| Objective:To compare the effects of TG(Tripterygium Glycosides tablet)from Hunan Qianjin Xieli and Zhejiang Deende on type II collagen-induced arthritis(CIA)in rats.Methods:A total of 72 SD rats,36 males and females,were randomly divided into normal group,model group,and the two times clinical equivalent dose of Qianjin tripterygium wilfordii glycosides(QJ-TG18mg·kg-1·d-1),the six times clinical equivalent dose(QJ-TG54mg·kg-1·d-1)group,the two times clinical equivalent dose of Deende tripterygium wilfordii glycosides(DED-TG18mg·kg-1·d-1)and six times clinical equivalent dose(DED-TG54mg·kg-1·d-1)group.QJ-TG and DED-TG were dissolved in 0.3%CMC-Na was administered intragastrically once a day.Observation of animal status,weighing once every three days.On the 21st and 42st day,A second booster on the 7th day after the first immunization.After the second immunization,the symptoms such as redness and swelling of joints were observed,and the clinical score and incidence of arthritis were evaluated.The inflammatory joints were examined by HE staining,Masson’s trichrome staining,and TRAP staining(anti-tartaric acid phosphatase staining).HE staining to observe the semi-quantitative analysis of synovial inflammation,pannus,cartilage erosion and bone destruction.Masson staining to observe the fibrosis of articular cartilage.TRAP staining to observe the number of osteoclasts in the subchondral bone and bone marrow cavity,comprehensive evaluation of the degree of joint disease;Blood routine test for rat red blood cell count(RBC),white blood cell count(WBC),platelet(PLT),hemoglobin(HGB),neutrophil ratio(NEUT%),and monocyte ratio(MONO%);Alkaline phosphatase(ALP),alanine aminotransferase(ALT),aspartate aminotransferase(AST),glutamyltransferase(GGT),total bilirubin in rat serum The contents of TBIL,creatinine(CRE)and urea(UREA)in serum were detected by enzymatic assay,as well as histopathological examination through organs.To evaluate the effects of TG from two different sources on organs such as liver and kidney in rats.The morphological changes of testis and ovary were observed by counting the abnormal sperm and HE staining.To evaluate the effects on sperm abnormality rate in epididymis,and testicular and ovarian tissue damage in rat ELISA was used to detect the effects of two different sources of TG on testosterone T,androgen synthesis-related rate-limiting enzymes CYP19A1 and inducible nitric oxide synthase(iNOS)in male CIA rats and effects of rate-limiting enzymes CYP19A1 and CYP17A1,serum estradiol(E2),luteinizing hormone(LH),and serum follicle stimulating hormone(FSH)levels in hormone synthesis in female CIA rats.Thus,the combined effects of TG from two different sources on the reproductive system of CIA model animals were evaluated.Result:1.Effects of two sources TG on the severity of arthritis in CIA model ratsBoth sources had different degrees of inhibition on joint swelling and malf ormation in CIA model rats after TG administration,and the higher the dose,t he more obvious the effect.The two dose groups of QJ-TG18mg·kg-1·d-1 and 54mg·kg-1·d-1 and the high dose group of DED-TG compared with the model group showed more obvious inhibition of clinical scores,and there was statistica 1 difference.Compared with the same dose of TG from the two sources,the DED-TG54 mg·kg-1·d-1 was significantly more effective on the 15th and 18th days than the QJ-TG same dose group,which was statistically significant(P<0.05),there was no statistical difference at other time points.2.Effects of two sources TG on joint histopathology and articular cart ilage morphology in CIA model ratsHE staining results showed that the joint tissue pathology improved after TG administration in two sources.QJ-TG18mg·kg-1·d-1 can significantly improve inflammatory cell infiltration(P<0.01)and bone destruction(P<0.05),QJ-TG54 mg·kg-1·d-1 can significantly inhibit joint pannus,cartilage and bone destruction(P<0.01);DED-TG18mg·kg-1·d-1 significantly improved bone destruction(P<0.05),and inhibition of inflammatory infiltration,pannus,cartilage destruction and bone destruction by DED-TG54mg·kg-1·d-1 Compared with the model group,t here was a significant statistical difference(P<0.01).There was no significant d ifference in the improvement of semi-quantitative scores of joint histopathology between CIA rats and the same dose of TG.By observing the Masson staining,the cartilage matrix and collagen fibers in the normal group were blue,uniform in coloration,gradually deepening blu e toward the calcification layer,clear tidal line.The boundary between the cartil age and the subchondral bone is clear,the dense bone and trabecular bone of the subchondral bone are blue,and the collagen is red;The surface of the arti cular cartilage in the model group was rough and absent,and the surface cartil age was degenerated and stained.The fibrosis was obvious.There were a lot o f flaming protuberances near the tidal line,the trabecular bone was lost,and t he boundary between cartilage and subchondral bone was not clear.Compared with the model group,the