Phosphorylation Of P38 MAPK Research Articles

Blocking the p38 MAPK Signaling Pathway in the Rat Hippocampus Alleviates the Depressive-like Behavior Induced by Spinal Cord Injury.

Patients with spinal cord injury (SCI) may develop depression, which can affect their rehabilitation. However, the underlying mechanism of depression in SCI patients remains unclear. Previous studies have revealed increased p38 MAPK phosphorylation in the rat hippocampus after SCI, accompanied by depression-like behaviors. However, the role of the p38 MAPK signaling pathway in SCI-induced depression remains unclear. In this study, we used an aneurysm clip-induced rat SCI model to investigate whether p38 MAPK phosphorylation in the hippocampus is associated with depression-like behaviors in rats after SCI. Behavioral testing revealed that SB203580, a p38 MAPK signaling inhibitor, reduced depression-like behaviors. Western blotting and morphological analyses showed that SB203580 inhibited the activation of microglia and astrocytes in the hippocampus after SCI. Additionally, SB203580 reduced the expression of tumor necrosis factor α and increased p38 MAPK phosphorylation and the number of bromodeoxyuridine-positive cells in the hippocampus. These findings suggest that SB203580 can inhibit hippocampal remodeling and the neuroimmune response in the rat hippocampus after SCI. Therefore, the phosphorylation of p38 MAPK in the hippocampus plays a key role in the depression-like behaviors induced by SCI. The inhibition of p38 MAPK phosphorylation may represent a mechanism to protect against hippocampal injury induced by SCI.

In vitro gastrointestinal digestion simulation screening of novel ACEI peptides from broccoli: mechanism in high glucose-induced VSMCs dysfunction

Many natural angiotensin-converting enzyme inhibitory (ACEI) peptides have been widely studied. However, their stability in vivo is poor in most cases. In this study, peptides were initially digested from broccoli in vitro, and absorption was simulated by Caco2 cells transport and then analyzed by Peptideomics and molecular docking. Subsequently, the mechanisms were verified using a high glucose-induced vascular smooth muscle cells (VSMCs) dysfunction model. Results showed that ACEI activity of broccoli crude peptide increased by 70.73 ± 1.42% after digestion. The enzymatic hydrolysates of crude broccoli peptides before and after digestion were detected by HPLC. The digested crude peptides were highly stable (with a stability level > 90%) in the intestine and possessed a strong absorptive potential. Five peptides with high stability and strong permeability were first identified, including HLEVR, LTEVR, LEHGF, HLVNK, and LLDGR, which exhibited high activity with IC50 values of 3.19 ± 0.23 mM, 17.07 ± 1.37 mM, 0.64 ± 0.02 mM, 0.06 ± 0.01 mM, and 2.81 ± 0.12 mM, respectively. When the VSMCs model was exposed to Ang II, the expressions of PCNA, MMP2, and Bcl2 were increased, while the expression of BAX was inhibited. When the VSMCs was exposed to high glucose (HG), the Ang II concentration significantly increased. This indicates that HG elevated Ang II levels. Finally, five peptides significantly attenuated Ang II-induced VSMCs proliferation and migration by down-regulating AT1R expression and inhibiting ERK and p38 MAPK phosphorylation. Notably, in exploring VSMCs dysfunction on a high glucose-induced model, ACEI peptides resulted in down-regulation of ACE and up-regulation of ACE2 expression. Therefore, it can be further referenced for the functional food against hypertension and cardiovascular diseases.

The Potential Role of Sanguinarine as an Inhibitor of Leishmania PP2C in the Induction of Apoptosis

Leishmania spp. cause a wide range of human diseases, localized skin lesions, mucocutaneous and visceral infections. In the present study, the aim was to investigate the potential role of sanguinarine as a specific inhibitor of Leishmania PP2C that can induce apoptosis in the parasite. The results demonstrated that sanguinarine inhibits, in a dose-dependent mode at 72 h, the growth and phosphatase activity of both Leishmania major and Leishmania mexicana promastigotes. Therefore, all assays were performed from this time period onwards. TUNEL assay was used to identify apoptosis and indicated apoptosis in L. major and L. mexicana promastigotes. Similarly, Western blot assay showed that PARP, a DNA damage indicator molecule, was present in L. major and L. mexicana promastigotes incubated with the inhibitor. In addition, differential expression of the proapoptotic protein Bax and the antiapoptotic protein Bcl-2 was observed in both Leishmania species. Finally, the protein phosphatase PP2C expression was not affected, whereas p38 MAPK phosphorylation was increased in L. major promastigotes than in L. mexicana promastigotes. Therefore, sanguinarine proved to be an inhibitor of the growth and PP2C enzymatic activity of L. major and L. mexicana promastigotes, and with it, this inhibition induced apoptosis.

