Peterson, Quinn

Stem cell-derived β (SC-β) cells are a promising therapy for type 1 diabetes (T1D), but their long-term efficacy is limited by immune-mediated graft rejection. Platelet-derived exosome product (PEP) has emerged as a novel immunomodulatory agent, although its effect on SC-β cell xenograft rejection remains unclear. Here, we evaluated PEP in stimulated human peripheral blood mononuclear cells (PBMCs) and in an immunocompetent mouse model of SC-β cell transplantation under the kidney capsule. Immune-related gene expression was assessed by quantitative PCR, immune cell infiltration by immunofluorescence, and graft function by circulating human insulin. In vitro, PEP reduced NK-cell-associated gene expression, including NK1.1, EOMES, and KLRK1, and increased IL-10 expression. In vivo, untreated SC-β cell xenografts showed progressive immune infiltration and loss of detectable human insulin by day 14, whereas PEP-treated grafts retained detectable insulin through day 14 across co-transplantation, pretreatment, and systemic administration strategies. PEP co-transplantation delayed, but did not prevent, xenograft rejection, with graft loss observed by day 21. This delay was associated with reduced NK1.1+ cell infiltration and lower expression of selected inflammatory and rejection-associated markers, including NK1.1, Nos2, and Nlrp3. These findings suggest that PEP modulates graft-associated immune responses and may serve as an adjunct immunomodulatory strategy for stem cell-based therapies for type 1 diabetes. Further studies are needed to evaluate sustained graft durability, safety, and translational efficacy.

Directed differentiation protocols have recently been developed to produce stem cell-derived α (SC-α) cells as a potential component of a complete cell-based therapy for T1D, to complement the more widely studied stem cell-derived β (SC-β) cells. Differentiation protocols for SC-β cells produce off-target cell populations implicated in the development of outgrowths in SC-β cell grafts, but outgrowths from SC-α cells have not been explored. This study identifies that engrafted SC-α cells generate outgrowths of similar composition to SC-β cell outgrowths. Both cell types share outgrowth-driving populations marked by SOX9, CDX2, or SOX2. Single-cell RNA sequencing was used to reveal an enrichment in cell-adhesion signaling events in outgrowth-driving populations. Small-molecule inhibition of the Notch pathway was insufficient to disrupt all three outgrowth-driving populations. A comprehensive disruption of cell-adhesion signaling via single-cell dispersion and reaggregation is found to reduce the outgrowth propensity in engrafted SC-α and SC-β cells. Together, these results suggest that disrupting residual progenitor cells with SC-α and SC-β cell clusters can enhance the safety profile of these cell therapy products for T1D therapy.

This review describes approaches for encapsulation of pluripotent stem cells, the biomaterials used, strategies for differentiation into hepatocytes and β cells, and the impact of microencapsulation on cellular therapy.

Human pluripotent stem cells (hPSCs) are capable of unlimited proliferation and can undergo differentiation to give rise to cells and tissues of the three primary germ layers. While directing lineage selection of hPSCs has been an active area of research, improving the efficiency of differentiation remains an important objective. In this study, we describe a two-compartment microfluidic device for co-cultivation of adult human hepatocytes and stem cells. Both cell types were cultured in a 3D or spheroid format. Adult hepatocytes remained highly functional in the microfluidic device over the course of 4 weeks and served as a source of instructive paracrine cues to drive hepatic differentiation of stem cells cultured in the neighboring compartment. The differentiation of stem cells was more pronounced in microfluidic co-cultures compared to a standard hepatic differentiation protocol. In addition to improving stem cell differentiation outcomes, the microfluidic co-culture system described here may be used for parsing signals and mechanisms controlling hepatic cell fate.