Comparison Of The Applied Value Of PCR-SSCP And Targeted Next-generation Sequencing In Diagnosis Of TSC-RAML

ObjectiveBy analyzing the The results of Polymerase Chain Reaction-Single-Strand Conformation Polymorphism(PCR-SSCP)and Next-generation Sequencing(NGS)in patients with Tuberous Sclerosis Complex Renal Angiomyolipoma(TSC-RAML),andclinical phenotypic characteristics of patients with different genotypes,this paper explored the application value of the two detection methods in TSC-RAML gene diagnosis,and provided reference for clinical decision-making and future research.MethodBoth PCR-SSCP and targeted NGS were used to detect the pathogenic genes TSC1 and TSC2 of TSC-RAML,so as to compare the accuracy of PCR-SSCP and NGS in genetic diagnosis of TSC-RAML patients.Patients were divided into groups according to the results of high-accuracy detection methods,and the differences between the general conditions and clinical phenotypes of patients in each group were compared to verify whether the clinical phenotype was consistent with genotype.Results(1)Among the 24 patients,mutations were detected in 18 cases by PCR-SSCP and 9 cases by NGS,and the mutation detection rate of PCR-SSCPwas significantly higher than that of NGS,with statistically significant difference(P < 0.05).There were 3 cases with consistent PCR-SSCP and NGS detection results,the proportion of which was significantly lower than that of the 21 cases with inconsistent results,and the difference was statistically significant(P < 0.05).The NGS mutation detection rate of the 18 patients with mutation detected by PCR-SSCP was lower than that of the 6 patients without mutation detected by PCR-SSCP(33.3% and 66.6%),but the difference was not statistically significant.All mutations detected by NGS were verified by Sanger sequencing,and the results were completely consistent.(2)There was no statistically significant difference in age and gender distribution between the TSC1/2 mutation group and the non-TSC1/2 mutation group(P < 0.05).The frequency of facial fibroadenoma,forehead plaque and pigment depigmentation in the TSC1/2 mutation group was higher than that in the non-TSC1/2 mutation group,and the difference was statistically significant(all P < 0.05).The frequency of tooth enamel point pits,finger(toe)nail fibroma and bone lesions in the two groups was significantly higher in the TSC1/2 mutation group than in the non-TSC1/2 mutation group,but the difference was not statistically significant(both P > 0.05).There was no statistically significant difference in the frequency of shark lesions,cortical nodules,Subependymal Nodules(SEN),Subependymal Giant cell Astrocytoma(SEGA),non-renal hamartomas or multiple renal cysts between the two groups(all P > 0.05).RAML related symptoms and signs of patients in both groups,the frequency of TSC1/2 mutation group was significantly higher than that of non-TSC1/2 mutation group,and the age of TSC1/2 mutation group at the time of occurrence was also significantly younger than that of non-TSC1/2 mutation group,with statistically significant differences(all P < 0.05).RAML diameter of the TSC1/2 mutant group was significantly larger than that of the non-TSC1/2 mutant group,with statistically significant difference(P<0.05).The proportion of patients in the TSC1/2 mutation group with RAML grading greater than grade 3 was significantly higher than that in the non-TSC1/2 mutation group,and the proportion of patients with RAML grading less than grade 3 was significantly lower than that in the non-TSC1/2 mutation group,with statistically significant differences(all P < 0.05).There was no statistical difference in the incidence of RAML rupture and massive bleeding between the two groups during the follow-up period(P < 0.05).Conclusion(2)PCR-SSCP is not reliable in the detection of pathogenic genes TSC1 and TSC2 in patients with TSC-RAML,and PCR-SSCP screening can not improve the mutation detection rate of NGS,so it is not recommended to promote clinical use.(2)The pathogenic genes TSC1 and TSC2 of patients withTSC-RAML detected by NGS are accurate and reliable,and the clinical phenotype of patients with TSC-RAML basically conforms to the genotype detected by NGS.(3)Patients with TSC-RAML mutation not detected by NGS have relatively mild clinical manifestations,and some patients may have genetic Mosaic phenomenon,which needs to be further studied.