Objective: To study cyclinD1/bcl-1 and p27/kip1 expression in gliomas and their correlation with pathological grade and prognosis. Methods: Immunohistochemical technique was used to detect the cyclinD1/bcl-1 and p...Objective: To study cyclinD1/bcl-1 and p27/kip1 expression in gliomas and their correlation with pathological grade and prognosis. Methods: Immunohistochemical technique was used to detect the cyclinD1/bcl-1 and p27/kip1 expression in 48 human brain glioma tissues of different malignant grades, and 12 normal non-neoplastic tissues collected from internal decompression. The data were analyzed quantitatively by the image system and also correlated retrospectively with the patients' clinical characteristics. Results: The immunohistochemical reaction for cyclinD1/bcl-1 and p27/kip1 was confined to the nuclei. The abnormal positive expression rates of both cyclinD1/bcl-1 and p27/kip1 in gliomas were found higher than that in non-neoplastic tissues(P<0.05). The number and staining intensity of cyclinD1/bcl-1 positive nuclei increased with malignant grades (P<0.05). On the contrary, the positive nuclei of p27/kip1 expression decreased in number and staining intensity with malignant grades(P<0.05). Higher expression of cyclinD1/bcl-1 or/and lower expression of p27/kip1 were associated with poor prognosis(P<0.05). Conclusion: The abnormal expression of both cyclinD1/bcl-1 and p27/kip1 might be closely related to the occurrence and development of gliomas and they might have synergistic effect. These data suggest that both cyclinD1/bcl-1 expression and p27/kip1 expression can act as an independent prognostic factor.展开更多
Objective: To study p27/Kipl expression in gliomas and its application value. Methods: Imo munohistochemical technique was used to detect the exprssion of p27/Kipl gene in 48 different malignant grade human brain gl...Objective: To study p27/Kipl expression in gliomas and its application value. Methods: Imo munohistochemical technique was used to detect the exprssion of p27/Kipl gene in 48 different malignant grade human brain glioma tissues categorized according to WHO classification and 12 normal human brain tissues,which were analyzed quantitatively by using the image system and compared retrospectively with the patients' clinical characteristics. Results: In this series, the immunohistochemical reaction for p27/Kipl was confined to the nuclei. The abnormal positive expression rate of p27/Kipl in gliomas was found to be higher than that in normal tissues (P〈0.05). The positive nuclei expression of p27/Kipl decreased in number and staining intensity with the increasing degree of histological malignancy (P〈0.05). Lower expression of p27/Kipl was associated with poor prognosis (P〈0.05). P27/Kipl expression could be regarded as an independent prognostic factor. Conclusion: The abnormal expression of p27/Kipl may be closely related to the occurrence and development of gliomas, and also can be used to evaluate the prognosis independently.展开更多
Background RhoA/ Rho kinase (ROCK) pathway is involved in pulmonary arterial hypertension (PAH) and pulmonary artery smooth muscle cell (PASMC) proliferation. Inhibition of ROCK has been proposed as a treatment ...Background RhoA/ Rho kinase (ROCK) pathway is involved in pulmonary arterial hypertension (PAH) and pulmonary artery smooth muscle cell (PASMC) proliferation. Inhibition of ROCK has been proposed as a treatment for PAH. But the mechanism of RhoA/ROCK pathway and its downstream signaling in proliferation of human PASMCs is unclear. We investigated the effect of fasudil, a selective ROCK inhibitor, on platelet-derived growth factor (PDGF) induced human PASMC proliferation, and the possible association between RhoA/ROCK and extracellular signal-regulated kinase (ERK),p27KiP1.Methods Human PASMCs were cultured with the stimulation of 10 ng/ml PDGF, and different concentrations of fasudil were added before the addition of mitogen. Cell viability and cell cycle were determined with MTT and flow cytometry respectively. ROCK activity, ERK activity