Effect of low level laser irradiation on VEGF gene expression in cultured endothelial cells

Background. Endothelial cells play a crucial role in the angiogenesis which is initiated by vascular endothelial growth factor (VEGF). Low level laser therapy (LLLT) stimulates repair processes which are based on the formation of new blood vessels.Aim. The aim of this study was to evaluate the impact of LLLT on VEGF gene expression in endothelial cells cultured in vitro.Material and methods. Freshly isolated endothelial cells from the human umbilical vein endothelial cells (HUVEC) line were used in the study. The cells were irradiated with a semiconductor laser emitting visiblelaser radiation at the wave length of 630 nm and the power of 30 mW, and radiation at the wavelength of 808 nm and the power of 60 mW in the infrared range. The study was performed with cell cultures subjected to four different procedures: I — control cells (not subjected to irradiation); II — cells subjected to an energy dose of 2 J/cm2; III — cells subjected to an energy dose of 4 J/cm2; and IV — cells subjected to an energy dose of 8 J/cm2. The cells were cultured for six days, and exposed to irradiation twice. The next step was to evaluate the VEGF gene expression by applying real-time PCR.Results. By using low power laser irradiation, we obtained a statistically significant increase in VEGF gene expression, particularly at doses of 2 and 8 J/cm2 in the wave length range of 630 nm. The wave length of 808 nm had a similar effect on increases in gene expression. However, the differences were statistically non-significant when compared to the control cells.Conclusions. This study has shown that low-power laser radiation in the visible light spectrum (630 nm) results in de novo formation of VEGF-A in the endothelial cells in culture

The influence of genetic polymorphisms of CYP2C19 and ABCB1 on ADP-induced platelet aggregation in clopidogrel-treated patients: A comparison between the index hospitalization for myocardial infarction and the 3-month follow-up visit

Background. Recent studies suggest that polymorphisms of genes involved in the clopidogrel metabolism may be associated with an impaired drug bioactivation and possibly with unfavourable clinical outcomes. The aim of this study was to assess the effect of selected genetic polymorphisms on adenosine diphosphate-induced platelet aggregation (ADP-PA) during the index hospitalization and after 3 months of clopidogrel therapy in patients presenting with myocardial infarction (MI). Materials and methods. The study was designed as a single-center cohort trial with the 3-month follow-up. Genotyping for CYP2C19*2, CYP2C19*17, ABCB1 alleles and platelet reactivity assessment using the Multiplate Analyzer were performed in 157 patients. Results. ADP-PA during the index hospitalization was significantly higher than at the 3-month follow-up visit regardless of the genotyp e [CYP2C19*1/*1 alleles (24.0 v. 15.5 U; p < 0.00001), CYP2C19*17 allele (CT: 25.0 v. 15.0 U; p = 0.000 2; TT: 35.0 v. 22.0 U; p = 0.02) and ABCB1 allele (CC: 27.0 v. 15.0 U; p < 0.0002; CT 24.0 v. 17.0 U; p < 0.0005)]. In univariate analysis we failed to demonstrate any impact of the analyzed genetic variants on both in-hospital and 3-month ADP-PA, except for CYP2C19*17/*17 homozygotes. Significantly higher values of ADP-PA were found in CYP2C19*17/*17 (TT homozygotes) allele carriers when compared with carriers of two wild alleles during the index hospitalization (CC: 20.0 U v. TT: 35.0 U; p = 0.02), but not at the 3-month follow-up visit. Multivariate regression analysis revealed increased mean platelet volume (β = 7.2), elevated platelet count (β = 0.2) and the presence of heart failure at discharge (β = 6.9), but not genetic polymorphisms, to be independent determinants of high ADP-PA during the index hospitalization. Similarly, elderly age (β = 3.3), high white blood cell count (β = 1.4), elevated platelet count (β = 0.4) and increased mean platelet volume (β = 0.1), but not genetic polymorphisms, were independently associated with the higher values of ADP-PA after 3 months of clopidogrel therapy. Conclusions. On-clopidogrel platelet reactivity significantly decreases beyond the acute phase of MI regardless of the genotype. Additionally, our study indicates that in clopidogrel-treated MI patients genetic polymorphisms are not the major determinants of the interindividual variability in platelet reactivity. However, due to a limited sample size, their minor contribution cannot be excluded

