Journal: Cells

Article Title: Gdnf Acts as a Germ Cell-Derived Growth Factor and Regulates the Zebrafish Germ Stem Cell Niche in Autocrine- and Paracrine-Dependent Manners

doi: 10.3390/cells11081295

Figure Lengend Snippet: A 3D model to predict the interaction between rhGDNF and zebrafish Gfrα1a. ( A ) Box 1 depicts the molecular components of the complex GDNF-GFRα1-RET. Boxes 2 and 3 show the predictive 3D model (template 4ux8.1) in which the structural similarities between zebrafish Gfrα1a and human GFRA1 are represented by orange and purple coloring and the identity of the structure formed at the binding sites is indicated in red. In box 2, green is used to indicate the conserved amino acid sequences between zebrafish Gfrα1a and human GFRA1 and blue indicates the GNDF protein. In box 3, we highlighted the interaction sites between human GDNF and zebrafish Gfrα1a/human GFRA1. ( B ) Alignment of zebrafish Gdnfa with rhGDNF. The blue lines indicate the conserved binding sites to zebrafish Gfrα1a or human GFRA1.

Article Snippet: To evaluate the effects of Gdnf on zebrafish spermatogenesis (see below), a rhGDNF purchased from PeproTech ® (London, UK) (reference no. 450-10; https://www.peprotech.com/en/recombinant-human-gdnf#productreviews )(accessed on 20 February 2020) was used.

Techniques: Binding Assay

Journal: Cells

Article Title: Gdnf Acts as a Germ Cell-Derived Growth Factor and Regulates the Zebrafish Germ Stem Cell Niche in Autocrine- and Paracrine-Dependent Manners

doi: 10.3390/cells11081295

Figure Lengend Snippet: Effects of Gdnf on germ cell composition and cellular proliferation, using a previously established primary testis tissue culture system. ( A , B ) BrdU immunodetection from zebrafish testicular explants incubated for 7 days in the absence (Basal) or presence of rhGDNF (100 ng/mL), demonstrating a higher proliferation activity for type A undifferentiated spermatogonia (A und ) and type A differentiated spermatogonia (A diff ) in the presence of rhGDNF. ( C ) Frequency of different germ cell cysts after 7 days of incubation in the absence (Basal) or presence of rhGDNF (100 ng/mL). Types A und , A diff and B spermatogonia (SPG B), spermatocytes (SPCs) and spermatids (SPTs) were identified according to morphological characteristics, as described by Leal and collaborators . ( D ) Mitotic indices of type A und and A diff spermatogonia after incubation in the absence (Basal) or presence of rhGDNF (100 ng/mL) for 7 days. ( E ) Mitotic indices of Sertoli cells in association with BrdU-negative or BrdU-positive type A und and A diff spermatogonia in the absence (Basal) or presence of rhGDNF (100 ng/mL) for 7 days. Sertoli cells were identified according to morphological characteristics, as described previously . In fish, Sertoli cells (SCs) have a triangular nuclear shape, dark chromatin and usually they appear surrounding spermatogenic cysts, as shown in . Bars represent the mean ± SEM ( n = 10). Paired t -test, * p < 0.05, ** p < 0.01, *** p < 0.001. Scale bars: 15 µm.

Article Snippet: To evaluate the effects of Gdnf on zebrafish spermatogenesis (see below), a rhGDNF purchased from PeproTech ® (London, UK) (reference no. 450-10; https://www.peprotech.com/en/recombinant-human-gdnf#productreviews )(accessed on 20 February 2020) was used.

Techniques: Immunodetection, Incubation, Activity Assay

Journal: Cells

Article Title: Gdnf Acts as a Germ Cell-Derived Growth Factor and Regulates the Zebrafish Germ Stem Cell Niche in Autocrine- and Paracrine-Dependent Manners

doi: 10.3390/cells11081295

Figure Lengend Snippet: Relative mRNA levels of genes related to Gdnf signaling ( gdnfa , gfrα1a and gfrα1b ) ( A–C ), Sertoli cell growth factors ( igf3 and amh ) ( D–E ), Fsh signaling ( fshr ) ( F ) and germ cell markers (undifferentiated spermatogonia— pou5f3 ( G ); differentiated spermatogonia and preleptotene spermatocytes— dazl ( H ); and primary spermatocytes— scyp3l ( I ) ). Testicular explants were cultivated for 7 days with rhGDNF, rzfFsh or both (rhGDNF + rzfFsh). The relative mRNA levels were normalized with the β-actin levels. Bars represent the mean ± SEM ( n = 20). One-way ANOVA followed by the Student–Newman–Keuls test, in which different letters denote significant differences ( p < 0.05) among treatment conditions.

Article Snippet: To evaluate the effects of Gdnf on zebrafish spermatogenesis (see below), a rhGDNF purchased from PeproTech ® (London, UK) (reference no. 450-10; https://www.peprotech.com/en/recombinant-human-gdnf#productreviews )(accessed on 20 February 2020) was used.

Techniques: