Expression and functions of cluster of differentiation 9 and 81 in rat mammary epithelial cells

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  • HORIGUCHI Kotaro
    Laboratory of Anatomy and Cell Biology, Department of Health Sciences, Kyorin University, Tokyo 181-8612, Japan
  • YOSHIDA Saishu
    Department of Biochemistry, The Jikei University School of Medicine, Tokyo 105-8461, Japan
  • TSUKADA Takehiro
    Department of Biomolecular Science, Faculty of Science, Toho University, Chiba 274-8510, Japan
  • NAKAKURA Takashi
    Department of Anatomy, Graduate School of Medicine, Teikyo University, Tokyo 173-8605, Japan
  • FUJIWARA Ken
    Department of Biological Science, Kanagwa University, Kanagawa 259-1293, Japan
  • HASEGAWA Rumi
    Laboratory of Anatomy and Cell Biology, Department of Health Sciences, Kyorin University, Tokyo 181-8612, Japan
  • TAKIGAMI Shu
    Laboratory of Anatomy and Cell Biology, Department of Health Sciences, Kyorin University, Tokyo 181-8612, Japan
  • OHSAKO Shunji
    Laboratory of Anatomy and Cell Biology, Department of Health Sciences, Kyorin University, Tokyo 181-8612, Japan

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Abstract

<p> Cluster of differentiation (CD) 9 and CD81 are closely-related members of the tetraspanin family that consist of four-transmembrane domain proteins. Cd9 and Cd81 are highly expressed in breast cancer cells; however, their expression in healthy mammary glands is unclear. In this study, we performed quantitative real-time PCR to analyze the expression levels of Cd9 and Cd81. Histological techniques were employed to identify Cd9- and Cd81-expressing cells in rat mammary glands during pregnancy and lactation. It was observed that Cd9 and Cd81 were expressed in the mammary glands, and their expression levels correlated with mammary gland development. To identify cells expressing Cd9 and Cd81 in the mammary glands, we performed double immunohistochemical staining for CD9 and CD81, prolactin receptor long form, estrogen receptor alpha, or Ki67. The results showed that CD9 and CD81 were co-expressed in proliferating mammary epithelial cells. Next, we attempted to isolate CD9-positive epithelial cells from the mammary gland using pluriBead cell-separation technology based on antibody-mediated binding of cells to beads of different sizes, followed by isolation using sieves with different mesh sizes. We successfully isolated CD9-positive epithelial cells with 96.8% purity. In addition, we observed that small-interfering RNAs against Cd9 and Cd81 inhibited estrogen-induced proliferation of CD9-positive mammary epithelial cells. Our current findings may provide novel insights into the proliferation of mammary epithelial cells during pregnancy and lactation as well as in pathological processes associated with breast cancer.</p>

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