Brain-derived neurotrophic factor serum concentration and BDNF Val66Met polymorphism in patients with peripheral artery disease: the importance of heart failure

Introduction: Brain-derived neurotrophic factor (BDNF) and BDNF Val66Met polymorphism have been associated with cardiovascular diseases such as atherosclerosis, congestive heart failure (CHF), hypertension and ischaemic heart disease (IHD). To the authors’ knowledge, such connections have not been described in peripheral artery disease (PAD) yet. Material and methods: 159 PAD subjects and 57 controls were included. All enrolled subjects underwent evaluation of clinical status. Information on comorbidities such as diabetes type 2, hypertension, IHD and CHF, was gathered. Serum concentrations of BDNF were measured by ELISA. Genotypes of the BDNF-AS SNP rs6265 were determined using TaqMan SNP Genotyping Assay. Results: PAD patients had significantly lower BDNF serum concentrations compared to controls (median values of 7.2 vs. 35.1 ng/mL, P < 0.001). Concentrations were significantly lower in patients with concomitant CHF (P < 0.05). The CHF subgroup was characterised by a greater prevalence of diabetes and ischaemic heart disease (P < 0.01). There was no significant difference between BDNF serum concentrations and other comorbidities, ABI, and medical history including disease duration and past interventions. No important correlations were found for BDNF Val66Met polymorphism. Conclusions: The present study adds to the body of evidence associating BDNF and atherosclerosis. The serum BDNF concentrations were lower in PAD, especially in a subgroup with comorbid CHF. These results suggest that a larger cardiovascular burden is connected with decreased BDNF serum concentrations. No evidence was found to support the hypothesis that BDNF gene polymorphism may be a contributing factor in the pathogenesis of cardiovascular diseases such as PAD

Impact of CYP2C19 polymorphisms on antiplatelet efficacy of clopidogrel in patients after myocardial infarction

Aim. The aim of this study was to evaluate the complex effects of polymorphisms of the gene encodingthe CYP2C19 enzyme on the antiplatelet efficacy of clopidogrel during follow-up visits.Material and methods. This study was designed as a prospective, single-centre observational clinical trialwith a one-year follow-up. Data from 178 patients on clopidogrel, taken during follow-up visits, was analysed.Results. Patients were divided into three groups according to the expected metabolic activity of theCYP2C19enzyme: ‘superior metabolisers’ (681 GG + 806 CT or TT), ‘neutral metabolisers’(681 GG + 806 CC and 681 GA or AA + 806 CT or TT) and ‘inferior metabolisers’ (681 GA or AA + 806 CC).The antiplatelet effect of clopidogrel was strongest in ‘superior metabolisers’ and weakest in ‘inferior metabolisers’,as assessed by adenosine diphosphate (ADP)-induced platelet aggregation and the VASP assay.Comparison of results of ADP-induced platelet aggregation measurements in pairs showed a significantdifference (p = 0.03) only between ‘superior metabolisers’ vs. ‘inferior metabolisers’. With the VASP assay,significant differences between ‘superior metabolisers’ and ‘neutral metabolisers’ (p = 0.013), ‘neutralmetabolisers’ and ‘inferior metabolisers’ (p = 0.015), and ‘superior metabolisers’ and ‘inferior metabolisers’(p = 0.00003) were found. No impact on clinical outcome was present. A tendency for an increasingneed for a clopidogrel-to-prasugrel switch was observed with a decrease in CYP2C19 metabolic activity.Conclusion. Multiple polymorphisms of the gene encoding the CYP2C19 enzyme have a significant impacton the antiplatelet efficacy of clopidogrel during follow-up visits; however, their influence on clinicaloutcome needs further clarification.Aim. The aim of this study was to evaluate the complex effects of polymorphisms of the gene encodingthe CYP2C19 enzyme on the antiplatelet efficacy of clopidogrel during follow-up visits.Material and methods. This study was designed as a prospective, single-centre observational clinical trialwith a one-year follow-up. Data from 178 patients on clopidogrel, taken during follow-up visits, was analysed.Results. Patients were divided into three groups according to the expected metabolic activity of theCYP2C19enzyme: ‘superior metabolisers’ (681 GG + 806 CT or TT), ‘neutral metabolisers’(681 GG + 806 CC and 681 GA or AA + 806 CT or TT) and ‘inferior metabolisers’ (681 GA or AA + 806 CC).The antiplatelet effect of clopidogrel was strongest in ‘superior metabolisers’ and weakest in ‘inferior metabolisers’,as assessed by adenosine diphosphate (ADP)-induced platelet aggregation and the VASP assay. Comparison of results of ADP-induced platelet aggregation measurements in pairs showed a significant difference (p = 0.03) only between ‘superior metabolisers’ vs. ‘inferior metabolisers’. With the VASP assay,significant differences between ‘superior metabolisers’ and ‘neutral metabolisers’ (p = 0.013), ‘neutralmetabolisers’ and ‘inferior metabolisers’ (p = 0.015), and ‘superior metabolisers’ and ‘inferior metabolisers’(p = 0.00003) were found. No impact on clinical outcome was present. A tendency for an increasingneed for a clopidogrel-to-prasugrel switch was observed with a decrease in CYP2C19 metabolic activity.Conclusion. Multiple polymorphisms of the gene encoding the CYP2C19 enzyme have a significant impacton the antiplatelet efficacy of clopidogrel during follow-up visits; however, their influence on clinicaloutcome needs further clarification

