Derivation of spermatogenic cells from human embryonic stem cells (hESCs)

To determine the spermatogenic differentiation potential of non-manipulated hESCs, MUSIe002-A were subjected to a spermatogenic differentiation procedure. At the end of culture, the hESC-derived spermatogenic cells expressed several human spermatogenic marker proteins, including the pan-germ cell marker (VASA), spermatogonia stem cell markers (PLZF, CD90 and GPR125), and a spermatocyte marker (PIWIL1) (Fig.1A) compared to non-differentiate hESCs on culture day 0 (Fig. S1). In accordance with their protein expression, hESC-derived spermatogenic cells were subjected to transcriptional analysis by qRT-PCR. Up-regulation of SOX17 and TFAP2C was observed, which indicated the development toward germ cell lineages35. Furthermore, an early PCG marker BLIMP1, the post-migratory gonocyte PIWIL4 and the germline-specific gene VASA were up-regulated on day 5 and were maintained to the end of culture. Spermatogonia markers, including PLZF, GFRA, and NANOS3, were found to be highly expressed on day 5 and to remain throughout the culture period, while expressions of GRP125, ID4, and c-Kit were found to be upregulated on day 5 after induction (Fig.1B). Furthermore, the expression levels of the meiotic marker PIWIL2, and the post-meiotic haploid marker ACR, in differentiated hESCs also increased steadily and reached their highest levels at the end of culture (Fig.1B).

Generation of hESC-derived spermatogenic cells. (A) Representative immunofluorescence micrographs show the expression of germ cell markers VASA, the hSSC markers PLZF, CD90, GPR125, and the spermatocyte marker PIWIL1 in hSSLC. Non-differentiated hESCs on culture day 0 and IgG staining served as a control. Scale bar: 100m in the first and middle panel, 20m in the last panel. The micrographs shown in the last column are the zoomed-in of the area indicated in red box (B). The graphs show the expression levels of the early germ cell markers (STELLA, TFAP2C, BLIMP1, SOX17, PIWIL4 and VASA), hSSC markers (PLZF, GFRA, GPR125, ID4, NANOS3 and c-Kit), spermatocyte (PIWIL2) and spermatid marker (ACR) during spermatogenic differentiation of hESCs relative to those on culture day 0. Data are presented as meanSD of 3 experiments. (C) Expression of CD90+ cells (top row), PLZF+ cells in the sorted CD90+ cell population determined by flow cytometry (middle row) and immunofluorescent staining (bottom row). The picture in the right panel showed the zoomed-in area indicated in red. (D) Analysis of DNA content by flow cytometry shows the percentages of hESC-derived haploid spermatid-like cells at the end of spermatogenic differentiation. Non-differentiated hESCs on culture day 0 serves as a negative control, while mature human sperm serves as a positive control. (E) Representative immunofluorescence micrographs show the expression of the haploid germ cell marker Acrosin in human sperm cells (top row), Acrosin and TNP1 in hESC-derived spermatid-like cells. (F) Representative micrographs show a single copy of chromosome 13, 21, X and Y chromosome in hESC-derived spermatid-like cells determined by fluorescent in situ hybridization (FISH). Non-differentiated hESCs, which served as a control, had two copies of these chromosomes.

Next, we determined the percentage of hESC-derived hSSLCs after spermatogenic differentiation using the cell surface marker CD90, which has previously been used to identify SSLC in human and mouse models20,36. The results show that 66% of differentiated hESCs expressed CD90 (Fig.1C; top row). We also found that more than 90% of the CD90+hSSLCs also co-expressed PLZF, another widely used SSLC marker, as determined by flow-cytometry and immunofluorescent staining (Fig.1C; middle and bottom rows, respectively). These results confirm that hESC-derived hSSLCs are present in the culture after spermatogenic differentiation and could be identified with a combination of CD90 and PLZF markers.

To determine whether the differentiated hESCs generated any haploid spermatid-like cells, a DNA content analysis was performed. The results show that 6.70.72% of the differentiated hESCs are haploid cells, while no haploid cell was detected in non-differentiated hESCs (Figs.1D and S2). The haploid spermatid-like cells derived from hESCs expressed human spermatid markers, Acrosin and TNP1 (Figs.1E and S2). Furthermore, fluorescent in situ hybridization (FISH) analysis confirmed that our sorted haploid spermatid-like cell derived from hESCs has a single copy of chromosome 13, 21 and sex chromosome compared to two copies of the same chromosomes in non-differentiated hESCs (Fig.1F). These results demonstrated that the spermatogenic differentiation procedure used in this study successfully generated haploid spermatid-like cells from hESCs. The pattern of marker expression during in vitro spermatogenic differentiation closely matched that of human testicular germ cells that underwent in vivo spermatogenesis (Supplementary Fig. S3AG)34. Collectively, these results confirm that we could derive male germ cells from hESCs in vitro using the protocol previously described by Easley et al.21. Next, we determine whether Hippo/YAP is expressed in human testicular germ cells. Analysis of scRNA sequencing data showed YAP expression during spermatogenesis (Supplementary Fig. S3H,I). Like PLZF, the expression level of YAP was higher in undifferentiated human spermatogonia compared to the later stages of human spermatogenic differentiation (Supplementary Fig. S3H,I). This result suggested that YAP might play a crucial role during the early stages of human spermatogenesis and could be involved in the expression of the undifferentiated spermatogonia marker PLZF.

To investigate the role of YAP during human spermatogenesis, the isogenic YAP-depleted hESCs (YAP-KD cells) were established by transducing hESCs (MUSIe002-A cells) with the YAP-targeted CRISPR/Cas9 plasmid33. Western blot analysis showed a 50% reduction in the YAP level in YAP-KD cells compared to their wild-type control (WT cells) (Fig.2A). The YAP-KD cells retained a typical morphology of non-differentiated wild-type hESCs even after being expanded for more than 25 passages (Figs.2B and S4). After spermatogenic differentiation, the YAP-KD cells generated several clusters of cells, which increased in size and number toward the end of the culture (indicated by black arrows in Figs. 2C and S5). These cell clusters were also observed in differentiated WT cells, but their number was less than in the differentiated YAP-KD cells (Figs. 2C and S5).

Spermatogenic differentiation of YAP knockdown hESCs (YAP-KD cells). (A) Western blot shows the level of YAP relative to -ACTIN in YAP-KD cells compared to wild-type hESCs (WT). (B) Representative micrograph showing the morphology of a non-differentiated YAP-KD cell colony. Scale bar: 200m. (C) Representative micrographs show the morphology of differentiated YAP-KD cells during their spermatogenic differentiation. Wild-type hESCs cultured under the same conditions (WT) serve as controls. Scale bar: 200m in micrographs, 50m in zoomed areas. (D) Representative immunofluorescence micrographs show the expression of the SSC marker PLZF in YAP-KD cells during their spermatogenic differentiation compared to WT cells. Scale bar: 200m. (E) Representative dot plots show the percentages of PLZF+ hSSLCs in YAP-KD cells during their spermatogenic differentiation compared to those of WT cells. (F) Graphs show the number of PLZF+ hSSLCs derived from YAP-KD cells during their spermatogenic differentiation compared to those of WT cells. Data are presented as meanSD from 3 experiments. ****P<0.0001. (G) Western blot shows the levels of the YAP and PLZF proteins relative to -ACTIN in YAP-KD cells during their spermatogenic differentiation compared to those of WT cells. (H) Graphs show the expression level of YAP protein relative to -ACTIN in YAP-KD cells during their spermatogenic differentiation compared to that of WT cells. Data are presented as meanSD of 3 experiments. *P<0.05, **P<0.01, ****P<0.0001. (I) Graphs show the expression level of PLZF protein relative to -ACTIN in YAP-KD cells during their spermatogenic differentiation compared to that of WT cells. Data are presented as meanSD of 3 experiments. **P<0.01, ***P<0.001. (J) Correlation analysis between YAP protein levels and hESC-derived PLZF+ hSSC numbers during spermatogenic differentiation of WT and YAP-KD cells. The uncropped gels associated with Fig.2A are shown in Supplementary Fig. S8.

YAP protein is highly expressed in spermatogonia residing in testicular germ cells10,11. Similar to WT cells, differentiated YAP-KD cells highly expressed PLZF, a spermatogonia marker. Immunofluorescence shows that the level of PLZF expression in differentiated YAP-KD cells increased steadily throughout the entire culture period and was much higher than that of WT cells (Fig.2D). Consistent with the level of PLZF expression, the percentages of PLZF-expressing hSSLCs (PLZF+ cells) derived from the differentiated YAP-KD cells increased steadily throughout the differentiation period and were 23 times higher than those derived from WT cells throughout the culture period (Figs. 2E and S6). Consistent with these data, when the absolute number of PLZF+ cells was determined by BD Truecount, the YAP-KD cell also generated a higher number of PLZF+ cells than the WT cells, resulting in an approximately threefold increase in the number of PLZF+ cells at the end of the culture (8.61105cells vs 2.19105 cells on culture day 12) (Fig.2F). This result indicated that the depletion of YAP in hESCs enhances their spermatogenic differentiation toward PLZF+ hSSLCs.

Next, we quantified the expression levels of YAP and PLZF proteins at different time points during the spermatogenic differentiation of YAP-KD cells compared to WT cells. The expression level of YAP in WT cells was initially downregulated on culture day 5, bounced back on culture day 10, and continued to increase towards the end of the culture (Fig.2G,H). Interestingly, YAP-KD cells, whose YAP expression was 50% lower than that of WT cells at the beginning of culture, rapidly increased their YAP expression on culture day 5 (Fig.2G,H). Similar to WT cells, the expression level of YAP in YAP-KD cells continued to increase toward the end of the culture. At the end of the culture (day 12), the expression level of YAP in YAP-KD cells is similar to that of WT cells. However, unlike WT cells, down-regulation of YAP during the early phase of spermatogenic differentiation (day 5) was not observed in differentiated YAP-KD cells (Fig.2G,H).

The up-regulation of YAP in WT and YAP-KD cells coincided with the expression of PLZF, whose level also increased steadily towards the end of the culture (Fig.2G,I). However, the differentiated YAP-KD cells expressed PLZF earlier than WT cells (day 5 vs day 10), and their levels of PLZF were significantly higher than those of WT cells throughout the culture period (Fig.2G,I).

Furthermore, the level of YAP protein in both WT and YAP-KD cells was positively correlated with the expression level of PLZF+ hSSLCs generated during their spermatogenic differentiation (R2=0.8577, P=0.001) (Fig.2J). These results suggested that depletion of YAP at the beginning of spermatogenic differentiation leads to higher and earlier expression of PLZF, resulting in a higher number of hESC-derived hSSLCs.

As shown above, early depletion of YAP in hESCs increased the percentage and absolute number of hSSLCs during spermatogenic differentiation. Next, we determine whether hESC-derived hSSLCs express c-Kit, a differentiated spermatogonia marker. We performed a flow cytometric analysis for the expression of c-Kit in hSSLCs derived from WT and YAP-KD cells at different time points during their spermatogenic differentiation. The results showed that both the percentage and the absolute number of c-Kit+differentiated spermatogonial cells were up-regulated in the WT and YAP-KD groups as differentiation progressed towards the end of the culture. In which the percentages and absolute numbers of c-Kit+cells in the YAP-KD groups are significantly higher than those of WT on days 7 and 12 of culture (Fig.3A). This pattern of c-kit expression is similar to that of PLZF expression and therefore confirms that YAP depletion increases the spermatogenic differentiation of hESCs.

Derivation of haploid spermatid-like cells from YAP-KD cells. (A) Representative dot plots show the percentages of c-Kit+ cells in differentiated WT and YAP-KD cells during their spermatogenic differentiation (left). The graph shows the absolute number of c-Kit+ differentiated spermatogonia derived from YAP-KD cells during their spermatogenic differentiation compared to those of WT cells (right). Data are presented as meanSEM from 3 replicates. *P<0.05, **P<0.01 (B) Western blot shows the levels of VASA proteins relative to -ACTIN in YAP-KD cells during their spermatogenic differentiation compared to those of WT cells. (C) Graphs show the expression level of VASA protein relative to -ACTIN in YAP-KD cells during their spermatogenic differentiation compared to that of WT cells. Data are presented as meanSD of 3 experiments. *P<0.05, ****P<0.0001. (D) Western blot shows the levels of Acrosin proteins relative to -ACTIN in YAP-KD cells during their spermatogenic differentiation compared to those of WT cells. (E) Graphs show the expression level of Acrosin protein relative to -ACTIN in YAP-KD cells during their spermatogenic differentiation compared to that of WT cells. Data are presented as meanSD of 3 experiments. **P<0.01, ****P<0.0001. (F) Analysis of DNA content by flow cytometry shows the percentages of haploid spermatid-like cells derived from YAP-KD cells during their spermatogenic differentiation compared to those of WT cells. (G) The graphs show the percentages of haploid spermatid-like cells derived from YAP-KD cells during their spermatogenic differentiation compared to those of WT cells. Data are presented as meanSD of 3 experiments. *P<0.05. The uncropped gels associated with Fig.3A,C are shown in Supplementary Fig. S9.

We then investigated the effect of YAP depletion during the early stages of spermatogenic differentiation by measuring the expression level of VASA, an early germ cell marker. We found that YAP-KD cells expressed a significantly higher level of VASA protein than WT cells throughout the entire culture period (Fig.3B,C).

