Erica P. Cai

Hepatocyte-specific Deletion of Janus Kinase 2 (JAK2) Protects against Diet-induced Steatohepatitis and Glucose Intolerance

Background: JAK2 mediates signaling by a number of cytokines in the liver. Results: Hepatic JAK2 KO mice developed spontaneous steatosis but were protected from high fat diet-induced steatohepaitits and insulin resistance. Conclusion: Hepatic JAK2 is required for the development of diet-induced steatohepatitis and glucose intolerance. Significance: Understanding the role of JAK2 in metabolism will provide insights into the pathogenesis of the metabolic syndrome. Non-alcoholic fatty liver disease (NAFLD) is becoming the leading cause of chronic liver disease and is now considered to be the hepatic manifestation of the metabolic syndrome. However, the role of steatosis per se and the precise factors required in the progression to steatohepatitis or insulin resistance remain elusive. The JAK-STAT pathway is critical in mediating signaling of a wide variety of cytokines and growth factors. Mice with hepatocyte-specific deletion of Janus kinase 2 (L-JAK2 KO mice) develop spontaneous steatosis as early as 2 weeks of age. In this study, we investigated the metabolic consequences of jak2 deletion in response to diet-induced metabolic stress. To our surprise, despite the profound hepatosteatosis, deletion of hepatic jak2 did not sensitize the liver to accelerated inflammatory injury on a prolonged high fat diet (HFD). This was accompanied by complete protection against HFD-induced whole-body insulin resistance and glucose intolerance. Improved glucose-stimulated insulin secretion and an increase in β-cell mass were also present in these mice. Moreover, L-JAK2 KO mice had progressively reduced adiposity in association with blunted hepatic growth hormone signaling. These mice also exhibited increased resting energy expenditure on both chow and high fat diet. In conclusion, our findings indicate a key role of hepatic JAK2 in metabolism such that its absence completely arrests steatohepatitis development and confers protection against diet-induced systemic insulin resistance and glucose intolerance.

In Vivo Role of Focal Adhesion Kinase in Regulating Pancreatic β-Cell Mass and Function Through Insulin Signaling, Actin Dynamics, and Granule Trafficking

Focal adhesion kinase (FAK) acts as an adaptor at the focal contacts serving as a junction between the extracellular matrix and actin cytoskeleton. Actin dynamics is known as a determinant step in insulin secretion. Additionally, FAK has been shown to regulate insulin signaling. To investigate the essential physiological role of FAK in pancreatic β-cells in vivo, we generated a transgenic mouse model using rat insulin promoter (RIP)–driven Cre-loxP recombination system to specifically delete FAK in pancreatic β-cells. These RIPcre+fakfl/fl mice exhibited glucose intolerance without changes in insulin sensitivity. Reduced β-cell viability and proliferation resulting in decreased β-cell mass was observed in these mice, which was associated with attenuated insulin/Akt (also known as protein kinase B) and extracellular signal–related kinase 1/2 signaling and increased caspase 3 activation. FAK-deficient β-cells exhibited impaired insulin secretion with normal glucose sensing and preserved Ca2+ influx in response to glucose, but a reduced number of docked insulin granules and insulin exocytosis were found, which was associated with a decrease in focal proteins, paxillin and talin, and an impairment in actin depolymerization. This study is the first to show in vivo that FAK is critical for pancreatic β-cell viability and function through regulation in insulin signaling, actin dynamics, and granule trafficking.

DJ-1 links muscle ROS production with metabolic reprogramming and systemic energy homeostasis in mice

Reactive oxygen species (ROS) have been linked to a wide variety of pathologies, including obesity and diabetes, but ROS also act as endogenous signalling molecules, regulating numerous biological processes. DJ-1 is one of the most evolutionarily conserved proteins across species, and mutations in DJ-1 have been linked to some cases of Parkinsons disease. Here we show that DJ-1 maintains cellular metabolic homeostasis via modulating ROS levels in murine skeletal muscles, revealing a role of DJ-1 in maintaining efficient fuel utilization. We demonstrate that, in the absence of DJ-1, ROS uncouple mitochondrial respiration and activate AMP-activated protein kinase, which triggers Warburg-like metabolic reprogramming in muscle cells. Accordingly, DJ-1 knockout mice exhibit higher energy expenditure and are protected from obesity, insulin resistance and diabetes in the setting of fuel surplus. Our data suggest that promoting mitochondrial uncoupling may be a potential strategy for the treatment of obesity-associated metabolic disorders.

