Elevated homocysteine and C-reactive protein levels independently predict worsening prognosis after stroke in Chinese patients

Increased plasma total homocysteine (tHcy) and high sensitivity C-reactive protein (hsCRP) levels are independent risk factors for cardiovascular disease.However, the predictive value of tHcy in combination with hsCRP in patients with stroke is not known.To determine the relationship between tHcy and hsCRP, we enrolled 291 patients with first-onset stroke (196 ischemic and 95 hemorrhagic).Plasma tHcy and hsCRP levels were measured and subsequent vascular events and deaths were determined over a 5-year period.Using the arbitrary cutoff for tHcy (<18 μmol/L and ≥18 μmol/L) and hsCRP (<1 mg/L, 1-3 mg/L and >3 mg/L), the patients were divided into 6 groups.Survival analysis showed that the probability of death or new vascular events during a 5-year follow-up increased according to tHcy and hsCRP levels (P<0.01).The relative risk (RR) of death or new vascular events was 4.67 (95% CI, 1.96 to 11.14, P=0.001) in patients with high tHcy (≥18 μmol/L) and hsCRP (>3 mg/L) compared with those with low tHcy (<18 μmol/L) and hsCRP (<1 mg/L).The increased tHcy level (≥18 μmol/L) combined with increased hsCRP level (>3 mg/L) was still significantly associated with the risk of death or new vascular events (RR, 4.10, 95% CI, 1.61 to 10.45, P=0.003) even when adjusted for other risk factors at inclusion.The combination of increased tHcy and hsCRP levels had a stronger predictive value than increased hsCRP alone or increased tHcy level alone.Further studies are required to evaluate the potential decrease in risks associated with lowering both Hcy and hsCRP levels in patients that present with both increased tHcy and hsCRP.     

Mortality and recurrence of vascular disease among stroke patients treated with combined TCM therapy

OBJECTIVE:To confirm the long-term outcomes of stroke patients and determine predicting factors for death,recurrence of vascular events and poor outcome(either recurrence or death) after the use of combined TCM therapy.METHODS:This was a retrospective hospital-based cohort study and was performed in the First Affiliated Hospital of Tianjin University of Traditional Chinese Medicine in Tianjin.All subjects with stroke consecutively admitted to an inpatient ward of the Acupuncture Department from January 1,2008,to December 31,2008 were retrospectively followed through one year.The main outcomes were either a recurrence of vascular events,,mortality or both.Risk factors were recorded from medical records.Multivariate regression models were used to analyze predictors.The following independent variables were used:age,gender,hypertension,ischemic heart disease,atrial fibrillation,diabetes mellitus,carotid arterial lesions and history of stroke.RESULTS:Four-hundred and five patients were included.The 1-year mortality rate was 11.11%.23.70% of the patients had a recurrent vascular event,and 30.86% suffered a poor outcome.Multiple logistic regression analysis found that previous stroke,and advanced age were predictors of death within one year,Recurrence of vascular events was associated with carotid arterial lesions,history of diabetes and previous stroke.Long-term poor outcome was predicted by advanced age,history of diabetes,and previous stroke.CONCLUSION:Age,previous stroke,carotid arterial lesions and diabetes history seem to have different impacts on the three outcomes within one year.Our findings provide important data for planning future hospital register studies of stroke patients in TCM hospitals.     

Comparison of Aspirin and Naoxintong Capsule (脑心通胶囊) with Adjusted-Dose Warfarin in Elderly Patients with High-Risk of Non-Valvular Atrial Fibrillation and Genetic Variants of Vitamin K Epoxide Reductase