and protein expression of proliferating cell nuclear angigen (PCNA) and p27Kip1 were measured by immunoblotting.Results By MTT assay, PDGF significantly increased the OD value that represented human PASMC proliferation, and pretreatment with fasudil significantly reversed this effect in a dose-dependent manner. After PDGF stimulation, the percentage of cells in S phase increased dramatically from 15.6% to 24.3%, while the percentage in G0/G1 phase was reduced from 80.6% to 59%. And pretreatment with fasudil reversed the cell cycle effect of PDGF significantly in a dose-dependent manner. PDGF markedly induced ROCK activity and ERK activity with a peak at 15 minutes, which were significantly inhibited by fasudil. In addition, fasudil significantly inhibited PDGF-induced PCNA expression and fasudil also upregulated p27Kip1 expression in human PASMCs, which decreased after PDGF stimulation.Conclusion RhoA/ROCK is vital for PDFG-induced human PASMC proliferation, and fasudil effectively inhibited PDGF-induced human PASMC proliferation by up-regulation of p27Kip1, which may be associated with inhibition of ERK activity.展开更多
AIM: To investigate p27 expression in hepatocellular carcinoma (HCC), adjacent nontumoral and normal liver tissues, and to verify whether the subcellular localization of p27 was altered in HCC.METHODS: The level of p2...AIM: To investigate p27 expression in hepatocellular carcinoma (HCC), adjacent nontumoral and normal liver tissues, and to verify whether the subcellular localization of p27 was altered in HCC.METHODS: The level of p27 in tumoral, nontumoral, and normal liver tissues were assessed by immunohistochemical (IHC) analysis. Parallel immunostaining was done for proliferating cell nuclear antigen (PCNA) to evaluate cell proliferation.RESULTS: The labeling index (LI) of p27 in tumoral lesions was significantly lower than that in adjacent nontumoral lesions (t=2.444, P=0.017) and normal controls (t=-2.268,P=0.029). The LI of p27 significantly decreased in patients with massive type (t=-2.227, P=0.037) and infiltration (t=2.197, P=0.036). The prognosis of patients with higher p27 LI was longer than that of patients with lower p27 LI (P=0.0247, log-rank test). The LI of PCNA was significantly higher in HCC than that in adjacent nontumoral lesions (t=2.092, P=0.041) and normal controls (t=-3.533, P=0.002).There was no significant correlation between p27 expression and cell proliferation in tumor samples. The level of p27 in the cytoplasmic fraction was higher in tumoral and nontumoral liver tissues, and was associated with clinical stage (t=-2.520, P=0.029) and the degree of invasion (t=2.640, P=0.019). Survival analysis showed that p27 was an independent prognosis marker for HCC patients.CONCLUSION: These results suggest that p27 underexpressing in patients with HCC is closely associated with infiltration, metastasis, and prognosis. Alterations in the subcellular localization of p27 protein may occur early during hepatocarcinogenesis.展开更多
AIM: To elucidate the effect of p27^KIP1 on cell cycle and apoptosis regulation in gastric carcinoma cells. METHODS: The whole length of p27^KIP1 cDNA was transfected into human gastric cancer cell line SCG7901 by l...AIM: To elucidate the effect of p27^KIP1 on cell cycle and apoptosis regulation in gastric carcinoma cells. METHODS: The whole length of p27^KIP1 cDNA was transfected into human gastric cancer cell line SCG7901 by lipofectamine. Expression of p27^KIP1 protein or mRNA was analyzed by Western blot and RNA dot blotting, respectively. Effect of p27^KIP1 on cell growth was observed by MTT assay and anchorage-independent growth in soft agar. Tumorigenicity in nude mice was used to assess the in vivo biological effect of p27^KIP1. Flow cytometry, TUNEL, and electron microscopy were used to assess the effect of p27^KIP1 on cell cycle and apoptosis. RESULTS: Expression of p27^KIP1 protein or mRNA increased evidently in SCG7901 cells transfected with p27^KIP1. The cell growth was reduced by 31% at 48 h after induction with zinc determined by