Next, we investigated the effect of YAP depletion during the later stages of spermatogenic differentiation by measuring the expression level of Acrosin, a specific marker of haploid spermatids, in differentiated YAP-KD cells. Similar to PLZF, differentiated YAP-KD cells up-regulated their acrosin expression earlier than WT cells (day 5 vs day 10) and their acrosin levels were significantly higher than those of WT cells on day 5 and day 10 of culture (Fig.3D,E). Consistent with the level of acrosin expression, the percentages of haploid spermatid cells in differentiated YAP-KD cells were significantly higher than those of their wild-type counterparts on day 5 (4.40.05% vs. 1.80.68%) and day 10 of culture (6.22.09% vs. 20.10%) (Fig.3F,G). These results suggest that YAP depletion at the beginning of spermatogenic differentiation could accelerate the progression of hESC-derived hSSLCs into haploid spermatid-like cells. However, we found that acrosin+spermatid-like cells could not be maintained in culture, since the level of acrosin in differentiated YAP-KD cells on day 12 of culture was lower than on day 10 (Fig.3D).

Due to the heterozygous nature of the mutated YAP gene in our YAP-KD cells, up-regulation of YAP expression was observed during the later stages of spermatogenic differentiation (days 512) (Fig.2G,H). This pattern of YAP expression in YAP-KD cells suggests that although YAP depletion at the beginning of spermatogenic differentiation appears to promote the germline specification of hESCs, up-regulation of YAP during the later stages of spermatogenic differentiation could be crucial for the expression of germ cell markers and the derivation of both hSSLCs and haploid spermatid-like cells.

To prevent the up-regulation of YAP in YAP-KD cells during the later stages of spermatogenic differentiation, a short hairpin RNA (shRNA) targeting the YAP transcript37 was transfected into YAP-KD cells to establish a YAP double knockdown hESCs (YAP-DKD cells). YAP-DKD cells showed a further reduction in YAP (Fig.4A; Supplementary Table S2) and YAP target genes, CTGF and CYR61, compared to YAP-KD cells (Fig.4B). Like YAP-KD cells, YAP-DKD cells exhibited a typical morphology of non-differentiated wild-type hESCs (Supplementary Fig. S4).

Spermatogenic differentiation of YAP double knockdown hESCs (YAP-DKD cells) (A). Western blot shows the level of YAP relative to -ACTIN in YAP-DKD cells compared to that of YAP-KD cells and wild-type hESCs (WT). ****P<0.0001. (B) Graphs show the expression levels of YAP target genes, CTGF and CYR61, in YAP-DKD cells compared to those of YAP-KD cells and WT. Data are presented as meanSD of 3 experiments. **P<0.01, ***P<0.001. (C) Western blot shows YAP, PLZF, VASA and acrosin protein levels relative to -ACTIN in WT, YAP-KD, and YAP-DKD cells during their spermatogenic differentiation. (D) Graphs show the level of PLZF protein relative to -ACTIN in YAP-DKD cells during their spermatogenic differentiation compared to YAP-KD cells and WT. (E) Graphs show the expression levels of spermatogenic cell markers (VASA, PLZF, GFRA1, GPR125 and ACR) during spermatogenic differentiation of YAP-DKD, compared to those of YAP-KD cells and WT. Data are presented as means SD of 3 experiments. *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001. (F) Representative micrographs show the morphology of YAP-DKD cells during their spermatogenic differentiation. White dots indicate areas of dead cells. Scale bar: 200m for the upper panel and 100m for the lower panel. (G) Western blot shows the level of the apoptotic marker cleaved Caspase3 relative to -ACTIN in WT and YAP-DKD cells during their spermatogenic differentiation. The uncropped membranes associated with this figure are provided in Supplementary Fig. S10.

When subjected to spermatogenic differentiation, different expression patterns of YAP and early germ cell markers of each cell type were observed. Unlike YAP-KD cells, the level of YAP expression in YAP-DKD cells was barely detected on day 10 and day 12 of culture (Fig.4C). This result confirmed that the shRNA successfully prevented the up-regulation of YAP in YAP-DKD cells during the later stages of their spermatogenic differentiation. Further analysis found that constitutive depletion of YAP in YAP-DKD inhibited the expression of the early germ cell marker VASA (Fig.4C), the SSC marker PLZF (Fig.4C,D), and the spermatid marker acrosin (Fig.4C). These results suggest that the up-regulation of YAP level during the later stages of spermatogenesis might be critical for the expression of germ cell markers and the derivation of hSSLCs and haploid spermatid-like cells from hESCs.

Consistent with the protein expression study, differentiated YAP-DKD cells expressed lower transcript levels of germ cell marker (VASA), hSSC marker genes (PLZF, GFRA1, and GPR125), and meiotic germ cell markers (ACR) compared to WT (Fig.4E). In contrast, YAP-KD cells, whose YAP was depleted only at the beginning of their spermatogenic differentiation, expressed higher levels of hSSC markers, PLZF, GPR125, and GFRA1 compared to WT cells (Fig.4E). Taken together, these results suggest that depleting YAP at the beginning of spermatogenic differentiation promotes germline specification of hESCs, and increasing the level of YAP during the later stages of spermatogenic differentiation is critical for the derivation and survival of hESC-derived hSSLCs and haploid spermatid-like cells.

Furthermore, we also found that the differentiated YAP-DKD cells contained several large areas of dead cells (dotted areas in Figs. 4F and S5). Consistent with this, western blots also showed a high level of cleaved-caspase 3 in YAP-DKD cells at the later stages of spermatogenic differentiation (D5-12) compared to D0 (Fig.4G; YAP-DKD), while the expression of cleaved-caspase 3 in WT did not show a sharp contrast compared to its control D0. These results confirm the higher level of apoptosis in differentiated YAP-DKD cells during the later stages of spermatogenesis.

To confirm that an initial depletion followed by an up-regulation of YAP promotes spermatogenic differentiation of hESCs, we modulated the expression level of YAP at specific time points during spermatogenic differentiation of wild-type hESCs using two small molecules. Dobutamine hydrochloride (DH), which has previously been shown to promote YAP phosphorylation and suppress YAP activity38,39, was used to inhibit YAP expression, while lysophosphatidic acid (LPA) activated YAP expression in wild-type hESCs. DH-treated hESCs showed a 50% reduction in YAP level compared to untreated hESCs (Supplementary Fig. S7B,C), and also expressed lower levels of YAP target genes, CTGF and CCND1, compared to untreated hESCs (Supplementary Fig. S7D). Our results demonstrate that DH treatment successfully inhibits the expression of YAP and its target genes in wild-type hESCs. Although LPA-treated hESCs increased CTGF, CCND1, and CYR61 expression levels compared to untreated hESCs (Supplementary Fig. S7D), their YAP protein level did not increase compared to the untreated cells (Supplementary Fig. S7B,C). These results suggest that although LPA did not increase YAP level in hESCs, it might enhance YAP activity resulting in higher levels of YAP target genes.

