TWEAK exacerbates allergic conjunctivitis via PKM2-dependent NF-κB activation and macrophage M1 polarization.

Abstract

BACKGROUND: Allergic conjunctivitis (AC) is a prevalent ocular inflammatory disease with unmet therapeutic needs. This study aimed to investigate whether TWEAK regulates NF-κB activation and macrophage polarization, and the underlying mechanism in AC. METHODS: In vivo, an ovalbumin-induced mouse model of AC was established to evaluate clinical scores, histopathology, and inflammatory markers. In vitro, macrophages were co-cultured with conjunctival cells subjected to TWEAK modulation to assess PKM2 expression/distribution, NF-κB activation, and polarization phenotypes in macrophages. Co-IP, WB, qRT-PCR, IHC, IF, and flow cytometry were employed for mechanistic studies. RESULTS: In AC mice, TWEAK knockdown reduced PKM2 expression, inhibited NF-κB signaling, and shifted macrophages from pro-inflammatory M1 to anti-inflammatory M2 phenotype, alleviating disease severity. In vitro, TWEAK overexpression enhanced PKM2 nuclear translocation, facilitated PKM2-NF-κB binding, activated NF-κB pathway and drove macrophage M1 polarization. Mechanistically, the inhibition of PKM2 nuclear translocation abrogated TWEAK-induced NF-κB activation and M1 polarization. Moreover, PKM2 overexpression reversed the anti-inflammatory effects of TWEAK knockdown in AC mice, whereas direct NF-κB activation bypassed the requirement for both TWEAK. CONCLUSION: In summary, our findings revealed that TWEAK promoted NF-κB activation and macrophage M1 polarization to exacerbate AC through PKM2-dependent mechanisms. This study identified the TWEAK-PKM2-NF-κB axis as a critical regulator of AC pathogenesis.