Pulsed-Ultrasound Irradiation Induces the Production of Itaconate and Attenuates Inflammatory Responses in Macrophages.

BACKGROUND

Itaconate is a key metabolite in the innate immune system and exerts strong anti-inflammatory effects in macrophages. For the production of itaconate in macrophages, immune-responsive gene 1 (IRG1) is an imperative enzyme, and activating the IRG1-itaconate pathway is reported to alleviate inflammatory diseases by upregulating nuclear factor-erythroid 2-related factor 2 (NRF2). However, there are very few reports on strategies to increase itaconate production. Ultrasound therapy is a widely used intervention for anti-inflammatory and soft-tissue regeneration purposes. Here we show the effect of ultrasound irradiation on the production of itaconate in macrophages.

METHODS

Murine bone marrow-derived macrophages (BMDMs) were exposed to pulsed ultrasound (3.0 W/cm) for 5 minutes. Three hours after irradiation, the intracellular levels of metabolites and mRNA expression levels of and were measured using CE/MS and qPCR, respectively. To evaluate macrophage inflammation status, 3 h after irradiation, the cells were stimulated with 100 ng/mL lipopolysaccharide (LPS) for 1.5 h and the mRNA expression levels of pro-inflammatory factors (, and ) were measured. Student's -test, one-way ANOVA and Tukey's multiple comparison test were used for statistical processing, and the significance level was set to less than 5%.

RESULTS

Ultrasound irradiation significantly increased the intracellular itaconate level and the expression levels of and in BMDMs. Upregulation of , and by LPS was significantly suppressed in BMDMs treated with ultrasound. Ultrasound irradiation did not affect cell viability and apoptosis.

CONCLUSION

Ultrasound irradiation induces the production of itaconate by upregulating expression and attenuates inflammatory responses in macrophages via . These results suggest that ultrasound is a potentially useful method to increase itaconate production in macrophages.