngest and most alive, as it works to firm, and add elasticity & texture to the skin. For best results, apply directly after the Nano-Peptide B5 Complex.

Directions: Apply a liberal amount on clean, dry face using fingertips, and massage into the skin. Let dry, and follow with a moisturizer and sun-block if used during the day, or the Vitamin A Facial Cream + III if used at night. Warning: For mature skin only. If redness occurs, lessen use to once or twice per week. If reactions persist, discontinue use.

Ingredients: Water (Aqua), Dipalmitoylhydroxyproline, Glycerin, Palmitoyl Tetrapeptide-7 (Rigin), Palmitoyl Oligopeptide (Biopeptide-CL), Butylene Glycol, Yeast (Faex Extract), Hydrocotyl Extract & Coneflower Extract, Aloe Barbadensis Leaf Extract, Palmitoyl Tripeptide-5 (Syn-Coll), Acetyl Hexapeptide-8 (Argireline), Palmitoyl Pentapeptide-4 (Matrixyl), Panthenol, Phenoxyethanol & Caprylyl Glycol & Ethylhexylglycerin & Hexylene Glycol, Triethanolamine, Carbomer, Decarboxy Carsonine HCI, Citrus Grandis (Grapefruit) Seed Extract, Copper PCA, Olea Europaea (Olive) Leaf Extract, Disodium EDTA

[ pH: 5.50 ]

Boasting a remarkable collection of natural and innovative ingredients from exotic plants and enhanced peptides, the neck firming cream has been designed & tested to firm and energize mature skin, while providing increased smoothness and elasticity to the often neglected neck area.

Directions: On clean dry skin, apply onto the neck area with fingertips in an upward motion. Apply twice a day, or as needed.

Key Ingredients: Bio-Bustyl: Stimulates cell metabolism, promotes collagen synthesis, and enhances fibroblast (collagen-producing cell) proliferation. INCI: Glyceryl Polymethacrylate, Soy Protein Ferment, PEG-8, & Palmitoyl Oligopeptide Polylift: Using a cross-linking technology, biopolymerization, Polylift reinforces the natural lifting effect of sweet almond proteins, providing a smooth firmness & radiance to the surface of the skin. INCI: Prunus Amygdalus Dulcis (Sweet Almond) Seed Extract.

Ingredients: Deionized Water, Prunus Amygdalus Dulcis (Sweet Almond Oil), Caprylic/Capric Triglycerides, Sesamum Indicum (Sesame) Seed Oil, Simmondsia (Jojoba) Seed Oil/ Buxus Chinensis, Cetearyl Alcohol, Dicetyl Phosphate, Ceteth-10 Phosphate, Palmitoyl Oligopeptide, Palmitoyl Tetrapeptide-7, Prunus Amygdalus Dulcis Seed Extract, Terminalia Catappa Leaf Extract & Sambucus Nigra Flower Extract & PVP & Tannic Acid, Glyceryl Polymethacrylate & Rahnella/ Soy Protein Ferment & PEG-8 & Palmitoyl Oligopeptide, Glycerin, Glyceryl Stearate & PEG 100 Stearate, Biosaccharide Gim-1, PPG-12/ SMDI Copolymer, Phyllanthus Emblica Fruit Extract, Stearic Acid, Centella Asiatica Extract & Darutosidetriethanolamine, Tocopheryl Acetate, Magnifera Indica (Mango) Seed Butter, Glycerin & Aqua & Lysolecithin & Perilla Frutescens Seed Oil, Xantham Gum, Retinyl Palmitate, Tetrahexyldecyl Ascorbate (Vitamin C Ester), Echinacea Purpurea Extract, Imperata Cylindrica (Root) Extract, Glycyrrhiza Glabra Root Extract, Magnesium, Aluminum Silicate, Disodium EDTA

[ pH: 6.25 ]

Hydration is the most essential way to keep our skin healthy feeling and healthy looking. While many of our products assist in hydrating the skin, hydration is the main focus for this product, making it an essential for all skin types. Fortified with Hyaluronic (30%) and Panthenol (Vitamin B5), the Nano-Peptide B5 Complex provides an especially deep and complete hydration. With the addition of peptides, it also assists in tightening and firming the skin while allowing for maximum absorption and effectiveness.

The Nano-Peptide B5 Complex should be applied directly after cleansing the skin, as the 2nd step in skin care regimens for all skin types (morning & night). For best results, age management regimens should follow with the Stem Cell Replenishing Serum and/or the Collagen Peptide Complex before moisturizing.

Directions: Apply a healthy amount on clean, dry skin. May be used around the eye area.

Key Ingredients: Palmitoyl Pentapeptide-4: Stimulates the skins fibroblasts to rebuild the extra-cellular matrix, including the synthesis of Collagen I and Collagen IV, fibronectin and of Glycosaminoglycans. It also stimulates the production of the dermal matrix (Collagen I & III) resulting in a significant reduction of wrinkles and fine lines. Acetyl Hexapeptide-8: Reduces facial wrinkle depth and the signs of skin aging resulting from facial movements and facial muscle contraction by halting the release of neurotransmitters from SNARE and catecholamine complexes, (which can also induce formation of wrinkles and fine lines to the skin). Hyaluronic Acid (30%): Penetrates deep into the skin, providing ample moisture Panthenol: Enhances formation of skin pigments for younger looking skin, and contains deep penetrating properties that allow a more complete hydration.

Other Ingredients: Water (Aqua), Hyaluronic Acid, Panthenol (Vitamin B5), MDI Complex, Palmitoyl Pentapeptide-4, Acetyl Hexapeptide-8, Phenoxyethanol, Hydrolyzed Wheat Protein, Butylene Glycol, Hydrocotyl & Coneflower Extract, Glycosaminoglycans.

[ pH: 5.5 ]

Designed for mature, sun damaged, and/or dehydrated skin, the Anti-Wrinkle Facial Cream is a peptide enriched moisturizer focused on increasing skin firmness & elasticity, and fortifying the skin with anti-oxidants & botanical extracts to facilitate healthy feeling and healthy looking skin.

Directions: Smooth a pearl size drop onto the face, massage into skin thoroughly. For use in the morning (recommended), follow with solar protection.

Ingredients: Water (Aqua), Glycerin, Dimethicone, Caprylic/Capric Triglycerides, C12-15 Alkyl Benzoate, Linoleic Acid & Glycine Soja (Soybean) Sterols & Phospholipids, Acetyl Hexapeptide-8, Butylene Glycol & Carbomer & Polysorbate 20 & Palmitoyl Pentapeptide-4, Cetearyl Alcohol & Dicetyl Phosphate & Ceteth-10 Phosphate, Glyceryl Stearate & PEG 100 Stearate, PPG-12/ SMDI Copolymer, Phyllanthus Emblica Fruit Extract, Darutoside, Cocoa Butter, Cetyl Alcohol, Butyrospermum Parkii (Shea Butter), Saccharomyces/Xylinum Black Tea Ferment & Glycerin & Hydroxyethylcellulose, Glucoseamine HCL & Algae Extract & Saccharomyces Cerevisiae (Yeast Extract) & Urea, Steareth-20 & Palmitoyl Tetrapeptide-7, Centella Asiatica Extract & Echinacea Purpurea Extract, Hydrolyzed Vegetable Protein, Imperata Cylindrica (Root) Extract & PEG-8 & Carbomer, Phenoxyethanol & Caprylyl Glycol & Ethylhexylglycerin & Hexylene Glycol, Polyglyceryl Methacrylate & Propylene Glycol & Palmitoyl Oligopeptide, Cyclopentasiloxane & Dimethicone, Stearic Acid, Mangifera Indica (Mango) Seed Butter, Tocopheryl Acetate, Glycyrrhiza Glabra Root Extract, Arctostaphylos Uva Ursi Leaf Extract, Chlorella Vulgaris Extract, Corallina Officinalis Extract, Dipotassium Glycyrrhizate, PEG-8 & Tocopherol & Ascorbyl Palmitate & Ascorbic Acid & Citric Acid, Disodium EDTA, Magnesium Aluminum Silicate, Xanthan Gum, Triethanolamine, Retinyl Palmitate, Lavandula Angustifolia (Lavender) Oil

[ pH: 5.75 ]

This advanced eye care treatment is expertly formulated to diminish the depth, increase firmness & elasticity, and to counteract skin slackening to the highly wrinkle prone and fragile eye area. Featuring (4) major peptides (Argireline, Matrixyl, Eyeliss, & Regu-age), the A&M Eye Recovery Therapy is our most potent eye treatment, and is recommended for mature skin.

Directions: Using fingertips, massage to surrounding eye areas affected by wrinkles due to muscle contractions. Also use in the nasal labial area. For best results, apply once per evening, followed by the A&M Facial Recovery Therapy, and/or the Vitamin A Facial Cream + III.

Ingredients Highlights: Palmitoyl Pentapeptide-4 (Matrixyl): Stimulates the skins fibroblasts to rebuild the extra-cellular matrix, including the synthesis of Collagen I and Collagen IV, fibronectin and of Glycosaminoglycans. It also stimulates the production of dermal matrix (Collagen I & III) resulting in a significant reduction of wrinkles and fine lines of the skin. Acetyl Hexapeptide-8 (Argireline): Reduces facial wrinkle depth and the signs of skin aging resulting from facial movements and facial muscle contraction by halting the release of neurotransmitters from SNARE and catecholamine complexes, (which can also induce formation of wrinkles and fine lines to the skin). Dipeptide-2 & Palmitoyl Tetrapeptide-7 (Eyeliss): Combats the effect of tiredness and hypertension, as well as the natural effects of aging, which contribute to the formation of bags under the eyes, Eyeliss is an outstanding anti-aging ingredient. Soy Peptides & Hydrolyzed Rice Bran Extract (Regu-Age): A highly active complex of specially purified soy and rice peptides and biotechnologically derived yeast protein, Regu-Age effectively addresses dark circles and puffiness around the eyes.

