Lipoxin A4 modulates the PKA/CREB and NF-κB signaling pathway to mitigate chondrocyte catabolism and apoptosis in temporomandibular joint osteoarthritis.

Temporomandibular joint osteoarthritis (TMJ-OA) is characterized by the degradation of the extracellular matrix (ECM) in cartilage and the apoptosis of chondrocytes, which is caused by inflammation and disruptions of chondrocyte metabolism and inflammation. Lipoxin A4 (LXA4), a specialized pro-resolving mediator, has been shown to inhibit inflammation and regulate the balance between ECM synthesis and degradation. However, the therapeutic effects of LXA4 on TMJ-OA and its underlying mechanisms remain unclear. Interleukin-1 beta (IL-1ß)-induced chondrocyte and surgically induced TMJ-OA rat models were established in this study. The viability of chondrocytes treated with LXA4 was evaluated with the cell counting kit-8 (CCK-8) assay, while protein levels were assessed by western blot analysis, and the apoptosis rate was evaluated with terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labelling (TUNEL) staining. Histological analysis was conducted to evaluate the impact of LXA4 on cartilage degradation in TMJ-OA rat models. In vitro, the qRT-PCR and western blot analysis demonstrated that LXA4 facilitated the upregulation of collagen proteins (Collagen II) and decreased expression of matrix metalloproteinases (MMP-3, and MMP-13) associated with ECM modulation. LXA4 enhanced the TMJ-OA chondrocyte viability and decreased apoptotic rate. In vivo, histology and immunohistochemistry (IHC) analysis revealed that intraperitoneal injection of LXA4 contributed to the amelioration of chondrocyte injuries and deceleration of TMJ-OA. Transcriptomic sequencing revealed that cAMP signaling pathway was up-regulated and NF-κB signaling pathway was down-regulated in LXA4 treated group. LXA4 inhibited the phosphorylation of P65 and inhibitor of nuclear factor kappa B (IκBα) proteins while enhancing the phosphorylation PKA and CREB. This study demonstrates the potential of LXA4 as a therapeutic agent for suppressing chondrocyte catabolism and apoptosis by increasing PKA/CREB activity and decreasing NF-κB signaling.

Biased receptor signalling and intracellular trafficking profiles of structurally distinct formylpeptide receptor 2 agonists.

BACKGROUND: There is increasing interest in developing FPR2 agonists (compound 43, ACT-389949 and BMS-986235) as potential pro-resolving therapeutics, with ACT-389949 and BMS-986235 having entered phase I clinical development. FPR2 activation leads to diverse downstream outputs. ACT-389949 was observed to cause rapid tachyphylaxis, while BMS-986235 and compound 43 induced cardioprotective effects in preclinical models. We aim to characterise the differences in ligand-receptor engagement and downstream signalling and trafficking bias profile. EXPERIMENTAL APPROACH: Concentration-response curves to G protein dissociation, ß-arrestin recruitment, receptor trafficking and second messenger signalling were generated using FPR2 ligands (BMS-986235, ACT-389949, compound 43 and WKYMVm), in HEK293A cells. Log(τ/KA) was obtained from the operational model for bias analysis using WKYMVm as a reference ligand. Docking of FPR2 ligands into the active FPR2 cryoEM structure (PDBID: 7T6S) was performed using ICM pro software. KEY RESULTS: Bias analysis revealed that WKYMVm and ACT-389949 shared a very similar bias profile. In comparison, BMS-986235 and compound 43 displayed approximately 5- to 50-fold bias away from ß-arrestin recruitment and trafficking pathways, while being 35- to 60-fold biased towards cAMP inhibition and pERK1/2. Molecular docking predicted key amino acid interactions at the FPR2 shared between WKYMVm and ACT-389949, but not with BMS-986235 and compound 43. CONCLUSION AND IMPLICATIONS: In vitro characterisation demonstrated that WKYMVm and ACT-389949 differ from BMS-986235 and compound 43 in their signalling and protein coupling profile. This observation may be explained by differences in the ligand-receptor interactions. In vitro characterisation provided significant insights into identifying the desired bias profile for FPR2-based pharmacotherapy.

Lipoxin A4 ameliorates knee osteoarthritis progression in rats by antagonizing ferroptosis through activation of the ESR2/LPAR3/Nrf2 axis in synovial fibroblast-like synoviocytes.

