| Sjogren syndrome(SS)is an autoimmune disease mainly involving exocrine glands.Its clinical manifestations are diverse and can involve multiple organs and systems such as skin,joints.lungs,kidneys,nerves and so on.The Huoxue Jiedu recipe guided by the dryness.toxin and blood stasis theory has achieved satisfactory curative effect in clinic.Starting from the dryness,toxin and Blood Stasis Theory and combined with the modern medical "immune metabolism" theory,this paper deeply discusses the relationship between dryness,toxin and blood stasis and autophagy,and confirms the correlation between them through clinical research and cell experiment,which provides micro dialectical evidence for dryness,toxin and Blood Stasis Theory and dryness,toxin and blood stasis syndrome type of SS patients(deficiency of yin and Tianjin,stagnation of evil toxin and blood stasis syndrome),The mechanism of Huoxue Jiedu recipe was explored from the perspective of autophagy.Theoretical discussionObjective:To explore the relationship between dryness,toxin and Blood Stasis Theory and autophagy in Sjogren’s syndrome.Methods:Through literature review,we sorted out the dryness toxin and Blood Stasis Theory of Sjogren’s syndrome,the clinical characteristics of patients with dryness toxin and blood stasis and the related literature research of autophagy.Taking the pathogenesis,etiology,pathogenesis,clinical manifestations and pathological characteristics of Sjogren’s syndrome as the node,we combed and discussed the context and relationship network between them.Conclusion:1.The abnormality of autophagy and autophagy is closely related to the disorder of autophagy and autophagy in traditional Chinese medicine.2.Autophagy and Sjogren’s syndrome have the characteristics of complexity and universality.3.The abnormality of autophagy can be regarded as the microscopic manifestation of abnormal gasification function caused by "dryness,toxin and blood stasis".Clinical researchStudy one Expression of LC3-Ⅱ in Treg cells of SS peripheral bloodObjective:To explore the expression of autophagy in Yin Jin deficiency,evil toxin stagnation and blood stasis syndrome group of SS,Yin Jin deficiency,evil toxin stagnation and blood stasis syndrome group of non SS and normal group,so as to provide Micro syndrome differentiation evidence for dryness,toxin and blood stasis theory.Methods:30 cases of SS deficiency of yin and fluid.20 cases of SS non deficiency of yin and fluid.2 0 cases of blood stasis and 10 cases of normal control group were included.The clinical information was collected and the peripheral blood mononuclear cells(PBMC)cells were extracted from the peripheral blood of each group.The expression of autophagy marker LC3-Ⅱ in peripheral blood Treg cells of each group was detected by flow cytometry,The correlation between the expression of LC3-II and disease activity in SS patients was analyzed.Results:1.The positive rate of anti-Ro52,anti SSA,anti SSB antibody,ESSDAI,SSDAI score and IgG level in SS Yin Jin deficiency evil toxin stagnation blood stasis group were significantly higher than those in SS non Yin Jin deficiency evil toxin stagnation blood stasis group(P<0.05);There was no significant difference between ESSPRI and TCM syndrome score(P>0.05).2.There was no significant difference in the proportion of peripheral blood Treg cells among the three groups(P>0.05);The expression of LC3-Ⅱ in peripheral blood Treg cells of SS patients was lower than that of normal group,the difference was statistically significant(P<0.05);The expression of LC3-Ⅱ in SS group with deficiency of yin and fluid,pathogenic toxin and blood stasis was higher than that in SS group without deficiency of yin and fluid,with significant difference(P<0.05).3.The expression of LC3-II was positively correlated with ESSDAI(r=0.470)and SSDAI(r=0.602)in SS patients with non Yin fluid deficiency,evil toxin stagnation and blood stasis(P<0.05);The expression of LC3-Ⅱ was positively correlated with ESSDAI(r=0.463),ssadi(r=0.486),IgG(r=0.559)and parotid gland swelling score(r=0.529)(P<0.05).4.The ROC curve analysis of IgG and LC3-Ⅱ shows that the areas under the surface are 0.697 and 0.692 respectively,and the dividing points are 12.35 and 30.85 respectively,which is of the greatest significance in the diagnosis of SS patients with Yin Tianjin deficiency,evil toxin stagnation and blood stasis syndrome.Conclusion:1.Under the guidance of dryness,toxin and Blood Stasis Theory,SS patients with Yin Tianjin deficiency,evil,toxin and blood stasis syndrome have higher disease activity and IgG expression;2.The autophagy activation of Treg cells was up-regulated in SS patients with Yin fluid deficiency,evil toxin stagnation and blood stasis syndrome.The expression of LC3-Ⅱ in peripheral blood Treg cells was positively correlated with disease activity score,IgG and parotid gland enlargement,suggesting that LC3-II was closely related to the function of Treg cells;3.The expression of IgG and LC3-II in peripheral blood Treg cells can be used as the microscopic syndrome differentiation index of SS patients with Yin and fluid deficiency,evil toxin stagnation and blood stasis syndrome.Study two Therapeutic effect of Huoxue Jiedu Recipe on SS and its intervention on autophagyObjective:To explore the clinical efficacy of Huoxue Jiedu recipe and its intervention on autophagy of Treg cells in peripheral blood of SS with Yin and fluid deficiency,evil toxin stagnation and blood stasis syndrome.Methods:30 patients with SS syndrome of yin and fluid deficiency,evil toxin stagnation and blood stasis were included and treated with Huoxue Jiedu formula granules.The TCM syndrome score,ESSDAI,SSDAI,ESSPRI,clinical laboratory indexes and safety indexes before and after treatment were compared,and the