Glabridin Suppresses Macrophage Activation by Lipoteichoic Acid In Vitro: The Crucial Role of MAPKs-IL-1β-iNOS Axis Signals in Peritoneal and Alveolar Macrophages

Inflammation, a fundamental response to infection and injury, involves interactions among immune cells and signaling molecules. Dysregulated inflammation contributes to diseases such as autoimmune disorders and cancer. Interleukin-1 beta (IL-1β), produced by macrophages in response to lipoteichoic acid (LTA) from Gram-positive bacteria, is a key inflammatory mediator. Glabridin (GBD), a bioactive compound from licorice root, exhibits anti-inflammatory properties. This study investigates GBD’s effects on LTA-induced proinflammatory signaling in RAW 264.7 murine macrophages and alveolar macrophages, MH-S, focusing on IL-1β expression and signaling pathways. Cell viability assays confirmed that 20 μM GBD was non-cytotoxic. Confocal microscopy and quantitative PCR showed that GBD significantly reduced IL-1β fluorescence intensity, mRNA, and protein levels. GBD also inhibited inducible nitric oxide synthase (iNOS) expression and nitric oxide (NO) production. Further analysis revealed that GBD suppressed NF-κB p65 nuclear translocation and selectively modulated MAPK pathway activation by reducing JNK and p38 MAPK phosphorylation without affecting ERK. Studies using specific inhibitors demonstrated that IL-1β production reduction was mechanistically linked to MAPK pathway inhibition. These findings highlight GBD’s potential as a therapeutic agent for inflammatory diseases through its ability to modulate critical inflammatory mediators and signaling pathways.

Adipose-derived stem cells promote the recovery of intestinal barrier function by inhibiting the p38 MAPK signaling pathway.

Intestinal barrier damage causes an imbalance in the intestinal flora and microbial environment, promoting a variety of gastrointestinal diseases. This study aimed to explore the mechanism by which adipose-derived stem cells (ADSCs) repair intestinal barrier damage. The human colon adenocarcinoma cell line Caco-2 and rats were treated with lipopolysaccharide (LPS) to establish in vitro and in vivo models, respectively, of intestinal barrier damage. The expression of inflammatory cytokines (TNF-α, HMGB1, IL-1β and IL-6), antioxidant enzymes (iNOS, SOD and CAT), and oxidative products (MDA and 8-iso-PGF2α) was detected using ELISA kits and related reagent kits. Apoptosis-related proteins (Bcl-2, Bax, Caspase-3 and Caspase-9), tight junction proteins (ZO-1, Occludin, E-cadherin, and Claudin-1) and p38 MAPK pathway-associated protein were detected by Western blotting. In addition, cell viability and apoptosis was determined by a CCK-8 kit and flow cytometry, respectively. Cell permeability was assayed by the transepithelial electrical resistance value and FITC-dextran concentration. The homing effect of ADSCs was detected by fluorescence labeling, and intestinal barrier tissue was observed by HE staining. After ADSC treatment, the level of phosphorylated p38 MAPK protein decreased, the expression of inflammatory factors, oxidative stress and cell apoptosis decreased, the expression of tight junction proteins increased, and cell permeability decreased in Caco-2 cells stimulated with LPS. In rats, ADSCs are directionally recruited to damaged intestinal tissue. ADSCs significantly decreased the levels of D-lactate, diamine oxidase (DAO) and FITC-dextran induced by LPS. ADSCs promoted tight junction proteins and inhibited oxidative stress in intestinal tissue. These effects were reversed after the use of a p38 MAPK activator. ADSCs can be directionally recruited to intestinal tissue, upregulate tight junction proteins, and reduce apoptosis and oxidative stress by inhibiting the p38MAPK signaling pathway. This study provides novel insights into the treatment of intestinal injury.

Deep-Sea-Derived Isobisvertinol Targets TLR4 to Exhibit Neuroprotective Activity via Anti-Inflammatory and Ferroptosis-Inhibitory Effects.