Other Ingredients: Water, Sodium Hyaluronate, Centella Asiatica Extract & Echinacea Purpurea Extract, Xanthan Gum-Chondrus Crispus & Glucose, Lecithin & Dipalmitoyl Hydroxyproline, Imperata Cylindrica Extract, PEG-8 Dimethicone, Cyclomethicone

[ pH: 6.25 ]

An advanced age management treatment that blends the most tried and true peptides and delivery systems, Argireline & Matrixyl, helping to prevent skin aging induced by repeated facial movement caused by excessive catecholamine release. Stimulating the deeper layers of the skin, the A&M Facial Recovery Therapy provides diminished wrinkle depth, as well as an increase in the elasticity and firmness of the skin. Recommend for mature skin types.

Directions: Using fingertips apply to facial areas and massage into skin once per evening, allowing it to absorb into the skin. Apply directly after the A&M Eye Recovery Therapy.

Ingredients Highlights: Palmitoyl Pentapeptide-4: Stimulates the skins fibroblasts to rebuild the extra-cellular matrix, including the synthesis of Collagen I and Collagen IV, fibronectin and of Glycosaminoglycans. It also stimulates the production of dermal matrix (Collagen I & III) resulting in a significant reduction of wrinkles and fine lines of the skin. Acetyl Hexapeptide-8: Reduces facial wrinkle depth and the signs of skin aging resulting from facial movements and facial muscle contraction by halting the release of neurotransmitters from SNARE and catecholamine complexes, (which can also induce formation of wrinkles and fine lines to the skin).

Other Ingredients: Deionized Water, Sodium Hyaluronate, Lecithin & Dipalmitoyl Hydroxyproline, Hydrocotyl & Coneflower Extracts, Glycosaminoglycans, Glucosamine HCI & Alagae Extract & Yeast Extract & Urea, Magnesium Ascorbyl Phosphate, Glycine HCL, Retinyl Palmitate

[ pH: 6.25 ]

Addressing the multiple problems of sun and age damaged skin, the Intensive Clarifying Facial Cream + III is a glycolic acid based moisturizer featuring three potent skin lighteners; Kojic Acid, Licorice, and Hydro- quinone (2%), which quickly & effectively treat hyperpigmentation & discolorations.

Vitamin C Ester (Tetrahexyldecyl Ascorbate) is a stable, oil-soluble form of Vitamin C, providing high level skin lightening, enhanced collagen synthesis, and increased DNA & UV protection with higher absorption capabilities and less irritating than Ascorbic Acid.

Because of how well it protects the skins collagen fibers, ascorbic acid based Vitamin C is widely considered one of the most effective antioxidants for skin rejuvenation & revitalization. The 20% Vitamin C Lightening drops combine a potent concentration of ascorbic acid with aloe, green tea leaf extract, and mushroom extract. *Also available is our original Vitamin C Serum, containing a milder blend of ascorbic acid (14%).

The Anti-Wrinkle Eye Cream contains a high potency blend of peptides, including EyelissTM & Regu-age (in addition to Argireline & Matrixyl) which work synergistically to improve firmness, elasticity, and reduce puffiness & dark circles around the eye area.

Addressing the multiple problems of sun and age damaged skin, the Intensive Clarifying Facial Cream + III moisturizer combines three powerful lightening. Agents: Hydroquinone, Kojic Acid, & Licorice, with Alpha Lipoic Acid, Vitamin C, & Co-enzyme Q10, minimizing fine lines, evening skin tone, and naturally exfoliating the outer layer of the skin while providing a 15 sun protection factor (SPF).

Directions: Smooth a pearl sized drop onto the face once or twice daily. Avoid eye area. If used during the day, apply additional sun protection if skin is in contact with the sun for an extended period (twenty minutes or more).

Active Ingredients: Octyl Methoxycinnamate - 7.5% Octyl Salcylate - 5% Glycolic Acid - 4% Benzophenone - 3% Hydroquinone - 2%

Inactive Ingredients: Deionized Water, Glyceryl Stearate & PEG-100 Stearate, Ascorbic Acid (Vitamin C), Alpha Lipoic Acid, Co-enzyme Q 10, Kojic Acid, Cetyl Alcohol, Licorice, Palmitic Acid, Octyl Salcylate, Phenoxyethanol, Tocopheryl Acetate, Essential Oil of Rosewood, Disodium tEDTA

[ pH: 4.5 ]

Vitamin C Ester is a stable, oil-soluble form of Vitamin C, providing high level Skin Lightening, enhanced Collagen Synthesis, and increased DNA & UV protection with higher absorption capabilities than Ascorbic Acid.

Directions: On clean, dry skin, apply four to five drops directly onto the face once a day, avoiding the eye area.

Ingredients: Cyclomethicone, Tetrahexyldecyl Ascorbate (Vitamin C Ester 10%), PPG-12/SMDI Copolymer, Santalum Album Extract, Phellodendrone Amurense Bark Extract, Barley Extract, Jojoba Seed Oil/Buxus Chinensis, Tocopheryl Acetate, Phenoxyethanol, Tricholoma Matsutake Singer (Mushroom Extract), Ascorbyl Palmitate, Bisabolol

[ pH: 7.0 ]

Ascorbic acid based Vitamin C is widely considered one of the most effective antioxidants for rejuvenating mature skin due to its ability to protect the skins collagen fibers, and for its ability to help inhibit melanin production, creating a lightening effect to the skin. The 20% Vitamin C Lightening Drops combine a potent concentration of ascorbic acid with aloe, green tea extract, and an exotic mushroom extract (Tricholoma Matsutake Singer) for additional lightening.

Directions: On clean, dry skin apply four to five drops directly onto the face once daily. Avoid the eye area. Thoroughly wash hands after use. Though a light tingling sensation is normal, if irritation (redness) results after application, discontinue or reduce the frequency of use of the product.

Ingredients: Water (Aqua), Ascorbic Acid -20%, Ethoxydiglycol, Hydroxyethylcellulose, Phenoxyethanol, Polysorbate 20, Camellia Sinensis Leaf Extract, Aloe Barbadensis Leaf Extract, Mushroom Extract (Tricholoma Matsutake Singer)-Enzymes- Alcohol, Sodium Sulfite, Disodium EDTA

[ pH: 3.00 ]

The Anti-Wrinkle Eye Cream is formulated to reduce puffiness, enhances firmness, strengthens connective tissues, and to help diminish dark circles around the eye area. In contrast to the A&M Eye Recovery Therapy, the Anti-Wrinkle Eye Cream concentrates on the upper layers of the skin, making it a great day moisturizer for the eyes.

Directions: Apply around the eye area with the ring finger once daily. For best results, follow with a moisturizer and solar protection.

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anti-aging stem cells - Lucrece

1. Introduction

Bone marrow is a complex tissue containing hematopoietic cell progenitors and their progeny integrated within a connective-tissue network of mesenchymal-derived cells known as the stroma (1). The stroma cells, or Mesenchymal stem cells (MSCs), are multi-potent progenitor cells that constitute a minute proportion of the bone marrow, represented as a rare population of cells that makes up 0.001 to 0.01% of the total nucleated cells. They represent 10-fold less abundance than the haematopoietic stem cells (2), which contributes to the organization of the microenvironment supporting the differentiation of hematopoietic cells (3). MSC are present in tissues of young, as well as, adult individuals (4, 5), including the adipose tissue, umbilical cord blood, amniotic fluid and even peripheral blood (1, 6-8). MSCs were characterized over thirty years ago as fibroblast-like cells with adhesive properties in culture (9, 10). The term MSC has become the predominant term in the literature since the early 90s (11), after which their research field has grown rapidly due to the promising therapeutic potential, resulting in an increased frequency of clinical trials in the new millennium at an explosive rate.

As data accumulated, there was a need to establish a consensus on the proper definition of the MSCs. The International Society for Cellular Therapy has recommended the minimum criteria for defining multi-potent human MSCs (12, 13). The criteria included: (i) cells being adherent to plastic under standard culture conditions; (ii) MSC being positive for the expression of CD105, CD73 and CD90 and negative for expression of the haematopoietic cell surface markers CD34, CD45, CD11a, CD19 or CD79a, CD14 or CD11b and histocompatibility locus antigen (HLA)-DR; (iii) under a specific stimulus, MSC differentiate into osteocytes, adipocytes and chondrocytes in vitro. These criteria presented properties to purify MSC and to enable their expansion by several-fold in-vitro, without losing their differentiation capacity. When plated at low density, MSCs form small colonies, called colony-forming units of fibroblasts (CFU-f), and which correspond to the progenitors that can differentiate into one of the mesenchymal cell lineages (14, 15). It has been reported lately that MSCs are able to differentiate into both mesenchymal, as well as, non-mesenchymal cell lineages, such as adipocytes, osteoblasts, chondrocytes, tenocytes, skeletal myocytes, neurones and cells of the visceral mesoderm, both in vitro and in vivo (16, 17).

