Macrophage recruitment and pro-inflammatory differentiation are hallmarks of various diseases, including infection and sepsis. Although studies suggest that mitochondria may regulate macrophage immune responses, it remains unclear whether mitochondrial mass affects macrophage pro-inflammatory differentiation. Here, we found that lipopolysaccharide (LPS)-activated macrophages possess higher mitochondrial mass than resting cells. Therefore, this study aimed to explore the functional role and molecular mechanisms of increased mitochondrial mass in pro-inflammatory differentiated macrophages. Results show that an increase in the mitochondrial mass of macrophages positively correlates with inflammatory cytokine generation in response to LPS. RNA-seq analysis revealed that LPS promotes signal transducers and activators of transcription 2 (Stat2) and dynamin-related protein 1 (Drp1) expression, which are enriched in positive mitochondrial fission regulation. Meanwhile, knockdown or pharmacological inhibition of Drp1 blunts LPS-induced mitochondrial mass increase and pro-inflammatory differentiation. Moreover, Stat2 boosts Drp1 phosphorylation at serine 616, required for Drp1-mediated mitochondrial fission. LPS also causes Stat2-and Drp1-dependent biogenesis, which contributes to the generation of additional mitochondria. However, these mitochondria are profoundly remodeled, displaying fragmented morphology, loose cristae, reduced Δψm, and metabolic programming. Furthermore, these remodeled mitochondria shift their function from ATP synthesis to reactive oxygen species (ROS) production, which drives NFκB-dependent inflammatory cytokine transcription. Interestingly, an increase in mitochondrial mass with constitutively active phosphomimetic mutant of Drp1 (Drp1 S616E ) boosted pro-inflammatory response in macrophages without LPS stimulation. In vivo, we also demonstrated that Mdivi-1 administration inhibits LPS-induced macrophage pro-inflammatory differentiation. Importantly, we observed Stat2 phosphorylation and Drp1-dependent mitochondrial mass increase in macrophages isolated from LPS-challenged mice. In conclusion, we comprehensively demonstrate that a Stat2-Drp1 dependent mitochondrial mass increase is necessary for pro-inflammatory differentiation of macrophages. Therefore, targeting the Stat2-Drp1 axis may provide novel therapeutic approaches for treating infection and inflammatory diseases.

Although macrophage (Mφ) polarization has been demonstrated to play crucial roles in cellular osteogenesis across the cascade of events in periodontal regeneration, how polarized Mφ phenotypes influence the cementoblastic differentiation of periodontal ligament stem cells (PDLSCs) remains unknown. In the present study, human monocyte leukemic cells (THP‐1) were induced into M0, M1, and M2 subsets, and the influences of these polarized Mφs on the cementoblastic differentiation of PDLSCs were assessed in both conditioned medium‐based and Transwell‐based coculture systems. Furthermore, the potential pathways and cyto‐/chemokines involved in Mφ‐mediated cementoblastic differentiation were screened and identified. In both systems, M2 subsets increased cementoblastic differentiation‐related gene/protein expression levels in cocultured PDLSCs, induced more PDLSCs to differentiate into polygonal and square cells, and enhanced alkaline phosphatase activity in PDLSCs. Furthermore, Akt and c‐Jun N‐terminal Kinase (JNK) signaling was identified as a potential pathway involved in M2 Mφ‐enhanced PDLSC cementoblastic differentiation, and cyto‐/chemokines (interleukin (IL)‐10 and vascular endothelial growth factor [VEGF]) secreted by M2 Mφs were found to be key players that promoted cell cementoblastic differentiation by activating Akt signaling. Our data indicate for the first time that Mφs are key modulators during PDLSC cementoblastic differentiation and are hence very important for the regeneration of multiple periodontal tissues, including the cementum. Although the Akt and JNK pathways are involved in M2 Mφ‐enhanced cementoblastic differentiation, only the Akt pathway can be activated via a cyto‐/chemokine‐associated mechanism, suggesting that players other than cyto‐/chemokines also participate in the M2‐mediated cementoblastic differentiation of PDLSCs. Stem Cells 2019;37:1567–1580

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Epithelial-to-mesenchymal transition (EMT) is critical to the progression of several disease processes including carcinoma metastasis and organ fibrosis. Recent studies show that reactive oxygen species (ROS) and mitochondrial dysfunction have been associated with EMT. However, the role of mitochondria in the EMT process remains to be elucidated. Through the induction of EMT using TGF-β1, we demonstrated that mitochondrial functions were abnormal by increasing ROS production and reducing mitochondrial membrane potential, ATP content and mitochondrial complex protein expression. Resveratrol, a mitochondria protective agent, was found to prevent EMT by preserving mitochondrial functions during the process. However, the inhibitory effects of resveratrol on EMT were abolished in mitochondrial DNA-depleted cells. These findings suggest a critical role for mitochondria in EMT and implicate the protection of mitochondria as a potential target to prevent EMT to treat tumour metastasis or tissue fibrosis, and other diseases involving with mitochondria.