BACKGROUND: Our previous studies have shown that lipoxin A4 (LXA4) can serve as a potential biomarker for assessing the efficacy of exercise therapy in knee osteoarthritis (KOA), and fibroblast-like synoviocytes (FLSs) may play a crucial role in KOA pain as well as in the progression of the pathology. OBJECTIVE: By analyzing the GSE29746 dataset and collecting synovial samples from patients with different Kellgren-Lawrence (KL) grades for validation, we focused on exploring the potential effect of LXA4 on ferroptosis in FLSs through the ESR2/LPAR3/Nrf2 axis to alleviate pain and pathological advancement in KOA. METHODS: The association between FLSs ferroptosis and chondrocyte matrix degradation was explored by cell co-culture. We overexpressed and knocked down LPAR3 in vitro to explore its potential mechanism in FLSs. A rat model of monosodium iodoacetate (MIA)-induced KOA was constructed and intervened with moderate-intensity treadmill exercise and intraperitoneal injection of PHTPP to investigate the effects of the LXA4 intracellular receptor ESR2 on exercise therapy. RESULTS: ESR2, LPAR3, and GPX4 levels in the synovium decreased with increasing KL grade. After LXA4 intervention in the co-culture system, GPX4, LPAR3, and ESR2 were upregulated in FLSs, collagen II was upregulated in chondrocytes, and MMP3 and ADAM9 were downregulated. LPAR3 overexpression upregulated the expression of GPX4, Nrf2, and SOD1 in FLSs, while downregulating the expression of MMP13 and MMP3; LPAR3 knockdown reversed these changes. Moderate-intensity platform training improved the behavioral manifestations of pain in KOA rats, whereas PHTPP treatment partially reversed the improvement in synovial and cartilage pathologies induced by platform training. CONCLUSION: LXA4 inhibited FLSs ferroptosis by activating the ESR2/LPAR3/Nrf2 axis, thereby alleviating the pain and pathological progression of KOA. This study brings a new target for the treatment of KOA and also leads to a deeper understanding of the potential mechanisms of exercise therapy for KOA.

Alterations in the pro-resolving lipid mediator machinery within first trimester maternal tissue: Implications in decidualization and miscarriage risk.

A pivotal event in uterine receptivity and human reproduction is the differentiation of endometrial stromal cells into decidual cells, known as decidualization. Decidualization is interlinked with its inflammatory environment. Our study aimed to investigate the presence and role of pro-resolving lipid mediators in first trimester maternal tissue. We assessed the levels of LXA4 and RvD1, along with their metabolic LOX enzymes, in elective (control) and sporadic miscarriage samples. We investigated the effects of LXA4 and RvD1 on decidualization using primary endometrial stromal cells and the immortalized endometrial stromal St-T1b cell line. The upregulation of 12- and 15-LOX expression was observed in pregnancy tissue after sporadic miscarriage, suggesting an inflammatory imbalance. Furthermore, incubation with these lipid mediators led to a decrease in decidualization biomarkers PRL and IGFBP-1, accompanied by morphological changes indicative of aberrant differentiation. The expression of LOX enzymes in decidual natural killer cells suggests their involvement in regulating the inflammatory surroundings and the extent of decidualization.

Association between arachidonate lipoxygenase 15,c.-292 C > T gene polymorphism and non-cystic fibrosis bronchiectasis in children: a pilot study on the effects on airway lipoxin A4 and disease phenotype.