All cells have half-lives and their natural expiration must be matched by their replacement; MSCs, by proliferating and differentiating, can be the proposed source of these new replacement cells as characterized in their differentiation capacity. This replacement hypothesis mimics the known sequence of events involved in the turnover and maintenance of blood cells that are formed from haematopoietic stem cells (HSCs) (18). Unlike HSCs, MSCs can be culture-expanded ex vivo in up to 40 or 50 cell doublings without differentiation (19). A dramatic decrease in MSC per nucleated marrow cell can be observed when the results are grouped by decade, thus showing a 100-fold decrease from birth to old age. Being pericytes present in all vascularized tissues, the local availability of MSC decreases substantially as the vascular density decreases by one or two orders of magnitude with age (20). In recent years, the discovery of MSCs with properties similar, but not identical, to BM-MSCs has been demonstrated in the stromal fraction of the connective tissue from several organs, including adipose tissue, trabecular bone, derma, liver and muscle (21-24). It is important to note that the origin of MSCs might determine their fate and functional characteristics (25). Studies of human bone marrow have revealed that about one-third of the MSC clones are able to acquire phenotypes of pre-adipocytes, osteocytes and chondrocytes (16). This is in concordance with data showing that 30% of the clones from bone marrow have been found to exhibit a trilineage differentiation potential, whereas the remainder display a bi-lineage (osteo-chondro) or uni-lineage (osteo) potential (26). Moreover, MSC populations derived from adipose tissue and derma present a heterogeneous differentiation potential; indeed, only 1.4% of single cells obtained from adipose-derived adult stem cell (ADAS) populations were tri-potent, the others being bi-potent or unipotent (27).

Mesenchymal Stem Cells have been shown to possess immunomodulatory characteristics, as described through the inhibition of T-cell proliferation in vitro (28-30). These observations have triggered a huge interest in the immunomodulatory effects of MSCs. The in vitro studies have been complemented in vivo, where both confirmed the immunosuppressive effect of MSC. MSC activating stimuli in vitro, appears to include the secretion of cytokines and the interaction with other immune cells in the presence of proinflammatory cytokines (Fig 1) (31). Primarily, the in vivo effect has been originally shown in a baboon model, in which infusion of ex vivoexpanded matched donor or third-party MSCs delayed the time to rejection of histo-incompatible skin grafts (29). The delay indicated a potential role for MSC in the prevention and treatment of graft-versus-host disease (GVHD) in ASCT, in organ transplantation to prevent rejection, and in autoimmune disorders. Recently, MSCs were used to successfully treat a 9-year-old boy with severe treatment-resistant acute GVHD, further confirming the potent immunosuppressive effect in humans (32). The immunosuppressive potential has no immunologic restriction, whether the MSCs are autologous with the stimulatory or the responder lymphocytes or the MSCs are derived from a third party. The degree of MSC suppression is dose dependent, where high doses of MSC are inhibitory, whereas low doses enhance lymphocyte proliferation in MLCs (33). Broadly, MSC modulate cytokine production by the dendritic and T cell subsets DC/Th1 and DC/Th2 (34), block the antigen presenting cell (APC) maturation and activation (35), and increase the proportion of CD4+CD25+ regulatory cells in a mixed lymphocyte reaction (36).

Potential mechanisms of the MSC interactions with immune cells. Mesenchymal stem cells (MSCs) can inhibit both the proliferation and cytotoxicity of resting natural killer (NK) cells, as well as, their cytokine production by releasing prostaglandin E2 (PGE2), indoleamine 2,3-dioxygenase (IDO) and soluble HLA-G5 (sHLA-G5). Killing of MSCs by cytokine-activated NK cells involves the engagement of cell-surface receptors (Thick blue line) expressed by NK cells with its ligands expressed on MSCs. MSCs inhibit the differentiation of monocytes to immature myeloid dendritic cells (DCs), bias mature DCs to an immature DC state, inhibit tumour-necrosis factor (TNF) production by DCs and increase interleukin-10 (IL-10) production by plasmacytoid DCs (pDCs). MSC-derived PGE2 is involved in all of these effects. Immature DCs are susceptible to activated NK cell-mediated lysis. MSC Direct inhibition of CD4+ T-cell function depends on their release of several soluble molecules, including PGE2, IDO, transforming growth factor-1 (TGF1), hepatocyte growth factor (HGF), inducible nitric-oxide synthase (iNOS) and haem-oxygenase-1 (HO1). MSC inhibition of CD8+ T-cell cytotoxicity and the differentiation of regulatory T cells mediated directly by MSCs are related to the release of sHLA-G5 by MSCs. In addition, the upregulation of IL-10 production by pDCs results in the increased generation of regulatory T cells through an indirect mechanism. MSC-driven inhibition of B-cell function seems to depend on soluble factors and cellcell contact. Finally, MSCs dampen the respiratory burst and delay the spontaneous apoptosis of neutrophils by constitutively releasing IL-6.

Dendritic cells have the elementary role of antigen presentation to nave T cells upon maturation, which in turn induce the proinflammatory cytokines. Immature DCs acquire the expression of co-stimulatory molecules and upregulate expression of MHC-I and II, as well as, other cell-surface markers (CD11c and CD83). Mesenchymal stem cells have profound effect on the development of DC, where in the presence of MSC, the percentage of DC with conventional phenotype is reduced, while that of plasmacytoid DC is increased, therefore biasing the immune system toward Th2 and away from Th1 responses in a PGE-2 dependent mechanism (37). Furthermore, MSCs inhibit the up-regulation of CD1a, CD40, CD80, CD86, and HLA-DR during DC differentiation and prevent an increase of CD40, CD86, and CD83 expression during DC maturation (38). When mature DCs are incubated with MSCs they have a decreased cell-surface expression of MHC class II molecules, CD11c, CD83 and co-stimulatory molecules, as well as, decreased interleukin-12 (Il-12) production, thereby impairing the antigen-presenting function of the DCs (Fig 1) (39, 40). MSCs can also decrease the pro-inflammatory potential of DCs by inhibiting their production of tumour-necrosis factor (TNF-) (40). Furthermore, plasmacytoid DCs (pDCs), which are specialized cells for the production of high levels of type-I IFN in response to microbial stimuli, upregulate production of the anti-inflammatory cytokine IL-10 after incubation with MSCs (34). These observations indicate a potent anti-inflammatory and immunoregulatory effect for MSC in vitro and potentially in vivo.

Natural killer (NK) cells are key effector cells of the innate immunity in anti-viral and anti-tumor immune responses through their Granzyme B mediated cytotoxicity and the production of pro-inflammatory cytokines (41). NK-mediated target cell lysis results from an antigen-ligand interaction realized by activating NK-cell receptors, and associated with reduced or absent MHC-I expression by the target cell (42). MSCs can inhibit the cytotoxic activity of resting NK cells by down-regulating expression of NKp30 and natural-killer group 2, member D (NKG2D), which are activating receptors involved in NK-cell activation and target-cell killing (Fig 1) (43). Resting NK cells proliferate and acquire strong cytotoxic activity when cultured with IL-2 or IL-15, but when incubated with MSC in the presence of these cytokines, resting NK-cell, as well as, pre-activated NK cell proliferation and IFN- production are almost completely abrogated (44, 45). It is worth noting that although the susceptibility of NK cells to MSC mediated inhibition is potent, the pre-activated NK cells showed more resistance to the immunosuppressive effect of MSC compared to resting NK cells (43). The susceptibility of human MSCs to NK-cell-mediated cytotoxicity depends on the low level of cell-surface expression of MHC class I molecules by MSCs and the expression of several ligands, that are recognized by activating NK-cell receptors. Autologous and allogeneic MSC were susceptible to lysis by NK cells (43), where NK cell-mediated lysis was inversely correlated with the expression of HLA class I on MSC (46). Incubation of MSCs with IFN- partially protected them from NK-cell-mediated cytotoxicity, through the up-regulation of expression of MHC-I molecules on MSCs (43). Taken together, a possible dynamic interaction between NK cells and MSC in vivo exists, where the latter partially inhibit activated MSC, without compromising their ability to kill MSC, reflecting on an interaction tightly regulated by IFN- concentration.

Neutrophils play a major role in innate immunity during the course of bacterial infections, where they are activated to kill foreign infectious agents and accordingly undergo a respiratory burst. MSCs have been shown to dampen the respiratory burst and to delay the spontaneous apoptosis of resting and activated neutrophils through an IL-6-dependent mechanism (47). MSC had no effect on neutrophil phagocytosis, expression of adhesion molecules, and chemotaxis in response to IL-8, f-MLP, or C5a (47). Stimulation with bacterial endotoxin induces chemokine receptor expression and mobility of MSCs, which secrete large amounts of inflammatory cytokines and recruit neutrophils in an IL-8 and MIF-dependent manner. Recruited and activated neutrophils showed a prolonged lifespan, an increased expression of inflammatory chemokines, and an enhanced responsiveness toward subsequent challenge with LPS, which suggest a role for MSCs in the early phases of pathogen challenge, when classical immune cells have not been recruited yet (48). Furthermore, MSC have shown the capability to mediate the preservation of resting neutrophils, a phenomenon that might be important in those anatomical sites, where large numbers of mature and functional neutrophils are stored, such as the bone marrow and lungs (49).

T-cells are major players of the adaptive immune response. After T-cell receptor (TCR) engagement, T cells proliferate and undergo numerous effector functions, including cytokine release and, in the case of CD8+ T cells (CTL), cytotoxicity. Abundant reports have shown that T-cell proliferation stimulated with polyclonal mitogens, allogeneic cells or specific antigen is inhibited by MSCs (28, 29, 50-56). The observation that MSCs can reduce T cell proliferation in vitro is mirrored by the in vivo finding through infusions of hMSCs that control GVHD following bone marrow transplantation. Nevertheless, there is no demonstrable correlation between the measured effects of MSCs in vitro and their counter effect in vivo due to the lack of universality of methodology correlating the in vitro findings with the in vivo therapeutic potential.