Enhanced antiplatelet effect of enteric- -coated acetylsalicylic acid in co-administration with pantoprazole

Background. Proton pump inhibitors (PPI) are recommended for patients receiving antiplatelet therapy. Several studies have revealed that PPI may attenuate the antiplatelet effect of ASA. However, our pilot study has indicated a positive interaction between pantoprazole and enteric-coated aspirin. The aim of the current study was to confirm that pantoprazole enhances the antiplatelet effect ofenteric-coated aspirin in patients with acute coronary syndrome (ACS) treated with dual antiplatelettherapy. Moreover, the influence of CYP2C19 polymorphism on the antiplatelet effect of aspirinwas assessed.Material and methods. Ninety three consecutive ACS patients were prospectively enrolled in a randomized, crossover, open-labeled study. Forty four patients were given orally 40 mg of pantoprazole for the initial fourdays while the remaining forty nine were treated with pantoprazole from the 5th to the 8th day of hospitalization. Blood samples were collected at 6.00 a.m., 10.00 a.m., 2.00 p.m., and 7.00 p.m. on the 4th and 8th daysof hospitalization. Aggregation in response to arachidonic acid was assessed by impedance aggregometry.Results. Lower mean platelet aggregation on pantoprazole was observed on the 8th day of hospitalization(p = 0.03). A cross-time analysis of platelet aggregation demonstrated statistical significance at two hoursand six hours after co-administration of pantoprazole and antiplatelet agents, with the highest absolutedifference observed two hours after drugs ingestion. No significant differences in aggregation betweenstudy groups were observed on the 4th day of hospitalization. No influence of CYP2C19 polymorphismon the antiplatelet effect of aspirin was observed.Conclusions. Co-administration of pantoprazole enhances the antiplatelet effect of enteric-coated aspirinin patients with ACS.Background. Proton pump inhibitors (PPI) are recommended for patients receiving antiplatelet therapy.Several studies have revealed that PPI may attenuate the antiplatelet effect of ASA. However, our pilot study has indicated a positive interaction between pantoprazole and enteric-coated aspirin.The aim of the current study was to confirm that pantoprazole enhances the antiplatelet effect ofenteric-coated aspirin in patients with acute coronary syndrome (ACS) treated with dual antiplatelettherapy. Moreover, the influence of CYP2C19 polymorphism on the antiplatelet effect of aspirinwas assessed.Material and methods. Ninety three consecutive ACS patients were prospectively enrolled in a randomized, crossover, open-labeled study. Forty four patients were given orally 40 mg of pantoprazole for the initial fourdays while the remaining forty nine were treated with pantoprazole from the 5th to the 8th day of hospitalization. Blood samples were collected at 6.00 a.m., 10.00 a.m., 2.00 p.m., and 7.00 p.m. on the 4th and 8th daysof hospitalization. Aggregation in response to arachidonic acid was assessed by impedance aggregometry.Results. Lower mean platelet aggregation on pantoprazole was observed on the 8th day of hospitalization(p = 0.03). A cross-time analysis of platelet aggregation demonstrated statistical significance at two hoursand six hours after co-administration of pantoprazole and antiplatelet agents, with the highest absolutedifference observed two hours after drugs ingestion. No significant differences in aggregation betweenstudy groups were observed on the 4th day of hospitalization. No influence of CYP2C19 polymorphismon the antiplatelet effect of aspirin was observed.Conclusions. Co-administration of pantoprazole enhances the antiplatelet effect of enteric-coated aspirinin patients with ACS

