More expression of BDNF associates with lung squamous cell carcinoma and is critical to the proliferation and invasion of lung cancer cells.

BACKGROUND: Brain-derived neurotrophic factor (BDNF) has been reported to promote tumorigenesis and progression in several human malignancies. The purpose of this study was to explore the function of BDNF in lung squamous cell carcinoma (SCC) and adenocarcinoma (ADC). METHODS: The expression of BDNF was examined in 110 samples of lung SCC and ADC by immunohistochemistry. The protein level of BDNF was examined in 25 lung SCC or ADC samples and paired non-tumors by western blot. BDNF expression was also evaluated in human bronchial epithelial cells (HBE) and 4 lung cancer cell lines using western blot. Three BDNF mRNA variants containing exons IV, VI and IX were evaluated in HBE, two SCC (SK, LK2) and two ADC (A549, LTE) cell lines by RT-PCR. The expression and secretion of BDNF were also determined in cells using western blot and ELISA. Then the shRNA specific for BDNF was transfected into LK2 or A549 cells to further elucidate the BDNF knockdown on cell proliferation, apoptosis and invasion, which were confirmed by MTT, flow cytometry and transwell examinations. RESULTS: 71.8 % (79 out of 110) of lung SCC and ADC samples were detected positive BDNF, and high expression of BDNF was significantly correlated with histological type and T stage. Compared with non-tumorous counterparts, BDNF was apparently overexpressed in SCC and ADC tissues. In cell studies, the extensive expression and secretion of BDNF were demonstrated in lung cancer cells compared with HBE cells. Interestingly, the expressions of BDNF mRNA variant IV and VI were identical in all cells examined. However, more expression of BDNF mRNA variant IX was found in SK and LK2 cells. The apoptotic cells were increased, and the cell proliferation and invasion were both attenuated once the expression of BDNF was inhibited. When retreated by rhBDNF, BDNF knockdown cells showed less apoptotic or more proliferative and invasive. CONCLUSIONS: Our data show that BDNF probably facilitates the tumorigenesis of lung SCC and ADC. The expression of BDNF mRNA variant IX is probably more helpful to the upregulation of BDNF in SCC, and intervening the production of BDNF could be a possible strategy to lung cancer therapy.

[The study about expression of Gli2 in vein grafts].

OBJECTIVE: To study the expression of cell cycle related factor Gli2 in autogenous vein graft and its relation with neointima formation. METHOD: Autogenous vein graft model were established in 36 male wistar rats of 8 weeks old, 140 g, by transplanting the left jugular vein to intra renal abdominal aorta with microsurgical technique. Graft veins were harvest at 14, 28 days after transplantation. The IF and W-B were used to detect the protein expression in the vein graft. At the same time Gli2- mRNA was measured by RT-PCR. RESULTS: Immunofluorescent staining showed that the Gli2+ cells was only 3.2% ± 0.4% in the normal vein, but was much more in the vein graft after surgery, was 41.3% ± 0.6%, 58.3 ± 0.6% respectively; The expression of Gli2 and PCNA were both elevated in the vein graft. There is a positive correlation between them which indicated by W-B, the relation index was 0.826; the Gli2 mRNA content was also increased in vein graft, was 8.9, 13.6 fold compared with normal vein as 1 respectively. CONCLUSION: Gli2 is upregulated in autogenous vein grafts and may correlated with the proliferation of vascular smooth muscle cells.

[The expression of sonic hedgehog in rat vein grafts].

OBJECTIVE: To study the expression of cell cycle related factor sonic hedgehog (SHH) in autogenous vein graft and its relation with neointima formation. METHODS: Autogenous vein graft model were established in 24 male Wistar rats of 8 weeks old and 140 g weight, by transplanting the left jugular vein to intra renal abdominal aorta with microsurgical technique. Graft veins were harvested at 14 d and 28 d after transplantation. The immunohistochemistry and Western blot were used to detect the SHH and PCNA expression in the vein graft. At the same time SHH mRNA was measured by quantitative real-time PCR. The opposite normal veins served as control. RESULTS: Histological staining showed that the percent of SHH+ cells was only (2.0 +/- 0.5)% in the normal vein, but was much more in the vein graft after surgery, as (39.4 +/- 0.4)% and (63.0 +/- 0.3)% respectively (P < 0.01). The expression of SHH and PCNA were both elevated in the vein graft. There was a positive correlation between them which indicated by Western blot (r = 0.808, P < 0.01). The SHH mRNA content also increased in vein graft to 9.5 and 23.8 folds of that in control. CONCLUSION: SHH is upregulated in autogenous vein grafts and may correlated with the proliferation of vascular smooth muscle cells.

[The sonic hedgehog induce vascular adventitial fibroblasts phenotypic modulation, proliferation and migration].

OBJECTIVE: To study the effect of shh on the migration, proliferation and phenotypic modulation of vascular adventitial fibroblasts. METHOD: Cultivate the vascular adventitial fibroblast in vitro. Use immunofluorescent, laser confocal microscopy, Western-blot and real-time PCR to detect the expression of mRNA and protein of related index. Estimate cell proliferation according to the expression of Ki67 and cell proliferation curve. Application of wound healing test to estimate migration of fibroblast. The expression of alpha-actin is thought to be marker of phenotypic modulation of fibroblast. RESULT: The expression of shh was detected in vascular adventitial fibroblast in vitro. After addition of exogenous shh (3.5 microg/ml), there were more Ki67(+) cells and the wounding area which was covered by cells became larger. The expression of alpha-actin was detected. After addition of cyclopamine (40 micromol/L), there were less Ki67(+) cells and the wounding area which was covered by cells became smaller. CONCLUSION: Shh can promote proliferation, migration and phenotypic modulation of vascular adventitial fibroblasts.

