TIFAB regulates the TIFA-TRAF6 signaling pathway involved in innate immunity by forming a heterodimer complex with TIFA.

Nuclear factor κB (NF-κB) is activated by various inflammatory and infectious molecules and is involved in immune responses. It has been elucidated that ADP-ß-D-manno-heptose (ADP-Hep), a metabolite in gram-negative bacteria, activates NF-κB through alpha-kinase 1 (ALPK1)-TIFA-TRAF6 signaling. ADP-Hep stimulates the kinase activity of ALPK1 for TIFA phosphorylation. Complex formation between phosphorylation-dependent TIFA oligomer and TRAF6 promotes the polyubiquitination of TRAF6 for NF-κB activation. TIFAB, a TIFA homolog lacking a phosphorylation site and a TRAF6 binding motif, is a negative regulator of TIFA-TRAF6 signaling and is implicated in myeloid diseases. TIFAB is indicated to regulate TIFA-TRAF6 signaling through interactions with TIFA and TRAF6; however, little is known about its biological function. We demonstrated that TIFAB forms a complex not with the TIFA dimer, an intrinsic form of TIFA involved in NF-κB activation, but with monomeric TIFA. The structural analysis of the TIFA/TIFAB complex and the biochemical and cell-based analyses showed that TIFAB forms a stable heterodimer with TIFA, inhibits TIFA dimer formation, and suppresses TIFA-TRAF6 signaling. The resultant TIFA/TIFAB complex is a "pseudo-TIFA dimer" lacking the phosphorylation site and TRAF6 binding motif in TIFAB and cannot form the orderly structure as proposed for the phosphorylated TIFA oligomer involved in NF-κB activation. This study elucidated the molecular and structural basis for the regulation of TIFA-TRAF6 signaling by TIFAB.

Precision treatment of viral pneumonia through macrophage-targeted lipid nanoparticle delivery.

Macrophages are integral components of the innate immune system, playing a dual role in host defense during infection and pathophysiological states. Macrophages contribute to immune responses and aid in combatting various infections, yet their production of abundant proinflammatory cytokines can lead to uncontrolled inflammation and worsened tissue damage. Therefore, reducing macrophage-derived proinflammatory cytokine release represents a promising approach for treating various acute and chronic inflammatory disorders. However, limited macrophage-specific delivery vehicles have hindered the development of macrophage-targeted therapies. In this study, we screened a pool of 112 lipid nanoparticles (LNPs) to identify an optimal LNP formulation for efficient siRNA delivery. Subsequently, by conjugating the macrophage-specific antibody F4/80 to the LNP surface, we constructed MacLNP, an enhanced LNP formulation designed for targeted macrophage delivery. In both in vitro and in vivo experiments, MacLNP demonstrated a significant enhancement in targeting macrophages. Specifically, delivery of siRNA targeting TAK1, a critical kinase upstream of multiple inflammatory pathways, effectively suppressed the phosphorylation/activation of NF-kB. LNP-mediated inhibition of NF-kB, a key upstream regulator in the classic inflammatory signaling pathway, in the murine macrophage cell line RAW264.7 significantly reduced the release of proinflammatory cytokines after stimulation with the viral RNA mimic Poly(I:C). Finally, intranasal administration of MacLNP-encapsulated TAK1 siRNA markedly ameliorated lung injury induced by influenza infection. In conclusion, our findings validate the potential of targeted macrophage interventions in attenuating inflammatory responses, reinforcing the potential of LNP-mediated macrophage targeting to treat pulmonary inflammatory disorders.

Notopterol Inhibits the NF-κB Pathway and Activates the PI3K/Akt/Nrf2 Pathway in Periodontal Tissue.