BACKGROUND: Persistent airway inflammation is a central feature of bronchiectasis. Arachidonate 15-lipoxygenase (ALOX-15) controls production of endogenous lipid mediators, including lipoxins that regulate airway inflammation. Mutations at various positions in ALOX-15 gene can influence airway disease development. We investigated association between ALOX-15,c.-292 C > T gene polymorphism and bronchiectasis unrelated to cystic fibrosis in Egyptian children. Also, lipoxin A4 (LXA4) level in bronchoalveolar lavage (BAL) was studied in relation to polymorphism genotypes and disease phenotypes determined by clinical, pulmonary functions, and radiological severity parameters. METHODS: This was an exploratory study that included 60 participants. Thirty children with non-cystic fibrosis bronchiectasis (NCFB) were compared with 30 age and sex-matched controls. ALOX-15,c.-292 C > T polymorphism was genotyped using TaqMan-based Real-time PCR. LXA4 was measured in BAL using ELISA method. RESULTS: There was no significant difference between patients and controls regarding ALOX-15,c.-292 C > T polymorphism genotypes and alleles (OR = 1.75; 95% CI (0.53-5.7), P = 0.35) (OR = 1; 95% CI (0.48-2), p = 1). BAL LXA4 level was significantly lower in patients, median (IQR) of 576.9 (147.6-1510) ng/ml compared to controls, median (IQR) of 1675 (536.8-2542) (p = 0.002). Patients with severe bronchiectasis had a significantly lower LXA4 level (p < 0.001). There were significant correlations with exacerbations frequency (r=-0.54, p = 0.002) and FEV1% predicted (r = 0.64, p = 0.001). Heterozygous CT genotype carriers showed higher LXA4 levels compared to other genotypes(p = 0.005). CONCLUSIONS: Low airway LXA4 in children with NCFB is associated with severe disease phenotype and lung function deterioration. CT genotype of ALOX-15,c.-292 C > T polymorphism might be a protective genetic factor against bronchiectasis development and/or progression due to enhanced LXA4 production.

Can essential fatty acids (EFAs) prevent and ameliorate post-COVID-19 long haul manifestations?

It is hypothesized that COVID-19, post-COVID and post-mRNA COVID-19 (and other related) vaccine manifestations including "long haul syndrome" are due to deficiency of essential fatty acids (EFAs) and dysregulation of their metabolism. This proposal is based on the observation that EFAs and their metabolites can modulate the swift immunostimulatory response of SARS-CoV-2 and similar enveloped viruses, suppress inappropriate cytokine release, possess cytoprotective action, modulate serotonin and bradykinin production and other neurotransmitters, inhibit NF-kB activation, regulate cGAS-STING pathway, modulate gut microbiota, inhibit platelet activation, regulate macrophage and leukocyte function, enhance wound healing and facilitate tissue regeneration and restore homeostasis. This implies that administration of EFAs could be of benefit in the prevention and management of COVID-19 and its associated complications.

LXA4 protected mice from renal ischemia/reperfusion injury by promoting IRG1/Nrf2 and IRAK-M-TRAF6 signal pathways.

Excessive inflammatory response and increased oxidative stress play an essential role in the pathophysiology of ischemia/reperfusion (I/R)-induced acute kidney injury (IRI-AKI). Emerging evidence suggests that lipoxin A4 (LXA4), as an endogenous negative regulator in inflammation, can ameliorate several I/R injuries. However, the mechanisms and effects of LXA4 on IRI-AKI remain unknown. In this study, A bilateral renal I/R mouse model was used to evaluate the role of LXA4 in wild-type, IRG1 knockout, and IRAK-M knockout mice. Our results showed that LXA4, as well as 5-LOX and ALXR, were quickly induced, and subsequently decreased by renal I/R. LXA4 pretreatment improved renal I/R-induced renal function impairment and renal damage and inhibited inflammatory responses and oxidative stresses in mice kidneys. Notably, LXA4 inhibited I/R-induced the activation of TLR4 signal pathway including decreased phosphorylation of TAK1, p36, and p65, but did not affect TLR4 and p-IRAK-1. The analysis of transcriptomic sequencing data and immunoblotting suggested that innate immune signal molecules interleukin-1 receptor-associated kinase-M (IRAK-M) and immunoresponsive gene 1 (IRG1) might be the key targets of LXA4. Further, the knockout of IRG1 or IRAK-M abolished the beneficial effects of LXA4 on IRI-AKI. In addition, IRG1 deficiency reversed the up-regulation of IRAK-M by LXA4, while IRAK-M knockout had no impact on the IRG1 expression, indicating that IRAK-M is a downstream molecule of IRG1. Mechanistically, we found that LXA4-promoted IRG1-itaconate not only enhanced Nrf2 activation and increased HO-1 and NQO1, but also upregulated IRAK-M, which interacted with TRAF6 by competing with IRAK-1, resulting in deactivation of TLR4 downstream signal in IRI-AKI. These data suggested that LXA4 protected against IRI-AKI via promoting IRG1/Itaconate-Nrf2 and IRAK-M-TRAF6 signaling pathways, providing the rationale for a novel strategy for preventing and treating IRI-AKI.

Exploring PGE2 and LXA4 Levels in Migraine Patients: The Potential of LXA4-Based Therapies.

