Toll-like receptor inflammatory cascade and the development of diabetic kidney disease in children and adolescents with type 1 diabetes.

AIM: This study aimed to evaluate the association of toll-like receptor (TLR) inflammatory cascade with the development of diabetic kidney disease (DKD) in children and adolescents with type 1 diabetes (T1D). METHODS: A total of 49 T1D patients and 49 normoglycaemic (NG) subjects aged 5-20 years old were recruited. TLR2, TLR4, MYD88, NFKB, MCP1/CCL2 and IL18 mRNA expressions were measured in peripheral blood mononuclear cells by reverse transcription-quantitative polymerase chain reaction. Fasting glucose, glycated haemoglobin, serum urea, serum creatinine and urinary albumin-to-creatinine ratio (ACR) were determined. RESULTS: The mRNA expressions of TLR2, TLR4, MYD88 and NFKB were significantly increased in the T1D group compared with the NG group. The mRNA expression levels of MCP1/CCL2 and IL18 were higher in 21 T1D patients (42.9%) (average of MCP1/CCL2: 6.6-fold and IL18: 5.8-fold) than in NG patients. Furthermore, ACR was increased in the T1D group compared with the NG group. CONCLUSION: The increased mRNA expression of TLR2, TLR4, MYD88, NFKB, MCP1/CCL2 and IL18 favours the development of an inflammatory process that may lead to a decline in renal function and consequently DKD in children and adolescents with T1D. This suggests that these genes are early mediators of onset DKD since the beginning of the lives of the paediatric T1D patients.

Functional analysis of PCSK9 3'UTR variants and mRNA­miRNA 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.

Statins differentially modulate microRNAs expression in peripheral cells of hyperlipidemic subjects: A pilot study.

AIM: Although statins are considered a cornerstone for the treatment of high cholesterol levels due to their powerful cholesterol-lowering effects, response to drug administration is still one of the main pitfalls of statin treatment. So far, the reasons underlying this undesired outcome are still poorly understood, but recently, various studies have suggested that miRNAs may be involved. Therefore, we aimed at evaluating the effect of short-term low-dose treatment with 2 statins on miRNAs expression in patients with hypercholesterolemia. METHODS: A total of 40 hypercholesterolemic (HC) subjects following 1 month of atorvastatin (10 mg/day; n = 20) or simvastatin (10 mg/day; n = 20) were included. Multiple available boinformatic algorithms (TargetScan, miRanda, DianaLab, MicroCosm and PicTar) were employed to select miRNAs regulating genes involved in cholesterol metabolism and statin response. Differential miRNAs expression was determined in peripheral cells using the miScript® miRNA PCR Array platform. Pathways involving differentially expressed miRNAs were explored using the Ingenuity Pathway Analysis software. RESULTS: Atorvastatin repressed miR-29a-3p, miR-29b-3p, miR-300, miR-33a-5p, miR-33b-5p and miR-454-3p in HC subjects. On the contrary, simvastatin did not show any effect on miRNAs expression. Network analysis indicated that atorvastatin-modulated miRNAs regulate key cholesterol genes (ABCA1, HMGCR, INSIG1, LDLR, LPL, SCAP and SREBF1). Further subgroups analyses showed that miR-106b-5p, miR-17-3p and miR-590-5p were repressed in HC subjects within the lower quartile of atorvastatin response (lower LDL-C reduction), while the expression of miR-106b-5p, miR-17-3p and miR-183-5p was higher in the upper quartile of simvastatin response (higher LDL-C reduction) (p < 0.05). CONCLUSION: We show that a miRNAs-mediated epigenetic mechanism is differentially affected by statins therapy in vivo, which could be implicated in the variable response to these drugs. Further studies are necessary to disclose their particular role in the cholesterol-reduction response to statins.

Influence of CYP3A4 and CYP3A5 polymorphisms on tacrolimus and sirolimus exposure in stable kidney transplant recipients.

