Cyclooxygenase Research Articles

NEDD4 Knockdown Suppresses Human Endometrial Stromal Cell Growth and Invasion by Regulating PTGS2-Mediated Ferroptosis in Endometriosis.

Endometriosis (EM) is a gynecological disease characterized by the benign growth of endometrial tissue outside the uterus. Upregulation of neuronally expressed developmentally downregulated 4 (NEDD4) has been reported to accelerate endometrial cancer progression. We explored whether abnormal expression of NEDD4 is correlated with EM. Endometrial tissue in patients without endometriosis was used to develop the original generation of endometrial stromal cells (ESCs). Different types of endometrial tissue of patients with endometriosis were used to measure the expression of NEDD4 by immunohistochemistry (IHC) and western blotting. Its biological functions in ESCs were investigated using a cell counting kit-8 assay, fluorescein diacetate (FDA) staining, and Transwell invasion assays. Additionally, its involvement in ferroptosis was assessed by measuring Fe2+, malondialdehyde (MDA), glutathione (GSH), and reactive oxygen species (ROS) levels and the expression of ferroptosis markers. Compared with normal controls, NEDD4 levels were significantly elevated in the endometrial tissue of patients with EM. Furthermore, NEDD4 expression was higher in the ectopic endometrium than in the eutopic endometrium. NEDD4 knockdown reduced the viability and invasive capacity of ESCs, increased Fe2+, MDA, and ROS levels, and decreased GSH content. Further analysis revealed that NEDD4 knockdown promoted ferroptosis in ESCs by increasing the expression of prostaglandin-endoperoxide synthase 2 (PTGS2). As an E3 ubiquitin ligase, NEDD4 reduced PTGS2 protein levels by accelerating its ubiquitination and subsequent proteasomal degradation. These findings suggest that inhibiting NEDD4 reduces ESC growth and invasion in EM by regulating PTGS2-dependent ferroptosis.

Petunidin attenuates vinclozolin instigated testicular toxicity in albino rats via regulating TLR4/MyD88/TRAF6 and Nrf-2/Keap-1 pathway: A pharmacodynamic and molecular simulation approach

Vinclozolin (VZN) is a widely used fungicide which exerts deleterious impacts on various organs including testis. Petunidin (PDN) is a polyphenolic compound that demonstrates a broad range of pharmacological activities. Thirty-two rats were divided into 4 groups including the control, VZN (100 mg/kg), VZN (100 mg/kg) + PDN (4 mg/kg) and PDN (4 mg/kg) treated group. The activities of antioxidant enzymes were assessed by using previously documented protocols. The gene expressions were determined by using qRT-PCR. The levels of hepatic function and apoptotic markers were evaluated by using standard ELISA technique. The histological analysis was carried out as per the standard protocol of histology. It was revealed that VZN disrupted the Nrf-2/Keap-1 pathway. Moreover, the activities of catalase (CAT), glutathione peroxidase (GPx), superoxide dismutase (SOD), heme-oxygenase-1 (HO-1) and glutathione reductase (GSR) were reduced whereas levels of reactive oxygen species (ROS) & malondialdehyde (MDA) were promoted following the VZN intoxication. Furthermore, VZN intoxication reduced total sperm count, viability, motility as well as luteinizing hormone (LH), follicle stimulating hormone (FSH), and plasma testosterone. Besides, administration of VZN decreased the expressions of 3β-Hydroxysteroid dehydrogenase (3β-HSD), steroidogenic acute regulatory protein (StAR) and 17β-Hydroxysteroid dehydrogenase (17β-HSD). Moreover, VZN exposure escalated the expressions of Bcl-2–associated X protein (Bax) and cysteine–aspartic acid protease-3 (Caspase-3) while reducing the expressions of B-cell lymphoma-2 (Bcl-2). Additionally, VZN administration increased the gene expression of toll-like receptor 4 (TLR4), tumor necrosis factor receptor-associated factor 6 (TRAF-6) and myeloid differentiation primary response 88 (MyD88). The levels of interleukin-6 (IL-6), nuclear factor kappa-B (NF-κB), interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), and the activity of cyclooxygenase-2 (COX-2) were promoted following the VZN administration. Furthermore, VZN intoxication disrupted the normal histology of testicular tissues. However, VZN + PDN treatment ameliorated testicular damage via regulating aforementioned dysregulations owing to its anti-inflammatory, antioxidative as well as anti-apoptotic potentials. Lastly, molecular docking (MD) was performed to assess the effectiveness of PDN as a curative compound by analyzing its binding affinity with the targeted proteins (Keap1, TLR4 and StAR). Our in-silico evaluations confirmed that PDN possesses the potential to interact with binding pockets of these proteins, emphasizing its capability as a curative compound to mitigate VZN-prompted reproductive damage.

