Controlled induction of human pancreatic progenitors produces functional beta-like cells in vitro.

Directed differentiation of human pluripotent stem cells into functional insulin-producing beta-like cells holds great promise for cell replacement therapy for patients suffering from diabetes. This approach also offers the unique opportunity to study otherwise inaccessible aspects of human beta cell development and function in vitro. Here, we show that current pancreatic progenitor differentiation protocols promote precocious endocrine commitment, ultimately resulting in the generation of non-functional polyhormonal cells. Omission of commonly used BMP inhibitors during pancreatic specification prevents precocious endocrine formation while treatment with retinoic acid followed by combined EGF/KGF efficiently generates both PDX1(+) and subsequent PDX1(+)/NKX6.1(+) pancreatic progenitor populations, respectively. Precise temporal activation of endocrine differentiation in PDX1(+)/NKX6.1(+) progenitors produces glucose-responsive beta-like cells in vitro that exhibit key features of bona fide human beta cells, remain functional after short-term transplantation, and reduce blood glucose levels in diabetic mice. Thus, our simplified and scalable system accurately recapitulates key steps of human pancreas development and provides a fast and reproducible supply of functional human beta-like cells.

CD28/B7 regulation of anti-CD3-mediated immunosuppression in vivo.

FcR-binding "classical" anti-CD3 mAb is a potent immunosuppressive drug that alters CD4(+) and CD8(+) T cell function in vivo via anergy induction and programmed cell death (PCD). Anti-CD3-mediated PCD was Fas independent but was mediated by the mitochondria-initiated apoptosis that was abrogated in Bcl-x(L)-transgenic T cells. The PCD was more pronounced in CD28-deficient mice consistent with defective Bcl-x(L) up-regulation. Residual T cells isolated from anti-CD3-treated wild-type, CD28(-/-), and Bcl-x(L)-transgenic mice were hyporesponsive. The hyporesponsiveness was more pronounced in CD28(-/-) and wild-type mice treated with anti-B7-2, suggesting that CD28 interaction with B7-2 regulates T cell responsiveness in anti-CD3-treated animals. Finally, anti-CD3 treatment led to indefinite cardiac allograft survival in wild-type but not Bcl-x(L) animals. Together these results implicate CD28/B7 signaling in the regulation of both anti-CD3-induced T cell depletion and hyporesponsiveness in vivo, but T cell depletion, not hyporesponsiveness, appears to be critical for anti-CD3 mAb-mediated long-term immune regulation.