HIV drug resistance detected during low-level viraemia is associated with subsequent virologic failure.

Background:The clinical implications of emergent HIV drug resistance on samples with low-level viraemia (LLV <1000 copies/ml) remain unclear. We undertook the present analysis to evaluate the impact of emergent HIV drug resistance at LLV on the risk of subsequent virologic failure. Methods:One thousand, nine hundred and sixty-five patients had genotype results at LLV. Risk of virologic failure (≥1000 copies/ml) after LLV was evaluated by Kaplan–Meier analysis and Cox proportional hazards regression. Resistance was assessed using the Stanford algorithm or virtual phenotypes. Patients were grouped into four susceptibility categories (’GSS’ or ‘vPSS’) during LLV, corresponding to the number of ‘active’ drugs prescribed: <1; 1–1.5; 2–2.5; and ≥3. Results:A total of 1702 patients with follow-up on constant therapy were eligible for analysis. Participants excluded due to changing therapy or loss to follow-up before their next observation had mostly similar characteristics to included participants. There was a ‘dose-dependent’ increase in the hazard ratio for virologic failure with susceptibility categories at LLV. Compared with a GSS of at least 3, hazard ratios for virologic failure were 1.4 for GSS 2–2.5; 2.0 for GSS 1–1.5; and 3.0 for GSS less than 1 (P < 0.001). Numerous sensitivity analyses confirmed these findings. Conclusion:Our results demonstrate that emergent HIV drug resistance at LLV is strongly associated with subsequent virologic failure. Furthermore, we uncovered a ‘dose-dependent’ increase in the hazard ratio for virologic failure with decreasing GSS estimated at the time of LLV. On the basis of these findings, we propose that resistance genotyping be encouraged for HIV-infected individuals on antiretroviral therapy experiencing low-level viraemia.

In Vivo Knockdown of Adipocyte Erythropoietin Receptor Does Not Alter Glucose or Energy Homeostasis

The growing prevalence of obesity and diabetes necessitate a better understanding of the role of adipocyte biology in metabolism. Increasingly, erythropoietin (EPO) has been shown to have extraerythropoietic and cytoprotective roles. Exogenous administration has recently been shown to have beneficial effects on obesity and diabetes in mouse models and EPO can modulate adipogenesis and insulin signaling in 3T3-L1 adipocytes. However, its physiological role in adipocytes has not been identified. Using male and female mice with adipose tissue-specific knockdown of the EPO receptor, we determine that adipocyte EPO signaling is not essential for the maintenance of energy homeostasis or glucose metabolism. Adipose tissue-specific disruption of EPO receptor did not alter adipose tissue expansion, adipocyte morphology, insulin resistance, inflammation, or angiogenesis in vivo. In contrast to the pharmacological effects of EPO, we demonstrate that EPO signaling at physiological levels is not essential for adipose tissue regulation of metabolism.

PTEN deletion in pancreatic α-cells protects against high-fat diet-induced hyperglucagonemia and insulin resistance.

An aberrant increase in circulating catabolic hormone glucagon contributes to type 2 diabetes pathogenesis. However, mechanisms regulating glucagon secretion and α-cell mass are not well understood. In this study, we aimed to demonstrate that phosphatidylinositol 3-kinase (PI3K) signaling is an important regulator of α-cell function. Mice with deletion of PTEN, a negative regulator of this pathway, in α-cells show reduced circulating glucagon levels and attenuated l-arginine–stimulated glucagon secretion both in vivo and in vitro. This hypoglucagonemic state is maintained after high-fat–diet feeding, leading to reduced expression of hepatic glycogenolytic and gluconeogenic genes. These beneficial effects protected high-fat diet–fed mice against hyperglycemia and insulin resistance. The data demonstrate an inhibitory role of PI3K signaling on α-cell function and provide experimental evidence for enhancing α-cell PI3K signaling for diabetes treatment.