Objective: To compared the therapeutic effect of a Chinese patent medicine Naoxintong Capsule(脑心通胶囊, NXT) and aspirin with adjusted-dose warfarin in Chinese elderly patients(over 65 years) with nonvalvular atrial fibrillation(NVAF) and genetic variants of vitamin K epoxide reductase(VKORC1), who are at high-risk of thromboembolism. Methods: A total of 151 patients, with NVAF and AA genotype of VKORC1-1639(a sensitive genotype to warfarin) and a CHA2 DS2-VASc clinical risk score of 2 or above, were chosen for this study. Patients were randomized into two groups and orally treated with a combination of aspirin(100 mg/day) and NXT(1.6 g thrice a day) or adjusted-dose warfarin [international normalized ratio 2.0–3.0). The primary end points including ischemic stroke and death as well as the secondary end points including hemorrhage events were followed up for at least 1 year. Results: Baseline clinical data and the rates of primary end points were similar between groups. However, the rate of serious bleeding(secondary event) in the combination therapy group was lower than that in the adjusted-dose warfarin group(0% vs. 7.9%, odds ratio: 0.921, 95% confidence interval: 0.862–0.984, P=0.028). Conclusions: Aspirin combined with NXT and warfarin displayed comparable rates of primary end point including ischemic stroke and all-cause death during the 1-year follow-up. However, as compared with warfarin, the combination therapy reduced the rate of serious bleeding. Therefore, aspirin combined with NXT might provide an alternative pharmacotherapy in preventing ischemic stroke for elderly patients with NAVF who cannot tolerate warfarin.(No. ChiC TR-TRC-13003596)     

Clinical study of PTAS therapy for patients with ischemia cerebrovascular disease caused by artery stenosis

The curative efficacy of percutaneous transluminal angioplasty and stenting(PTAS) in the treatment of patients with ischemia cerebrovascular disease caused by artery stenosis was explored.The clinical data of 111 patients with ischemia cerebrovascular disease receiving PTAS in Guangdong Province General Hospital from Aug.2007 to Nov.2009 were retrospectively analyzed.In total 132 stents were implanted in the 111 patients.The mortality and rate of neural and non-neural complications were assessed perioperatively.Outcomes [including the frequency of transient ischemic attack(TIA),stroke,or death from vascular diseases) were assessed after operation.NIHSS rating was performed in all cases before and at first week,6th month and 12th month after the operation.The PTAS success rate was 100%.The degree of stenosis was reduced after PTAS.The total complication rate during perioperative period was 15.3%(the rate of neural complications was 3.6%).Sixty-seven patients were followed up.Three patients(4.48%) developed cerebrovascular events within 1 month,containing one case of TIA,one case of ipsilateral mild stroke and one case of contralateral mild stroke.No severe stroke or death was observed.During a follow-up period of 12 months 7 patients had cerebrovascular events(10.44%),including 2 cases of ipsilateral TIA(2.99%),2 cases of ipsilateral mild stroke and 2 cases of contralateral mild stroke(2.99%),one case of severe stroke(1.49%).In 13 patients receiving DSA re-examination one year after PTAS,2 patients(15.38%) had in-stent restenosis.NIHSS scores were obviously decreased during a follow-up period as compared with those pre-operation(P<0.05).It was concluded that PTAS could significantly alleviate the neural function deficit of the patients with ischemia cerebrovascular disease.The success rate of PTAS was high,and the rate of complications was lower and the clinical outcomes were satisfactory.PTAS is a safe and effective therapeutic method,though the long-term outcomes need further study.     

Atherosclerosis, vascular aging and therapeutic strategies

<正>With the arrival of the era of global population aging,we strive for healthy aging and a healthy senior life rattier than simple prolongation of the physical age.For the past 50 years,cardiovascular diseases (CVD) have been the most common cause of death among the elderly people globally.In China,there has been an exponential increase in the incidence of heart disease and stroke in the elderly population.Atherosclerosis is the pathological change in the coronary artery disease,stroke,and peripheral vascular disease.Despite the significant benefit demonstrated,control of classic risk factors alone,such as lifestyle change or drug therapy, was shown to have limitations in reducing the incidence of cardiovascular events.Vascular aging has been shown to be an important independent predictor of CVD events.Interventions targeting vascular aging have emerged as a new paradigm in conjunction with control of risk factors for the prevention of CVD.Vascular aging and atherosclerosis are two distinct pathological changes and difficult to distinguish clinically.Recent research with Chinese medicine(CM) has shown encouraging observations,linking the clinical benefit of delaying vascular aging and treating atherosclerosis.These results demonstrate great potential of CM in the prevention and treatment of CVD.     