We then supplemented DH and LPA at various time points during spermatogenic differentiation of wild-type hESCs (Fig.5A). Consistent with the results generated from YAP-KD cells, the highest number of hESC-derived hSSLCs was observed under the condition that DH was added to inhibit YAP expression during the first 5days of spermatogenic differentiation before being removed for the rest of the culture period (Fig.5B, D12: DH-ET in Group 2). On the contrary, hESCs treated with LPA to increase YAP activity during the first 5days of their spermatogenic differentiation generated a lower number of hSSLCs than both D12: DH-ET and untreated cells (Fig.5B, D12: LPA-ET in Group 2).

Modulation of YAP expression during the various stages of spermatogenic cell differentiation of wild-type hESCs by using DH and LPA. (A) Diagrams show the timeline of DH and LPA supplementation during spermatogenic differentiation of wild-type hESCs. (B) Graphs show the total number of differentiated hESCs and the number of PLZF+ hSSCs derived from various conditions described in (A) at the end of the culture. Data are presented as meanSD of 3 experiments. **P<0.01, ***P<0.001, ****P<0.0001 and ### P<0.001 indicated the statistically significant of spermatogonia number from DH treated cells between each group. (C) Representative immunofluorescence micrographs show the level of PLZF expression in DH-ET and LPA-ET cells compared to that of their untreated counterpart. Scale bar: 200m. (D) Western blot shows the expression of YAP, PLZF, Acrosin, and -ACTIN proteins in DH-ET cells during their spermatogenic differentiation compared to those of DH-AT cells, untreated control cells, and non-differentiated hESCs. Red numbers indicate the expression level of each sample relative to -ACTIN. (E) Analysis of DNA content by flow cytometry shows the percentage of haploid spermatid-like cells derived from DH-ET cells at the end of their spermatogenic differentiation compared to that of LPA-ET and untreated cells. (F) The graph shows the percentage of haploid spermatid-like cells derived from DH-ET cells at the end of their spermatogenic differentiation compared to that of LPA-ET and untreated cells. Data are presented as meanSD of 3 experiments. **P<0.01, ***P<0.001. The uncropped membranes associated with Fig.5D are provided in Supplementary Fig. S11.

Unlike YAP suppression during the early stage of spermatogenic differentiation (days 05), suppressing YAP during the middle phase (days 510) of spermatogenic differentiation reduced the number of hESC-derived hSSLCs compared to untreated hESCs (Fig.5B, D12: DH-MT in Group 3). Moreover, suppressing YAP during the middle stage (DH-MT in Group 3) or throughout the spermatogenic differentiation (DH-AT in Group 1) also reduced the total number of differentiated hESCs in culture (Fig.5B), which could result in the decrease in the number of hESC-derived hSSLCs at the end of culture. This is consistent with our previous result with YAP-DKD cells showing that continued suppression of YAP during the later stages of spermatogenic differentiation increased the apoptotic level of differentiated hESCs (Fig.4F,G).

Furthermore, our immunofluorescence study shows that the level of PLZF expression in DH-ET cells on day 12 was higher than that of the untreated control, while the level of PLZF expression in LPA-ET cells was lower than that of the untreated control (Fig.5C). Western blot analysis further confirmed that DH-ET cells expressed higher levels of PLZF than the untreated control at the end of the culture (Fig.5D), while LPA-ET cells expressed lower levels of PLZF than the untreated control at the end of the culture (Supplementary Fig. S7E,F). Unlike suppression of YAP during the early stage of spermatogenic differentiation (DH-ET), suppression of YAP throughout spermatogenic differentiation (DH-AT) decreased the expression level of PLZF compared to the untreated control (Fig.5D). Although DH-ET treatment produces a higher number of haploid germ cells than the control, the level of Acrosin expression in the DH-ET group is lower than that of the control (Fig.5D). Because the level of Acrosin is associated with spermatid maturity, it is possible that while DH-ET treatment increases the number of hESC-derived spermatids, these spermatids may be less mature than those derived from the untreated control.

We further investigated haploid germ cell production using cell cycle analysis to observe DNA content of the cells. The DH-ET cells also generate significantly higher percentages of haploid spermatid-like cells at the end of culture than their untreated counterpart (14.162.61% vs 8.302.86%) (Fig.5E,F). These results support our hypothesis that inhibition of YAP activity during the early stage of spermatogenic differentiation facilitates the derivation of hSSLCs and spermatid-like cells from hESCs. In contrast, inhibition of YAP activity during the later stages of spermatogenic differentiation reduced the number of hESC-derived hSSLCs and spermatid-like cells, possibly by increasing the apoptotic rate of hSSLCs and preventing their maturation into haploid spermatid-like cells.

View post:
The dynamic expression of YAP is essential for the development of male germ cells derived from human embryonic stem cells - Nature.com

Kolios, G. & Moodley, Y. Introduction to stem cells and regenerative medicine. Respiration. 85(1), 310 (2013).

Article PubMed Google Scholar

Wisniewski, D., Affer, M., Willshire, J. & Clarkson, B. Further phenotypic characterization of the primitive lineage- CD34+CD38-CD90+CD45RA- hematopoietic stem cell/progenitor cell sub-population isolated from cord blood, mobilized peripheral blood and patients with chronic myelogenous leukemia. Blood Cancer J. 1(9), e36 (2011).

Article CAS PubMed PubMed Central Google Scholar

Cheng, H., Zheng, Z. & Cheng, T. New paradigms on hematopoietic stem cell differentiation. Protein Cell. 11(1), 3444 (2020).

Article PubMed Google Scholar

Walasek, M. A., van Os, R. & de Haan, G. Hematopoietic stem cell expansion: Challenges and opportunities. Ann. N. Y. Acad. Sci. 1266, 138150 (2012).

Article ADS CAS PubMed Google Scholar

Yu, B. et al. Co-expression of Runx1, Hoxa9, Hlf, and Hoxa7 confers multi-lineage potential on hematopoietic progenitors derived from pluripotent stem cells. Front Cell Dev. Biol. 10, 859769 (2022).

Article PubMed PubMed Central Google Scholar

Lee, Y. et al. FGF signalling specifies haematopoietic stem cells through its regulation of somitic Notch signalling. Nat. Commun. 5, 5583 (2014).

Article ADS CAS PubMed Google Scholar

Hofmeister, C. C., Zhang, J., Knight, K. L., Le, P. & Stiff, P. J. Ex vivo expansion of umbilical cord blood stem cells for transplantation: Growing knowledge from the hematopoietic niche. Bone Marrow Transplant. 39(1), 1123 (2007).