Phenotyping vs. genotyping for prediction of clopidogrel efficacy and safety: the PEGASUS-PCI study

Background: Prognostic values of genotyping and phenotyping for assessment of clopidogrel responsiveness have been shown in independent studies. Objectives: To compare different assays for prediction of events during long-term follow-up. Methods: In this prospective cohort study polymorphisms of CYP2C19*2 and CYP2C19*17 alleles, vasodilator- stimulated phosphoprotein phosphorylation (VASP) assay, multiple electrode aggregometry (MEA), cone and platelet analyser (CPA) and platelet function analyser (PFA- 100) were performed in 416 patients undergoing percutaneous coronary intervention. The rates of events were recorded during a 12-month follow-up. Results: Platelet aggregation by MEA predicted stent thrombosis (2.4%) better (c-index = 0.90; P < 0.001; sensitivity = 90%; specificity = 83%) than the VASP assay, CPA or PFA-100 (c-index 0.05; sensitivity < 70%; specificity < 70% for all) or even the CYP2C19*2 polymorphism (c-index 0.05; sensitivity = 30%; specificity = 71%). Survival analysis indicated that patients classified as poor responders by MEA had a substantially higher risk of developing stent thrombosis or MACE than clopidogrel responders (12.5% vs. 0.3%, P < 0.001, and 18.5% vs. 11.3%, P = 0.022, respectively), whereas poor metabolizers (CYP2C19*1/*2 or *2/*2 carriers) were not at increased risks (stent thrombosis, 2.7% vs. 2.5%, P > 0.05; MACE, 13.5% vs. 12.1%, P = 0.556). The incidence of major bleedings (2.6%) was numerically higher in patients with an enhanced vs. poor response to clopidogrel assessed by MEA (4% vs. 0%) or in ultra-metabolizers vs. regular metabolizers (CYP2C19*17/*17 vs. CYP2C19*1/*1; 9.5% vs. 2%). The classification tree analysis demonstrated that acute coronary syndrome at hospitalization and diabetes mellitus were the best discriminators for clopidogrel responder status. Conclusions: Phenotyping of platelet response to clopidogrel was a better predictor of stent thrombosis than genotyping

Searching for association of the CAG repeat polymorphism in the mitochondrial DNA polymerase gamma gene (POLG) with colorectal cancer

Mitochondrial DNA polymerase gamma (POLG) is the only DNA polymerase involved in maintaining the mitochondrial genome. Recent studies demonstrated an association of CAG repeat polymorphism in the second exon of POLG gene with the risk of cancer. We investigated the CAG repeat variability in the POLG gene in tumor and non-tumor tissues from colorectal cancer patients and in DNA samples isolated from blood obtained from age-matched healthy persons. Somatically occuring CAG-repeat alterations in cancer tissues have been observed in 10% of patients, but no association has been found between the CAG repeat variants in the POLG gene and colorectal cancer risk

