Beta-cell replication dramatically declines with age. Here, we report that the level of CENP-A, a protein required for cell division, declines precipitously with age in an islet-specific manner. CENP-A is essentially undetectable after age 29 in humans. However, exocrine cells retain CENP-A expression. The decline in islet-cell CENP-A expression is more striking in humans than in mice, where CENP-A expression continues to be detectable at low levels even in elderly mice. The mechanism by which CENP-A declines appears to be post-transcriptional, as there was no correlation between CENP-A mRNA levels and age or islet purity. This finding has implications for efforts to induce beta-cell replication as a treatment for diabetes.

Different factors may lead to hepatitis. Among which are liver inflammation and poisoning. We chose two hepatitis models, typical for these two underlying causes. Thus, we aimed to characterize the role of protease-activated receptor 2 (Par2) in liver regeneration and inflammation to reconcile Par2 conflicting role in many damage models, which sometimes aggravates the induced damage and sometimes alleviates it.

The maternal embryonic leucine zipper kinase (MELK) is expressed in stem/progenitor cells in some adult tissues, where it has been implicated in diverse biological processes, including the control of cell proliferation. Here, we described studies on its role in adult pancreatic regeneration in response to injury induced by duct ligation and β-cell ablation. MELK expression was studied using transgenic mice expressing GFP under the control of the MELK promoter, and the role of MELK was studied using transgenic mice deleted in the MELK kinase domain. Pancreatic damage was initiated using duct ligation and chemical beta-cell ablation. By tracing MELK expression using a MELK promoter-GFP transgene, we determined that expression was extremely low in the normal pancreas. However, following duct ligation and β-cell ablation, it became highly expressed in pancreatic ductal cells while remaining weakly expressed in α-cells and β- cells. In a mutant mouse in which the MELK kinase domain was deleted, there was no effect on pancreatic development. There was no apparent effect on islet regeneration, either. However, following duct ligation there was a dramatic increase in the number of small ducts, but no change in the total number of duct cells or duct cell proliferation. In vitro studies indicated that this was likely due to a defect in cell migration. These results implicate MELK in the control of the response of the pancreas to injury, specifically controlling cell migration in normal and transformed pancreatic duct cells.

The Wnt pathway effector gene TCF7L2 has been linked to type II diabetes, making it important to study the role of Wnt signaling in diabetes pathogenesis. We examined the expression of multiple Wnt pathway components in pancreases from normal individuals and type II diabetic individuals. Multiple members of the Wnt signaling pathway, including TCF7L2, Wnt2b, beta-catenin, pGSK3beta, TCF3, cyclinD1, and c-myc, were undetectable or expressed at low levels in islets from nondiabetic individuals, but were also upregulated specifically in islets of type II diabetic patients. Culture of pancreatic tissue and islet isolation led to Wnt activation that was reversed by the Wnt antagonist sFRP, demonstrating that Wnt activation in that setting was due to soluble Wnt factors. These data support a model in which the Wnt pathway plays a dynamic role in the pathogenesis of type II diabetes and suggest manipulation of Wnt signaling as a new approach to beta-cell-directed diabetes therapy.

We report the discovery of strong HNF4α agonists and their use to uncover a previously unknown pathway by which HNF4α controls the level of fat storage in the liver. This involves the induction of lipophagy by dihydroceramides, the synthesis and secretion of which is controlled by genes induced by HNF4α. The HNF4α activators are N-trans caffeoyltyramine (NCT) and N-trans feruloyltyramine (NFT), which are structurally related to the known drugs alverine and benfluorex, which we previously showed to be weak HNF4α activators. In vitro, NCT and NFT induced fat clearance from palmitate-loaded cells. In DIO mice, NCT led to recovery of hepatic HNF4α expression and reduction of steatosis. Mechanistically, increased dihydroceramide production and action downstream of HNF4α occurred through increased expression of HNF4α downstream genes, including SPNS2 and CYP26A1. NCT was completely nontoxic at the highest dose administered and so is a strong candidate for an NAFLD therapeutic.