Article CAS PubMed Google Scholar

Reya, T. et al. A role for Wnt signalling in self-renewal of haematopoietic stem cells. Nature. 423(6938), 409414 (2003).

Article ADS CAS PubMed Google Scholar

Muzzey, D., Evans, E. A. & Lieber, C. Understanding the basics of NGS: From mechanism to variant calling. Curr. Genet. Med. Rep. 3(4), 158165 (2015).

Article PubMed PubMed Central Google Scholar

Wang, Y. & Blelloch, R. Cell cycle regulation by microRNAs in stem cells. Results Probl. Cell Differ. 53, 459472 (2011).

Article CAS PubMed Google Scholar

Lima, J. F., Cerqueira, L., Figueiredo, C., Oliveira, C. & Azevedo, N. F. Anti-miRNA oligonucleotides: A comprehensive guide for design. RNA Biol. 15(3), 338352 (2018).

Article PubMed PubMed Central Google Scholar

Ajami, M., Soleimani, M., Abroun, S. & Atashi, A. Comparison of cord blood CD34 + stem cell expansion in coculture with mesenchymal stem cells overexpressing SDF-1 and soluble /membrane isoforms of SCF. J Cell Biochem. 120(9), 1529715309 (2019).

Article CAS PubMed Google Scholar

Albayrak, E. & Kocaba, F. Therapeutic targeting and HSC proliferation by small molecules and biologicals. Adv. Protein Chem. Struct. Biol. 135, 425496 (2023).

Article PubMed Google Scholar

Nakamura-Ishizu, A. Thrombopoietin regulates mitochondria homeostasis for hematopoietic stem cell maintenance. Rinsho Ketsueki. 62(5), 521527. https://doi.org/10.11406/rinketsu.62.521 (2021) (Japanese).

Article PubMed Google Scholar

Gao, P. et al. Transcriptional regulatory network controlling the ontogeny of hematopoietic stem cells. Genes Dev. 34(1314), 950964 (2020).

Article CAS PubMed PubMed Central Google Scholar

Kimura, T. & Yamazaki, S. Development of low-cost ex vivo hematopoietic stem cell expansion. Rinsho Ketsueki. 63(10), 14221429 (2022).

PubMed Google Scholar

Gao, L., Decker, M., Chen, H. & Ding, L. Thrombopoietin from hepatocytes promotes hematopoietic stem cell regeneration after myeloablation. Elife. 31(10), e69894. https://doi.org/10.7554/eLife.69894.PMID:34463253;PMCID:PMC8457823 (2021).

Article Google Scholar

Li, J. et al. Development and clinical advancement of small molecules for ex vivo expansion of hematopoietic stem cell. Acta Pharm Sin B. 12(6), 28082831 (2022).

Article CAS PubMed Google Scholar

Nakamura-Ishizu, A. & Suda, T. Multifaceted roles of thrombopoietin in hematopoietic stem cell regulation. Ann. N. Y. Acad. Sci. 1466(1), 5158. https://doi.org/10.1111/nyas.14169 (2020).

Article ADS PubMed Google Scholar

Nakamura-Ishizu, A. et al. Prolonged maintenance of hematopoietic stem cells that escape from thrombopoietin deprivation. Blood. 137(19), 26092620 (2021).

Article CAS PubMed PubMed Central Google Scholar

Papa, L., Djedaini, M. & Hoffman, R. Ex vivo HSC expansion challenges the paradigm of unidirectional human hematopoiesis. Ann. N. Y. Acad. Sci. 1466(1), 3950 (2020).

Article ADS PubMed Google Scholar

Lynch, J. et al. Hematopoietic stem cell quiescence and DNA replication dynamics maintained by the resilient -catenin/Hoxa9/Prmt1 axis. Blood. 143(16), 15861598. https://doi.org/10.1182/blood.2023022082.PMID:38211335;PMCID:PMC11103100 (2024).

Article CAS PubMed PubMed Central Google Scholar

Sakamaki, T. et al. Hoxb5 defines the heterogeneity of self-renewal capacity in the hematopoietic stem cell compartment. Biochem. Biophys. Res. Commun. 539, 3441 (2021).

Article CAS PubMed Google Scholar

Schirripa, A., Sexl, V. & Kollmann, K. Cyclin-dependent kinase inhibitors in malignant hematopoiesis. Front Oncol. 12, 916682 (2022).

Article CAS PubMed PubMed Central Google Scholar

Singh, A. K., Althoff, M. J. & Cancelas, J. A. Signaling pathways regulating hematopoietic stem cell and progenitor aging. Curr. Stem Cell Rep. 4(2), 166181 (2018).

Article CAS PubMed PubMed Central Google Scholar

Wang, Y. & Sugimura, R. Ex vivo expansion of hematopoietic stem cells. Exp. Cell Res. 427(1), 113599 (2023).

Article CAS PubMed Google Scholar

Chen, Z., Guo, Q., Song, G. & Hou, Y. Molecular regulation of hematopoietic stem cell quiescence. Cell Mol. Life Sci. 79(4), 218. https://doi.org/10.1007/s00018-022-04200-w.PMID:35357574;PMCID:PMC11072845 (2022).

Article CAS PubMed PubMed Central Google Scholar

Zhang, L., Mack, R., Breslin, P. & Zhang, J. Molecular and cellular mechanisms of aging in hematopoietic stem cells and their niches. J. Hematol. Oncol. 13(1), 157 (2020).

Article PubMed PubMed Central Google Scholar

Pinto, J. P. et al. StemChecker: A web-based tool to discover and explore stemness signatures in gene sets. Nucleic Acids Res. 43(W1), W72W77 (2015).

Article CAS PubMed PubMed Central Google Scholar

Kanehisa, M., Furumichi, M., Sato, Y., Kawashima, M. & Ishiguro-Watanabe, M. KEGG for taxonomy-based analysis of pathways and genomes. Nucleic Acids Res. 51(D1), D587D592 (2023).

Article CAS PubMed Google Scholar

Shannon, P. et al. Cytoscape: A software environment for integrated models of biomolecular interaction networks. Genome Res. 13(11), 24982504 (2003).

Article CAS PubMed PubMed Central Google Scholar

Takagi, S. et al. Membrane-bound human SCF/KL promotes in vivo human hematopoietic engraftment and myeloid differentiation. Blood J. Am. Soc. Hematol. 119(12), 27682777 (2012).

CAS Google Scholar

Sigurjonsson, O. E., Gudmundsson, K. O., Haraldsdttir, V., Rafnar, T. & Gudmundsson, S. Flt3/Flk-2-ligand in synergy with thrombopoietin delays megakaryocyte development and increases the numbers of megakaryocyte progenitor cells in serum-free cultures initiated with CD34+ cells. J. Hematotherapy Stem Cell Res. 11(2), 389400 (2002).

