RESUMO
AIMS: Obesity associated prolonged hyperinsulinemia followed by ß-cell failure is well established as the pathology behind type 2 diabetes mellitus (T2DM). However, studies on nonobese T2DM have reported it to be a distinct clinical entity with predominant insulin secretory defect. We, therefore, hypothesized that compensatory hyperinsulinemia in response to weight gain is impaired in nonobese subjects. METHODS: This was a cross-sectional study from a community-based metabolic health screening program. Adiposity parameters including body mass index (BMI), waist circumference (WC), body fat percentage, plasma leptin concentration and metabolic parameters namely fasting insulin, glucose, total cholesterol, and triglycerides were measured in 650 individuals (73% healthy, 62% nonobese with a BMI <25). RESULTS: In contrast to obese T2DM, nonobese T2DM patients did not exhibit significant hyperinsulinemia compared with the nonobese healthy group. Age, sex, and fasting glucose adjusted insulin levels, homeostatic model assessment of insulin resistance (HOMA-IR) and HOMA-beta cell function (HOMA-B) were increased in obese T2DM compared with nonobese T2DM. Although adiposity parameters showed strong correlation with fasting insulin in obese healthy (r = 0.38, 0.38, and 0.42, respectively; all p values < 0.001) and T2DM (r = 0.54, 0.54, and 0.66, respectively; all p < 0.001), only BMI and leptin showed a weak correlation with insulin in the nonobese healthy group (0.13 and 0.13, respectively; all p < 0.05) which were completely lost in the nonobese T2DM. CONCLUSIONS: Compensatory hyperinsulinemia in response to weight gain is impaired in the nonobese population making insulin secretory defect rather than IR the major pathology behind nonobese T2DM.
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Hepatocellular death or ballooning distinguishes the transition of simple steatosis to irreversible nonalcoholic steatohepatitis (NASH). However, the molecular mechanism of hepatocellular apoptosis in NASH is largely unclear, and discovery of endogenous mediators that could prevent or inhibit cell death is thereby critical in intercepting NASH progression. Here, we identified pigment epithelium-derived factor (PEDF), a secreted, moonlighting hepatokine as 1 hepatoprotective agent in mice with diet-induced NASH. Hepatic PEDF expression is induced by IL-1ß, which is derived from inflammasome activation in liver-resident Kupffer cells, an effect that is negatively regulated by TNF-α and predominantly secreted by monocyte-derived, recruited, hepatic macrophages. Mechanistically, reciprocal and opposing roles for IL-1ß and TNF-α in PEDF expression are mediated by differential activation of NF-κB. Although augmented TNF-α production leads to temporal reduction of PEDF expression in NASH, PEDF conversely abrogates TNF-α-mediated hepatocyte death by modulating the extrinsic apoptosis pathway. Thus, our study highlights PEDF as a functionally important hepatokine in NASH progression by linking inflammasome activation and hepatocellular death.-Adak, M., Das, D., Niyogi, S., Nagalakshmi, C., Ray, D., Chakrabarti, P. Inflammasome activation in Kupffer cells confers a protective response in nonalcoholic steatohepatitis through pigment epithelium-derived factor expression.
RESUMO
Modification of protein and other biopolymers by labeling them with small or macromolecules has become a very powerful research tool in biochemistry, molecular biology, diagnostics, and therapeutics. However, current methodologies available for their preparations are not straightforward and take several hours of incubation time. In this paper, we describe a new filtration-assisted technique for covalent conjugation between the reactive functional groups of two different molecules (small or macromolecules). Compared to the current method, this new approach significantly reduces the total reaction time from several hours to just a few minutes. The technique has been used for the preparation of conjugates of a small molecule to a protein such as biotin-BSA conjugate or small molecules to a small molecule such as biotin-tyramine conjugate or protein-protein conjugation such as antibody-horseradish peroxidase conjugate. The procedure consists of filtering the reaction mixture multiple times through membrane micropores with the help of two syringes, which make the cross filtration process less laborious. The method saves time, allows conjugation of less than 1mg protein and produces conjugates better than those obtained by the current methods. Although the present technique has been applied on some common conjugation methods, it provides a potentially general method, and may further be expanded for the synthesis of several other macromolecular conjugates.
