Regulation Of Proprotein Convertase Research Articles

Far-infrared irradiation increases the uptake of LDL cholesterol by downregulating PCSK9 through the activation of TRPV4 calcium channels in HepG2 cells

This study investigated the effects of far-infrared (FIR) irradiation on low-density lipoprotein cholesterol (LDL-C) uptake by human hepatocellular carcinoma G2 (HepG2) cells via the regulation of proprotein convertase subtilisin/kexin type 9 (PCSK9). FIR irradiation for 30 min significantly decreased PCSK9 expression (p < 0.01) in HepG2 cells. FIR irradiation substantially increased the low-density lipoprotein receptor (p < 0.0001) and LDL-C uptake (p < 0.01). Activation of transient receptor potential vanilloid (TRPV) channels mimicked the effects of FIR irradiation, significantly decreasing the protein expression of PCSK9 (p < 0.05). Conversely, inhibition of TRP channels using ruthenium red reversed the reduction in PCSK9 protein expression following FIR irradiation (p < 0.01). The specific activation of TRPV4 using 4α-PDD mimicked the effect of FIR irradiation (p < 0.01), whereas PCSK9 reduction by FIR irradiation was significantly reversed by the inhibition of TRPV4 using RN1734 (p < 0.05). These findings implied that FIR irradiation emitted from a ceramic lamp specifically increased TRPV4 activity. These findings provide insights into a novel therapeutic approach using FIR irradiation for LDL-C regulation and its implications for cardiovascular health.

Role of miRNA-27a and miRNA-224 in posttranscriptional regulation of PCSK9 gene in Iraqi patient with coronary artery disease

AimThe aim of this study was to assess miRNA-27a and miRNA-224 expression levels in the blood samples from CAD patients and their role in the regulation of Proprotein Convertase Subtilisin/Kexin Type 9 Gene (PCSK9) expression. MethodsThis case-control study included (150) participants ranged in age from 30 to 65 years old, both male and female. Patients with atherosclerosis (n = 100) were chosen based on the clinical appearance of atherosclerotic coronary artery disease (ASCAD), as well as a medical and physical examination and documentation via coronary angiography, while the healthy group (n = 50) were chosen at random, and the health questionnaire and clinical examination revealed no signs of cardiovascular disease (CVD). For estimation of the expression levels of PCSK9, MiRNA-27a, and MiRNA-224 genes utilize reverse transcription-quantitative polymerase chain reaction (qRT-PCR) method was used. ResultsA current study found miRNA-27a gene expression level was 5.01 ± 0.75 in the ASCAD patients group based on 2-∆∆Ct and 1.00 ± 0.58 in the healthy control group, while miRNA-224 gene expression level of two study groups was 0.31 ± 0.15 vs 1.00 ± 0.58. In addition, analysis of PCSK9 expression level indicated that was 6.06 ± 1.84 in patient group and 1.00 ± 0.19 in healthy control group. ConclusionsThe fact that miRNA-27a and PCSK9 gene expression levels increased while miRNA-224 gene expression levels decreased indicated they have opposite activity in the development of CAD.

Loss of HNF1α Function Contributes to Hepatocyte Proliferation and Abnormal Cholesterol Metabolism via Downregulating miR-122: A Novel Mechanism of MODY3.

PurposeMutations in hepatocyte nuclear factor 1α (HNF1α) are the cause of maturity-onset diabetes of the young type 3 (MODY3) and involved in the development of hepatocellular adenoma and abnormal lipid metabolism. Previously, we have found that the serum microRNA (miR)-122 levels in MODY3 patients were lower than those in type 2 diabetes mellitus and healthy controls. This study aimed to investigate the mechanism of decreased miR-122 levels in patients with MODY3 and whether low levels of miR-122 mediate tumorigenesis and abnormal lipid metabolism associated with HNF1α deficiency in human hepatocytes.MethodsThe expression of miR-122 was examined by real-time PCR. Dual-luciferase reporter assay was performed to confirm the transcriptional regulation of miR-122 by HNF1α. HepG2 cells were transfected with siRNA or miRNA mimic to downregulate or upregulate the expression of HNF1α or miR-122, respectively. CCK-8 and colony formation assay were used to determine cell proliferation. Lipid accumulation was examined by Oil Red O staining and intracellular triglyceride and cholesterol quantification assays.ResultsHNF1α regulated the expression of miR-122 by directly binding to its promoter. Knockdown of HNF1α in HepG2 cells reduced the expression of miR-122, increased proliferation and promoted intracellular cholesterol accumulation. Overexpression of miR-122 partially rescued the phenotypes associated with HNF1α deficiency in human hepatocytes. Mechanistically, HNF1α modulated cholesterol homeostasis via miR-122-dependent activation of sterol regulatory element-binding protein-2 (SREBP-2) and regulation of proprotein convertase subtilisin/kexin type 9 (PCSK9). Moreover, circulating miR-122 levels were associated with serum cholesterol levels.ConclusionLoss of HNF1α function led to hepatocyte proliferation and abnormal cholesterol metabolism by downregulating miR-122. Our findings revealed a novel mechanism that low levels of miR-122 mediate tumorigenesis and abnormal lipid metabolism associated with MODY3. MiR-122 may be a potential therapeutic target for the treatment of MODY3.

