K-Mer Analyses Reveal Different Evolutionary Histories of Alpha, Beta, and Gamma Papillomaviruses.

Papillomaviruses (PVs) are a heterogeneous group of DNA viruses that can infect fish, birds, reptiles, and mammals. PVs infecting humans (HPVs) phylogenetically cluster into five genera (Alpha-, Beta-, Gamma-, Mu- and Nu-PV), with differences in tissue tropism and carcinogenicity. The evolutionary features associated with the divergence of Papillomaviridae are not well understood. Using a combination of k-mer distributions, genetic metrics, and phylogenetic algorithms, we sought to evaluate the characteristics and differences of Alpha-, Beta- and Gamma-PVs constituting the majority of HPV genomes. A total of 640 PVs including 442 HPV types, 27 non-human primate PV types, and 171 non-primate animal PV types were evaluated. Our analyses revealed the highest genetic diversity amongst Gamma-PVs compared to the Alpha and Beta PVs, suggesting reduced selective pressures on Gamma-PVs. Using a sequence alignment-free trimer (k = 3) phylogeny algorithm, we reconstructed a phylogeny that grouped most HPV types into a monophyletic clade that was further split into three branches similar to alignment-based classifications. Interestingly, a subset of low-risk Alpha HPVs (the species Alpha-2, 3, 4, and 14) split from other HPVs and were clustered with non-human primate PVs. Surprisingly, the trimer-constructed phylogeny grouped the Gamma-6 species types originally isolated from the cervicovaginal region with the main Alpha-HPV clade. These data indicate that characterization of papillomavirus heterogeneity via orthogonal approaches reveals novel insights into the biological understanding of HPV genomes.

Association of waterpipe smoking with myocardial infarction and determinants of metabolic syndrome among catheterized patients.

BACKGROUND: Waterpipe smoking is a rising global public health epidemic perceived by many users to be less harmful, though its toxicity overlaps or even exceeds that of cigarette smoking. Short-term cardiovascular changes due to waterpipe smoking are well established, but longer-term health impacts are still not fully elucidated. OBJECTIVE: We aim to investigate the association of waterpipe smoking with myocardial infarction among patients undergoing cardiac catheterization. METHODS: The study was performed on Lebanese patients referred for cardiac catheterization. Patient's blood was collected for metabolic measures and questionnaires were filled out to include socio-demographic, behavioral and pertinent medical characteristics of the study subjects. RESULTS: Myocardial infarction is significantly and independently associated with waterpipe smoking, with odds ratio (OR) of 1.329 (95% CI: [1.04-1.68]; p = .021), which is lower than that for cigarette smoking (OR = 1.87, 95% CI: [1.63-2.15]; p < .001). Only diabetes showed significant association with waterpipe smoking among MI enrollees (OR = 1.66, 95%CI: [1.04-2.63]; p = .032). CONCLUSION: The study provides yet another evidence for the adverse cardiovascular effects of waterpipe smoking on a clinical level. The harmful effects of waterpipe smoking should be underscored by health care professionals.

Impact of inflammation, gene variants, and cigarette smoking on coronary artery disease risk.

BACKGROUND: The role of inflammation in coronary artery disease (CAD) pathogenesis is well recognized. Moreover, smoking inhalation increases the activity of inflammatory mediators through an increase in leukotriene synthesis essential in atherosclerosis pathogenesis. AIM: The aim of this study is to investigate the effect of "selected" genetic variants within the leukotriene (LT) pathway and other variants on the development of CAD. METHODS: CAD was detected by cardiac catheterization. Logistic regression was performed to investigate the association of smoking and selected susceptibility variants in the LT pathway including ALOX5AP, LTA4H, LTC4S, PON1, and LTA as well as CYP1A1 on CAD risk while controlling for age, gender, BMI, family history, diabetes, hyperlipidemia, and hypertension. RESULTS: rs4769874 (ALOX5AP), rs854560 (PON1), and rs4646903 (CYP1A1 MspI polymorphism) are significantly associated with an increased risk of CAD with respective odds ratios of 1.53703, 1.67710, and 1.35520; the genetic variant rs9579646 (ALOX5AP) is significantly associated with a decreased risk of CAD (OR 0.76163). Moreover, a significant smoking-gene interaction is determined with CYP1A1 MspI polymorphism rs4646903 and is associated with a decreased risk of CAD in current smokers (OR 0.52137). CONCLUSION: This study provides further evidence that genetic variation of the LT pathway, PON1, and CYP1A1 can modulate the atherogenic processes and eventually increase the risk of CAD in our study population. Moreover, it also shows the effect of smoking-gene interaction on CAD risk, where the CYP1A1 MspI polymorphism revealed a decreased risk in current smokers.