Neurogenic inflammation plays a significant role in the pathogenesis of migraines. This study aimed to investigate the serum levels of prostaglandin E2 (PGE2), lipoxin A4 (LXA4), and other inflammatory biomarkers (C-reactive protein, fibrinogen) in migraine patients. In total, 53 migraine patients and 53 healthy controls were evaluated. Blood serum samples were collected during both attack and interictal periods and compared with the control group. In both the attack and interictal periods, PGE2 and LXA4 values were significantly lower in migraine patients compared to the control group (p < 0.001). Additionally, PGE2 values during the attack period were significantly higher than those during the interictal period (p = 0.016). Patients experiencing migraine attacks lasting ≥ 12 h had significantly lower serum PGE2 and LXA4 levels compared to those with attacks lasting < 12 h (p = 0.028 and p = 0.009, respectively). In ROC analysis, cut-off values of 332.7 pg/mL for PGE2 and 27.2 ng/mL for LXA4 were determined with 70-80% sensitivity and specificity. In conclusion, PGE2 and LXA4 levels are significantly lower in migraine patients during both interictal and attack periods. Additionally, the levels of LXA4 and PGE2 decrease more with the prolongation of migraine attack duration. Our findings provide a basis for future treatment planning.

BML-111 inhibit H2O2-induced pyroptosis and osteogenic dysfunction of human periodontal ligament fibroblasts by activating the Nrf2/HO-1 pathway.

BACKGROUND: Periodontitis is a common and harmful chronic inflammatory oral disease, characterized by the destruction of periodontal soft and hard tissues. The NLRP3 inflammasome-related pyroptosis and human periodontal ligament fibroblasts (hPDLFs) osteogenic dysfunction are involved in its pathogenesis. Studies have shown that lipoxin A4 is an endogenous anti-inflammatory mediator and BML-111 is a lipoxin A4 analog, which was found to have potent and durable anti-inflammatory effects in inflammatory diseases, but the mechanism remains unclear. The purpose of this study was to investigate whether BML-111 inhibits H2O2-induced dysfunction of hPDLFs, attenuates inflammatory responses, and identifies the underlying mechanisms. METHODS: The oxidative stress model was established with H2O2, and the cell proliferation activity was measured by CCK-8. ALP staining and alizarin red staining were used to detect the osteogenic differentiation capacity of cells; flow cytometry and ELISA were used to detect cell pyroptosis; we explored the effect of BML-111 on hPDLFs under oxidative stress by analyzing the results of PCR and Western blotting. The Nrf2 inhibitor ML385 was added to further identify the target of BML-111 and clarify its mechanism. RESULTS: BML-111 can alleviate the impaired cell proliferation viability induced by H2O2. H2O2 treatment can induce NLRP3 inflammasome-related pyroptosis, impairing the osteogenic differentiation capacity of hPDLFs. BML-111 can effectively alleviate H2O2-induced cellular dysfunction by activating the Nrf2/HO-1 signaling pathway. CONCLUSION: The results of this study confirmed the beneficial effects of BML-111 on H2O2-induced NLRP3 inflammasome-related pyroptosis in hPDLFs, and BML-111 could effectively attenuate the impaired osteogenic differentiation function. This beneficial effect is achieved by activating the Nrf2/HO-1 signaling pathway, therefore, our results suggest that BML-111 is a potential drug for the treatment of periodontitis.

Protective Effects of Lipoxin A4 and B4 Signaling on the Inner Retina in a Mouse Model of Experimental Glaucoma.

Glaucoma is a common neurodegenerative disease characterized by progressive degeneration of retinal ganglion cells (RGCs) and the retinal nerve fiber layer (RNFL), resulting in a gradual decline of vision. A recent study by our groups indicated that the levels of lipoxins A4 (LXA4) and B4 (LXB4) in the retina and optic nerve decrease following acute injury, and that restoring their function is neuroprotective. Lipoxins are members of the specialized pro-resolving mediator (SPM) family and play key roles to mitigate and resolve chronic inflammation and tissue damage. Yet, knowledge about lipoxin neuroprotective activity remains limited. Here we investigate the in vivo efficacy of exogenous LXA4 and LXB4 administration on the inner retina in a mouse model of chronic experimental glaucoma. To investigate the contribution of LXA4 signaling we used transgenic knockout (KO) mice lacking the two mouse LXA4 receptors (Fpr2/Fpr3-/-). Functional and structural changes of inner retinal neurons were assessed longitudinally using electroretinogram (ERG) and optical coherence tomography (OCT). At the end of the experiment, retinal samples were harvested for immunohistological assessment. While both lipoxins generated protective trends, only LXB4 treatment was significant, and consistently more efficacious than LXA4 in all endpoints. Both lipoxins also appeared to dramatically reduce Müller glial reactivity following injury. In comparison, Fpr2/Fpr3 deletion significantly worsened inner retinal injury and function, consistent with an essential protective role for endogenous LXA4. Together, these results support further exploration of lipoxin signaling as a treatment for glaucomatous neurodegeneration.