Chitosan oligosaccharides: A potential therapeutic agent for inhibiting foam cell formation in atherosclerosis

Foam cell formation is a key hallmark in atherosclerosis and associated cardiovascular diseases (CVDs). The potential anti-atherosclerotic potential of chitosan oligosaccharides (COS) was investigated using oxLDL-treated RAW264.7 murine cells. COS treatment led to a significant inhibition of lipid accumulation, as demonstrated by Oil Red O staining, and reduced levels of total cholesterol, free cholesterol, cholesterol esters, and triglycerides in.oxLDL-treated RAW264.7 cells. COS blocked cholesterol influx through down-regulating class A1 scavenger receptors (SR-A1) and cluster of differentiation 36 (CD36) expression and stimulated cholesterol efflux through up-regulating ABC transporters ABCA-1 and ABCG-1 expressions. Additionally, COS treatment stimulated nuclear signaling pathways involving peroxisome proliferator-activated receptor-γ (PPAR-γ) and liver X receptor α (LXR-α), and also led to the phosphorylation of AMP-activated protein kinase (AMPK). COS further demonstrated anti-inflammatory effects by inhibiting the production of pro-inflammatory cytokines and the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in oxLDL-treated RAW264.7 cells, through suppression of NF-κB signaling. Furthermore, COS alleviated oxidative stress induced by oxLDL by activating Nrf2 signaling and enhancing the expression of antioxidant genes, including heme oxygenase-1 (HO-1), superoxide dismutase (SOD), glutathione peroxidase (Gpx), and catalase (CAT). In conclusion, COS can be beneficial in preventing atherosclerosis and related diseases.

The role of the cyclooxygenase-2 pathway in tissue ischemia and revascularization following skeletal muscle injury induced by bothropic snake venom

Bothrops asper venom (Bav) contains metalloproteinases that disrupt the microvascular system, impairing muscle tissue regeneration after injury. This study investigated the impact of the cyclooxygenase-2 (COX-2) pathway on vascular injury and revascularization in muscle injuries induced by Bav. Mice were injected with Bav into the gastrocnemius muscle and treated with lumiracoxib, a selective COX-2 inhibitor, 30 min, 2 days, and 6 days post-Bav injection. Muscle tissue was analyzed at 24 h, 7 days, and 21 days post-injection. A decrease in COX-2 expression at 24 h post-Bav injection indicated significant necrosis and tissue loss. Both Bav injection and lumiracoxib treatment influenced the decrease of prostaglandin (PG)D2 and PGE2 production. Seven and 21 days post-Bav injections, COX-2 expression increased, along with PGDs levels unaffected by lumiracoxib, indicating that the other isoform COX-1 pathway could contribute to the release of PGs. Bav/lumiracoxib treated animals presented exacerbated limb ischemia, implying that COX-2-derived prostaglandins preserve vessel integrity. CD31, an angiogenesis marker, initially (24 h) decreased post-Bav injection but increased at 7 and 21 days in Bav/lumiracoxib mice, suggesting a down-modulatory role for COX-2-derived prostaglandins in early angiogenesis and tissue regeneration. Vascular endothelial growth factor (VEGF) production rose 7 days post-Bav injection, supporting its role in angiogenesis. Previous treatment with lumiracoxib promoted release of VEGF levels 21 days post-Bav injury showing that the inhibition of COX-2 pathway in the early stage of revascularization stimulates the neovascularization regulated by elevated release of VEGF. Similarly, metalloproteinases (MMPs), such as MMP-9, MMP-10, and MMP-13, crucial for vascular remodeling, were elevated 21 days after Bav/lumiracoxib treatment. In conclusion, the COX-2 pathway is essential to decrease the high grade of ischemia caused by acute injury induced by Bav. However, the decrease of activity in the COX-2 pathway in the first stages of revascularization contributes to the elevated production of key pro-angiogenic mediators that up-regulate the restoration of microvasculature and blood flow in muscle tissue injured by botropic venoms.

