Effects of HMG on revascularization and follicular survival in heterotopic autotransplants of mouse ovarian tissue Yanrong Wang, Qing Chang, Jing Sun,

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Effects of HMG on revascularization and follicular survival in heterotopic autotransplants of mouse ovarian tissue Yanrong Wang, Qing Chang, Jing Sun, Ling Dang, Wenzhi Ma, Changchun Hei, Xinsheng Shen, Chengjun Zhao, Yufang Cai, Xiuying Pei, Xiaoguang Zhang, Yin Wang, Xiaohua Jiang Reproductive BioMedicine Online Volume 24, Issue 6, Pages 646-653 (June 2012) DOI: 10.1016/j.rbmo.2012.02.025 Copyright © 2012 Reproductive Healthcare Ltd. Terms and Conditions

Figure 1 The flow chart of the study. Reproductive BioMedicine Online 2012 24, 646-653DOI: (10.1016/j.rbmo.2012.02.025) Copyright © 2012 Reproductive Healthcare Ltd. Terms and Conditions

Figure 2 The immunohistochemical results of the expression and integrated optical density of vascular endothelial growth factor (VEGF). Mouse ovaries were cultured in vitro for 3h with or without different concentrations of human menopausal gonadotrophin (HMG) and then stained with VEGF antibody: (A) not cultured control (CG); (B) no HMG culture control (HMG0.00); (C) cultured with HMG 0.15IU/ml (HMG0.15); (D) cultured with HMG 0.30IU/ml (HMG0.30); (E) cultured with HMG 0.60IU/ml (HMG0.60); (F) negative control. Arrows indicate theca folliculi; asterisks indicate follicles. Bar=100μm. The histogram shows VEGF expression, based on six hemi-ovaries per group. ∗ P<0.001 compared with other groups. Reproductive BioMedicine Online 2012 24, 646-653DOI: (10.1016/j.rbmo.2012.02.025) Copyright © 2012 Reproductive Healthcare Ltd. Terms and Conditions

Figure 3 Bloodstream reperfusion in grafts after avascular anastomotic transplantation of ovaries with intervention of human menopausal gonadotrophin (HMG) and immunohistochemical staining of CD34: (A) not cultured control: no ink perfusion in uncultured ovarian tissue 48h after transplantation; (B) no HMG cultured control: ink granules in the ovarian blood vessels 48h after transplantation; (C) culture with 0.15IU/ml HMG: ink granules in ovarian blood vessels 36h after transplantation; (D) culture with 0.30IU/ml HMG: numerous ink granules in the ovarian blood vessels 36h after transplantation; (E) culture with 0.60IU/ml HMG: ink granules in the ovarian blood vessels 24h after transplantation; (F) culture with 0.60IU/ml HMG: CD34 immunohistochemisty at 24h (inset is the negative control). Black triangles indicate kidneys; stars indicate ovaries; asterisks indicate follicles; arrows indicate blood vessels with ink granules; unfilled triangle indicates CD34-positive cell. Bars=50μm (A–E) and 200μm (F). Reproductive BioMedicine Online 2012 24, 646-653DOI: (10.1016/j.rbmo.2012.02.025) Copyright © 2012 Reproductive Healthcare Ltd. Terms and Conditions

Figure 4 Micrographs and measurement of tissue fluid exudation from autologous renal subcapsular grafts 36h after transplantation of ovarian tissue cultured with different concentrations of HMG before transplantation, showing that exudation increases with intervention concentration of HMG. (A) no HMG culture control (HMG0.00): almost no exudation existed around the graft; (B–D) different amounts of exudation after culture with HMG 0.15IU/ml (HMG0.15; B), HMG 0.30IU/ml (HMG0.30; C) or HMG 0.60IU/ml (HMG0.60; D). Black triangles indicate kidneys; stars indicate ovaries; asterisks indicate follicles; arrows show exudation. Bar=100μm. The histogram represents the numerical areas of tissue fluid exudation according to different HMG concentrations. Six mice in each treatment group. ∗ P<0.05 compared with other groups. Reproductive BioMedicine Online 2012 24, 646-653DOI: (10.1016/j.rbmo.2012.02.025) Copyright © 2012 Reproductive Healthcare Ltd. Terms and Conditions