Notopterol, an active component isolated from the traditional Chinese medicine Notopterygium incisum Ting ex H.T. Chang, exerts anti-inflammatory activity in rheumatoid arthritis. However, its roles in suppression of inflammatory insults and halting progression of tissue destruction in periodontitis remain elusive. In this study, we reveal that notopterol can inhibit osteoclastogenesis, thereby limiting alveolar bone loss in vivo. In vitro results demonstrated that notopterol administration inhibited synthesis of inflammatory mediators such as IL-1ß, IL-32, and IL-8 in LPS-stimulated human gingival fibroblasts. Mechanistically, notopterol inhibits activation of the NF-κB signaling pathway, which is considered a prototypical proinflammatory signaling pathway. RNA sequencing data revealed that notopterol activates the PI3K/protein kinase B (Akt)/NF-E2-related factor 2 (Nrf2) signaling pathway in LPS-stimulated human gingival fibroblasts, a phenomenon validated via Western blot assay. Additionally, notopterol treatment suppressed reactive oxygen species levels by upregulating the expression of antioxidant genes, including heme oxygenase 1 (HO-1), NAD(P)H quinone oxidoreductase 1 (NQO1), catalase (CAT), and glutathione reductase (GSR), indicating that notopterol confers protection against oxidative stress. Notably, inhibition of Akt activity by the potent inhibitor, MK-2206, partially attenuated both anti-inflammatory and antioxidant effects of notopterol. Collectively, these results raise the possibility that notopterol relieves periodontal inflammation by suppressing and activating the NF-κB and PI3K/AKT/Nrf2 signaling pathways in periodontal tissue, respectively, suggesting its potential as an efficacious treatment therapy for periodontitis.

Acute pulpitis promotes purinergic signaling to induce pain in rats via P38MAPK/NF-κB signaling pathway.

Toothache is one of the most common types of pain, but the mechanisms underlying pulpitis-induced pain remain unknown. The ionotropic purinergic receptor family (P2X) is reported to mediate nociception in the nervous system. This study aims to investigate the involvement of P2X3 in the sensitisation of the trigeminal ganglion (TG) and the inflammation caused by acute pulpitis. An acute tooth inflammation model was established by applying LPS to the pulp of SD rats. We found that the increased expression of P2X3 was induced by acute pulpitis. A selective P2X3 inhibitor (A-317491) reduced pain-like behavior in the maxillofacial region of rats and depressed the activation of neurons in the trigeminal ganglion induced by pulpitis. The upregulated MAPK signaling (p-p38, p-ERK1/2) expression in the ipsilateral TG induced by pulpitis could also be depressed by the application of the P2X3 inhibitor. Furthermore, the expression of markers of inflammatory processes, such as NF-κB, TNF-α and IL-1ß, could be induced by acute pulpitis and deduced by the intraperitoneal injection of P2X3 antagonists. Our findings demonstrate that purinergic P2X3 receptor signaling in TG neurons contributes to pulpitis-induced pain in rats and that P2X3 signaling may be a potential therapeutic target for tooth pain.

Bone-Targeted Biomimetic Nanogels Re-Establish Osteoblast/Osteoclast Balance to Treat Postmenopausal Osteoporosis.

Insufficient bone formation and excessive bone resorption caused by estrogen deficiency are the major factors resulting in the incidence of postmenopausal osteoporosis (PMOP). The existing drugs usually fail to re-establish the osteoblast/osteoclast balance from both sides and generate side-effects owing to the lack of bone-targeting ability. Here, engineered cell-membrane-coated nanogels PNG@mR&C capable of scavenging receptor activator of nuclear factor-κB ligand (RANKL) and responsively releasing therapeutic PTH 1-34 in the bone microenvironment are prepared from RANK and CXCR4 overexpressed bone mesenchymal stem cell (BMSC) membrane-coated chitosan biopolymers. The CXCR4 on the coated-membranes confer bone-targeting ability, and abundant RANK effectively absorb RANKL to inhibit osteoclastogenesis. Meanwhile, the release of PTH 1-34 triggered by osteoclast-mediated acid microenvironment promote osteogenesis. In addition, the dose and frequency are greatly reduced due to the smart release property, prolonged circulation time, and bone-specific accumulation. Thus, PNG@mR&C exhibits satisfactory therapeutic effects in the ovariectomized (OVX) mouse model. This study provides a new paradigm re-establishing the bone metabolic homeostasis from multitargets and shows great promise for the treatment of PMOP.

miR-708-5p deficiency involves the degeneration of mandibular condylar chondrocytes via the TLR4/NF-κB pathway.

OBJECTIVE: Ageing and aberrant biomechanical stimulation are two major risk factors for osteoarthritis (OA). One of the main characteristics of aged cartilage is cellular senescence. One of the main characteristics of osteoarthritic joints is cartilage degeneration. The cells in the temporomandibular joint (TMJ) cartilage are zonally arranged. The deep zone cells are differentiated from the superficial zone cells (SZCs). The purpose of the present study was to investigate whether degenerative shear stress (SS) stimulates the senescence programme in TMJ SZCs, and to determine which miRNA is involved in this process. METHOD: SZCs were isolated from the TMJ condyles of 3-week-old rats and treated with continuous passaging or SS. RNA sequencing was conducted to identify miRNA(s) that overlap with those involved in the replication senescence process and the SS-induced degeneration programme. Unilateral anterior crossbite (UAC), which is TMJ-OA inducible, was applied to 2-month-old and 12-month-old mice for 3 weeks. The effect of TMJ local injection of agomiR-708-5p was evaluated histologically. RESULTS: Both replication and SS treatment induced SZC senescence. miR-708-5p was identified. Knocking down miR-708-5p in SS-treated SZCs led to more severe senescence by alleviating the inhibitory impact of miR-708-5p on the TLR4/NF-κB pathway. miR-708-5p expression in mouse TMJ cartilage decreased with age. UAC induced more severe osteoarthritic cartilage lesions in 12-month-old mice than in 2-month-old mice. Injection of agomiR-708-5p suppressed UAC-induced osteoarthritic cartilage lesions. CONCLUSIONS: Age-related miR-708-5p deficiency is involved in the mechanically stimulated OA process. Intra-articular administration of agomiR-708-5p is a promising new strategy for OA treatment.

Inflammatory Periodontal Ligament Stem Cells Drive M1 Macrophage Polarization via Exosomal miR-143-3p-Mediated Regulation of PI3K/AKT/NF-κB Signaling.

Macrophage polarization plays an important role in the progression of inflammation. Exosomes derived from stem cells are promising candidates for macrophage immunoregulation. However, how exosomes derived from periodontal ligament stem cells (PDLSCs) in an inflammatory environment influence macrophage polarization has yet to be fully elucidated. In this study, inflammatory PDLSCs were found to downregulate M2 macrophage polarization at the mRNA and protein levels in a Transwell coculture system of PDLSCs and THP-1-derived M0 macrophages. Furthermore, inflammatory PDLSC-derived exosomes shifted macrophages toward the M1 phenotype. The inhibition of inflammatory PDLSC-derived exosomes by GW4869 weakened inflammatory PDLSC-mediated M1 macrophage polarization. A miRNA microarray was used to determine the differential miRNAs shuttled by healthy and inflammatory PDLSC-derived exosomes. Compared with healthy exosomes, miR-143-3p was enriched in inflammatory PDLSC-derived exosomes, which targeted and inhibited the expression of PI3Kγ and promoted M1 macrophage polarization by suppressing PI3K/AKT signaling and activating NF-κB signaling, while an agonist of the PI3K pathway reversed this effect. Moreover, exosome-shuttled miR-143-3p from PDLSCs drove M1 macrophage polarization and aggravated periodontal inflammation in a mouse periodontitis model. In conclusion, these results demonstrate that inflammatory PDLSCs facilitate M1 macrophage polarization through the exosomal miR-143-3p-mediated regulation of PI3K/AKT/NF-κB signaling, providing a potential new target for periodontitis treatment.

Galectin-3 inhibition alleviated LPS-induced periodontal inflammation in gingival fibroblasts and experimental periodontitis mice.

