AIM: To evaluate of Cx26 in correlation with Bcl-xL and Bax proteins in colorectal cancer. METHODS: Immunohistochemical staining using specific antibodies was performed to evaluate the protein expression of Cx26, Bax ...AIM: To evaluate of Cx26 in correlation with Bcl-xL and Bax proteins in colorectal cancer. METHODS: Immunohistochemical staining using specific antibodies was performed to evaluate the protein expression of Cx26, Bax and Bcl-xL in 152 colorectal cancer samples and the correlations among studied proteins as well as the relationships between the expression of Cx26, Bax, Bcl-xL and clinicopathological features were analyzed. RESULTS: Both normal epithelial cells and carcinoma cells expressed Cx26, Bax and Bcl-xL, but Cx26 in cancer cells showed aberrant, mainly cytoplasmic staining. Expression of Cx26, Bax and Bcl-xL was observed in 55.9%, 55.5% and 72.4% of evaluated colorectal cancers respectively. We found the positive correlation between Cx26 and Bax expression (r= 0.561, P<0.0001), Cx26 and Bcl-xL (P=0.409, P<0.0001) as well as between Bax and Bcl-xL (P=0.486, P<0.0001). Association of Cx26, Bax and Bcl-xL expression with histological G2 grade of tumors was noted (P<0.005, P<0.001 and P<0.002 respectively). CONCLUSION: Cytoplasmic presence of Cx26 and its association with apoptotic markers could indicate a distinct role from physiological functions of Cx26 in cancer cells and it could suggest that connexins might be a target point for modulations of apoptosis with therapeutic implications.展开更多
AIM: To determine the inhibitory effect of the vectorgenerated small interfering RNAs (siRNAs) on the expression of the BcI-XL gene in established human esophageal cancer cells, and to investigate the effect of the...AIM: To determine the inhibitory effect of the vectorgenerated small interfering RNAs (siRNAs) on the expression of the BcI-XL gene in established human esophageal cancer cells, and to investigate the effect of the BcI-XL siRNAs on cell growth and apoptosis in esophageal cancer cells. METHODS: Three siRNA-expressing vectors targeting different sites of the Bcl-XL gene were constructed from pTZ-U6+I vector. Cultured esophageal cancer cells were transfected with the siRNA-expressing vector (or the control vector) using lipofectamine 2000. BcI-XL gene expression was determined with semiquantitative RT- PCR assay and Western blotting. Among the three siRNA- expressing vectors, the most highly functional vector and its effect on cell growth and apoptosis in esophageal cancer ceils was further analyzed. RESULTS: Of the three siRNA-expressing vectors, siRNA- expressing vector No.1 was the most potent one which suppressed Bcl-XL mRNA production to 32.5% of that in the untreated esophageal cancer cells. Western blotting analysis showed that siRNA-expressing vector No.1 markedly down-regulated the expression of Bcl-XL in human esophageal cancer cells. Treatment of esophageal cancer cells with siRNA-expressing vector No.1 resulted in inhibition of cell growth and induction of apoptosis. CONCLUSION: Down-regulation of BcI-XL by vectorgenerated small interfering RNAs can suppress cell growth and induce apoptosis in human esophageal cancer cells.展开更多
AIM: We optimized a rapid and efficient tissue lysis method using the MagNA Lyser (Roche, Germany). Using this novel method combined with immunoblot analysis, we investigated the correlation between abnormal Bcl-XL...AIM: We optimized a rapid and efficient tissue lysis method using the MagNA Lyser (Roche, Germany). Using this novel method combined with immunoblot analysis, we investigated the correlation between abnormal Bcl-XL expression and clinicopathological characteristics in colorectal cancer. METHODS: Tissue samples from Sprague-Dawley rats were tested to determine optimal lysis conditions for use with MagNA Lyser. We next used the new method to extract tissue proteins from the tumor tissue of a colorectal cancer patient. The availability of extractable tissue proteins for proteomic study was demonstrated by two-dimensional (2D) gel electrophoresis and subsequent matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. In addition, we prepared tissue lysates from paired tumor tissues and adjacent nontumor tissues of 50 colorectal carcinoma patients. Ensuing immunoblot analyses were performed to detect the level of Bcl-X, expression. RESULTS: The optimal sample sizes processed were found to be around 200 mg, with oscillation frequency of 6 500 r/rain for 80 s. Test of the first human tissue lysate confirmed that the MagNA Lyser method was adequate for protein extraction and subsequent identification by current proteomic protocols. The method was also applicable to immunoblot analysis. Thirty of 50 (60%) colorectal patients exhibited higher level of Bcl-XL expression in their tumor tissues. Raised level of Bcl-XL expression correlated with patients' gender and tumor cell proliferation index (P= 0.037 and P〈0.001, respectively), but was independent of clinicopathological characteristics and overall survival. CONCLUSION: We report a novel tissue lysis method applicable to proteomic and immunoblot analyses, which can facilitate the discovery and detection of cancer protein alterations.展开更多