HNF4α has been implicated in IBD through a number of genome-wide association studies. Recently, we developed potent HNF4α agonists, including N-trans caffeoyltyramine (NCT). NCT was identified by structural similarity to previously the previously identified but weak HNF4α agonists alverine and benfluorex. Here, we administered NCT to mice fed a high fat diet, with the goal of studying the role of HNF4α in obesity-related diseases. Intestines from NCT-treated mice were examined by RNA-seq to determine the role of HNF4α in that organ. Surprisingly, the major classes of genes altered by HNF4α were involved in IBD and Paneth cell biology. Multiple genes downregulated in IBD were induced by NCT. Paneth cells identified by lysozyme expression were reduced in high fat fed mice. NCT reversed the effect of high fat diet on Paneth cells, with multiple markers being induced, including a number of defensins, which are critical for Paneth cell function and intestinal barrier integrity. NCT upregulated genes that play important role in IBD and that are downregulated in that disease. It reversed the loss of Paneth cell markers that occurred in high fat diet fed mice. These data suggest that HNF4α could be a therapeutic target for IBD and that the agonists that we have identified could be candidate therapeutics.

We report here that the potent HNF4α agonist N-trans-caffeoyltyramine (NCT) promotes weight loss by inducing an increase in mitochondrial mass and function, including fatty acid oxidation. Previously, we found in a short term trial in obese mice that NCT promoted reversal of hepatic steatosis through a mechanism involving the stimulation of lipophagy by dihydroceramides. NCT led to increased dihydroceramide levels by inhibiting dihydroceramide conversion to ceramides. Here, we were able to administer NCT orally, permitting longer term administration. Mice fed NCT mixed with high fat diet exhibited decreased weight. Examination of RNA-seq data revealed an increase in PPARGC1A, a central regulator of mitochondrial biogenesis. In addition to the decreased hepatic steatosis that we found previously, mice fed a high fat diet containing NCT mice weighed substantially less than control mice fed high fat diet alone. They had increased mitochondrial mass, exhibited increased fatty acid oxidation, and had an increased level of NAD. Markers of liver inflammation such as interleukin-6 (IL-6) and tumor necrosis factor alpha (TNFα), which are important in the progression of non-alcoholic fatty liver disease to non-alcoholic steatohepatitis were decreased by NCT. There was no evidence of any toxicity from NCT consumption. These results indicate that HNF4α is an important regulator of mitochondrial mass and function and support that use of HNF4α to treat disorders of fatty acid excess, potentially including obesity, NAFLD, and NASH.

The nature and even existence of adult pancreatic endocrine stem or progenitor cells is a subject of controversy in the field of beta-cell replacement for diabetes. One place to search for such cells is in the nonendocrine fraction of cells that remain after islet isolation, which consist of a mixture of epithelia and mesenchyme. Culture in G418 resulted in elimination of the mesenchymal cells, leaving a highly purified population of nonendocrine pancreatic epithelial cells (NEPECs). To evaluate their differentiation potential, NEPECs were heritably marked and transplanted under the kidney capsule of immunodeficient mice. When cotransplanted with fetal pancreatic cells, NEPECs were capable of endocrine differentiation. We found no evidence of beta-cell replication or cell fusion that could have explained the appearance of insulin positive cells from a source other than NEPECs. Nonendocrine-to-endocrine differentiation of NEPECs supports the existence of endocrine stem or progenitor cells within the epithelial compartment of the adult human pancreas.

Understanding the mechanisms by which cells sense and respond to injury is central to developing therapies to enhance tissue regeneration. Previously, we showed that pancreatic injury consisting of acinar cell damage+β-cell ablation led to islet cell transdifferentiation. Here, we report that the molecular mechanism for this requires activating protease-activated receptor-2 (PAR2), a G-protein-coupled receptor. PAR2 modulation was sufficient to induce islet cell transdifferentiation in the absence of β-cells. Its expression was modulated in an islet cell type-specific manner in murine and human type 1 diabetes (T1D). In addition to transdifferentiation, PAR2 regulated β-cell apoptosis in pancreatitis. PAR2's role in regeneration is broad, as mice lacking PAR2 had marked phenotypes in response to injury in the liver and in digit regeneration following amputation. These studies provide a pharmacologically relevant target to induce tissue regeneration in a number of diseases, including T1D.