MSC inhibition of T-cell proliferation is not MHC restricted, since it can be mediated by both autologous and allogeneic MSCs and depends on the arrest of T-cells in the G0/G1 phase of the cell cycle (55, 57). Thus, MSCs do not promote T-cell apoptosis, but instead maintain T cell survival upon subjection to overstimulation through the TCR and upon commitment to undergo CD95CD95-ligand-dependent activation-induced cell death (57). MSC effects on T cell proliferation in vitro appear to have both contact-dependent and contact-independent components (58). Inhibition of T-cell proliferation by MSCs leads to decreased IFN- secretion in vitro and in vivo associated with increased IL-4 production by T helper 2 (TH2) cells (34, 59). Taken together, there is an implication for a shift from a pro-inflammatory state characterized by IFN- secretion to an anti-inflammatory state characterized by IL-4 secretion (Fig 1). An imperative role for effector T-cell is the MHC restricted killing of virally-infected or of allogeneic cells mediated via CD8+ CTLs, and which is down-regulated by MSC (60).

Regulatory T cells (Tregs), a subpopulation of T cells, are vital to keep the immune system in check, help avoid immune-mediated pathology and contain unrestricted expansion of effector T-cell populations, which results in maintaining homeostasis and tolerance to self antigens. Tregs are currently identified by co-expression of CD4 and CD25, expression of the transcription factor FoxP3, production of regulatory cytokines IL-10 and TGF-, and ability to suppress proliferation of activated CD4+CD25+ T cells in co-culture experiments. Differential expression of CD127 (-chain of the IL-7 receptor) enable flow cytometry-based separation of human Tregs from CD127+ non-regulatory T-cells (61). MSCs have been reported to induce the production of IL-10 by pDCs, which, in turn, trigger the generation of regulatory T cells (Fig 1) (34, 40). Furthermore, Tregs secrete TGF- and when used in vitro, TGF- in combination with IL-2 directs the differentiation of T-cells into Tregs, while Tregs suppress the proliferation of TCR-dependent proliferation of effector CD25null or CD25low T-cells in a non-autologous fashion. Also TGF- alters angiogenesis following injury in experiments using MSC (62). In addition, after co-culture with antigen-specific T-cells, MSCs can directly induce the proliferation of regulatory T-cells through release of the immunomodulatory HLA-G isoform HLA-G5 (Fig 1) (63). Taken together, MSCs can modulate the intensity of an immune response by inhibiting antigen-specific T-cell proliferation and cytotoxicity and promoting the generation of regulatory T-cells.

Antibody producing B-cells constitute the second main cell type involved in adaptive immunity. Interactions between MSCs and B-cells have produced controversial results attributable to the inconsistent experimental conditions used (31, 55, 64). Initial reports on mice suggested that MSC exercise a dampening effect on the proliferation of B-cells (64), which is in concordance with most published works to date (31, 55, 64). Furthermore, MSCs can also inhibit B-cell differentiation and constitutive expression of chemokine receptors via the release of soluble factors and cell-cell contact mediated possibly by the Programmed Cell Death 1 (PD-1) and its ligand (31, 64). The addition of MSCs, at the beginning of a mixed lymphocyte reaction (MLC), considerably inhibited immunoglobulin production in standard MLC, irrespective of the MSC dose employed, which suggests that third-party MSC are able to suppress allo-specific antibody production, consequently, overcoming a positive cross-match in sensitized transplant recipients (65). However, other in vitro studies have shown that MSCs could support the survival, proliferation and differentiation to antibody-secreting cells of B-cells from normal individuals and from pediatric patients with systemic lupus erythematosus (66, 67). A major mechanism of B-cell suppression was hMSC production of soluble factors, as indicated by transwell experiments, where hMSCs inhibited B-cell differentiation shown as significant impairment of IgM, IgG, and IgA production. CXCR4, CXCR5, and CCR7 B-cell expression, as well as chemotaxis to CXCL12, the CXCR4 ligand, and CXCL13, the CXCR5 ligand, were significantly down-regulated by hMSCs, suggesting that these cells affect chemotactic properties of B-cells (Fig 1). B-cell costimulatory molecule expression and cytokine production were unaffected by hMSCs (64). Regardless of the controversial in vitro effects, B-cell response is mainly a T-cell dependent mechanism, and thus its outcome is significantly influenced by the MSC-mediated inhibition of T-cell functions. More recently, Corcione et al have shown that systemic administration of MSCs to mice affected by experimental autoimmune encephalomyelitis (EAE), a prototypical disease mediated by self-reactive T cells, results in striking disease amelioration mediated by the induction of peripheral tolerance (52). In addition, it has been shown that tolerance induction by MSCs may occur also through the inhibition of dendritic-cell maturation and function (34, 35), thus suggesting that activated T cells are not the only targets of MSCs.

Low concentrations of IFN- upregulate the expression of MHC-II molecules by MSCs, which indicates that they could act as antigen presenting cells (APCs) early in an immune response, when the level of IFN- are low (68, 69). However, this process of MHC-II expression by MSCs, along with the potential APC characteristics, was reversed as IFN- concentrations increased. These observations could suggest that MSCs function as conditional APC in the early phase of an immune response, while later switch into an immunosuppressive function (68). Since bone marrow might be a site for the induction of T-cell responses to blood-borne antigens (70), and since MSC are derived from the stromal progenitor cells that reside in the bone marrow, therefore, MSC express a yet unidentified role in the control of the immune response physiology of the bone marrow. Dendritic cells are the main APC for T-cell responses, and MSC-mediated suppression of DC maturation would prohibit efficient antigen presentation and thus, the clonal expansion of T-cells. Direct interactions of MSCs with T-cells in vivo could lead to the arrest of T-cell proliferation, inhibition of CTL-mediated cytotoxicity and generation of CD4+ regulatory T-cells. As a consequence, impaired CD4+ T-cell activation would translate into defective T-cell help for B-cell proliferation and differentiation to antibody-secreting cells.

The hMSCs express few to none of the B7-1/B7-2 (CD80/CD86) costimulatorytype molecules; this appears to contribute, at least in part, to their immune privilege characteristic. Mechanisms that lead to immune tolerance rely on interrelated pathways that involve complex cross talk and cross regulation of T-cells and APCs by one another. Both soluble mediators and modulation exerted via complex networks of cytokines and costimulatory molecules appear to play a role in MSC regulation of T cells, and these mechanisms invoke both contact-dependent and -independent pathways.

Although many of the studies use MSC-conditioned medium, both contact-dependent and -independent mechanisms are probably invoked in the therapeutic use of MSCs (20, 71). In addition to cytokines, the network of costimulatory molecules is hypothesized to play a prominent role in modulating tolerance and inflammation. MSCs down-regulate the expression of costimulatory molecules (30, 72, 73). The discovery of new functions for B7 family members, together with the identification of additional B7 and CD28 family members, is revealing new ways in which the B7:CD28 family may regulate T-cell activation and tolerance. Not only do CD80/86:CD28 interactions promote initial T-cell activation, they also regulate self-tolerance by supporting CD4+CD25+ Treg homeostasis (74-76). Cytotoxic T-lymphocyte antigen 4 (CTLA-4) can exert inhibitory effects in both B7-1/B7-2dependent and independent fashions. B7-1 and B7-2 can signal bi-directionally through engaging CD28 and CTLA-4 on T cells and by delivering signals into B7-expressing cells (77). The B7 family membersinducible co-stimulator (ICOS) ligand, PD-L1 (B7-H1), PD-L2 (B7-DC), B7-H3, and B7-H4 (B7x/B7-S1)are expressed on professional APC cells, while B7-H4 and B7-H1 are expressed on hMSCs and on cells within non-lymphoid organs. These observations may provide a new means for regulating T-cell activation and tolerance in peripheral tissues (31, 71, 78). ICOS and PD-1 are expressed upon T-cell-induction, and they regulate previously activated T-cells (79). Both the ICOS:ICOSL pathway and the PD-1:PD-L1/PD-L2 pathway play a critical role in regulating T-cell activation and tolerance (79). There is consensus that both CTLA-4 and PD-1 inhibit T-cell and B-cell activation and may play a crucial role in peripheral tolerance (79, 80). Both CTLA-4 and PD-1 functions are associated with Rheumatoid Arthritis (RA) and other autoimmune diseases. PD-1 is over expressed on CD4+ T cells in the synovial fluid of RA patients (81). Whether or not these costimulatory molecules are critical mediators of MSC-mediated immune suppression and/or tolerance in vivo is still under current investigation.

Studies have shown that MSCs escape the immune system, and this makes them a potential therapeutic tool for various transplantation procedures. MSCs express intermediate levels of HLA major histocompatibility complex (MHC) class I molecules (16, 50, 82, 83), while they do not express HLA class II antigens of the cell surface. However, HLA class II is readily detectable by Western blot on whole-cell lysates of unstimulated adult MSCs, thus suggesting that MSCs contain intracellular deposits of HLA class II allo-antigens (83). Cell-surface expression can be induced by treatment of the cells with IFN- for 1 or 2 days. Unlike adult MSCs, the fetal liver derived cells have no intracellular nor cell surface HLA class II expression (84), but incubation with IFN- initiated their intracellular expression followed by surface expression. Differentiation of MSCs into their mesodermal lineages of bone, cartilage, or adipose tissue, both in adult and fetal MSCs continued to express HLA-I, but not class II (84). Undifferentiated MSCs in vitro fail to elicit a proliferative response from allogeneic lymphocytes, thus suggesting that the cells are not inherently immunogenic (28, 30, 50). When pre-cultured with IFN- for full HLA class II expression, MSCs still escape recognition by allo-reactive T-cells, (83, 84) as is the case with MSCs differentiated adipocytes, osteoblasts, and chondrocytes. Limited in vivo data demonstrate the persistence of allogeneic MSCs into immunocompetent hosts after transplantation. In one patient treated with MSCs, DNA of donor MSC could not be detected in any organ at autopsy few weeks after the infusion, while in another patient receiving MSCs from two donors, the donor DNA from both donors was detected in lymph node and colon, the target organs of GVHD, within weeks after infusion (85). Data from our lab indicated that MSC were undetectable after two weeks in an allogeneic system (86). Therefore, the question of whether MSCs are recognized by an intact allogeneic immune system in vivo remains open, although the in vitro data support the theory that MSCs escape the immune system. MSCs do not express FAS ligand or costimulatory molecules, such as B7-1, B7-2, CD40, or CD40L (50). When costimulation is inadequate, T-cell proliferation can be induced by the addition of exogenous costimulation. However, MSCs differ from other cell types, and no T-cell proliferation can be observed when they are cultured with HLA-mismatched lymphocytes in the presence of a CD28-stimulating antibody (50). However, in agreement with the in vitro data, infusion or implantation of allogeneic and MHC-mismatched MSCs into baboons has been well tolerated (87-89). Unique immunologic properties of MSCs were also suggested by the fact that engraftment of human MSCs occurred after intra-uterine transplantation into sheep, even when the transplantation was performed after the fetuses became immunocompetent (90). MSC mainly fail to activate T-cells and show to be targets for CD8+ T cell-cytotoxicity, althought controversial (60). Phyto-hemagglutinin (PHA) blasts, generated to react against a specific donor, will lyse chromium-labeled mononuclear cells from that individual but it will not lyse MSCs derived from the same donor. Furthermore, killer cell inhibitory receptor (KIR ligand)mismatched natural killer cells do not lyse MSCs (60). Thus, MSCs, although incompatible at the MHC, tend to escape the immune system.