articular cartilage morphology of TG of the two ma nufacturers group was improved.The surface of the articular cartilage in the lo w dose group was smooth,uniform in coloration,a small amount of fibrosis,a nd a small amount of flaming protrusions near the tidal line.The high-dose gr oup dose of articular cartilage surface was smooth,uniform coloration,no fibro sis.3.Effects of two sources TG on bone destruction of rat joints in CIA modelTRAP staining showed that there was no osteoclast infiltration in the artic ular cartilage and a small amount of osteoclasts in the subchondral bone in the normal group.The model group showed a large area of positive staining area,and there were a large number of wine red granules around the trabecular bo ne marrow,indicating obvious bone resorption.Compared with the model grou p,the number of osteoclasts in the articular subchondral bone ofTG of the two manufacturers group was significantly reduced.The positive staining area and the number of osteoclasts in the QJ-TG54mg·kg-1·d-1 group and the DED-TG54 mg·kg-1·d-1 group were significantly less than those in the QJ-TG18mg·kg-1·d-1 group and DED-TG18mg·kg-1·d-1 group,bone destruction is lighter.4.Effects of two sources TG on general condition,blood and organ co efficient of CIA model rats(1)There was no significant difference in the body weight of CIA model rats between the two sources.(2)Blood test results of rats after 21 days and 42 days showed blood rou tine indicators of each dose group after two TG administrations:red blood cell count(RBC),white blood cell count(WBC),platelet(PLT),hemoglobin(HG B),monocyte ratio(MONO%)and neutral cell ratio(NEUT%)did not change significantly.After the TG administration of the two manufacturers,the numbe r of white blood cells was significantly increased in the dose group of DED-T G54g·kg-1·d-1(P<0.01),and no other changes were observed(3)Compared with the model group,there was no obvious abnormality i n the diet and defecation of the rats after TG administration.DED-TG54mg·kg-1·d-1 significantly increased spleen index(P<0.01),and no significant changes were observed in other drug-administered groups.HE staining of gastric section s of rats in the TG administration group showed no damage such as bleeding.5.Effects of two sources TG on liver and kidney function in rats with CIA modelThe liver and kidney function of the rats were measured after 21 days an d 42 days of administration.The results showed that alanine aminotransferase(ALT),aspartate aminotransferase(AST),γ-transpeptidase(GGT),total bilirubin(TBIL),blood Creatinine(CRE)and urea(UREA)showed no obvious abnorm alities.After TG administration in two manufacturers,the HE staining results o f liver and kidney tissue sections at the same time point showed that the liver cell structure and morphology were not abnormal and no abnormalities were f ound in renal tissue structure,glomeruli and renal tubules.6.Effects of two sources TG on the reproductive system of CIA mode 1 rats(1)Two dose groups of QJ-TG and DED-TG can produce higher sperm d eformity rate and a certain degree of testicular damage after 21 days and 42 d ays of administration.The spermatogenic cells are disorderly arranged,the inter stitial cells are damaged,the volume of the seminiferous tubules is reduced,th e gap between the seminiferous tubules is increased,the lumen is empty,and s ome are not lumens.And the toxicity increased with the increase of dose,and the toxic effect was stronger than 21 days in 42 days.Compared with the T G of the two manufacturers,the degree of testicular lesions and sperm deformi ty of DED-TG was higher than that of the same dose of QJ-TG.The levels of testosterone T,CYP19A1 and iNOS in the TG intervention group of the two manufacturers at 21d and 42d after administration were not significantly change d by the detection of serum reproductive hormone levels.TG does not have a role in lowering testosterone T in CIA rats and increasing the rate-limiting en zymes CYP19A1 and iNOS associated with androgen synthesis.(2)In the QJ-TG and DED-TG54mg·kg-1·d-1 dose groups,the vasculature of the ovary was scattered and showing a decreasing trend,and the corpus lut eum volume was reduced and the boundary was not clear at 21 and 42 days of administration.There is no significant difference between the two manufactu rers.Other abnormal changes have not been observed.The effect of TG on th e expression of estrogen and hormone synthesis-related rate-limiting enzymes C YP19A1 and CYP17A1 in CIA rats was not significant and there was no stati stical difference.The difference between the TG groups of two different manuf acturers was also not obvious.Conclusion:Two manufacturers TG2 times(18mg·kg-1·d-1)and 6 times(54mg·kg-1·d-1 clinical equivalent doses can delay the onset of CIA in rats,reduc e the clinical score of arthritis,and improve the pathological changes of joints.And there is a certain degree of male reproductive toxicity.The high-dose D ED-TG is more toxic than the QJ-TG. |
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