Inflammatory cytokines disrupt LDL-receptor feedback regulation and cause statin resistance: a comparative study in human hepatic cells and mesangial cells (original) (raw)
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Effects of different LDL particles on inflammatory molecules in human mesangial cells
Diabetologia, 2008
Aims/hypothesis Inflammation is a mechanism of glomerular damage in chronic glomerulopathies. LDL may increase the production of inflammatory cytokines in renal tissues. However, the relative role of native, oxidised and glycated LDL in promoting this process has been only partially elucidated. Methods We tested the inflammatory and proapoptotic effects of native, oxidised and glycated LDL in human mesangial cells (HMCs) by measuring levels of IL6, CD40 and macrophage migration inhibitory factor (MIF) genes, MIF protein, release of IL6, soluble CD40, fibronectin and laminin, early and late apoptosis, and extracellular regulated kinases (ERK) 1/2 and c-Jun N-terminal kinase (JNK) activation. Results IL6 and CD40 mRNA were dose-dependently upregulated by all three species; this was closely paralleled by their increased release. MIF mRNA was potently stimulated by modified LDL, as confirmed by immunostaining. Fibronectin and laminin release was stimulated by both oxidised and glycated, but not native, LDL. All LDL species induced some increase in late, but not early, apoptosis, and similarly activated JNK2/3 phosphorylation; in contrast, ERK1/2 phosphorylation was more strongly upregulated by oxidised than either native or glycated LDL.
Cytokines regulation of low-density lipoprotein receptor gene transcription in human mesangial cells
Nephrology Dialysis Transplantation, 1998
scription; human mesangial cell; LDL receptor; LDL receptor gene promoter Background. The intracellular transport of lipids through regulation of the LDL receptor (LDLr) may be important in the progression of renal dysfunction. The present study was undertaken to investigate Introduction whether cytokines have any major effects on LDLr regulation and lipid-mediated glomerular injury in In recent years a number of studies have shown that human mesangial cells (HMC). the histologic features of glomerulosclerosis are similar Methods. We explored the effects of 50 ng/ml of to the changes observed in atherosclerosis, and the tumour necrosis factor a (TNFa), 5 ng/ml of transterm 'glomerular atherosclerosis' has been proposed forming growth factor b (TGFb), platelet-derived [1,2]. The similarities between the two include abnorgrowth factor (PDGF), and interleukin-1b (IL-1b) on mal lipid metabolism, initial endothelial injury, subthe regulation of LDLr gene transcription in a human sequent mesangial or smooth muscle cell proliferation mesangial cell line (HMCL) using cell proliferation, and expansion of the extracellular matrix [2]. There LDL binding, northern blot and LDLr promoter are significant abnormalities in lipid metabolism in activity assays. renal disease, renal failure and renal transplant Results. TNFa, TGFb, PDGF or IL-1b did not patients. Lipid deposition can be found in mesangial significantly stimulate HMCL proliferation at the cells and extracellularly in mesangial matrix in concentrations given above, but maximally stimulated advanced glomerular lesions. There is increasing evid-LDLr mRNA expression and increased LDLr proence that mesangial lipid accumulation is an important moter activity by 167.48±23.56%, 150.47±24.41%, risk factor for glomerulosclerosis. While hyperlipidae-127.71±24.65% and 163.01±31.91% respectively, at mia is often striking in the majority of nephrotic 24 h. An increased LDL binding was observed in patients, plasma lipids are normal in some patients parallel with increased LDLr mRNA. The tyrowith progressive renal disease. This suggests that sine kinase transduction pathway was involved in intracellular transport of lipids through regulation of LDLr upregulation induced by all four cytokines. the LDL receptor (LDLr) may be important in the Additionally, TGFb involved serine/threonine kinase progression of renal dysfunction. and G-protein pathways, and IL-1b involved calmodu-The LDLr is the primary receptor for the binding lin, serine/threonine kinase and