Functional analysis of PCSK9 3'UTR variants and mRNA-miRNA interactions in patients with familial hypercholesterolemia.

Aim: Functional analysis of PCSK9 3'UTR variants and mRNA-miRNA interactions were explored in patients with familial hypercholesterolemia (FH). Materials & methods:PCSK9 3'UTR variants were identified by exon-targeted gene sequencing. Functional effects of 3'UTR variants and mRNA-miRNA interactions were analyzed using in silico and in vitro studies in HEK293FT and HepG2 cells. Results: Twelve PCSK9 3'UTR variants were detected in 88 FH patients. c.*75C >T and c.*345C >T disrupted interactions with miR-6875, miR-4721 and miR-564. Transient transfection of the c.*345C >T decreased luciferase activity in HEK293FT cells. miR-4721 and miR-564 mimics reduced PCSK9 expression in HepG2 cells. Conclusion:PCSK9 c.*345C >T has a possible role as loss-of-function variant. miR-4721 and miR-564 downregulate PCSK9 and may be useful to improve lipid profile in FH patients.

High serum miR-421 is associated with metabolic dysregulation and inflammation in patients with metabolic syndrome.

Aim: To explore the association of circulating miRNAs with adiposity, metabolic status and inflammatory biomarkers in patients with metabolic syndrome (MetS). Methods: Serum levels of 372 miRNAs were measured in patients with (n = 6) and without MetS (n = 6) by quantitative PCR array, and dysregulated miRNAs were validated in a larger cohort (MetS, n = 89; non-MetS, n = 144). Results: In the screening study, seven miRNAs were dysregulated in patients with MetS, and miR-421 remained increased in the validation study. miR-421 was associated with a high risk of MetS and insulin resistance and hypertension and correlated with glycated hemoglobin, triacylglycerols, high-sensitivity CRP, IL-6, resistin and adiponectin (p < 0.05). Conclusion: Circulating miR-421 is a potential biomarker for insulin resistance, metabolic dysregulation and inflammatory status in patients with MetS.

High serum miR-421 is associated with metabolic dysregulation and inflammation in patients with metabolic syndrome

ABSTRACT: To explore the association of circulating miRNAs with adiposity, metabolic status and inflammatory biomarkers in patients with metabolic syndrome (MetS). METHODS: Serum levels of 372 miRNAs were measured in patients with (n = 6) and without MetS (n = 6) by quantitative PCR array, and dysregulated miRNAs were validated in a larger cohort (MetS, n = 89; non-MetS, n = 144). RESULTS: In the screening study, seven miRNAs were dysregulated in patients with MetS, and miR-421 remained increased in the validation study. miR-421 was associated with a high risk of MetS and insulin resistance and hypertension and correlated with glycated hemoglobin, triacylglycerols, high-sensitivity CRP, IL-6, resistin and adiponectin (p < 0.05). CONCLUSION: Circulating miR-421 is a potential biomarker for insulin resistance, metabolic dysregulation and inflammatory status in patients with MetS.

Association of the PPP3CA c.249G>A variant with clinical outcomes of tacrolimus-based therapy in kidney transplant recipients.

BACKGROUND: The effects of genetic variants related to the pharmacodynamic mechanisms of immunosuppressive drugs on their therapeutic efficacy and safety have been poorly explored. This study was performed to investigate the influence of the PPP3CA c.249G>A variant on the clinical outcomes of kidney transplant recipients. PATIENTS AND METHODS: A total of 148 Brazilian patients received tacrolimus (TAC)-based immunosuppressive therapy for 90 days post-kidney transplantation. The PPP3CA rs3730251 (c.249G>A) polymorphism was determined by real-time polymerase chain reaction. Single-nucleotide polymorphism (SNP) data for CYP3A5 rs776746 (CYP3A5*3C; g.6986A>G) were used to eliminate the confounding effects of this variant. RESULTS: The PPP3CA c.249G>A SNP did not influence early TAC exposure, renal function, or other laboratory parameters, including levels of urea, creatinine, glucose, and lipids, and blood counts. This variant also did not account for the cumulative incidence of biopsy-confirmed acute rejection or delayed graft function. Regarding adverse events, PPP3CA c.249A allele carriers initially had a 3.05-fold increased probability of treatment-induced blood and lymphatic system disorders compared with c.249GG genotype individuals (95% confidence interval: 1.10-8.48, p=0.032). However, this result was not maintained after adjusting for body weight and CYP3A5*3C SNP status (p=0.086). CONCLUSION: The PPP3CA c.249G>A variant does not influence the clinical outcomes of Brazilian patients in the early phase of TAC-based immunosuppressive regimen.