Neuroinflammation and neuronal cell death are leading causes of death in the elderly and underlie various neurodegenerative diseases. These diseases involve complex pathophysiological mechanisms, including inflammatory responses, oxidative stress, and ferroptosis. Compounds derived from deep-sea fungi exhibit low toxicity and potent neuroprotective effects, offering a promising source for drug development. In this study, we isolated 44 natural products from deep-sea-derived fungi and identified isobisvertinol (17) as a compound with anti-inflammatory and ferroptosis-inhibiting effects. Using LPS-induced microglial inflammation and RSL3-induced neuronal ferroptosis models, we found that 17 targets TLR4 to provide neuroprotection. Molecular docking studies revealed that 17 inhibits TLR4 activation by occupying the hydrophobic pocket at the TLR4-MD2 binding site. Additionally, 17 suppresses TLR4, reducing p38 MAPK phosphorylation, and inhibits ferroptosis by decreasing lipid peroxidation and modulating mitochondrial membrane potential. Metabolomic analysis showed that 17 rescues alterations in multiple metabolic pathways induced by RSL3 and increases levels of antioxidant metabolites, including glutamine, glutamate, and glutathione. In summary, our results indicate that isobisvertinol (17) targets TLR4 in neural cells to reduce inflammation and inhibit p38 MAPK phosphorylation, while regulating metabolic pathways, mainly GSH synthesis, to provide antioxidant effects and prevent ferroptosis in neurons.

Targeting p38 MAPK signaling pathway and neutrophil extracellular traps: An important anti-inflammatory mechanism of Huangqin Qingre Chubi Capsule in rheumatoid arthritis.

Rheumatoid arthritis (RA) is a common chronic autoimmune disease. Neutrophils release and their extracellular traps (NETs) tend to result in synovial inflammation and cartilage damage. Huangqin Qingre Chubi Capsule (HQC) is an important herbal formulation for RA treatment and has been used for many years. This study applied an interdisciplinary and integrated strategy combining clinical data, integrative bioinformatics, molecular modeling, and both in vitro and in vivo experiments to elucidate the efficacy and potential anti-inflammatory mechanisms of HQC for RA. Retrospective clinical data mining demonstrated that patients with RA treated with HQC exhibited recovery of immuno-inflammatory markers and self-perception. By applying network pharmacological analysis, molecular docking, molecular dynamics simulation, and cellular thermal shift assays, we determined that HQC can directly bind to MAPK14 and target p38 MAPK signaling pathway and NETs formation. Treatment of an adjuvant-induced arthritis rat model combining damp-heat patterns using HQC demonstrated it's effectiveness in reducing the severity of inflammatory arthritis in a dose-dependent manner and downregulated phosphorylation of p38 MAPK and NETs release. In vitro co-culture system mimicking inflammatory microenvironment of RA in vivo revealed that crosstalk between neutrophils (PMNs) and fibroblast-like synoviocytes (FLSs) was manifested by activation of p38 MAPK signaling pathway, release of NETs, and amplification of inflammation, which was blocked by HQC. Mechanistic validation using the p38 MAPK agonist diprovocim demonstrated that HQC inhibited NET formation by modulating the p38 MAPK pathway (mainly by inhibiting its phosphorylation) to exert anti-inflammatory effects. In conclusion, our interdisciplinary and integrated study demonstrated that HQC can target the p38 MAPK signaling pathway to inhibit NET formation and inflammatory response, thereby blocking the crosstalk between PMNs and FLSs to ameliorating RA.

Postoperative Tongqi Formula ameliorates postoperative ileus via p38 MAPK signaling pathway and metabolic disorder.

This study investigated the mechanism by which the Postoperative Tongqi Formula (PTQF) treats postoperative ileus (POI) through regulation of the p38 MAPK signaling pathway, Zona occludens 1 (ZO-1) protein, and metabolism. The primary components of PTQF were characterized using UHPLC-Q-TOF-MS/MS. The identified compounds subsequently employed network pharmacology to predict the signaling pathways associated with the inflammatory phase of POI. The anti-inflammatory effects of PTQF were evaluated in vitro using RAW264.7cells. A rat model of POI was used to assess efficacy based on the spleen index and charcoal powder propulsion rat in the small intestine. Furthermore, pathological damage to the small intestine was analyzed using hematoxylin and eosin (HE) staining as well as immunofluorescence to evaluate ZO-1 protein expression. Inflammatory cytokine levels were quantified using enzyme-linked immunosorbent assay (ELISA). Subsequently, Western blot analysis was performed to examine the p38 MAPK signaling pathway. Finally, a metabolomics approach was employed to analyze serum samples to identify potential metabolic pathways. A total of 130 chemical constituents were identified in PTQF. Following the network pharmacology analysis of these compounds, the p38 MAPK signaling pathway was chosen for further investigation. In vitro, PTQF effectively inhibited inflammatory responses in RAW264.7cells. Results from the spleen index and charcoal powder propulsion rate indicated that PTQF alleviated the inflammatory phase of POI in rats by mitigating systemic and intestinal inflammation. This was supported by reduced levels of inflammatory factors, modulation of ZO-1 protein expression, and a decrease in p38 MAPK phosphorylation levels. Furthermore, serum metabolomics revealed nine differential metabolites linked to intestinal inflammation. PTQF mitigates inflammation and intestinal damage in POI rats by modulating inflammatory factors, ZO-1 protein expression, the p38 MAPK signaling pathway, and metabolic disturbances.

