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OBJECTIVE：To study the effects of couplet medicine of Rheum p almatum-Salvia miltiorrhiza on the contents of enterogenous urotoxin and intestinal barrier function in chronic renal failure （CRF）model rats. METHODS ：Totally 55 male Wistar rats were randomly divided into sham operation group （10 rats）and modeling group （45 rats）. In sham operation group ，the kidneys were isolated but not removed ；CRF model was reproduced by 5/6 nephrectomy in modeling group. After modeling （excluding 5 dead and non-modeling rats ），modeling rats were divided into model group （water），Niaoduqing granules group （2.5 g/kg），couplet medicine of R. palmatum -S. miltiorrhiza groups（6，3 g/kg，by crude drug ），with 10 rats in each group. Sham operation group and model group were given constant volume of water intragastrically. Administration groups were given relevant medicine intragastrically ，once a day ，for consecutive 12 weeks. After last administration ，the contents of creatinine （Scr）and urea nitrogen（BUN）in serum ，the content of urinary creatinine （Ucr） in urine were determined by automatic biochemical analyzer；creatinine clearance rate （Ccr）was calculated. The contents of enterogenous urotoxin [trimethylamine-N-oxide （TMAO），indoxyl sulfate （IS）and p-cresyl sulphate （PCS）] were determined by UPLC-ESI-MS/MS. Real-time RT-PCR and immunofluorescence assay were used to detect the mRNA and protein expression of Occludin and ZO-1 in the ileum tissue. HE staining and Masson staining were used to observe the pathologi cal changes of renal tissue. The ultrastructural changes of rat colon were observed by transmission electron microscope. RESULTS ：Compared with sham operation group ，serum contents of Scr，BUN，TMAO，PCS and IS were increased significantly in model group （P＜0.01），while urine content of Ucr ，Ccr，mRNA and protein expression of Occludin and ZO- 1 in ileum tissue were decreased significantly （P＜0.01）；renal glomerulosclerosis ， renal tubules dilation and inflammatory invasion and fibrosisin the interstitium were all found ；the intestinal epithelial barrier structure of colon tissue was severely damaged. Compared with model group ，serum contents of Scr ，BUN，TMAO，PCS and IS were decreased significantly in administration groups （P＜0.05 or P＜0.01）；the mRNA and protein expression of Occludin and ZO-1 in the ileum tissue were increased significantly （except for mRNA expression of ZO- 1 in R. palmatum -S. miltiorrhiza low-dose group （P＜0.05 or P＜0.01）；the infiltration of inflammatory cells in renal interstitium ，the degree of fibrosis and the damage of intestinal epithelial barrier structure in colon tissue were reduced. CONCLUSIONS ：Couplet medicine of R. palmatum -S. miltiorrhiza can effectively protect the residual renal function of CRF model rats ，the mechanism of which may be associated with reducing the serum contents of enterogenous urotoxin ，up-regulating mRNA and protein expresssion of Occludin and ZO- 1 in the ileum tissue so as to improve intestinal barrier function.
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Objective To observe the host response and the expression of interleukin-4 (IL-4) in different cross-linked hyaluronic acid composite gels at different time points after the implantation in vivo, and to explore the significance of biocompatibility and macrophage polarization in post-implantation inflammatory response and tissue remodeling. Methods New Zealand white rabbits were respectively injected with crosslinked hyaluronic acid-crosslinked hydroxypropyl methylcellulose gel (sample 1), crosslinked hyaluronic acid-hydroxypropyl methylcellulose gel (sample 2) and commercially available modified sodium hyaluronate gel (control) in subcutaneous tissue at both sides of the spine. Then the rabbits were dissected at 1, 4 and 12 weeks after the implantation. The tissues were fixed with 10％formaldehyde solution, embedded in paraffin and sliced. Hematoxylin-eosin (HE) staining was performed to observe the degree of inflammation and fibrosis. Masson staining was performed to observe the formation of collagen fibers. Immunohistochemical staining was performed to observe the expression of IL-4. Results The results of HE staining showed that the inflammatory reaction in the sample 1 and sample 2 groups was significantly higher than that in the control group 1 and 4 weeks after the implantation. The inflammatory cells aggregated, and the wall of capsule and microcapsule was thick. The sample 1 group was more obvious, and the result was mild stimulation. For all the groups, the results were all non-irritating at 12 weeks after the implantation. The results of Masson staining showed that the collagen fibers in the sample 1 and sample 2 groups were increased compared with the control group, mainly distributed around the implantation site, and a small amount among the gels after 1 and 4 weeks. After 12 weeks, the collagen fibers were further increased, especially among the gels, which were consistent with the control group. The results of immunohistochemical staining showed that, at the same time point, the expression of IL-4 in sample 1 and sample 2 groups was higher than that in the control group, and the expression of IL-4 increased gradually with time. The expression of IL-4 in the control and sample 1 group at 12 weeks after the implantation was higher than that at 1 and 4 weeks respectively, and the differences were statistically significant (all P<0.05). In the sample 1 group, the expression of IL-4 at 12 weeks after the implantation was higher than that at 1 week, and the difference was statistically significant (P<0.05). The expression of IL-4 in the sample 1 group was significantly higher than that in the control group at 4 weeks after the implantation, and the difference was statistically significant (P<0.05). Conclusions The two different cross-linked sodium hyaluronate composite gels have good biocompatibility. The formation of collagen fiber and the expression of IL-4 can gradually increased within 12 weeks after the subcutaneous implantation, which is beneficial to the tissue remodeling.