Although MSCs are transplantable across allogeneic barriers, a delayed type hypersensitivity reaction can lead to rejection in xenogenic models of human MSCs injected into immunocompetent rats (91). In this study, MSCs were identified in the heart muscle of severe compromised immune deficiency rats, in contrast to that of immunocompetent rats. In the latter group, peripheral blood lymphocytes proliferated after re-stimulation with human MSCs in vitro, thus suggesting cellular immunization. Such a proliferative response in vitro has not been detected in humans treated with intravenous (IV) infusion of allogeneic MSCs (Le Blanc and Ringdn, unpublished data, 2004).

Several studies have acknowledged the immunosuppressive activities of MSCs, but the underlying mechanisms are far from being fully characterized. The initial step in the interaction between MSCs and their target cells involves cellcell contact mediated by adhesion molecules, in concordance with studies showing the dependence of T-cell proliferation on the engagement of PD-1 by its ligand (31). Several soluble immunosuppressive factors, either produced constitutively by MSCs or released following cross-talk with target cells have been reported, including nitric oxide and indoleamine 2,3-dioxygenase (IDO), which are only released by MSC after IFN- stimulation with target cells (92, 93), and thus not in a constitutive manner. IDO induces the depletion of tryptophan from the local environment, which is an essential amino acid for lymphocyte proliferation. MSC-derived IDO was reported as a requirement to inhibit the proliferation of IFN--producing TH1 cells (92) and together with prostaglandin E2 (PGE-2) to block NK-cell activity (Fig 1) (44). In addition, IFN-, alone or in combination with TNF-, IL-1 or IL-1, stimulates the production of chemokines by mouse MSCs that attract T-cells and stimulate the production of inducible nitric-oxide synthase (iNOS), which in turn inhibits T-cell activation through the production of nitric oxide (56). It is worth noting that MSCs from IFN- receptor (IFN--R1) deficient mice do not have immunosuppressive activity, which highlights the vital role of IFN- in the immunosuppressive function of MSC (56).

Additional soluble factors, such as transforming growth factor-1 (TGF-1), hepatocyte growth factor (HGF), IL-10, PGE-2, haem-oxygenase-1 (HO1), IL-6 and soluble HLA-G5, are constitutively produced by MSCs (28, 34, 51, 63, 94) or secreted in response to cytokines released by target cells upon interacting with MSC. TNF- and IFN- have been shown to stimulate the production of PGE-2 by MSCs above constitutive level (34). Furthermore, IL-6 was shown to dampen the respiratory burst and to delay the apoptosis of human neutrophils by inducing phosphorylation of the transcription factor signal transducer and activator of transcription 3 (47), and to inhibit the differentiation of bone-marrow progenitor cells into DCs (95).

The failure to reverse suppression, when neutralizing antibodies against IL-10, TGF- and IGF were added to MLR reactions does point to the possibility that MSC may secrete as yet uncharacterized immunosuppressive factors (93). Galectin-1 and Galectin-3, newly characterized lectins, are constitutively expressed and secreted by human bone marrow MSC. Inhibition of galectin-1 and galectin-3 gene expression with small interfering RNAs abrogated the suppressive effect of MSC on allogeneic T-cells (Fig 1) (96). The restoration of T-cell proliferation in the presence of - lactose indicates that the carbohydrate-recognition domain of galectins is responsible for the immunosuppression of T-cells and highlights an extracellular mechanism of action for the MSC-secreted galectins. In this respect, the inhibition of T-cell proliferation could result from either direct effects of galectin-1 and galectin-3 on T cells and/or through a direct or an indirect on effect on dendritic cells (97).

HLA-G5 represents another important molecule involved in MSC mediated regulation of the immune response, where its production has been shown to suppress T-cell proliferation, as well as NK-cell and T-cell cytotoxicity, while promoting the generation of Tregs (63, 98). HLA-G protein expression is constitutive and the level is not modified upon stimulation by allogeneic lymphocytes in MSC/MLR. HLA-G5 is detected on MSCs by real-time reverse-phase polymerase chain reaction, immune-fluorescence, flow cytometry and enzyme-linked immunosorbent assay in the supernatant (99). Cell contact between MSCs and activated T-cells induces IL-10 production, which, in turn, stimulates the release of soluble HLA-G5 by MSCs (63). It is worth nothing that none of these molecules have an exclusive role and that MSC-mediated immune-regulation is a redundant system that is mediated by several molecules.

One important characteristic of hMSCs is their ability to suppress inflammation resulting from injury, as well as, resulting from allogeneic solid organ transplants, and autoimmune disease. Consistent with in vitro studies, murine allogeneic MSCs are effective cellular therapy models in the treatment of murine models of human disease (52, 100-102). Several studies have documented the substantial clinical improvements observed in animal models, when MSC were systemically introduced as a therapy in mouse models of multiple sclerosis (102, 103), inflammatory bowel disease (104-106), infarct, stroke, and other neurologic diseases (107, 108), as well as diabetes (109). These findings strongly suggest that xenogeneic hMSCs are not immunologically recognized by various immunocompetent mouse models of disease and are able to home to sites of inflammation. However, the mechanisms behind the immunosuppressive actions at the site of inflammation and its association with the homing activity have not yet been completely elucidated.

Nitric Oxide (NO) mediate its effect partly through phosphorylation of Stat-5, which results in suppression of T- cell proliferation, partly through the inhibition of NO synthase or the inhibition of prostaglandin synthesis. This reveals the MSC-dependent effects on proliferation. Although indoleamine-2, 3-dioxygenase (IDO) has been hypothesized to be critical in mediating the effect of NO, neutralizing IDO by using a blocking antibody did not interfere with NOs suppressive effects (93, 110).

Within an in vivo setting, injury, inflammation, and/or foreign cells can lead to T-cell activation, which results in those T-cells producing proinflammatory cytokines including, but not limited to, TNF-, IFN-, IL-1, and IL-1. Combinations of cytokines may also induce cell production of chemokines, some of which bind to CXCR3-R expressing cells (including T cells) that co-localize with MSCs. MSCs then produce NO, which inhibits Stat-5 phosphorylation, thereby leading to cell-cycle arrest (and thus halting T cell proliferation) (Fig 1) (110). In addition, iNOS appears to be important in mouse MSC in vivo effects. MSCs from mice that lack iNOS (or IFN-R1) are unable to suppress GVHD. In contrast to mouse MSCs that use NO in mediating their immune-suppressive effect, hMSCs and MSCs from non-human primates appear to mediate their immune-suppressive effects via IDO (56). There is some controversy about whether the effect of IDO results from local depletion of tryptophan, or from the accumulation of tryptophan metabolites (which is suggested by data showing that use of a tryptophan antagonist, 1-methyl-L tryptophan, restored allo-reactivity that would otherwise have been suppressed by MSCs). In addition to its effect on the JAK-STAT pathway, NO may also influence mitogen activated protein kinase and nuclear factor B, which would cause a reduction in the gene expression of proinflammatory cytokines.

The clinical experience with and the safety of MSCs is of utmost interest for their wide therapeutic applications. The pioneering in vivo studies with MSC focused on the engraftment facilitation for the haematopoietic stem cells (111). Further work also focused on the regenerative functions of MSC in terms of functional repair of damaged tissues (112). Hypoimmunogenicity of MSC provided a critical advantage in their use for clinical and therapeutic purposes in vitro (50), followed by pre-clinical studies (29) and reaching the human clinical studies (32) with the use of allogeneic donors. Allogeneic MSC have proved to be an option with major advantages in clinical use, since the use of autologous MSC is hindered by the limited time frame for clonal expansion and the costly in vitro proliferation. However, some sub-acute conditions, such as autoimmune diseases, might allow the use of autologous MSCs and their culture in vitro. It is worth noting that some reports have recently challenged the belief that allogeneic MSCs are poorly immunogenic (113, 114), indicating that in some cases an autologous MSC source could be advantageous. Recent reports have shown that MSCs from patients with autoimmune disease have a normal capability to support hematopoiesis, (115) and to exercise immunomodulatory functions (116), and to show a normal phenotypic characteristics (117).

