It is possible that PSN632408 could have improved blood glucose levels and increased pancreatic b-cell mass by a 1370468-36-2 direct action on b-cells and/or by stimulating GLP-1 production by intestinal L cells. These alternatives might be answered by using GLP-1 receptor knockout mice. Multiple factors may contribute to the effects on pancreatic bcell mass including b-cell regeneration , hypertrophy and apoptosis. To assess b-cell regeneration in these mice, we continuously labeled mice with BrdU to track replicating cells and measured the proliferation rates in terms of percentage of insulin and BrdU co-positive cells in total islet bcells. We found that PSN632408 and sitagliptin combination treatment significantly increased the numbers of replicating b-cells compared with vehicle or PSN632408 treatment alone. It has been suggested that BrdU incorporation is associated with a DNA damage response, not replication, in human pancreatic b-cells . Therefore, we did not solely rely upon BrdU incorporation as evidence of b-cell replication. We used Ki67, a cellular marker for replication, to further determine b-cell replication. Ki67 is strictly associated with cell replication and is expressed during all phases of the cell cycle tracking active dividing cells. Although Ki67 staining would have identified the cells undergoing cell division during the last fraction of the treatment period, our results corroborated a similar trend observed with insulin and BrdU staining. Using insulin and Ki67 staining as reliable evidence of bcell replication, we found that treatment with PSN632408 alone or sitagliptin alone could stimulate b-cell replication; however, PSN632408 and sitagliptin combination was significantly better than either alone. Whether the replication of these b-cells was from self-renewal of mature b cells or was from specialized progenitors in islets needs to be further investigated. There is compelling evidence supporting b-cell neogenesis from Quercitrin precursors/stem cells in the ductal epithelium of the pancreas as a mechanism of b-cell regeneration in several diabetic models . Exendin-4, a GLP-1 analogue, has been shown to stimulate not only b-cell replication, but also b-cell neogenesis . In this study, we observed
