title: Fsh Stimulates Spermatogonial Proliferation and Differentiation in Zebrafish via Igf3.
abstract: Growth factors modulate germ line stem cell self-renewal and differentiation behavior. We investigate the effects of Igf3, a fish-specific member of the igf family. Fsh increased in a steroid-independent manner the number and mitotic index of single type A undifferentiated spermatogonia and of clones of type A differentiating spermatogonia in adult zebrafish testis. All 4 igf gene family members in zebrafish are expressed in the testis but in tissue culture only igf3 transcript levels increased in response to recombinant zebrafish Fsh. This occurred in a cAMP/protein kinase A-dependent manner, in line with the results of studies on the igf3 gene promoter. Igf3 protein was detected in Sertoli cells. Recombinant zebrafish Igf3 increased the mitotic index of type A undifferentiated and type A differentiating spermatogonia and up-regulated the expression of genes related to spermatogonial differentiation and entry into meiosis, but Igf3 did not modulate testicular androgen release. An Igf receptor inhibitor blocked these effects of Igf3. Importantly, the Igf receptor inhibitor also blocked Fsh-induced spermatogonial proliferation. We conclude that Fsh stimulated Sertoli cell production of Igf3, which promoted via Igf receptor signaling spermatogonial proliferation and differentiation and their entry into meiosis. Because previous work showed that Fsh also released spermatogonia from an inhibitory signal by down-regulating anti-Müllerian hormone and by stimulating androgen production, we can now present a model, in which Fsh orchestrates the activity of stimulatory (Igf3, androgens) and inhibitory (anti-Müllerian hormone) signals to promote spermatogenesis.
authors.affiliation: Department of Morphology (R.H.N.), Institute of Bioscience, São Paulo State University, 18618-970 Botucatu, Brazil; Reproductive Biology Group (R.H.N., R.D.V.d.S.M., D.C., P.P.d.W., R.W.S., J.B.), Division of Developmental Biology, Department of Biology, Faculty of Science, Utrecht University, 3584 CH Utrecht, The Netherlands; and Laboratory of Cellular Biology (L.R.d.F.), Department of Morphology, Institute of Biological Sciences, Federal University of Minas Gerais, 31270-901 Belo Horizonte, Brazil.
authors.forename: Rafael Henrique
authors.initials: RH
authors.lastname: Nóbrega
authors.affiliation: Department of Morphology (R.H.N.), Institute of Bioscience, São Paulo State University, 18618-970 Botucatu, Brazil; Reproductive Biology Group (R.H.N., R.D.V.d.S.M., D.C., P.P.d.W., R.W.S., J.B.), Division of Developmental Biology, Department of Biology, Faculty of Science, Utrecht University, 3584 CH Utrecht, The Netherlands; and Laboratory of Cellular Biology (L.R.d.F.), Department of Morphology, Institute of Biological Sciences, Federal University of Minas Gerais, 31270-901 Belo Horizonte, Brazil.
authors.forename: Roberto Daltro Vidal de Souza
authors.initials: RD
authors.lastname: Morais
authors.affiliation: Department of Morphology (R.H.N.), Institute of Bioscience, São Paulo State University, 18618-970 Botucatu, Brazil; Reproductive Biology Group (R.H.N., R.D.V.d.S.M., D.C., P.P.d.W., R.W.S., J.B.), Division of Developmental Biology, Department of Biology, Faculty of Science, Utrecht University, 3584 CH Utrecht, The Netherlands; and Laboratory of Cellular Biology (L.R.d.F.), Department of Morphology, Institute of Biological Sciences, Federal University of Minas Gerais, 31270-901 Belo Horizonte, Brazil.
authors.forename: Diego
authors.initials: D
authors.lastname: Crespo
authors.affiliation: Department of Morphology (R.H.N.), Institute of Bioscience, São Paulo State University, 18618-970 Botucatu, Brazil; Reproductive Biology Group (R.H.N., R.D.V.d.S.M., D.C., P.P.d.W., R.W.S., J.B.), Division of Developmental Biology, Department of Biology, Faculty of Science, Utrecht University, 3584 CH Utrecht, The Netherlands; and Laboratory of Cellular Biology (L.R.d.F.), Department of Morphology, Institute of Biological Sciences, Federal University of Minas Gerais, 31270-901 Belo Horizonte, Brazil.
authors.forename: Paul P
authors.initials: PP
authors.lastname: de Waal
authors.affiliation: Department of Morphology (R.H.N.), Institute of Bioscience, São Paulo State University, 18618-970 Botucatu, Brazil; Reproductive Biology Group (R.H.N., R.D.V.d.S.M., D.C., P.P.d.W., R.W.S., J.B.), Division of Developmental Biology, Department of Biology, Faculty of Science, Utrecht University, 3584 CH Utrecht, The Netherlands; and Laboratory of Cellular Biology (L.R.d.F.), Department of Morphology, Institute of Biological Sciences, Federal University of Minas Gerais, 31270-901 Belo Horizonte, Brazil.
authors.forename: Luiz Renato
authors.initials: LR
authors.lastname: de França
authors.affiliation: Department of Morphology (R.H.N.), Institute of Bioscience, São Paulo State University, 18618-970 Botucatu, Brazil; Reproductive Biology Group (R.H.N., R.D.V.d.S.M., D.C., P.P.d.W., R.W.S., J.B.), Division of Developmental Biology, Department of Biology, Faculty of Science, Utrecht University, 3584 CH Utrecht, The Netherlands; and Laboratory of Cellular Biology (L.R.d.F.), Department of Morphology, Institute of Biological Sciences, Federal University of Minas Gerais, 31270-901 Belo Horizonte, Brazil.
authors.forename: Rüdiger W
authors.initials: RW
authors.lastname: Schulz
authors.affiliation: Department of Morphology (R.H.N.), Institute of Bioscience, São Paulo State University, 18618-970 Botucatu, Brazil; Reproductive Biology Group (R.H.N., R.D.V.d.S.M., D.C., P.P.d.W., R.W.S., J.B.), Division of Developmental Biology, Department of Biology, Faculty of Science, Utrecht University, 3584 CH Utrecht, The Netherlands; and Laboratory of Cellular Biology (L.R.d.F.), Department of Morphology, Institute of Biological Sciences, Federal University of Minas Gerais, 31270-901 Belo Horizonte, Brazil.
authors.forename: Jan
authors.initials: J
authors.lastname: Bogerd
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D002454:Cell Differentiation / Q000187:drug effects* / Q000235:genetics; D049109:Cell Proliferation / Q000187:drug effects* / Q000235:genetics; D005640:Follicle Stimulating Hormone / Q000235:genetics / Q000494:pharmacology*; D015870:Gene Expression / Q000187:drug effects; D057809:HEK293 Cells; D006801:Humans; D008297:Male; D008540:Meiosis / Q000187:drug effects / Q000235:genetics; D018613:Microscopy, Confocal; D009154:Mutation; D011401:Promoter Regions, Genetic / Q000235:genetics; D017526:Receptor, IGF Type 1 / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012708:Sertoli Cells / Q000187:drug effects / Q000378:metabolism; D013002:Somatomedins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D013091:Spermatogenesis / Q000187:drug effects / Q000235:genetics; D013093:Spermatogonia / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D013737:Testis / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D046509:Tissue Culture Techniques; D015027:Zebrafish / Q000235:genetics / Q000378:metabolism; D029961:Zebrafish Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology
chemical_list: D013002:Somatomedins; D029961:Zebrafish Proteins; C558670:insulin-like growth factor 3, zebrafish; D005640:Follicle Stimulating Hormone; D017526:Receptor, IGF Type 1