Dyslipidemia and Outcome in Patients with Acute Ischemic Stroke

Objective To study the relationship between dyslipidemia and outcome in patients with acute ischemic stroke. Methods Data about 2 568 patients with acute ischemic stroke were collected from 4 hospitals in Shandong Province from January 2006 to December 2008. National Institute of Health Stroke Scale （NIHSS） 〉10 at discharge or death was defined as the outcome. Effect of dyslipidemia on outcome in patients with acute ischemic stroke was analyzed by multivariate logistic regression analysis and propensity score-adjusted analysis, respectively. Results The serum levels of TC, LDL-C, and HDL-C were significantly associated with the outcome in patients with acute ischemic stroke. Multivariate logistic regression analysis and propensity score-adjusted analysis showed that the ORs and 95% CIs were 3.013 （1.259, 7.214）/2.655 （1.298, 5.43）, 3.157 （1.306, 7.631）/3.405 （1.621, 7.154）, and 0.482 （0.245, 0.946）/0.51 （0.282, O.921）, respectively, for patients with acute ischemic stroke. Hosmer-Lemeshow goodness-of-fit test showed no significant difference in observed and predicted risk in patients with acute ischemic stroke （chi-square=8.235, P=0.411）. Conclusion Serum levels of TC, LDL-C, and HDL-C are positively related with the outcome in patients with acute ischemic stroke.     

Association of polymorphisms in angiotensin II receptor genes with aldosterone-producing adenoma

This study examined the association of polymorphisms in angiotensinⅡreceptor genes(AT1R and AT2R) with the risk for aldosterone-producing adenoma(APA) in a Chinese Han population.Four polymorphisms including rs5182(573T/C) in exon 4,rs5186(1166A/C) in 3’-untranslated region(3’-UTR) in AT1R gene and rs5194(2274G/A) in 3’-UTR,rs1403543(1675G/A) in intron 1 in AT2R gene were detected in 148 APA patients and 192 normal subjects(serving as control) by using a MGB-Taqman probe.The distribution of genotypes of each locus was in accordance with Hardy-Weinberg Equilibrium(HWE) in the APA and control groups(P>0.05).The allele A frequency at rs5194 was significantly higher in the APA group(0.49) than in the control group(0.35)(χ2=12.08,P=0.001).Subjects with homozygotic genotype AA and heterozygotic genotype GA were at an increased risk for APA as compared to those with GG genotype(OR=2.66,95% CI=1.45-4.87;OR=1.67,95% CI=1.02-2.74).Furthermore,rs5194 single-nucleotide polymorphism(SNP) at AT2R gene was significantly associated with APA in additive(OR=1.64,95% CI=1.21-2.20,P=0.001),dominant(OR=1.94,95% CI=1.23-3.06,P=0.003),and recessive model(OR=2.01,95% CI=1.17-3.45,P=0.01).It was concluded that rs5194 polymorphism at AT2R gene was associated with the risk for APA,which may constitute a genetic marker of APA.     

GENE POLYMORPHISM OF APOLIPOPROTEIN E AND CORTICAL CEREBRAL INFARCTION

The polymorphism of apoE, an important determinant in the lipid metabolism, may be related to the atherosclerotic events. In this study, the apoE genotypes, by means of PCR-AFLP assay in 42 patients with cortical cerebral infarction and 85 healthy controls were detected. The results showed that the apoE ε 3/3 genotypa was more prevalent in the controls (55.29% ), and ε 3/4 genotype was more common in the patients (57.14%). The frequency of ε 4 allele was more predominate in the patients than that in the controls. The ε 3/4 genotype or ε 4 allele was associated with 2.44- or 2. 31-fold increase in the risk of cortical cerebral infarction. Therefore, it was suggested that apoE ε 4 allele is a genetic factor of susceptibility to cerebral infarction, which supported the hypothesis of the association between apoE polymorphism and atherosclerotic cardio-cerebrovascular disorders.     

