In vitro culture of ovine mammary gland cells expressing beta-lactoglobulin and beta-casein

  • Mariana Ianello Giassetti Universidade de São Paulo, Faculdade de Medicina Veterinária, Departamento de Reprodução Animal
  • Flávia Regina Oliveira de Barros Universidade de São Paulo, Faculdade de Medicina Veterinária, Departamento de Reprodução Animal The Hospital for Sick Children
  • Camilla Mota Mendes Universidade de São Paulo, Faculdade de Medicina Veterinária, Departamento de Reprodução Animal
  • Marcelo Demarchi Goissis Universidade de São Paulo, Faculdade de Medicina Veterinária, Departamento de Reprodução Animal
  • Fernanda Sevciuc Maria Universidade de São Paulo, Faculdade de Medicina Veterinária, Departamento de Reprodução Animal
  • Adriano Felipe Perez Siqueira Universidade de São Paulo, Faculdade de Medicina Veterinária, Departamento de Reprodução Animal
  • Renata Simões Universidade de São Paulo, Faculdade de Medicina Veterinária, Departamento de Reprodução Animal Universidade Federal do ABC, Centro de Ciências Naturais e Humanas
  • Mayra Elena Ortiz Avila Assumpção Universidade de São Paulo, Faculdade de Medicina Veterinária, Departamento de Reprodução Animal
  • José Antônio Visintin Universidade de São Paulo, Faculdade de Medicina Veterinária, Departamento de Reprodução Animal
Keywords: Mammary cell culture, Milk proteins, Beta-casein, Ovine

Abstract

The expression of milk proteins in vitro is essential to exploit the mammary gland cells as a biological model. Enzymatic tissue disaggregation has been widely used to establish mammary cell culture, but its effect in long-term ovine mammary cell culture is not completely elucidated. This study aimed at comparing mechanical/enzymatic and mechanical dissociation methods to establish ovine mammary cell culture. We compared cellular differentiation induced by lactating ewe sérum or fetal bovine serum based on the gene expression levels of milk proteins (beta-lactoglobulin, alpha s1-casein, and betacasein). Mechanically dissociated cells were positive immunostaining for cytokeratin 8.13, such as mammary epithelial cells. These cells are responsible for milk protein expression and they are low immunostaining for vimentin, mesenchymal marker. Mechanical/enzymatic dissociation cells were positive for vimentin. The fastest cell growth (cell/hour) was observed in the mechanical dissociation group cultured with 10% fetal bovine serum medium. Mechanically and mechanically/enzymatically derived cells were able to express beta-casein and beta-lactoglobulin, but not alpha s1-casein. The relative expression of beta-lactoglobulin was not affected by the tissue dissociation method or culture media, beta-casein relative expression was down regulated in mechanically dissociated cells cultured in the presence of lactating ewe serum, (P = 0.019). Beta-casein relative expression was also down regulated in mechanically/enzymatically dissociated cells cultured with fetal bovine sérum (P = 0.021). In the present conditions, we conclude that mechanical dissociation followed by culture with 10% of fetal bovine serum was the most efficient method to induce milk proteins’ mRNA expression by ovine mammary epithelial cells in vitro.

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Author Biography

Mariana Ianello Giassetti, Universidade de São Paulo, Faculdade de Medicina Veterinária, Departamento de Reprodução Animal
Departamento de Reprodução Animal
Published
2017-08-18
How to Cite
Giassetti, M., Barros, F. R., Mendes, C., Goissis, M., Maria, F., Siqueira, A. F., Simões, R., Assumpção, M. E., & Visintin, J. A. (2017). In vitro culture of ovine mammary gland cells expressing beta-lactoglobulin and beta-casein. Brazilian Journal of Veterinary Research and Animal Science, 54(2), 188-196. https://doi.org/10.11606/issn.1678-4456.bjvras.2017.125590
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