基金Supported by the Polish State Committee for Scientific Research (3 PO5B 07922)
文摘AIM: To evaluate of Cx26 in correlation with Bcl-xL and Bax proteins in colorectal cancer. METHODS: Immunohistochemical staining using specific antibodies was performed to evaluate the protein expression of Cx26, Bax and Bcl-xL in 152 colorectal cancer samples and the correlations among studied proteins as well as the relationships between the expression of Cx26, Bax, Bcl-xL and clinicopathological features were analyzed. RESULTS: Both normal epithelial cells and carcinoma cells expressed Cx26, Bax and Bcl-xL, but Cx26 in cancer cells showed aberrant, mainly cytoplasmic staining. Expression of Cx26, Bax and Bcl-xL was observed in 55.9%, 55.5% and 72.4% of evaluated colorectal cancers respectively. We found the positive correlation between Cx26 and Bax expression (r= 0.561, P<0.0001), Cx26 and Bcl-xL (P=0.409, P<0.0001) as well as between Bax and Bcl-xL (P=0.486, P<0.0001). Association of Cx26, Bax and Bcl-xL expression with histological G2 grade of tumors was noted (P<0.005, P<0.001 and P<0.002 respectively). CONCLUSION: Cytoplasmic presence of Cx26 and its association with apoptotic markers could indicate a distinct role from physiological functions of Cx26 in cancer cells and it could suggest that connexins might be a target point for modulations of apoptosis with therapeutic implications.
基金Supported by Science and Technology Fund of SichuanProvince, No. 2003A067
文摘AIM: To determine the inhibitory effect of the vectorgenerated small interfering RNAs (siRNAs) on the expression of the BcI-XL gene in established human esophageal cancer cells, and to investigate the effect of the BcI-XL siRNAs on cell growth and apoptosis in esophageal cancer cells. METHODS: Three siRNA-expressing vectors targeting different sites of the Bcl-XL gene were constructed from pTZ-U6+I vector. Cultured esophageal cancer cells were transfected with the siRNA-expressing vector (or the control vector) using lipofectamine 2000. BcI-XL gene expression was determined with semiquantitative RT- PCR assay and Western blotting. Among the three siRNA- expressing vectors, the most highly functional vector and its effect on cell growth and apoptosis in esophageal cancer ceils was further analyzed. RESULTS: Of the three siRNA-expressing vectors, siRNA- expressing vector No.1 was the most potent one which suppressed Bcl-XL mRNA production to 32.5% of that in the untreated esophageal cancer cells. Western blotting analysis showed that siRNA-expressing vector No.1 markedly down-regulated the expression of Bcl-XL in human esophageal cancer cells. Treatment of esophageal cancer cells with siRNA-expressing vector No.1 resulted in inhibition of cell growth and induction of apoptosis. CONCLUSION: Down-regulation of BcI-XL by vectorgenerated small interfering RNAs can suppress cell growth and induce apoptosis in human esophageal cancer cells.
文摘AIM: We optimized a rapid and efficient tissue lysis method using the MagNA Lyser (Roche, Germany). Using this novel method combined with immunoblot analysis, we investigated the correlation between abnormal Bcl-XL expression and clinicopathological characteristics in colorectal cancer. METHODS: Tissue samples from Sprague-Dawley rats were tested to determine optimal lysis conditions for use with MagNA Lyser. We next used the new method to extract tissue proteins from the tumor tissue of a colorectal cancer patient. The availability of extractable tissue proteins for proteomic study was demonstrated by two-dimensional (2D) gel electrophoresis and subsequent matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. In addition, we prepared tissue lysates from paired tumor tissues and adjacent nontumor tissues of 50 colorectal carcinoma patients. Ensuing immunoblot analyses were performed to detect the level of Bcl-X, expression. RESULTS: The optimal sample sizes processed were found to be around 200 mg, with oscillation frequency of 6 500 r/rain for 80 s. Test of the first human tissue lysate confirmed that the MagNA Lyser method was adequate for protein extraction and subsequent identification by current proteomic protocols. The method was also applicable to immunoblot analysis. Thirty of 50 (60%) colorectal patients exhibited higher level of Bcl-XL expression in their tumor tissues. Raised level of Bcl-XL expression correlated with patients' gender and tumor cell proliferation index (P= 0.037 and P〈0.001, respectively), but was independent of clinicopathological characteristics and overall survival. CONCLUSION: We report a novel tissue lysis method applicable to proteomic and immunoblot analyses, which can facilitate the discovery and detection of cancer protein alterations.