A83-01 and DMH1 effects in the zebrafish spermatogonial niche: Unraveling the roles of TGF-ß and BMP signaling in the Fsh-mediated spermatogonial fate.

Transforming growth factor-ß (TGF-ß) and bone morphogenetic protein (BMP) signaling has fundamental roles in the regulation of the stem cell niche for both embryonic and adult stem cells. In zebrafish, male germ stem cell niche is regulated by follicle-stimulating hormone (Fsh) through different members of the TGF-ß superfamily. On the other hand, the specific roles of TGF-ß and BMP signaling pathways are unknown in the zebrafish male germ stem cell niche. Considering this lack of information, the present study aimed to investigate the pharmacological inhibition of TGF-ß (A83-01) and BMP (DMH1) signaling pathways in the presence of recombinant zebrafish Fsh using testicular explants. We also reanalyzed single cell-RNA sequencing (sc-RNA-seq) dataset from adult zebrafish testes to identify the testicular cellular sites of smad expression, and to understand the physiological significance of the changes in smad transcript levels after inhibition of TGF-ß or BMP pathways. Our results showed that A83-01 potentiated the pro-stimulatory effects of Fsh on spermatogonial differentiation leading to an increase in the proportion area occupied by differentiated spermatogonia with concomitant reduction of type A undifferentiated (Aund) spermatogonia. In agreement, expression analysis showed lower mRNA levels for the pluripotency gene pou5f3, and increased expression of dazl (marker of type B spermatogonia and spermatocyte) and igf3 (pro-stimulatory growth factor) following the co-treatment with TGF-ß inhibitor and Fsh. Contrariwise, the inhibition of BMP signaling nullified the pro-stimulatory effects of Fsh, resulting in a reduction of differentiated spermatogonia and increased proportion area occupied by type Aund spermatogonia. Supporting this evidence, BMP signaling inhibition increased the mRNA levels of pluripotency genes nanog and pou5f3, and decreased dazl levels when compared to control. The sc-RNA-seq data unveiled a distinctive pattern of smad expression among testicular cells, primarily observed in spermatogonia (smad 2, 3a, 3b, 8), spermatocytes (smad 2, 3a, 8), Sertoli cells (smad 1, 3a, 3b), and Leydig cells (smad 1, 2). This finding supports the notion that inhibition of TGF-ß and BMP signaling pathways may predominantly impact cellular components within the spermatogonial niche, namely spermatogonia, Sertoli, and Leydig cells. In conclusion, our study demonstrated that TGF-ß and BMP signaling pathways exert antagonistic roles in the zebrafish germ stem cell niche. The members of the TGF-ß subfamily are mainly involved in maintaining the undifferentiated state of spermatogonia, while the BMP subfamily promotes spermatogonial differentiation. Therefore, in the complex regulation of the germ stem cell niche by Fsh, members of the BMP subfamily (pro-differentiation) should be more predominant in the niche than those belonging to the TGF-ß (anti-differentiation). Overall, these findings are not only relevant for understanding the regulation of germ stem cell niche but may also be useful for expanding in vitro the number of undifferentiated spermatogonia more efficiently than using recombinant hormones or growth factors.

Increased levels of soluble triggering receptor expressed on myeloid cells-1 in patients with acute pancreatitis.

OBJECTIVE: To determine the contribution of triggering receptor expressed on myeloid cells (TREM)-1 in acute pancreatitis (AP). DESIGN: Prospective study. SETTING: General intensive care unit at Kobe University Hospital. PATIENTS: Forty-eight patients with AP and seven patients as control. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: We measured serum concentrations of soluble TREM-1 (sTREM-1) at the time of admission by enzyme-linked immunoadsorbent assay. Serum sTREM-1 levels increased significantly in AP (63 +/- 11 pg/mL) and correlated with Ranson score (R = .628, p < .001) and Acute Physiology and Chronic Health Evaluation II score (R = .504, p < .001). Serum TREM-1 levels were higher in patients with early organ dysfunction (which occurred within 7 days after onset) than those without early organ dysfunction (101 +/- 19 vs. 25 +/- 4 pg/mL, p < .001). Incidences of early organ dysfunction in patients whose serum sTREM-1 levels were < or = 40 and > 40 pg/mL were 17% and 83%, respectively (p < .001). The usefulness of serum sTREM-1 in detecting early organ dysfunction was superior to that of C-reactive protein, interleukin-6, interleukin-8, Ranson score, and Acute Physiology and Chronic Health Evaluation II score. Serum sTREM-1 levels decreased with resolution of early organ dysfunction. CONCLUSIONS: Serum sTREM-1 levels were significantly increased and correlated with disease severity and early organ dysfunction in patients with AP. Serum sTREM-1 level may be a useful marker for early organ dysfunction in AP.