cell viability assay. The alteration of cell malignant phenotype was evidently indicated by the loss of anchorage-independent growth ability in soft agar. The tumorigenicity in nude mice was reduced evidently (0.55±0.14 cm vs 1.36±0.13cm, P〈0.01). p27^KIP1 overexpression caused cell arrest with 36% increase (from 33.7% to 69.3%, P〈0.01) in G1 population. Prolonged p27^KIP1 expression induced apoptotic cell death reflected by pre-G1 peak in the histogram of FACS, which was also confirmed by TUNEL assay and electron microscopy. CONCLUSION: p27^KIP1 can prolong cell cycle in G1 phase and lead to apoptosis, p27^KIP1 may be a good candidate for cancer gene therapy.展开更多
AIM:To investigate the effects of macrophage migration inhibitory factor (MIF) on proliferation of human gastric cancer MGC-803 cells and expression of cyclin D1 and p27Kip1 in them,and further determine whether the e...AIM:To investigate the effects of macrophage migration inhibitory factor (MIF) on proliferation of human gastric cancer MGC-803 cells and expression of cyclin D1 and p27Kip1 in them,and further determine whether the effects are related to the PI3K/Akt signal transduction pathway. METHODS:Gastric cancer MGC-803 cells were cultured and then treated with 50 μg/L recombinant human MIF (rhMIF) with and without a PI3K inhibitor,LY294002 (25 μmol/L). MTT assay was used to detect the prolifer-ation of MGC-803 cells. Cell cycle was detected by flow cytometry. Expression of cyclin D1 and p27Kip1 mRNA was by reverse transcription-polymerase chain reaction. Protein expression of phosphorylated Akt (p-Akt),Akt,cyclin D1 and p27Kip1 was examined by immunocyto-chemistry and Western blotting. RESULTS:rhMIF signifi cantly stimulated the prolifera-tion of MGC-803 cells and cell cycle progression from G1 phase to S phase in a concentration-and time-de-pendent manner. After the MGC-803 cells were treated with rhMIF for 24 h,the expression of cyclin D1 was signifi cantly up-regulated compared with the cells not treated with rhMIF at both mRNA and protein levels(0.97 ± 0.02 vs 0.74 ± 0.01,P = 0.002; 0.98 ± 0.05 vs 0.69 ± 0.04,P = 0.003). The p27Kip1 was down-regulated but only statistically significant at the protein level. rhMIF significantly increased the expression of p-Akt,which reached the peak at 30 min,but did not affect the expression of Akt. However,LY294002 inhibited all the effects of rhMIF.CONCLUSION:Macrophage MIF increases the proliferation of gastric cancer cells,induces the expression of cyclin D1 at the transcriptional level and inhibits the expression of p27Kip1 at the post-transcriptional level via the PI3K/Akt pathway.展开更多
文摘Objective: To study cyclinD1/bcl-1 and p27/kip1 expression in gliomas and their correlation with pathological grade and prognosis. Methods: Immunohistochemical technique was used to detect the cyclinD1/bcl-1 and p27/kip1 expression in 48 human brain glioma tissues of different malignant grades, and 12 normal non-neoplastic tissues collected from internal decompression. The data were analyzed quantitatively by the image system and also correlated retrospectively with the patients' clinical characteristics. Results: The immunohistochemical reaction for cyclinD1/bcl-1 and p27/kip1 was confined to the nuclei. The abnormal positive expression rates of both cyclinD1/bcl-1 and p27/kip1 in gliomas were found higher than that in non-neoplastic tissues(P<0.05). The number and staining intensity of cyclinD1/bcl-1 positive nuclei increased with malignant grades (P<0.05). On the contrary, the positive nuclei of p27/kip1 expression decreased in number and staining intensity with malignant grades(P<0.05). Higher expression of cyclinD1/bcl-1 or/and lower expression of p27/kip1 were associated with poor prognosis(P<0.05). Conclusion: The abnormal expression of both cyclinD1/bcl-1 and p27/kip1 might be closely related to the occurrence and development of gliomas and they might have synergistic effect. These data suggest that both cyclinD1/bcl-1 expression and p27/kip1 expression can act as an independent prognostic factor.