PKC pathways in and internalization of plasma-derived LDL-cholesterol upregulating LDLr. A high concentration of LDL and thereby regulates plasma LDL [3,4]. Expression (250 mg/ml) inhibited promoter activity, but TNFa, of the LDLr gene may be regulated by factors such as TGFb, PDGF and IL-1b co-incubated with LDL could intracellular levels of cholesterol, oxysterols and several override transcriptional inhibition by LDL. growth factors [5-8]. Recently, it has been shown Conclusion. TNFa, TGFb, PDGF and IL-1b increased that tumour necrosis factor a (TNFa), transforming LDLr gene expression by increasing sterol-independent growth factor b (TGFb), platelet-derived growth and mitogenesis-independent gene transcription. This factor (PDGF), fibroblast growth factor (FGF), insuprocess may contribute to lipid deposition and foam lin, oncostatin M and interleukin 1b (IL-1b) promoted cell formation in HMC. LDL binding in HepG2 and fibroblasts, presumably as a consequence of their mitogenic activity. Cell Key words: cytokine; glomerulosclerosis; gene trandivision increases the cellular demand for cholesterol, a major component in membrane biosynthesis [9-11]. The LDLr is present on human mesangial cells (HMC)
Cardiovascular Research, 2007
In the extracellular intima, extracellular matrix proteoglycans favor LDL retention and aggregation (agLDL). In contrast to native LDL (nLDL), agLDL induces high intracellular cholesteryl ester (CE) accumulation in macrophages. It has been suggested that LDL receptor-related protein (LRP1) is involved in agLDL binding and internalization by macrophages. The aim of this work was to analyze whether sterol regulatory element binding proteins (SREBPs) modulate LRP1 expression and LRP1-mediated agLDL uptake by human monocyte-derived macrophages (HMDM). Methods and results: The treatment of HMDM with small anti-LRP1 interfering RNA (siRNA-LRP1) led to the specific inhibition of LRP1 mRNA expression and also to the inhibition of LRP1 protein expression in these cells. In siRNA-LRP1-treated HMDM, CE accumulation from agLDL uptake (84.66 ± 5 μg CE/mg protein) was reduced by 95.76 ± 5.22%. This suggests that LRP1 plays a pivotal role in agLDL uptake by HMDM. N-acetyl-leucyl-leucyl-norleucinal (ALLN), an inhibitor of SREBP catabolism, maintained high levels of active SREBP-2 and SREBP-1 even in the presence of nLDL and agLDL. Therefore, ALLN induced LDL receptor (LDLR) upregulation. Concomitantly, a strong downregulation of LRP1 mRNA and LRP1 protein was observed in ALLN-treated macrophages. By decreasing LRP1 expression levels, ALLN reduced CE accumulation from agLDL at all tested concentrations. Conclusions: These results suggest that high levels of active SREBPs downregulate LRP1 expression and intracellular CE accumulation in HMDM.
Sterol dependent LDL-receptor gene transcription in lymphocytes from normal and CML patients
Cancer Letters, 1996
Sterol regulatory element (SRE) has been recognized to regulate various key genes coding for especially low density lipoprotein (LDL)-receptor, 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase and HMG-CoA synthase known to play a crucial role in the cholesterol feedback mechanism. The deranged cholesterol feedback mechanism has been widely recognised in initiation as well as progression of various types of cancers including chronic myeloid leukaemia (CML). Consequently, the present study was addressed to understand this phenomenon and revealed the existence of a unique 47 kDa protein factor having affinity for this SRE sequence in lymphocytes from normal subjects as well as its absence in lymphocytes from untreated CML patients. However, this factor appeared when the CML patients achieved complete haematological remission (CHR) through a-interferon therapy. Further, an inverse relationship was also observed between sterol modulated LDL-receptor gene transcription and the binding affinity of this 47 kDa factor to the SRE sequence. Based upon these results we propose that o-interferon through its receptor initiates phosphatidic acid dependent signalling which in turn regulates the affinity of 47 kDa sterol regulatory element binding factor as well as LDL-receptor gene transcription in lymphocytes from CML patients.
The Journal of Lipid Research, 2014
The expression of liver LDL receptor (LDLR) regulates human plasma LDL-cholesterol (LDL-C) homeostasis ( 1-3 ). Increased hepatic LDLR expression results in improved clearance of plasma LDL-C through receptor-mediated endocytosis, which is strongly associated with a decreased risk of developing cardiovascular disease in humans ( 4, 5 ). Thus far, many studies have demonstrated that intracellular cholesterol levels play a primary role in determination of hepatic LDLR expression levels through a negative feedback mechanism that controls gene transcription mediated by the sterolregulatory element (SRE) located in LDLR promoter and SRE binding proteins (SREBPs) ( 6, 7 ).
Archives of Biochemistry and Biophysics, 1997
noblotting; transcription; cholesterol; lovastatin; sterol response element binding proteins. The mechanism by which dietary cholesterol regulates expression of the hepatic low-density lipoprotein (LDL) receptor was investigated. In a previous study (Arch. Biochem. Biophys. 325, 242-248, 1996), we dem-Regulation of low-density lipoprotein (LDL) 2 receptor onstrated that dietary cholesterol reduces the rate of gene expression by cholesterol is thought to occur ex-LDL receptor protein degradation without affecting clusively at the transcriptional level (1). The LDL resteady-state levels of receptor protein. In view of these findings, it was expected that dietary cholesterol ceptor provides cholesterol to cells by binding and interwould decrease the rate of transcription of the hepatic nalizing LDL. When cholesterol demands are high, the LDL receptor gene, resulting in lower mRNA levels LDL receptor gene is transcribed actively, and conseand lower rates of synthesis of LDL receptor protein. quently more LDL is internalized. However, when cho-Surprisingly, neither the rate of transcription nor the lesterol accumulates within the cell, transcription of level of LDL receptor mRNA was reduced in response this gene is repressed causing a decrease in LDL interto dietary cholesterol, even though hepatic cholesterol nalization. levels were increased twofold. This suggests that un-
Journal of Clinical Investigation, 1994
The liver plays a key regulatory role in cholesterol metabolism. Two proteins are central in this role; the LDL receptor and 3-hydroxy-3-methylglutaryl CoA reductase (HMG CoA reductase), the rate-limiting enzyme in cholesterol biosynthesis. In the current investigation, we have used a sensitive nonradioactive method to study the regulation of LDL receptor and HMG CoA reductase mRNA levels in liver biopsy samples and freshly isolated mononuclear leukocytes from 13 patients who underwent cholecystectomy for gallstones. mRNA copy numbers were determined by PCR amplification of reversetranscribed RNA using synthetic RNA as an internal standard. Incorporation of digoxigenin-11-dUTP during amplification allowed direct detection and quantitation of mRNA levels by chemiluminescence. These experiments showed that the average number of LDL receptor mRNA molecules in liver (21±3 X 104/,gg of RNA) and mononuclear leukocytes (24±3 X 104/ ,ug of RNA) are indistinguishable, whereas the number of HMG CoA reductase molecules in liver (107±15 X 10'/,ug of RNA) is smaller than that in mononuclear leukocytes (158±21 x 104/,g of RNA, P < 0.05). These numbers correspond to an average of 1-6 copies of LDL receptor mRNA and 542 copies of HMG CoA reductase mRNA per cell. There was a significant correlation between the numbers of LDL receptor (P = 0.0005) and HMG CoA reductase (P = 0.003) mRNA molecules in liver and mononuclear leukocytes. Furthermore, the numbers of copies of HMG CoA reductase and LDL receptor mRNA were correlated with each other in both liver (P = 0.02) and mononuclear leukocytes (P = 0.01), consistent with coordinate regulation. These data demonstrate that the mechanisms which regulate mRNA levels in liver and mononuclear cells are similar and suggest that freshly isolated mononuclear cells can be used to predict HMG CoA reductase and LDL receptor mRNA levels in liver. (
Atherosclerosis, 2005
Hepatic lipase (HL) not only plays an important role in plasma lipoprotein transport, but may also affect intracellular lipid metabolism. We hypothesize that HL expression is regulated as an integral part of intracellular lipid homeostasis. Addition of oleate (1 mM) to HepG2 cells increased HL secretion to 134 ± 14% of control (p < 0.02), and increased the transcriptional activity of a 698-bp HL promoter-reporter construct two-fold. Atorvastatin (10 M) abolished the oleate stimulation. The transcriptional activity of a sterol-regulatory-element binding protein (SREBP)-sensitive HMG-CoA synthase promoter construct was reduced 50% by oleate, and increased 2-3-fold by atorvastatin. Cotransfection with an SREBP-2 expression vector reduced HL promoter activity and increased HMG-CoA synthase promoter activity. Upstream stimulatory factors (USF) are also implicated in maintenance of lipid homeostasis. Co-transfection with a USF-1 expression vector stimulated HL promoter activity 4-6-fold. The USF-stimulated HL promoter activity was not further enhanced by oleate, but almost completely prevented by atorvastatin or co-transfection with the SREBP-2 vector. Opposite regulation by USF-1 and SREBP-2 was also observed with a 318-bp HL promoter construct that lacks potential SRE-like and E-box binding motifs. We conclude that the opposite regulation of HL expression by fatty acids and statins is mediated via SREBP, possibly through interaction with USF.