Evaluation of TGF-Beta 2 and VEGF α

Purpose. To evaluate the expression profiles of the VEGFα and TGFβ in the ERMs and ILMs in retinal disorders. Methods. In this nonrandomized prospective study, 75 patients (34 females and 41 males) referred to pars plana vitrectomy (PPV) due to different retinal diseases were enrolled to the study. The samples of ERMs and ILMs collected during PPV were immediately put in TRIzol® Reagent (Life Technologies, USA) and stored at −70°C until RNA extraction. Gene expression analysis was done with TaqMan® Gene Expression Assays (Applied Biosystems, USA) following the manufacturer’s instructions. Results. The gene expression levels of VEGFα as well as of TGFβ2 were significantly higher in ERMs than in ILMs in all studied groups. The level of TGFβ2 expression exhibits a significantly lower values in iERMs as compared with the RRD group (p=0.043). There were differences in TGFβ2 expression in ILM in groups studied: DR versus RRD, p=0.003; DR versus iERM, p=0,047; and iERM versus RRD, p=0.004. Conclusions. Our results revealed that factors associated with angiogenesis and wound healing processes in eyes with RRD, PDR, iERM, and MH were more upregulated in ERMs than in ILMs. This may indicate that ILM is not responsible for reproliferation and its peeling should be avoided in routine PPV

Association between Genetic Variants on Chromosome 15q25 Locus and Several Nicotine Dependence Traits in Polish Population: A Case-Control Study

Tobacco smoking continues to be a leading cause of disease and mortality. Recent research has confirmed the important role of nicotinic acetylcholine receptor (nAChR) gene cluster on chromosome 15q 24-25 in nicotine dependence and smoking. In this study we tested the association of smoking initiation, age at onset of daily smoking, and heaviness of smoking with five single nucleotide polymorphisms (SNPs) within the CHRNA5-CHRNA3-CHRNB4 cluster. The group of 389 adult subjects of European ancestry from the north of Poland, including 212 ever (140 current and 72 former) and 177 never smokers with mean age 49.26, was genotyped for rs16969868, rs1051730, rs588765, rs6495308, and rs578776 polymorphisms. Distributions of genotypes for rs16969868 and rs1051730 were identical so they were analyzed together. Further analysis revealed the association between rs16969868-1051730 (OR = 2.66; 95% CI: 1.30–5.42) and number of cigarettes smoked per day (CPD) with heaviness of nicotine addiction measured by the Fagerström Test for Nicotine Dependence (FTND) (OR = 2.60; 95% CI: 1.24–5.43). No association between these polymorphisms and other phenotypes was found. Similarly, the association between rs588765, rs6495308, rs578776, and analyzed phenotypes was not confirmed. This study provides strong evidence for the role of the CHRNA5-CHRNA3-CHRNB4 cluster in heaviness of nicotine addiction

Interplay between genetic and clinical variables affecting platelet reactivity and cardiac adverse events in patients undergoing percutaneous coronary intervention.

Several clinical and genetic variables are associated with influencing high on treatment platelet reactivity (HTPR). The aim of the study was to propose a path model explaining a concurrent impact among variables influencing HTPR and ischemic events. In this prospective cohort study polymorphisms of CYP2C19*2, CYP2C19*17, ABCB1, PON1 alleles and platelet function assessed by Multiple Electrode Aggregometry were assessed in 416 patients undergoing percutaneous coronary intervention treated with clopidogrel and aspirin. The rates of major adverse cardiac events (MACE) were recorded during a 12-month follow up. The path model was calculated by a structural equation modelling. Paths from two clinical characteristics (diabetes mellitus and acute coronary syndrome (ACS)) and two genetic variants (CYP2C19*2 and CYP2C19*17) independently predicted HTPR (path coefficients: 0.11 0.10, 0.17, and -0.10, respectively; p<0.05 for all). By use of those four variables a novel score for prediction of HTPR was built: in a factor-weighted model the risk for HTPR was calculated with an OR of 3.8 (95%CI: 3.1-6.8, p<0.001) for a score level of ≥1 compared with a score of <1. While MACE was independently predicted by HTPR and age in the multivariate model (path coefficient: 0.14 and 0.13, respectively; p<0.05), the coexistence of HTPR and age ≥75 years emerged as the strongest predictor of MACE. Our study suggests a pathway, which might explain indirect and direct impact of variables on clinical outcome: ACS, diabetes mellitus, CYP2C19*2 and CYP2C19*17 genetic variants independently predicted HTPR. In turn, age ≥75 years and HTPR were the strongest predictors of MACE