Association of hypertension with coronary artery disease onset in the Lebanese population.

The onset of coronary artery disease (CAD) is influenced by cardiovascular risk factors that often occur in clusters and may build on one another. The objective of this study is to examine the relationship between hypertension and CAD age of onset in the Lebanese population. This retrospective analysis was performed on data extracted from Lebanese patients (n = 3,753). Logistic regression examined the association of hypertension with the age at CAD diagnosis after controlling for other traditional risk factors. The effect of antihypertensive drugs and lifestyle changes on the onset of CAD was also investigated. Results showed that hypertension is associated with late onset CAD (OR=0.656, 95% CI=0.504-0.853, p=0.001). Use of antihypertensive drugs showed a similar association with delayed CAD onset. When comparing age of onset in CAD patients with traditional risk factors such as hypertension, diabetes, hyperlipidemia, obesity, smoking and family history of CAD, the age of onset was significantly higher for patients with hypertension compared to those with any of the other risk factors studied (p < 0.001). In conclusion, hypertension and its treatment are associated with late coronary atherosclerotic manifestations in Lebanese population. This observation is currently under investigation to clarify its genetic and/or environmental mechanisms.

Genetic and environmental influences on total plasma homocysteine and its role in coronary artery disease risk.

BACKGROUND: Elevated levels of total plasma homocysteine are a risk factor for atherosclerotic disease. AIMS: The rationale behind this study is to explore the correlation between degree and site of coronary lesion and hyperhomocysteinemia in Lebanese CAD patients and assess environmental and genetic factors for elevated levels of total plasma homocysteine. METHODS: A total of 2644 patients were analyzed for traditional CAD risk factors. Logistic regression was performed to determine the association of hyperhomocysteinemia with degree and site of coronary lesions controlling for risk factors. Environmental and genetic factors for hyperhomocysteinemia were analyzed by logistic regression using a candidate gene approach. RESULTS: Traditional risk factors were correlated with stenosis. Hyperhomocysteinemia associated with increased risk of overall stenosis, and risk of mild and severe occlusion in major arteries. Hyperhomocysteinemia and hypertension were highly correlated suggesting that hyperhomocysteinemia acts as a hypertensive agent leading to CAD. Diuretics and genetic polymorphisms in MTHFR and SLCO1B1 were associated with hyperhomocysteinemia. CONCLUSIONS: Hyperhomocysteinemia is a medical indicator of specific vessel stenosis in the Lebanese population. Hypertension is a major link between hyperhomocysteinemia and CAD occurrence. Genetic polymorphisms and diuretics' intake explain partly elevated homocysteine levels. This study has important implications in CAD risk prediction.

Large scale association analysis identifies three susceptibility loci for coronary artery disease.

Genome wide association studies (GWAS) and their replications that have associated DNA variants with myocardial infarction (MI) and/or coronary artery disease (CAD) are predominantly based on populations of European or Eastern Asian descent. Replication of the most significantly associated polymorphisms in multiple populations with distinctive genetic backgrounds and lifestyles is crucial to the understanding of the pathophysiology of a multifactorial disease like CAD. We have used our Lebanese cohort to perform a replication study of nine previously identified CAD/MI susceptibility loci (LTA, CDKN2A-CDKN2B, CELSR2-PSRC1-SORT1, CXCL12, MTHFD1L, WDR12, PCSK9, SH2B3, and SLC22A3), and 88 genes in related phenotypes. The study was conducted on 2,002 patients with detailed demographic, clinical characteristics, and cardiac catheterization results. One marker, rs6922269, in MTHFD1L was significantly protective against MI (OR=0.68, p=0.0035), while the variant rs4977574 in CDKN2A-CDKN2B was significantly associated with MI (OR=1.33, p=0.0086). Associations were detected after adjustment for family history of CAD, gender, hypertension, hyperlipidemia, diabetes, and smoking. The parallel study of 88 previously published genes in related phenotypes encompassed 20,225 markers, three quarters of which with imputed genotypes The study was based on our genome-wide genotype data set, with imputation across the whole genome to HapMap II release 22 using HapMap CEU population as a reference. Analysis was conducted on both the genotyped and imputed variants in the 88 regions covering selected genes. This approach replicated HNRNPA3P1-CXCL12 association with CAD and identified new significant associations of CDKAL1, ST6GAL1, and PTPRD with CAD. Our study provides evidence for the importance of the multifactorial aspect of CAD/MI and describes genes predisposing to their etiology.

