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  • Title: Functional analysis of recombinant single-chain Japanese eel Fsh and Lh produced in FreeStyle 293-F cell lines: Binding specificities to their receptors and differential efficacy on testicular steroidogenesis.
    Author: Suzuki H, Kazeto Y, Gen K, Ozaki Y.
    Journal: Gen Comp Endocrinol; 2020 Jan 01; 285():113241. PubMed ID: 31400434.
    Abstract:
    Pituitary gonadotropins, follicle-stimulating hormone (Fsh) and luteinizing hormone (Lh), play central roles in the control of gonadal development of vertebrates. In mammals, Fsh and Lh exclusively activate their respective cognate receptors: Fsh receptor (Fshr) in the Sertoli cell and Lh/choriogonadotropin receptor (Lhcgr) in the Leydig cell. In teleosts, the distinct functions of Fsh and Lh and information on cellular localization of their receptors are still poorly understood. Recently we established FreeStyle 293-F cell lines producing recombinant Japanese eel Fsh and Lh (reFsh and reLh), which form a single chain consisting of a common α-subunit and β-subunits. In this study, we conducted functional analyses of reFsh and reLh, focusing on the binding specificities to their receptors and effects on testicular steroidogenesis in vitro. Assays with gonadotropin receptors-expressing COS-7 cells indicated reFsh stimulated its cognate receptor, meanwhile reLh activated both receptors. Although results of in vitro incubations showed that reFsh and reLh induced testicular 11-ketotestosterone production in a dose and time-dependent manner by upregulating expression of steroidogenic enzymes, the effective doses of reLh were apparently lower and the effects of reLh emerged faster in comparison with reFsh. Results of quantitative real-time PCR using testicular cell fractions showed that fshr and lhcgr1 mRNA were detected both in Sertoli and Leydig cells. These analyses revealed that reFsh and reLh were biologically active and hence will be useful for future studies. Moreover, our data showed that both eel Fsh and Lh acted as steroidogenic hormones through their receptors in testicular somatic cells; however, Lh was more potent on androgen production, implying differential functions on spermatogenesis.
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