Domain 2 of Hepatitis C Virus Protein NS5A Activates Glucokinase and Induces Lipogenesis in Hepatocytes

Hepatitis C virus (HCV) relies on cellular lipid metabolism for its replication, and actively modulates lipogenesis and lipid trafficking in infected hepatocytes. This translates into an intracellular accumulation of triglycerides leading to liver steatosis, cirrhosis and hepatocellular carcinoma, which are hallmarks of HCV pathogenesis. While the interaction of HCV with hepatocyte metabolic pathways is patent, how viral proteins are able to redirect central carbon metabolism towards lipogenesis is unclear. Here, we report that the HCV protein NS5A activates the glucokinase (GCK) isoenzyme of hexokinases through its D2 domain (NS5A-D2). GCK is the first rate-limiting enzyme of glycolysis in normal hepatocytes whose expression is replaced by the hexokinase 2 (HK2) isoenzyme in hepatocellular carcinoma cell lines. We took advantage of a unique cellular model specifically engineered to re-express GCK instead of HK2 in the Huh7 cell line to evaluate the consequences of NS5A-D2 expression on central carbon and lipid metabolism. NS5A-D2 increased glucose consumption but decreased glycogen storage. This was accompanied by an altered mitochondrial respiration, an accumulation of intracellular triglycerides and an increased production of very-low density lipoproteins. Altogether, our results show that NS5A-D2 can reprogram central carbon metabolism towards a more energetic and glycolytic phenotype compatible with HCV needs for replication.

STUDY OF THE EFFECT OF CHITOSAN EXTRACTED FROM THE MUSHROOM ON THE EXPERIMENTALLY INDUCED HYPERLIPIDEMIA IN MALE RABBITS

The present study aimed to identify the therapeutic evaluation of chitosan extracted from the fungus cushroom and pure chitosan on glucose and lipid profile in the blood of 35 male rabbits with hyperlipidemia induced experimentally by cholesterol. The tests included estimation of glucose levels, total cholesterol, triglycerides, high-density lipoproteins, low-density lipoproteins, and very low-density lipoproteins. hyperlipidemia was induced in the male rabbits used in the study which was administered orally with cholesterol 150mg/kg body weight for a week. rabbits were divided into seven groups: control, cholesterol, pure chitosan, mushroom chitosan, cholesterol and pure chitosan, cholesterol and mushroom chitosan and cholesterol and simvastatin. The results of the study showed, the hyperlipidemia induced experimentally resulted a significant increase (P<0.05) in TC, TG, LDL, and VLDL, while no significant difference in HDL compared with control group, on the otherwise the glucose level significantly increase than control. Also, groups of animals treatment with pure chitosan and mushroom chitosan showed a significant decrease (P<0.05) in glucose, TC, TG, LDL, and VLDL, and no significant difference in HDL compared with control group. While, the groups showed treatment with cholesterol and pure chitosan, cholesterol and mushroom chitosan, cholesterol and simvastatin a significant decrease (P<0.05) in glucose, TC, TG, LDL, and VLDL, and a significant increase (P<0.05) in HDL compared with the cholesterol group. The research study revealed that chitosan extracted from mushroom can control the levels of fat concentrations and their complications, in addition to its important role in biochemical variables, and treatment of most disease cases, especially cardiovascular disease.

Caveolae mediated endocytosis of VLDL particles in macrophages requires NPC1 and STARD3 for further lysosomal processing

Macrophages accumulate triglycerides under certain pathological conditions such as atherosclerosis. Triglycerides are carried in the bloodstream as part of very low-density lipoproteins (VLDL) and chylomicrons. How macrophages take up and process VLDL-lipids is not very well known. Here, using VLDL-sized triglyceride-rich emulsion particles, we aimed to study the mechanism by which VLDL-triglycerides are taken up, processed, and stored in macrophages. Our results show that macrophage uptake of emulsion particles mimicking VLDL (VLDLm) is dependent on lipoproteins lipase (LPL) and requires the lipoprotein-binding C-terminal domain of LPL but not the catalytic N-terminal domain. Subsequent internalization of VLDLm-triglycerides by macrophages is carried out by caveolae-mediated endocytosis, followed by triglyceride hydrolysis catalyzed by lysosomal acid lipase. Transfer of lysosomal fatty acids to the ER for subsequent storage as triglycerides is mediated by Stard3, whereas NPC1 was found to promote the extracellular efflux of fatty acids from lysosomes. Our data provide novel insights into how macrophages process VLDL-derived triglycerides and suggest that macrophages have the remarkable capacity to excrete part of the internalized triglycerides as fatty acids.

Metabolic Components of the Combined Course of Osteoarthritis and Obesity

According to statistics provided by the World Health Organization, in 2020 there will be about 2 billion adults, 41 million children under the age of 5 and 340 million children and adolescents aged 5 to 19 who are obese. It is proved that quite often obesity is a predictor of the formation of diseases of the musculoskeletal system. The purpose of the study was to determine the state of lipid and carbohydrate metabolism in patients with different stages of obesity and to establish their influence on the course of osteoarthritis. Materials and methods. The study involved 75 patients with osteoarthritis, which proceed on the background of obesity (main group). 50 patients had manifestations of osteoarthritis without changes in body mass index and 37 almost healthy individuals were included in the control group. Anthropometric data and body mass index were calculated. The state of lipid metabolism was determined by indicators of total cholesterol, triglycerides, high, low and very low density lipoproteins, atherogenic factor. The rate of carbohydrate metabolism was assessed by the presence of insulin resistance by calculating the Homeostasis Model Assessment of Insulin Resistance index. Body mass index was calculated by the Kettle formula. Other anthropometric parameters were determined by measuring the volume of the thigh, waist volume and calculated the ratio of volume of the thigh to waist volume. As markers of lipid metabolism we studied the content of total cholesterol, triglycerides, high-density lipoproteins, low-density lipoproteins and very low-density lipoproteins in the blood system with peroxide. The atherogenicity index was determined by the formula of Klimov A. M. Statistical analysis was performed using the software package "Statistica 10.0" and Excel 2010. To quantify the results, the results were presented as the median with a quarterly interval [Q25%; Q75%] taking into account the lack of normal distribution. Quantitative and ordinal changes were compared using the Mann-Whitney test. The correlation was calculated using Spearman's rank correlations. In all procedures of statistical analysis, the significance level p was assumed to be equal to or less than 0.05 (p <0.05). Results and discussion. We divided the patients of the main group into subgroups depending on the body mass index: overweight patients with I and II obesity degree. The comparative analysis of indicators of lipid metabolism allowed to establish increase of indicators of all atherogenic classes in comparison with control. A statistically significant difference in the rate of total cholesterol was determined only in the case of its comparison between the group with elevated body weight and I degree of obesity. Conclusion. In patients with osteoarthritis, occurring on the background of overweight or obesity, there are shifts in lipid and carbohydrate metabolism, which contribute to metabolic disorders in the body and, in particular, cartilage as a type of connective tissue. Patients with isolated osteoarthritis also have hyperlipidemia and in some cases insulin resistance, which is a negative basis for the progression of the pathological process. One of the factors in the progression of osteoarthritis in obese patients can be considered the activation of free radical oxidation of lipids, which occurs both as a result of osteoarthritis and the presence of concomitant pathology

