Overexpression of Hepatocyte Nuclear Factor-4 alpha Initiates Cell Cycle Entry, but Is not Sufficient to Promote beta-Cell Expansion in Human Islets

摘要

The transcription factor HNF4 alpha ( hepatocyte nuclear factor-4 alpha) is required for increased beta-cell proliferation during metabolic stress in vivo. We hypothesized that HNF4 alpha could induce proliferation of human beta-cells. We employed adenoviral-mediated overexpression of an isoform of HNF4 alpha (HNF4 alpha 8) alone, or in combination with cyclin-dependent kinase (Cdk) 6 and Cyclin D3, in human islets. Heightened HNF4 alpha 8 expression led to a 300-fold increase in the number of beta-cells in early S-phase. When we overexpressed HNF4 alpha 8 together with Cdk6 and Cyclin D3, beta-cell cycle entry was increased even further. However, the punctate manner of bromodeoxyuridine incorporation into HNF4 alpha(High) beta-cells indicated an uncoupling of the mechanisms that control the concise timing and execution of each cell cycle phase. Indeed, in HNF4 alpha 8-induced bromodeoxyuridine(+,punctate) beta-cells we observed signs of dysregulated DNA synthesis, cell cycle arrest, and activation of a double stranded DNA damage-associated cell cycle checkpoint mechanism, leading to the initiation of loss of beta-cell lineage fidelity. However, a substantial proportion of beta-cells stimulated to enter the cell cycle by Cdk6 and Cyclin D3 alone also exhibited a DNA damage response. HNF4 alpha 8 is a mitogenic signal in the human beta-cell but is not sufficient for completion of the cell cycle. The DNA damage response is a barrier to efficient beta-cell proliferation in vitro, and we suggest its evaluation in all attempts to stimulate beta-cell replication as an approach to diabetes treatment. (Molecular Endocrinology 26: 1590-1602, 2012)