Rhynchophylline Alleviates Hyperactivity and Cognitive Flexibility Impairment Associated With Inhibition of Inflammatory Responses in Mice That Partly Lack the Dopamine Transporter Protein.

Rhynchophylline (RHY) can alleviate some cognitive flexibility impairment and stereotyped behavior for attention-deficit hyperactivity disorder (ADHD) and Tourette syndrome (TS) patients as one of a keyextract and an active ingredient in Ningdong granule (NDG), which is a Traditional Chinese medicine (TCM) preparation widely used in the treatment of ADHD and TS children in China; however, the underlying mechanism is not well understood. Therefore, this study aimed to evaluate how RHY alleviates hyperactivity and cognitive flexibility impairment while inhibiting inflammatory responses in mice that partly lack dopamine transporter protein (DAT- mice). Male DAT- mice were randomly divided into the RHY group (n = 8) and administered RHY (30mg/kg) in the DAT- group (n = 8) and administered saline (i.p., 10mL/kg) in wild-type (WT) mice as the WT control group (n = 8). Hyperactivity and cognitive flexibility impairment were evaluated by the open field test (OFT) and the Morris water maze (MWM) test. The levels of the inflammatory factors of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) in cortical homogenates were tested by enzyme-linked immunosorbent assays (ELISA) after 8 weeks of treatment with RHY. In vitro, primary microglia and astrocytes extracted from the cortices of DAT- neonatal mice and WT neonatal mice were treated with lipopolysaccharide (LPS) (1mg/mL) to induce neuroinflammatory responses and with RHY (20mM) for 48h. The levels of the inflammatory factors TNF-α, IL-1β, inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX2) in the culture medium were measured at 6h, 24h, and 48h after treatment with LPS and RHY. RHY ameliorated hyperactivity and cognitive flexibility impairment in DAT- mice and inhibited the expression of the inflammatory factors TNF-α, IL-1β, iNOS, and COX-2 in microglia and astrocytes in vitro, and also inhibited the expression of TNF-α and IL-1β in cortical homogenates after 8 weeks of treatment. RHY improved hyperactivity and cognitive flexibility impairment through inhibiting inflammatory responses in DAT- mice.

Ginkgetin alleviates polystyrene microplastics-instigated liver injury in rats through Nrf-2/Keap-1 pathway activation

Polystyrene microplastics (PS-MPs) are potential toxicants that are reported to instigate oxidative stress (OS) in the liver. Ginkgetin (GK) is a natural biflavonoid with potential therapeutic activities. This experiment was executed to access the putative effect of GK against PS-MPs provoked hepatotoxicity. Four groups were formed from 48 rats including control, PS-MPs (0.01 mg/kg), PS-MPs (0.01 mg/kg) + GK (25 mg/kg) and GK (25 mg/kg) alone. The inebriation of PS-MPs markedly decreased the expressions of antioxidant genes and Nrf-2, besides escalating Keap-1 expression. It also decreased antioxidants i.e., glutathione (GSH), glutathione S-transferase (GST), catalase (CAT), glutathione peroxidase (GPx), heme oxygenase-1 (HO-1), superoxide dismutase (SOD), glutathione reductase (GSR) activities, while remarkably upsurged reactive oxygen species (ROS) along with malondialdehyde (MDA) contents. Additionally, a notable escalation in hepatic serum markers i.e., alkaline phosphatase (ALP), alanine transaminase (ALT) and aspartate aminotransferase (AST) level was observed. Furthermore, PS-MPs exposure escalated inflammatory biomarkers, tumor necrosis factor-α (TNF-α), interleukin- 6 (IL-6), nuclear factor-kappa B (NF-kB), interleukin-β (IL-1β) level and cyclooxygenase-2 (COX-2) activity as well as augmented Caspase-3 along with Bax expression and decreased Bcl-2 expression. Nevertheless, GK treatment notably abated PS-MPs prompted liver injuries owing to its hepatoprotective efficacy.