BACKGROUND: Polymorphisms in genes encoding for drug-metabolizing enzymes and drug transporters are among multiple factors that modulate the pharmacokinetic variability of tacrolimus (TAC) and sirolimus (SRL). This study aimed to evaluate the influence of single nucleotide polymorphisms (SNPs) on TAC and SRL dose-adjusted concentrations (C0/D) in stable kidney transplant recipients. METHODS: This is an exploratory and prospective study, which includes 46 stable kidney transplant recipients. These patients were monitored from the 3rd to the 24th month after transplantation. The SRL group consisted of 25 patients receiving TAC, prednisone (PRED), and mycophenolate sodium (MPS), which were converted from TAC to SRL at 3rd month after transplantation. The TAC group consisted of 21 patients who underwent treatment with TAC, PRED, and MPS. Both groups were genotyped for CYP3A4 rs2242480 (g.20230G>A), CYP3A5 rs15524 (g.31611C>T), CYP2C8 rs10509681 (c.1196A>G) and ABCB1 rs1045642 (c.3435C>T), rs1128503 (c.1236C>T), and rs2032582 (c.2677G>T/A) polymorphisms. RESULTS: In the TAC group, CYP3A4 rs2242480 A allele carriers were associated with lower TAC C0/D. For CYP3A5 rs15524 SNP, C0/D was higher among patients carrying TT genotype when compared with CT and CC genotype carriers in the SRL and, more consistently, in the TAC groups. For ABCB1 rs1045642 SNP, TT genotype was associated with reduced SRL C0/D, but only at month 15. CONCLUSIONS: CYP3A4 rs2242480 and CYP3A5 rs15524 SNPs resulted in significant changes in SRL and TAC C0/D at different times after transplantation.

Zinc supplementation reduces RANKL/OPG ratio and prevents bone architecture alterations in ovariectomized and type 1 diabetic rats.

Type 1 diabetes mellitus (T1DM) and estrogen deficiency are associated with several alterations in bone turnover. Zinc (Zn) is required for growth, development, and overall health. Zinc has been used in complementary therapy against bone loss in several diseases. We hypothesized that Zn supplementation represents a potential therapy against severe bone loss induced by the combined effect of estrogen deficiency and T1DM. We evaluated the protective effect of Zn against bone alterations in a chronic model of these disorders. Female Wistar rats were ramdomized into 3 groups (5 rats each): control, OVX/T1DM (ovariectomized rats with streptozotocin-induced T1DM), and OVX/T1DM+Zn (OVX/T1DM plus daily Zn supplementation). Serum biochemical, bone histomorphometric, and molecular analyses were performed. Histomorphometric parameters were similar between the control and OVX/T1DM+Zn groups, suggesting that Zn prevents bone architecture alterations. In contrast, the OVX/T1DM group showed significantly lower trabecular width and bone area as well as greater trabecular separation than the control. The OVX/T1DM and OVX/T1DM+Zn groups had significantly higher serum alkaline phosphatase activity than the control. The supplemented group had higher levels of serum-ionized calcium and phosphorus than the nonsupplemented group. The RANKL/OPG ratio was similar between the control and OVX/T1DM+Zn groups, whereas it was higher in the OVX/T1DM group. In conclusion, Zn supplementation prevents bone alteration in chronic OVX/T1DM rats, as demonstrated by the reduced RANKL/OPG ratio and preservation of bone architecture. The findings may represent a novel therapeutic approach to preventing OVX/T1DM-induced bone alterations.

Influence of ABCC2, CYP2C8, and CYP2J2 Polymorphisms on Tacrolimus and Mycophenolate Sodium-Based Treatment in Brazilian Kidney Transplant Recipients.

