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OBJECTIVE:To study the effect of Zuogui pill on the expression of gap junction protein 43(Cx43)in ovarian tis-sue of mice with premature ovarian failure(POF)induced by intraperitoneal injection of cyclophosphamide(CTX),and explore its mechanism in the treatment of chemotherapy-induced POF. METHODS:72 mice were randomly divided into blank group,model group,Estradiol valerate tablet group (positive control,0.00013 g/kg),Zuogui pill low-dose,medium-dose,high-dose groups (13.65,40.95,122.85 g/kg)by body mass,10 in each group. Except for blank group,other groups were reduce POF model by ip CTX,once a day,for 20 d. Meanwhile,the mice were intragastrically administrated related medicines,once a day,for 30 d. After 2 h of last administration,reverse transcription-polymerase chain reaction method and Western blot method were used to detect the Cx43 mRNA and protein expressions in ovarian tissue respectively,and immunohistochemistry method was adopted to detect the distribution of Cx43 protein in ovarian tissue. RESULTS:Compared with blank group,Cx43 mRNA and protein expressions in ovarian tissue of mice in model group were obviously weakened,and the distribution of Cx43 protein in follicle and granulocytes were obviously reduced(P<0.01). Compared with model group,Cx43 mRNA and protein expressions in ovarian tissue of mice in Estradiol valerate tablet group and Zuogui pill medium-dose,high-dose groups were obviously strengthened,and the distribution of Cx43 protein in follicle and granulocytes were obviously increased(P<0.01). CONCLUSIONS:Zuogui pill can increase the Cx43 mRNA and protein expressions in ovarian tissue of CTX-induced POF mice,increase the distribution of Cx43 in follicle and granu-locytes and gap junction function,which may be one of the treatment mechanism of POF.
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Objective To explore the effects of bioactive parts of Xiongma Decoction (parts of ethyl acetate and n-butanol extract) on the CGRP-CRLR/RAMP1 signal pathway so as to clarify its therapeutic mechanism on migraine.Methods We randomly divided 36 male SD rats into 6 groups with 6 in each:blank group,model group,groups of low-,medium-and high-dose Xiongma Decoction bioactive parts,and Sumatriptan group.By giving hypodermic injection of 10 mg/kg nitroglycerin,the migraine rat model was copied;Only 18 rats were found to have positive expressions of CGRP,CRLR,and RAMP1 in TCC with immunohistochemistry staining after heart perfusion.For the remaining 18 rats,TCC was stripped directly from the whole brain and divided into two parts,one part used to detect CGRP,CRLR,RAMP1 mRNA expressions by qPCR,and the other part to detect CGRP,CRLR,RAMP1 protein expressions by Western blot.Results The number of CGRP,CRLR and RAMP1 immunoreactive cells,the mRNA and protein expressions on TCC in model group were effectively increased,compared with those in the blank group (P＜0.05),indicating that the model copying was successful.Compared with those in the model group,the number of CGRP,CRLR and RAMP1 immunoreactive cells in Xiongma Decoction bioactive parts was significantlv decreased,and the expressions of CGRP,CRLR and RAMP1 mRNA and protein were reduced (P＜0.05).Conclusion The bioactive parts of Xiongma Decoction can reduce the activity of CGRP-CRLR/RAMP1 signal pathway in TCC of migraine rats.
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Objective To observe the influence of silent information regulator factor 1(SIRT1)on TNF-αinduced intestinal epi-thelial Caco-2 cell barrier function destroy and to investigate its molecular machenism.Methods Caco-2 cells were randomly divided into three groups:normal control group (control),TNF-αgroup (TNF-α,100 ng/mL for 24 h)and 100 ng/mL plus 40μm resvera-trol group (TNF-α+Res).Transepithelial electrical resistance (TER)was determined.SIRT1 and the protein expressions of ZO-1 , occludin were examined by using Western blot.Results The relative expression amounts of SIRT1 protein were 0.81 ± 0.02, 0.43±0.04 and 0.60±0.03 respectively.TER of three groups were (154.00±5.00),(97.00±4.00)and(128.00±6.00)Ohm/cm2 respectively.Compared with the control group,the expression of SIRT1 protein was reduced by 47% and TER was decreased by 37.00% in the TNF-αgroup.After resveratrol precondition,TER was increased by 32.00% compared with the TNF-αgroup. The relative expression amounts of ZO-1 and occludin protein in the control group,TNF-αgroup and TNF-α+Res group were (0.62±0.06,0.57±0.03),(0.23±0.05,0.33±0.04)and(0.41±0.03,0.50±0.02)respectively.After TNF-αtreatment,the ex-pressions of ZO-1 and occludin protein were significantly deceased(P<0.05),but the resveratrol precondition could attenuate this phenomenon,compared with TNF-αgroup,the protein expression was increased by 78.00% and 51.00% respectively (P<0.05). Conclusion Under the condition of TNF-αtreatment,the SIRT1 level is decreased,but increasing SIRT1 level could increase the in-testinal tight j unction protein ZO-1 and occludin protein expression,thus alleviate the damage of TNF-αon the epithelial barrier function constituted by Caco-2 cells.