Bioactive lipid-based therapeutic approach to COVID-19 and other similar infections.

COVID-19 is caused by SARS-CoV-2 infection. Epithelial and T, NK, and other immunocytes release bioactive lipids especially arachidonic acid (AA) in response to microbial infections to inactivate them and upregulate the immune system. COVID-19 (coronavirus) and other enveloped viruses including severe acute respiratory syndrome (SARS-CoV-1 of 2002-2003) and Middle East respiratory syndrome (MERS; 2012-ongoing) and hepatitis B and C (HBV and HCV) can be inactivated by AA, γ-linolenic acid (GLA, dihomo-GLA (DGLA), eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), which are precursors to several eicosanoids. Prostaglandin E1, lipoxin A4, resolvins, protectins and maresins enhance phagocytosis of macrophages and leukocytes to clear debris from the site(s) of infection and injury, enhance microbial clearance and wound healing to restore homeostasis. Bioactive lipids modulate the generation of M1 and M2 macrophages and the activity of other immunocytes. Mesenchymal and adipose tissue-derived stem cells secrete LXA4 and other bioactive lipids to bring about their beneficial actions in COVID-19. Bioactive lipids regulate vasomotor tone, inflammation, thrombosis, immune response, inactivate enveloped viruses, regulate T cell proliferation and secretion of cytokines, stem cell survival, proliferation and differentiation, and leukocyte and macrophage functions, JAK kinase activity and neutrophil extracellular traps and thus, have a critical role in COVID-19.

Lipoxin A4 Ameliorates Imiquimod-Induced Psoriasis-Like Dermatitis via Promoting the Regression of Inflammation.

Introduction: As a mediator of inflammation resolution, lipoxin A4 (LXA4) mainly plays an anti-inflammatory role and promotes inflammation resolution. LXA4 plays an inhibiting inflammatory role in a variety of diseases, tissues and cells, including keratinocytes. Psoriasis is a chronic inflammatory skin disease mediated by dysregulation of inflammation of immune cells and keratinocytes. However, the expression and role of LXA4 in psoriasis-like mouse models are still unclear. Methods: Imiquimod (IMQ) topical treatment of dorsal skin induces psoriasis-like dermatitis in BALB/c mice, pretreated intraperitoneally with or without LXA4 prior to IMQ application. Severity of dorsal lesions is assessed by using a modified human scoring system and histopathology. The concentration of LXA4 and the expression of ALOX15 (a key gene in LXA4 metabolic synthesis) in lesional skins were detected by ELISA and Western blot. Quantitative PCR and ELISA were conducted to detect the mRNA and secretion levels of inflammatory cytokines. The proportion of IL-17A-producing γδT cells in skin and skin draining cervical lymph nodes and helper (Th) 17 cells in spleens was evaluated by flow cytometry. Western blotting was used to analyze the expressions of p-STAT3 and TRAF6. Results: The concentration of LXA4 and the expression of ALOX15 were decreased in IMQ-induced lesional skin. LXA4 significantly relieved psoriasis-like lesions in IMQ-induced mouse models. Furthermore, LXA4 decreased IMQ-induced systemic inflammation, including reduced the proportion of IL-17A-producing gdT cells in skin and skin draining cervical lymph nodes and Th17 cells in spleens, the secretion and expression of CCL20, IL-17A, IL-1ß, and TNF-α in skin and serum. LXA4 markedly inhibited IMQ-induced expression of TRAF6 and p-STAT3. Conclusion: LXA4 significantly ameliorates IMQ-induced psoriasis-like inflammation, and LXA4 can be used as a target for psoriasis treatment.