Bushen Daozhuo Granules Alleviate Chronic Non-Bacterial Prostatitis in Rats through p38 MAPK and Akt Signaling Pathways Based on Tandem Mass Tag-Based Quantitative Proteomics and Network Pharmacology Analyses.

The traditional Chinese medicine formula, Bushen Daozhuo Granules (BSDZG), is used to treat chronic non-bacterial prostatitis (CNP) clinically. However, its mechanism of action is unclear. The aim of our study was to determine the effect of BSDZG on CNP and its underlying mechanisms. Male Wistar rats were randomly assigned to control, CNP, and BSDZG groups. CNP was induced using purified prostaglandin solution and Freund's complete adjuvant, after which the BSDZG group received 1.54 g/kg/d of BSDZG for 30 days. Prostate tissues were used to determine apoptosis and inflammatory cytokines. The herb-composition-target network and functional signaling pathways were built using a network pharmacology approach, which was also confirmed in vivo. Treatment with BSDZG significantly alleviated the histopathological lesions, inflammation, and apoptosis in the prostate of CNP rats. The herb-composition-target network comprising 42 active compounds and 32 targets of 11 herbs was illustrated, and KEGG pathways analysis identified the Akt and MAPK pathways as related to the effects of BSDZG. Phosphorylation of p38 MAPK, NF-кB, and Bax expression was significantly enhanced and phosphorylated Akt and Bcl-2 levels were decreased in CNP rats, which could be reversed by BSDZG. This study presented for the first time that BSDZG effectively alleviated CNP symptoms in rats and elucidated the underlying mechanisms mediated by the Akt and MAPK pathways, providing the theoretical basis for the clinical use and promotion of BSDZG.

Sargassum plagiophyllum Ethanolic Extract Enhances Wound Healing by Modulating FAK/Src/Akt/p38 and Rac1 Signaling in Keratinocytes HaCaT Cells

Recently, seaweed extracts have been found to have potential in skin benefits. This study, therefore, aimed to explore phytochemical analysis, antimicrobial, antioxidant, and wound healing properties of brown seaweed Sargassum plagiophyllym ethanolic extract (SPEE) on human skin keratinocyte HaCaT cells and the possible mechanism involved. Our results indicated that SPEE contained flavonoid, phenolic, and carotenoid as the major active constituents. The HPLC chromatogram revealed C‐phycocyanin and fucoidan presented in SPEE. SPEE demonstrated the antioxidant capability and significantly reduced wound space at 24 and 48 h in wound‐healing assay. Treatment with SPEE (50 and 100 μg/mL) increased FAK and Src phosphorylation in western blotting. Moreover, SPEE also upregulated Akt and p38 MAPK phosphorylation but not ERK1/2. SPEE increased Rac1 protein expression. Interestingly, hyaluronan synthase (HAS1 and HAS2) as well as collagen type I and elastin were also significantly upregulated when compared with the control upon exposure to SPEE. In conclusion, our data suggested that SPEE promotes cutaneous wound healing by regulating FAK/Src‐mediated Akt, p38 MAPK, and Rac1 signaling. These findings suggest the potential use of SPEE for skin wound treatment.

MAPK pathways regulated apoptosis and pyroptosis in respiratory epithelial cells of a primitive vertebrate model during bacterial infection.

