The Cardiactoxicity Effect And Molecular Mechanism In Response To Atmospheric PM_2.5 Inhalation

In the past three decades,the health problems caused by air pollution have been the focus of the world and the hotspot of the scientific research.As the main component of smog,atmospheric fine particulate matter PM_2.5,that is,particulate matter with aerodynamic diameter less than or equal to 2.5 μm,is the main indicator of its health effects.PM_2.5 is mainly derived from daily power generation,chemical production,automobile exhaust emissions,etc.,with large surface area and complex composition,these complex atmospheric fine particles reacts with gaseous pollutants such as sulfur oxides（SOx）and nitrogen oxides（NOx）to generate the secondary particles in the atmosphere,which has various effects on human health.Numerous epidemiological and experimental studies have shown that exposure to PM_2.5 not only causes damage to the respiratory system,inducing lung diseases,but also affects the nervous system and the cardiovascular system（CVS）through blood circulation,leading to cardiovascular and cerebrovascular diseases,especially in children and elderly.Recent studies suggest that,besides traditional directly exposure,fetal exposure may be a process that cannot be ignored,but animal studies and related regulation process are lack.Therefore,based on the source analysis and health risk assessment of heavy metal,this study establish a PM_2.5 exposure-cytotoxicity model to evaluate the seasonal differences in regionally specific PM_2.5 on myocardial cell injury and reveal related biological processes;further focusing on the heart damage induced by PM_2.5 according to the “horizontal” susceptibility of different ages and the “longitudinal” effect of maternal-child inheritance;and then investigates the heart damage caused by combined exposure of atmospheric pollutants.1.As one of the toxic components of PM_2.5,heavy metals have a significant contribution to the morbidity and mortality of cardiovascular diseases（CVDs）due to their non-degradability and bioaccumulation effects.The detection and source analysis of PM_2.5-heavy metals in typical areas is of great significance for studying the effects of cardiac damage.Therefore,in the first part of our study,based on the monitoring of PM_2.5 concentration in Taiyuan,a coal-fired city in northern China,the content of 8 heavy metals（Cr,Ni,Cu,Cd,Pb,Zn,Mn and Co）in PM_2.5 was analyzed by inductively coupled plasma mass spectrometry（ICP-MS）,the enrichment factor（EF）and principal component analysis（PCA）were used to analyze the main source of these heavy metals,and non-cancer risk assessment of these heavy metals were also did.The results showed that the concentration of PM_2.5 and the heavy metals showed the same seasonal trend.The heavy metals from coal combustion contributed the most to the non-carcinogenic risk value,and the risk for children is greater than adults.Based on the significant contribution of heavy metal components in PM_2.5 to CVD morbidity and mortality,and the important role of cardiomyocyte apoptosis in the pathogenesis of CVD,we established an in vitro cell-infected model to investigate the apoptosis from multiple aspects with PM_2.5（10 μg/m L）from different seasons,then explained the related molecular mechanism from the aspects of oxidative stress and inflammatory reaction by exposed cardiomyocytes to winter PM_2.5（0,1,3,and 10 μg/m L）.The results indicated that PM_2.5 can induce season-dependent apoptosis,with increased p53 and bax,and decreased bcl-2 expression,the significant difference occurred in spring and winter,oxidative damage caused by elevated ROS and the inflammatory response characterized by ICAM-1 and i NOS were participated in the molecular regulation of apoptosis.The results of this part provide the experimental evidence for the analysis of heavy metal components in geographical specificity PM_2.5 in order to achieve the effective control of significant contribution components.2.Epidemiological and experimental evidence indicates that PM_2.5 enters the body via breathing and affects the CVS through blood circulation,leading to arrhythmia,hypertension,coronary heart disease,heart failure and even sudden death,especially in the elderly and children.Therefore,it is particularly important to investigate the susceptibility of PM_2.5 exposure on heart injury at different life stages,as well as the in-depth analysis of related molecular regulatory processes.In this study,C57BL/6 mice at different life stages（young（4 weeks old）,adult（4 months old）,middle-aged（10 months old））received PM_2.5 at 3 mg/kg every other day for 4 weeks.Echocardiography was used to investigate the changes of cardiac structure and function of mice at different ages,quantitative reverse transcription-polymerase chain reaction（q RT-PCR）was used to elucidate the susceptibility mechanism of young mice,and the changes of cardiac metabolic profiles in middle-aged mice were analyzed by gas chromatography-mass spectrometer（GC-MS）.The results indicated that compared with adult mice,middle-aged mice were more susceptible to PM_2.5,followed by young children mice.Further molecular regulation studies showed that PM_2.5 exposure induced cardiac