Mitogen-activated protein kinases (MAPKs), including extracellular signal-regulated kinases (ERKs), c-Jun NH2-terminal kinases (JNKs) and p38 MAPK, play an important role in transducting environmental stimuli to the t...Mitogen-activated protein kinases (MAPKs), including extracellular signal-regulated kinases (ERKs), c-Jun NH2-terminal kinases (JNKs) and p38 MAPK, play an important role in transducting environmental stimuli to the transcriptional machinery in the nucleus in mammalian cells by virtue of their ability to phosphorylate and regulate the activity of various transcription factors. It was recently found that the changes in activity of MAPKs occurred during ischemia/reperfusion, but the biological significance of the changes was still controversial.展开更多
Objective To detect if Fas is expressed in human renal interstitial fibroblasts (hRIFs) and apoptosis of hRIFs can be induced by specific anti-Fas antibody. Methods hRIFs were cultured from isolated papillae of huma...Objective To detect if Fas is expressed in human renal interstitial fibroblasts (hRIFs) and apoptosis of hRIFs can be induced by specific anti-Fas antibody. Methods hRIFs were cultured from isolated papillae of human kidney, and identified by morphologic examination, assay of antigenic components and culture in D-valine selective medium. Fas expression in normal hRIFs was detected by RT-PCR and immunocytochemistry staining. After hRIFs were incubated with interferon gamma (γ-IFN 500 U/ml, 1000 U/ml, 1500 U/ml and 2000 U/ml, respectively) for 48 hours, Fas expression was determined by Northern blot, Western blot and flow cytometry. hRIFs pre-stimulated with γ-IFN (500 U/ml, 48 hours) were incubated with anti-Fas antibody (IgM, 0.5 μg/ml) for 12 hours. And apoptosis was identified by morphologic examination, DNA ladder assay and flow cytometry.Results The cultured hRIFs showed a shuttle-like shape and were positively stained by labeled anti-vimentin antibody but negatively stained by anti-epithelial membrane antibody. They could not grow in the D-valine selective medium and died partly in a week. Fas mRNA and protein were expressed in normal hRIFs and markedly upregulated by stimulation with γ-IFN. Apoptosis in γ-IFN pre-stimulated hRIFs was induced by anti-Fas antibody, showing cell nuclear shrinkage and condensation in morphologic feature, internucleosomal DNA fragmentation in DNA ladder assay and a pick of hypo-diploid nuclei by flow cytometry. Conclusion Fas is normally expressed in hRIFs and can be markedly upregulated by γ-IFN. Anti-Fas antibodies can induce apoptosis of hRIFs pre-stimulated with γ-IFN.展开更多
文摘Mitogen-activated protein kinases (MAPKs), including extracellular signal-regulated kinases (ERKs), c-Jun NH2-terminal kinases (JNKs) and p38 MAPK, play an important role in transducting environmental stimuli to the transcriptional machinery in the nucleus in mammalian cells by virtue of their ability to phosphorylate and regulate the activity of various transcription factors. It was recently found that the changes in activity of MAPKs occurred during ischemia/reperfusion, but the biological significance of the changes was still controversial.
文摘Objective To detect if Fas is expressed in human renal interstitial fibroblasts (hRIFs) and apoptosis of hRIFs can be induced by specific anti-Fas antibody. Methods hRIFs were cultured from isolated papillae of human kidney, and identified by morphologic examination, assay of antigenic components and culture in D-valine selective medium. Fas expression in normal hRIFs was detected by RT-PCR and immunocytochemistry staining. After hRIFs were incubated with interferon gamma (γ-IFN 500 U/ml, 1000 U/ml, 1500 U/ml and 2000 U/ml, respectively) for 48 hours, Fas expression was determined by Northern blot, Western blot and flow cytometry. hRIFs pre-stimulated with γ-IFN (500 U/ml, 48 hours) were incubated with anti-Fas antibody (IgM, 0.5 μg/ml) for 12 hours. And apoptosis was identified by morphologic examination, DNA ladder assay and flow cytometry.Results The cultured hRIFs showed a shuttle-like shape and were positively stained by labeled anti-vimentin antibody but negatively stained by anti-epithelial membrane antibody. They could not grow in the D-valine selective medium and died partly in a week. Fas mRNA and protein were expressed in normal hRIFs and markedly upregulated by stimulation with γ-IFN. Apoptosis in γ-IFN pre-stimulated hRIFs was induced by anti-Fas antibody, showing cell nuclear shrinkage and condensation in morphologic feature, internucleosomal DNA fragmentation in DNA ladder assay and a pick of hypo-diploid nuclei by flow cytometry. Conclusion Fas is normally expressed in hRIFs and can be markedly upregulated by γ-IFN. Anti-Fas antibodies can induce apoptosis of hRIFs pre-stimulated with γ-IFN.
