Consistent with earlier reports 
, we observed reduced β-cells and increased α-cells in patients with type 2 diabetes. Interestingly, we detected a striking increase in the number of α- and β-cells that co-stained with the proliferation marker, PCNA, suggesting that both α- and β -cells are attempting to enter the cell cycle. These data may have important implications for therapeutic efforts to counter hyperglycemia in patients with type 1 or type 2 diabetes that are focused on the regenerative potential of β-cells.
Insulin/IGF-I signaling mediates diverse pathways to modulate proliferation and anti-apoptosis in most mammalian cells including pancreatic islets 
. Recent studies in humans and in rodent models of insulin resistance, diabetes and obesity implicate an important role for insulin/IGF-I signaling in β-cell biology 
. The significant decrease in insulin receptor expression in islets from patients with type 2 diabetes indicates that blunted insulin signaling in β-cells may make them susceptible to apoptosis 
. Alterations in phosphorylation of the pro-apoptotic protein BAD are known to modulate apoptosis. Thus, a reduced phosphorylation of BAD in the diabetic pancreas is consistent with an increased apoptosis in type 2 diabetes that possibly occurs, in part, secondary to hyperglycemia 
. In contrast, the higher α-cell mass may reflect the resistance of α-cells to apoptosis in patients with type 2 diabetes and is associated with higher circulating levels of glucagon 
. We have previously reported that human islets that have been transplanted in streptozotocin-treated diabetic SCID mice exhibit α-cells that are resistant to hyperglycemia-induced apoptosis, in contrast to the apoptosis-susceptible β-cells 
Studies in rodents indicate that β-cell replication is a major mechanism that contributes to maintaining adult β-cell mass 
Our observations of a striking increase in the number of PCNA+ cells clearly indicates that human islet cells are also capable of entering the cell cycle. The significant increase in the number of PCNA+ β-cells in the diabetic group indicates that either the β-cells are attempting to replicate as a compensatory response to peripheral insulin resistance and/or that the increase in PCNA expression is a DNA repair response to overcome the effects of pro-apoptotic stimuli including elevated circulating levels of glucose and free fatty acids - a consistent pathological feature of type 2 diabetes 
. It is worth noting that replicating β-cells are more susceptible to cell death induced by islet amyloid polypeptide that accumulates in β-cells in patients with type 2 diabetes 
. Furthermore, consistent with the findings regarding an increase in PCNA expression in conditions of cell stress, in Affymetrix gene expression studies, we have observed a 40-fold increase in the expression of PCNA mRNA in human islets of Langerhans that have been cultured for five days as compared to 24 hours (Folli F, Perego L, Davalli A, unpublished observations), a condition in which significant β-cell apoptosis can be detected 
. The lack of significant differences in PCNA mRNA in islet samples () may be due to presence of multiple cell types in islets and/or differential regulation of PCNA at the transcriptional versus post-translational levels in β-cells 
. While these possibilities are not mutually exclusive, the reduced β-cell mass clearly indicates an abortive attempt of the PCNA+ cells to progress through the cell cycle and develop into functional β-cells with a normal life span. The down regulation of key cell cycle proteins including p27-kip1 and cdk2 in the diabetic pancreas provides additional evidence for altered islet cell cycle dynamics that could promote a default pathway towards apoptosis in β-cells. For example, p27-kip1, in addition to inhibiting cyclins also acts as an anti-apoptotic factor and the low expression of the protein in β-cells may accelerate the apoptotic process 
. This is compounded by a concomitant reduction in the expression of CDK2, CDK4 and cyclin E proteins, which are essential for multiple steps in the transition from G1 to S phase of the cell cycle.
In addition to anti-apoptosis, insulin signaling regulates the transcription factor FoxO1 that, in turn, interacts with PDX-1 to modulate β-cell proliferation 
. The near complete reversal of nuclear restriction of FoxO1 and rescue of blunted proliferation by re-expression of the insulin receptor in βIRKO cells 
indicates a direct link between insulin signaling and β-cell-cycle control. FoxO proteins, including FoxO1, have been implicated in cell cycle regulation 
. For example, stress-induced FoxO activation has been reported to alter the expression of genes that contribute to cell cycle arrest 
. Additional studies are necessary to investigate the proteins that are directly activated by FoxO1 to modulate islet cell cycle progression. Free fatty acids are also known to modulate expression of insulin signaling proteins in vitro 
. Although we did not observe alterations in expression of enzymes involved in lipid metabolism in diabetic islets (data not shown), it is possible that ectopic lipid deposition in islets could produce some of the changes observed in the diabetic group and requires further study.
In conclusion, we propose that β-cells in patients with T2DM are able to enter the cell-cycle, but fail to proliferate successfully to compensate for peripheral insulin resistance due to dysfunctional insulin signaling and cell-cycle arrest (). Restoration of insulin signaling and cell-cycle control in β-cells may be one approach to plan therapeutic strategies to counter β-cell loss in T2DM.
Schematic linking the growth factor pathway to defects in cell cycle progression and apoptosis in diabetic β-cells.