A Randomized Trial to Assess the Long-term Safety of NeuroAiD among Caucasian Patients with Acute Ischemic Stroke

Objective：To assess the long-term（up to 6 months）safety profile of a 3-month regimen of NeuroAiD for acute ischemic stroke.Methods：A total of 190 patients with acute ischemic stroke were identified for eligibility in a randomized,double-blind,placebo-controlled clinical trial,of which 150 patients allocated to either receiving NeuroAiD（80 cases）or placebo（70 cases）were analyzed after dropouts due to absence of baseline data,early death,or noncompliance.Both groups received treatment for three months and followed up for another three months after the completion of the treatment.Occurrence of clinical adverse events and laboratory parameters were assessed at 1 month,3 months（while under treatment）and 6 months（3 months after the completion of treatment）.Statistical comparisons between groups were performed using chi-square test or t-test whenever appropriate.Results：The two groups had comparable baseline characteristics.Mild nausea was more commonly reported in patients taking NeuroAid compared with placebo（P=0.01）,of which 9 out of10 were observed only during the first month of treatment.However,none of the adverse events reported were considered severe or required discontinuation of the study drug.There was no significant change observed in mean arterial blood pressure,haemoglobin,renal and liver laboratory parameters during treatment with NeuroAid and up to 3 months after completion of a 3-month regimen.Conclusion：NeuroAiD is safe and does not affect hematologic,hepatic,and renal functions during and long after completion of treatment.     

The association analysis of FcεRⅠβ with allergic asthma in a Chinese population

Objective To investigate the link between the polymorphism of -109 and Glu237 in the high-affinity IgE receptor β （FcεRⅠβ） gene and susceptibilty to allergic asthma in a Chinese population.Method Blood samples from 216 allergic asthma patients and 198 age- and sex-matched controls were studied. A-109C/T and a coding variant Glu237Gly in FcεRⅠβ were detected with polymerase chain reaction-restriction fragment length polymorphism(PCR-RFLP). Results The genotype frequencies were 0.403 for -109T/T, 0.491 for －109T/C and 0.106 for －109C/C in allergic asthma in a Chinese population. No significant difference in the distribution of －109C/T polymorphism was found between allergic asthma subjects and healthy controls, however, homozygosity for the －109T allele was associated with increased total plasma IgE levels in subjects with allergic asthma (F=4.020,P&lt;0.05). The allele frequency of Gly237 in the patients and control was 0.236 and 0.136 respectively. There was a significant association between the Gly/Gly genotype and allergic asthma. Among allergic asthma patients Gly237 was significantly associated with high IgE levels.Conclusions These results suggest that the Gly237 variant of the FcεRⅠβ gene is involved in the development of allergic asthma. The-109C/T and Glu237Gly polymorphisms are two of the genetic factor identified thus far, which affect total plasma IgE levels of allergic asthma patients in a Chinese population.     

Association between fifteen risk factors and progressing ischemic stroke in the Han population of northeast China

Background The mortality and disability associated with progressing ischemic stroke are much higher than general ischemic stroke. This study was conducted to determine the risk factors for progressing ischemic stroke in the Han population of northeast China. Methods A total of 2511 patients with ischemic stroke within 24 hours admitted to Department of Neurology, First Affiliated Hospital of Harbin Medical University were studied, from November 2007 to May 2009. All of the patients were classified into the progressing or non-progressing group according to the scores of the Scandinavian Neurological Stroke Scale. Fifteen putative risk factors were evaluated. The influence of risk factors for progressing ischemic stroke was analyzed with the simple Logistic analysis, the multiple Logistic analysis, and the stepwise Logistic regression model. All the statistical analysis was performed by SAS 9.1. Results Totally 359 （14.3%） patients met the criteria for progressing ischemic stroke. The Logistic analysis showed that age, family stroke history, smoking history, hypertension on admission, a drop in blood pressure after admission to the hospital, high serum glucose on admission, and fever were related to progressing ischemic stroke in the Han population of northeast China. Conclusion People of the ischemic stroke with these factors are more likely to develop progressing ischemic stroke.     

