Expression And Significance Of IL-22 And Its Inhibitory Protein IL-22bp In Peripheral Blood Of Patients With Type 2 Diabetes Mellitus

Objective:To investigate the changes and mechanism of interleukin-22(IL-22)and interleukin-22 binding proteins(IL-22BP)in peripheral blood of patients with type 2 diabetes mellitus.To understand the effect of IL-22 on the proliferation and apoptosis of islet beta cells under diabetic conditions.Methods:90 patients with T2DM admitted to the Department of Endocrinology,Jingzhou Hospital,from September 2017 to September 2018 were selected as the T2DM group.In addition,40 patients from the medical examination center of the hospital were selected as the control(NC)group.Collect general demographic data of patients,such as gender,age,height,weight,waist circumference,hip circumference,course of disease,calculate body mass index(BMI),waist-to-hip ratio(WHR).Enzyme-linked immunosorbent assay was used to detect the content of IL-22 and IL-22BP in each group,and to determine the general biochemical indicators,such as fasting blood glucose(FPG),total cholesterol(TC),triacylglycerol(TG),low-density lipoprotein cholesterol(LDL-C),glycated hemoglobin(HbAlC),fasting blood insulin(FINS),determination of urinary albumin and creatinine,urinary albumin creatinine ratio(UACR)was calculated and homeostasis model homa-IR was used to assess insulin sensitivity.Patients in the T2DM group were screened for diabetic nephropathy,diabetic retinopathy,diabetic peripheral neuropathy and diabetic peripheral vascular disease.MIN6 cells in logarithmic growth stage were divided into control group,diabetic condition+IL-22 group and diabetic condition+control group.50ng/ml IL-22 was used.CCK-8 assay was used to detect cell proliferation.Annexin V-FITC/PI double staining was utilized to measure the apoptosis rate of MIN6 cells.Western blot was used to analyse the expression levels of Egr-1,PCNA,FasL,Bcl-2 and Bax.Results:1.Comparison between general demographic data and laboratory results(1)Comparison of IL-22 and IL-22BP:compared with the normal control group,the expression level of IL-22 in the T2DM group decreased[12.39(7.7,18.49)vs19.55(15.89,28.65)](P<0.001),while the level of IL-22BP increased[3.65(2.81,5.66)vs3.04(2.63,3.39)](P<0.01).(2)Comparison of other clinical features:Compared with the control group,BMI and WHR in the T2DM group increased[24.27± 3.08vs22.42± 2.54;0.91(0.88,0.96)vs0.86(0.83,0.89)],(P<0.01),LDL-c,FPG and HbA1C in T2DM group were all higher than those in control group[2.28(1.87,2.89)vs2.07(1.8,2.3);10.0±3.26vs5.05±0.62;8.8(7.08,11.13)vs5.15(4.7,5.48)](P<0.001).There was no significant difference in age,gender,TC and TG between T2DM group and control group.2.Correlation analysis(1)Correlation analysis of IL-22 and general indicators:Spearman correlation analysis showed that the expression level of IL-22 in peripheral serum of patients in T2DM group was negatively correlated with ABI and IL-22BP,Which was positively correlated with WHR,FPG,HbA1C,HOMA-IR.Spearman correlation analysis showed that the level of IL-22 in the peripheral blood of patients in the T2DM group was negatively correlated with ABI and IL-22 BP(p=-0.312,-0.273,P<0.01),and positively correlated with WHR,FPG,HbA1C,and HOMA-IR(p=0.209,0.282,0.243,0.241,P<0.05).(2)Correlation analysis of IL-22BP and general indicators:Spearman correlation analysis showed that IL-22BP in peripheral serum of T2DM group was negatively correlated with HbA1C and LDL-C(p＝-0.290,-0.208,P<0.05).3.Logistic regression analysis of plasma IL-22,IL-22BP levels and type 2 diabetes incidence:With or without diabetes as the dependent variable,IL-22,IL-22BP,FPG,WHR,BMI,LDL-C,gender As an independent variable,logistic regression analysis showed that IL-22(OR=0.891,95%CI:0.812-0.978,P<0.05),IL-22BP(OR=1.930,95%CI:1.028-3.626,P<0.05),FPG(OR=3.966,95%CI:2.060-7.633,P<0.001)was a risk factors for T2DM patients.4.The relationship between plasma IL-22,IL-22BP levels and chronic complications in patients with type 2 diabetes:In patients with diabetic nephropathy,compared with patients without diabetic nephropathy,serum IL-22 expression level decreased and IL-22BP expression level increased[10.09(5.50,16.32)vs13.54(9.46,20.97);4.84(3.07,10.0)vs3.22(2.76,5.01)](P<0.05).There was no significant difference in the expression levels of IL-22 and IL-22BP in patients with or without diabetic retinopathy,diabetic peripheral neuropathy and diabetic macrovascular disease(P>0.05).5.The results of cck-8 experiment showed that the A450 light absorption value of islet β cells MIN6 of control group,diabetes condition control group and IL-22 group were 0.86±0.02,0.44±0.03 and 0.68±0.01,respectively.Compar ed with the control group,the proliferation rate of β cells in the diabetic condit ion control group decreased significantly.While β cells treated with IL-22,the proliferation rate in the diabetic condition was significantly improved.Western blot analysis showed that Egr-1 protein level was respectively 0.10±0.02,0.72±0.07 and 0.35±0.09 in three groups,and PCNA expression was 0.59±0.09,0.08±0.01 and 0.26±0.01,respectively.Egr-1 expression was increased and PCNA pro tein level was decreased in the diabetic condition control group related to contr ol group.After IL-22 treatment,the expression of Egr-1 was significantly decre ased and the expression of PCNA was significantly increased(P<0.01).6.The apoptosis rate of MIN6 cells in the control group,the diabetic control group and the IL-22 group was respectively 3.76%±0.11%,28.00%±1.55%and 9.94%%±1.01%,.Compared with the control group,the β cells apoptosis rate in the diabetic condition control group was significantly increased and the apoptosis of isletβ cells could be recovered to a certain extent,and the difference was statistically significant(P<0.01)after the treatment with IL-22.Western blot analysis showed that the expression of pro-apoptotic protein Fas-L in the control group,diabetes control group and IL-22 group was 0.06±0.01,0.43±0.04 and 0.21 ±0.04,Bax protein levels were 0.06±0.01,0.49±0.02 and 0.22±0.02,and the expressions of anti-apoptotic protein Bcl-2 were respectively 0.72±0.10,0.10±0.03 and 0.30±0.06 Compared with the control group,the expression of pro-apoptotic proteins FasL and Bax in β-cells in the diabetic condition control group was significantly increased,and Bcl-2 expression was decreased.After IL-22 treatment,the expression of β cells apoptosis related protein FasL and Bax was inhibited,while Bcl-2 expression significantly increased,and the differences were statistically significant(P<0.01).Conclusions:1.The levels of IL-22 expression was decreased and IL-22BP expression was increased in type 2 diabetic patients.IL-22 expression was related to ABI,WHR,HOMA-IR,while IL-22BP expression was related to HbA1C,LDL-C.IL-22 and IL-22BP may be involved in the development of type 2 diabetes.2.Serum IL-22 expression level decreased and IL-22BP expression level increased in patients with diabetic nephropathy,which may be related to the occurrence of diabetic nephropathy.3.IL-22 could improve the proliferation and inhibit apoptosis of islet cells under diabetic conditions.