Parental consanguinity and family history of coronary artery disease strongly predict early stenosis.

BACKGROUND: Coronary artery disease (CAD) is a multifactorial disease with acquired and inherited components. AIM: We investigated the roles of family history and consanguinity on CAD risk and age at diagnosis in 4284 patients. The compounded impact of diabetes, hyperlipidemia, hypertension, smoking, and BMI, which are known CAD risk factors, on CAD risk and age at diagnosis was also explored. METHODS: CAD was determined by cardiac catheterization. Logistic regression and stratification were performed to determine the impact of family history and consanguinity on risk and onset of CAD, controlling for diabetes, hyperlipidemia, hypertension, smoking, and BMI. RESULTS: Family history of CAD and gender significantly increased the risk for young age at diagnosis of CAD (p<0.001). Consanguinity did not promote risk of CAD (p=0.38), but did affect age of disease diagnosis (p<0.001). The mean age at disease diagnosis was lowest, 54.8 years, when both family history of CAD and consanguinity were considered as unique risk factors for CAD, compared to 62.8 years for the no-risk-factor patient category (p<0.001). CONCLUSIONS: Family history of CAD and smoking are strongly associated with young age at diagnosis. Furthermore, parental consanguinity in the presence of family history lowers the age of disease diagnosis significantly for CAD, emphasizing the role of strong genetic and cultural CAD modifiers. These findings highlight the increased role of genetic determinants of CAD in some population subgroups, and suggest that populations and family structure influence genetic heterogeneity between patients with CAD.

Skeletal muscle channelopathies: new insights into the periodic paralyses and nondystrophic myotonias.

PURPOSE OF REVIEW: To summarize advances in our understanding of the clinical phenotypes, genetics, and molecular pathophysiology of the periodic paralyses, the nondystrophic myotonias, and other muscle channelopathies. RECENT FINDINGS: The number of pathogenic mutations causing periodic paralysis, nondystrophic myotonias, and ryanodinopathies continues to grow with the advent of exon hierarchy analysis strategies for genetic screening and better understanding and recognition of disease phenotypes. Recent studies have expanded and clarified the role of gating pore current in channelopathy pathogenesis. It has been shown that the gating pore current can account for the molecular and phenotypic diseases observed in the muscle sodium channelopathies, and, given that homologous residues are affected in mutations of calcium channels, it is possible that pore leak represents a pathomechanism applicable to many channel diseases. Improvements in treatment of the muscle channelopathies are on the horizon. A randomized controlled trial has been initiated for the study of mexiletine in nondystrophic myotonias. The class IC antiarrhythmia drug flecainide has been shown to depress ventricular ectopy and improve exercise capacity in patients with Andersen-Tawil syndrome. SUMMARY: Recent studies have expanded our understanding of gating pore current as a disease-causing mechanism in the muscle channelopathies and have allowed new correlations to be drawn between disease genotype and phenotype.

Peroxynitrite increases VEGF expression in vascular endothelial cells via STAT3.

Increased expression of vascular endothelial growth factor (VEGF) has been correlated with increased oxidative stress and formation of peroxynitrite in numerous disease conditions, including diabetic microangiopathy, tumor angiogenesis, and atherosclerosis. In this study we tested the hypothesis that peroxynitrite stimulates VEGF expression. Treatment of microvascular endothelial cells with exogenous peroxynitrite induced a time- and dose-dependent increase in VEGF mRNA, which peaked within 1 h of treatment at a concentration of 100 muM. The increase in VEGF mRNA was followed by a significant increase in VEGF protein. To define the molecular mechanisms involved, the effect of peroxynitrite was determined on the activation of two transcription factors known to regulate VEGF expression during hypoxia and tumor angiogenesis-signal transducer and activator of transcription 3 (STAT3) and hypoxia-inducible factor-1 (HIF-1). Peroxynitrite caused activation and nuclear translocation of STAT3, but not HIF-1. Moreover, transduction of endothelial cells with dominant-negative STAT3 abrogated the peroxynitrite-induced increase in VEGF mRNA. The increase in VEGF mRNA was also blocked by inhibitors of transcription and was unaffected by the inhibition of protein synthesis. These results indicate that peroxynitrite causes increased expression of VEGF in vascular endothelial cells by a process that requires the activation of STAT3.

