Interleukin 7 and its receptor promote cell proliferation and induce lymphangiogenesis in non-small cell lung cancer / 中华病理学杂志

<p><b>OBJECTIVE</b>To study the mechanism of interleukin 7/interleukin 7 receptor (IL-7/IL-7R) in promoting cell proliferation and inducing lymphangiogenesis of non-small cell lung cancer (NSCLC) in vivo and in vitro.</p><p><b>METHODS</b>Immunohistochemical study for IL-7, IL-7R, cyclin D1 and vascular endothelial growth factor-D (VEGF-D) was carried out in NSCLC tissues from 95 patients. The relationship between IL-7/IL-7R expression and various parameters was analyzed. The mechanism of IL-7/IL-7R in promoting cell proliferation and inducing lymphangiogenesis was studied by methylthiazolyldiphenyl-tetrazolium bromide, fluorescence-activated cell sorting, reverse transcriptase-PCR, Western blot, co-immunoprecipitation, chromatin immunoprecipitation and nude mice experiments with xenograft tumors.</p><p><b>RESULTS</b>IL-7 (63.2%, 60/95), IL-7R (61.1%, 58/95), cyclin D1 (52.6%, 50/95) and VEGF-D (58.9%, 56/95) showed that high level of expression in NSCLC. IL-7/IL-7R over-expression correlated with cyclin D1 expression (P < 0.01, P < 0.01), VEGF-D expression (P < 0.01, P < 0.01), increased lymphovascular density (P = 0.005, P = 0.013), advanced clinical stage (P = 0.008, P = 0.005) and presence of lymph node metastasis (P < 0.01, P < 0.01). IL-7/IL-7R could promote proliferation of A549 cell, increase cyclin D1 and VEGF-D expression, and enhance c-Fos/c-Jun expression and phosphorylation, resulting in formation of heterodimer. Furthermore, IL-7/IL-7R could induce binding of c-Fos/c-Jun to cyclin D1/VEGF-D promoters and regulate their transcription. IL-7/IL-7R could also promote proliferation and lymphangiogenesis of lung cancer xenograft tumors.</p><p><b>CONCLUSIONS</b>IL-7/IL-7R promotes c-Fos/c-Jun expression and activity in NSCLC. This further facilitates cyclin D1 expression and accelerates proliferation of cells and VEGF-D-induced lymphovascular formation.</p>

Expression of SOCS3 and Pyk2 and their correlation in non-small cell lung cancer / 中华病理学杂志

<p><b>OBJECTIVE</b>To investigate the expression of SOCS3 and Pyk2 and their correlations in non-small cell lung cancer (NSCLC).</p><p><b>METHODS</b>The expression of SOCS3 and Pyk2 was detected in 100 cases of NSCLC, human bronchial epithelial cells (HBE) and 6 lung cancer cell lines by immunohistochemistry and immunofluorescence staining. The methylation status of SOCS3 was investigated in A549 cells by methylation-specific PCR. A549 cells were either treated with a demethylation agent 5-aza-2'-deoxycytidine (5-aza) or transfected with three SOCS3 mutants with various functional domains deleted. Besides, the cells were pretreated with a proteasome inhibitor β-lactacystin where indicated. The effects of SOCS3 on Pyk2 expression, Pyk2 Tyr 402 and ERK1/2 phosphorylations were assessed by Western blot. RT-PCR was used to estimate Pyk2 mRNA levels. Transwell experiments were performed to evaluate cell migration.</p><p><b>RESULTS</b>SOCS3 (43.0%, 43/100) and Pyk2 (65.0%, 65/100) were expressed in NSCLC. A significant negative correlation was found between SOCS3 and Pyk2 in both NSCLC tissues and cell lines. SOCS3 was aberrantly methylated and 5-aza restored SOCS3 expression. Transfection studies indicated that exogenous SOCS3 interacted with Pyk2, and both Src homology 2 (SH2) and kinase inhibitory region (KIR) domains contributed to Pyk2 binding. Furthermore, SOCS3 was found to inhibit Pyk2-associated ERK1/2 activity in A549 cells. SOCS3 possibly promoted degradation of Pyk2 in a SOCS-box-dependent manner and interfered with cell migration.</p><p><b>CONCLUSIONS</b>The data indicates that SOCS3 definitely plays roles in regulating Pyk2 signaling, and cell motility. Decreased SOCS3 induced by methylation may confer a migration advantage to A549 cells.</p>

Effects of heparanase inhibition by RNA interference on proliferation, invasiveness and apoptosis of lung cancer cells / 中华病理学杂志

<p><b>OBJECTIVE</b>To investigate the effects of heparanase expression inhibition on the proliferation, invasiveness and apoptosis of human lung adenocarcinoma cell line A549 cells.</p><p><b>METHODS</b>Recombinant eukaryotic expression plasmid pshRNA-Hpa targeting human heparanase gene was constructed. A549 cells were cultured in DMEM and transfected with pshRNA-Hpa. The expression of heparanase mRNA and protein were examined by RT-PCR and Western blot. The proliferation, invasiveness and apoptotic rates of A549 cells were determined by MTT method, matrigel invasion assays and flow cytometry respectively.</p><p><b>RESULTS</b>The expression levels of heparanase mRNA and protein were down-regulated in A549 transfected with pshRNA-Hpa. The number of cells penetrating matrigel and the proliferation ability of A549 cells transfected with pshRNA-Hpa were reduced significantly compared to the control cells. The apoptotic rate of A549 cells transfected with pshRNA-Hpa was 12.53% +/- 0.34%, being significantly higher than that of the control cells (both P < 0.01). Western-blot showed that inhibition of heparanase expression led to reduced Akt phosphorylation.</p><p><b>CONCLUSIONS</b>The recombinant plasmid pshRNA-Hpa effectively inhibited the expression of heparanase, thus suppressing the proliferation and invasion and inducing apoptosis of A549 cells. The effects may be due to the down-regulation of Akt phosphorylation level.</p>