Changes in circulating pro-protein convertase subtilisin/kexin type 9 levels - experimental and clinical approaches with lipid-lowering agents.

Regulation of pro-protein convertase subtilisin/kexin type 9 (PCSK9) by drugs has led to the development of a still small number of agents with powerful activity on low-density lipoprotein cholesterol levels, associated with a significant reduction of cardiovascular events in patients in secondary prevention. The Further Cardiovascular Outcomes Research with PCSK9 Inhibition in Subjects with Elevated Risk (FOURIER) and Evaluation of Cardiovascular Outcomes After an Acute Coronary Syndrome During Treatment With Alirocumab (ODYSSEY OUTCOMES) studies, with the two available PCSK9 antagonists, i.e. evolocumab and alirocumab, both reported a 15% reduction in major adverse cardiovascular events. Regulation of PCSK9 expression is dependent upon a number of factors, partly genetic and partly associated to a complex transcriptional system, mainly controlled by sterol regulatory element binding proteins. PCSK9 is further regulated by concomitant drug treatments, particularly by statins, enhancing PCSK9 secretion but decreasing its stimulatory phosphorylated form (S688). These complex transcriptional mechanisms lead to variable circulating levels making clinical measurements of plasma PCSK9 for cardiovascular risk assessment a debated matter. Determination of total PCSK9 levels may provide a diagnostic tool for explaining an apparent resistance to PCSK9 inhibitors, thus indicating the need for other approaches. Newer agents targeting PCSK9 are in clinical development with a major interest in those with a longer duration of action, e.g. RNA silencing, allowing optimal patient compliance. Interest has been expanded to areas not only limited to low-density lipoprotein cholesterol reduction but also investigating other non-lipid pathways raising cardiovascular risk, in particular inflammation associated to raised high-sensitivity C-reactive protein levels, not significantly affected by the present PCSK9 antagonists.

Black Raspberry Extract Enhances LDL Uptake in HepG2 Cells by Suppressing PCSK9 Expression to Upregulate LDLR Expression.

Black raspberry extract (BRE) has been widely used for treating prostate and urinary diseases and hyperlipidemia in Asia due to its significant lipid-lowering effects. The aim of this investigation was to evaluate the antihypercholesterolemia activity of BRE and the potential molecular mechanisms responsible for its antihypercholesterolemia activity by regulation of proprotein convertase subtilisin/kexin type 9 (PCSK9) expression in the human liver cell line HepG2. Reporter-based functional assay was used to identify herbal extracts that suppress PCSK9 expression in the HepG2 cells. Quantitative real-time polymerase chain reaction, Western blot analysis, and immunofluorescence staining were used to evaluate whether BRE modulates low-density lipoprotein receptor (LDLR) expression by repressing the hepatic expression of PCSK9. The LDLR activity of the HepG2 cells was determined using an LDL uptake assay. Our finding revealed that BRE modulates LDLR expression by suppressing the hepatic expression of PCSK9. We found that the combination of simvastatin and BRE caused the synergic induction of LDLR expression and LDL-C uptake, whereas simvastatin alone increased the expression of PCSK9 in the HepG2 cells. These results clearly demonstrated that the BRE from black raspberry suppressed simvastatin-induced PCSK9 expression and improved LDL-C uptake by hepatocytes through the induction of LDLR expression. These results suggest that the suppression of PCSK9 expression by BRE may potentiate the hypolipidemic effect of statins.

Role of Insulin in the Regulation of Proprotein Convertase Subtilisin/Kexin Type 9.

Proprotein convertase subtilisin/kexin type 9 (PCSK9), which binds the low-density lipoprotein receptor and targets it for degradation, has emerged as an important regulator of serum cholesterol levels and cardiovascular disease risk. Although much work is currently focused on developing therapies for inhibiting PCSK9, the endogenous regulation of PCSK9, particularly by insulin, remains unclear. The objective of these studies was to determine the effects of insulin on PCSK9 in vitro and in vivo. Using rat hepatoma cells and primary rat hepatocytes, we found that insulin increased PCSK9 expression and increased low-density lipoprotein receptor degradation in a PCSK9-dependent manner. In parallel, hepatic Pcsk9 mRNA and plasma PCSK9 protein levels were reduced by 55% to 75% in mice with liver-specific knockout of the insulin receptor; 75% to 88% in mice made insulin-deficient with streptozotocin; and 65% in ob/ob mice treated with antisense oligonucleotides against the insulin receptor. However, antisense oligonucleotide-mediated knockdown of insulin receptor in lean, wild-type mice had little effect. In addition, we found that fasting was able to reduce PCSK9 expression by 80% even in mice that lack hepatic insulin signaling. Taken together, these data indicate that although insulin induces PCSK9 expression, it is not the sole or even dominant regulator of PCSK9 under all conditions.

Recent advances in the understanding and care of familial hypercholesterolaemia: significance of the biology and therapeutic regulation of proprotein convertase subtilisin/kexin type9.

