Relationship Between PM_2.5 Components And Emergency Hospital Visits For Hypertensive Cardio-cerebrovascular Diseases And Its Influence On RAS Genes Expression

Fine Particulate Matter with aerodynamic diameter less then2.5micrometers (PM2.5) is the major component of air pollution. Many epidemiological surveys have shown that the increased concentration of PM2.5could lead to serious cardiovascular events, such as hypertension, or even cerebral hemorrhage. However, the components of PM2.5are so complicated that it is not clear whether there is any correlation between the concentration of each specific PM2.5component and the emergency hospital visits(EHVs) for hypertension. Meanwhile, it is also worth investigating whether different PM2.5component has any effect on the expression of renin-angiotensin system (RAS) genes.Aims1. To investigate the risk effect of PM2.5and its metal components on hypertensive cardio-cerebrovascular diseases induced emergency hospital visits in Changsha, China.2. To clarify the effect of different PM2.5component on the gene expression of vascular endothelial cell ACE-Angll-AT1R and ACE2-Ang(1-7)-Mas axes.Methods1. Daily EHVs for hypertensive cardiovascular and cerebrovascular diseases, including hypertension, hemorrhagic stroke, ischemic stroke, transient ischemic attack (TIA), coronary heart disease, were gathered from the third Xiangya hospital of central south university between June1st to October31st,2009. During the above research period, the local concentrations of fine particulate matter less than2.5micron in aerodynamic diameter (PM2.5) and its metal components were measured in the Surveillance Spots of Central South University, while the data of local particulate matter less than10micron in aerodynamic diameter (PM10), sulfur dioxide (SO2), nitrogen dioxide (NO2), daily temperature, relative humidity, atmospheric pressure and wind velocity were collected from Changsha environmental protection agency. A time-stratified case-crossover design with distributed lag model was used to evaluate associations between ambient air pollutants and hypertensive cardiovascular diseases, and to calibrate the influence of various meteorological factors, such as daily mean temperature, atmospheric pressure and wind velocity. 2. PM2.5samples were collected from non-industrial area of Changsha between October2010and December2011. HUVEC cells were used for in vitro studies and were randomly divided into6groups, including blank control group, low concentration group (PM2.5100μg/ml), median concentration group (PM2.5200μg/ml), high concentration group (PM2.5400μg/ml), ACEI pre-treated group (PM2.5400μg/ml+perindopril10μmol/L) and positive control group (angiotensin I0.01μmol/ml). Each group was treated with corresponding doses of the soluble or insoluble components of PM2.5, while the cell and supernatant fractions were collected24hours after the exposure. Furthermore, HUVEC cells were treated with400μg/ml soluble or insoluble PM2.5component, respectively, and the cell and supernatant fractions were collected at0h.12h.24h and48h. RT-PCR and Western Blot were used to analyze the mRNA and protein expression of ACE, AT1R, ACE2and Mas, respectively. ELISA was used to determine the protein level of AngII and Ang(1-7) in the supernatant.Results1. There were1,027cases of hypertensive cardiovascular diseases induced EHVs collected during the research period, including86cases of hypertension,99cases of hypertensive hemorrhagic stroke,353cases of hypertensive ischemic stroke,242cases of hypertensive TIA, and246cases of hypertensive coronary heart disease. During the research period, the daily average concentration of PM2.5in Changsha was87.79μg/m3. After calibration of the influences of meteorological factors (daily temperature, atmospheric pressure and wind velocity), NO? and SO2, an increase in10μg/m3in PM2.5is associated with hypertension induced EHVs with odds ratios of1.080(95%confidence interval (CI):0.960,1.215), with hypertensive hemorrhagic stroke induced EHVs with odds ratios of1.177(95%confidence interval (CI):1.006,1.376), with hypertensive ischemic stroke induced EHVs with odds ratios of1.040(95%confidence interval (CI):0.953,1.135), with hypertensive TIA induced EHVs with odds ratios of1.028(95%confidence interval (CI):0.901,1.172), and with hypertensive coronary heart disease induced EHVs with odds ratios of1.021(95%confidence interval (CI):0.901,1.156), respectively. The increase in10μg/m3in PM2.5is significantly correlated with hypertensive hemorrhagic stroke induced EHVs (P=0.042). However, there was no statistically significant correlation between daily average PM10concentration and hypertensive cardiovascular diseases induced EHVs (P>0.05).2. An daily average increase of one IQR (Interquartile Range) in Na, K, Ni, Zn, Pb component of PM2.5is associated with hypertensive cerebral hemorrhage induced EHVs with odds ratios of2.993(95%CI:1.176-7.618, P=0.021),2.160(95%CI:1.092-4.271, P=0.027),1.826(95%CI:1.031-3.233, P=0.039),1.568(95%CI:1.015-2.423, P=0.043),1.682(95%CI:1.010-2.800, P=0.046). While other component has no relationship with EHVs for hypertension (P>0.05).3. After HUVEC cells were exposed to the soluble component of PM2.5for24hours, both the mRNA and protein expression levels of ACE and AT1R in low concentration group, median concentration group, or high concentration group were significantly higher than those in controls (P<0.05), and this effect was dose-dependent. However, the protein expression of AngII did not differ between the various groups. On the other hand, when compared with those of controls,24hours exposure to the insoluble component of PM25markedly increased the mRNA and proteins expression of ACE, AT1R, and AngII in low concentration group, median concentration group, and high concentration group in a dose-dependent manner (P<0.05). ACEI pre-treated group showed a lower level of ACE mRNA and protein than that in high concentration group (P<0.05). Moreover, the expressions of ACE, Angll, and AT1R are induced more significantly in high concentration group after insoluble PM2.5component treatment compared to those after soluble PM2.5component treatment (P>0.05).4. HUVEC cells were treated with400μg/ml of either soluble or insoluble component of PM2.5, and the mRNA and protein levels of ACE were found higher at12h,24h, and48h compared to those at Oh, while the peak was reached at24h (P<0.05). Meanwhile, the mRNA and protein expressions of AT1R were induced in a time-dependent manner by the treatment of400μg/ml of either soluble or insoluble component of PM2.5and peaked at48h (P<0.05). At all time points, insoluble PM2.5component induction led to higher expressions of ACE and AT1R (P<0.05), and statistically no different expression of Angll (P>0.05) compared to soluble PM2.5component induction. The protein expression of Angll was markedly increased with the peak reached at24h after treatment of insoluble PM2.5component (P0.05).5.24hours exposure to either soluble or insoluble component of PM2.5led to a significant dose-dependent increase of the mRNA and protein expressions of ACE2, Ang(1-7) and Mas in low concentration, median concentration and high concentration compared to those of blank controls (P<0.05). Mas mRNA level was significantly lower in perindopril pre-treated group than that in high concentration group (P<0.05). There were no statistical differences in the expression of ACE2, Ang(1-7), and Mas between PM2.5soluble component induction and PM2.5insoluble component induction (P>0.05).6. After treatment with400μg/ml of either soluble or insoluble PM2.5component in HUVEC cells, the mRNA and protein levels of ACE2, Ang(1-7) and Mas were significantly increased in a dose-related manner at12h,24h, and48h compared to those at Oh, with the peak reached at48h (P<0.05). At all time points, insoluble PM2.5component induced statistically significant increase in the expressions of ACE2and Mas (P<0.05). However, the induction of Ang(1-7) expression by insoluble PM2.5component or that by soluble PM2.5component showed no statistical difference (P>0.05).Conclusions1. The elevated PM2.5in urban gaseous air pollution was associated with increased hypertensive hemorrhagic stroke induced EHVs in Changsha, China. But there was no correlation between PM10concentration and the EHVs for hypertension.2. The concentration of Na, K, Ni, Zn, Pb in PM2.5is related to the EHVs for hypertensive hemorrhagic stroke.3. Both the soluble and insoluble components of PM2.5could increase the expression of ACE-AngII-ATIR and ACE2-Ang(1-7)-Mas axes genes in HUVEC cells, and the effect is more significantly after insoluble PM2.5components induction.4. ACE is the initial gene of RAS after PM2.5exposure. While the ACEI could decrease the over-activation of RAS genes induced by the insoluble components of PM2.5.