The perspective role of adult stem cells in degenerative disease conditions, where there is progressive tissue damage and an inability to repair, possibly due to the depletion of stem cell populations or functional alteration, has been considered. In cases of osteoarthritis, a disease of the joints where there is progressive and irreversible loss of cartilage characterized by changes in the underlying bone, Murphy et al showed that the proliferative capacity of the MSC was substantially reduced, and this was independent of the harvest site from patients with end-stage OA undergoing joint replacement surgery (118). In this study the marrow samples were harvested both from the site of surgery (either the hip or the knee) and also from the iliac crest. These effects were apparently disease-related, and not age-related. However, the data suggest that susceptibility to OA and perhaps other degenerative diseases may be due to the reduced mobilization or proliferation of stem cells. In addition, successfully recruited cells may have a limited capacity to differentiate, leading to defective tissue repair. Alternatively, the altered stem cell activity may be in response to the elevated levels of inflammatory cytokines seen in OA, which was confirmed by several other investigators (119, 120).

Similarly, the functional impairment of the anti-proliferative effect of MSCs derived from patients with aplastic anaemia (121) or multiple myeloma (122) might be resulting from an intrinsic abnormality in the microenvironment of the bone marrow, which is consistent with the possible use of autologous MSC for therapeutic purposes.

With the knowledge of the homing capacity of MSC and their capacity to engraft into the recipients bone after systemic administration, MSCs have been utilized to treat children with severe osteogenesis imperfecta, leading to improved parameters of increased growth velocity and total body mineral content associated with fewer fractures (123). Systemic infusion of allogeneic MSCs also led to encouraging bone marrow recovery in patients with tumors following chemotherapy (123). The immunosuppressive effect of infused MSCs has been successfully shown in acute, severe graft-versus-host disease (GvHD) (32). The probable effect of MSC was the inhibition of donor T-cell reactivity to histocompatibility antigens of the recipient tissue. Currently, there is no successful therapy for steroid-refractory acute GVHD. The possible role of MSCs in this context is therefore of potential interest. Le Blanc et al reported a case of grade IV acute GVHD of the gut and liver in a patient who had undergone ASCT with cells from an unrelated female donor (32). The patient was unresponsive to all types of immunosuppression drugs. When the patient was infused with 2x 106 MSCs per kilogram from his HLA-haploidentical mother, his GVHD responded with a decline in bilirubin and normalization of stools. After the MSC infusion, DNA analysis of his bone marrow showed the presence of minimal residual disease (124). When immunosuppression was discontinued, the patient again developed severe acute GVHD, with its associated symptoms within a few weeks.

Modulation of host allo-reactivity led to accelerated bone-marrow recovery in patients co-transplanted with MSCs and haplo-identical HSCs (125). Clinical trials are being conducted on the immunomodulatory potential of MSCs in the treatment of Crohns disease, with the potential for those cells to contribute to the regeneration of gastrointestinal epithelial cells (126).

As described previously, MSCs are characterized by their hypoimmunogenicity. In 2000, data from several research groups demonstrated long-term allo-MSC engraftment in a variety of non-cardiac tissues in the absence of immunosuppression (88, 90). On the basis of these observations, investigators began to look into the possibility of allo-MSCs engraftment into affected myocardium in rats, and later in swine, where allo-MSCs were found to readily engraft in necrotic myocardium and favorably alter ventricular function (2). The allo-MSC engraftment occurred without evidence of immunologic rejection or lymphocytic infiltration in the absence of assisted immunosuppressive therapy emphasizing some of the apparent advantages of these cells over other cell populations for cellular cardiomyoplasty. The immunologically privileged status of MSCs was also observed in xenogeneic setting, where Saito et al injected MSC intravenously from C57BL/6 mice into immunocompetent adult Lewis rats (127). When these animals were later subjected to MIs, murine MSCs could be identified in the region of necrosis, and these cells expressed muscle specific proteins not present before coronary ligation.

Consistent with results from in vitro studies, murine allogeneic MSCs are effective in the treatment of murine models of human disease (52, 103, 128). Several studies have reported clinical improvements in mouse models of multiple sclerosis and amyotrophic lateral sclerosis, inflammatory bowel disease, stroke, diabetes, infarct and GVHD using I.V. injected xenogeneic hMSCs rather than allogeneic MSCs (108, 109). A major advantage in using hMSCs in mouse models of human disease is that the possibility of gathering mechanistic data through measuring biomarkers from body fluids or using noninvasive imaging technology, which may prove to be an advantage in clinical studies when applied on humans.

In experiments designed to study the trafficking of hMSCs, investigators used mouse models of severe erosive polyarthritis characterized by an altered transgene allele that results in chronic over-expression of TNF- and which resemble human RA patients (60, 72). The motive behind utilizing these mice models was to investigate similarities in MSC homing with mouse models of chronic asthma and acute lung injury. Injected hMSC revealed a reduction in ankle arthritis parameters associated with decrease appendage related erythema, possibly due to the MSC localization to ankle joints as revealed by bioluminescence (129). Similar observations for inducing tolerance were made using adipose-derived MSC, where Treg were induced in RA PBMC and in mouse models of arthritis (36, 130). Furthermore, studies of rheumatoid arthritis T-cells showed a down-regulation of effector response using adipose-derived MSCs (131). Variations in this potential described by the capability of MSCs to down-regulate collagen-induced arthritis, and in the ability to induce Tregs, depend on the source of MSC (mouse vs. human) and its characteristics (primary isolate of MSC line), which reflect on difference in function compared to primary expanded MSC (132). Other studies reported that in the collagen-induced model of arthritis, mice infused with MSCs have increased numbers of CD4+CD25+ cells that express FoxP3 and thus reveal a Treg phenotype (20). Recent data on collagen-induced arthritis model, where murine MSCs did not reveal therapeutic benefits against arthritis in vivo, but did show anti-proliferative effect in vitro suggest that there is no appropriate in vitro measures that can be accurately extrapolated into a potential therapeutic utility of MSCs in vivo, and that mouse MSCs show difference in functional characteristics to hMSC in terms of immunoregulatory capacity (133).

MSCs immunological properties appeared to have potential therapeutic advantages in other forms of autoimmune diseases, especially in type 1 diabetes. In NOD mouse model, several physiological defects that aim to maintain peripheral and central tolerance contribute to the development of autoimmune diabetes. These defects are summed up as a combination of immune cell dysfunction (including T-cell, NK cells, B-cells, and dendritic cells), associated with the presence of inflammatory cytokine milieu (134). MSCs possess specific immunomodulatory properties capable of halting autoimmunity through immunomodulation processes described in this chapter. The processes might be through a direct effect via the presentation of differential levels of negative costimulatory molecules and the secretion of regulatory cytokines that affect regulatory T-cells/autoreactive T-cells. Furthermore, MSCs could modulate the immunological dysregulation observed in antibody producing B-cells and cytotoxic NK cells. Dendritic cells have been shown to be defective in NOD mice characterized by higher levels of costimulation with a potential capability to shift to a TH-1 type of immune response.

In an experimental mouse model of diabetes induced by streptozotocin, it was observed that MSCs promoted the endogenous repair of pancreatic islets and renal glomeruli (109). Similarly, co-infusion of MSCs and bone-marrow cells inhibited the proliferation of -cell-specific T-cells isolated from the pancreas of diabetic mice and restored insulin and glucose levels through the induction of recipient-derived pancreatic -cell regeneration in the absence of trans-differentiation of MSCs (135). These studies show that the in vivo administration of MSCs is clinically efficacious through the modulation of pathogenic - and T-cell responses and through potent bystander effects on the target tissue.

The timing of MSC infusion seems to be a critical parameter in their therapeutic efficacy. In the EAE mouse model of multiple sclerosis, MSC systematically injected at disease onset ameliorated myelin oligodendrocyte glycoprotein (MOG)-induced EAE and further decreased the infiltration T-cells, B-cells and macrophages into the central nervous system (CNS). Furthermore, T cells isolated from the lymph nodes of MSC-treated mice did not proliferate after in vitro re-challenge with MOG peptide, which is an indication of the induction of T-cell anergy (52). Systematic injection of MSCs was found to inhibit the in vivo production of pathogenic plp-specific antibodies and to suppress the encephalitogenic potential of plp-specific T cells in passive-transfer experiments. In this model, the MSCs migrated to the lymphoid organs, as well as, to the inflamed CNS, where they exercised a protective effect on the neuronal axons in situ (135, 136). In these studies, the therapeutic effect of MSCs depended on the release of anti-apoptotic, anti-inflammatory and trophic molecules, as occurred in the case of stroke in rats (137), and, possibly, on the recruitment of local progenitors and their subsequent induction to differentiate into neural cells (138). As trophic effect, the MSCs appeared to favor oligo-dendrogenisis by neural precursor cells (139).

Several other studies have provided insights into the effects of MSCs mediated by cytokines. In a model of acute renal failure, the administration of MSCs increased the recovery of renal function through the inhibition of production of proinflammatory cytokines, such as Il-1, TNF and IFN, and through an anti-apoptotic effect on target cells (140). Along the same line, the anti-inflammatory activity of MSCs was revealed in a mouse model of lung fibrosis, where they inhibited the effects of IL-1-producing T cells and TNF-producing macrophages through the release of IL-1 receptor antagonist (IL-1RA) (141). The release of trophic factors such as the WNT-associated molecule secreted frizzled-related protein 2 (SFRp2), which leads to the rescue of ischemic cardiomyocytes and the restoration of ventricular functions represent another important function for MSC (142).

