Expression Of The Active Factor Of Angiogenesis In The Tumors Caused By Marek’s Disease

Marek’s Disease (MD) is a malignant disease of poultry caused by Marek’s disease virus (MDV), has a higher visceral tumor incidence. In recent years, with the emergence of virulent strain, the vaccine can not play a very good protective role. Growth and metastasis of malignant tumors must rely on a rich supply of nutrients, the angiogenesis ensure that nutrients and oxygen are offered for the tumor tissue as well as laying foundation for tumor cells migration to the target organ. If angiogenesis doesn’t happen, tumor volume would be no more than1-2mm3.However angiogenesis is inextricably associated with the active factor of endothelial cells. vascular endothelial cell grown factor(VEGF) and basic fibroblast growth factor(bFGF) in combination with specific receptors located the cell surface of vascular endothelial increase vascular permeability, stimulate the proliferation and migration of vascular endothelial cells, promote angiogenesis, and eventually lead to the continuous growth and metastasis of tumor. The study is to establish the MD pathological model and then validate the role of VEGD,VEGFR2and bFGF in the MD tumor formation process and mechanism through analyzing the difference expression of MD liver, spleen, kidney tumors,as well as lay a foundation for the conforming of target for the treatment of MD tumor. The main contents of the test and results are as follows:1. Successfully established Marek’s Disease models of Hy brown chickens.According to the VEGF, VEGFR2and bFGF cDNA sequence of the Gallus logged in NCBI, we found the conserved region of the gene by analyzing software, and then using design specific primers. We successfully amplified VEGF, VEGFR2and bFGF gene sequence which were96bp,131bp,169bp respectively by PCR..By comparison with the NCBI, it is found that the PCR products we got have a high homology with the Gallus in VEGF, VEGFR2, and bFGF gene sequences. Therefore, we determine that they are Hyline Brown chicken’s VEGF, VEGFR2, bFGF gene sequences.2. We detected the mRNA expression levels of VEGF, of VEGFR2and bFGF in liver, spleen, kidney tumor of MD group and liver, spleen, kidney tissue of normal group by QRT-PCR. The results showed that:(1) VEGF, VEGFR2, bFGF gene expression in the liver tumor were1.97、5.66、2.06times than the liver tissue of the normal group (P<0.01).(2) VEGF, VEGFR2, bFGF gene expression in spleen tumor were2.60、2.67、1.84times than the spleen tissue of the normal group, virus group and the normal group (P<0.01, P<0.05).(3) VEGF, VEGFR2, bFGF gene expression in the kidney tumor were2.38、1.96、1.85times than kidney tissue in the normal group (P<0.01,P<0.05).3. The expression levels of VEGF protein was studied by Western Blot. The results showed that:the liver, spleen, kidney tumors in MD group and liver, spleen, kidney tissue in normal group had the protein bands which was sized approximately45kD that could combine with rabbit anti-VEGF polyclonal antibody. By determining the gray values of protein bands and (3-actin as internal reference for semi-quantitative analyzes, showed that (1) VEGF’s average protein content in the liver tumor(0.8962±0.0320) was more than the normal group (0.1582±0.0804)(P<0.01);(2) VEGF’s average protein content in spleen tumor (0.9020±0.0891)was more than the normal group (0.1979±0.1123)(P<0.01);(3) VEGF’s average protein content in the kidney tumor(0.5122±0.1543) was more than the normal group(0.2419±0.0932)(P<0.05).In summary, the VEGF, VEGFR2and bFGF expression levels in MD group’ liver, spleen, kidney tumors were significantly higher than normal tissue. In the MD tumor formation process, VEGF specifically combines with VEGFR2and collaborates with bFGF and its receptor promotes vascular endothelial cell proliferation, migration into the tube, while inhibiting the process of apoptosis of endothelial cells, and then supplying nutrients for the tumor cells, prompting a continuous tumor growth and metastasis, and ultimately lead to a malignant tumor.