Therapeutic activity of lipoxin A4 in TiO2-induced arthritis in mice: NF-κB and Nrf2 in synovial fluid leukocytes and neuronal TRPV1 mechanisms.

Background: Lipoxin A4 (LXA4) has anti-inflammatory and pro-resolutive roles in inflammation. We evaluated the effects and mechanisms of action of LXA4 in titanium dioxide (TiO2) arthritis, a model of prosthesis-induced joint inflammation and pain. Methods: Mice were stimulated with TiO2 (3mg) in the knee joint followed by LXA4 (0.1, 1, or 10ng/animal) or vehicle (ethanol 3.2% in saline) administration. Pain-like behavior, inflammation, and dosages were performed to assess the effects of LXA4 in vivo. Results: LXA4 reduced mechanical and thermal hyperalgesia, histopathological damage, edema, and recruitment of leukocytes without liver, kidney, or stomach toxicity. LXA4 reduced leukocyte migration and modulated cytokine production. These effects were explained by reduced nuclear factor kappa B (NFκB) activation in recruited macrophages. LXA4 improved antioxidant parameters [reduced glutathione (GSH) and 2,2-azino-bis 3-ethylbenzothiazoline-6-sulfonate (ABTS) levels, nuclear factor erythroid 2-related factor 2 (Nrf2) mRNA and Nrf2 protein expression], reducing reactive oxygen species (ROS) fluorescent detection induced by TiO2 in synovial fluid leukocytes. We observed an increase of lipoxin receptor (ALX/FPR2) in transient receptor potential cation channel subfamily V member 1 (TRPV1)+ DRG nociceptive neurons upon TiO2 inflammation. LXA4 reduced TiO2-induced TRPV1 mRNA expression and protein detection, as well TRPV1 co-staining with p-NFκB, indicating reduction of neuronal activation. LXA4 down-modulated neuronal activation and response to capsaicin (a TRPV1 agonist) and AITC [a transient receptor potential ankyrin 1 (TRPA1) agonist] of DRG neurons. Conclusion: LXA4 might target recruited leukocytes and primary afferent nociceptive neurons to exert analgesic and anti-inflammatory activities in a model resembling what is observed in patients with prosthesis inflammation.

Lipoxin A4 (LXA4) Reduces Alkali-Induced Corneal Inflammation and Neovascularization and Upregulates a Repair Transcriptome.

PURPOSE: To investigate the anti-inflammatory and anti-angiogenic effects of the bioactive lipid mediator LXA4 on a rat model of severe corneal alkali injury. METHODS: To induce a corneal alkali injury in the right eyes of anesthetized Sprague Dawley rats. They were injured with a Φ 4 mm filter paper disc soaked in 1 N NaOH placed on the center of the cornea. After injury, the rats were treated topically with LXA4 (65 ng/20 µL) or vehicle three times a day for 14 days. Corneal opacity, neovascularization (NV), and hyphema were recorded and evaluated in a blind manner. Pro-inflammatory cytokine expression and genes involved in cornel repair were assayed by RNA sequencing and capillary Western blot. Cornea cell infiltration and monocytes isolated from the blood were analyzed by immunofluorescence and by flow cytometry. RESULTS: Topical treatment with LXA4 for two weeks significantly reduced corneal opacity, NV, and hyphema compared to the vehicle treatment. RNA-seq and Western blot results showed that LXA4 decreased the gene and protein expression of pro-inflammatory cytokines interleukin (IL)-1ß and IL-6 and pro-angiogenic mediators matrix metalloproteinase (MMP)-9 and vascular endothelial growth factor (VEGFA). It also induces genes involved in keratinization and ErbB signaling and downregulates immune pathways to stimulate wound healing. Flow cytometry and immunohistochemistry showed significantly less infiltration of neutrophils in the corneas treated with LXA4 compared to vehicle treatment. It also revealed that LXA4 treatment increases the proportion of type 2 macrophages (M2) compared to M1 in blood-isolated monocytes. CONCLUSIONS: LXA4 decreases corneal inflammation and NV induced by a strong alkali burn. Its mechanism of action includes inhibition of inflammatory leukocyte infiltration, reduction in cytokine release, suppression of angiogenic factors, and promotion of corneal repair gene expression and macrophage polarization in blood from alkali burn corneas. LXA4 has potential as a therapeutic candidate for severe corneal chemical injuries.