Inhibition of NAD(P)H oxidase activity blocks vascular endothelial growth factor overexpression and neovascularization during ischemic retinopathy.

Because oxidative stress has been strongly implicated in up-regulation of vascular endothelial growth factor (VEGF) expression in ischemic retinopathy, we evaluated the role of NAD(P)H oxidase in causing VEGF overexpression and retinal neovascularization. Dihydroethidium imaging analyses showed increased superoxide formation in areas of retinal neovascularization associated with relative retinal hypoxia in a mouse model for oxygen-induced retinopathy. The effect of hypoxia in stimulating superoxide formation in retinal vascular endothelial cells was confirmed by in vitro chemiluminescence assays. The superoxide formation was blocked by specific inhibitors of NAD(P)H oxidase activity (apocynin, gp91ds-tat) indicating that NAD(P)H oxidase is a major source of superoxide formation. Western blot and immunolocalization analyses showed that retinal ischemia increased expression of the NAD(P)H oxidase catalytic subunit gp91phox, which localized primarily within vascular endothelial cells. Treatment of mice with apocynin blocked ischemia-induced increases in oxidative stress, normalized VEGF expression, and prevented retinal neovascularization. Apocynin and gp91ds-tat also blocked the action of hypoxia in causing increased VEGF expression in vitro, confirming the specific role of NAD(P)H oxidase in hypoxia-induced increases in VEGF expression. In conclusion, NAD(P)H oxidase activity is required for hypoxia-stimulated increases in VEGF expression and retinal neovascularization. Inhibition of NAD(P)H oxidase offers a new therapeutic target for the treatment of retinopathy.

Vascular endothelial growth factor and diabetic retinopathy: role of oxidative stress.

Retinal neovascularization and macular edema are central features of diabetic retinopathy, a major cause of blindness in working age adults. The currently established treatment for diabetic retinopathy targets the vascular pathology by laser photocoagulation. This approach is associated with significant adverse effects due the destruction of neural tissue and is not always effective. Characterization of the molecular and cellular processes involved in vascular growth and hyperpermeability has led to the recognition that the angiogenic growth factor and vascular permeability factor VEGF (vascular endothelial growth factor) play a pivotal role in the retinal microvascular complications of diabetes. Thus, VEGF represents an important target for therapeutic intervention in diabetic retinopathy. Agents that directly inhibit the actions of VEGF and its receptors show considerable promise, but have not proven to be completely effective in blocking pathological angiogenesis. Therefore, a better understanding of the molecular events that control VEGF expression and mediate its downstream actions is important to define more precise therapeutic targets for intervention in diabetic retinopathy. This review highlights the current understanding of the process by which VEGF gene expression is regulated and how VEGF's biological effects are altered during diabetes. In particular, cellular and molecular alterations seen in diabetic models are considered in the context of high glucose-mediated oxidative stress effects on VEGF expression and action. Potential therapeutic strategies for preventing VEGF overexpression or blocking its pathological actions in the diabetic retina are considered.

Vascular endothelial growth factor and diabetic retinopathy: pathophysiological mechanisms and treatment perspectives.

Retinal neovascularization and macular edema are central features of diabetic retinopathy, the major cause of blindness in the developed world. Current treatments are limited in their efficacy and are associated with significant adverse effects. Characterization of the molecular and cellular processes involved in vascular growth and permeability has led to the recognition that the angiogenic growth factor and vascular permeability factor vascular endothelial growth factor (VEGF) plays a pivotal role in the retinal microvascular complications of diabetes. Therefore, VEGF represents an exciting target for therapeutic intervention in diabetic retinopathy. This review highlights the current understanding of the mechanisms that regulate VEGF gene expression and mediate its biological effects and how these processes may become altered during diabetes. The cellular and molecular alterations that characterize experimental models of diabetes are considered in relation to the influence of high glucose-mediated oxidative stress on VEGF expression and on the mechanisms of VEGF's actions under hyperglycemic induction. Finally, potential therapeutic strategies for preventing VEGF overexpression or blocking its pathological effects in the diabetic retina are considered.