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Objeetive To improve the method for the isolation and purification of rat hepatic stellate(HSC) cells and to provide a stable cell source for the research on liver-related diseases.Methods Rat liver was digested in situ by a two-step infusion assay under a strict control of the infusion temperature,flow rate and time with a combined utilization of Pronase E and Collagenase Ⅳ.And then,the HSC cells were separated by Percoll density gradient centrifugation.The cell growth curve and survival rate were measured by CCK-8 and trypan blue staining,respectively.The HSC cells were identified by flow cytometry and immunofluorescence cytochemistry.Results With the improved methods,there were (2.1 ± 0.2) × 107 HSC cells isolated from one rat and the survival rate was (96.2 ± 0.8) ％.The percentage of HSC cells with a spontaneous fluorescent characteristic from the isolated cells was 96.3％.The immunofluorescence cytochemistry was used to detect the expressions of the surface antigens α-SMA and Desmin in the isolated HSC cells.Conclusion By strict control of infusion temperature,flow rate and perfusion time as well as the combined application of Pronase E and Collagenase Ⅳ,there is an increased harvest of HSC cells with improved cell viability and purity,which is helpful for further research on HSC cells.
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<p><b>OBJECTIVE</b>To study the effects of different dietary fatty acid on the expression of nuclear receptor genes in the breast cancer of rats.</p><p><b>METHODS</b>Fifty-day-old female Sprague-Dawley rats were fed on eight different diets containing following fatty acids: saturated fatty acid (SFA); monounsaturated fatty acid (MUFA); n-6 polyunsaturated fatty acid (PUFA); n-3 PUFA; 1:1 n-6/n-3; 5:1 n-6/n-3; 10:1 n-6/n-3; 1:2:1 S/M/P (n-6/n-3 at 1:1). The rats were given a single intraperitoneal injection of methyl-nitrosourea (MNU) at 50 mg/kg body weight to establish the rat model of mammary carcinogenesis, the ultrastructure changes of mammary gland cells in rats were observed by transmission electron microscope, the cell proliferation activity was detected by BrdU-labeled immunocytochemistry, and the expression of PPARbeta and PPARgamma mRNA were assayed by RT-PCR.</p><p><b>RESULTS</b>There was no breast cancer occurring in control groups and the MNU-treated n-3 PUFA group, and the ultrastructure and proliferation activity of mammary gland cells in these groups were normal. In contrast, there appeared obvious marker of adenocarcinomas in mammary gland cells of MNU-induced breast cancer, and a high cell proliferation activity was found in tumor growth-enhancing groups (SFA, MUFA, n-6 PUFA, 5:1 n-6/n-3, 10:1 n-6/n-3 and S/M/P, 21% - 22% of BrdU-labeled cells), while a low cell proliferation activity was detected in rats fed with 1:1 n-6/n-3 diet (13% of BrdU-labeled cells, P < 0.05). Moreover, peroxisome proliferator-activated receptors (PPARs), as important nuclear receptor genes of relating lipid metabolism, the expressions of PPARbeta and PPARgamma mRNA were significantly up-regulated in mammary adipose tissues of MNU-induced breast cancer as compared with the control groups, but the expression levels of peroxisome proliferator-activated receptors (PPARs) in rats fed with 1:1 n-6/n-3 group were lowest (P < 0.05).</p><p><b>CONCLUSION</b>The different dietary fatty acid compositions should diversely adjust the expression of PPARs gene in rats, which maybe have an important role in affecting incidence of breast cancer.</p>
Subject(s)
Animals , Female , Rats , Fatty Acids , Pharmacology , Fatty Acids, Omega-3 , Pharmacology , Fatty Acids, Omega-6 , Pharmacology , Fatty Acids, Unsaturated , Pharmacology , Gene Expression , Gene Expression Regulation, Neoplastic , Mammary Neoplasms, Experimental , Genetics , PPAR gamma , Genetics , RNA, Messenger , Genetics , Rats, Sprague-DawleyABSTRACT
<p><b>OBJECTIVE</b>To study the effect of lovastatin on the expression of IkappaBalpha and cell-cycle regulating proteins in MCF-7 cells.</p><p><b>METHODS</b>MCF-7 cells were treated with 4, 8 and 16 micro mol/L lovastatin for 48 - 72 h. The distribution of cell cycles was assayed by flow cytometry (FCM). The protein expression of IkappaBalpha, CDK4, p16, pRb in cytoplasm and IkappaBalpha in the nucleus were detected by Western blot.</p><p><b>RESULTS</b>Lovastatin could arrest cellcycle in the G(0)/G(1) phase in a dose- and time-dependent manner, obviously lowering the expression of IkappaBalpha, CDK4 and pRb protein level in the cytoplasm and increasing IkappaBalpha in the nucleus, but not on p16 protein level.</p><p><b>CONCLUSION</b>Lovastatin can induce the arrest of cell cycle in G(0)/G(1) phase by affecting the expression of IkappaBalpha and cell-cycle regulating protein in MCF-7 cells.</p>