Intra-Airway Treatment with Synthetic Lipoxin A4 and Resolvin E2 Mitigates Neonatal Asthma Triggered by Maternal Exposure to Environmental Particles.

Particulate matter in the air exacerbates airway inflammation (AI) in asthma; moreover, prenatal exposure to concentrated urban air particles (CAPs) and diesel exhaust particles (DEPs) predisposes the offspring to asthma and worsens the resolution of AI in response to allergens. We previously tested the hypothesis that such exposure impairs the pathways of specialized proresolving mediators that are critical for resolution and found declined Lipoxin A4 (LxA4) and Resolvin E2 (RvE2) levels in the "at-risk" pups of exposed mothers. Here, we hypothesized that supplementation with synthetic LxA4 or RvE2 via the airway can ameliorate AI after allergen exposure, which has not been tested in models with environmental toxicant triggers. BALB/c newborns with an asthma predisposition resultant from prenatal exposure to CAPs and DEPs were treated once daily for 3 days with 750 ng/mouse of LxA4 or 300 ng/mouse of RvE2 through intranasal instillation, and they were tested with the intentionally low-dose ovalbumin protocol that elicits asthma in the offspring of particle-exposed mothers but not control mothers, mimicking the enigmatic maternal transmission of asthma seen in humans. LxA4 and RvE2 ameliorated the asthma phenotype and improved AI resolution, which was seen as declining airway eosinophilia, lung tissue infiltration, and proallergic cytokine levels.

LXA4 inhibits TGF-ß1-induced airway smooth muscle cells proliferation and migration by suppressing the Smad/YAP pathway.

The aims of the present study were to examine the signaling mechanisms for transforming growth factor-ß1 (TGF-ß1)-induced rat airway smooth muscle cells (ASMCs) proliferation and migration and to determine the effect of lipoxin A4 (LXA4) on TGF-ß1-induced rat ASMCs proliferation and migration and its underlying mechanisms. TGF-ß1 upregulated transcriptional coactivator Yes-associated protein (YAP) expression by activating Smad2/3 and then upregulated cyclin D1, leading to rat ASMCs proliferation and migration. This effect was reversed after treatment with the TGF-ß1 receptor inhibitor SB431542. YAP is a critical mediator of TGF-ß1-induced ASMCs proliferation and migration. Knockdown of YAP disrupted the pro-airway remodeling function of TGF-ß1. Preincubation of rat ASMCs with LXA4 blocked TGF-ß1-induced activation of Smad2/3 and changed its downstream targets, YAP and cyclin D1, resulting in the inhibition of rat ASMCs proliferation and migration. Our study suggests that LXA4 suppresses Smad/YAP signaling to inhibit rat ASMCs proliferation and migration and therefore has potential value in the prevention and treatment of asthma by negatively modulating airway remodeling.

Lipoxin A4 methyl ester attenuated ketamine-induced neurotoxicity in SH-SY5Y cells via regulating leptin pathway.

Ketamine, the widely used intravenous anesthetic, has been reported to cause neurotoxicity and disturbs normal neurogenesis. However, the efficacy of current treatment strategies targeting ketamine's neurotoxicity remains limited. Lipoxin A4 methyl ester (LXA4 ME) is relatively stable lipoxin analog, which serves an important role in protecting against early brain injury. The purpose of this study was to investigate the protective effect of LXA4 ME on ketamine-caused cytotoxicity in SH-SY5Y cells, as well as the underlying mechanisms. Cell viability, apoptosis and endoplasmic reticulum stress (ER stress) were detected by adopting experimental techniques including CCK-8 assay, flow cytometry, western blotting and transmission electron microscope. Furthermore, examining the expression of leptin and its receptor (LepRb), we also measured the levels of activation of the leptin signaling pathway. Our results showed that LXA4 ME intervention promoted the cell viability, inhibited cell apoptosis, and reduced the expression of ER stress related protein and morphological changes induced by ketamine. In addition, inhibition of leptin signaling pathway caused by ketamine could be reversed by LXA4 ME. However, as the specific inhibitor of leptin pathway, leptin antagonist triple mutant human recombinant (leptin tA) attenuated the cytoprotective effect of LXA4 ME against ketamine-induced neurotoxicity. In conclusion, our findings demonstrated LXA4 ME could exert a neuroprotective effect on ketamine-induced neuronal injury via activation of the leptin signaling pathway.

