Ghrelin cells fill the void
Beta (?-) cell differentiation depends, amongst a variety of staged events, on the activities of transcription factors Nkx2.2 and Pax4 (Schwitzgebel 2001; Sosa-Pineda 2004; Sosa-Pineda et al. 1997; Sussel et al. 1998; Wang et al. 2004). In the murine model, deletion of Nkx2.2 leads to a unique and total ablation of insulin-producing ?-cells, as well as a significant reduction of a- and PP-cells. Similarly, the loss of Pax4 prevents the expression of Pdx1, HB9 and insulin, markers of ?-cell precursors (Wang et al. 2004). Interestingly, Nkx2.2-/- islets contain a population of cells that do not produce any of the four major islet hormones (Sussel et al. 1998). Dr. Sussel''s group has confirmed that these cells are a unique ghrelin-producing endocrine cell population present in wild type islets.
Panel 1. Wild Type Pancreatic islet of Langerhans. In the center of the panel are shown the four major cell types along with a fifth cell type ? the ghrelin-producing ε-cell...
Ghrelin is a peptide hormone that promotes appetite and release of growth hormone. It exists predominantly in the stomach and to less extent in other tissues such as the pancreas (Kageyama et al. 2005; Kojima et al. 1999; Ueno et al. 2005). It is an endogenous ligand to growth hormone secretagogues receptors (GHS-R). Expression of ghrelin in pancreatic islets has remained controversial since it has been located in a-cells (Date et al. 2002), ?-cells (Volante et al. 2002) and in a unique cell type first hypothesized to be ?ghrelin cells? (Wierup et al. 2002).
Through a comparative microarray analysis of wild type and knockout mutants of Nkx2.2 and Pax4 in mice, it has been shown that (1) Nkx2.2 and Pax4 are independently required to specify the fate of islet ?-cells, (2) the loss of Nkx2.2 and Pax4 leads to ?-cell depletion and ?replacement? with ghrelin-producing ε-cells, and (3) that these newly-identified ε-cells define a unique population of cells within mouse islets (Panel 1) (Prado et al. 2004).
Key notes on methodology
Nkx2.2 (Swiss Black background) and Pax4 (NMRI background) heterozygous mice were generated by homologous recombination.
For microarray analyses, staged embryo embryonic tissues were collected at e12.5-e13.5, e15.5 and e18.5. The Affymetrix Mu19KA gene chip was used to analyze total RNA. Whole pancreata were PCR-genotyped. Heterozygous and homozygous (WT) results were combined since there were no observed phenotypic differences.
All immunochemistry was confirmed by RNA in situ analysis, at e12.5, e14.5, e15.5, e16.5, e18.5 and in neonates. Double immunofluorescence on adjacent pancreas sections using three different antibodies were to used to confirm ghrelin expression.