Subject(s)
Female , Humans , Antineoplastic Agents , Pharmacology , Breast Neoplasms , Metabolism , Pathology , Cell Cycle , Cell Cycle Proteins , Metabolism , Cell Line, Tumor , Cyclin-Dependent Kinase 4 , Cyclin-Dependent Kinase Inhibitor p16 , Metabolism , Cyclin-Dependent Kinases , Metabolism , I-kappa B Proteins , Metabolism , Lovastatin , Pharmacology , NF-KappaB Inhibitor alpha , NF-kappa B , Proto-Oncogene Proteins , Metabolism , Retinoblastoma Protein , MetabolismABSTRACT
Objective To explore the metabolism of micronutri ents related to dark adaptation of radar operators through nutritional intervent ion. Methods A total of 34 male radar operators aged between 18 ～29 years old were randomly divided into control and experimental groups. The c ontrol group were on normal diet, and the experimental group received the supple ment of VA, Zn and Se in additional to normal diet. The experiment lasted 4 week s. The levels of serum VA, Zn and Se were measured before and after the experime nt. Results The levels of serum VA, Zn and Se in the experime ntal group were significantly higher than that in the control group after the experiment (P<0.01). Conclusion The supplement of VA, Zn an d Se for 4 weeks may elevate, the levels of serum VA, Zn and Se significantly (reached or surpassed normal levels) and suggests that VA, Zn and Se su pplementation may effectively enhance the dark adaptation of radar operators.
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Objective To investigate the expression of TGF-β and TGF-β receptor in human breast cancer cell Bcap-37 inhibited by soybean isoflavones. Methods mRNA and protein of TGF-β1、TGF-βRⅠ in Bcap-37 cells were examined with reverse transcription ploymerase chain reaction(RT-PCR) and immunohistochemistry after cells were treated with daidein or genistein for 1-4 d.The expression of TGF-β1 and TGF-β2 was determined with TGF-β resistance test. Results The TGF-β1, TGF-β2 and TGF-β recepor increased in Bcap-37 cells at a concentration of 3×10-5 mol/L of genistein. No changes was found when treated with daidzein. Conclusion Genistein may inhibit the proliferation of Bcap-37 cells and accompany with increasing expression of TGF-β1, TGF-β2 and TGF-β receptor.
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Objective To explore the metabolism of micronutri ents related to dark adaptation of radar operators through nutritional intervent ion. Methods A total of 34 male radar operators aged between 18 ～29 years old were randomly divided into control and experimental groups. The c ontrol group were on normal diet, and the experimental group received the supple ment of VA, Zn and Se in additional to normal diet. The experiment lasted 4 week s. The levels of serum VA, Zn and Se were measured before and after the experime nt. Results The levels of serum VA, Zn and Se in the experime ntal group were significantly higher than that in the control group after the experiment (P<0.01). Conclusion The supplement of VA, Zn an d Se for 4 weeks may elevate, the levels of serum VA, Zn and Se significantly (reached or surpassed normal levels) and suggests that VA, Zn and Se su pplementation may effectively enhance the dark adaptation of radar operators.
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Objective To investigate the expression of TGF-β and TGF-β receptor in human breast cancer cell Bcap-37 inhibited by soybean isoflavones. Methods mRNA and protein of TGF-β1、TGF-βRⅠ in Bcap-37 cells were examined with reverse transcription ploymerase chain reaction(RT-PCR) and immunohistochemistry after cells were treated with daidein or genistein for 1-4 d.The expression of TGF-β1 and TGF-β2 was determined with TGF-β resistance test. Results The TGF-β1, TGF-β2 and TGF-β recepor increased in Bcap-37 cells at a concentration of 3×10-5 mol/L of genistein. No changes was found when treated with daidzein. Conclusion Genistein may inhibit the proliferation of Bcap-37 cells and accompany with increasing expression of TGF-β1, TGF-β2 and TGF-β receptor.