Mechanisms of dysregulation of low-density lipoprotein receptor expression in vascular smooth muscle cells by inflammatory cytokines

Objective - Although inflammation is a recognized feature of atherosclerosis, the impact of inflammation on cellular cholesterol homeostasis is unclear. This study focuses on the molecular mechanisms by which inflammatory cytokines disrupt low-density lipoprotein (LDL) receptor regulation.Methods and Results - IL-1 beta enhanced transformation of vascular smooth muscle cells into foam cells by increasing uptake of unmodified LDL via LDL receptors and by enhancing cholesterol esterification as demonstrated by Oil Red O staining and direct assay of intracellular cholesterol concentrations. In the absence of IL-1 beta, a high concentration of LDL decreased LDL receptor promoter activity, mRNA synthesis and protein expression. However, IL-1 beta enhanced LDL receptor expression, overriding the suppression usually induced by a high concentration of LDL and inappropriately increasing LDL uptake. Exposure to IL-1 beta also caused overexpression of the sterol regulatory element binding protein ( SREBP) cleavage-activating protein ( SCAP), and enhanced its translocation from the endoplasmic reticulum to the Golgi, where it is known to cleave SREBP, thereby enhancing LDL receptor gene expression.Conclusions - These observations demonstrate that IL-1 beta disrupts cholesterol-mediated LDL receptor feedback regulation, permitting intracellular accumulation of unmodified LDL and causing foam cell formation. The implication of these findings is that inflammatory cytokines may contribute to intracellular LDL accumulation without previous modification of the lipoprotein

Nonesterified free fatty acids enhance the inflammatory response in renal tubules by inducing extracellular ATP release

In proteinuric renal diseases, excessive plasma nonesterified free fatty acids bound to albumin can leak across damaged glomeruli to be reabsorbed by renal proximal tubular cells and cause inflammatory tubular cells damage by as yet unknown mechanisms. The present study was designed to investigate these mechanisms induced by palmitic acid (PA; one of the nonesterified free fatty acids) overload. Our results show that excess PA stimulates ATP release through the pannexin 1 channel in human renal tubule epithelial cells (HK-2), increasing extracellular ATP concentration approximately threefold compared with control. The ATP release is dependent on caspase-3/7 activation induced by mitochondrial reactive oxygen species. Furthermore, extracellular ATP aggravates PA-induced monocyte chemoattractant protein-1 secretion and monocyte infiltration of tubular cells, enlarging the inflammatory response in both macrophages and HK-2 cells via the purinergic P2X7 receptor-mammalian target of rapamycin-forkhead box O1-thioredoxin-interacting protein/NOD-like receptor protein 3 inflammasome pathway. Hence, PA increases mitochondrial reactive oxygen species-induced ATP release and inflammatory stress, which cause a “first hit,” while ATP itself is a “second hit” in amplifying the renal tubular inflammatory response. Thus, inhibition of ATP release or the purinergic P2X7 receptor may be an approach to reduce renal inflammation and improve renal function

Paradoxical effect of rapamycin on inflammatory stress-induced insulin resistance in vitro and in vivo

Insulin resistance is closely related to inflammatory stress and the mammalian target of rapamycin/S6 kinase (mTOR/S6K) pathway. The present study investigated whether rapamycin, a specific inhibitor of mTOR, ameliorates inflammatory stress-induced insulin resistance in vitro and in vivo. We used tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) stimulation in HepG2 hepatocytes, C2C12 myoblasts and 3T3-L1 adipocytes and casein injection in C57BL/6J mice to induce inflammatory stress. Our results showed that inflammatory stress impairs insulin signaling by reducing the expression of total IRS-1, p-IRS-1 (tyr632), and p-AKT (ser473); it also activates the mTOR/S6K signaling pathway both in vitro and in vivo. In vitro, rapamycin treatment reversed inflammatory cytokine-stimulated IRS-1 serine phosphorylation, increased insulin signaling to AKT and enhanced glucose utilization. In vivo, rapamycin treatment also ameliorated the impaired insulin signaling induced by inflammatory stress, but it induced pancreatic β-cell apoptosis, reduced pancreatic β-cell function and enhanced hepatic gluconeogenesis, thereby resulting in hyperglycemia and glucose intolerance in casein-injected mice. Our results indicate a paradoxical effect of rapamycin on insulin resistance between the in vitro and in vivo environments under inflammatory stress and provide additional insight into the clinical application of rapamycin

