The Study On The Relationship Between Cigarette Smoking And Diabetes Mellitus And Its Related Mechanism

Diabetes mellitus is a complex chronic endocrine and metabolic disease,mainly characterized by persistent hyperglycemia.The etiology of diabetes is not yet fully elucidated,it has proved that lots of risks were correlated to diabetes,such as heredity,autoimmune,lifestyle and environmental factors.The development of diabetes involve in a number of physiological processes of mutiple organs and systems,including insulin secretion defects of beta-cell of islet and the reduced insulin sentivity of peripherl tissue(such as skeletal muscle and adipose tissue).A nationwide survey in 2010 showed that the prevalence of diabetes in China has reached to 11.6%,therefore,the estimated diabetes cases up may be as many as 113 million people.Smoking is one of the major risk factors for four major chronic noncommunicable diseases,including diabetes mellitus,cardiovascular and cerebrovascular diseases,cancer and chronic respiratory diseases.China has a large population of smokers.The latest monitoring showed that there were 396 million adult smokers,which means that cigarette smoking present a severe epidemic situation in China.The study on the relationship between cigarette smoking and diabetes has been the focus recently.Epidemiological research maintains heat degree recently,however mechansim research has nothing good or unusual to report.MAPKs signal pathway is an important signal transduction pathway of transmitting extracellular signaling from the cell membrane to the cytoplasm and finally get into the nucleus of DNA.It is involved in the regulation of many biological processes such as proliferation,growth,differentiation and death.In this article,epidemiology study is based on the monitor of adult chronic diseases and risk factors in Jiangsu Province,and the object of study is male,to explore the relationship between cigarette smoking and diabetes.Meanwhile,in vitro,the affects of cigarette smoke extract on pancreatic beta-cell and the role of MAPKs signal pathway in it was also observed,and in order to explore the partial molecular mehansims among cigarette smoking and diabetes.Part I Epidemiological study of the association between smoking and diabetes Objective:To explore the association between cigarette smoking and diabetes mellitus.Methods:Based on 1047 adult male surveyed by the monitor of adult chronic diseases and risk factors in Jiangsu Province,the association between cigarette smoking and the related indices of diabetes was analyzed by univariate analysis and multivariate analysis.The relationship between cigarette smoking and diabetes was analyzed by adjusting the confounding factors in the binary logistic regression model.Results:The prevalence of diabetes was significantly increased in heavy smoking men.The prevalence of diabetes was 12.80%in the>20 cigarettes daily smokers and the prevalence of diabetes in the>30 pack-years smokers was 14.54%,which were all higher than that of non-smokers.The insulin resistance index(HOME-IR of 0～15 pack-years smokers was significantly higher than that of non-smokers(2.09 vs 1.70,P=0.002),and HOME-β index of>30 pack-years smokers was significantly lower than that of non-smokers(53.71 vs 67.46,P=0.003).When age and BMI were not stratified,>20 cigarettes daily smoking was significantly associated with diabetes(AOR=1.730,95%Cl:1.144-2.615,P=0.009),and>30 pack-years smoking was also significantly associated with diabetes(AOR=1.738,95%CI:1.101-2.742,P=0.018).After stratification of age and BMI,≥20 cigarettes daily smoking was signficantly postive associated with diabetes(AOR= 1.891,95%CI:1.039-3.442,P=0.037)in age ≥ 55 years group,besides,0～20 cigarettes daily smoking was significantly negative associated with diabetes(AOR= 0.389,95%CI:0.155-0.976,P=0.044)in age<55 years group;in BMI<25 kg/m2 group,the risk of diabetes was signficantly higher not only in>20 cigarettes daily smokers(AOR=2.464,95%CI:1.290-4.707,P=0.006),but also in>30 pack-years smokers(AOR=1.991,95%CI:1.003-3.951,P=0.049)Conclusion:Heavy smoking was signicifant associated with diabetes in male,especially in age>55 years and BMI<25 kg/m2 people.Part Ⅱ p38MAPK mediate beta-cell damage induced by cigarette smoke extractObjective:To explore the effects of cigarette smoke extract on the proliferation,apoptosis and function of islet cells and the expression of MAPKs signal pathway proteins and their roles.Methods:(1)MIN6 cells were exposed to different concentrations of cigarette smoke extract(0、20、50、100、200 μg/mL)for 24h,and then the effects of cigarette smoke extract on the morphology of MIN6 cells were observed under inverted microscope.The effects of cigarette smoke extract on cell proliferation were detected by CCK-8 kit.(2)After MIN6 cells were treated by(0、20、50、100 μg/mL)for 24h,Annexin V-FITC double staining methods was used to detect the effect of cigarette smoke extract on apoptosis of MIN6 cells.Western blots were used to detect the levels of apoptosis-related protein cleavd caspase-3 and anti-apoptotic protein Bcl-2.And the effects of cigarette smoke extract on insulin secretion and intracellular ATP level in MIN6 cells were detected by ELISA and Luminometer.(3)After MIN6 cells were treated by different concentrations of cigarette smoke extract(0、20、50、100μg/mL)for 24h,we detect the expression levels of total proteins and phosphorylation proteins of MAPKs signal pathway(JNK、p-JNK、ERK、p-ERK、p38MAPK、p-p38MAPK)by western blots.Finally,we were going to investgate the role of p38MAPK and JNK by using their specific inhibition,and then the effects of cigarette smoke extract on apoptosis and insulin secetion in MIN6 cells were observed by western blots.Results:(1)With the increasing of the concentration of cigarette smoke extract,abnormal morphological change of MIN6 cells was observed.Cells were elongated and spindle-like changes were observed under high-concentration cigarette smoke extract,and lots of cell debris were found,accompanied by decreased cell viability.When MIN6 cells were treated by 100μg/mL and above cigaratte smoke extract,the cell viability was significantly lower than negative control group(P<0.05).(2)Cigarette smoke extract induced the apoptosis of MIN6 cells,and the expression levels of Bcl-2 were decreased obviously,while the expression levels of cleaved caspase-3 were increaed obviously,and both two protein levels at the concentration of 50μg/mL and 100μg/mL cigarette smoke extract was significantly different from the control.After treatment of cigarette smoke extract,glucose induced insulin secretion levels and intracelluar ATP levels and the expression levels of PDX-1 mRNA were all declined obviously compared to the control(P<0.05).(3)Western blots revealed that when MIN6 cells were treated by cigarette smoke extract,the phosphorylated p38MAPK and JNK were activated obviously,and the levels of p-p38MAPK and p-JNK were significantly increased compared to the control at the concentration of 500μg/mL and above.After pretreatment with p38MAPK selective inhibitor,cigarette smok extract induced apoptosis and decreased insulin secretion of MIN6 cells were restrained.