Study On Changes Of INKT Cells And Their Subpopulations In Peripheral Blood In Patients With Moderate To Severe Thyroid-associated Ophthalmopathy

Background:Thyroid-associated ophthalmopathy（TAO）is an organ-specific autoimmune disease,the pathogenesis has not been fully clarified.The currently recognized pathogenesis is mainly related to immune factors,and a variety of immune cells are involved,such as T lymphocytes,B lymphocytes cell.With the continuous exploration of immune cells,a new type of lymphocyte NKT cells has received extensive attention.Type I NKT cells（iNKT cells）have different expressions in different thyroid diseases and play an immunoregulatory role.Moderate and severe thyroid-associated ophthalmopathy is harmful and difficult to treat,so it is particularly important to further explore its mechanism.At present,the relationship between moderate and severe thyroid-associated ophthalmopathy and iNKT cells is still unclear,and further in-depth research and discussion are needed.Objective:To observe the changes of iNKT cells and their subpopulations in the peripheral blood of patients with moderate to severe thyroid-associated ophthalmopathy,to explore the correlation of iNKT cells and subpopulations with clinical indicators in patients with moderate to severe TAO,and to compare the changes of iNKT cells and subpopulations in the peripheral blood of patients with thyroid-associated ophthalmopathy and subacute thyroiditis.Methods:In this study,12 patients with moderate to severe thyroid-associated ophthalmopathy（TAO）who were newly diagnosed or who had not received hormone and otherimmunosuppressive therapy within 3 months,subacute thyroiditis control group（SAT）8 cases and healthy control group（HC）12 cases were selected from the First Hospital of Jilin University from July 2015 to October 2021.The gender,age,white blood cells,lymphocytes,and thyroid function of the research subjects,as well as the exophthalmos and CAS scores of the TAO group were collected.Flow cytometry was used for cell isolation and phenotype identification.Flowjo 7.6software was used to determine the cell frequency of iNKT cells and their subpopulations,and the absolute value of cells was converted.SPSS 26.0 software was used for statistical analysis,and the differences between each of the three groups were analyzed.The measurement data that conformed to the normal distribution were tested by independent samples t test,and those that did not conform to the normal distribution were analyzed by the Mann-Whitney U test.Pearson test was used for correlation analysis of measurement data that conformed to normal distribution,and Spearman test was used for correlation analysis of measurement data that did not conform to normal distribution.The significance test criterion was p<0.05.Result:1.Compared with the HC group,the absolute values of TCRVβ11+iNKT cells,TCRVα24+iNKT cells and TCRVα24Jα18+iNKT cells in the TAO group were significantly decreased（p<0.05）.The frequencies and absolute value of CD4-CD8-TCRVβ11+iNKT cells and CD4-CD8-TCRVα24Jα18+iNKT cells were significantly decreased（p<0.05）,and the frequencies of CD4-CD8-TCRVα24+iNKT cells was significantly decreased（p<0.05）.The frequencies and absolute value of CD8+TCRVα24Jα18+iNKT cells were significantly decreased（p<0.05）,and the absolute value of CD8+TCRVα24+iNKT cells was significantly decreased（p<0.05）.The frequencies and absolute value of CD4+TCRVα24Jα18+iNKT cells were significantly increased（p<0.05）.2.Compared with the HC group,the absolute values of TCRVα24+iNKT cells and TCRVα24Jα18+iNKT cells in the SAT group were significantly decreased（p<0.05）.The absolute value of CD4-CD8-TCRVα24Jα18+iNKT cells was significantly decreased（p<0.05）.The frequencies and absolute value of CD8+TCRVα24+iNKT cells and CD8+TCRVα24Jα18+iNKT cells were significantly decreased（p<0.05）.The frequency of CD4+TCRVα24Jα18+iNKT cells was significantly increased（p<0.05）.3.The frequencies of CD8+TCRVα24+iNKT cells in TAO group（r=0.741,p=0.006）was positively correlated with TPOAb,and the frequencies of CD4+TCRVα24Jα18+iNKT cells（r=-0.600,p=0.040）was negatively correlated with TPOAb.4.Comparing the TAO and SAT groups,it was found that the absolute value of CD4-CD8-TCRVβ11+iNKT cells in the TAO group decreased more significantly（p<0.05）,and the frequencies of CD8+TCRVα24+iNKT cells in the SAT group decreased more significantly（p<0.05）.Conclusion:1.The number of iNKT cells are decreased,mainly CD4-CD8-iNKT cells and CD8+iNKT cell subpopulations decreased in peripheral blood of patients with moderate to severe thyroid-associated ophthalmopathy,suggesting that iNKT cells play a special immunoregulatory role in moderate and severe thyroid-associated ophthalmopathy.2.With the increase of TPOAb in the peripheral blood of patients with moderate and severe thyroid-associated ophthalmopathy,different iNKT cell subpopulations showed different changes,CD8+iNKT cell subpopulations increased accordingly,and CD4+iNKT cell subpopulations showed a downward trend,indicating that there is a certain relationship between different subpopulations of iNKT cells and TPOAb.3.Compared with the SAT group,there were differences in the subpopulations of iNKT cells in the peripheral blood between the TAO group and the SAT group,indicating that there were specific changes in the iNKT cell subpopulations in the peripheral blood of patients with moderate to severe thyroid-associated ophthalmopathy.