Exogenous Lipoxin A4 attenuates IL4-induced Mucin Expression in Human Airway Epithelial Cells.

Introduction: The proinflammatory cytokine interleukin-4 (IL-4) induces mucus hypersecretion by human airway epithelial cells and the MAP kinase signalling pathway may be important in terms of IL-4-induced MUC5AC gene expression. Lipoxin A4 (LXA4) is an arachidonic acid-derived mediator that promotes inflammation by binding to the anti-inflammatory receptors (ALXs) or the formyl-peptide receptor like-1 (FPRL1) protein expressed by airway epithelial cells. Here, we explore the effects of LXA4 on IL-4-induced mucin gene expression in, and secretion from, human airway epithelial cells. Methods: We co-treated cells with IL-4 (20 ng/mL) and LXA4 (1 nM) and measured the expression levels of mRNAs encoding MUC5AC and 5B via real-time polymerase chain reaction; protein expression levels were determined by Western blotting and immunocytofluorescence. The ability of IL-4 and LXA4 to suppress protein expression was determined by Western blotting. Results: IL-4 increased MUC5AC and 5B gene and protein expression. LXA4 suppressed IL-4-induced MUC5AC and 5B gene and protein expression by interacting with the IL4 receptor and mitogen-activated protein kinase (MAPK) pathway, including both phospho-p38 MAPK and phospho-extracellular signal-regulated kinase (phospho-ERK). IL-4 and LXA4 increased and decreased, respectively, the number of cells that stained with anti-MUC5AC and 5B antibodies. Conclusions: LXA4 may regulate mucus hypersecretion induced by IL4 in human airway epithelial cells.

Cathelicidin promotes liver repair after acetaminophen-induced liver injury in mice.

Background & Aims: Acetaminophen (APAP)-induced acute liver injury (AILI) is a leading cause of acute liver failure (ALF). N-acetylcysteine (NAC) is only effective within 24 h after APAP intoxication, raising an urgent need for alternative approaches to treat this disease. This study aimed to test whether cathelicidin (Camp), which is a protective factor in chronic liver diseases, protects mice against APAP-induced liver injury and ALF. Methods: A clinically relevant AILI model and an APAP-induced ALF model were generated in mice. Genetic and pharmacological approaches were used to interfere with the levels of cathelicidin in vivo. Results: An increase in hepatic pro-CRAMP/CRAMP (the precursor and mature forms of mouse cathelicidin) was observed in APAP-intoxicated mice. Upregulated cathelicidin was derived from liver-infiltrating neutrophils. Compared with wild-type littermates, Camp knockout had no effect on hepatic injury but dampened hepatic repair in AILI and reduced survival in APAP-induced ALF. CRAMP administration reversed impaired liver recovery observed in APAP-challenged Camp knockout mice. Delayed CRAMP, CRAMP(1-39) (the extended form of CRAMP), or LL-37 (the mature form of human cathelicidin) treatment exhibited a therapeutic benefit for AILI. Co-treatment of cathelicidin and NAC in AILI displayed a stronger hepatoprotective effect than NAC alone. A similar additive effect of CRAMP(1-39)/LL-37 and NAC was observed in APAP-induced ALF. The pro-reparative role of cathelicidin in the APAP-damaged liver was attributed to an accelerated resolution of inflammation at the onset of liver repair, possibly through enhanced neutrophil phagocytosis of necrotic cell debris in an autocrine manner. Conclusions: Cathelicidin reduces APAP-induced liver injury and ALF in mice by promoting liver recovery via facilitating inflammation resolution, suggesting a therapeutic potential for late-presenting patients with AILI with or without ALF. Impact and implications: Acetaminophen-induced acute liver injury is a leading cause of acute liver failure. The efficacy of N-acetylcysteine, the only clinically approved drug against acetaminophen-induced acute liver injury, is significantly reduced for late-presenting patients. We found that cathelicidin exhibits a great therapeutic potential in mice with acetaminophen-induced liver injury or acute liver failure, which makes up for the limitation of N-acetylcysteine therapy by specifically promoting liver repair after acetaminophen intoxication. The pro-reparative role of cathelicidin, as a key effector molecule of neutrophils, in the APAP-injured liver is attributed to an accelerated resolution of inflammation at the onset of liver repair, possibly through enhanced phagocytic function of neutrophils in an autocrine manner.