Retinoblastoma tumor suppressor protein in pancreatic progenitors controls α- and β-cell fate

Pancreatic endocrine cells expand rapidly during embryogenesis by neogenesis and proliferation, but during adulthood, islet cells have a very slow turnover. Disruption of murine retinoblastoma tumor suppressor protein (Rb) in mature pancreatic β-cells has a limited effect on cell proliferation. Here we show that deletion of Rb during embryogenesis in islet progenitors leads to an increase in the neurogenin 3-expressing precursor cell population, which persists in the postnatal period and is associated with increased β-cell mass in adults. In contrast, Rb-deficient islet precursors, through repression of the cell fate factor aristaless related homeobox, result in decreased α-cell mass. The opposing effect on survival of Rb-deficient α- and β-cells was a result of opposing effects on p53 in these cell types. As a consequence, loss of Rb in islet precursors led to a reduced α- to β-cell ratio, leading to improved glucose homeostasis and protection against diabetes.

FAK signalling controls insulin sensitivity through regulation of adipocyte survival

Focal adhesion kinase (FAK) plays a central role in integrin signalling, which regulates growth and survival of tumours. Here we show that FAK protein levels are increased in adipose tissue of insulin-resistant obese mice and humans. Disruption of adipocyte FAK in mice or in 3T3 L1 cells decreases adipocyte survival. Adipocyte-specific FAK knockout mice display impaired adipose tissue expansion and insulin resistance on prolonged metabolic stress from a high-fat diet or when crossed on an obese db/db or ob/ob genetic background. Treatment of these mice with a PPARγ agonist does not restore adiposity or improve insulin sensitivity. In contrast, inhibition of apoptosis, either genetically or pharmacologically, attenuates adipocyte death, restores normal adiposity and improves insulin sensitivity. Together, these results demonstrate that FAK is required for adipocyte survival and maintenance of insulin sensitivity, particularly in the context of adipose tissue expansion as a result of caloric excess.

The redundant role of JAK2 in regulating pancreatic β-cell mass.

Janus kinase (JAK) 2 is a non-receptor tyrosine kinase that mediates the downstream effects of various growth factors, including growth hormone,1 prolactin,2 placental lactogen3 and erythropoietin (EPO).4 EPO is a hematopoietic growth factor that is largely known for its role in promoting proliferation, differentiation and survival of cells in the erythroid lineage. Global loss of the EPO receptor (EPO-R) has been shown to be embryonically lethal in mice due to anemia attributed to defects in erythropoiesis.5 Interestingly, mice with global deficiency of JAK2 share a similar developmental phenotype as the EPO-R knockout mice,6 demonstrating that JAK2 is essential in eliciting the biological effects of EPO, particularly in erythrocytosis. Recent studies from our group have shown that exogenous EPO protects mice against diabetes through direct effects on pancreatic β-cells, and these protective effects are dependent on the presence of JAK2 in the β-cells.7 Here, we briefly highlight the cytoprotective effects of exogenous EPO in the pancreatic β-cells, as well as our new findings on the redundant role of JAK2 in β-cell expansion after high-fat feeding in mice.

Dichotomous role of pancreatic HUWE1/MULE/ARF-BP1 in modulating beta cell apoptosis in mice under physiological and genotoxic conditions

Aims/hypothesisDiabetes mellitus represents a significant burden on the health of the global population. Both type 1 and type 2 diabetes share a common feature of a reduction in functional beta cell mass. A newly discovered ubiquitination molecule HECT, UBA and WWE domain containing 1, E3 ubiquitin protein ligase (HUWE1 [also known as MULE or ARF-BP1]) is a critical regulator of p53-dependent apoptosis. However, its role in islet homeostasis is not entirely clear.MethodsWe generated mice with pancreas-specific deletion of Huwe1 using a Cre-loxP recombination system driven by the Pdx1 promoter (Pdx1cre+Huwe1fl/fl) to assess the in vivo role of HUWE1 in the pancreas.ResultsTargeted deletion of Huwe1 in the pancreas preferentially activated p53-mediated beta cell apoptosis, leading to reduced beta cell mass and diminished insulin exocytosis. These defects were aggravated by ageing, with progressive further decline in insulin secretion and glucose homeostasis in older mice. Intriguingly, Huwe1 deletion provided protection against genotoxicity, such that Pdx1cre+Huwe1fl/fl mice were resistant to multiple-low-dose-streptozotocin-induced beta cell apoptosis and diabetes.Conclusion/interpretationHUWE1 expression in the pancreas is essential in determining beta cell mass. Furthermore, HUWE1 demonstrated divergent roles in regulating beta cell apoptosis depending on physiological or genotoxic conditions.