STUDY OBJECTIVE: To investigate the influence of single nucleotide polymorphisms (SNPs) in genes encoding metabolizing enzymes (CYP2C8, CYP2J2, and UGT2B7) and transporters (ABCC2 and ABCG2) on dose and dose-adjusted trough blood concentrations (C:D ratio), clinical outcomes, and occurrence of adverse events of tacrolimus and mycophenolate sodium in Brazilian kidney transplant recipients. DESIGN: Pharmacogenetic analysis of patients enrolled in a previously published study. PATIENTS: One hundred forty-eight adult kidney transplant recipients treated with tacrolimus, enteric-coated mycophenolate sodium, and prednisone for 90 days posttransplantation. MEASUREMENTS AND MAIN RESULTS: ABCC2 c.-24C>T and c.3972C>T, ABCG2 c.421C>A, CYP2C8*3, CYP2J2 c.-76G>T, and UGT2B7 c.372A>G SNPs were determined by real-time polymerase chain reaction. The CYP3A5*3C SNP data were used to eliminate the confounding effect of this variant on the results. ABCC2 c.3972T allele carriers showed higher tacrolimus C:D values than did carriers of the c.3972CC genotype. The CYP2C8*3 variant was also associated with slightly higher tacrolimus C:D values and higher estimated glomerular filtration rate but only in CYP3A5-nonexpressing patients (CYP3A5*3C/*3C carriers). None of the SNPs were associated with mycophenolate sodium dose or episodes of biopsy-confirmed acute rejection or delayed graft function. The CYP2J2 c.-76T allele was associated with increased risk for treatment-induced nausea and/or vomiting (OR: 5.30, 95% confidence interval 1.49-18.79, p<0.05). CONCLUSION: The ABCC2 c.3972C >T polymorphism affected tacrolimus C:D in Brazilian kidney transplant recipients. Further, CYP2C8*3 and CYP2J2 c.-76G>T SNPs influenced the renal function of these patients and the occurrence of adverse events during treatment with tacrolimus and mycophenolate sodium.

Modulation of miR-26a-5p and miR-15b-5p Exosomal Expression Associated with Clopidogrel-Induced Hepatotoxicity in HepG2 Cells.

Clopidogrel is an essential antiplatelet drug used to prevent thrombosis complications associated with atherosclerosis. However, hepatotoxicity is a potential adverse effect related to clopidogrel therapy. Exosome-derived miRNAs may be useful for improved monitoring of drug response and hepatotoxicity risk. In the present study, the expression of several exosomal miRNAs (miR-26a-5p, miR-145-5p, miR-15b-5p, and miR-4701-3p) and cell-derived mRNA targets (PLOD2, SENP5, EIF4G2, HMGA2, STRADB, and TLK1) were evaluated in HepG2 cells treated with clopidogrel (6.25, 12.5, 25, 50, and 100 µM) for 24 and 48 h. Then, clopidogrel cytotoxicity was evaluated by analyzing DNA fragmentation and the cell cycle profile using flow cytometry. Differential expression of exosome-derived miRNAs and cell-derived mRNAs was analyzed by RT-qPCR. Exposure of HepG2 cells to high concentrations of clopidogrel (50 and 100 µM) for 24 h caused significant DNA fragmentation (17.6 and 44.4%, respectively; p < 0.05) and 48 h (26.8 and 48.9%, respectively; p < 0.05), indicating cellular toxicity. Upregulation of miR-26a-5p and downregulation of miR-15b-5p was observed in cells exposed to 100 µM clopidogrel for 24 and 48 h. The miR-26a-5p target mRNAs HMGA2, EIF4G2, STRADB, and SENP5 were downregulated in HepG2 cells following exposure to cytotoxic concentrations of clopidogrel (50 and 100 µM) for 24 h, and HMGA2 levels remained low after 48 h of treatment. TLK1, a target of miR-15b-5p, was downregulated by 50 and 100 µM clopidogrel at 24 h. In conclusion, our results suggest that exposure to high concentrations of clopidogrel modulates the expression of exosomal miR-26a-5p and miR-15b-5p and their target mRNAs in HepG2 cells. Dysregulation of these miRNAs maybe modulate the regulatory pathways involved in clopidogrel-induced liver injury.