Respiratory diseases caused by bacterial and viral infection have seriously affected human health. The invaginated lung structure in mammals caused difficulties in relevant research, here we evaluated the regulatory roles of MAPK pathways in apoptosis and pyroptosis during bacterial infection in an evaginated respiratory organ model for the first time. F. columnare was adopted for bacterial infection in rainbow trout in vivo and RTgill-W1 cells in vitro. Infected trout gills were separated for histological analysis, transcriptomic sequencing, TUNEL, RT-qPCR and enzyme activity assay. RTgill-W1 cells were treated with different inhibitors of MAPK pathway for evaluating apoptosis and pyroptosis. Bacterial infection induced serious histological changes and apoptosis in trout gill, accompanied with p38MAPK/ JNK pathway activation, while pyroptosis were induced after secondary infection along with ERK pathway activation. In vitro study confirmed pro-apoptotic roles of bacterial infection, accompanied with the increased phosphorylation of p38 MAPK and JNK. Moreover, p38 MAPK inhibition significantly decreased the F. columnare infection-induced apoptosis of RTgill-W1 cell via affecting Bcl2 protein expression and mitochondrial membrane potential. Therefore, our study indicated that MAPK pathways regulated apoptosis and pyroptosis in teleost respiratory organ during bacterial infection, which will benefit developing strategies in fighting against bacterial disease in aquaculture practice.

Tetrabromobisphenol A induced p38-MAPK/AMPKα activation downstream-triggered CHOP signal contributing to neuronal apoptosis and death

Tetrabromobisphenol A (TBBPA), a brominated flame retardant (BFR), has been implicated as the neurotoxic effects in mammalian. However, the exact mechanisms underlying TBBPA-induced neurotoxicity remain unclear. In the present study, Neuro-2a cells, a mouse neural crest-derived cell line, were used to examine the mechanism of TBBPA-induced neuronal cytotoxicity. TBBPA exposure caused alterations in cell viability and mitochondrial membrane potential (MMP) and induction of apoptotic events, such as increased apoptotic cell population and cleaved caspase-3, -7, -9, and poly (ADP-ribose) polymerase (PARP) protein expression). TBBPA exposure triggered CCAAT/enhancer-binding protein (C/EBP) homologous protein (CHOP) activation. Transfection with CHOP-specific small interfering RNA (siRNA) obviously prevented the expression of CHOP protein and markedly attenuated MMP loss, and caspase-3 and -7 activation in TBBPA-exposed Neuro-2a cells. In addition, TBBPA exposure significantly evoked the phosphorylation of c-Jun N-terminal kinase (JNK), extracellular-signal regulated kinase1/2 (ERK1/2), p38-mitogen-activated protein kinase (p38-MAPK), and AMP-activated protein kinase (AMPK)α proteins. Pretreatment of cells with pharmacological inhibitors of p38-MAPK (SB203580) and AMPK (compound C), but not inhibitors of JNK (SP600125) or ERK1/2 (PD98059), effectively prevented the increase in caspase-3 activity, MMP loss, and activated CHOP and cleaved caspase-3 and -7 protein expression in TBBPA-treated cells. Notably, transfection with either p38α-MAPK- or AMPKα1/2-specific siRNAs markedly attenuated the expression of CHOP, and cleaved caspase-3 and -7. Interestingly, transfection with each siRNA significantly reduced the TBBPA-induced phosphorylation of p38-MAPK and AMPKα proteins. Collectively, these findings suggest that CHOP activation-mediated mitochondria-dependent apoptosis contributes to TBBPA-induced neurotoxicity. An interdependent p38-MAPK and AMPKα signaling-regulated apoptotic pathway may provide new insights into the mechanism understanding TBBPA-elicited neurotoxicity.

Lactoferrin targeting INTL1 receptor inhibits hepatocellular carcinoma progression via apoptosis and cell cycle signaling pathways

Hepatocellular carcinoma (HCC) constitutes 90% of liver cancer cases and ranks as the third leading cause of cancer-related mortality, necessitating urgent development of alternative therapies. Lactoferrin (LF), a natural iron-binding glycoprotein with reported anticancer effects, is investigated for its potential in liver cancer treatment, an area with limited existing studies. This study focuses on evaluating LF’s anti-liver cancer effects on HCC cells and assessing the preventive efficacy of oral LF administration in a murine model. Data showed that LF exerted anti-proliferative effects on HepG2, Hep3B, and SK-Hep1 cells while having no cytotoxicity on healthy liver cells (FL83B). Mechanistically, LF induces mitochondrial-mediated apoptosis and G0/G1 cell cycle arrest in HepG2 cells, associated with increased phosphorylation of p38 MAPK and JNK for apoptosis, and ERK phosphorylation for cell cycle arrest. Intelectin-1 (INTL1) is identified as the receptor facilitating LF endocytosis in HepG2 cells, and downregulation of INTL1 inhibits LF-induced signaling pathways. Notably, oral LF administration prevents HCC development in nude mice with orthotopic HepG2 cell injection. This study unveils the mechanistic basis of LF action in HepG2 cells, showcasing its potential in HCC prevention. Importantly, we report the novel identification of INTL1 as the LF receptor in HepG2 cells, providing valuable insights for future exploration of LF and its derivatives in liver cancer therapy.

