PP2Ac knockdown attenuates lipotoxicity­induced pancreatic ß­cell dysfunction and apoptosis.

Protein phosphatase 2A (PP2A) is one of the most common serine/threonine phosphatases in mammalian cells, and it primarily functions to regulate cell signaling, glycolipid metabolism and apoptosis. The catalytic subunit of PP2A (PP2Ac) plays an important role in the functions of the protein. However, there are few reports on the regulatory role of PP2Ac in pancreatic ß-cells under lipotoxic conditions. In the present study, mouse insulinoma 6 (MIN6) pancreatic cells were transfected with short hairpin RNAs to generate PP2Ac knockdown cells and incubated with palmitate (PA) to establish a lipotoxicity model. Serine/threonine phosphatase assay system, Cell Counting Kit-8, flow cytometry, enzyme-linked immunosorbent assay and western blotting were used to measure PP2A activity, cell viability, apoptosis, oxidative stress and insulin secretion in the cells. In addition, a mouse model of lipotoxicity was established with a high-fat diet (HFD) and the knockdown of PP2Ac using adeno-associated viruses to interfere with PP2Ac expression in the pancreatic tissues. The activity of PP2A in the mouse pancreatic tissue and the serum insulin level were measured. Furthermore, the proliferation of mouse pancreatic ß-cells was assessed using pancreatic tissue immunofluorescence. PP2Ac knockdown inhibited lipotoxicity-induced PP2A hyperactivation, increased the resistance of pancreatic ß-cells to lipotoxicity and attenuated PA-induced apoptosis in MIN6 cells. It also protected the endoplasmic reticulum and mitochondria, and ameliorated insulin secretion. The results of mRNA sequencing and western blotting analysis suggested that the protective effects of PP2Ac knockdown in MIN6 cells may be mediated via the MAPK pathway. Moreover, the results of the animal experiments suggested that specific knockdown of pancreatic PP2Ac effectively attenuated HFD-induced insulin resistance and reduced the compensatory proliferation of pancreatic ß-cells in mice. In summary, the present study revealed the effects of interfering with PP2Ac gene expression on pancreatic ß-cells in vivo and in vitro and the underlying mechanisms, which may provide insights for the treatment of type 2 diabetes mellitus in the clinic.

FUNDC1 modulates mitochondrial defects and pancreatic ß-cell dysfunction under lipotoxicity.

Insulin resistance and many metabolic disorders are causally linked to mitochondrial dysfunction or defective mitochondrial quality control. Mitophagy is a highly selective mechanism that recognizes and removes damaged mitochondria to maintain mitochondrial homeostasis. Here, we addressed the potential role of FUNDC1, a mediator of mitophagy, in pancreatic ß-cell dysfunction under lipotoxicity. In pancreatic MIN6 cells, FUNDC1 deficiency aggravated palmitate-induced mitochondrial dysfunction, which led to cell death and insulin insensitivity. Interestingly, FUNDC1 overexpression prevented these cellular harms brought on by palmitate. In mice models, pancreatic-specific FUNDC1 overexpression alleviated high-fat diet (HFD)-induced insulin resistance and obesity. Mechanistically, pancreatic-specific overexpression of FUNDC1 ameliorated mitochondrial defects and endoplasmic reticulum (ER) stress upon HFD. Our research indicates that FUNDC1 plays an essential role in apoptosis and dysfunction of pancreatic ß-cells via modulating lipotoxicity-induced mitochondrial defects.

Hepatic IDH2 regulates glycolysis and gluconeogenesis.

BACKGROUND AND AIMS: The liver plays a central role in controlling glucose and lipid metabolism. IDH2, a mitochondrial protein, controls TCA cycle flux. However, its role in regulating metabolism in obesity is still unclear. This study intends to investigate the impact of hepatic IDH2 expression on overnutrition-regulated glucose and lipid metabolism. METHODS: Hepatic IDH2 was knocked-out in mice by the approach of CRISPR-Cas9. Mice were subjected to starvation and refeeding for hepatic glucose and lipid studies in vivo. Primary hepatocytes and mouse normal liver cell line, AML12 cells were used for experiments in vitro. RESULTS: This study found that IDH2 protein levels were elevated in the livers of obese people and mice with high-fat diet consumption or hepatic steatosis. Liver IDH2-deletion mice (IDH2LKO) were resistant to high-fat diet-induced body weight gain, with lower serum glucose and TG levels, increased insulin sensitivity, and higher FGF21 secretion, despite the higher TG content in the liver. Consistently, overexpression of IDH2 in hepatocytes promoted gluconeogenesis and enhanced glycogenesis. By performing mass spectrometry and proteomics analyses, we further demonstrated that IDH2-deficiency in hepatocytes accelerated ATP production by increasing forward TCA cycle flux, thus promoting glycolysis pathway and decreasing glycogen synthesis at refeeding state, and inhibiting hepatic gluconeogenesis, increasing ß-oxidation during starvation. Moreover, experiments in vivo demonstrated that IDH2-knockout might not exacerbate hepatic inflammatory responses in the NASH model. CONCLUSIONS: Elevated hepatic IDH2 under over-nutrition state contributes to elevated gluconeogenesis and glycogen synthesis. Inhibition of IDH2 in the liver could be a potential therapeutic target for obesity and diabetes.

Nr2e1 ablation impairs liver glucolipid metabolism and induces inflammation, high-fat diets amplify the damage.

Nonalcoholic fatty liver disease (NAFLD) is a common and complex metabolic disorder. Despite the widespread concern, there are still few effective treatments except lifestyle interventions. Nuclear receptor subfamily 2 group E member 1 (Nr2e1) is a transcription factor which regulates many biological processes, including development, growth, and differentiation of nerve cells. However, its specific function in hepatocyte is still unknown. In the present study, we found that the expression of Nr2e1 decreased in the livers of high-fat diet-fed mice. We generated Nr2e1 knockout (KO) mice and studied whether Nr2e1 ablation was related to NAFLD. We found that typical pathological features of NAFLD, including insulin resistance, hepatic steatosis, and inflammation, were present in Nr2e1-KO mice or high-fat diet-induced mice models. In conclusion, Nr2e1 ablation promotes liver steatosis and systemic insulin resistance. Nr2e1 may play a protective role in the formation of NAFLD and may serve as a worthy therapeutic target for NAFLD.