| Objective: Flow cytometry and enzyme-related immunosorbent assay(ELISA)were used to detect the expression levels of CD3+,CD4+,CD8+ T lymphocytes,Treg cells and related cytokines in peripheral blood of patients with hemophilia A,and to identify the differences between patients with positive FVIII inhibitor and patients with negative FVIII inhibitor.To investigate its possible mechanism in FVIII inhibitor formation.Methods: A total of 80 hemophilia A patients with complete medical history who had been registered and managed in our hospital(Anhui Provincial Hospital)from December 2003 to December 2022 were collected and divided into two groups according to the presence or absence of inhibitor formation,one group was inhibitor positive group,the other group was inhibitor negative group.At the same time,a questionnaire of risk factors related to the production of inhibitors was developed,and the general data,clinical characteristics and molecular basic data of patients were recorded.Finally,the summary and statistical analysis were carried out.Results: 1.All the 80 patients included in this study were male,45 cases(56%)in the inhibitor-negative group,with a median age of 3(0,52)years,and 35 cases(44%)in the inhibitor-positive group,with a median age of 4(1,55)years.2.The detection rate of inhibitors in hemophilia A patients in our hospital was 44%,and among the positive cases of inhibitors,there were 22 cases with low titer,accounting for63%;13 cases with high titer,accounting for 37%;3.The statistical results of the relevant factors affecting the formation of inhibitors suggested that the differences of gender,age of onset,family history,form of first bleeding and treatment between the two groups were statistically insignificant(P>0.05);It was correlated with FVIII activity(P=0.017).The disease severity was correlated with inhibitor formation(P=0.026).4.There was no significant difference in exposure days between the two groups(P>0.05);5.A total of 35 immune cell sets were tested,including 14 cases(40%)in the inhibitor-positive group and 21 cases(60%)in the inhibitor-negative group.The proportion of CD4+ T lymphocytes in the inhibitor-positive group was(33.99±3.99)%,which was significantly lower than that in the negative group(37.64±4.09)%.The difference was statistically significant(P=0.018).The percentage of CD8+ T lymphocytes in the inhibitor-positive group was(28.22±4.29)%,which was significantly higher than that in the negative group(24.49±5.02)%,and the difference was statistically significant(P=0.029).The ratio of Treg cells in the inhibitor-positive group(2.53±1.07)% was significantly lower than that in the negative group(4.45±2.21)%,and the difference was statistically significant(P=0.002).There was no significant difference in CD3+ T lymphocytes and Breg cells between the two groups,which was statistically significant(P>0.05);6.Cytokine analysis: The median IL-10 level in the inhibitor-positive group was 2.74(1.66-2.92)pg/ml,which was significantly higher than that in the negative group(1.88(1.69-2.13)pg/ml,the difference was statistically significant(P=0.037),while the IL-6expression level was not significantly different between the two groups(P>0.05).7.Genetic analysis: A total of 6 genetic tests were performed,among which 2 patients were followed up in the outpatient department.The gene mutation types found included intron 22 inversion,missense mutation and deletion mutation,and no new gene mutation types were found this time.Conclusion: 1.The detection rate of HA inhibitors in our hospital was 44%;2.The disease severity of HA patients was significantly correlated with the formation of inhibitors;3.The production of HA inhibitors was not correlated with gender,age of onset,family history,bleeding pattern,exposure date and treatment.4.The production of inhibitors in HA patients was correlated with the proportion of CD4+,CD8+ T lymphocytes and the level of Treg cells in peripheral blood;There was no significant correlation with CD3+ T lymphocyte and Breg cell levels in peripheral blood.5.The formation of inhibitors in HA patients was correlated with the expression level of IL-10 in peripheral blood,but had no significant correlation with IL-6 level. |
