| ObjectiveFrom the three aspects of ancient literature research, clinical research and basic research to investigate rheumatoid arthritis (RA) pathogenesis of blood stasis and its molecular basis.Methods1. By using the method of literature research, summarize the ancient literature of TCM, which is relevanted to the etiology of RA. Search terms about bi syndrome, li-jie disease, rheumatism, arthroncus of knee, white tiger were selected to screening related literatures, then took further screening according to the description related to blood stasis, which include bi, blood stasis, astringent, impassability, accumulation, numbness, unfavorable and pain. Analyze the articles which had been retrieved, investigate RA pathogenesis of blood stasis by the understanding of etiology of ancient doctors.2. The four diagnostic information of inpatients with rheumatoid arthritis (n=168) in Guangdong Povincial Hospital of Chinese Medicine were collected and subsequently analyzed by using the factor analysis method. The factors and loading associated with different symptoms were analyzed accordingly to make the corresponding syndromes clear, so to reveal the features of syndrome type of RA in Guangdong Povincial Hospital of Chinese Medicine. Then further reference the latest diagnostic criteria of blood stasis syndrome, and analyze the correlation of each syndrome and the pathogenesis of blood stasis3. Take classical Chinese and Western medicine of RA as a tool, to investigate the molecular basis of RA pathogenesis of blood stasis. Tumor necrosis factor alpha (TNF-a) was used to induce fibroblast-like synovial cell (MH7A) of RA, so to construct RA in vitro mode. Then utilize methotrexate (MTX) and sinomenine hydrochloride (SIN), the classical drugs for the treatment of RA, applied to MH7A cells respectively. By means of MTS method, detect the effect of MTX and SIN for the proliferation of MH7A cells, and acquire their optimal working concentration and optimal working time. Use western blotting method to detect different concentrations of MTX and SIN, for their effects of the expression of thromboxane synthase (TxAS), the marker of blood stasis, and alpha auxiliary actin-1(ACTN1), the marker of cell proliferation and migration. The expression of cyclooxygenase-2(COX-2) was detected as a positive control in this experiment. Finally, select articular synovial tissue in patients of three common clinical syndrome types of RA, include dampness-heat blockage, cold-dampness blockage, deficiency of liver and kidney. By western blotting method, detect the expression difference of TxAS, ACTN1in synovial tissue of different syndrome types.Results1. Ancient literature research There were altogether42more relevant articles been screened out in the study. Further analysis about internal causes, external causes, and non-external and non-internal causes draw a conclusion, various of causes of RA can lead to blood stasis, and the stasis can occur in each phase of RA. External cause (wind, cold, dampness, heat, etc) lead to blood stasis is the main early performance. External evil causes meridian blocking, Qi and blood circulation blocked. Internal cause, eficiency syndrome or excess syndrome, lead to blood stasis, when disease progression. Deficiency of Qi, blood, Yin and Yang, lead to difficulties in nourishing blood or promoting circulation of blood. Or phlegm and blood stasis alternate, lead to phlegm blocking meridian. When time delay, persistent illness entering collaterals, Qi and blood have difficult in circumfluence. The causes of blood stasis, also include traumatism, emotions, disharmony of thoroughfare and conception vessels, etc.2. Clinical research There are15common factors had been extracted from38symptoms of RA patients, by using the factor analysis method, among which11typical common factors correspond to the three syndromes of rheumatoid arthritis, dampness-heat blockage type, cold-dampness blockage type and deficiency of liver and kidney type. Make further analysis by referencing diagnostic criteria of blood stasis syndrome, it was found that all the three syndromes of RA contain blood stasis symptoms.3. Basic research (1) Results of MTS showed, MTX and SIN, of which concentrations include50μg/ml,200μg/ml,400μg/ml, could inhibit the proliferation of MH7A, whether the work time was24hours,48hours and72hours. There were significant differences compared with control group (P<0.05), while SIN of400μg/ml made the most significant inhibition effect after72hours.(2) The western blotting results of MH7A showed, the expression of TxAS, ACTN1and COX-2was inhibited by MTX and SIN, while the effect was more significantly in the model group which had been induced by TNF-α.Compared with the control group, the expression of COX-2, ACTN1and TxAS, which was the biomarker of blood stasis, was higher in the model group. And the expression trend of TxAS, ACTN1and COX-2had consistency.(3) The synovial tissues derived from patients of RA were detected by western blotting, of which the results showed, the expression of TxAS and ACTN1was higher in cold-dampness blockage type and deficiency of liver and kidney type, and their expression trend was coincident.Conclusion1. A variety of causes of RA can lead to blood stasis, and as a basic pathogenesis, blood stasis occurs in each phase of RA.2. Dampness-heat blockage type, cold-dampness blockage type and deficiency of liver and kidney type were the most common syndrome types of patients with in Guangdong Povincial Hospital of Chinese Medicine. And they all related to blood stasis, which supported the point that blood stasis was the pathogenesis syndrome factor throughout different kinds of syndromes of RA.3. TxAS could be adjusted by classical anti-rheumatism medicine, MTX and SIN, and its expression was coincident with ACTN1, the marker of proliferation and invasive to synovial cell of RA, all these indicated that TxAS may be one of molecular basis to the RA pathogenesis of blood stasis. |
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