A Study Of RANKL Protein Expression And Gene Polymorphisms And Bone Mass Abnormalities In Postmenopausal Women With T2DM

Objective:This study investigates the relationship between protein expression levels,gene polymorphisms and mutations of Receptor activator of nuclear factorκB ligand（RANKL）and bone mineral density（BMD）and bone metabolism markers in postmenopausal women with type 2 diabetic mellium（T2DM）,and provides a scientific basis for the prevention and treatment of osteoprosis（OP）in postmenopausal women with T2DM.Methods:1.200 postmenopausal women were enrolled in the study population（48 years≤age≤80 years）and were divided into group A:normal glucose tolerance（NGT）+normal bone mass;group B:NGT+abnormal bone mass;group C:T2DM+normal bone mass;and group D:T2DM+abnormal bone mass according to the results of oral glucose tolerance test（OGTT）and dual energy X-ray（DEXA）;2.The baseline data were obtained at the menopausal transition age,body mass index（BMI）and waist-to-hip ratio（WHR）;the biochemical analyzer（Roche automatic）was used to determine fasting blood glucose（FPG）,triglycerides（TG）,blood Calcium（Ca）and other biochemical parameters of bone metabolism and glucose and lipids;the high pressure liquid chromatograph（HPLC）was used to determine glycated hemoglobin（HbA1c%）;the DEXA was used to determine femoral neck BMD（FN BMD）and lumbar spinal _1-4BMD(LS_1-4BMD);Detecting polymorphisms at the RANKL locus by time-of-flight mass spectrometry.Detecting RANKL protein expression level by ELISA（Enzyme-linked immunosorbent assay）.3.Data were processed by IBM SPSS v.25.0 software.Theχ2 goodness-of-fit Hardy-Weinberg equilibrium（HWE）was used to perform the RANKL genotype concordance test.Data conforming to normal distribution were expressed as mean±standard deviation（（?）±s）,Analysis of variance（ANOVA）was used to compare baseline data and biochemical indexes between groups,otherwise non-parametric tests were used,Spearman correlation analysis for association between bivariates,multiple linear regression analysis for factors influencing LS_1-4BMD and FN BMD,and ROC curves for RANKL protein were plotted to determine their clinical diagnostic value,and P<0.05 was considered a statistically significant difference.Results:1.Compared with group A,the levels of FPG and HbAlc in groups C and D were higher than those in group A{（10.32±2.46,9.38±2.63 vs 5.21±0.75）,（9.05±1.60,8.63±2.02 vs 6.14±1.04）,P<0.01};The levels of LS_1-4BMD and FN BMD in groups B and D were lower than those in group A{（0.93±0.13,0.93±0.07vs 1.16±0.18）,（0.78±0.12,0.77±0.11 vs 1.02±0.24）,P<0.01}.2.Compared with group A,RANKL protein expression concentrations were higher in groups B,C and D（P<0.05）;Spearman correlation analysis,RANKL protein expression level was positively correlated with age,year of menopause and FPG（P<0.001）;and negatively correlated with HDL-C,blood P,LS_1-4BMD and FN BMD（P<0.05）.3.The optimal concentration of RANKL protein for diagnosing OP was 86.17 pg/ml,with a sensitivity of 68.3%and specificity of 83.3%（P<0.001）.4.Comparison of genetic polymorphisms:at the rs9594738 locus,there was a statistical difference in the frequency distribution of genotypes in group D compared with group A（P<0.05）.5.Comparison of gene mutations:1)Bone metabolism indexes:rs9525641 locus,mutant blood Ca and ALP were higher than wild type in groups A and C（P<0.05）;rs1021188 locus,mutant blood Ca was lower than wild type in groups B and C（P<0.05）;2)BMD comparison:rs9594738 locus in group C and in rs9525641 locus in group B mutant LS_1-4BMD was lower than wild type（P<0.05）;mutant FN BMD was higher than wild type in group D at rs9594738 locus（P<0.05）.3)Lipid metabolism index:mutant HDL was higher than wild type in group D at rs9525641 locus（P<0.05）.6.Multiple linear regression analysis showed that high RANKL protein levels,age at menopause and rs9525641 and rs1021188 locus polymorphisms and low TG were risk factors for decreased LS_1-4BMD;high RANKL protein levels,age at menopause and ALP and low blood P and HDL were risk factors for decreased FN BMD.Conclusions:1.Up-regulated expression of RANKL protein and polymorphism of rs9594738,rs9525641 and rs1021188are associated with decreased bone mass in postmenopausal T2DM women in Shihezi.2.Diagnosis of OP in postmenopausal women with T2DM is valuable when the serum RANKL protein concentration is greater than 86.17 pg/ml;3.Mutations in the RANKL genes rs9525641,rs1021188 and rs9594738 loci were associated with abnormal bone mass in this population,and mutations in the rs9525641 locus were associated with disorders of lipid metabolism.