Hyperphosphatemia in chronic kidney disease exacerbates atherosclerosis via a mannosidases-mediated complex-type conversion of SCAP N-glycans

Blood phosphate levels are linked to atherosclerotic cardiovascular disease in patients with chronic kidney disease (CKD), but the molecular mechanisms remain unclear. Emerging studies indicate an involvement of hyperphosphatemia in CKD accelerated atherogenesis through disturbed cholesterol homeostasis. Here, we investigated a potential atherogenic role of high phosphate concentrations acting through aberrant activation of sterol regulatory element-binding protein (SREBP) and cleavage-activating protein (SCAP)-SREBP2 signaling in patients with CKD, hyperphosphatemic apolipoprotein E (ApoE) knockout mice, and cultured vascular smooth muscle cells. Hyperphosphatemia correlated positively with increased atherosclerotic cardiovascular disease risk in Chinese patients with CKD and severe atheromatous lesions in the aortas of ApoE knockout mice. Mice arteries had elevated SCAP levels with aberrantly activated SCAP-SREBP2 signaling. Excess phosphate in vitro raised the activity of α-mannosidase, resulting in delayed SCAP degradation through promoting complex-type conversion of SCAP N-glycans. The retention of SCAP enhanced transactivation of SREBP2 and expression of 3-hydroxy-3-methyl-glutaryl coenzyme A reductase, boosting intracellular cholesterol synthesis. Elevated α-mannosidase II activity was also observed in the aortas of ApoE knockout mice and the radial arteries of patients with uremia and hyperphosphatemia. High phosphate concentration in vitro elevated α-mannosidase II activity in the Golgi, enhanced complex-type conversion of SCAP N-glycans, thereby upregulating intracellular cholesterol synthesis. Thus, our studies explain how hyperphosphatemia independently accelerates atherosclerosis in CKD

CD36 and lipid metabolism in the evolution of atherosclerosis

Background: CD36 is a multi-functional class B scavenger receptor, which acts as an important modulator of lipid homeostasis and immune responses. Sources of data: This review uses academic articles. Areas of agreement: CD36 is closely related to the development and progression of atherosclerosis. Areas of controversy: Both persistent up-regulation of CD36 and deficiency of CD36 increase the risk for atherosclerosis. Abnormally up-regulated CD36 promotes inflammation, foam cell formation, endothelial apoptosis, macrophage trapping and thrombosis. However, CD36 deficiency also causes dyslipidemia, subclinical inflammation and metabolic disorders, which are established risk factors for atherosclerosis. Growing points: There may be an 'optimal protective window' of CD36 expression. Areas timely for developing research: In addition to traditionally modulating protein functions using gene overexpression or deficiency, the modulation of CD36 function at post-translational levels has recently been suggested to be a potential therapeutic strategy

