Background Diabetic cutaneous ulcers (DCU) are a complication of diabetes with diabetic foot ulcers being the most common, and the wounds are difficult to heal, increasing the risk of bacterial infection. Cell-based therapy utilizing mesenchymal stem cells (MSCs) is currently being investigated as a therapeutic avenue for both chronic diabetic ulcers and severe burns. Wharton’s jelly mesenchymal stem cell (WJMSC) with PF-127 hydrogel and sodium ascorbyl phosphate (SAP) improved skin wound healing in mice. Whether this combination strategy is helpful to diabetic ulcers wound healing remains to be explored. Methods Firstly, the WJMSCs embedded in PF-127 and SAP combination were transplanted onto excisional cutaneous wound bed in type 2 diabetic Sprague Dawley (SD) rats. Two weeks after transplantation, the skin tissue was collected for histological and immunohistochemical analysis. Further, overexpressing-EGFP WJMSCs were performed to investigate cell engraftment in the diabetic cutaneous ulcer. The apoptosis of WJMSCs which encapsulated with combination of PF-127 and SAP was detected by TUNEL fluorescence assay and RT-PCR in vitro. And the mitochondrial damage induced by oxidative stress assessed by MitoTracker and CMH2DCFDA fluorescence assay. Results In diabetic cutaneous wound rat model, PF-127 plus SAP-encapsulated WJMSCs transplantation promoted diabetic wound healing, indicating improving dermis regeneration and collagen deposition. In diabetic wound healing, less pro-inflammatory M1 macrophages, more anti-inflammatory M2 tissue-healing macrophages, and neovascularization were observed in PF-127 + SAP + WJMSCs group compared with other groups. SAP supplementation alleviated the apoptosis ratio of WJMSCs embedded in the PF-127 in vitro and promoted cell survival in vivo. Conclusion PF-127 plus SAP combination facilitates WJMSCs-mediated diabetic wound healing in rat through promoting cell survival, the macrophage transformation, and angiogenesis. Our findings may potentially provide a helpful therapeutic strategy for patients with diabetic cutaneous ulcer.

Background: Factors such as poor engraftment, retention, and survival of the transplanted stem cells are deemed to limit their therapeutic efficacy for wound regeneration. Hence, it is necessary to explore these issues in order to resolve them. In this study, we aim to investigate the role of Pluronic F-127 (PF-127) hydrogel plus antioxidant sodium ascorbyl phosphate (SAP) in enhancing Wharton's jelly mesenchymal stem cell (WJMSC)-mediated effectiveness on full-thickness skin wound healing in mice. Methods: First, the cytotoxicity of PF-127 and the biological effect of SAP on the survival of WJMSCs were tested in vitro using cell viability and proliferation assays. Next, a cell suspension containing WJMSCs, PF-127, and SAP was topically administered onto an 8-mm diameter excisional full-thickness wound bed. Eight days after transplantation, the mice were sacrificed and the skin tissue was excised for histological and immunohistochemical analysis. Finally, in vivo distribution of transplanted WJMSCs was traced to investigate cell engraftment and the potential therapeutic mechanism. Results: PF-127 was found to be cytotoxic to WJMSCs while SAP significantly improved the survival of PF-127embedded WJMSCs. When this combination was topically transplanted onto the wound bed, wound healing was facilitated and dermis regeneration was achieved on the 8th day after surgery, as evidenced by an increase in dermal thickness, newly developed hair follicles, and collagen fiber deposition accompanied by a reduction in scar width. Further, immunohistochemical analysis demonstrated a higher number of anti-inflammatory M2 macrophages, proliferating cells, and newly formed blood vessels in the WJMSCs/PF-127/SAP group relative to all other groups. In addition, in vivo tracking results revealed a highly enhanced engraftment of WJMSCs accumulated in the dermis in the WJMSCs/PF-127/SAP group. (Continued on next page)Conclusions: SAP significantly improves the survival of WJMSCs in PF-127 encapsulation. Further, PF-127 plus SAP is an effective combination that enhances WJMSC engraftment in the dermis, which then promotes full-thickness wound healing through potential M2 macrophage formation and angiogenesis.

Human embryonic stem cells (hESCs) have long been considered as a promising source for cell replacement therapy. However, one major obstacle for the use of these cells is immune compatibility. Histocompatible human parthenogenetic ESCs have been reported as a new method for generating human leukocyte antigen (HLA)-matched hESCs. To further investigate the possibility of obtaining histocompatible stem cells from uniparental embryos, we tried to produce androgenetic haploid human embryos by injecting a single spermatozoon into enucleated human oocyte, and establish human androgenetic embryonic stem (hAGES) cell lines from androgenetic embryos. In the present study, a diploid hAGES cell line has been established, which exhibits typical features of human ESCs, including the expression of pluripotency markers, having differentiation potential in vitro and in vivo, and stable propagation in an undifferentiated state (>P40). Bisulfite sequencing of the H19, Snrpn, Meg3, and Kv imprinting control regions suggested that hAGES cells maintained to a certain extent a sperm methylation pattern. Genome-wide single nucleotide polymorphism, short tandem repeat, and HLA analyses revealed that the hAGES cell genome was highly homozygous. These results suggest that hAGES cells from spermatozoon could serve as a useful tool for studying the mechanisms underlying genomic imprinting in humans. It might also be used as a potential resource for cell replacement therapy as parthenogenetic stem cells.

Y-27632 is a specific inhibitor of Rho-associated protein kinases (ROCKs), which are downstream effectors of Rho GTPase. The present study aimed to determine the effect of the specific ROCK inhibitor, Y-27632, on fresh human embryos and on single blastomeres obtained from discarded human embryos. A total of 784 poor-quality embryos were included, of which 526 were allocated to blastocyst culture directly and the remaining 258 were allocated to blastomere isolation. Embryos and single blastomeres were cultured either with, or without, Y-27632. Embryonic development was observed and recorded daily from day 5 onwards. Y-27632 did not affect the ratio of blastocyst formation or the quality of the human embryos. The duration of blastocyst formation was compared between the two groups in the embryo culture. On day 5, the blastocyst formation ratio in the experimental group was 11.4% (26/228), which was significantly (P=0.015) lower than the corresponding rate (19.7%; 44/223) in the control group. Survival analysis of the blastocyst formation duration showed that the median formation duration in the experimental group was significantly higher than that of the control group. The present study also obtained 1,192 blastomeres from 258 discarded day 3 embryos, and sibling blastomeres of similar sizes were equally allocated to experimental and control groups (n=596 in each). Treatment with Y-27632 increased the blastocyst formation ratio of human individual blastomeres, with 82 blastocysts of 596 blastomeres (13.8%), and 51 blastocysts of 596 blastomeres (8.6%) formed in the presence and absence of Y-27632, respectively (P=0.004). Compared with the control group, the mRNA and protein expression levels of E-cadherin in the blastocysts from blastomeres were enhanced by Y-27632 (P=0.022). In conclusion, the present study demonstrated that Y-27632 has different effects on the cleavage-stage of embryos and single blastomeres. Y-27632 increases the ratio of formation of blastocysts from single human blastomeres, but inhibits the direct formation of blastocysts from discarded human embryos.