Influence of ABCC2, CYP2C8, and CYP2J2 polymorphisms on tacrolimus and mycophenolate sodium-based treatment in brazilian kidney transplant recipients

STUDY OBJECTIVE: To investigate the influence of single nucleotide polymorphisms (SNPs) in genes encoding metabolizing enzymes (CYP2C8, CYP2J2, and UGT2B7) and transporters (ABCC2 and ABCG2) on dose and dose-adjusted trough blood concentrations (C:D ratio), clinical outcomes, and occurrence of adverse events of tacrolimus and mycophenolate sodium in Brazilian kidney transplant recipients. DESIGN: Pharmacogenetic analysis of patients enrolled in a previously published study. PATIENTS: One hundred forty-eight adult kidney transplant recipients treated with tacrolimus, enteric-coated mycophenolate sodium, and prednisone for 90 days posttransplantation.MEASUREMENTS AND MAIN RESULTS:ABCC2 c.-24C>T and c.3972C>T, ABCG2 c.421C>A, CYP2C8*3, CYP2J2 c.-76G>T, and UGT2B7 c.372A>G SNPs were determined by real-time polymerase chain reaction. The CYP3A5*3C SNP data were used to eliminate the confounding effect of this variant on the results. ABCC2 c.3972T allele carriers showed higher tacrolimus C:D values than did carriers of the c.3972CC genotype. The CYP2C8*3 variant was also associated with slightly higher tacrolimus C:D values and higher estimated glomerular filtration rate but only in CYP3A5-nonexpressing patients (CYP3A5*3C/*3C carriers). None of the SNPs were associated with mycophenolate sodium dose or episodes of biopsy-confirmed acute rejection or delayed graft function. The CYP2J2 c.-76T allele was associated with increased risk for treatment-induced nausea and/or vomiting (OR: 5.30, 95% confidence interval 1.49-18.79, p<0.05)...

Modulation of miR-26a-5p and miR-15b-5p exosomal expression associated with Clopidogrel-induced hepatotoxicity in HepG2 Cells

Clopidogrel is an essential antiplatelet drug used to prevent thrombosis complications associated with atherosclerosis. However, hepatotoxicity is a potential adverse effect related to clopidogrel therapy. Exosome-derived miRNAs may be useful for improved monitoring of drug response and hepatotoxicity risk. In the present study, the expression of several exosomal miRNAs (miR-26a-5p, miR-145-5p, miR-15b-5p, and miR-4701-3p) and cell-derived mRNA targets (PLOD2, SENP5, EIF4G2, HMGA2, STRADB, and TLK1) were evaluated in HepG2 cells treated with clopidogrel (6.25, 12.5, 25, 50, and 100 μM) for 24 and 48 h. Then, clopidogrel cytotoxicity was evaluated by analyzing DNA fragmentation and the cell cycle profile using flow cytometry. Differential expression of exosome-derived miRNAs and cell-derived mRNAs was analyzed by RT-qPCR. Exposure of HepG2 cells to high concentrations of clopidogrel (50 and 100 μM) for 24 h caused significant DNA fragmentation (17.6 and 44.4%, respectively; p < 0.05) and 48 h (26.8 and 48.9%, respectively; p < 0.05), indicating cellular toxicity...

Zinc supplementation reduces RANKL/OPG ratio and prevents bone architecture alterations in ovariectomized and type 1 diabetic rats

Type 1 diabetes mellitus (T1DM) and estrogen deficiency are associated with several alterations in bone turnover. Zinc (Zn) is required for growth, development, and overall health. Zinc has been used in complementary therapy against bone loss in several diseases. We hypothesized that Zn supplementation represents a potential therapy against severe bone loss induced by the combined effect of estrogen deficiency and T1DM. We evaluated the protective effect of Zn against bone alterations in a chronic model of these disorders. Female Wistar rats were ramdomized into 3 groups (5 rats each): control, OVX/T1DM (ovariectomized rats with streptozotocin-induced T1DM), and OVX/T1DM+Zn (OVX/T1DM plus daily Zn supplementation). Serum biochemical, bone histomorphometric, and molecular analyses were performed. Histomorphometric parameters were similar between the control and OVX/T1DM+Zn groups, suggesting that Zn prevents bone architecture alterations. In contrast, the OVX/T1DM group showed significantly lower trabecular width and bone area as well as greater trabecular separation than the control...

