Objective To be expressed human vascular endothelial growth factor (VEGF) recombinant protein in Escherichia Coli in high level. Methods VEGF was amplified from human fetal brain cDNA library, the amplified fragment was inserted into M13mP18 and confirmed to be VEGF165cDNA by restriction mapping and DNA sequencing, then it was combined with an expression vector PRL621. This recombinant plasmid overexpressed a 20kd recombinant protein in E.Coli(TG1), the protein was isolated and purifed from E.Coli, and initially renatured. Results The overexpressed recombinant protein was 35% of the total cell protein, the sequence of its first 15-N terminal amino acid was identrical to that of the human natural VEGF protein, Chorioallantoic membrane(CAM) assay showed that the rhVEGF promated new capillary vessels formation. Conclusion The genetic engineering Escherichia Coli can express human vascular endothelial growth factor in high level.
Objective To investigate clinical significance of serum VEGF-C level and C-erbB-2 protein expression in patients with breast cancer. Methods Sixty-two female patients with breast invasive ductal cancer and breast benign lesion were respectively selected. Serum VEGF-C level was detected by enzyme-linked immunosorbent assay (ELISA) before operation and at one month after operation, and C-erbB-2 protein expression in tissues of breast cancer was detected by immunohistochemistry. Then, the relationship between serum VEGF-C level and clinicopathologic characteristics and C-erbB-2 protein expressions wereas analyzed. Results The serum VEGF-C level before operation in breast cancer patients〔(279.65±17.34) pg/ml〕 was significantly higher than that in breast benign lesions patients 〔(167.26±12.15) pg/ml〕, P＜0.01. In breast cancer patients, the serum VEGF-C level before operation was higher than that at one month after operation 〔(209.45±15.23) pg/ml〕, P＜0.01. The serum VEGF level was related to tumor stage (P＜0.05) but not to patient age, tumor size, menopause status , lymph node metastasis or not and ER and PR expression (Pgt;0.05). The positive expression rate of C-erbB-2 protein in breast cancer patients (54.84%, 34/62) was significantly higher than that in breast benign lesion patients (11.29%, 7/62), P＜0.01. Moreover, the positive expression rate of C-erbB-2 protein in breast cancer patients with axilla lymph node metastasis (69.44%) was significantly higher than that without axilla lymph node metastases (34.62%), P＜0.05. The serum VEGF level increased with increasing expression intensity of C-erbB-2 protein and there was positive correlation between them (r=0.813，P＜0.05). Conclusions The serum VEGF-C level in breast cancer may be conducted as an assisted marker to differential diagnosis of breast tumor. C-erbB-2 is related to lymph node metastasis of breast cancer patients. There is synergistic effect between VEGF-C and C-erbB-2 in the lymph node metastasis way of breast cancer.