The receptor, c-Kit, and its ligand, stem cell factor (SCF), are critical for hematopoietic stem cell differentiation and have been implicated in the development, function and survival of rodent islets. Previously we reported that exogenous SCF treatments of cultured human fetal (14-16 wk fetal age) islet-epithelial clusters enhanced islet cell differentiation and proliferation (Int J Biochem Cell Biol 38:961-972, 2006). In the present study, we examined the expression pattern of c-Kit in early to mid-gestation human fetal pancreata and the relevance of c-Kit receptor tyrosine kinase for insulin gene expression and -cell survival. c-Kit is expressed in the intact pancreas in a cell-specific manner, with a significant decrease in immunoreactivity in the duct regions from 8-21 weeks fetal age, paralleled by a significant increase in expression within endocrine regions. These c-Kit⁺ cells are highly proliferative and show frequent co-expression with insulin and glucagon. Treatment of islet-epithelial clusters with anti-ACK45 antibody stimulates c-Kit phosphorylation paralleled by a significant increase in PDX-1 and insulin expression, increased cell proliferation and reduced -cell death. In contrast, transient transfection with c-Kit siRNA results in a 3-4 fold decrease in c-Kit, PDX-1 and insulin expression and decreased cell proliferation. This study describes important changes in the distribution and dynamics of c-Kit-expressing cells during human fetal pancreatic neogenesis, suggesting that c-Kit may be a marker for human pancreatic islet progenitor cells. Functional analysis of the c-Kit receptor tyrosine kinase provides evidence that phosphorylation of c-Kit receptor may be involved in mediating early -cell differentiation and survival.