With all the promising therapeutic potential of MSC, there seems to be a growing concern about their association with tumors. The immunoregulatory and anti-proliferative effects of MSCs led to several studies investigating the inhibitory effect of MSCs on tumor growth. Inhibition or, more frequently, stimulation of tumor-cell proliferation in vitro and/or tumor growth in vivo by MSCs has been reported (143-145). The heterogeneous nature of the MSC populations and the different experimental tumor models used, contribute to the effect of tumors on MSC in which the microenvironment generated by tumors influence the behavior of MSCs (146). Two main mechanisms are probably involved in the enhancement of tumor growth by MSCs. First, the cell-to-cell cross-talk between MSCs and tumor cells contribute to tumor progression, thus integrating within the tumor stroma (147), and second, the suppressive effects of MSCs on the immune system of tumor-bearing hosts might facilitate tumorigenesis, as shown for the inhibition of melanoma rejection, possibly mediated by regulatory CD8+ T cells (144). Irrespective of the possible interactions between cancer cells, immune cells and MSCs, the potential risk of stimulating the growth cancer by MSCs must be considered.

As a whole, the data accumulated from preclinical and clinical data indicate that bone marrow-derived MSCs have, in addition to their therapeutic purposes in regenerative medicine, effects that can result from other characteristics, such as their anti-proliferative and anti-inflammatory properties. The immuno suppressive activity of MSCs provides means for inducing peripheral tolerance following systemic injection mediated through the inhibition of cell division, thereby preventing their responsiveness to antigenic triggers while maintaining them in a quiescent state. In addition, the clinical efficacy of MSCs in different experimental model seems to occur almost only during the acute phase of disease associated with limited trans-differentiation, which indicates that the therapeutic effectiveness of MSCs relies heavily on their ability to modify microenvironments. These modifications occur through the release of anti-inflammatory cytokines, and anti-apoptotic and trophic molecules that promote the repair and protection of damaged tissues, as well as, maintain the integrity of the immune cells.

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Mesenchymal Stem Cells: Immunology and Therapeutic ...

Mesenchymal stem cells, or MSCs, are multipotent stromal cells that can differentiate into a variety of cell types,[1] including: osteoblasts (bone cells),[2]chondrocytes (cartilage cells),[3]myocytes (muscle cells)[4] and adipocytes (fat cells). This phenomenon has been documented in specific cells and tissues in living animals and their counterparts growing in tissue culture.

While the terms mesenchymal stem cell and marrow stromal cell have been used interchangeably, neither term is sufficiently descriptive:

The youngest, most primitive MSCs can be obtained from the umbilical cord tissue, namely Wharton's jelly and the umbilical cord blood. However the MSCs are found in much higher concentration in the Whartons jelly compared to the umbilical cord blood, which is a rich source of hematopoietic stem cells. The umbilical cord is easily obtained after the birth of the newborn, is normally thrown away, and poses no risk for collection. The umbilical cord MSCs have more primitive properties than other adult MSCs obtained later in life, which might make them a useful source of MSCs for clinical applications.

An extremely rich source for mesenchymal stem cells is the developing tooth bud of the mandibular third molar. While considered multipotent, they may prove to be pluripotent. The stem cells eventually form enamel, dentin, blood vessels, dental pulp, and nervous tissues, including a minimum of 29 different unique end organs. Because of extreme ease in collection at 810 years of age before calcification, and minimal to no morbidity, they will probably constitute a major source for personal banking, research, and multiple therapies. These stem cells have been shown capable of producing hepatocytes.

Additionally, amniotic fluid has been shown to be a rich source of stem cells. As many as 1 in 100 cells collected during amniocentesis has been shown to be a pluripotent mesenchymal stem cell.[9]

Adipose tissue is one of the richest sources of MSCs. There are more than 500 times more stem cells in 1 gram of fat than in 1 gram of aspirated bone marrow. Adipose stem cells are actively being researched in clinical trials for treatment of a variety of diseases.

The presence of MSCs in peripheral blood has been controversial. However, a few groups have successfully isolated MSCs from human peripheral blood and been able to expand them in culture.[10] Australian company Cynata also claims the ability to mass-produce MSCs from induced pluripotent stem cells obtained from blood cells using the method of K. Hu et al.[11][12]

Mesenchymal stem cells are characterized morphologically by a small cell body with a few cell processes that are long and thin. The cell body contains a large, round nucleus with a prominent nucleolus, which is surrounded by finely dispersed chromatin particles, giving the nucleus a clear appearance. The remainder of the cell body contains a small amount of Golgi apparatus, rough endoplasmic reticulum, mitochondria, and polyribosomes. The cells, which are long and thin, are widely dispersed and the adjacent extracellular matrix is populated by a few reticular fibrils but is devoid of the other types of collagen fibrils.[13][14]

The International Society for Cellular Therapy (ISCT) has proposed a set of standards to define MSCs. A cell can be classified as an MSC if it shows plastic adherent properties under normal culture conditions and has a fibroblast-like morphology. In fact, some argue that MSCs and fibroblasts are functionally identical.[15] Furthermore, MSCs can undergo osteogenic, adipogenic and chondrogenic differentiation ex-vivo. The cultured MSCs also express on their surface CD73, CD90 and CD105, while lacking the expression of CD11b, CD14, CD19, CD34, CD45, CD79a and HLA-DR surface markers.[16]

MSCs have a great capacity for self-renewal while maintaining their multipotency. Beyond that, there is little that can be definitively said. The standard test to confirm multipotency is differentiation of the cells into osteoblasts, adipocytes, and chondrocytes as well as myocytes and neurons. MSCs have been seen to even differentiate into neuron-like cells,[17] but there is lingering doubt whether the MSC-derived neurons are functional.[18] The degree to which the culture will differentiate varies among individuals and how differentiation is induced, e.g., chemical vs. mechanical;[19] and it is not clear whether this variation is due to a different amount of "true" progenitor cells in the culture or variable differentiation capacities of individuals' progenitors. The capacity of cells to proliferate and differentiate is known to decrease with the age of the donor, as well as the time in culture. Likewise, whether this is due to a decrease in the number of MSCs or a change to the existing MSCs is not known.[citation needed]

Numerous studies have demonstrated that human MSCs avoid allorecognition, interfere with dendritic cell and T-cell function, and generate a local immunosuppressive microenvironment by secreting cytokines.[20] It has also been shown that the immunomodulatory function of human MSC is enhanced when the cells are exposed to an inflammatory environment characterised by the presence of elevated local interferon-gamma levels.[21] Other studies contradict some of these findings, reflecting both the highly heterogeneous nature of MSC isolates and the considerable differences between isolates generated by the many different methods under development.[22]

The majority of modern culture techniques still take a colony-forming unit-fibroblasts (CFU-F) approach, where raw unpurified bone marrow or ficoll-purified bone marrow Mononuclear cell are plated directly into cell culture plates or flasks. Mesenchymal stem cells, but not red blood cells or haematopoetic progenitors, are adherent to tissue culture plastic within 24 to 48 hours. However, at least one publication has identified a population of non-adherent MSCs that are not obtained by the direct-plating technique.[23]

Other flow cytometry-based methods allow the sorting of bone marrow cells for specific surface markers, such as STRO-1.[24] STRO-1+ cells are generally more homogenous, and have higher rates of adherence and higher rates of proliferation, but the exact differences between STRO-1+ cells and MSCs are not clear.[25]

Methods of immunodepletion using such techniques as MACS have also been used in the negative selection of MSCs.[26]

The supplementation of basal media with fetal bovine serum or human platelet lysate is common in MSC culture. Prior the use of platelet lysates for MSC culture, the pathogen inactivation process is recommended to prevent pathogen transmission.[27]

Mesenchymal stem cells have been shown to contribute to cancer progression in a number of different cancers, particularly the Hematological malignancies because they contact the transformed blood cells in the bone marrow.[28]

The mesenchymal stem cells can be activated and mobilized if needed. However, the efficiency is very low. For instance, damage to muscles heals very slowly but further study into mechanisms of MSC action may provide avenues for increasing their capacity for tissue repair.[29][30]

Many of the early clinical successes using intravenous transplantation have come in systemic diseases like graft versus host disease and sepsis. However, it is becoming more accepted that diseases involving peripheral tissues, such as inflammatory bowel disease, may be better treated with methods that increase the local concentration of cells.[31] Direct injection or placement of cells into a site in need of repair may be the preferred method of treatment, as vascular delivery suffers from a "pulmonary first pass effect" where intravenous injected cells are sequestered in the lungs.[32] Clinical case reports in orthopedic applications have been published, though the number of patients treated is small and these methods still lack rigorous study demonstrating effectiveness. Wakitani has published a small case series of nine defects in five knees involving surgical transplantation of mesenchymal stem cells with coverage of the treated chondral defects.[33]

At least 218 clinical trials investigating the efficacy of mesenchymal stem cells in treating diseases have been initiated - many of which being autoimmune diseases.[34] Promising results have been shown in a variety of conditions, such as graft versus host disease, Crohn's disease, multiple sclerosis, systemic lupus erythematosus, and systemic sclerosis.[35] While their anti-inflammatory/immunomodulatory effects appear to greatly ameliorate autoimmune disease severity, the durability of these effects remain to be seen.

Scientists have reported that MSCs when transfused immediately within few hours post thawing may show reduced function or show decreased efficacy in treating diseases as compared to those MSCs which are in log phase of cell growth, so cryopreserved MSCs should be brought back into log phase of cell growth in in vitro culture before these are administered for clinical trials or experimental therapies, re-culturing of MSCs will help in recovering from the shock the cells get during freezing and thawing. Various clinical trials on MSCs have failed which used cryopreserved product immediately post thaw as compared to those clinical trials which used fresh MSCs.[36]

In 1924, Russian-born morphologist Alexander A. Maximow used extensive histological findings to identify a singular type of precursor cell within mesenchyme that develops into different types of blood cells.[37]

Scientists Ernest A. McCulloch and James E. Till first revealed the clonal nature of marrow cells in the 1960s.[38][39] An ex vivo assay for examining the clonogenic potential of multipotent marrow cells was later reported in the 1970s by Friedenstein and colleagues.[40][41] In this assay system, stromal cells were referred to as colony-forming unit-fibroblasts (CFU-f).