Ninety-day administration of dl-3-n-butylphthalide for acute ischemic stroke: a randomized, double-blind trial

Background DI-3-n-butylphthalide （NBP）, first isolated from the seeds of celery, showed efficacy in animal models of stroke. This study was a clinical trial to assess the efficacy and safety of NBP with a continuous dose regimen among patients with acute ischemic stroke. Methods A randomized, double-blind, double-dummy trial enrolled 573 patients within 48 hours of onset of ischemic stroke in China. Patients were randomly assigned to receive a 14-day infusion of NBP followed by an NBP capsule, a 14- day infusion of NBP followed by aspirin, or a 14-day infusion of ozagrel followed by aspirin. The efficacy measures were Barthel index score and the modified Rankin scale （mRS） at day 90. Differences among the three groups on mRS were compared using X2 test of proportions （with two-sided e=0.05） and Logistic regression analysis was conducted to take the baseline National Institutes of Health Stroke Scale （NIHSS） score into consideration. Results Among the 535 subjects included in the efficacy analysis, 90-day treatment with NBP was associated with a significantly favorable outcome than 14-day treatment with ozagrel as measured by mRS （P 〈0.001）. No significant difference was found among the three groups on Barthel index at day 90. The rate of adverse events was similar among the three groups. Conclusions The 90-day treatment with NBP could improve outcomes at the third month after stroke. The NBP treatment （both intravenous and oral） is safe （ChiCTR-TRC-09000483）.     

The role of high-sensitivity C-reactive Protein,interleukin-6 and cystatin C in ischemic stroke complicating atrial fibrillation

This study examined the role of high-sensitivity C-reactive protein (hsCRP), interleukin-6 (IL-6) and cystatin C in ischemic stroke complicating atrial fibrillation (AF) and the relationship of systemic inflammation with this disease in order to identify AF patients who are at high risk of stroke and need optimal anticoagulant therapy.A total of 103 AF patients, simple (n=75) or complicated by ischemic stroke (n=28), and 112 control subjects were recruited.IL-6 level was detected by using enzyme linked immunosorbent assay.Cystatin C and hsCRP levels were measured by means of a particle-enhanced immunonephelometric assay.The results showed that the AF patients had higher levels of hsCRP (P=0.004), IL-6 (P=0.000), and cystatin C (P=0.000) than control subjects.Plasma hsCRP level was increased in the AF patients with ischemic stroke as compared to the patients with simple AF (P=0.036).The AF patients who had the level of hsCRP exceeding 3.83 mg/L were at a higher risk than those with hsCRP level lower than 3.83 mg/L (P=0.030).After adjusting for other factors, cystatin C remained positively associated with IL-6 (r=0.613) and hsCRP (r=0.488).It was concluded that hsCRP is positively correlated with ischemic stroke complicating AF and may be a risk factor independent of other risk factors for AF.Elevated cystatin C level is also indicative of the increased risk of AF.     

Definition,prediction, prevention and management of patients with severe ischemic stroke and large infarction

Severe ischemic stroke carries a high rate of disability and death. The severity of stroke is often assessed by the degree of neurological deficits or the extent of brain infarct, defined as severe stroke and large infarction, respectively. Critically severe stroke is a life-threatening condition that requires neurocritical care or neurosurgical intervention, which includes stroke with malignant brain edema, a leading cause of death during the acute phase, and stroke with severe complications of...     

Distribution of Hepatitis B Virus Genotypes and Its Clinical Significance in Hubei Province, China

The distribution of hepatitis B virus genotype in Hubei province and its clinical signifi- cance were investigated. HBV genotypes of 276 patients were detected by PCR-microplate sandwich hybrization-ELISA technique. The level of HBV DNA was detected by using PCR-fluorescence quantification test. Among 276 patients, there were 78 cases of HBV asymptomatic carriers, 110 cases of chronic hepatitis B (CHB), 62 cases of severe hepatitis (SH) or liver cirrhosis (LC) and 26 cases of hepatocellular carcinoma (HCC). The genotypes of HBV included C, B, mixtures (B C, B D, C D) and D, accounting for 55.8%, 25.4%, 16.7% and 2.1% respectively. The average level of HBV DNA in genotypes C, B, mixtures and D was 1.20×106, 7.81×104, 3.26×105 and 5.01×104 cop- ies/mL respectively. The ratio of SH, LC and HCC in genotype B, C and mixtures was 20%, 30% and 48% respectively. Statistical analysis revealed the percentage of genotype mixtures infection was sig- nificantly higher than that of genotype B infection. There was no significant difference in the per- centage between genotype B and genotype C or between genotype C and mixtures. The distribution of genotype B, C and mixtures in SH, LC and HCC was significantly different. The frequency of HCC was zero in patients with co-infection. Genotype D was only related with SH and LC. The in- creased ALT could be converted to categorical grades of severity. From mild, moderate to severity, the prevalence of genotype C showed an opposite trend, although no statistically significant differ- ence was observed. The HBeAg positive rate was higher in patients with genotype C infection than in those with genotype B, especially in the patients whose ages were from 31 to 40 years old. Compared with genotype B, genotype C showed a higher HBeAg positive rate in patients with SH and LC. The percentage of SH, LC and HCC was higher in patients with genotype C and mixtures infection. On the contrary, the percentage of genotype B was lower. The HBeAg positive rate in patients with genotype C infection was higher than those with genotype B infection. Genotype C and mixtures may be associated with development of severe liver disease.     