The NIH-NIAID Filariasis Research Reagent Resource Center

Filarial worms cause a variety of tropical diseases in humans; however, they are difficult to study because they have complex life cycles that require arthropod intermediate hosts and mammalian definitive hosts. Research efforts in industrialized countries are further complicated by the fact that some filarial nematodes that cause disease in humans are restricted in host specificity to humans alone. This potentially makes the commitment to research difficult, expensive, and restrictive. Over 40 years ago, the United States National Institutes of Health–National Institute of Allergy and Infectious Diseases (NIH-NIAID) established a resource from which investigators could obtain various filarial parasite species and life cycle stages without having to expend the effort and funds necessary to maintain the entire life cycles in their own laboratories. This centralized resource (The Filariasis Research Reagent Resource Center, or FR3) translated into cost savings to both NIH-NIAID and to principal investigators by freeing up personnel costs on grants and allowing investigators to divert more funds to targeted research goals. Many investigators, especially those new to the field of tropical medicine, are unaware of the scope of materials and support provided by the FR3. This review is intended to provide a short history of the contract, brief descriptions of the fiilarial species and molecular resources provided, and an estimate of the impact the resource has had on the research community, and describes some new additions and potential benefits the resource center might have for the ever-changing research interests of investigators

Inflammatory Stress Sensitizes the Liver to Atorvastatin-Induced Injury in ApoE-/- Mice

Statins, which are revolutionized cholesterol-lowing agents, have been reported to have unfavorable effects on the liver. Inflammatory stress is a susceptibility factor for drug-induced liver injury. This study investigated whether inflammatory stress sensitized the liver to statin-induced toxicity in mice and explored the underlying mechanisms. We used casein injection in ApoE-/- mice to induce inflammatory stress. Half of the mice were orally administered atorvastatin (10mg/kg/d) for 8 weeks. The results showed that casein injection increased the levels of serum pro-inflammatory cytokines (IL-6 and TNFα). Atorvastatin treatment increased serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in casein injection mice. Moreover, atorvastatin treatment exacerbated hepatic steatosis, inflammation and fibrosis, as well as increased hepatic reactive oxygen species (ROS) and malondialdehyde in casein injection mice. However, above changes were not observed in atorvastatin treated alone mice. The protein expression of liver nuclear factor erythroid 2-related factor 2 (Nrf2) and the mRNA expressions of Nrf2 target genes were increased, together with the enhancement of activities of hepatic catalase and superoxide dismutase in atorvastatin treated alone mice, but these antioxidant responses were lost in mice treated with atorvastatin under inflammatory stress. This study demonstrates that atorvastatin exacerbates the liver injury under inflammatory stress, which may be associated with the loss of adaptive antioxidant response mediated by Nrf2

CD36 palmitoylation disrupts free fatty acid metabolism and promotes tissue inflammation in non-alcoholic steatohepatitis

BACKGROUND AND AIMS: Fatty acid translocase CD36 (CD36) is a membrane protein with multiple immuno-metabolic functions. Palmitoylation has been suggested to regulate the distribution and functions of CD36, but little is known about its significance in NASH. METHODS: Human liver tissue samples were obtained from patients undergoing liver biopsy for diagnostic purposes. CD36 knockout mice were injected with lentivirus vectors to express wild type CD36 and palmitoylation sites mutated CD36 in the livers. Liver histology, immunofluorescence, mRNA expression profile, subcellular distributions and functions of CD36 protein were assessed. RESULTS: The localization of CD36 on the plasma membrane of hepatocytes was markedly increased in patients with NASH compared to patients with normal liver and those with simple steatosis. Increased CD36 palmitoylation and increased localization of CD36 on the plasma membrane of hepatocytes were also observed in livers of mice with NASH. Furthermore, inhibition of CD36 palmitoylation protected mice from developing NASH. The absence of palmitoylation decreased CD36 protein hydrophobicity reducing its localization on the plasma membrane as well as in lipid raft of hepatocytes. Consequently, a lack of palmitoylation decreased fatty acid uptake and CD36/Fyn/Lyn complex in HepG2 cells. Inhibition of CD36 palmitoylation not only ameliorated intracellular lipid accumulation via activating the AMPK pathway, but also inhibited inflammatory response through the inhibition of the JNK signaling pathway. CONCLUSIONS: Our findings demonstrate the key role of palmitoylation in regulating CD36 distributions and its functions in NASH. Inhibition of CD36 palmitoylation may represent an effective therapeutic strategy in patients with NASH. LAY SUMMARY: Fatty acid translocase CD36 (CD36) is a multifunctional membrane protein which contributes to the development of liver steatosis. In the present study, we demonstrated that the localization of CD36 on the plasma membrane of hepatocytes is increased in patients with NASH. Blocking the palmitoylation of CD36 reduces CD36 distribution in hepatocytes plasma membrane and protects mice from NASH. The inhibition of CD36 palmitoylation not only improved fatty acid metabolic disorders but also reduced the inflammatory response in vitro and in vivo. The present study suggests that CD36 palmitoylation is important for NASH development and inhibition of CD36 palmitoylation could be used to cure NASH