Glutathione system activity in the blood of overweight postmenopausal women

One of the important components of the antioxidant defense system is the glutathione system, the activity of which, when overweight, changes direction depending on gender and ethnicity. The results of studies involving overweight menopausal women are mixed. The study involved 61 postmenopausal women, who, after clinical and anamnestic examination, were divided into 2 groups: control (BMI = 19-24.9 kg / m2) and overweight group (BMI = 25-29.9 kg/m2). The use of hormone replacement therapy; the use of antioxidant drugs; diseases of endocrine genesis; exacerbation of chronic diseases; premature early menopause; surgical menopause was the exclusion criteria for women from the study. The lipid profile parameters with the calculation of the atherogenic coefficient; reduced and oxidized glutathione levels with the calculation of their ratio, the glutathione S-transferase and glutathione reductase activities were determined in the blood. Overweight women showed an increase in the triacylglycerols (p = 0.041) and cholesterol in very low density lipoproteins levels (p = 0.044). When assessing the glutathione system activity in women of the main group, compared with the control, an increase in the glutathione-S-transferase (p = 0.023) and glutathione reductase (p = 0.022) activities was noted, however, the reduced and oxidized glutathione levels, as well as their ratio did not differ from the control values. The results obtained indicate the activation of the glutathione system enzymatic link in response to changes in lipid status in postmenopausal women with overweight.

Physiological Responses to Hypoxia on Triglyceride Levels

Hypoxia is a condition during which the body or specific tissues are deprived of oxygen. This phenomenon can occur in response to exposure to hypoxic environmental conditions such as high-altitude, or because of pathophysiological conditions such as obstructive sleep apnea. Circumstances such as these can restrict supply or increase consumption of oxygen, leading to oxyhemoglobin desaturation and tissue hypoxia. In certain cases, hypoxia may lead to severe health consequences such as an increased risk of developing cardiovascular diseases and type 2 diabetes. A potential explanation for the link between hypoxia and an increased risk of developing cardiovascular diseases lies in the disturbing effect of hypoxia on circulating blood lipids, specifically its capacity to increase plasma triglyceride concentrations. Increased circulating triglyceride levels result from the production of triglyceride-rich lipoproteins, such as very-low-density lipoproteins and chylomicrons, exceeding their clearance rate. Considerable research in murine models reports that hypoxia may have detrimental effects on several aspects of triglyceride metabolism. However, in humans, the mechanisms underlying the disturbing effect of hypoxia on triglyceride levels remain unclear. In this mini-review, we outline the available evidence on the physiological responses to hypoxia and their impact on circulating triglyceride levels. We also discuss mechanisms by which hypoxia affects various organs involved in the metabolism of triglyceride-rich lipoproteins. This information will benefit scientists and clinicians interested in the mechanistic of the regulatory cascade responsible for the response to hypoxia and how this response could lead to a deteriorated lipid profile and an increased risk of developing hypoxia-related health consequences.

Characterization of lipoprotein lipase storage vesicles in 3T3-L1 adipocytes

Lipoprotein lipase (LPL) is a secreted triglyceride lipase involved in the clearance of very-low-density lipoproteins and chylomicrons from circulation. LPL is expressed primarily in adipose and muscle tissues and transported to the capillary lumen. LPL secretion is regulated by insulin in adipose tissue, however few studies have examined the regulatory and trafficking steps involved in secretion. Here we describe the intracellular localization and insulin-dependent trafficking of LPL in 3T3-L1 adipocytes. We compared LPL trafficking to the better characterized trafficking pathways taken by leptin and GLUT4. We show that LPL trafficking shares some characteristics of these other pathways, but that LPL subcellular localization and trafficking are distinct from GLUT4 and leptin. LPL secretion occurs slowly in response to insulin and rapidly in response to the calcium ionophore ionomycin. This regulated trafficking is dependent on Golgi protein kinase D and the ADP-ribosylation factor GTPase ARF1 localized to caveolar membrane domains. Together, these data give support to a new trafficking pathway for soluble cargo active in adipocytes.