HMAGEA2 as a potential diagnostic and therapeutic target for melanoma progression and metastasis.

The incidence of melanoma, a highly aggressive skin cancer, continues to increase worldwide, particularly among populations with lighter skin tones. The diagnostic challenge of melanoma lies in the absence of a distinctive clinical presentation, as its characteristics vary based on anatomical location, growth type, and histopathology. The melanoma-associated antigen (MAGE) gene family is differentially expressed in various human cancers, including melanoma. In this study, we explored the association between human MAGEA2 (hMAGEA2) expression and melanoma. Using a human melanoma tissue array, we confirmed that hMAGEA2 expression was higher in melanoma and metastatic melanoma than in normal tissues. Additionally, we used SK-MEL-5 and SK-MEL-28 cell lines to investigate the cellular and molecular mechanisms underlying melanoma progression and invasiveness. In SK-MEL-5 and SK-MEL-28 cells, hMAGEA2 overexpression accelerated cell proliferation. Conversely, the knockdown of hMAEGA2 reduced cell proliferation, colony formation, and migration significantly and induced arrest at the G2/M phase of the cell cycle. With respect to the molecular mechanism, the knockdown of hMAGEA2 decreased the phosphorylation of Akt, JNK, and p38 MAPK. Additionally, hMAGEA2 knockdown reduced tumor formation significantly at the in vivo level. Collectively, the robust correlation between hMAGEA2 and melanoma metastasis supports the potential utility of hMAGEA2 as both a diagnostic marker and novel therapeutic target for patients with melanoma metastasis.

G(1-5)-EM2, a multi-targeted agonist to opioid and growth hormone secretagogue receptors exhibited nontolerance forming antinociceptive effects in a mouse model of burn pain

Burn induced-pain (BIP) is one of the most common pain symptoms, which seriously affects the quality of sufferer life. Researches show that multi-targeted drug therapies offer superior efficacy and fewer side effects compared to single-target drug therapies. Consequently, in this study, we developed G(1-5)-EM2, a multi-targeted peptide designed to target μ-opioid receptor and the growth hormone secretagogue receptor 1α (GHS-R1α), and explored its antinociceptive effects on burn injury pain. Calcium mobilization experiments revealed that G(1-5)-EM2 demonstrated weak multi-agonist activities to μ-opioid receptor and κ-opioid receptor as well as GHS-R1α in vitro. Near-infrared fluorescence imaging experiments demonstrated that G(1-5)-EM2 could penetrate the blood-brain barrier (BBB) and access the brain following intravenous injection. The enzymatic stability of G(1-5)-EM2 was significantly enhanced compared to EM2. Our results indicated that intrathecal administration of G(1-5)-EM2 mitigated mechanical allodynia and thermal hyperalgesia in BIP. These antinociceptive effects of G(1-5)-EM2 were partially mediated through μ-opioid receptor and GHS-R1α. Moreover, intrathecal administration of G(1-5)-EM2 significantly decreased burn-induced up-regulation of phosphorylated p38 MAPK, phosphorylated NF-κBp65 and TRPV1 in the ipsilateral spinal cord, reduced the levels of IL-1β, IL-6 and TNF-α in serum, and enhanced wound healing in burned skin. Repeated intrathecal administration of G(1-5)-EM2 produced a non-tolerance-forming antinociception in BIP. These results suggest that the multi-targeted peptide G(1-5)-EM2 exhibits a novel role in alleviating BIP with fewer side effects and may represent a promising strategy for developing new analgesic drugs.

Maresin-1 Ameliorates Sepsis-Induced Microglial Activation Through Modulation of the P38 MAPK Pathway.

Sepsis is a life-threatening disease characterized by a dysregulated immune response to infection, often leading to neuroinflammation. As a known immunomodulator, Maresin-1 (MaR1) may have potential applications in the treatment of sepsis-induced neuroinflammation, but its effects in this context are unknown. We used a mouse cecum ligation and puncture (CLP)-induced sepsis model and an in vitro lipopolysaccharide (LPS)-induced neuroinflammatory model of BV2 microglia. Expression of microglial cell markers (IBA1, CD11B, CD68, CD86 and CD206) and pro-inflammatory markers (iNOS and COX2) was assessed. The role of MaR1 in regulating the P38 MAPK pathway was explored using the P38 MAPK inhibitor SB203580. In the CLP model, an increased proportion of M1-type microglia was observed, and MaR1 was able to reverse it. However, the combination of SB203580 and MaR1 did not enhance the therapeutic effect compared to SB20580 alone. In vitro experiments, MaR1 inhibited LPS-induced P38 MAPK nuclear translocation and decreased the expression of pro-inflammatory markers such as iNOS and COX2. As with the animal results, no stacking effect could be obtained with the co-administration of SB203580 and MaR1. Our findings suggest that MaR1 attenuates sepsis-induced neuroinflammation mainly by inhibiting phosphorylation of P38 MAPK in microglial cells. This suggests that MaR1 may have a potential therapeutic role in the treatment of sepsis neuroinflammation.

