The Effects of Di-(2-ethylhexyl) Phthalate (DEHP) in Testosterone Synthesis and Its Molecular Mechanisms in the Fetal Testis of Male Mouse by Organ Culture in vitro

XU Hua, HUANG Jie, LI Min, GAO Zhi-bin, ZHU Yan-feng, LI Yun

Abstract

To investigate the effects of DEHP in testosterone synthesis and the related genes expression in the fetal testis of male mouse by organ culture in vitro. Methods The testis tissues were cultured in infiltrating type rotating device for 72 hours. The culture and gas were changed every 24 hours. The testis tissues were divided into the DMSO control group and four DEHP groups (the terminal concentration were 0.1, 1.0, 10.0, 100.0 μmol/L). The testosterone levels in the cultured medium were measured by radioimmunoassay, the INHBβ levels were measured by Elisa, the gene expressions related with testosterone synthesis were detected by real-time PCR, the morphological changes of cultured testis were observed by HE staining under optical microscope, the expressions of related proteins were measured by immunohistochemistry. Results Compared to the control group, the levels of testosterone synthesis in 0.1, 1.0, 10.0 μmol/L groups were increased, but decreased in 100.0 μmol/L group which were exposed in DEHP for 48 hours and 72 hours. There was an increase of INHBβ synthesis in 0.1 and 1.0 μmol/L groups, but a decrease in 10.0 and 100.0 μmol/L groups. The gene expressions of 3β-HSD, P450c17, P450Scc, vimentin were significantly decreased compared to that of control group especially in 1.0 and 10.0 μmol/L groups (P<0.05), but the expressions of INHBβ had no significant changes. There were no apparent morphological changes in testis tissue by HE staining. A significant increase of the three proteins (3β-HSD, P450c17, P450Scc) and a significant decrease of vimentin were observed in 10.0 and 100.0 μmol/L groups (P<0.05). Conclusion DEHP exposure can affect the testosterone synthesis in the fetal testis of male mouse. The regulation of gene expression involving in testosterone synthesis might be the mechanism.

 

Keywords: Organ culture in vitro, Di-(2-ethylhexyl) phthalate, Fetal mouse, Testis, Real-time PCR

 

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References


Wozniak SE, Gcc LL, Wachtel MS, et al. Adipose tissue; the new endocrine organ? A review article. Dig Dis Sci,2009;54 (9): 1847-1856.

. Jing W, Xiao J. Xiong Z. et al. Explant culture: an efficient method to isolate adipose-derived stromal cells for tissue engineering. Artif Organs,2011;35(2): 105-112.

Zhang Y, Li Y, Qiu F, et al. Comprehensive analysis of low- abundance proteins in human urinary exosomes using peptide ligand library technology, peptide OFFGEL fractionation and nanoHPLC-chip-MS/MS. Electrophoresis, 2010; 31 ( 23-24 ): 3797-3807.

Faulconnier Y, Thcvenct M, Flechet J, et al. Lipoprotein lipase and metabolic activities in incubated bovine adipose tissue explants: effects of insulin, dexamethasonc, and fetal bovine serum. J Anim Sci, 1994;72( 1): 184-191.

van der Valk J, Mcllor D, Brands R, et al. The humane collection of fetal bovine serum and possibilities for serum-free cell and tissue culture. Toxicol In Vitro,2004; 18( 1): 1-12.

Galic S, Oakhill JS, Steinberg GR. Adipose tissue as an endocrine organ. Mol Cell Endocrinol,2010;316(2): 129-139.

Kheterpal I, Ku G, Coleman L, et al. Proteome of human subcutaneous adipose tissue stromal vascular fraction cells versus mature adipocytes based on DIGE. J Proteome Res, 2011;10(4):1519-1527.

Permana PA, Mengc C, Reaven PD. Macrophage-secreted factors induce adipocyte inflammation and insulin resistance. Biochem Biophys Res Commun,2006?341(2) :507-514.

Zhao T, Hou M, Xia M, et al. Globular adiponectin decreases leptin-induced tumor necrosis factor-alpha expression by murine macrophages: involvement of cAMP-PKA and МАРК pathways. Cell Immunol,2005;238(1): 19-30.

Sarkanen JR, Kaila V, Mannerstrom B, et al. Human adipose tissue extract induces angiogenesis and adipogenesis in vitro. Tissue Eng Fart A,2012; 18( 1-2): 17-25.

Farmer SR. Transcriptional control of adipocyte formation. Cell Metab, 2006; 4(4): 263-273.

Zhang YY. Li X, Qian SW, et al. Transcriptional activation of histone 114 by C/EBPbeta during the mitotic clonal expansion of 3T3-L1 adipocyte differentiation. Mol Biol Cell, 2011; 22 (13):2165-2174.


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