Article CAS Google Scholar

Aizenman, Y. & de Vellis, J. Brain neurons develop in a serum and glial free environment: Effects of transferrin, insulin-insulin-like growth factor-I and thyroid hormone on neuronal survival, growth and differentiation. Brain Res. 406(12), 3242 (1987).

Article CAS PubMed Google Scholar

Yadav, P., Vats, R., Bano, A. & Bhardwaj, R. Hematopoietic stem cells culture, expansion and differentiation: an insight into variable and available media. Int. J. Stem Cells. 13(3), 326334 (2020).

Article PubMed PubMed Central Google Scholar

Tsiftsoglou, A. S. Erythropoietin (EPO) as a key regulator of erythropoiesis, bone remodeling and endothelial transdifferentiation of multipotent mesenchymal stem cells (MSCs): Implications in regenerative medicine. Cells. 10(8), 2140 (2021).

Article CAS PubMed PubMed Central Google Scholar

Nogueira-Pedro, A. et al. -Tocopherol induces hematopoietic stem/progenitor cell expansion and ERK1/2-mediated differentiation. J. Leukocyte Biol. 90(6), 11111117 (2011).

Article CAS PubMed Google Scholar

Hu, Q. et al. GPX4 and vitamin E cooperatively protect hematopoietic stem and progenitor cells from lipid peroxidation and ferroptosis. Cell Death Dis. 12(7), 706 (2021).

Article CAS PubMed PubMed Central Google Scholar

Ren, Y., Cui, Y. N. & Wang, H. W. Effects of different concentrations of nicotinamide on hematopoietic stem cells cultured in vitro. World J. Stem Cells. 16(2), 163175 (2024).

Article PubMed PubMed Central Google Scholar

Brewer, G. J., Torricelli, J. R., Evege, E. K. & Price, P. J. Optimized survival of hippocampal neurons in B27-supplemented neurobasal, a new serum-free medium combination. J. Neurosci. Res. 35(5), 567576 (1993).

Article CAS PubMed Google Scholar

Phuc, P. V. et al. Isolation of three important types of stem cells from the same samples of banked umbilical cord blood. Cell Tissue Bank. 13(2), 341351 (2012).

Article CAS PubMed Google Scholar

Grassinger, J. et al. Differentiation of hematopoietic progenitor cells towards the myeloid and B-lymphoid lineage by hepatocyte growth factor (HGF) and thrombopoietin (TPO) together with early acting cytokines. Eur. J. Haematol. 77(2), 134144 (2006).

Article CAS PubMed Google Scholar

Rayner, K. J. & Moore, K. J. The plaque micro environment: microRNAs control the risk and the development of atherosclerosis. Curr. Atheroscler. Rep. 14(5), 413421 (2012).

Article CAS PubMed PubMed Central Google Scholar

Alexander, T., Greco, R. & Snowden, J. A. Hematopoietic stem cell transplantation for autoimmune disease. Annu. Rev. Med. 72, 215228 (2021).

Article CAS PubMed Google Scholar

Zhu, X., Tang, B. & Sun, Z. Umbilical cord blood transplantation: Still growing and improving. Stem Cells Transl. Med. 10(Suppl 2), S62-s74 (2021).

Article PubMed PubMed Central Google Scholar

Gudauskait, G., Kairien, I., Ivakien, T., Rascon, J. & Mobasheri, A. Therapeutic perspectives for the clinical application of umbilical cord hematopoietic and mesenchymal stem cells: Overcoming complications arising after allogeneic hematopoietic stem cell transplantation. Adv. Exp. Med. Biol. 1409, 111126 (2023).

Article PubMed Google Scholar

Wilkinson, A. C., Igarashi, K. J. & Nakauchi, H. Haematopoietic stem cell self-renewal in vivo and ex vivo. Nat. Rev. Genet. 21(9), 541554 (2020).

Article CAS PubMed PubMed Central Google Scholar

Amiri, F., Kiani, A. A., Bahadori, M. & Roudkenar, M. H. Co-culture of mesenchymal stem cell spheres with hematopoietic stem cells under hypoxia: A cost-effective method to maintain self-renewal and homing marker expression. Mol. Biol. Rep. 49(2), 931941 (2022).

Article CAS PubMed Google Scholar

Shirdare, M., Amiri, F., Samiee, M. P. & Safari, A. Influential factors for optimizing and strengthening mesenchymal stem cells and hematopoietic stem cells co-culture. Mol. Biol. Rep. 51(1), 189 (2024).

Article CAS PubMed Google Scholar

Elahimanesh, M., Shokri, N., Mohammadi, P., Parvaz, N. & Najafi, M. Step by step analysis on gene datasets of growth phases in hematopoietic stem cells. Biochem. Biophys. Rep. 1(39), 101737 (2024).

Google Scholar

Read the original:
Exploring the potential of predicted miRNAs on the genes involved in the expansion of hematopoietic stem cells - Nature.com

BORDEAUX, France, July 9, 2024 /PRNewswire/ -- TreeFrog Therapeutics, the French biotech company advancing a pipeline of regenerative medicine cell therapies based on a disruptive proprietary technology platform will present at several events at the global stem cell event in Hamburg, including a poster session with data from their lead program in Parkinson's Disease and an innovation showcase with a deep dive on their transformational technology, C-Stem.

Regenerative medicine holds immense potential in addressing some of the largest unmet needs in diseases of the major organs such as the central nervous system, the liver, pancreas and heart. However, despite major advances in the last 70 years, there are still bottlenecks holding up innovation, particularly, the ability to produce the required amount of high-quality cells, efficiently.

TreeFrog's lead program in Parkinson's disease has proven efficacy in pre-clinical models using a unique approach of grafting 3D format microtissues containing dopaminergic progenitors and mature dopaminergic neurons, as opposed to single-cell suspensions. The poster, presented by Maxime Feyeux, Chief Scientific Officer and Co-founder of TreeFrog will highlight the therapeutic potential of the 3D approach, and delve deeper into the characterization of the product through complementary methods including qPCR, RNAseq, flow cytometry and microscopy.

The pipeline of TreeFrog is based on their C-Stemtechnology, the culmination of over 20 years of research bringing the best in biophysics and stem cell biology together. This breakthrough technology addresses the challenges of scale and quality, and the closed system enables both the amplification and differentiation of cells. Maxime will be joined by two senior scientists Joffrey Mianne, Head of iPSC Research and Clement Rieu, Head of Technologies R&D - during an innovation showcase about C-Stem.