Association of the PPP3CA c.249G>A variant with clinical outcomes of tacrolimus-based therapy in kidney transplant recipients

BACKGROUND:The effects of genetic variants related to the pharmacodynamic mechanisms of immunosuppressive drugs on their therapeutic efficacy and safety have been poorly explored. This study was performed to investigate the influence of the PPP3CA c.249G>A variant on the clinical outcomes of kidney transplant recipients.PATIENTS AND METHODS:A total of 148 Brazilian patients received tacrolimus (TAC)-based immunosuppressive therapy for 90 days post-kidney transplantation. The PPP3CA rs3730251 (c.249G>A) polymorphism was determined by real-time polymerase chain reaction. Single-nucleotide polymorphism (SNP) data for CYP3A5 rs776746 (CYP3A5*3C; g.6986A>G) were used to eliminate the confounding effects of this variant.

Altered microRNome Profiling in Statin-Induced HepG2 Cells: A Pilot Study Identifying Potential new Biomarkers Involved in Lipid-Lowering Treatment

PURPOSE:Statins are widely prescribed drugs to manage hypercholesterolemia. Despite they are considered effective lipid-lowering agents, significant inter-individual variability has been reported in relation to drug response. Among the reasons explaining this variation, genetic factors are known to partially contribute. Nonetheless, poor evidence exists regarding epigenetic factors involved.METHODS:We investigated if atorvastatin can modulate the cholesterol related miR-33 family. Furthermore, we analyzed the microRNA expression profiles in HepG2 cells treated for 24 hours with atorvastatin or simvastatin using a microarray platform.RESULTS:Our results indicate that atorvastatin does not influence the expression of the miR-33 family. In addition, microarray examination revealed that atorvastatin modulated thirteen miRs, whilst simvastatin only affected two miRs. All significantly modulated miRs after simvastastin therapy were also modulated by atorvastatin. In addition, four novel miRs with previously unreported functions were identified as statin-modulated...

Polymorphisms in genes encoding metalloproteinase 9 and Lymphotoxin-Alpha can influence warfarin treatment

Objectives: Warfarin treatment is influenced by environmental and genetic factors. The influence of polymorphisms in genesencoding metalloproteinase 9 (MMP9), lymphotoxin-alpha (LTA) andTNFSF14 (LIGHT), related to the inflammatory process ofcoronary artery disease, on warfarin dose and time to reach target was investigated in this study.Methods: Outpatients on warfarin treatment (n=227), 20 to 92 years, were enrolled at the Institute Dante Pazzanese of Cardiology(IDPC). Genomic DNA was obtained from peripheral whole blood to evaluate MMP9 rs17576 (Gln279Arg, A>G), LTA rs1041981(Thr60Asn, C>A) and rs909253 (c.252T>C) and TNFSF14rs2291668 (c.147C>T) and rs344560 (Lys214Glu, G>A) polymorphismsby pyrosequencing in Q24PyroMark.Results: The patients carrying MMP9 rs17576GG genotype were more likely to require a lower warfarin weekly dose (OR:2.73, 95% CI: 1.01-7.41, p=0.048). Also, LTA rs909253 variant was associated with a longer time to reach the target internationalnormalized ratio (INR) (OR: 1.98, 95% CI: 1.02-3.86, p=0.043). Age was inversely correlated with the target INR (r=-0.387, p<0.001),and dose was directly correlated with time to reach target INR (r=0.244, p<0.001).Conclusion: MMP9 rs17576 variant may have an important influence on warfarin weekly dose, and that LTA rs909253polymorphism may also influence the time to reach the target INR.