OBJECTIVES: Clinical studies have confirmed that galectin-3 (Gal-3) levels are significantly elevated in periodontitis patients. The present study aimed to explore the effects of Gal-3 inhibition on periodontal inflammation in vitro and in vivo. METHODS: Human gingival fibroblasts (HGFs) with or without Gal-3 knockdown were stimulated by lipopolysaccharide (LPS), and a ligation-induced mouse periodontitis model treated with a Gal-3 inhibitor was established. Hematoxylin-eosin (H&E) and immunohistochemistry (IHC) staining were used to evaluate Gal-3 levels in gingival tissues. Quantitative real-time polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA) were used to detect Gal-3, interleukin (IL)-6, IL-8, and C-C motif ligand 2 (CCL2) expression. Immunofluorescence and western blotting were used to detect NF-κB and ERK signaling pathway activation. Micro-computed tomography was used to analyse the degree of bone loss. RESULTS: Gal-3 was significantly up-regulated in inflamed gingival tissues and LPS-induced HGFs. Gal-3 knockdown markedly decreased LPS-induced IL-6, IL-8, and CCL2 expression and blocked NF-κB and ERK signaling pathway activation in HGFs. In the mouse periodontitis model, Gal-3 inhibition significantly alleviated IL-1ß and IL-6 infiltration in gingival tissue and mitigated periodontal bone loss. CONCLUSIONS: Gal-3 inhibition notably alleviated periodontal inflammation partly through blocking NF-κB and ERK signaling pathway activation.

MST1/2 exerts a pivotal role in inducing neuroinflammation and Coxsackievirus-A10 replication by interacting with innate immunity.

Coxsackievirus-A10 (CV-A10), responsible for the hand, foot and mouth disease (HFMD) pandemic, could cause serious central nervous system (CNS) complications. The underlying molecular basis of CV-A10 and host interactions inducing neuropathogenesis is still unclear. The Hippo signaling pathway, historically known for a dominator of organ development and homeostasis, has recently been implicated as an immune regulator. However, its role in host defense against CV-A10 has not been investigated. Herein, it was found that CV-A10 proliferated in HMC3 cells and promoted the release of inflammatory cytokines. Moreover, pattern recognition receptors (PRRs)-mediated pathways, including TLR3-TRIF-TRAF3-TBK1-NF-κB axis, RIG-I/MDA5-MAVS-TRAF3-TBK1-NF-κB axis and TLR7-MyD88-IRAK1/IRAK4-TRAF6-TAK1-NF-κB axis, were examined to be elevated under CV-A10 infection. Meanwhile, it was further uncovered that Hippo signaling pathway was inhibited in HMC3 cells with CV-A10 infection. Previous studies have been reported that there exist complex relations between innate immune and Hippo signaling pathway. Then, plasmids of knockdown and overexpression of MST1/2 were transfected into HMC3 cells. Our results showed that MST1/2 suppressed the levels of inflammatory cytokines via interacting with TBK1 and IRAK1, and also enhanced virus production via restricting IRF3 and IFN-ß expressions. Overall, these data obviously pointed out that CV-A10 accelerated the formation of neuroinflammation by the effect of the Hippo pathway on the PRRs-mediated pathway, which delineates a negative immunoregulatory role for MST1/2 in CV-A10 infection and the potential for this pathway to be pharmacologically targeted to treat CV-A10.

Polyethylene microplastics induced inflammation via the miR-21/IRAK4/NF-κB axis resulting to endoplasmic reticulum stress and apoptosis in muscle of carp.

As a widespread environmental pollutant, microplastics pose a great threat to the tissues and organs of aquatic animals. The carp's muscles are necessary for movement and survival. However, the mechanism of injury of polyethylene microplastics (PE-MPs) to carp muscle remains unclear. Therefore, in this study, PE-MPs with the diameter of 8 µm and the concentration of 1000 ng/L were used to feed carp for 21 days, and polyethylene microplastic treatment groups was established. The results showed that PE-MPs could cause structural abnormalities and disarrangement of muscle fibers, and aggravate oxidative stress in muscles. Exposure to PE-MPs reduced microRNA (miR-21) in muscle tissue, negatively regulated Interleukin-1 Receptor Associated Kinase 4 (IRAK4), activated Nuclear Factor Kappa-B (NF-κB) pathway, induced inflammation, and led to endoplasmic reticulum stress and apoptosis. The present study provides different targets for the prevention of muscle injury induced by polyethylene microplastics.