Vegfa/vegfr2 signaling is necessary for zebrafish islet vessel development, but is dispensable for beta-cell and alpha-cell formation.

The mechanisms underlying zebrafish pancreatic islet vascularization have not been well characterized. We sought to determine the angiogenic factors responsible for islet vascularization and assess whether an absence of endothelial cells affects beta-cell and alpha-cell formation. We used a double transgenic zebrafish Tg(fli1:EGFP; insa:tagRFP) to label endothelial cells and beta-cells, respectively. Beta-cells developed adjacent to endothelial cells and by 72 hours post fertilization (hpf) the zebrafish pancreatic islet was highly vascularized. Zebrafish beta-cells express vascular endothelial growth factors (vegf), vegfaa and vegfab. Double knockdown of vegfaa and vegfab or the primary Vegfa receptors (Vegfr2), kdr and kdrl, resulted in vessel deficient islets. While beta-cell and alpha-cell numbers remained unchanged in vessel deficient islets, insulina expression was downregulated relative to controls. Vegfaa/Vegfab-Vegfr2 signaling is necessary for proper islet vessel development, but not for the initial formation of beta-cells and alpha-cells.

Contribution of a non-ß-cell source to ß-cell mass during pregnancy.

ß-cell mass in the pancreas increases significantly during pregnancy as an adaptation to maternal insulin resistance. Lineage tracing studies in rodents have presented conflicting evidence on the role of cell duplication in the formation of new ß-cells during gestation, while recent human data suggest that new islets are a major contributor to increased ß-cell mass in pregnancy. Here, we aim to: 1) determine whether a non-ß-cell source contributes to the appearance of new ß-cells during pregnancy and 2) investigate whether recapitulation of the embryonic developmental pathway involving high expression of neurogenin 3 (Ngn3) plays a role in the up-regulation of ß-cell mass during pregnancy. Using a mouse ß-cell lineage-tracing model, which labels insulin-producing ß-cells with red fluorescent protein (RFP), we found that the percentage of labeled ß-cells dropped from 97% prior to pregnancy to 87% at mid-pregnancy. This suggests contribution of a non-ß-cell source to the increase in total ß-cell numbers during pregnancy. In addition, we observed a population of hormone-negative, Ngn3-positive cells in islets of both non-pregnant and pregnant mice, and this population dropped from 12% of all islets cells in the non-pregnant mice to 5% by day 8 of pregnancy. Concomitantly, a decrease in expression of Ngn3 and changes in its upstream regulatory network (Sox9 and Hes-1) as well as downstream targets (NeuroD, Nkx2.2, Rfx6 and IA1) were also observed during pregnancy. Our results show that duplication of pre-existing ß-cells is not the sole source of new ß-cells during pregnancy and that Ngn3 may be involved in this process.