changes of LC3-II expression in peripheral blood Treg cells before and after treatment were observed.Results:1.A total of 28 patients with SS syndrome of Yin Tianjin deficiency,evil toxin stagnation and blood stasis were observed.One case lost follow-up and one case had abnormal liver function.The total completion rate was 93.3%.2.Cases used nimodipine method to calculate the curative effect index of traditional Chinese medicine syndrome before and after treatment.It was concluded that the clinical control was 0 cases,the significant effect was 5 cases-the effective was 20 cases and the ineffective was 3 cases.The total effective rate was 89.3%.3.Compared with before treatment,the scores of ESSDAI.SSDIA,ESSPRI and TCM syndrome decreased after treatment,and the difference was statistically significant(P<0.05).4.Compared with before treatment,the levels of ESR.IgG and IgA in patients’ laboratory indexes decreased,and the difference was statistically significant(P<0.05).5.Compared with before treatment,the proportion of Treg cells in peripheral blood did not change significantly(P>0.05);The expression of LC3-Ⅱ in Treg cells decreased,and the difference was statistically significant(P<0.05).Conclusion:1.Huoxue Jiedu recipe can significantly improve the TCM syndrome score and disease activity of SS Yin Jin deficiency,evil toxin stagnation and blood stasis syndrome.and has high safety.2.Huoxue Jiedu recipe can improve the disease activity and reduce the expression of LC3-II in Treg cells,suggesting that Huoxue Jiedu recipe can affect the function of Treg cells by regulating autophagy.3.The Micro syndrome differentiation under the theory of dryness,toxin and blood stasis has been confirmed by clinical treatment,forming a complete chain of evidence.Experimental studyExperiment one IFNy Effect on autophagy of HSG cellsObjective:To explore IFNy Effect on autophagy of HSG cellsMethods:IFNy was applied as a microenvironment simulating SGEC injury in SS patients,IFN y was explored by autophagy inhibitors effect on autophagy of HSG cells.Results:1.Under light microscope,compared with the normal group.IFNγ In group B,the number of cells decreased,the adhesion was poor,the cell shape was long spindle,the cell edge was bright,the intracellular material was unclear,vacuoles of different sizes were visible,the cell gap was increased,a large number of vacuolar particles were visible,and the nucleus was concentrated.2.Baf Al+IFNγ The expression of LC3-Ⅱ in group was higher than that in IFNγ Group,comparison of LC3-Ⅱ difference between groups,Baf A1 group(0.046)-blank group(0.010)<Baf A1+ IFNγ Group(0.082)-IFNγ Group(0.019).3.Compared with the blank group,BAF A1+IFNγ Decreased expression of histone p62 protein,IFNy The expression of p62 protein in group A was significantly higher than that in the blank group(P<0.05);And IFNγ Compared with BAF A1 group and Baf A1+ IFNγ The expression of p62 protein decreased in group A(P<0.05).Conclusion:IFNγ It can promote HSG autophagy,which may be the cause of sgec injury and apoptosis.Experiment two Effect of Huoxue Jiedu Recipe on IFNγ Role of induced autophagy in HSG cellsObjective:To explore the effect of Huoxue Jiedu Recipe on IFNy Role of induced autophagy in HSG cells.Methods:IFNγ was applied as a microenvironment inducer simulating sgec injury in SS patients.The experimental groups included blank group,Baf A1 group,model group,traditional Chinese medicine group and chloroquine group.The morphology and quantity of cells in each group were observed under light microscope;CCK8 was used to detect the cell survival rate of each group;WB was used to detect the protein expressions of LC3-Ⅱ,p62,PI3Kp110α,AMPK,p-AMPK,mTOR,p-mTOR,pro-caspase-3 and cleaved-caspase-3 in each group.Results:1.Under light microscope,the number of cells in the model group and Baf A1 group was less than that in the blank group,the gap was increased,the nucleus was concentrated,the cell shape was long spindle,and the number of cells in the chloroquine group was more than that in the model group.2.CCK8 was used to detect the cell viability of each group.The cell viability of model group and Baf A1 group was significantly lower than that of normal group,and the OD value was lower than that of normal group(P<0.05);The cell viability of chloroquine group and traditional Chinese medicine group was higher than that of model group,and the OD value was higher than that of model group(P<0.05);Compared with chloroquine group,the cell viability of traditional Chinese medicine group was higher,and the OD value was higher than that of chloroquine group(P<0.05);Compared with the model group,the levels of LC3-Ⅱ,PI3Kp110α and p-mTOR protein in the traditional Chinese medicine group and chloroquine group were significantly higher than those in the model group(P<0.05);Compared with the model group,the expression of p62 protein in traditional Chinese medicine group and chloroquine group decreased,and the difference was statistically significant(P<0.05).Conclusion:1.Huoxue Jiedu recipe can inhibit IFNy The mechanism of induced autophagy of HSG cells may be related to the activation of PI3K/ATK/mTOR.2.This study believes that dryness,toxin and blood stasis are the general pathological basis of SS.This theory is verified by clinical experiment and cell experiment through one of the micro indexes of autophagy.The "method" and "formula" formed under the guidance of "reason"have been further verified through clinical and experimental research,and the relationship between dryness,toxin and blood stasis and autophagy has been inversely verified through Huoxue Jiedu formula.Finally,the establishment of Micro syndrome differentiation under the guidance of dryness,toxin and blood stasis theory has been formed. |
PDF Full Text Request