Directed differentiation of human embryonic stem cells towards a pancreatic cell fate.

AIMS/HYPOTHESIS: The relative lack of successful pancreatic differentiation of human embryonic stem cells (hESCs) may suggest that directed differentiation of hESCs into definitive endoderm and subsequent commitment towards a pancreatic fate are not readily achieved. The aim of this study was to investigate whether sequential exposure of hESCs to epigenetic signals that mimic in vivo pancreatic development can efficiently generate pancreatic endodermal cells, and whether these cells can be further matured and reverse hyperglycaemia upon transplantation. MATERIALS AND METHODS: The hESCs were sequentially treated with serum, activin and retinoic acid (RA) during embryoid body formation. The patterns of gene expression and protein production associated with embryonic germ layers and pancreatic endoderm were analysed by RT-PCR and immunostaining. The developmental competence and function of hESC-derived PDX1-positive cells were evaluated after in vivo transplantation. RESULTS: Sequential treatment with serum, activin and RA highly upregulated the expression of the genes encoding forkhead box protein A2 (FOXA2), SRY-box containing gene 17 (SOX17), pancreatic and duodenal homeobox 1 (PDX1) and homeobox HB9 (HLXB9). The population of pancreatic endodermal cells that produced PDX1 was significantly increased at the expense of ectodermal differentiation, and a subset of the PDX1-positive cells also produced FOXA2, caudal-type homeobox transcription factor 2 (CDX2), and nestin (NES). After transplantation, the PDX1-positive cells further differentiated into mature cell types producing insulin and glucagon, resulting in amelioration of hyperglycaemia and weight loss in streptozotocin-treated diabetic mice. CONCLUSIONS/INTERPRETATION: Our strategy allows the progressive differentiation of hESCs into pancreatic endoderm capable of generating mature pancreatic cell types that function in vivo. These findings may establish the basis of further investigations for the purification of transplantable islet progenitors derived from hESCs.

Prognostic factors affecting graft and patient survival in cadaveric and living kidney transplantation.

Numerous studies have reported various prognostic factors that affect graft and patient survival in living and cadaveric donor kidney transplantation (KT). The purpose of this study was to evaluate the clinical outcomes and prognostic factors affecting graft and patient survivals in living and cadaveric donor KT. Between February 1995 and December 2001, 421 patients who had undergone cadaveric donor KT (group I: 216 cases, 51.3%) or living donor KT (group II: 205 cases, 48.7%), were retrospectively analyzed. Five-year overall graft survival rates in living was significantly better than that in cadaveric donor KT, respectively (P = .0234). There was no difference in patient survival rates between the two groups. Such factors as absence of rejection, female donor, female recipient, adult KT according to recipient age (>14 years), and donor serum creatinine level just before transplantation (< 2.5 mg/dL) were significantly associated with good graft survival among cadaveric donor KT, whereas two factors-absence of rejection and adult KT according to recipient age (>14 years)-influenced graft survival in living donor KT. In multivariate analysis, the only significant prognostic factor related to graft survival was the presence of rejection. In conclusion, we suggest that the presence of rejection is the only factor that impairs graft survival in both cadaveric and living donor KT, while other factors affected graft survival differently in the two groups.

Prevention of restenosis by a herpes simplex virus mutant capable of controlled long-term expression in vascular tissue in vivo.

Neointimal hyperplasia resulting from vascular smooth muscle cell (SMC) proliferation and luminal migration is the major cause of autologous vein graft failure following vascular coronary or peripheral bypass surgery. Strategies to attenuate SMC proliferation by the delivery of oligonucleotides or genes controlling cell division rely on the use of high concentrations of vectors, and require pre-emptive disruption of the endothelial cell layer. We report a genetically engineered herpes simplex virus (HSV-1) mutant that, in an in vivo rabbit model system, infects all vascular layers without prior injury to the endothelium; expresses a reporter gene driven by a viral promoter with high efficiency for at least 4 weeks; exhibits no systemic toxicity; can be eliminated at will by administration of the antiviral drug acyclovir; and significantly reduces SMC proliferation and restenosis in vein grafts in immunocompetent hosts.