The Relationship of Insulin—receptor Gene Polymorphism to Ischemic Stroke？

Objective To investigate the role of mutation of insulin-receptor(INSR) gene in the development of ischemic stroke.Methods The base-variations at exon 17 and 20 of INSR gene,by means of PCR-SSCP were determined in 68 cases of atherothrombotic cerebral infarction (ACI),81 cases of lacunar infarction(LI) and 62 healthy controls(HC).Results There were 2 alleles of T and C at exon 17 of INSR gene.The prevalence of mutant of t allele in ACI patients was more common than that in the controls.the blood pressure and the parameters of lipid metabolism in the patients with mutant were higher than those in the controls with wild-type gene.However,the correlative analysis showed that the polymorphism of INSR gene was not related statistically to the blood pressure.No base-variation at exon 20 was found in the study.Conclusion The mutation at exon 17 of INSR gene,by promoting the development of atherosclerosis,may participate in the occurrence of ischemic stroke.     

Severe acute respiratory syndrome-associated coronavirus genotype and its characterization

Objective To study the severe acute respiratory syndrome (SARS)-associated coronavirus genotype and its characteristics.Methods A SARS-associated coronavirus isolate named ZJ01 was obtained from throat swab samples taken from a patient in Hangzhou, Zhejing province. The complete genome sequence of ZJ01 consisted of 29 715 bp ( GenBank accession : AY297028, version : gi : 30910859). Seventeen SARS-associated coronavirus genome sequences in GenBank were compared to analyze the common sequence variations and the probability of co-occurrence of multiple polymorphisms or mutations.Phylogenetic analysis of those sequences was done.Results By bioinformatics processing and analysis, the 5 loci nucleotides at Z J01 genome were found being T, T, G, T and T, respectively. Compared with other SARS-associated coronavirus genomes in the GenBank database, an A/G mutation was detected besides the other 4 mutation loci( C: G: C: C/T: T: T: T) involved in this genetic signature. Therefore a new definition was put forward according to the 5 mutation loci. SARS-associated coronavirus strains would be grouped into two genotypes (C:G:A: C: C/T: T: G: T: T), and abbreviated as SARS coronavirus C genotype and T genotype. On the basis of this new definition, the ZJ01 isolate belongs to SARS-associated coronavirus T genotype, first discovered and reported in mainland China. Phylogenetic analysis of the spike protein gene fragments of these SARS-associated coronavirus strains showed that the GZ01 isolate was phylogenetically distinct from other isolates, and compared with groups F1and F2 of the T genotype, the isolates of BJ01and CUHK-Wl were more closely related to the GZ01 isolate. It was interesting to find that two(A/G and C/T) of the five mutation loci occurred in the spike protein gene, which caused changes of Asp to Gly and Thr to lie in the protein, respectively.Conclusion Attention should be paid to whether these genotype and mutation patterns are related to the virus‘ s biological activities,epidemic characteristics and host clinical svmotoms.     