CD36 plays a negative role in the regulation of lipophagy in hepatocytes through an AMPK-dependent pathway

Fatty acid translocase CD36 (CD36) is a multifunctional membrane protein that facilitates the uptake of long-chain fatty acid (LCFA). Lipophagy is autophagic degradation of lipid droplets. Accumulating evidence suggests that CD36 is involved in the regulation of intracellular signal transduction that modulates fatty acid storage or usage. However, little is known about the relationship between CD36 and lipophagy. In this study, we found that increasedCD36 expression was coupled with decreasedautophagy in the livers of mice treated with a high-fat diet. Overexpressing CD36 in HepG2 and Huh7 cells inhibited autophagy, while knocking down CD36 expression induced autophagy due to the increased autophagosome formation in autophagic flux. Meanwhile, knockout of CD36 in mice increased autophagy while reconstruction of CD36 expression in CD36-knockout mice reduced autophagy. CD36 knockdown in HepG2 cells increased lipophagy and β-oxidation, which contributed to improving lipid accumulation. In addition, CD36 expression regulated autophagy through the AMPK pathway with phosphorylation of ULK1/Beclin1 also involved in the process. These findings suggest that CD36 is a negative regulator of autophagy and induction of lipophagy by ameliorating CD36 expression can be a potential therapeutic strategy for the treatment of fatty liver diseases through attenuating lipid over-accumulation

CD36 deficiency aggravates macrophage infiltration and hepatic inflammation by up-regulating MCP-1 expression of hepatocytes through HDAC2-dependant pathway

AIMS: Cluster of differentiation 36 (CD36) is involved in the development of non-alcoholic steatohepatitis (NASH). Excess CD36 facilitates liver cells taking fatty acid and activates inflammatory signals to promote hepatic steatosis and inflammation. However, CD36-deficiency paradoxically promotes non-alcoholic fatty liver disease by unknown mechanisms. We explored the probable molecular mechanism of hepatic inflammation induced by CD36 deficiency. RESULTS: CD36 deletion in mice(CD36-/- mice) specifically increased MCP-1 in hepatocytes, promoted macrophage migration to liver and aggravated hepatic inflammatory response and fibrosis. The nuclear expression of histone deacetylase 2 (HDAC2) which highly expresses in wild-type hepatocytes and has an inhibitory effect on acetyl H3 was reduced in CD36 deficiency hepatocytes. Consequently, the level of acetyl H3 binding to MCP-1 promoters was increased in CD36 deficient hepatocytes, causing hepatic specific MCP-1 transcriptional activation. Reduction of nuclear HDAC2 in both CD36-/- mice liver and cultured hepatocytes was due to reduction of intracellular reactive oxygen species (ROS) level, while supplement of low concentration hydrogen peroxide (H2O2) overcame the suppression of HDAC2 caused by CD36 deficiency, decreasing MCP-1 gene transcription and microphage migration. INNOVATION: Our results provide first evidence that decreased ROS production by CD36 deletion was also harmful for livers. The fine balance of CD36 plays an important role in maintaining balances of hepatic ROS and nuclear HDAC2 which could be potential new therapeutic strategy for the prevention of NASH development. CONCLUSION: CD36 deficiency promoted the development of NASH by facilitating transcription of MCP-1 in hepatocytes, due to the reduction of ROS and nuclear HDAC2