| Steroid-induced osteonecrosis of the femoral head is a non-traumatic osteonecrosis of the femoral head after the use of steroids,also known as avascular necrosis of the femoral head,which is related to high triglyceride,cholesterol,family history of osteonecrosis,coagulation and other high risk factors.After the use of hormones,there may be increased osteoclast production,decreased osteocyte production,muscle atrophy,reduced calcium absorption and other changes,followed by decreased microcirculation,osteoporosis,and finally progressed to pathological fracture.Glucocorticoid is one of the main drugs for the treatment of neuroimmune diseases,and neuroimmune diseases account for at least 2.31% of all primary diseases with SONFH,which brings a heavy burden to society and families,and early prevention and treatment is of great significance.In traditional Chinese medicine,SONFH is classified as "bone arthralgia",which is located in bones and joints,which is characterized by deficiency of qi and blood,deficiency of liver and kidney,spleen,kidney,liver and other viscera,marked as wind,cold,dampness,heat,phlegm and blood stasis.It is considered that it is the result of the interaction between congenital heredity and acquired drugs,and the disease has a specific physical basis.Kidney is the congenital foundation,which corresponds to genetic polymorphism and plays a decisive role in epigenetics,transcription,translation and so on.Spleen and stomach is the acquired foundation,corresponding to diet,climate and other external environment can be affected by epigenetic modification and gene mutation and passed on to the next generation.This is also in line with the saying that "nature promotes nurture,nurture nature".Modern medicine tends that SONFH is a polygenic genetic disease,which involves many mechanisms such as hormone receptor,metabolic transport,oxidative stress,heredity,nerve synapse,lipid and bone metabolism.The epigenetic modification of drugs can interact with genetic gene polymorphism.Based on the study of the genetic structure of the disease,we can accurately grasp the law of the development of the disease.It was found that the genetic polymorphisms of NR3C1,ABCB1,NOS3,MTHFR,CYP3A4,SYN2,IGFBP3,PON1 and APOA5 were closely related to the pathogenesis of SONFH in Han population.In this study,blood samples of 193 cases of Han nationality(79 cases of SONFH and 114 cases of intervention group without osteonecrosis treated with glucocorticoid)were collected.18 SNP loci on the above 9 genes were detected,and the DNA methylation level of positive sites was determined.Objective: Through the study of the genetic structure of the interaction between SONFH-related genes and their DNA methylation levels,to explore the pathogenesis of the disease,with a view to establishing a gene network model of SONFH in the future,and providing a holographic picture for the combination of Chinese and Western diagnosis and treatment of the disease.Methods: 1.Eighteen SNP loci(rs100529579,rs1045642,rs1128503,rs1799983,rs1801133,rs2032582 C,rs2032582T,rs227980,rs2279750,rs2453839,rs308952,rs3110697,rs3755724,rs3817004,rs41423247,rs662,rs662799,rs99365)of 79 patients with SONFH in the case group and 114 patients without osteonecrosis in the verified intervention group were detected by i MLDR method.The gene data of 208 normal subjects in the Han population genome database were also included as normal controls for statistical analysis.2.The methylation levels on 11 Cp G islands of IGFBP3,ABCB1,NR3C1 and MTHFR gene amplifiers in 59 patients with SONFH and 85 patients without osteonecrosis of the femoral head were detected by Methyl Target technique,and the methylation levels of loci,fragments and genes were statistically analyzed.Finally,e QTLD technique was used to analyze the interaction between SNP and locus methylation level.Results: 1.General data: there were 79 cases(female 45,male 34)in the case group,aged 50.65 ±13.95 years old,and 114 cases(female 65,male 49)in the verified intervention group,with an average age of 42.63 ±19.27 years.Compared with the verification group,the age t test P = 0.001,the difference was statistically significant,while the sex t test P = 0.8989,the difference was not statistically significant.2.All samples Hardy-Weinberg equilibrium(Hardy-Weinberg equilibrium,HWE)quality control: all samples 18 SNPs(rs100529579,rs1045642,rs1128503,rs1799983,rs1801133,rs2032582 C,rs2032582T,rs227980,rs2279750,rs2453839,rs308952,rs3110697,rs3755724,rs3817004,rs41423247,rs662,rs662799,rs99365)HWE test P >0.05,consistent with equilibrium.3.SNP analysis of all samples: compared with the validated intervention group,it was found that the risk of rs2242480 allele T was lower in the case group,and the risk of rs1801133 allele G was higher than that in the normal group.Compared with the validated intervention group,the risk of rs3110697 genotype A was lower in the validated intervention group,and the risk of rs1801133 genotype Ghand A was higher and that of rs3110697 genotype A was lower than that of the normal group.Compared with the validated intervention group,the risk of rs2032582 T allele T was relatively lower under the dominant genetic model,and the risk of rs3110697 allele A was lower under the recessive genetic model.Compared with the normal group,rs10052957 allele An and rs1801133 allele G had a higher risk under the dominant genetic model,while rs3110697 allele A had a lower risk under the recessive genetic model.Compared with verification group,ABCB-1(rs2032582T and rs1128503),SYN2(rs3755724,rs2279750,rs3817004)had stronger linkage,ABCB-1(rs1045642,rs2032582T),SYN2(rs3755724 and other SNP)had stronger linkage than normal group.Compared with verification group and normal group,there was interaction between 34 groups and 16 groups of SNP.4.Analysis of methylation level and its interaction with SNP in case group and verification intervention group: high methylation level of ABCB1＿1 POS.192,ABCB1＿2 POS.43,IGFBP3＿2 POS.143,MTHFR＿2 POS.42 site and high risk of disease,high methylation level of MTHFR＿1 POS.36,MTHFR＿1 POS.77,MTHFR＿1 POS.139,NR3C1＿2 POS.163,NR3C1＿4 POS.47 site and low risk of disease.The methylation level of ABCB1＿2 fragment was 0.024,and there was statistical difference.The methylation level of ABCB1 gene was 0.04,and there was statistical difference.There were significant differences in ABCB1＿2ctttttttttttttttttttttttt,IGFBP3＿1tttttttttcttt,IGFBP3＿3tttttttctttttttt,IGFBP3＿2 ttttttttttttctttt,IGFBP3＿2ctttttttttttttttt,NR3C1＿4 ttttctttttttt,NR3C1＿1 tttttttttttttctt,MTHFR＿1 ttttttcttttttttttttttttttt,NR3C1＿4ttttttttttttt,MTHFR＿1tttttttttttttctttttttttttt,ABCB1＿2tcttttttttttttttttttttttt,ABCB1＿1ttctttttttttt,MTHFR＿1tctttttttttttttttttttttttt,MTHFR＿1ttttttttttttttttcttttttttt and NR3C1＿1 cttttttttttttttt haplotypes.There were significant differences in the close(cis)effect of 22 on SNP and site methylation level,the SNP of 139 pairs and the long distance(trans)effect of site methylation level.Conclusion: Steroid-induced osteonecrosis of the femoral head is a polygenic disease,the extensive interaction between SNP and DNA methylation level or the dominant disease process,its occurrence and development has a certain complexity and inevitability,which is consistent with the congenital and acquired theories of traditional Chinese medicine.It can include various factors and draw lessons from the syndrome differentiation system of traditional Chinese medicine to establish the gene regulatory network model of SONFH in the future,which will provide a holographic picture for the diagnosis and treatment of diseases under the concept of accurate medical treatment. |