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Objective To explore the mechanism of rapamycin-eluted stent in vascular endothelial injury. Meth?ods (1)Rapamycin (rapamycin group) was injected to rabbit dorsum muscle to simulate rapamycin-eluted stent implanta?tion into muscles. Control group and acetone control group were established at the same time. Morphological change in mus?cle was observed and serum calcium levels were measured after rapamycin injection.(2)HUVECs were incubated with 0.1, 1, 10 and 100μg/L rapamycin for 48 h respectively or with 1μg/L rapamycin for 6, 12, 24 and 48 h respectively.Cell via?bility was examined by MTT and its relationship with drug concentration and treatment time were analyzed.(3)HUVECs were divided into control group and 1μg/L rapamycin group. After 48 h,morphological changes of HUVECs were assessed by HE staining,the production of nitric oxide was examined by Nitric Oxide Assay Kit and the intracellular calcium ion con?centration was tagged with Fluo-3/AM. Results (1) Organizational morphology in local muscle with rapamycin injection represent stent implantation of rabbit,and calcium content in local muscle increased significantly in rapamycin group com?pared with nomal control group and acetone control group(P<0.05). (2) Cell survival rate decreased significantly upon ad?ministration of rapamycin in both concentration and time dependent manner(P<0.01). (3) In rapamycin group, cytoplasm vacuoles, nucleus pycnosis and nuclear fragmentation were observed;compared with control group,the levels of intracellular free Ca2+increased while the levels of nitric oxide was reduced. Conclusion Rapamycin treatment lead of injury to vascular endothelial cells which might through up-regulating intracellular Ca2+level.
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Objective To investigate the effects of ulinastatin (UTI) combined with magnesium isoglycyrrhizinate (MgIG) on the expression of transforming growth factor-β1 (TGF-β1) and connective tissue growth factor (CTGF) in lung tis?sue of rats with pulmonary fibrosis induced by bleomycin (BLM). Methods Ninety Wistar rats were randomly divided into five groups: BLM group, methylprednisolone (MTH) group, UTI group, MgIG group and UTI combined with MgIG (UTI+MgIG) group, n=18 for each group. The rat model of pulmonary fibrosis was established by injecting bleomycin through tra?chea in five groups. Twenty-four hours after treatment with BLM,rats were treated with normal saline every day in BLM group, and rats were treated by corresponding drugs in other groups. Six rats of each group were killed at the 7th,14th and 28th day respectively. The pathological changes of alveolitis and pulmonary fibrosis were evaluated by HE staining, and ex?pression levels of TGF-β1 and CTGF in lung tissues were detected by immunohistochemistry method. Results (1) Com?pared with BLM group, the degree of alveolitis and pulmonary fibrosis was reduced in other groups. There was significant dif?ference in alveolitis at the 7th and 14th day between UTI+MgIG group and BLM group. And there was significant difference in pulmonary fibrosis at the 14th and 28th day between UTI+MgIG group and BLM group (P<0.05). (2) Compared with BLM group, the expression levels of TGF-β1 and CTGF were decreased in other groups. In UTI+MgIG group, the expres?sion levels of TGF-β1 were significantly lower at the 7th and 14th day compared with those in UTI group and MgIG group, and which were significantly lower at the 28th day than those in MTH group, UTI group and MgIG group (P<0.05). The ex?pression levels of CTGF were significantly lower at the 7th day in UTI + MgIG group than those in UTI group and MgIG group, and which were significantly lower at the 14th and 28th day than those in MTH group, UTI group and MgIG group (P<0.05). Conclusion The combination of UTI and MgIG can alleviate alveolitis and fibrosis in BLM-induced pulmonary fibrosis rats, which might related with the down-regulation of TGF-β1 and CTGF expressions.