Apolipoprotein E mRNA expression in mononuclear cells from normolipidemic and hypercholesterolemic individuals treated with atorvastatin

Background Apolipoprotein E (apoE) is a key component of the lipid metabolism. Polymorphisms at the apoE gene (APOE) have been associated with cardiovascular disease, lipid levels and lipid-lowering response to statins. We evaluated the effects on APOE expression of hypercholesterolemia, APOE å2/å3/å4 genotypes and atorvastatin treatment in Brazilian individuals. The relationship of APOE genotypes and plasma lipids and atorvastatin response was also tested in this population.MethodsAPOE å2/å3/å4 and plasma lipids were evaluated in 181 normolipidemic (NL) and 181 hypercholesterolemic (HC) subjects. HC individuals with indication for lowering-cholesterol treatment (n = 141) were treated with atorvastatin (10 mg/day/4-weeks). APOE genotypes and APOE mRNA in peripheral blood mononuclear cells (PBMC) were analyzed by TaqMan real time PCR.ResultsHC had lower APOE expression than NL group (p < 0.05) and individuals with low APOE expression showed higher plasma total and LDL cholesterol and apoB, as well as higher apoAI (p < 0.05). Individuals carrying å2 allele have reduced risk for hypercholesterolemia (OR: 0.27, 95% I.C.: 0.08-0.85, p < 0.05) and NL å2 carriers had lower total and LDL cholesterol and apoB levels, and higher HDL cholesterol than non-carriers (p < 0.05). APOE genotypes did not affect APOE expression and atorvastatin response. Atorvastatin treatment do not modify APOE expression, however those individuals without LDL cholesterol goal achievement after atorvastatin treatment according to the IV Brazilian Guidelines for Dyslipidemia and Atherosclerosis Prevention had lower APOE expression than patients with desirable response after the treatment (p < 0.05).Conclusions...

Influence of SCARB1 polymorphisms on serum lipids of hypercholesterolemicindividuals treated with atorvastatin

Background: The SR-BI is a key component on the cholesterol metabolism. Polymorphisms in the SR-BI gene (SCARB1) were related with variations on plasma lipoprotein profile and other risk factors for cardiovasculardisease. We tested the relationship of 3 SCARB1 single nucleotide polymorphisms (SNPs) with hypercholesterolemia in a Brazilian population and whether these variants can influence lipid-lowering responseto atorvastatin. Methods: c.4GNA, c.726+54CNT and c.1050CNT SNPs and serum concentrations of lipid and apolipoproteins were evaluated in 147 hypercholesterolemic (HC) and 185 normolipidemic (NL) unrelated Brazilian subjects. HC patients were treated with atorvastatin (10 mg/day/4 weeks).Results: Frequencies of SCARB1 polymorphisms were similar between the HC and NL groups (pN0.05). TheT allele for c.726+54CNT was associated with higher LDL-c in NL and with higher apoB and apoB/apoAI inHC (pb0.05). HC individuals carrying c.1050C allele carriers (CC and CT genotypes) had lower change of totalcholesterol, LDL-c, apoB and apoB/apoAI ratio (pb0.05) than the TT genotype carriers in response to atorvastatin. Conclusion: The SCARB1 polymorphisms are relatedwith variations in serumlipids in the Brazilian population and c.1050CNT SNP is associated with lipid-lowering atorvastatin response.

ABCB1 and ABCC1 expression in peripheral mononuclear cells is influenced by gene polymorphisms and atorvastatin treatment

This study investigated the effects of atorvastatin on ABCB1 and ABCC1 mRNA expression on peripheral blood mononuclear cells (PBMC) and their relationship with gene polymorphismsand lowering-cholesterol response. One hundred and thirty-six individuals withhypercholesterolemia were selected and treated with atorvastatin (10 mg/day/4 weeks). Blood samples were collected for serum lipids and apolipoproteins measurements and DNA and RNA extraction. ABCB1 (C3435T and G2677T/A) and ABCC1 (G2012T) gene polymorphisms were identified by polymerase chain reaction-restriction (PCR)-RFLP and mRNA expression was measured in peripheral blood mononuclear cells by singleplex real-time PCR. ABCB1 polymorphisms were associated with risk for coronary artery disease (CAD) ( p 0.085: LDL-c = 40.3 14.3%; apoB = 32.5 10.7%; p < 0.05). ABCB1 substrates or inhibitors did not affect the baseline expression, while ABCB1 inhibitors reversedthe effects of atorvastatin on both ABCB1 and ABCC1 transporters. In conclusion, ABCB1 and ABCC1 mRNA levels in PBMC are modulated by atorvastatin and ABCB1 G2677T/A polymorphism and ABCB1 baseline expression is related to differences in serum LDL cholesterol and apoB in response to atorvastatin.