A single nucleotide polymorphism in the human BMP-2 gene (109T>G) affects Smad signaling pathway and the predisposition to ossification of posterior longitudinal ligament of spine

Background Although various systemic and local factors such as abnormal carbohydrate or calcium metabolism, aging and hormonal disturbances have been suggested as causes of Ossification of the posterior longitudinal ligament (OPLL), the etiology of OPLL is not fully understood. The purpose of this study was to investigate whether BMP-2 is a candidate gene to modify the susceptibility of OPLL and the mechanism of ossification in signal transduction. Methods A total of 420 OPLL patients and 506 age- and sex-matched controls were studied. The complete coding sequence of the human BMP-2 gene were analysed through PCR and direct sequencing, all the SNPs were detected and genotype. The BMP-2 expression vectors containing positive polymorphism were constructed and transfected to the C3H10T1/2 cell. The expression of BMP-2 and Smad signal pathway in positive cell clones were detected by western blotting. The ALP activity was detected by quantitative detection kits. Results The frequencies for the 109T>G and 570A>T polymorphisms were difference between the case and control group. The “TG’ genotype in 109T>G polymorphism is associated with the occurrence of OPLL, the frequency of the “G” allele is significantly higher in patients with OPLL than in control subjects (P<0.001). The “AT’ genotype in 570A>T polymorphism is associated with the occurrence of OPLL, the frequency of the “T” allele is significantly higher in patients with OPLL than in control subjects (P=0.005). Western blot analysis showed that the expression of P-Smad1/5/8 protein transfected by wild-type or mutant expression vectors were significantly higher than control groups (P<0.05), but there was no statistical difference in each experimental group (P>0.05). The expression of Smad4 protein transfected by wild-type or mutant expression vectors were significantly higher than control groups (P<0.05), the expression of Smad4 protein transfected by pcDNA3.1-BMP2 (109G) and pcDNA3.1-BMP2 (109G, 570T) were significantly higher than the other experimental groups(P<0.05). The ALP activity increase has been detected in pcDNA3.1-BMP2 (109G) and pcDNA3.1-BMP2 (109G, 570T) transfected cells to 4 weeks after stably transfection. The activity ALP results were (30.56±0.46) nmol/minute/mg protein and (29.62±0.68) nmol/minute/mg protein, respectively. There was a statistical difference compared with the other experimental groups (P<0.05). Conclusions The BMP-2 is the predisposing gene of OPLL. The “TG’ genotype in the 109T>G and the “AT’ genotype in the 570A>T polymorphisms are associated with the occurrence of OPLL. In the intracellular signalling transduction, the 109T>G polymorphism in exon-2 of BMP-2 gene is positively associated with the level of Smad4 protein expression and the activity of ALP. Smad mediated signalling pathway plays an important role during the pathological process of OPLL induced by SNPs of BMP-2 gene.     

Inhibitive effects of glucose and free fatty acids on proliferation of human vascular endothelial cells in vitro

Objectives To investigate the effects of glucose and free fatty acids (FFAs) on the proliferation and cell cycle of human vascular endothelial cells in vitro , and to examine whether the combined presence of elevated FFAs and glucose may cross-amplify their individual injurious effects.Methods Cultured human vascular endothelial cells (ECV304) were incubated with various concentrations of glucose and/or FFAs (palmitate and/or oleate) for 24-96 h. Morphologic alterations were observed using a phase contrast microscope and an electron microscope. Inhibition of proliferation was measured by a colorimetric 3-［4, 5-dimethyl thiazol-2-yl］-2, 5-diphenyltetrazolium bromide (MTT) assay. Cell viability was determined using trypan blue exclusion. Distribution of cells along phases of the cell cycle was analyzed by flow cytometry. Results Glucose 15 or 30 mmol/L, palmitate (PA) 0.25 or 0.5 mmol/L, and oleate (OA) 0.5 mmol/L inhibited proliferation and accelerated death of endothelial cells in a dose-and-time-dependent manner. After treatment with elevated glucose and/or FFAs, the G(0)/G(1) phase cells increased, whereas S phase cells decreased, suggesting that high glucose and/or FFAs mainly arrested endothelial cells at G(0)/G(1) phase. The inhibitive rates of proliferation and population of dead cells in endothelial cells incubated with glucose plus FFAs (glucose 30 mmol/L+PA 0.25 mmol/L, glucose 30 mmol/L+OA 0.5 mmol/L, glucose 30 mmol/L+PA 0.25 mmol/L+OA 0.5 mmol/L) increased more markedly than those treated with high glucose or FFAs (PA and/or OA) alone. Conclusion Both high ambient glucose and FFAs can inhibit proliferation and accelerate death of endothelial cells in vitro. These changes were cross-amplified in the combined presence of high levels of glucose and FFAs.