| Objective:1.To explore the pathogenetic genes of autism spectrum disorder complicated with mental retardation,and explore the mechanism of ASD associated with mental disorders.2.To explore the cytokine and it’s pathogenesis related to ASD.Methods:1.To explore the susceptibility genes of ASD by the whole exome sequencing:(1)To examine the subjects,including physical examination,nerve examination and mental assessment etc.(2)To extract peripheral blood of patients with ASD.(3)To extract the whole genome DNA.(4)Using the high-throughput sequencing technology to carry out the sequenced sequencing of exons,and the sequencing results were verified by Sanger sequencing.2.The Milliplex multifactor detection technique is used to explore the cytokines related to the pathogenesis of ASD:(1)We collected the clinical data,including the general data,the Autistic Behavior Checklist(ABC),the Modified Checklist for Autism in Toddlers(M-CHAT)etc.(2)To extracte the peripheral blood of ASD patients and control groups.(3)To apply the Milliplex multifactor detection technique determine the concentrations of 11 kinds of cytokines related to helper T cells 17(Th17),helper T cell1(Th1)and helper T cell 2(Th2)in the serum of ASD patients and control group.(4)Peripheral blood RNA was extracted for real-time quantitative PCR(rt-pcr)experiment to detect the expression of TNF-PCR and hnRNPL immunoregulatory LincRNA(THRIL).Results:1.By high-throughput exome sequencing,we found 3 candidate genes,including DIAPH3,RELN,SETD2,and verified by Sanger sequencing.The RELN gene found a synonymous mutations;DIAPH3 gene mutation in 18 exon of c.2156T>C(p.I719T),which occurred in the FH2 conserved proline rich domain,is a missense mutation,the parents were not found the mutation,so ti is considered a new mutation;The mutant c.6895G>A(p.G2299R)of the SETD2 gene is also identified as a new missense mutation.2.(1)Tumor Necrosis factor-α(TNF-α),interleukin-1(IL-1)and interleukin-17a(IL-17a)were significantly increased in peripheral blood of ASD patients(P<0.05).(2)TNF-α expression was positively correlated with the severity of the five clinical symptom dimensions of the autism behavior rating scale(ABC scale).(3)THRIL expression was significantly reduced in ASD patients(P<0.01).Conclusion:1.The mutations of DIAPH3,SETD2 may increase the risk of suffering from autism spectrum disorders.2.Abnormal expression of cytokines such as TNF-α radiation,IL-1,and IL-17 a may also increase the risk of ASD.3.The expression of TNF-α was significantly increased in ASD patients,and was positively correlated with the severity of clinical symptoms. |