Features of vasoregulatory function of vascular endothelium and correlations between them, hemodynamic and metabolic parameters in patients with coronary heart disease and type 2 diabetes mellitus

The features of the vasoregulating function of the vascular endothelium have been determined and correlations between haemodynamic and metabolic indices have been established in patients with coronary heart disease associated with type II diabetes mellitus. In patients with coronary heart disease associated with type II diabetes mellitus in comparison with patients without an association with diabetes mellitus significantly higher indices of the content of the blood fibrinogen the diameter of the brachial artery (after decompression) were observed. Direct correlations between fasting glycemia and microalbuminuria and reverse correlations with the endothelial dependent vasodilatation (EDVA), fasting glucemia, the blood content of glycosylated hemoglobin and very low density lipoproteins have been found.

The Importance of Lipoprotein Lipase Regulation in Atherosclerosis

Lipoprotein lipase (LPL) plays a major role in the lipid homeostasis mainly by mediating the intravascular lipolysis of triglyceride rich lipoproteins. Impaired LPL activity leads to the accumulation of chylomicrons and very low-density lipoproteins (VLDL) in plasma, resulting in hypertriglyceridemia. While low-density lipoprotein cholesterol (LDL-C) is recognized as a primary risk factor for atherosclerosis, hypertriglyceridemia has been shown to be an independent risk factor for cardiovascular disease (CVD) and a residual risk factor in Atherosclerosis development. In this review, we focus on the lipolysis machinery and discuss the potential role of triglycerides, remnant particles, and lipolysis mediators in the onset and progression of Atherosclerotic cardiovascular disease (ASCVD). This review details a number of important factors involved in the maturation and transportation of LPL to the capillaries, where the triglycerides are hydrolyzed, generating remnant lipoproteins. Moreover, LPL and other factors involved in intravascular lipolysis are also reported to impact the clearance of remnant lipoproteins from plasma and promote lipoprotein retention in capillaries. Apolipoproteins (Apo) and angiopoietin-like proteins (ANGPTLs) play a crucial role in regulating LPL activity and recent insights into LPL regulation may elucidate new pharmacological means to address the challenge of hypertriglyceridemia in Atherosclerosis development.

Sirtuin 3 Restores Synthesis and Secretion of Very Low-Density Lipoproteins in Cow Hepatocytes Challenged with Nonesterified Fatty Acids In Vitro

Fatty liver is closely associated with elevated concentrations of nonesterified fatty acids (NEFA) and a low level of very low-density lipoproteins (VLDL) in blood of dairy cows. High NEFA inhibit the VLDL synthesis and assembly, and cause hepatic triacylglycerol (TAG) deposition. Sirtuin 3 (SIRT3), a mitochondrial deacetylase, antagonizes NEFA-induced TAG accumulation through modulating expressions of fatty acid synthesis and oxidation genes in cow hepatocytes. However, the role of SIRT3 in the VLDL synthesis and assembly was largely unknown. Here we aimed to test whether SIRT3 would recover the synthesis and assembly of VLDL in cow hepatocytes induced by high NEFA. Primary cow hepatocytes were isolated from 3 Holstein cows. Hepatocytes were infected with SIRT3 overexpression adenovirus (Ad-SIRT3), SIRT3-short interfering (si) RNA, or first infected with Ad-SIRT3 and then incubated with 1.0 mM NEFA (Ad-SIRT3 + NEFA). Expressions of key genes in VLDL synthesis and the VLDL contents in cell culture supernatants were measured. SIRT3 overexpression significantly increased the mRNA abundance of microsomal triglyceride transfer protein (MTP), apolipoprotein B100 (ApoB100) and ApoE (p < 0.01), and raised VLDL contents in the supernatants (p < 0.01). However, SIRT3 silencing displayed a reverse effect in comparison to SIRT3 overexpression. Compared with NEFA treatment alone, the Ad-SIRT3 + NEFA significantly upregulated the mRNA abundance of MTP, ApoB100 and ApoE (p < 0.01), and increased VLDL contents in the supernatants (p < 0.01). Our data demonstrated that SIRT3 restored the synthesis and assembly of VLDL in cow hepatocytes challenged with NEFA, providing an in vitro basis for further investigations testing its feasibility against hepatic TAG accumulation in dairy cows during the perinatal period.