Effects of 1,25(OH)2 vitamin D3 on cytokine production by endometrial cells of women with recurrent spontaneous abortion Maryam Tavakoli, M.D., Mahmood.

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Effects of 1,25(OH)2 vitamin D3 on cytokine production by endometrial cells of women with recurrent spontaneous abortion Maryam Tavakoli, M.D., Mahmood Jeddi-Tehrani, Ph.D., Alireza Salek-Moghaddam, Ph.D., Samira Rajaei, M.D., Afsaneh Mohammadzadeh, M.D., Shahrzad Sheikhhasani, M.D., Golnaz-Ensieh Kazemi-Sefat, M.Sc., Amir Hassan Zarnani, Ph.D. Fertility and Sterility Volume 96, Issue 3, Pages 751-757 (September 2011) DOI: 10.1016/j.fertnstert.2011.06.075 Copyright © 2011 American Society for Reproductive Medicine Terms and Conditions

Figure 1 Immunofluorescence staining of vimentin and cytokeratin in endometrial stromal cells. The endometrial stromal cells were purified from endometrial samples and their identity was determined as vimentin+ (A) and cytokeratin− (C) by immunofluorescent staining. (B, D) Negative reagent controls of vimentin and cytokeratin stainings, respectively. Magnification, ×200. Fertility and Sterility 2011 96, 751-757DOI: (10.1016/j.fertnstert.2011.06.075) Copyright © 2011 American Society for Reproductive Medicine Terms and Conditions

Figure 2 Immunocytochemical analysis of vitamin D3 receptor (VDR) and 1α-hydroxylase expression by endometrial stromal cells (ESCs). Expression of VDR and 1α-hydroxylase in purified ESCs was assessed by immunocytochemistry. The ESCs exhibited high levels of specific VDR immunoreactivity in their nucleus with some scattered pattern of reactivity in their cytoplasm (A). The 1α-hydroxylase was expressed predominantly in the cytoplasm of ESCs (C). Negative controls (B, D) were shown to be negative. Magnification, ×200. Fertility and Sterility 2011 96, 751-757DOI: (10.1016/j.fertnstert.2011.06.075) Copyright © 2011 American Society for Reproductive Medicine Terms and Conditions

Figure 3 Effect of 1,25(OH)2 vitamin D3 (1,25(OH)2D3) on cytokine production by whole endometrial cells (WECs) and endometrial stromal cells (ESCs). The WECs and ESCs from patients with unexplained recurrent spontaneous abortion (URSA) and control subjects were treated with 1,25(OH)2 vitamin D3 and the levels of interleukin (IL)-8, IL-6, transforming growth factor β (TGF-β), IL-10, and interferon γ (IFN-γ) production and the IFN-γ:IL-10 ratio were measured by capture ELISA. ∗P<.05. Fertility and Sterility 2011 96, 751-757DOI: (10.1016/j.fertnstert.2011.06.075) Copyright © 2011 American Society for Reproductive Medicine Terms and Conditions

Figure 4 The 1α-hydroxylase activity in whole endometrial cells (WECs) and endometrial stromal cells (ESCs) of patients with unexplained recurrent spontaneous abortion (URSA) and controls. The capacity of WECs and ESCs from women with URSA and controls to produce the active form of vitamin D3 were measured and compared after addition of 1,25(OH)2 vitamin D3 (1,25(OH)2D3) to the cell culture media. ∗P<.05. Fertility and Sterility 2011 96, 751-757DOI: (10.1016/j.fertnstert.2011.06.075) Copyright © 2011 American Society for Reproductive Medicine Terms and Conditions