Atorvastatin attenuation of ABCB1 expression is mediated by microRNA miR-491-3p in Caco-2 cells.

AIM: Atorvastatin, a HMG-CoA reductase inhibitor, used in the treatment of hypercholesterolemia, has been previously shown to regulate ABCB1 expression in vivo and in vitro. We hypothesized that the statin could regulate gene expression of ABCB1 transporter via microRNAs. METHODS: Expression of microRNAs and ABCB1 mRNA was examined in atorvastatin-treated and control cells using real-time PCR. miR-491-3P mimic and inhibitor were transfected in Caco-2 and ABCB1 expression was monitored by western blot and real-time PCR. RESULTS: In HepG2 cells, none of the microRNAs predicted to target ABCB1 3'UTR was regulated by atorvastatin treatment. In agreement with this, ABCB1 3'UTR activity was not modulated in HepG-2 cells after 48h-treatment as measured by luciferase assay. In Caco-2 cells, atorvastatin treatment provoked a decrease in luciferase activity and, accordingly, miR-491-3p was upregulated about 2.7 times after 48h-statin treatment. Luciferase analysis of miR-491-3p with a mimetic or inhibitor of miR-491-3p revealed that this microRNA could target ABCB1 3'UTR, as after miR-491-3p inhibition, ABCB1 levels were increased by two-fold, and miR-491-3p superexpression decreased ABCB1 3'UTR activity. Finally, functional analysis revealed that treatment with miR-491-3p inhibitor could reverses atorvastatin attenuation of ABCB1 (Pg-p) protein levels. CONCLUSION: Our results suggest atorvastatin control ABCB1 expression via miR-491-3p in Caco-2 cells. This finding may be an important mechanism of statin drug-drug interaction, since common concomitant drugs used in the prevention of cardiovascular diseases are ABCB1 substrates.

Influence of the CYP3A4/5 genetic score and ABCB1 polymorphisms on tacrolimus exposure and renal function in Brazilian kidney transplant patients.

BACKGROUND: Polymorphisms in genes encoding transport proteins and metabolizing enzymes involved in tacrolimus (TAC) disposition may be important sources of individual variability during treatment. OBJECTIVE: The aim of this study was to investigate the effect of combined CYP3A4 and CYP3A5 variants, using a CYP3A4/5 genetic score, and ABCB1 polymorphisms on therapeutic TAC monitoring and their relationship with clinical outcomes. MATERIAL AND METHODS: Brazilian kidney transplant recipients (n=151), who received TAC over 3 months after transplantation, were genotyped for CYP3A4 rs2242480 (g.20230G>A), CYP3A5 rs15524 (g.31611C>T) and rs776746 (g.6986A>G), ABCB1 rs1128503 (c.1236C>T), rs1045642 (c.3435C>T), and rs2032582 (c.2677G>T/A) polymorphisms. RESULTS: Frequencies of CYP3A4 g.20230A, CYP3A5 g.31611C, and g.6986A were 0.37, 0.26, and 0.28, respectively. These alleles were associated with TAC rapid metabolization and were used for CYP3A4/5 genetic score construction. A higher CYP3A4/5 genetic score was associated with higher TAC dose and lower concentrations for dose administered (Co/D, P<0.05). Ninety days after transplantation, the presence of two or more rapid metabolization alleles contributed toward 27.7% of Co/D variability and was associated with a lower estimated glomerular filtration rate values (P<0.05). For ABCB1, the frequencies of c.1236T, c.3435T, and c.2677T/A alleles were 0.42, 0.42, and 0.33/0.04. At 30 days after transplantation, patients carrying ABCB1 c.1236TT+c.3435TT+(c.2677TT+TA) genotypes had higher TAC Co/D than those with common or heterozygous genotypes (P<0.05). CONCLUSION: The results show the impact of the CYP3A4/5 genetic score on TAC exposure and renal function in Brazilian patients. Furthermore, ABCB1 polymorphisms, in a combined analysis, influenced TAC Co/D at 30 days after transplantation.