Clinical significance of heparanase and basic fibroblast growth factor expression in human non-small cell lung cancer / 中华病理学杂志

<p><b>OBJECTIVE</b>To assess protein and mRNA expression levels of heparanase and basic fibroblast growth factor (bFGF) genes in human non-small cell lung cancer (NSCLC) and their roles in tumor invasion, metastasis and prognosis.</p><p><b>METHODS</b>A total of 115 paraffin-embedded and 45 fresh-frozen tissue specimens of NSCLC were studied by immunohistochemistry, Western Blot and in situ hybridization to evaluate the protein and mRNA expression status of heparanase and bFGF genes. The data was analyzed by SPSS statistical software.</p><p><b>RESULTS</b>Both human heparanase and bFGF were highly expressed in NSCLC cells, in contrast to none or a low expression in normal lung tissue. Expression of heparanase also showed a significantly higher than that in the normal tissue by Western blot (P = 0.041). Immunohistochemistry showed that heparanase expression was both cytoplasmic and membranous. The agreement between heparanase and bFGF was significant. A significant correlation was found between the expression of either protein and TNM stage, vascular invasion, lymphatic metastasis and microvascular density (MVD). Co-expression of the two proteins demonstrated an even higher correlation with the tumor stage and MVD. In addition, expression of bFGF correlated with tumor cell differentiation. Data of a multivariate analysis indicated that tumor cell differentiation, vascular invasion, lymphatic metastasis and expression of bFGF were identified as significant prognostic parameters.</p><p><b>CONCLUSIONS</b>Both heparanase and bFGF may play important roles in tumor angiogenesis, metastasis, and prognosis of NSCLC.</p>

Expression of p120ctn and its significance in non-small cell lung cancer / 中华病理学杂志

<p><b>OBJECTIVE</b>To investigate the expression of p120(ctn) in non-small cell lung cancer (NSCLC) and its correlation with the clinical and pathologic parameters.</p><p><b>METHODS</b>Immunohistochemistry (S-P method) was used to detect the expression of p120(ctn) in 143 NSCLC cases. The variation of protein expression was further analyzed in 36 cases by Western blot. The correlation with clinical and pathologic parameters was studied.</p><p><b>RESULTS</b>Immunohistochemically, normal bronchial cells showed membranous expression for p120(ctn), while NSCLC was characterized by cytoplasmic or diminished membranous staining. The rate of abnormal p120(ctn) expression was 79.7% (114/143). There was a significant correlation between abnormal expression of p120(ctn) and tumor differentiation, clinical stage, lymph node metastasis and poor prognosis (< 0.05), but not histologic typing. Western blot showed that the total amount of p120(ctn) in normal bronchial cells was significantly higher than that in NSCLC. The p120(ctn) isoform 1 (120,000) and isoform 3 (100,000) were expressed in normal lung tissue, while there was a reduced expression or absence of isoform 1 in NSCLC.</p><p><b>CONCLUSION</b>The expression of p120(ctn) is abnormal in NSCLC; p120(ctn) may serve as a useful prognostic marker for NSCLC.</p>

Expression of vascular endothelial growth factor (VEGF) C and VEGF receptor 3 in non-small cell lung cancer / 中华病理学杂志

<p><b>OBJECTIVE</b>To study the relationship between angiogenesis and lymphangiogenesis with the expression of vascular endothelial growth factor C (VEGF-C) and VEGFR-3 in human non-small cell lung cancer (NSCLC).</p><p><b>METHODS</b>Samples of 76 NSCLC cases with the neighboring noncancerous tissue were studied using anti- VEGF-C, VEGFR-3 and CD34 antibodies. Assessment of lymphatic vessel density and microvessel density (MVD) were performed.</p><p><b>RESULTS</b>VEGF-C expression in NSCLC was associating with the differentiation of tumor cells (P = 0.009). Expression of VEGF-C and VEGFR-3 was significantly associated with lymph node metastasis (P = 0.008 and P = 0.013 respectively) and lymphatic invasion (P = 0.027 and P = 0.020 respectively). A significant positive correlation was found between VEGF-C in cancer cells and VEGFR-3 in lymphatic endothelial cells (P = 0.009). The number of lymphatic vessels (P = 0.006) and microvascular (P = 0.046) in VEGF-C positive tumors was significantly larger than in VEGF-C-negative tumors. Lymphatic vessel density was closely related to lymph node metastasis (P = 0.010), lymphatic invasion (P = 0.019) and clinical stages (P = 0.015). MVD was closely related to blood metastasis (P < 0.001) and clinical stages (P < 0.001). Patients with positive VEGF-C expression had a worse prognosis than those with a negative VEGF-C expression (P < 0.001).</p><p><b>CONCLUSIONS</b>VEGF-C/VEGF-D in NSCLCs, are related to lymphangiogenesis and angiogenesis, as well as to the occurrence and the development of lung cancers. VEGF-C promotes intratumoral lymphangiogenesis via VEGFR-3, resulting facilitated invasion of cancer cells into the lymphatic vessels. VEGF-C expression can be a useful predictor of poor prognosis in NSCLC.</p>