Presentations details:Poster Presentation: #190 Clinical Applications (CA) SessionWednesday, July 10, 2024 @ 6:45 PM 7:45 PM Room: Poster & Exhibit hall

Innovation Showcase Thursday July 11th, 2024 @ 6:00PM 6:30PM Room: Hall 3, entrance level

AboutTreeFrog TherapeuticsTreeFrog Therapeutics is a biotech company set to unlock access to cell therapies for millions of patients bringing together biophysicists, cell biologists and bioproduction engineers to address the challenges of producing and differentiating cells of quality at unprecedented scale, cost-effectively. To realize their mission of Cell Therapy for all, TreeFrog has their own therapeutic programs and partnerships with leading biotech and industry players in other areas.

http://www.treefrog.fr

Contact: Rachel Mooney Chief Communications Officer TreeFrog Therapeutics [emailprotected]

Logo - https://mma.prnewswire.com/media/2272329/treefrog_Logo.jpg

SOURCE TreeFrog Therapeutics

Excerpt from:
TREEFROG THERAPEUTICS PARTICIPATES IN AN INNOVATION SHOWCASE & POSTER SESSION AT THE INTERNATIONAL SOCIETY FOR STEM CELL RESEARCH (ISSCR) ANNUAL...

S.BIOMEDICS Dopamine Cell Therapy for Parkinsons Disease with TED-A9 Shows Promising Results at 12 months in Phase I/IIa Clinical Trial

S.BIOMEDICS(KOSDAQ: 304360) announced positive one-year assessment data from the first 3 participants (first low dose cohort) of Phase I/IIa clinical trial for Parkinsons disease with TED-A9, hESC (human embryonic stem cell)-derived midbrain dopaminergic progenitors. The data demonstrates that 3 participants showed safety and efficacy of TED-A9 in a one-year follow-up.

This press release features multimedia. View the full release here: https://www.businesswire.com/news/home/20240709317387/en/

Dong-Wook Kim, Professor at Yonsei University College of Medicine and CTO of S.BIOMEDICS (Photo: S.BIOMEDICS)

The clinical trial was conducted at Severance Hospital of Yonsei University in South Korea led by Prof. Jin-Woo Chang, a neurosurgeon and Prof. Phil Hyu Lee, a neurologist.

For the 3 participants who received initial low dose (3.15 million cells), MRI and CT scans after one year revealed no adverse effects related to the cell transplantation or surgery.

Moreover, the MDS-UPDRS Part III (off) evaluation, which objectively measures motor functions, showed a mean decrease of 12.7 points, from baseline 61.7 to 49.0 one-year post treatment. The MDS-UPDRS Part III (off) evaluation indicated significant improvement in their motor abilities. Improvement was also observed in symptoms such as wearing off and freezing of gait.

DAT brain imaging (FP-CIT-PET), conducted one-year post transplantation, revealed an increase in dopamine transporters (DAT), suggesting the potential engraftment of dopamine neurons, which correlated with improvements in the patients' Parkinson's symptoms.

Although this clinical evaluation only targets the first 3 low-dose patients, but not all 12 subjects, no adverse issues related to transplant surgery or cell safety were observed in one-year post transplantation. Importantly, the clinical results demonstrated very promising efficacy, said Prof. Dong-Wook Kim at Yonsei University College of Medicine and CTO of S.BIOMEDICS. The results are believed to align closely with the findings from our preclinical in vitro and in vivo studies. We are excited that TED-A9 could be a fundamental treatment that directly replaces dopaminergic neurons lost in patients with Parkinson's disease.

Assessment data of further participants (first high dose cohort) will be announced in September or October of 2024 after a one-year follow-up.

About TED-A9 and Phase I/IIa clinical trial

TED-A9 is an investigational cell therapy designed to replace ventral midbrain-specific dopaminergic cells lost in patients with Parkinson's disease. These ventral midbrain-specific dopaminergic cells are derived from hESCs by exclusively utilizing small molecules only. TED-A9 represents a significant advancement in the field, offering highly purified dopamine cells derived from hESCs. Through surgical procedure, these hESC-derived dopaminergic progenitor (precursor) cells (TED-A9) are transplanted to three segments of the putamen; the anterior, middle, and posterior sections, with three tracks per each putamen. Bilateral putamina received cell transplantation in a single surgical procedure, with cells injected at three points within each track. After transplantation, the expectation is that the transplanted dopaminergic progenitor cells will mature into dopaminergic neurons which will supply the dopamine that Parkinsons patients are lacking, restoring the motor function of patients.

The Phase I/IIa clinical trial is conducted on 12 participants who have been diagnosed with Parkinson's disease for more than 5 years and exhibited motor complications such as wearing-off, freezing of gait or dyskinesia. The participants age was between 50 and 75 years old. TED-A9 were administered to 6 participants in the low-dose group (3.15 million cells) and to another 6 participants in the high-dose group (6.30 million cells).

An initial cohort of three patients was enrolled at a low dose to assess initial safety, including dose-limiting toxicity (DLT) evaluation, over a period of up to 3 months post-transplantation. There were no safety concerns within this timeframe. Thus, an additional 3 patients were enrolled at a high dose for evaluation over another 3-month period post-transplantation. As no safety issues arose during this extended period, the clinical trial continued by adding three further patients to each of the low-dose and high-dose groups, totaling 12 patients. The final participant was administered with TED-A9 on February 2024.

The primary objective of the Phase I/IIa trial is to assess the safety and exploratory efficacy of TED-A9 transplantation over two years post-transplant. Safety will be monitored for additional 3 years, making it a total 5 years.

About S.BIOMEDICS

Established in 2005, S.BIOMEDICS is at the forefront of stem cell therapy, focusing on regenerative medicine powered by data-driven biology. Based on two core platform technologies, S.BIOMEDICS currently develops seven cell therapy programs, targeting non-curable diseases. Its leading programs are under clinical stage accelerating the journey of cell medicine as shown below:

For more information about S.BIOMEDICS, visit http://www.sbiomedics.com. S.BIOMEDICS is listed on the Korea Exchange (KOSDAQ: 304360) and is also the founder and controller of S.THEPHARM (www.sthepharm.com), a corporation specializing in anti-aging products such as HA-Filler. More Information about the Phase 1/2a clinical trial for Parkinsons disease is available at ClinicalTrials.gov (NCT05887466).

S.BIOMEDICS Jong-Wan Kim jwkim@sbiomedics.com

View source version on businesswire.com: https://www.businesswire.com/news/home/20240709317387/en/

See the original post:
S.BIOMEDICS Dopamine Cell Therapy for Parkinsons Disease with TED-A9 Shows Promising Results at 12 months in Phase I/IIa Clinical Trial - Morningstar

CAMBRIDGE, Mass., July 9, 2024 /PRNewswire/ --Factor Bioscience Inc., a Cambridge-based biotechnology company focused on developing mRNA and cell-engineering technologies,announced its participation in the International Society for Stem Cell Research (ISSCR) 2024 Annual Meeting to be held in Hamburg, Germany from July 10-13, 2024. Factor will deliver six presentations covering the latest preclinical data from Factor's cell engineering platforms.