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<p><b>OBJECTIVE</b>To observe the effect of SIRT1 on intestinal barrier function of epithelial Caco-2 cells under hypoxia and investigate its mechanism.</p><p><b>METHODS</b>Caco-2 cells were randomly divided into three groups: normoxia group (Nx), hypoxia group (Hx,1%O2 for 6 h) and hypoxia plus 40 μmol/L Resveratrol (agonist of SIRT1) group (Hx+Res). Transepithelial electrical resistance (TER) was determined. mRNA and protein expressions of SIRT1 and tight junctions (ZO-1, Occludin, Claudin-1) were examined by reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting.</p><p><b>RESULTS</b>Both mRNA and protein expressions of SIRT1 were significantly reduced in Hx group as compared with Nx group (0.40±0.02 vs. 0.70±0.07, P=0.001; 0.37±0.03 vs. 0.76±0.03, P=0.001). The mRNA and protein expressions of SIRT1 were significantly increased in Hx+Res group as compared with Hx group(0.50±0.02 vs. 0.40±0.02, P=0.026; 0.54±0.02 vs. 0.37±0.03, P=0.011). The expression levels of ZO-1, Occludin and Claudin-1 in Hx group were lower than those in Nx group (P<0.05), however, pretreatment with Resveratrol could attenuate the decreased expression of above 3 molecules under hypoxia(P<0.05). TERs of Nx group, Hx group and Hx+Res group were (142±7) Ohm/cm(2), (94±3) Ohm/cm(2) and (119±7) Ohm/cm(2) respectively. Compare with the Nx group, the TER of Hx group was significantly decreased(P<0.05). TER of Hx+Res group was significantly increased compare with Hx group, but it was still significantly lower than that in Nx group(P<0.05).</p><p><b>CONCLUSIONS</b>Expression of SIRT1 is significantly reduced under hypoxia. Activation of SIRT1 can maintain the epithelial barrier function through regulating the expression of tight junctions under hypoxia.</p>
Subject(s)
Humans , Caco-2 Cells , Cell Hypoxia , Claudin-1 , Metabolism , Epithelial Cells , Metabolism , Intestinal Mucosa , Cell Biology , Occludin , Metabolism , Sirtuin 1 , Metabolism , Zonula Occludens-1 Protein , MetabolismABSTRACT
Objective To observe the effects of mesenchymal stem cells (MSCs)injection from tail-vein on the expression of glial cell line derived neurotrophic factor (GDNF) after the spinal cord injury (SCI) of rats.MethodMSCs were cultured from the thighbone of adult wistar rats.The T10 level spinal cord injury was execute by the modify-Allen's device. The MSCs labeled by bromodeoxyuridine were transplanted into the vein of tail by injection immediately after spinal cord injury. Adult Wistar rats were randomly divided into three groups: SCI cured with MSCs transplantation (group A), SCI received normal saline(group B),and control group (group C). Then MSCs were detected and the expressions of GDNF of the lesion and neighbor areas were examined by Reversetranscription polymerase chain reaction (RT-PCR) and immunohistochemistry.ResultImmunohistochemistry: MSCs labeled by bromodeoxyuridine could be detected in the injuried spinal cord after transplantation. It was located at the nucleolus of MSCs. The cells survived and migrated rostrallyand caudally from the injection sites. With the time passed, the number of MSCs labeled with bromodeoxyuridine was decreased. RT-PCR: The expression of Group A was increased on the followed day, and the expression of Group B was increased on the 1st day but decrease from the 5th day then on.compared with group B, transplantation of MSCs significantly enhanced the expression of GDNF mRNA than group A on the 1st, 3rd, 5th day after cell transplantation(P