Reversing metabolic reprogramming by CPT1 inhibition with etomoxir promotes cardiomyocyte proliferation and heart regeneration via DUSP1 ADP-ribosylation-mediated p38 MAPK phosphorylation

The neonatal mammalian heart has a remarkable regenerative capacity, while the adult heart has difficulty to regenerate. A metabolic reprogramming from glycolysis to fatty acid oxidation occurs along with the loss of cardiomyocyte proliferative capacity shortly after birth. In this study, we sought to determine if and how metabolic reprogramming regulates cardiomyocyte proliferation. Reversing metabolic reprogramming by carnitine palmitoyltransferase 1 (CPT1) inhibition, using cardiac-specific Cpt1a and Cpt1b knockout mice promoted cardiomyocyte proliferation and improved cardiac function post-myocardial infarction. The inhibition of CPT1 is of pharmacological significance because those protective effects were replicated by etomoxir, a CPT1 inhibitor. CPT1 inhibition, by decreasing poly(ADP-ribose) polymerase 1 expression, reduced ADP-ribosylation of dual-specificity phosphatase 1 in cardiomyocytes, leading to decreased p38 MAPK phosphorylation, and stimulation of cardiomyocyte proliferation. Our present study indicates that reversing metabolic reprogramming is an effective strategy to stimulate adult cardiomyocyte proliferation. CPT1 is a potential therapeutic target for promoting heart regeneration and myocardial infarction treatment.

Protective effect of 6'-Sialyllactose on LPS-induced macrophage inflammation via regulating Nrf2-mediated oxidative stress and inflammatory signaling pathways.

Macrophages play a central role in cardiovascular diseases, like atherosclerosis, by accumulating in vessel walls and inducing sustained local inflammation marked by the release of chemokines, cytokines, and matrix-degrading enzymes. Recent studies indicate that 6'-sialyllactose (6'-SL) may mitigate inflammation by modulating the immune system. Here, we examined the impact of 6'-SL on lipopolysaccharide (LPS)-induced acute inflammation using RAW 264.7 cells and a mouse model. In vivo, ICR mice received pretreatment with 100 mg/kg 6'-SL for 2 h, followed by intraperitoneal LPS injection (10 mg/kg) for 6 h. In vitro, RAW 264.7 cells were preincubated with 6'-SL before LPS stimulation. Mechanistic insights were gained though Western blotting, qRT-PCR, and immunofluorescence analysis, while reactive oxygen species (ROS) production was assessed via DHE assay. 6'-SL effectively attenuated LPS-induced p38 MAPK and Akt phosphorylation, as well as p65 nuclear translocation. Additionally, 6'-SL inhibited LPS-induced expression of tissue damage marker MMP9, IL-1β, and MCP-1 by modulating NF-κB activation. It also reduced ROS levels, mediated by p38 MAPK and Akt pathways. Moreover, 6'-SL restored LPS-suppressed Nrf2 and HO-1 akin to specific inhibitors SB203580 and LY294002. Consistent with in vitro results, 6'-SL decreased oxidative stress, MMP9, and MCP-1 expression in mouse endothelium following LPS-induced macrophage activation. In summary, our findings suggest that 6'-SL holds promise in mitigating atherosclerosis by dampening LPS-induced acute macrophage inflammation.

