RESUMO
Calcium (Ca2+) is an important element for many physiological functions of the uterus, including embryo implantation. Here, we investigated the possible involvement of altered intracellular Ca2+ levels in decidualization in human endometrial stromal cells (hEMSCs). hEMSCs showed high levels of mesenchymal stem cell marker expression (CD73, CD90, and CD105) and did not express markers of hematopoietic progenitor cells (CD31, CD34, CD45, and HLA-DR). Decidualization is a process of ovarian steroid-induced endometrial stromal cell proliferation and differentiation. Several types of ion channels, which are regulated by the ovarian hormones progesterone and estradiol, as well as growth factors, are important for endometrial receptivity and embryo implantation. The combined application of progesterone (1⯵M medroxyprogesterone acetate) and cyclic AMP (0.5â¯mM) for 6 days not only elevated inositol 1,4,5-triphosphate receptor (IP3R)-mediated Ca2+ release and IP3R expression, it also promoted ORAI and STIM expression as well as cyclopiazonic acid-induced Ca2+ release. Finally, intracellular Ca2+ levels and ion channel gene expression influenced hEMSC proliferation. These results suggest that cytosolic Ca2+ dynamics, mediated by specific ion channels, serve as an important step in the decidualization of hEMSCs.
Assuntos
Cálcio/metabolismo , Decídua/citologia , Decídua/metabolismo , Endométrio/citologia , Endométrio/metabolismo , Células Estromais/metabolismo , Adulto , Antígenos CD/metabolismo , Canais de Cálcio Ativados pela Liberação de Cálcio/metabolismo , Sinalização do Cálcio , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Retículo Endoplasmático/metabolismo , Feminino , Humanos , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Moléculas de Interação Estromal/metabolismoRESUMO
In this report, we introduce a standard protocol for stem cell self-renewal in vitro. Both fundamental and major procedures of stem cell manipulation, which are required for somatic cell coculture and self-renewal, are briefly described since they are important for stabilization and data normalization. In this chapter, information on the basic preparation of stem cell culture such as labware washing, equipment sanitization, microbe control, and mycoplasmosis prevention is provided. In addition, protocols for cell retrieval and preservation, proliferation assays, and basic manipulation techniques for the coculture of stem cells with somatic cells are described.