Şap Virüsünün Hücreye Girişinde Sindekan 1’in Rolünün Araştırılması

Abstract

Foot and mouth disease is an acute and highly contagious viral disease of hoofed that causes epidemics and significant economic losses in many countries, including Turkey. There are 7 serotypes and many variants of the foot-and-mouth disease virus (FMDV) , which is the causative agent of the disease. Vaccination remains the most effective solution in the fight against FMDV. Viruses initiate the infection process by binding to specific receptors located on the cell surfaces of susceptible host cells. Field isolates of the FMDV use integrin αvβ1, αvβ3, αvβ6, and αvβ8 receptors for entry into the host cell. It has been observed that heparan sulfate proteoglycans have a role in binding to the host cell surface in many viruses, including the foot-and-mouth disease virus, but the information on which of this family plays a role for the FMDV has not been found in the literature. In order to contribute to this deficiency in the literature, the role of Syndecan 1 (SDC1), which is in the heparan sulfate proteoglycan family, was investigated in this thesis study. In this thesis, it is shown that human colon carcinoma cell (SW480) which is not susceptible to FMDV becomes susceptible to FMDV after transfection and expression of the plasmid containing the human syndecan 1 (SDC1) gene. Integrin αvβ6, which is known to be a receptor for the FMDV, was used as a positive control group. OTUR/17 virus strain adapted in susceptible baby hamster kidney cell (BHK-21) was used because heparan sulfate prefers the O serotype during the adaptation to cell culture. In the first phase of the studies, sterility tests of cells were performed, growth curves were drawn, cell stocks were prepared and the susceptibility of SW480 and BHK-21 cells to FMDV was checked comparatively. In the second step, SDC1, integrin β6 and control plasmids were individually transformed into competent bacteria. Transformed bacteria were propagated, plasmid DNAs were re-isolated and stored by measuring their concentrations. In the third step, the transfection application dose was determined, transfection procedures were performed, and RNA isolations were made and gene expressions were measured by polymerase chain reaction (PCR). At the last stage of the study, the susceptibility of the transfected cells was examined by inoculating the virus. As a result of transfection processes, an increase in gene expressions was observed at the expected level. As a consequence of the incubation of the transfected cells with the FMDV, cell shedding was observed as a result of a gradual cytopathic effect in the wells transfected with β6 and SDC1, more so in the wells transfected with the β6 gene.