Combined in vitro microcurrent stimulation and celecoxib suppress NF-κB/NLRP3 signaling and cartilage catabolism in human chondrocytes.

Osteoarthritis is characterized by persistent inflammatory signaling, oxidative stress, chondrocyte dysfunction, and extracellular-matrix degradation. This study investigated whether controlled in vitro microcurrent stimulation could enhance the anti-inflammatory and chondroprotective effects of celecoxib in interleukin-1β (IL-1β)-challenged human chondrocytes. Immortalized C28/I2 chondrocytes and primary human knee articular chondrocytes were treated with celecoxib, microcurrent stimulation, or their combination. A factorial concentration-current matrix, Bliss-independence analysis, and two-way analysis of variance were used to evaluate interaction effects on IL-6 secretion. Selected inflammatory, anabolic, catabolic, NF-κB, COX-2/PGE₂, NLRP3-associated, oxidative-stress, mitochondrial, and apoptosis-related endpoints were examined at the submaximal combination of 2.5 µM celecoxib and 25 µA microcurrent stimulation. PGE₂ add-back and nigericin challenge experiments were performed to investigate the involvement of PGE₂- and NLRP3-associated signaling. IL-1β induced a pronounced inflammatory and catabolic phenotype. Celecoxib and microcurrent stimulation individually reduced selected inflammatory mediators, NF-κB signaling, matrix-catabolic markers, oxidative stress, and pyroptosis-associated endpoints. The celecoxib × microcurrent interaction for IL-6 inhibition was significant (F[12,40] = 3.67, P = 0.0009). Bliss analysis identified synergy in eight of the twelve nonzero combinations, whereas four combinations were not distinguishable from Bliss additivity. The greatest Bliss excess occurred with 25 µA microcurrent stimulation plus 2.5 µM celecoxib (18.0% points; 95% bootstrap confidence interval, 12.7-23.7). At this condition, combined treatment produced greater IL-6 and PGE₂ inhibition and more favorable MMP13 and COL2A1 responses than either monotherapy. Because formal factorial synergy analysis was performed only for IL-6, improvements in the other endpoints were interpreted as enhanced combined responses rather than demonstrated synergy. Combined treatment was associated with reduced MYD88 expression, NF-κB pathway phosphorylation and nuclear p65 localization, together with suppression of COX-2/PGE₂-associated signaling, lower NLRP3/caspase-1/GSDMD-related responses and oxidative stress, and preservation of selected mitochondrial indices. Exogenous PGE₂ and nigericin partially reversed selected responses, supporting pathway involvement without establishing causal necessity or dependency. Selected treatment responses were reproduced in primary human knee articular chondrocytes. Controlled in vitro microcurrent stimulation combined with celecoxib produced concentration- and current-specific Bliss synergy for IL-6 inhibition and enhanced selected inflammatory and matrix-protective responses. These findings support further evaluation in physiologically relevant cartilage and in vivo osteoarthritis models but should not be interpreted as evidence of clinical efficacy or equivalence to clinical electroacupuncture.