An investigation of the short-medium-long term effects of LOW molecular weight heparins and growth factors on angiogenesis BY chick chorioallantoic membrane method

Angiogenesis is process that occurs new capillaries from pre-existings. During angiogenesis vascular endothelial growth factor (VEGF) and epidermal growth factor (EGF) are responsible of activation, migration and proliferation of endothelial and epidermal cells respectively. Low molecular weight heparins (LMWHs) influence angiogenesis growth factor by regulating basic fibroblaste growth factor and synthesise of its inhibitors. Chick chorioallantoic membrane (CAM) is extraembryonic membrane that consist of embryonic chorion and allantoic. Chorioallantoic membrane occurs during forth day of fertilization. It is a common method in angiogenesis and antiangiogenesis research. This study claims to investigate antiangiogenic or angiogenic effects of LMWHs tinzaparin, enoxaparin and bemiparin by CAM method. This study consits of six groups and seven fertilized eggs for each one. We used agarose as control group, bevacizumab as VEGF inhibitor, cetuximab as EGF inhibitor and bemiparin, enoxaparin and tinzaparin as LMWHs. This comparison was made according to vessel lenght and junction changes at immediately after topical application in a time period of (0 hour) short term (12 hours) medium term (24 hours) and long term (36 hours). Calculation of vessel lenght and junctions carried out by automatic measurement with Angioquant software and manual measurement with ImageJ software. After 12 hours of drug application, vessel length changes were statistically significant in enoxaparin compare to agarose, bemiparin and cetuximab (p=0.029, p=0.035, p=0.047 respectively). After 36 hours of drug application, vessel length changes were statistically significant in bevacuzimab, cetuximab and enoxaparin compare to control group (p=0.015, p=0.035, p=0.002 respectively). In 12th and 36th hours, vessel junction changes were statistically significant in enoxaparin compare to control (p=0.025, p=0.04 respectively). The results of this study showed that LMWHs may have antiangiogenic effects and we concluded that antiangiogenic effects of LMWHs do not depends on molecular weight. In addition, the most effective time for study of antiangiogenic effect is 36 hours after drug application.