Exercise Training Modulates TLR4/NF-κB-Mediated Inflammatory Signaling in Rat Testes after Tramadol Withdrawal

Chronic tramadol (TRA) exposure has been reported to induce inflammatory responses in testicular tissue, whereas exercise training exerts immunomodulatory effects within the testes. Accordingly, this study investigated whether exercise training protocols (ETPs) during the post-withdrawal period modulate testicular inflammation via the TLR4/NF-κB-mediated inflammatory signaling pathway.

In this experimental study, adult Wistar rats (n=36; sex: male; age: 8 weeks; body weight: 180-220 g) were randomly assigned to a control group and tramadol-treated groups. The control group received normal saline. TRA was administered at a dose of 40 mg/kg, via intraperitoneal route, for 60 days. TRA withdrawal was induced by stopping TRA administration after 60 days and maintained for an additional 60 days. Following withdrawal, tramadol-exposed rats were allocated to a withdrawal-only group or to low-, moderate-, or high-intensity continuous exercise training groups. At the end of the experimental period, testicular tissue samples were collected. The expression or levels of IL-6, IL-10, COX-II, TLR4, NF-κB, TNF-α, and iNOS parameters were evaluated using quantitative reverse transcription polymerase chain reaction (qRT-PCR), enzyme-linked immunosorbent assay (ELISA), immunohistochemistry, immunofluorescence staining, and Western blotting methods.

The TRA administration increased the expression of TLR4, NF-κB, IL-6, and TNF-α. These elevations were reduced after withdrawal and were more markedly attenuated in exercise-trained groups. IL-10 levels were decreased in the tramadol-only group but were restored during withdrawal with exercise training. TRA exposure was associated with increased numbers of iNOS+ and COX-II+ somatic and germ cells; these alterations were reversed after withdrawal, particularly in exercised animals. Despite TRA cessation, residual immune dysregulation persisted, characterized by elevated TLR4, NF-κB, IL-6, and TNF-α expression.

Exercise training following TRA withdrawal could restore inflammatory balance by suppressing TLR4 and NF-ΚB signaling, normalizing pro- and anti-inflammatory cytokine profiles, and possibly attenuating the iNOS/NO/COXII pathway in Wistar rats.