Familial hypercholesterolaemia (FH) is an autosomal co-dominant disorder that markedly raises plasma low-density lipoprotein-cholesterol (LDL-C) concentration, causing premature atherosclerotic coronary artery disease (CAD). FH has recently come under intense focus and, although there is general consensus in recent international guidelines regarding diagnosis and treatment, there is debate about the value of genetic studies. Genetic testing can be cost-effective as part of cascade screening in dedicated centres, but the full mutation spectrum responsible for FH has not been established in many populations, and its use in primary care is not at present logistically feasible. Whether using genetic testing or not, cholesterol screening of family members of index patients with an abnormally raised LDL-C must be used to determine the need for early treatment to prevent the development of CAD. The metabolic defects in FH extend beyond LDL, and may affect triacylglycerol-rich and high-density lipoproteins, lipoprotein(a) and oxidative stress. Achievement of the recommended targets for LDL-C with current treatments is difficult, but this may be resolved by new drug therapies. Lipoprotein apheresis remains an effective treatment for severe FH and, although expensive, it costs less than the two recently introduced orphan drugs (lomitapide and mipomersen) for homozygous FH. Recent advances in understanding of the biology of proprotein convertase subtilisin/kexin type 9 (PCSK9) have further elucidated the regulation of lipoprotein metabolism and led to new drugs for effectively treating hypercholesterolaemia in FH and related conditions, as well as for treating many patients with statin intolerance. The mechanisms of action of PCSK9 inhibitors on lipoprotein metabolism and atherosclerosis, as well as their impact on cardiovascular outcomes and cost-effectiveness, remain to be established.

Expression and regulation of proprotein convertase subtilisin/kexin type 9 in vascular smooth muscle cells

Expression and regulation of proprotein convertase subtilisin/kexin type 9 in vascular smooth muscle cells

Pituitary protein 7B2 plasma levels in patients with liver disease: Comparisons with other hormones and neuropeptides

7B2, a protein initially isolated from the porcine pituitary gland, has been identified in numerous animal and human tissues, with the highest concentrations in the pituitary and hypothalamus. The 7B2 molecule is highly evolutionarily conserved and is considered to be indispensable in the function and regulation of proprotein convertase 2 (PC2). In the present study, the plasma 7B2 immunoreactivity (7B2-IR) of 18 patients with liver disease was studied. Of these patients, seven (three male and four female), aged 37–67 [54.6±13.5 (SD)] years, suffered from liver cirrhosis of cryptogenic (n=2) or alcoholic (n=5) aetiology. The remaining 11 patients (four male and seven female), aged 22–76 [56.1±17.6 (SD)] years, suffered from miscellaneous liver abnormalities. The clinical diagnosis was confirmed in the majority of patients by the histological examination of a percutaneous liver biopsy or by appropriate radiological investigations. Plasma bilirubin, alkaline phosphatase, aspartate aminotransferase, albumin, prothrombin time, electrolytes, urea and creatinine were measured. The plasma 7B2-IR levels were estimated using a sensitive radioimmunoassay (RIA), and the elution position of 7B2-IR was verified by gel chromatography. The mean plasma 7B2-IR concentration in patients with liver disease was 99.44±15.9 pmol/l. In the patients with hepatocellular damage due to metastatic tumours [Ca bronchus, carcinoid (n=6)], the 7B2-IR concentrations were significantly higher [185±36.9 pmol/l, (P<0.05)] compared with the overall subjects with liver damage. The results of the present study demonstrate that 7B2-IR is increased in liver disease, with the highest levels detected in patients with tumourous liver conditions.

The Localization and Regulation of Proprotein Convertase Subtilisin/Kexin (PCSK) 6 in Human Ovary

The aim of this study is to evaluate the expression and regulation of proprotein convertase subtilisin/kexin (PCSK) 6, which is known to be an important factor in the production of bone morphogenetic protein (BMP) cytokines in human ovary. The localization of PCSK 6 protein in normal human ovaries was examined by immunohistochemistry. Human granulosa cells (GC), obtained from 34 patients undergoing ovarian stimulation for in vitro fertilization, were cultured with BMP-2, BMP-6, BMP-7, BMP-15, growth differentiation factor (GDF)-9, and activin-A with or without FSH. PCSK 6 mRNA expression level was evaluated by quantitative real-time reverse transcription and polymerase chain reaction (RT-PCR). An immunohistochemistry study revealed that GC expressed PCSK 6 throughout follicular development, beginning in the primary follicle stage, while oocytes expressed PCSK 6 from the primordial follicle stage onwards. An in vitro study demonstrated that BMP-2, BMP-6, BMP-7, and BMP-15, not activin-A and GDF-9, decreased PCSK 6 gene expression in human GC. FSH induced PCSK 6 mRNA in the presence of activin-A or GDF-9. GDF-3, which is an inhibitor of BMP cytokines, also induced PCSK 6 mRNA expression. PCSK 6, which is a critical factor to produce BMP cytokines, was suppressed with BMP stimulation in human GC, suggesting the presence of a negative feedback system in the follicular development process.