title: Purification and identification of 25-hydroxycholesterol in a reptile: Seasonal variation and hormonal regulation.
abstract: The present in vitro study, for the first time, demonstrates the production of 25-hydroxycholestrol (25-HC) by testicular macrophages of a non-mammalian vertebrate. The ether extracts of testicular macrophage-conditioned medium (TMCM) were fractionated on a C18 reversed phase high-performance liquid chromatography (HPLC) column using methanol as the mobile phase. The mass spectrometry (MS) fragmentation pattern of HPLC-purified 25-HC was found to be identical to that of authentic 25-HC. Further, a significant seasonal variation in 25-HC concentration was observed with maximal level in regressed and minimal during breeding phase. To understand the hormonal control of 25-HC production, testicular macrophages from regressed phase testes were incubated with 0.5μg/ml of ovine follicle stimulating hormone (FSH) and 0.1, 1 and 10μg/ml of testosterone (T). FSH considerably enhanced 25-HC production by testicular macrophages. In contrast, T markedly inhibited 25-HC production in a dose-dependent manner. In addition, T significantly inhibited FSH-induced 25-HC production, though pre-treatment with T was more effective as compared to post-treatment with T to FSH. Our findings on production, seasonal variation and hormonal control of 25-HC suggest the functional significance of 25-HC in the testis of reptiles.
authors.affiliation: Department of Zoology, Kalindi College, University of Delhi, New Delhi 110008, India.
authors.forename: Varsha
authors.initials: V
authors.lastname: Singh
authors.affiliation: Comparative Immuno-endocrinology Laboratory, Department of Zoology, University of Delhi, North Campus, Chhatra Marg, Delhi 110007, India.
authors.forename: Manisha
authors.initials: M
authors.lastname: Priyam
authors.affiliation: Comparative Immuno-endocrinology Laboratory, Department of Zoology, University of Delhi, North Campus, Chhatra Marg, Delhi 110007, India.
authors.forename: Mamta
authors.initials: M
authors.lastname: Tripathy
authors.affiliation: Comparative Immuno-endocrinology Laboratory, Department of Zoology, University of Delhi, North Campus, Chhatra Marg, Delhi 110007, India. Electronic address: rai_u@rediff.mail.com.
authors.forename: Umesh
authors.initials: U
authors.lastname: Rai
mesh_terms: D000818:Animals; D002851:Chromatography, High Pressure Liquid; D005640:Follicle Stimulating Hormone / Q000494:pharmacology; D006728:Hormones / Q000494:pharmacology*; D006888:Hydroxycholesterols / Q000737:chemistry / Q000302:isolation & purification*; D008116:Lizards / Q000378:metabolism*; D008297:Male; D013058:Mass Spectrometry; D012098:Reproduction / Q000187:drug effects; D012621:Seasons; D012756:Sheep; D013737:Testis / Q000187:drug effects / Q000378:metabolism; D013739:Testosterone / Q000494:pharmacology
chemical_list: D006728:Hormones; D006888:Hydroxycholesterols; D013739:Testosterone; C007997:25-hydroxycholesterol; D005640:Follicle Stimulating Hormone
keywords: 25-Hydroxycholesterol; Follicle stimulating hormone; Testicular macrophage; Testosterone