hypertrophy by activating the expression of cardiac-specific transcription factors Nkx2.5 and GATA4,further stimulate their downstream target genes ANP,BNP and β-MHC;PM_2.5 exposure significantly affected the energy metabolism,fatty acid metabolism and amino acid metabolic pathway in middle-aged mice.This part of the study provides experimental evidence for the diagnosis of cardiac injury at different life stages under PM_2.5 exposure conditions.3.The theory of fetal origins of adult disease（FOAD）believes that adult diseases may be derived from the changes of the uterine environment during the fetal period,leading to impaired growth and development,and may even increase the risk of illness in adulthood.Previous studies have shown that PM_2.5 can across the placental barrier,further affecting the fetal development.Epidemiological evidence have shown that maternal exposure to PM can cause adverse cardiac outcomes such as left ventricular dysplasia,ventricular septal defect（VSD）,and cardiovascular malformation,resulting in cardiac dysfunction in children and increasing the risk of various CVDs in childhood.However,such researches mainly focuses on the epidemiological studies or biological effects at a certain developmental time point after birth,lack systemic studies tracking postnatal cardiac development and investigating related molecular regulation mechanisms at different time points.Therefore,in this part,pregnant C57BL/6 mice received 3 mg/kg PM_2.5 or vehicle control every other day throughout the gestation period.Cardiac function and biological testing of offspring were investigated on postnatal days（PNDs）1,7,14,and 21 by echocardiography,histopathological observations,hematoxylin and eosin（H&E）staining,and q RT-PCR analysis.Our results found that maternal PM_2.5 exposure can induce gender-dependent cardiac hypertrophy and cardiac dysfunction in PND1 and PND7 offspring mice,accompanied by an increase in the m RNA expression of ANP,BNP and β-MHC,only female showed the significant differences.In addition,the cardiac specific transcription gene GATA4 that activates the above marker factors is also highly expressed in the exposed PND1 and PND7 female mice,and the micro RNA（mi R-200c-3p）that regulates GATA4 expression also shows a downward trend.This result provides systematic evidence for the study of maternal PM_2.5 exposure and gender-susceptible heart injury,and suggests that micro RNA may be involved in this regulation.4.Air particulate pollution is a serious environmental health problem and can lead to a variety of CVDs.According to the current research,PM_2.5,SOx and NOx are the most common pollutants.Study of atmospheric pollutants co-exposure as much as possible is of great significance for the overall control of pollutants,as well as the prevention and diagnosis of CVDs.In this part,based on the secondary concentration limits of China’s ambient air quality standards,we designed two different concentration groups of PM_2.5,SO₂ and NO₂ for C57BL/6 mice(dynamic inhalation of 0.5 mg/m3 SO₂ and 0.2 mg/m3 NO₂（6 h/d,28d）,received 1 mg/kg PM_2.5 every other day;or 3.5 mg/m3 SO₂,2 mg/m3 NO₂,and 3 mg/kg PM_2.5 every other day).Based on heart-to-body-weight statistics,BP-2010 A was used to investigate the changes of heart rate（HR）and blood pressure（BP）in mice,and cardiac ultrastructure,inflammatory reaction and endothelial injury were further conducted to investigate the cardiotoxicity.The results showed that co-exposure of PM_2.5,SO₂ and NO₂ could induce the increase of HR and decrease of BP in mice with a dose-dependent manner,accompanied by ultrastructural changes such as mitochondrial membrane damage,cristae rupture and mitochondrial lysis.At the same time,the heart tissue of the exposed group showed a dose-dependent disorder of fiber arrangement,widening of the gap,swelling of the nucleus,inflammatory cell infiltration,and high levels of inflammatory factors such as IL-6,TNF-α,COX-2 and i NOS,and the significant difference were found at the lower concentrations.In addition,the expression of endothelial cell factor ET-1 and e NOS also showed an imbalance.Our results suggest that under the condition of combined atmospheric pollution,biological damage can be caused even if the concentration of single pollutant is at a low,which provides experimental basis for the control of complex atmospheric pollution.In this study,we first established a PM_2.5 exposure-cytotoxicity evaluation model based on heavy metal toxicity components,evaluated the seasonal differences and investigated molecular regulation mechanisms of cardiomyocytes injury induced by geographical specificity PM_2.5.Further explored the susceptibility of cardiac damage induced by PM_2.5 exposure and investigated the related molecular biological processes in mice at different life stages using the whole experimental animal model;in addition,based on the FOAD theory,studied the structural and functional damage,as well as the possible molecular regulation mechanism in offspring exposed by PM_2.5 during pregnancy;finally,conducted a preliminary inquiry of the heart damage in response to PM_2.5,SO₂ and NO₂ co-exposure,providing new ideas for pollution control and health protection in polluted areas.