ABSTRACT
This study aimed to develop a bovine mammary epithelial (BME) cell line model, which provides a possibility to determine functional properties of the bovine mammary gland. The primary cell culture was derived from bovine mammary gland tissues and processed enzymatically to obtain cell colonies with epithelial-like morphology. The cultures of BME cells were purified and optimally cultured at 37 °C in DMEM/F12 medium supplemented with 10% fetal bovine serum. The BME cells were identified as epithelial cell line by the evaluating the expression of keratin-18 using immunofluorescence staining. A novel gene expression system strongly enhances the expression of telomerase, has been used to immortalize BME cell line termed hTBME cell line. Interestingly, telomerase remained active even after over 60 passages of hTBME cell line, required for immortalization of BME cells. In addition, the hTBME cell line was continuously subcultured with a spontaneous epithelial-like morphology, with a great proliferation activity, and without evidence of apoptotic and necrotic effects. Further characterization showed that hTBME cell line can be continuously propagated in culture with constant chromosomal features and without tumorigenic properties. Finally, established hTBME cell line was evaluated for mammary gland specific functions. Our results demonstrated that the hTBME cell line was able to retain functional-morphological structure, and functional differentiation by expression of beta (ß)-casein as in the bovine mammary gland in vivo. Taken together, our findings suggest that the established hTBME cell line can serve as a valuable tool for the study of bovine mammary gland functions.
ABSTRACT
The formation of vertebrate skeletal muscles widely thought to be under the control of hierarchy of regulatory genes. MYF5 is one of the myogenic determination gene expressed in the developing mouse dermomyotome which control skeletal muscle differentiation. In the current work, we had obtained the cDNA sequence including the full coding region of the bos taurus myogenic factor MYF5 by reverse transcription polymerase chain reaction. Furthermore, we examined whether fibroblast cell derived from mouse and bos taurus can be transduced using plasmid vectors carrying bos taurus MYF5. Bos taurus MYF5 activates MYF5 and MYOD1 expression after 1 day culture. The concerted upregulation of the myogenic regulatory factors enhanced myosin (skeletal fast) expression. These observation show that MYF5 is essential for myogenic differentiation and provides candidates for regulation bos taurus skeletal muscle development.
