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Objective:To explore and optimize the preparation method of specimen about mouse liver tissue for scanning electron microscopy.Methods: The specimens of mouse liver using for scanning electron microscope were prepared through improving the fixed location of perfusion, optimizing material and procedure of perfusion, adopting fixation of perfusion via portal vein and changing the process of fixation and so on. And the results of morphological observation of histocyte were compared between the improved method and traditional method.Results: The improved preparation method could save the liver tissue and the living state of the cells in preferably situation. By using this method, the ultra-micro morphological structure of liver tissue and cell could be clearly observed and the ideal effect of observation could be achieved. Besides, its operation was simple and its success rate was high. Therefore, it was superior to traditional method.Conclusion: The preparation of specimen about the fixation of mouse liver tissue via perfusion of portal vein for scanning electron microscope is a superior method, which has certain applied value.
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Objective To establish the models with human rheumatoid arthritis (RA) synovial fibroblasts (RASFs)-cartilage-severe combined immune deficient (SCID) mice for the study of the RASFs in the pathogenesis of RA.Methods The 4th passage RASFs were marked with 5-bromodexyuridine (5-Brdu)and injected into a cavity of inert sterile gel sponge,then with the normal human cartilage co-implanted in the back subcutaneously of SCID mice to set up a novel model of RA.Osteoarthritis synovial fibroblasts (OASFs)were injected as control group.Thirty days after the surgery,the mice were killed,the grafts and the knee joints were proceed with histological study and immunohistochemistry to detect the expression of 5-Brdu and Vimentin in synoviocytes.The serum level of interleukin (IL)-6 and matrix metalloproteinase (MMP)-3 were detected with enzyme-linked immunosorbent assay (ELISA).Results Twenty-three mice survived except for one mouse in the RASFs group died of anesthesia.① Only in one case in the RASFs group,the IL-6 was detected,the others were unable to be detected.The MMP-3 in the OASFs group was (40±17) pg/ml,but in RASFs group only one case was detected.② There were 4 and 3 implanted cartilages loss in the RASFs group and OASFs group respectively.The histological scores of cartilage invasion by synoviocytes and cartilage degradation in grafts were higher in RASFs groups than in OASFs groups (0.6±0.7 vs 0.3±0.5,2.3±0.8 vs 1.7±1.0 respectively).③ The histological scores of synovial hyperplasia and cartilage invasion in the knee joints was significantly higher in the RASFs group than in the OASFs group (3.1±0.8 vs 1.7±1.0,P＜0.01,1.6±1.7 vs 0.6±1.4,P＜0.05 respectively).④ During the grafts,a lot of 5-Brdu and Vimentin markers positive synovio-cytes were found in the mice subcutaneous tissue,but manipulus positive synoviocytes were found on the area of cartilage invasion in both groups.In knee joints,single positive synoviocytes could be detected in bone marrow and hyperplasic area of the synovial tissue in both groups.Conclusion The isolated RASFs can survival and have the ability to invade and degrade the cartilage in vivo without the limitation of immunity induced inflammations,and can also migrate to the synovial joints in distance and induce arthritis.
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Aim To establish and evaluate the simple and effective animal model of rheumatoid arthritis in SD rats induced by heat-killed mycobacterium tuberculosis H37Ra(Mtb).Methods SD rats were immunized s.c.at the base of the tail with Mtb(1 mg/rat) in mineral oil,and then body weight,blood routine,clinical signs of arthritis were observed regularly.And also hind paw volume in SD rats with AA was tested.Meanwhile,T-lymphocyte subset was determined by flow cytometry in peripheral blood,the level of TNF-?in serum and IL-1 and IL-6 and vascular endothe-lial growth factor(VEGF) in joint tissue were tested by ELISA.The ankle pathological change and radiographic evidence of the hind paws of SD rats were observed to evaluate the AA model effects and availability in SD rats.Results When compared with normal control rats,the total number of white blood cells,platelets and monocytes in peripheral blood and ratio of CD4/CD8 was markedly raised after immunization with Mtb.But body weight of model rats was significantly low from day 14 to 28 after injection of Mtb.The inflammatory arthritic lesions such as edema and erythema in the paws appeared after 9～10 d.The peak of inflammation was 14～16 d.The level of TNF-?in serum and IL-1,IL-6 and VEGF in joint tissue was significantly higher in SD rats with Mtb than that in normal control rats at d 28.The histopathological examination revealed cellular infiltration,synovial and cartilage hyperplasia in ankle joints of SD rats.Conclusions The model induced by Mtb in SD rats is more similar in clinical characterization of rheumatoid arthritis in human being.It is a more efficient and simpler manipulative procedure for screening of new anti-arthritis drugs and experimental studying of anti-rheumatoid arthritis.