Inhibition of Neointima Formation and Migration of Vascular Smooth Muscle Cells by Anti-vascular Endothelial Growth Factor Receptor-1 (Flt-1) Peptide in Diabetic Rats / 대한흉부외과학회지

BACKGROUND: Vascular endothelial growth factor (VEGF) plays an important role in angiogenesis, including stimulating the proliferation and migration of vascular smooth muscle cells (VSMCs). It has been known that diabetes is associated with accelerated cellular proliferation via VEGF, as compared to that under a normal glucose concentration. We investigated the effects of selective blockade of a VEGF receptor by using anti-Flt-1 peptide on the formation and hyperplasia of the neointima in balloon injured-carotid arteries of OLETF rats and also on the in vitro VSMCs' migration under high glucose conditions. MATERIAL AND METHOD: The balloon-injury method was employed to induce neointima formation by VEGF. For 14 days beginning 2 days before the ballon injury, placebo or vascular endothelial growth factor receptor-1 (VEGFR-1) specific peptide (anti-Flt-1 peptide), was injected at a dose of 0.5 mg/kg daily into the OLETF rats. At 14 days after balloon injury, the neointimal proliferation and vascular luminal stenosis were measured, and cellular proliferation was assessed by counting the proliferative cell nuclear antigen (PCNA) stained cells. To analyze the effect of VEGF and anti-Flt-1 peptide on the migration of VSMCs under a high glucose condition, transwell assay with a matrigel filter was performed. And finally, to determine the underlying mechanism of the effect of anti-Flt-1 peptide on the VEGF-induced VSMC migration in vitro, the expression of matrix metalloproteinase (MMP) was observed by performing reverse transcription-polymerase chain reaction (RT-PCR). RESULT: Both the neointimal area and luminal stenosis associated with neointimal proliferation were significantly decreased in the anti-Flt-1 peptide injected rats, (0.15+/-0.04 mm2 and 36.03+/-3.78% compared to 0.24+/-0.03 mm2 and 61.85+/-5.11%, respectively, in the placebo-injected rats (p<0.01, respectively). The ratio of PCNA(+) cells to the entire neointimal cells was also significantly decreased from 52.82+/-4.20% to 38.11+/-6.89% by the injected anti-Flt-1 peptide (p<0.05). On the VSMC migration assay, anti-Flt-1 peptide significantly reduced the VEGF-induced VMSC migration by about 40% (p<0.01). Consistent with the effect of anti-Flt-1 peptide on VSMC migration, it also obviously attenuated the induction of the MMP-3 and MMP-9 mRNA expressions via VEGF in the VSMCs. CONCLUSION: Anti-Flt-1 peptide inhibits the formation and hyperplasia of the neointima in a balloon-injured carotid artery model of OLETF rats. Anti-Flt-1 peptide also inhibits the VSMCs' migration and the expressions of MMP-3 and MMP-9 mRNA induced by VEGF under a high glucose condition.

Effects of Tobacco-Specific Carcinogen on Protein Kinase C Isoforms / 대한흉부외과학회지

BACKGROUND: Cigarette smoking is the leading cause of the lung cancer. However, mechanism of action underlying the carcinogenesis in the lung still remains to be elucidated. The present study attempted to look into the carcinogenic potential of tobacco-specific nitrosamine, NNK (4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone) and the effects of protein kinase C (PKC) isoforms in an immortalized human epithelial cell model. MATERIAL AND METHOD: Immortalized human epithelial cells were exposed with NNK and examined for its carcinogenic potential as measured by saturation density, soft-agar colony formation, and cell aggregation assay. The specific isoform of PKCs involved in the cellular transformation was analysed through western blot with monoclonal antibody and measured separately in cytosolic fraction and membrane fraction. RESULT: Human epithelial cells exposed with NNK showed prominent carcinogenic potential in saturation density, soft agar colony formation, and cell aggregation assay. PKC isoform analysis results are as follows: PKC-alpha showed significant translocation of protein levels from cytosolic fraction to membrane fraction, as analyzed by immunoblot. PKC-epsilon showed a dose-dependent increase of translocation. PKC-lambda was not affected by NNK treatment. CONCLUSION: The study demonstrated that there was a certain specificity in the patterns of isoform induction following chemical carcinogen exposure. Thus, it is suggested that identification of specific isoform be a clue to find target molecules in the carcinogenesis.