Arteriosclerosis, Thrombosis, and Vascular Biology, 2013
Objective— The risk of cardiovascular disease is increased by up to 33 to 50× in chronic inflammatory states and convention doses of statins may not provide the same cardiovascular protection as in noninflamed patients. This study investigated whether the increase in 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCoA-R)–mediated cholesterol synthesis observed under inflammatory stress was resistant to the action of statins and if so, whether this was because of interference with the sterol regulatory element binding protein cleavage–activating protein pathway. Approach and Results— Inflammatory stress was induced by adding cytokines (interleukin-1β, tumor necrosis factor-α, and interleukin-6) and lipopolysaccharides to vascular smooth muscle cells in vitro and by subcutaneous casein injection in apolipoprotein E/scavenger receptors class A/CD36 triple knockout mice in vivo. Inflammatory stress exacerbated cholesterol ester accumulation and was accompanied in vitro and in vivo by incre...
Biochimica et Biophysica Acta (BBA) - Molecular and Cell Biology of Lipids, 2006
Conjugated linoleic acids (CLA) have attracted scientific interest due to their potential beneficial effects on atherosclerosis. Recently, a mixture of CLA isomers was demonstrated to upregulate LDL receptor expression in the human hepatoma cell line HepG2. However, the underlying mechanisms remain to be resolved. Thus, the aim of this study was to elucidate how CLA mediates upregulation of LDL receptor in HepG2 cells and whether this upregulation is isomer-specific. The results revealed that LDL receptor promoter activity and mRNA expression were strongly induced upon treatment with t10c12-CLA (P < 0.05), whereas c9t11-CLA and linoleic acid (LA) had no effect. In addition, only treatment with t10c12-CLA markedly induced mRNA expression of SREBP-2 and HMG-CoA reductase and slightly induced that of SREBP-1 (P < 0.05). Using SREBP-2 knockdown cells, we could demonstrate that the effect of t10c12-CLA on LDL receptor gene transcription was significantly reduced when compared to control cells (P < 0.05). When using SREBP-1 knockdown cells the effect of t10c12-CLA on LDL receptor mRNA only slightly decreased compared to control cells. In addition, using different deletion constructs of the LDL receptor gene promoter we showed that the induction of the LDL receptor by t10c12-CLA is independent of the AP-1 motif in the LDL receptor promoter. In conclusion, the present study revealed that transcriptional activation of the LDL receptor gene by t10c12-CLA is dependent on the upregulation of SREBP-2 and is probably due to the activation of the SRE-1 in the LDL receptor gene promoter in HepG2 cells. Thus, the decreased plasma cholesterol levels in response to CLA as observed in a limited number of animal and human studies might be explained by an enhanced uptake of VLDL and LDL cholesterol via hepatic LDL receptors. However, it provides no explanation for the outcome of most human studies reporting unaltered or even increased plasma and LDL cholesterol concentrations in response to supplementation with CLA.