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. CONCLUSION: We identified several novel miRs affected by statin treatment. Additional research is needed to determine the biological significance of differentially expressed miRs identified in statins-induced HepG2 cells.

Antimicrobial photodynamic effect to treat residual pockets in periodontal patients: a randomized controlled clinical trial.

AIM: A randomized controlled clinical trial was designed to evaluate the efficacy of the photodynamic therapy (PDT) in the treatment of residual pockets of chronic periodontitis patients. MATERIAL AND METHODS: Thirty-four patients with at least four residual periodontal pockets undergoing maintenance care were included and randomly assigned to test group (PDT, n = 18) or control group (sham procedure, n = 16). The intervention was performed at baseline, 3, 6 and 12 months. Clinical parameters such as pocket probing depth (PPD), clinical attachment level (CAL), bleeding on probing (BoP) and plaque index (PI) were measured before intervention and after 3, 6 and 12 months. Subgingival samples were obtained at baseline, and after 7 days, 3, 6 and 12 months to quantify Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Treponema denticola and Tannerella forsythia by real-time polimerase chain reaction (PCR). RESULTS: All clinical variables showed significant improvement during the study, but there was no significant difference between test and control groups. The microbiological analyses showed no differences between groups at any time during the study. CONCLUSION: Within the limits of this clinical trial and considering the laser and photosensitizer protocol used, PDT failed to demonstrate additional clinical and bacteriological benefits in residual pockets treatment.

Antimicrobial photodynamic effect to treat residual pockets in periodontal patients: a randomized controlled clinical trial

AIM:A randomized controlled clinical trial was designed to evaluate the efficacy of the photodynamic therapy (PDT) in the treatment of residual pockets of chronic periodontitis patients.MATERIAL AND METHODS:Thirty-four patients with at least four residual periodontal pockets undergoing maintenance care were included and randomly assigned to test group (PDT, n = 18) or control group (sham procedure, n = 16). The intervention was performed at baseline, 3, 6 and 12 months. Clinical parameters such as pocket probing depth (PPD), clinical attachment level (CAL), bleeding on probing (BoP) and plaque index (PI) were measured before intervention and after 3, 6 and 12 months. Subgingival samples were obtained at baseline, and after 7 days, 3, 6 and 12 months to quantify Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Treponema denticola and Tannerella forsythia by real-time polimerase chain reaction (PCR).RESULTS:All clinical variables showed significant improvement during the study, but there was no significant difference between test and control groups. The microbiological analyses showed no differences between groups at any time during the study.CONCLUSION:Within the limits of this clinical trial and considering the laser and photosensitizer protocol used, PDT failed to demonstrate additional clinical and bacteriological benefits in residual pockets treatment.

Association between Pro12Ala, Pvull, Avall, Sstl and ADIPOQ single-nucleotide polymorphisms with lipid and glycemic profiles of patients with Berardinelli-Seip syndrome.