DLK1 and DLK2, two non-canonical ligands of NOTCH receptors, differentially modulate the osteogenic differentiation of mesenchymal C3H10T1/2 cells

BackgroundC3H10T1/2 is a mesenchymal cell line capable of differentiating into osteoblasts, adipocytes and chondrocytes. The differentiation of these cells into osteoblasts is modulated by various transcription factors, such as RUNX2. Additionally, several interconnected signaling pathways, including the NOTCH pathway, play a crucial role in modulating their differentiation into mature bone cells. We have investigated the roles of DLK1 and DLK2, two non-canonical inhibitory ligands of NOTCH receptors, in the osteogenic differentiation of C3H10T1/2 cells.ResultsOur results corroborate existing evidence that DLK1 acts as an inhibitor of osteogenesis. In contrast, we demonstrate for the first time that DLK2 enhances this differentiation process. Additionally, our data suggest that NOTCH2, 3 and 4 receptors may promote osteogenesis, as indicated by their increased expression during this process, whereas NOTCH1 expression, which decreases during cell differentiation, might inhibit osteogenesis. Moreover, treatment with DAPT, a NOTCH signaling inhibitor, impeded osteogenic differentiation. We have confirmed the increase in ERK1/2 MAPK and p38 MAPK phosphorylation in C3H10T1/2 cells induced to differentiate to osteoblasts. Our new findings reveal increased ERK1/2 MAPK phosphorylation in differentiated C3H10T1/2 cells with a decrease in DLK1 expression or an overexpression of DLK2, which is coincident with the behavior of those transfectants where we have detected an increase in osteogenic differentiation. Additionally, p38 MAPK phosphorylation increases in differentiated C3H10T1/2 cells with reduced DLK1 levels.ConclusionsOur results suggest that DLK1 may inhibit osteogenesis, while DLK2 may promote it, by modulating NOTCH signaling and the phosphorylation of ERK1/2 and p38 MAPK pathways. Given the established inhibitory effect of DLK proteins on NOTCH signaling, these new insights could pave the way for developing future therapeutic strategies aimed at treating bone diseases.

Novel role of human epicardial adipocyte-derived SPARCL1 in suppression of postoperative atrial fibrillation in patients undergoing cardiovascular surgery

Abstract Background Postoperative atrial fibrillation (POAF) occurs in 20-50% after cardiovascular surgery and is associated with the short and long-term morbidity/mortality. The epicardial adipose tissue (EAT) may have a potential role to regulate the incidence of POAF. Purpose We explored the possible role of human epicardial adipocyte-derived adipokines in the regulation of the myocardial redox state and POAF. Methods We prospectively evaluated 149 patients without known AF who underwent scheduled open-heart cardiovascular surgery between May 2020 and November 2022. They were divided into POAF or Non-POAF group and followed up after discharge (476.6 ± 290.0 days). POAF was defined as AF lasting more than 30 seconds after the surgery. Human EAT samples were obtained from these patients during surgery and the preadipocytes were isolated. Preadipocytes were terminally differentiated to mature adipocytes and mRNAs were extracted. Genome-wide expression profiling of 6 vs. 6 matched samples of the POAF and Non-POAF groups was performed using Microarray analysis, and the results were validated with qPCR in all the cohort samples. The effects of secreted adipokines on the cardiomyocytes were assessed in terms of oxidative stress. Superoxide and whole reactive oxygen species (ROS) were evaluated with luminometry and live-cell fluorescent probes respectively. As the oxidative stress associated signalling, the phosphorylation of ERK and p38-MAPK were evaluated with western blotting. Angiotensin II (Ang-II) was used to induce oxidative stress and phosphorylation of MAPK. Results POAF was observed in 53 patients (36%). Microarray analysis (n = 6) revealed that there were 132 down- or up-regulated cording genes in epicardial adipocytes of the POAF group compared to the Non-POAF group. Of these genes, 11 genes cording secretable molecules were validated by qPCR (n = 69), showing that only SPARCL1 had significantly downregulated in the POAF group compared to the Non-POAF group (P = 0.005). Superoxide and ROS were induced by Ang II (P < 0.01) and attenuated by SPARCL1 dose-dependent manner (P < 0.01) in neonatal rat cardiomyocytes. The phosphorylation of ERK and p38-MAPK were induced by Ang II (P < 0.01) and attenuated by SPARCL1 (P < 0.01) dose-dependently. Co-culture of human induced pluripotent stem cell-derived atrial cardiomyocytes (iPS-ACM) and human epicardial adipocytes revealed that the adipocytes derived from Non-POAF group suppressed the superoxide (P = 0.002) and ROS (P = 0.02) in iPS-ACM compared to POAF group. Kaplan-Meier survival analysis revealed that the POAF group (P = 0.01) and low SPARCL1 expression in epicardial adipocytes (P = 0.018) were associated with a higher incidence of long-term adverse cardiovascular events after surgery. Conclusion: SPARCL1 derived from epicardial adipocytes may play an important role in the suppression of POAF and long-term cardiovascular events via improving the myocardial redox state.