| Background:Sj?gren’s syndrome(SS)is a diffuse connective tissue disease,which is characterized by damage of exocrine glands,such as lacrimal and salivary glands,and massive infiltration of lymphocytes.The most common symptoms are dry mouth and dry eyes.Lung involvement is one of the most common visceral lesions in SS.γδ T cells play roles of immune surveillance and immune defense in tumors and infections.Activated γδ T cells can exhibit multiple functions including cytotoxic effects,production of cytokines and chemokines,presentation of antigens,and modulation of immunity.therefore,activated γδ T cells participate in a range of disease processes including anti-infection,relief of allergies,autoimmunity,and cancer-killing.The role of γδ T cells in sjogren’s syndrome is unknown.Mesenchymal stem cells(MSCs),which exhibit low immunogenicity and unique immunomodulatory effects,has benefited the treatment of several autoimmune diseases.However,the effect and mechanism of UC-MSCs in treating Primary Sjogren’s syndrome(pSS)with pulmonary inflammation are still incompletely understood.The clinical practices showed that dendrobium officinale polysaccharide(DOP)could generate a satisfied immunity effect.The effects of UC-MSCs and DOP on γδ T cells in the treatment of Sjogren’s syndrome need to be further explored.Objective:This study aims to investigate the role of γδ T cells and their subsets in the development of SS,to explore the efficacy and effects UC-MSCs and UC-MSCs combined with DOP on γδ T cells in the treatment of SS.Methods:1)The changes of the frequencies and absolute numbers of peripheral γδ T cells,γδT17(IL-17~+γδ T cells),γδT1(INF-γ~+γδ T cells)were detected by flow cytometry,and correlation analysis was performed between these results and related clinical indicators.The total number of γδ T cells,were detected by flow cytometry with full venous blood tests before and 24 hours after the UC-MSCs transplantation.2)Saliva flow of 8-week-old NOD/Ltj mice was measured after anesthesia,and submandibular gland,tear gland and lung were taken for HE pathological staining after sacrifice to get its’ pathological scores.And the frequencies and absolute numbers of γδ T cells and subgroups Vγ1,Vγ4,Vγ5 and Vy6 in peripheral blood and lung were detected by utilizing the flow cytometry.Related data was analyzed with application software.3)For the NOD/Ltj mice groups of 6,8 and 12 weeks old,saliva flow was measured,HE staining was performed,and the frequencies and absolute numbers of γδ T cells were detected.4)The γδ T cells,which sorted out from the spleen of ICR mice,were injected into 5-week-old NOD/Ltj mice through tail vein,the mice were raised for 3 weeks after tail vein injection and received surveillance of saliva flow rate and observation of the pathological changes as wells as the changes of γδ T cell by flow cytometry.5)UC-MSCs were injected into the tail vein of 8-week-old NOD/Ltj mice,and the mice were sacrificed after four weeks of treatment.The saliva flow rate was detected.the submandibular gland and lung were taken and HE pathological staining was performed to observe and assess the inflammation degree of the submandibular gland in each group.The frequencies and absolute numbers of γδ T cells and Vγ4~+T cells in the spleen,lung and peripheral blood were detected by flow cytometry.6)After 4 weeks treatment,pathological changes between treatments of DOP alone and DOP combining UC-MSCs were compared.Flow cytometry was used to detect the expression of γδT cells in the peripheral blood,spleen and lung in each group.Results:1)The frequencies and absolute numbers of peripheral blood γδ T cells in pSS patients were lower than those in the healthy controls,and the proportions and absolute numbers of γδ T1 cells in pSS patients were higher than that of healthy controls.The frequencies and the absolute numbers of Vδ2~+T cells in peripheral blood of patients with pSS were significantly decreased,and those of Vδ1~+T cells did not show a significantly change.The proportions of γδ T cells in the peripheral blood of pSS patients was positively correlated with ESSDAI score,ESR,IgG and negatively correlated with C4 level.The frequencies and absolute numbers of γδ T cells decreased in pSS patients 24h after UC-MSCs transplantation.2)Compared with ICR mice,the salivary flow rate of NOD/Ltj mice was significantly reduced,and the submandibular glands were significantly swollen.A large amount of lymphocyte infiltration was observed in the submandibular gland and the lacrimal gland using HE staining.The proportions and absolute numbers of γδ T cells,the absolute numbers of γδ T17(IL-17A~+ γδ T)cells,the proportions of Vγ4~+ T cells and the proportions and absolute numbers of Vγ4~+ IL-17A~+ T cells in peripheral blood decreased significantly.The proportions and absolute numbers of γδ T cells,γδ T17 cells,Vγ4~+ T cells and Vγ4~+ IL-17A~+ T cells in the lung increased significantly.Compared with 6-week-old NOD/Ltj mice,8-week-old and 12-week-old NOD/Ltj mice had more lymphocytes infiltrated and larger infiltrated area in the lungs with the increase of the age of the mice,the percentage and absolute number of γδ T cells in peripheral blood gradually decreased,and the percentage and absolute number of γδ T cells in lung gradually increased.The salivary flow rate was significantly decreased after 5 weeks of the injection of γδ T cells into the tail vein of NOD/Ltj mice,and the infiltration of lymphocytes was significantly increased in the lung.γδ The frequencies of γδ T cells and the frequencies and absolute numbers of γδ T17 cells,Vy4~+T cells,Vγ4~+ IL-17A~+ T cells in the lung were increased.3)Compared with the control group and NOD group,the saliva flow rate was markedly increased in the UC-MSCs treatment group,and lymphocyte infiltration reduction could be observed by HE staining of the submandibular gland and lung.The proportions and absolute numbers of γδ T17,Vγ4~+T,Vγ4~+IL-17A~+T cells in the spleen were reduced.The absolute absolute numbers of γδ T cells,γδ T17 cells and Vγ4~+ T cells were reduced,and the proportions and absolute absolute numbers of Vγ4~+ IL-17A~+ T cells in peripheral blood were reduced.The proportions and absolute absolute numbers of γδ T cells,γδ T17 cells,Vγ4~+ T and Vγ4~+IL-17A~+T cells in the lung were notably reduced.4)Compared with the UC-MSCs group,the salivary flow rate remained unchanged in dendrobium polysaccharides group,but the pathology of the submandibular gland in DOP and UC-MSCs combined group were dramatically improved and the salivary flow rate increased more markedly.Compared with UC-MSCs group,DOP and UC-MSCs combined treatment did not exhibit significant effect on the level of γδ T cells.Conclusions:γδ T cells and Vγ4~+T cells are involved in the pathogenesis of Sjogren’s syndrome.UC-MSCs may treat Sjogren’s syndrome by regulating γδ T cells.The combined treatment of DOP and UC-MSCs could improve the therapeutic effect of Sjogren’s syndrome.In view of the different links in the course of Sjogren’s syndrome,we should give full play to the overall concept of "treatment based on syndrome differentiation" of traditional Chinese medicine and western medicine,which provides a new treatment way for Sjogren’s syndrome. |
PDF Full Text Request