"We are excited to showcase our recent progress on developing next-generation therapies based on cutting-edge stem cell science at ISSCR 2024," said Dr. Matt Angel, Co-Founder, Chairman and CEO of Factor. "The work that we will be presenting this week represents more than a decade of focused effort. We are committed to developing these new medicines to enable a brighter future for patients and their families."

The work that we will be presenting this week represents more than a decade of focused effort.

Dr. Kyle Garland, Factor's Director of Translational Science, added, "Our six presentations at ISSCR 2024 will cover several novel and unique stem cell technologies, including iPSC-derived macrophages engineered with mRNA to enhance T cell cytotoxicity to solid tumor cells. We are excited to share these and other advances in Hamburg over the next few days."

Details of the presentations are below:

For more information about the International Society for Stem Cell Research (ISSCR) 2024 Annual Meeting, visit http://www.isscr2024.org.

About Factor BioscienceFounded in 2011, Factor Bioscience engineers cells to promote health and improve lives. Factor Bioscience is privately held and headquartered in Cambridge, MA. For more information, visit http://www.factorbio.com.

SOURCE Factor Bioscience

Original post:
Factor Bioscience to Deliver Six Presentations at the International Society for Stem Cell Research (ISSCR) 2024 Annual Meeting - PR Newswire

UC San Diego scientists have found the most common form of liver cancer could be targeted and treated more effectively using a stem cell-derived therapy, according to a report published Tuesday.

The report, published in the scientific journal Cell Stem Cell, focuses on hepatocellular carcinoma, a cancer with a high mortality rate.

While not yet studied in patients, the treatment which involves the lab engineering of natural killer white blood cells to battle HCC could be mass-produced and ready for deployment rapidly, the researchers said.

Unlike a treatment called chimeric antigen receptor-expressing T-cell therapy, which requires patient personalization, the NK-cell therapy could be more acceptable to more bodies.

To some extent all tumor cells perhaps hepatocellular carcinoma more so inhibit immune cells that try to kill them, said UCSD School of Medicine Professor Dr. Dan Kaufman, lead author on the study and director of the Sanford Advanced Therapy Center at the universitys Sanford Stem Cell Institute and Moores Cancer Center member.

This is one key reason why some immunotherapies like CAR T cells have been less successful for solid tumors than for blood cancers the immunosuppressive tumor microenvironment.

Kaufman and his team produced the NK cells in which a protein that impairs immune function was disabled. Hepatocellular carcinoma tumors and the liver in general contain large amounts of the substance, which both inhibits the immune cell activity and allows cancer to proliferate, the authors write.

Typical NK cells without the disabled receptor, like CAR T cells, were not very effective in battling the cancer.

These are pretty resistant tumors when we put them in mice, they grow and kill the mice, Kaufman said.

The five-year survival rate for HCC in humans is less than 20% and is responsible for more than 12,000 deaths in the United States annually.

However, when researchers tested the modified NK cells against the cancer, we got very good anti-tumor activity and significantly prolonged survival.

These studies demonstrate that it is crucial to block transforming growth factor beta at least for NK cells, but I also think its true for CAR T cells, Kaufman said. If you unleash NK cells by blocking this inhibitory pathway, they should kill cancer quite nicely.

He said his teams discovery will likely be reflected in clinical trials for many groups working with the various therapies or solid tumors.

City News Service, Inc.

Read the original here:
UCSD Team Tests Stem Cell-Derived Therapy On Liver Cancer; Results Promising - Times of San Diego

Our teeth cant repair themselvesbut what if they could? The future of dentistry lies in the captivating field of regenerative medicine, where stem cell research is diving deep into the potential to repair damaged teeth with living fillings. But how far are we from ditching fillings for specialized tooth restoration? While the research is science fact, getting a living filling from your dentist is still science fictionfor now.

I find this field really fascinating, says New York cosmetic dentist Victoria Veytsman, DDS. The field of tissue engineering and regenerative medicine in dentistry is really at the forefront of where healthcare is going.

Stem cells are those super useful specialized cells (found in adult body tissues and in embryos) that can be guided towards becoming many different cell types and can self-replicate. That makes them immensely useful in regenerative medicine, where the goal is to get the bodys repair processes engaged to handle damaged, diseased or otherwise unwell tissues. According to the California Institute for Regenerative Medicine, the most commonly used stem cell-based therapy is for bone marrow transplants.

When it comes to filling a cavity with them, stem cells alone arent enough to complete the process of tooth restoration, explains Dallas, TX cosmetic dentist Salvator La Mastra, DDS. They would need a framework of some kind in order to form in the correct manner.

Dr. Veytsman explains that current research is focused on creating that framework, creating a kind of living filling.

We dont want enamel to grow in a petri dish; we want it to grow on your tooth, Dr. Veytsman says. So the process requires a scaffold or matrix to support that growth.

When a tooth develops a cavity, the first step is to remove the decay and stop the process of damage. Cavities are caused by bacteria, Dr. La Mastra explains. That acid producing bacteria is what causes the cavitation of the tooth, which is the cavity itself and the decay. Its basically necrotic tissue that we have to drill out.

Then, you have to fill in whats lost. We do things like crowns and fillings to replace the chief structure that was lost or decayed, Dr. Veytsman explains. Its called restorative dentistry because were trying to restore whats been lost.

Those fillings are made of amalgam (a mixture of metals) or composite resin filling materials (made from polymers and glass particles), and we know theyre safe, functional and that they wont decay in the future. Thats something we cant say about these living fillings.

One thing about our current implants and fillings is that we know they wont develop cavities down the line, La Mastra says. There are complications that could arise from the regenerative method that could cause more than just aesthetic consequences; your bite can also be impacted.

I think were just at the beginning of this technology, Dr. Veytsman says. But it definitely has the potential to change the way we approach cavities in the years to come.

Stem cells could also be utilized outside of living fillings to benefit oral health. Aside from repairing enamel, stem cells could be used to encourage the growth of dentin, restore pulp, even regenerate lost gum tissues.

Youre seeing the rise of stem cell banking now for these purposes, Dr. Veytsman explains. Harvesting and banking stem cells for future applications and to use as a preventative measure are growing in popularity.

I think were multiple decades away from a changeover to regenerative medicine in dentistry, La Mastra says. I already have patients who ask me if they can just regrow their tooth, and we are nowhere near being able to do that.

While living fillings arent going to enter your dentists office in the immediate future, theres still reason to be excited.

The advent of AI technologies is really accelerating this research, Dr. Veytsman says. And its letting us ask a ton of questions about possible applications. Can regenerative medicine deal with prevention? Can it help stop decay in the very early stages? Were still so early in this process, but AI and regenerative medicine are really at the forefront of healthcare right now.

View post:
"Living Fillings" Could be the Future Thanks to Stem Cells - NewBeauty Magazine