BACKGROUND/AIMS: Berardinelli-Seip syndrome (BSS) is a recessive autosomal genetic disorder characterized by the near loss of adipose tissue with disturbance in lipid metabolism. METHODS: Biochemical and hormonal parameters and Pro12Ala, Pvull, Avall, Sstl and ADIPOQ polymorphisms in 22 patients with BSS were analyzed and examined for a possible association with lipid profiles. RESULTS: Parental consanguinity, insulin resistance and diabetes mellitus were observed in 63.6, 81.8 and 59.1% of patients, respectively. All individuals presented high triglyceride levels, and 68.1% of patients showed high cholesterol levels. The Pro/Pro genotype of the Pro12Ala polymorphism of the PPARγ2 gene was found in 86.3% of patients; the Ala/Ala variant was not observed in any patient. The PvuII polymorphism of the LPL gene showed a frequency of 50% for the P1P2 variant. The AvaII polymorphism of the LDLR gene showed a similar frequency of 40.9% for both CT and TT variants. The S1S1 genotype of the Sstl polymorphism of the APOC3 gene had a frequency of 86.3%. The CC allele of the ADIPOQ polymorphism of the adiponectin gene was found in 54.6% of patients. CONCLUSIONS: No association was found between lipid parameters and the relevant Pvull, Avall and Sstl polymorphisms. However, we did observe an association of the Pro12Ala and ADIPOQ polymorphisms with higher lipid levels, suggesting a close relationship between these factors.

Influence of PCSK9 polymorphisms on plasma lipids and response to atorvastatin treatment in Brazilian subjects.

BACKGROUND: The proprotein convertase subtilisin/kexin type 9 (PCSK9) has a key role in the regulation of plasma low-density lipoprotein (LDL) cholesterol by enhancing the degradation of LDL receptor. Functional variants in PCSK9 have been associated with differences in plasma lipids and may contribute to the variability of the response to cholesterol-lowering drugs. OBJECTIVE: To investigate the influence of PCSK9 variants on plasma lipid profile and response to atorvastatin in Brazilian subjects. METHODS: PCSK9 E670G, I474V, and R46L single nucleotide polymorphisms (SNPs) and plasma lipids were evaluated in 163 hypercholesterolemics (HC) and 171 normolipidemics (NL). HC patients with indication for cholesterol-lowering drug therapy (n = 128) were treated with atorvastatin (10 mg/d/4 wk). PCSK9 SNPs were analyzed by real time polymerase chain reaction. RESULTS: Frequencies of the PCSK9 SNPs were similar between the HC and NL groups. Logistic regression analysis showed a trend of association between PCSK9 E670G and hypercholesterolemia after adjustment for covariates (P = .059). The 670G allele was associated with high basal levels of LDL cholesterol (P = .03) in HC patients using the extreme discordant phenotype method. No association tests were performed for R46L variant because of its very low frequency, whereas the I474V polymorphism and PCSK9 haplotypes were not related to hypercholesterolemia or variability on plasma lipids in both NL and HC groups (P > .05). LDL cholesterol reduction in response to atorvastatin was not influenced by PCSK9 genotypes or haplotypes. CONCLUSIONS: PCSK9 E670G polymorphism but not I474V contributes to the variability on plasma LDL cholesterol levels in hypercholesterolemic subjects. Both PCSK9 variants have no influence on cholesterol-lowering response to atorvastatin.

Detection and quantification of periodontal pathogens in smokers and never-smokers with chronic periodontitis by real-time polymerase chain reaction.

BACKGROUND: The purpose of the present investigation is to compare the presence and number of periodontal pathogens in the subgingival microbiota of smokers versus never-smokers with chronic periodontitis and matched probing depths (PDs) using real-time polymerase chain reaction (RT-PCR). METHODS: Forty current smokers and 40 never-smokers, matched for age, sex, and mean PD of sampling site, were included in this investigation. A full-mouth periodontal examination was performed, and a pooled subgingival plaque sample was collected from the deepest site in each quadrant of each participant. To confirm smoking status, expired carbon monoxide (CO) concentrations were measured with a CO monitor. The presence and quantification of Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Tannerella forsythia, and Treponema denticola were determined using RT-PCR. RESULTS: Smokers had greater overall mean PD (P = 0.001) and attachment loss (P = 0.006) and fewer bleeding on probing sites (P = 0.001). An association was observed between smoking status and the presence of A. actinomycetemcomitans (P <0.001). The counts of A. actinomycetemcomitans (P <0.001), P. gingivalis (P = 0.042), and T. forsythia (P <0.001) were significantly higher in smokers. CONCLUSIONS: Smokers showed significantly greater amounts of P. gingivalis, A. actinomycetemcomitans, and T. forsythia than never-smokers. There was a significant association between smoking and the presence of A. actinomycetemcomitans.