The first clinical trials of MSCs were completed in 1995 when a group of 15 patients were injected with cultured MSCs to test the safety of the treatment. Since then, over 200 clinical trials have been started. However, most are still in the safety stage of testing.[7]

Subsequent experimentation revealed the plasticity of marrow cells and how their fate could be determined by environmental cues. Culturing marrow stromal cells in the presence of osteogenic stimuli such as ascorbic acid, inorganic phosphate, and dexamethasone could promote their differentiation into osteoblasts. In contrast, the addition of transforming growth factor-beta (TGF-b) could induce chondrogenic markers.[citation needed]

Statistical-based analysis of MSC therapy for osteo-diseases inferred that most studies are still under investigation. There are different follow-up times that indicate we are still far from reaching the final conclusion. [42]

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Mesenchymal stem cell - Wikipedia, the free encyclopedia

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The term stem cell by itself can be misleading. In fact, there are many different types of stem cells, each with very different potential to treat disease.

Stem Cell Pluripotent Embryonic Stem Cell Adult Stem Cell iPS Cell Cancer Stem Cell

By definition, all stem cells:

Pluripotent means many "potentials". In other words, these cells have the potential of taking on many fates in the body, including all of the more than 200 different cell types. Embryonic stem cells are pluripotent, as are induced pluripotent stem (iPS) cells that are reprogrammed from adult tissues. When scientists talk about pluripotent stem cells, they mostly mean either embryonic or iPS cells.

Embryonic stem cells come from pluripotent cells, which exist only at the earliest stages of embryonic development. In humans, these cells no longer exist after about five days of development.

When isolated from the embryo and grown in a lab dish, pluripotent cells can continue dividing indefinitely. These cells are known as embryonic stem cells.

James Thomson, a professor in the Department of Cell and Regenerative Biology at the University of Wisconsin, derived the first human embryonic stem cell lines in 1998. He now shares a joint appointment at the University of California, Santa Barbara, a CIRM-funded institution.

Adult stem cells are found in the various tissues and organs of the human body. They are thought to exist in most tissues and organs where they are the source of new cells throughout the life of the organism, replacing cells lost to natural turnover or to damage or disease.

Adult stem cells are committed to becoming a cell from their tissue of origin, and cant form other cell types. They are therefore also called tissue-specific stem cells. They have the broad ability to become many of the cell types present in the organ they reside in. For example:

Unlike embryonic stem cells, researchers have not been able to grow adult stem cells indefinitely in the lab, but this is an area of active research.

Scientists have also found stem cells in the placenta and in the umbilical cord of newborn infants, and they can isolate stem cells from different fetal tissues. Although these cells come from an umbilical cord or a fetus, they more closely resemble adult stem cells than embryonic stem cells because they are tissue-specific. The cord blood cells that some people bank after the birth of a child are a form of adult blood-forming stem cells.

CIRM-grantee IrvWeissman of the Stanford University School of Medicine isolated the first blood-forming adult stem cell from bone marrow in 1988 in mice and later in humans.

Irv Weissman explains the difference between an adult stem cell and an embryonic stem cell (video)

An induced pluripotent stem cell, or iPS cell, is a cell taken from any tissue (usually skin or blood) from a child or adult and is genetically modified to behave like an embryonic stem cell. As the name implies, these cells are pluripotent, which means that they have the ability to form all adult cell types.

Shinya Yamanaka, an investigator with joint appointments at Kyoto University in Japan and the Gladstone Institutes in San Francisco, created the first iPS cells from mouse skin cells in 2006. In 2007, several groups of researchers including Yamanaka and James Thomson from the University of Wisconsin and University of California, Santa Barbara generated iPS cells from human skin cells.

Cancer stem cells are a subpopulation of cancer cells that, like stem cells, can self-renew. However, these cellsrather than growing into tissues and organspropagate the cancer, maturing into the many types of cells that are found in a tumor.

Cancer stem cells are a relatively new concept, but they have generated a lot of interest among cancer researchers because they could lead to more effective cancer therapies that can treat tumors resistant to common cancer treatments.

However, there is still debate on which types of cancer are propelled by cancer stem cells. For those that do, cancer stem cells are thought to be the source of all cells that make up the cancer.

Conventional cancer treatments, such as chemotherapy, may only destroy cells that form the bulk of the tumor, leaving the cancer stem cells intact. Once treatment is complete, cancer stem cells that still reside within the patient can give rise to a recurring tumor. Based on this hypothesis, researchers are trying to find therapies that destroy the cancer stem cells in the hopes that it truly eradicates a patients cancer.

John Dick from the University of Toronto first identified cancer stem cells in 1997. Michael Clarke, then at the University of Michigan, later found the first cancer stem cell in a solid tumor, in this case, breast cancer. Now at Stanford University School of Medicine, Clarke and his group have found cancer stem cells in colon cancer and head and neck cancers.

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Catriona Jamieson talks about therapies based on cancer stem cells (4:32)

Stanford Publication: The true seeds of cancer

UCSD Publication: From Bench to Bedside in One Year: Stem Cell Research Leads to Potential New Therapy for Rare Blood Disorder

Updated 2/16

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Stem Cell Key Terms | California's Stem Cell Agency

Stem cells are cells in the body that have the potential to turn into anything, such as a skin cell, a liver cell, a brain cell, or a blood cell. Stem cells that turn into blood cells are called hematopoietic (heh-mat-uh-poy-EH-tik)stem cells. These cells are capable of developing into the three types of blood cells:

Hematopoietic stem cells can be found in bone marrow (the spongy tissue inside bones), the bloodstream, and the umbilical cord blood of newborn babies.

A stem cell transplant (sometimes called a bone marrow transplant) can replenish a child's supply of healthy hematopoietic stem cells after they have been depleted. It's used to treat a wide range of diseases, including cancers like leukemia, lymphoma, neuroblastoma, Wilms tumor, and certain testicular or ovarian cancers; blood disorders; immune system diseases; and bone marrow syndromes.

Transplanted hematopoietic stem cells are put into the bloodstream through an intravenous (IV) line, much like a blood transfusion. Once in the body, they can produce healthy new blood and immune system cells.

The two main types of stem cell transplants are autologous (aw-TAHL-uh-gus)and allogeneic (al-uh-juh-NEE-ik). The type of transplant needed will depend on the child's specific medical condition and the availability of a matching donor.

This procedure may be done once or many times, depending on the need. Sometimes doctors will use extra-high doses of chemotherapy during treatment (to kill as many cancer cells as possible) if they know a patient will be getting a stem cell transplant soon after.

Unlike with an autologous transplant, there is a risk of a child's body rejecting the donated cells. This means that the body's ownimmune cells destroy the transplanted stem cells because they sense they are foreign.Sometimes, despite the donor being a good match, the transplant simply may not take. Other times, the donor cells can begin to make immune cells that attack the recipient's body. This condition is called graft-versus-host disease, and can be quite serious. Fortunately, most cases are successfully treated with steroids and other medicines.

Sometimes, an upside of graft-versus-host disease is that the newly transplanted cells recognize the body's cancer cells as different or foreign, and actually work to fight them.

Stem cell transplantation is a very complex process that may span several months. A team of doctors is usually involved in determining if a child is a candidate and, if so, whether the transplant will be autologous or allogeneic.

For an allogeneic transplant, a compatible donor will be sought among family members or through a national registry of volunteers. Once a match is found, the donor's stem cells will be harvested. Three different types of hematopoietic stem cells can be collected or harvested:

While all three types can replenish a patient's blood and bone marrow cells, there are advantages and disadvantages to each. The doctor will suggest the best type of stem cell for your child's illness.

The next step in the transplantation process is conditioning therapy, which kills unhealthy cells (like cancer cells) to make room for stem cells to grow and/or weakens the immune system so that theres less chance of the body rejecting the new cells.

One type of conditioning therapy delivers high doses of chemotherapy and/or radiation to kill cells, destroy the bone marrow, and weaken the immune system. Most kids will get this type of therapy. Another type of conditioning therapy delivers lower doses of chemotherapy, radiation, or another treatment to weaken the immune system. The doctor will decide which type of conditioning therapy is best.

Soon after the conditioning phase, the transplant itself will be done through intravenous (IV)infusion, and healthy stem cells will be introduced to the child's body. After the infusion, the child will be watched very closely to make sure the new stem cells are settling into the marrow and beginning to make new blood cells (called engrafting). Doctors will watch for any signs of rejection as well as graft-verses-host disease in kids with allogeneic transplants.

Engrafting takes an average of 2 weeks, but can be as quick as 1 week or as long as 6 weeks. Your child will receive medicines to promote engrafting and prevent rejection and graft-versus-host disease.

Kids who receive stem cell transplants have a high risk of infection. During conditioning therapy and while the transplant is engrafting, their immune systems are weakened and unable to fight bacteria and other germs that enter the body. Children who receive an allogeneic transplant have an even greater risk of infection because they require medicines to further suppress their immune systems to reduce the chance of rejection.

Because of these risks, a child who's had a stem cell transplant will not be released from the hospital until doctors are sure the transplant has successfully engrafted and the child is otherwise doing well.

Once released, a child needs very close monitoring and follow-up care. School and other public indoor areas may be off limits for 3 months to a year, and other places might be restricted as well. This is because for kids with a compromised immune system, even a simple infection like a common cold can be serious and even life-threatening if untreated.

The stress of having a child who is being treated for cancer or another serious conditioncan be overwhelming for a family. That stress can grow when treatment requires a long "isolation period," as is necessary with a stem cell transplant.

To find out what support is available to you and your child, talk to your doctor, a hospital social worker, or child life specialist. Many resources are available that can help you get through this difficult time.

Date reviewed: August 2015

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Stem Cell Transplants (For Parents) - KidsHealth