The Investigation of Effects of Medicinal Leech Saliva Extract on the Breast Fibroblast Cell Line In Vitro: An Experimental Study

Abstract

Medicinal leeches benefit by delivering various bioactive substances from their saliva during sucking. It was aimed to investigate the effects of leech saliva extract (LSE) on breast fibroblast cell line (Hs 578Bst) and HUVEC (CRL-1730) line in vitro. Material and Methods: Leech saliva was obtained from medicinal leeches of the Hirudo verbana species. The LSE was obtained from the Hirudo verbana. LSE dose amounts (400 µg/mL, 200 µg/mL, 100 µg/mL and 50 µg/mL) were adjusted for protein concentration. In cell culture; cell viability, apoptosis, cell migration and gene expressions responsible for these effects were evaluated. The effect of LSE on the cell viability was evaluated with MTT assay. The Scratching Test was used to evaluate the effects of LSE on the cell migration. The effect of LSE on mRNA expression levels of fibroblast growth factor (FGF), vascular endothelial growth factor (VEGF) and epidermal growth factor (EGF) was determined with polymerase chain reaction. The effect of LSE on apoptosis was investigated with flow cytometry. Results: The doses of LSE (400-50 µg/mL) significantly increased the cell viability compared with the control group for 24 h and 48 h (p<0.05). In the scratching test, it was observed that the 50 µg/mL dose induced the cell migration and covered all areas with cells at 24 h. It was observed that 200 µg/mL LSE up-regulated the mRNA expression of FGF, VEGF and EGF compared with the control and other groups (p<0.001). All doses of LSE shows no apoptotic or necrotic effects in the cell lines. Conclusion: In this study, it was found that the LSE applied at different doses had significant effects on the cell viability, cell migration and gene expression responsible for these effects in the breast fibroblast cell line.