Low bone mineral density is associated to poor glycemic control and increased OPG expression in children and adolescents with type 1 diabetes.

AIMS: To investigate early alterations on bone mineral density (BMD) and RANK, RANKL and OPG mRNA expression in peripheral blood leukocytes (PBL) in children and adolescents with type 1 diabetes (T1D) and the relationship with glycemic control and bone biomarkers. METHODS: This cross-sectional study included 75 children and adolescents with T1D and 100 individuals without diabetes (normoglycemic-NG) aged 6-20 years old. T1D individuals were considered to have good (T1DG) or poor (T1DP) glycemic control according to the values of HbA1c. Phosphorus, magnesium, total and ionized calcium, osteocalcin, alkaline phosphatase and tartaric-resistant acid phosphatase (TRAP) values were determined in blood samples. BMD was measured by DEXA. RANK, RANKL and OPG mRNA expression was measured in PBL by real-time PCR. RESULTS: Osteocalcin values were decreased in diabetic groups in comparison to NG group (p<0.05), and a negative correlation with both serum glucose (r=-0.265, p<0.01) and Hb1Ac (r=-0.252, p<0.01) in T1D group was found. BMD was lower in diabetic groups in comparison with NG group (p<0.05) and a negative correlation was observed between BMD and both serum glucose (r=-0.357, p<0.01) and HbA1c (r=-0.351, p<0.01) in T1D group. OPG mRNA expression was significantly increased in T1D and T1DP groups in comparison with NG group (p<0.05). In conclusion, children and adolescents with early onset T1D presented low bone mineral density associated to unsatisfactory glycemic control, increased OPG mRNA expression and low osteocalcin concentration.

Reduced ABCG2 and increased SLC22A1 mRNA expression are associated with imatinib response in chronic myeloid leukemia.

Imatinib mesylate (IM) has become a standard of care in chronic myeloid leukemia (CML) therapy. Single nucleotide polymorphisms (SNPs) and altered expression in drug transporter genes may influence IM response. In order to investigate whether mRNA expression and SNPs in drug transporters are associated with IM resistance, we studied 118 chronic-phase CML patients receiving the standard dose of IM (400 mg/day). They were assigned as responders and non-responders according to European LeukemiaNet criteria (2009). mRNA expression in samples at diagnosis (without IM therapy) and outcomes after IM failure were also evaluated in subgroups of patients. Major molecular response (MMR), complete molecular response and primary and secondary resistance were all assessed. BCR-ABL1, ABCB1, ABCG2, SLC22A1 and SLCO1A2 mRNA expression and SNPs in ABCG2 and SLC22A1 genes were analyzed. ABCG2 mRNA expression in the non-responders was higher before and during IM therapy. Furthermore, ABCG2 was overexpressed in those who did not achieve MMR (P=0.027). In a subgroup of patients who switched to second-generation tyrosine kinase inhibitors, high mRNA expression of ABCG2 was associated with a risk of 24 times that of not achieving complete cytogenetic response (OR 24.00, 95% CI 1.74-330.80; P=0.018). In the responder group, patients who achieved MMR (P=0.009) presented higher mRNA levels of SLC22A1. The SNPs were not associated with mRNA expression of ABCG2 and SLC22A1. Our data suggest that elevated ABCG2 expression (an efflux transporter) could be associated with IM resistance and could impact on second-generation TKI response, whereas high SLC22A1 expression (an influx transporter) may be associated with a successful IM therapy in CML patients.