Expression of T cell factor-4 in non-small-cell lung cancer.

BACKGROUND: T cell factor-4 (TCF-4) plays an important role in development and carcinogenesis. Recently, the role of TCF-4 has been described in colon cancer and other cancers. However, whether TCF-4 plays a similar role in lung cancer is unknown. To answer this question, we studied the expression of TCF-4 protein and mRNA in non-small-cell lung cancer (NSCLC) and the relation of TCF-4 expression pattern to histological type and cell differentiation. METHODS: Tissue samples from sixty cases of pathologically diagnosed NSCLC and eight normal tissue samples were obtained between September 2001 and March 2003. Immunohistochemistry was used to investigate the distribution of TCF-4 protein. The staining patterns of the tumors were divided into 4 categories: nuclear staining alone or nuclear staining greater than cytoplasmic staining; cytoplasmic staining or cytoplasmic staining greater than nuclear staining; equal nuclear and cytoplasmic staining; no nuclear staining or cytoplasmic staining. The integrated optical density (OD) values of all sections were analyzed by UIC MetaMorph image analysis software. The expression of TCF-4 mRNA was detected by one-step reverse transcription-polymerase chain reaction (RT-PCR). The integrated density values of the PCR products were analyzed semi-quantitatively. RESULTS: Immunohistochemistry showed that there was no expression of TCF-4 in normal tissue. However, TCF-4 was expressed in 86.7% (52/60) of NSCLC samples, mainly in the nuclei of tumor cells. Furthermore, there was a significant difference in TCF-4 localization patterns between squamous cell carcinomas and adenocarcinomas (P < 0.05). The integrated OD values of TCF-4 expression was significantly higher in tumors with moderate-poor cell differentiation than in well differentiated tumors (51.63 +/- 6.67 vs 46.13 +/- 12.31, P < 0.01). There was no TCF-4 mRNA expression in normal tissue. However, 63.9% (23/36) of carcinoma samples expressed TCF-4 mRNA. TCF-4 mRNA expression was significantly higher in tumors with moderate-poor cell differentiation than in well differentiated tumors (P < 0.05). There were no significant differences in mRNA expression in comparison with histological type. CONCLUSIONS: The sub-cellular distribution of TCF-4 may correlate with NSCLC histological type. High expression of TCF-4 mRNA and protein may be associated with the degree of cell differentiation in NSCLC.

[Association of genetic polymorphism in the DNA repair gene XRCC1 with susceptibility to lung cancer in non-smoking women].

OBJECTIVE: To assess the relationship between the XRCC1 polymorphism and susceptibility to lung cancer in non-smoking female on the basis of a hospital-based case-control study. METHODS: Genotypes were determined by PCR-restriction fragment length polymorphism in 50 patients with lung cancer and 50 controls. The adjusted odds ratios (OR) and 95% confidence intervals (CI) were calculated using logistic regression model to study the relationship between different genotypes and risk of lung cancer in non-smoking women. Furthermore, a multiplicative interaction between exposure to cooking oil smoke and the variant XRCC1 399Gln allele on risk of lung adenocarcinoma was evaluated. RESULTS: Individuals carrying Gln/Gln genotype were at an increased risk to suffer from lung adenocarcinoma as compared with those with the Arg/Arg genotype (OR: 14.12; 95% CI: 2.14 approximately 92.95, adjusted for age and cooking oil smoke). The OR of lung adenocarcinoma for the variant XRCC1 399Gln allele with exposure to cooking oil smoke was 6.29 (95% CI 1.99 approximately 19.85). CONCLUSION: The above described findings indicate that Arg 399Gln polymorphism in the XRCC1 is associated with risk of lung adenocarcinoma but not with risk of squamous-cell carcinoma of the lung in non-smoking women.

Effect of cyclin G2 on proliferative ability of SGC-7901 cell.

AIM: To study the effect of cyclin G2 on proliferation of gastric adenocarcinoma cell line-SGC-7901 cell in vitro. METHODS: By use of cation lipofectamine transfection reagent, the pIRES-G2 and pIRESneo plasmids were transferred into SGC-7901cell line. Anticlones were selected by G418. Positive clones were observed and counted using Giemsa staining. Cell proliferative ability was assayed by MTT. RESULTS: (1) The clone number of pIRES-G2 group decreased, clone volume reduced. The number of cell clones in pIRESneo group was 87+/-3, that of pIRES-G2 group was 53+/-4, occupying 60.1% of pIRESneo group, there was significant difference obviously (P<0.01, t=15.45). (2) The average absorbance of clone cell obtained by stable transfection of pIRES-G2 at 570 nm was 1.6966+/-0.2125, the average absorbance of clone cell obtained by stable transfection of pIRESneo at 570 nm was 2.1182+/-0.3675, there was significant difference between them (P<0.01, t=3.412). CONCLUSION: Cyclin G2 can inhibit SGC-7901cell proliferative ability obviously, it may be a negative regulator in cell cycle regulation.