Pro-inflammatory agents nitric oxide and TNFα arrest GC-1 spermatogonia cell cycle through different mechanism

AMARILLA MARÍA SOFÍA; FERREIRO ME; GLIENKE L; GONZÁLEZ C; JACOBO PV; SOBARZO CM; DE LAURENTIIS A; FERRARIS M J; THEAS M S

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Congreso; LXIV Reunión de las Pro-inflammatory agents nitric oxide and TNFα arrest GC-1 spermatogonia cell cycle through different mechanism; 2019

Pro-inflammatory agents nitric oxide and TNFα arrest GC-1 spermatogonia cell cycle through different mechanism

Nitric oxide (NO) and Tumor Necrosis Factor alpha (TNFα) are pro-inflammatory agents able to interfere with cell cycle. Experimental autoimmune orchitis (EAO) is a model of chronic inflammation associated to infertility. In EAO high levels of NO and TNFα are produced by testicular macrophages and pre-meiotic germ cells (spermatogonia and pre-leptotene spermatocytes) proliferation is reduced. We propose that NO and TNFα arrest spermatogonial cell cycle in EAO. To evaluate this hypothesis we explored the effect of DETA-NO (a NO donor) and TNFα on cell cycle and death on GC-1 spermatogonia cell line by flow cytometry. Both TNF-α (50ng/ml) and DETA-NO (2.0mM), significantly increased the percentage of GC-1 cells in the S-phase and significantly reduced the percentage in the G1-phase of the cell cycle (Propidium Iodide incorporation, IP) also inducing cell apoptosis (Annexin V-FITC-IP assay) after 24 and 18 h of incubation respectively. Pre-incubation of GC-1 cells with a general antioxidant, N-acetyl-L-cysteine (NAC, 2.5 and 5.0 mM) significantly reduced DETA-NO effect on cell cycle arrest and apoptosis while NAC did not modify TNFα action. DETA-NO induced GC-1 cell cycle arrest and apoptosis was reverted after DETA-NO withdrawal unlike TNFα.Our results showed that NO and TNFα arrest the cell cycle of GC-1 spermatogonia. While NO effect on GC-1 cells was mediated by oxidative stress and reversible, TNFα action was independent of oxidative stress and irreversible. We conclude that NO and TNFα might control spermatogonial cycle progression through different pathways in the testis under chronic inflammation.