文摘Objective: To study p27/Kipl expression in gliomas and its application value. Methods: Imo munohistochemical technique was used to detect the exprssion of p27/Kipl gene in 48 different malignant grade human brain glioma tissues categorized according to WHO classification and 12 normal human brain tissues,which were analyzed quantitatively by using the image system and compared retrospectively with the patients' clinical characteristics. Results: In this series, the immunohistochemical reaction for p27/Kipl was confined to the nuclei. The abnormal positive expression rate of p27/Kipl in gliomas was found to be higher than that in normal tissues (P〈0.05). The positive nuclei expression of p27/Kipl decreased in number and staining intensity with the increasing degree of histological malignancy (P〈0.05). Lower expression of p27/Kipl was associated with poor prognosis (P〈0.05). P27/Kipl expression could be regarded as an independent prognostic factor. Conclusion: The abnormal expression of p27/Kipl may be closely related to the occurrence and development of gliomas, and also can be used to evaluate the prognosis independently.
基金This study was supported by grants from the National Natural Science Foundation of China (No. 30972958), Beijing Natural Science Foundation (No. 7112046), Beijing Municipal Education Commission (No. PXM2011_014226 07 000060).
文摘Background RhoA/ Rho kinase (ROCK) pathway is involved in pulmonary arterial hypertension (PAH) and pulmonary artery smooth muscle cell (PASMC) proliferation. Inhibition of ROCK has been proposed as a treatment for PAH. But the mechanism of RhoA/ROCK pathway and its downstream signaling in proliferation of human PASMCs is unclear. We investigated the effect of fasudil, a selective ROCK inhibitor, on platelet-derived growth factor (PDGF) induced human PASMC proliferation, and the possible association between RhoA/ROCK and extracellular signal-regulated kinase (ERK),p27KiP1.Methods Human PASMCs were cultured with the stimulation of 10 ng/ml PDGF, and different concentrations of fasudil were added before the addition of mitogen. Cell viability and cell cycle were determined with MTT and flow cytometry respectively. ROCK activity, ERK activity and protein expression of proliferating cell nuclear angigen (PCNA) and p27Kip1 were measured by immunoblotting.Results By MTT assay, PDGF significantly increased the OD value that represented human PASMC proliferation, and pretreatment with fasudil significantly reversed this effect in a dose-dependent manner. After PDGF stimulation, the percentage of cells in S phase increased dramatically from 15.6% to 24.3%, while the percentage in G0/G1 phase was reduced from 80.6% to 59%. And pretreatment with fasudil reversed the cell cycle effect of PDGF significantly in a dose-dependent manner. PDGF markedly induced ROCK activity and ERK activity with a peak at 15 minutes, which were significantly inhibited by fasudil. In addition, fasudil significantly inhibited PDGF-induced PCNA expression and fasudil also upregulated p27Kip1 expression in human PASMCs, which decreased after PDGF stimulation.Conclusion RhoA/ROCK is vital for PDFG-induced human PASMC proliferation, and fasudil effectively inhibited PDGF-induced human PASMC proliferation by up-regulation of p27Kip1, which may be associated with inhibition of ERK activity.
文摘AIM: To investigate p27 expression in hepatocellular carcinoma (HCC), adjacent nontumoral and normal liver tissues, and to verify whether the subcellular localization of p27 was altered in HCC.METHODS: The level of p27 in tumoral, nontumoral, and normal liver tissues were assessed by immunohistochemical (IHC) analysis. Parallel immunostaining was done for proliferating cell nuclear antigen (PCNA) to evaluate cell proliferation.RESULTS: The labeling index (LI) of p27 in tumoral lesions was significantly lower than that in adjacent nontumoral lesions (t=2.444, P=0.017) and normal controls (t=-2.268,P=0.029). The LI of p27 significantly decreased in patients with massive type (t=-2.227, P=0.037) and infiltration (t=2.197, P=0.036). The prognosis of patients with higher p27 LI was longer than that of patients with lower p27 LI (P=0.0247, log-rank test). The LI of PCNA was significantly higher in HCC than that in adjacent nontumoral lesions (t=2.092, P=0.041) and normal controls (t=-3.533, P=0.002).There was no significant correlation between p27 expression and cell proliferation in tumor samples. The level of p27 in the cytoplasmic fraction was higher in tumoral and nontumoral liver tissues, and was associated with clinical stage (t=-2.520, P=0.029) and the degree of invasion (t=2.640, P=0.019). Survival analysis showed that p27 was an independent prognosis marker for HCC patients.CONCLUSION: These results suggest that p27 underexpressing in patients with HCC is closely associated with infiltration, metastasis, and prognosis. Alterations in the subcellular localization of p27 protein may occur early during hepatocarcinogenesis.
文摘AIM: To elucidate the effect of p27^KIP1 on cell cycle and apoptosis regulation in gastric carcinoma cells. METHODS: The whole length of p27^KIP1 cDNA was transfected into human gastric cancer cell line SCG7901 by lipofectamine. Expression of p27^KIP1 protein or mRNA was analyzed by Western blot and RNA dot blotting, respectively. Effect of p27^KIP1 on cell growth was observed by MTT assay and anchorage-independent growth in soft agar. Tumorigenicity in nude mice was used to assess the in vivo biological effect of p27^KIP1. Flow cytometry, TUNEL, and electron microscopy were used to assess the effect of p27^KIP1 on cell cycle and apoptosis. RESULTS: Expression of p27^KIP1 protein or mRNA increased evidently in SCG7901 cells transfected with p27^KIP1. The cell growth was reduced by 31% at 48 h after induction with zinc determined by cell viability assay. The alteration of cell malignant phenotype was evidently indicated by the loss of anchorage-independent growth ability in soft agar. The tumorigenicity in nude mice was reduced evidently (0.55±0.14 cm vs 1.36±0.13cm, P〈0.01). p27^KIP1 overexpression caused cell arrest with 36% increase (from 33.7% to 69.3%, P〈0.01) in G1 population. Prolonged p27^KIP1 expression induced apoptotic cell death reflected by pre-G1 peak in the histogram of FACS, which was also confirmed by TUNEL assay and electron microscopy. CONCLUSION: p27^KIP1 can prolong cell cycle in G1 phase and lead to apoptosis, p27^KIP1 may be a good candidate for cancer gene therapy.
基金Supported by Grant from Hunan Provincial Science and Technology Department (2008 FJ 3088), China
文摘AIM:To investigate the effects of macrophage migration inhibitory factor (MIF) on proliferation of human gastric cancer MGC-803 cells and expression of cyclin D1 and p27Kip1 in them,and further determine whether the effects are related to the PI3K/Akt signal transduction pathway. METHODS:Gastric cancer MGC-803 cells were cultured and then treated with 50 μg/L recombinant human MIF (rhMIF) with and without a PI3K inhibitor,LY294002 (25 μmol/L). MTT assay was used to detect the prolifer-ation of MGC-803 cells. Cell cycle was detected by flow cytometry. Expression of cyclin D1 and p27Kip1 mRNA was by reverse transcription-polymerase chain reaction. Protein expression of phosphorylated Akt (p-Akt),Akt,cyclin D1 and p27Kip1 was examined by immunocyto-chemistry and Western blotting. RESULTS:rhMIF signifi cantly stimulated the prolifera-tion of MGC-803 cells and cell cycle progression from G1 phase to S phase in a concentration-and time-de-pendent manner. After the MGC-803 cells were treated with rhMIF for 24 h,the expression of cyclin D1 was signifi cantly up-regulated compared with the cells not treated with rhMIF at both mRNA and protein levels(0.97 ± 0.02 vs 0.74 ± 0.01,P = 0.002; 0.98 ± 0.05 vs 0.69 ± 0.04,P = 0.003). The p27Kip1 was down-regulated but only statistically significant at the protein level. rhMIF significantly increased the expression of p-Akt,which reached the peak at 30 min,but did not affect the expression of Akt. However,LY294002 inhibited all the effects of rhMIF.CONCLUSION:Macrophage MIF increases the proliferation of gastric cancer cells,induces the expression of cyclin D1 at the transcriptional level and inhibits the expression of p27Kip1 at the post-transcriptional level via the PI3K/Akt pathway.