Therapeutic role of physalin A in the pathogenesis of Graves’ orbitopathy

Background Graves’ orbitopathy (GO) is an autoimmune condition that causes serious ocular symptoms; its treatment strategies are limited. Physalin A is a phytosterol that has shown various therapeutic properties, including anti-inflammatory and anti-fibrotic effects. In this study, we investigated whether physalin A could inhibit inflammation, fibrosis, hyaluronan (hyaluronic acid) production, and adipogenesis, which are crucial to the pathogenesis of GO. Methods Orbital tissue explants were obtained from patients with GO during orbital decompression surgery and healthy controls. Orbital fibroblasts (OFs) were isolated and treated with different concentrations of physalin A. Using western blot and ELISA analyses, we determined the effects of physalin A on OFs. Results Physalin A treatment suppressed the production of interleukin (IL)-1β-induced prostaglandin E2 (PGE2) and pro-inflammatory molecules, including cyclooxygenase (COX)-2, IL-6, IL-8, and intercellular adhesion molecule (ICAM)-1. We discovered that physalin A attenuated hyaluronan production induced by IL-1β or insulin-like growth factor (IGF)-1. Moreover, physalin A reduced lipid droplet formation and production of peroxisome proliferator activator (PPAR) γ, CCAAT-enhancer-binding protein (C/EBP) α, C/EBP β, sterol regulatory element binding protein (SREBP)-1, leptin, and adiponectin proteins. Physalin A suppressed the phosphorylation of extracellular signal-related kinase (ERK), nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB), and suppressor of mothers against decapentaplegic (SMAD) 2 signaling protein. Conclusions Our study suggests that the major mechanisms by which physalin A suppresses GO include reducing inflammation, fibrosis, hyaluronan production, and adipogenesis in OFs. The findings of this study provide evidence of the therapeutic effect of physalin A in GO.

Activation of mixed lineage kinase 3 by fine particulate matter induces skin inflammation in human keratinocytes.

Fine particulate matter (PM2.5) induces a range of diseases, including skin disorders, through inflammatory responses. In this study, we investigated the novel mechanisms by which PM2.5 causes skin inflammation in human keratinocytes HaCaT. We observed increased protein expression of cyclooxygenase-2 (COX-2) and the production of prostaglandin E2 (PGE2) in PM2.5-treated HaCaT cells. To identify the pathways promoting the expression of these inflammatory proteins, we conducted a phospho-kinase antibody array and confirmed that the phosphorylation levels of JNK and p38 were increased by PM2.5-treated HaCaT cells. Further investigation of the phosphorylation levels of mitogen-activated protein kinases (MAPKs) and upstream signals revealed that PM2.5 activated the MKK4/7-JNK-c-Jun and MKK3/6-p38-p70S6K signaling pathways, while the phosphorylation level of ERK1/2 remained unchanged. HaCaT cells treated with PM2.5 phosphorylated Mixed-lineage kinase 3 (MLK3), an upstream regulator of p38 and JNK. Furthermore, inhibition of ROS production by N-Acetylcysteine (NAC) treatment inhibited MLK3 phosphorylation. Taken together, ROS production induced by PM2.5 activated the MLK3 signaling pathway and induced skin inflammation.

Simulated microgravity environment inhibits matrix mineralization during the osteoblast to osteocyte differentiation

This study investigates the effects of microgravity on the differentiation and mineralization of IDG-SW3 osteocyte-like cells to understand the response of bone cells to microgravity and develop strategies to mitigate bone loss in astronauts. IDG-SW3 cells were cultured in collagen-coated dishes and subjected to a 3D clinostat to simulate microgravity 14 days after initiating differentiation. The static group remained under normal gravity. Cells were analyzed on days 14, 18, 22, and 26. Alizarin red staining demonstrated a substantial and time-dependent increase in mineralization in the static group, whereas the microgravity group exhibited little detectable mineralization throughout the experimental period. Quantitative RT-PCR revealed significant upregulation of Rankl, Alpl, Dmp1, and Fgf23 and downregulation of Sost and Phex in the microgravity group. RNA sequencing on day 26 showed distinct gene expression profiles between conditions. Heatmaps highlighted upregulated genes (Ptgs2, Alpl, Comp, Atf4, Lox) and downregulated genes (Rspo2, Ank, Ptn, Mmp13, Aspn, Spp1) under microgravity. Gene ontology (GO) enrichment analysis indicated that upregulated genes were associated with cytoskeletal organization and receptor activities, while downregulated genes were linked to extracellular matrix components and immune response. These findings provide insights into the molecular mechanisms of bone loss in space and emphasize the importance of gravity in bone remodeling. Future studies should validate these genes' functions in osteocyte biology under microgravity.

A potent dual inhibitor targeting COX-2 and HDAC of acute myeloid leukemia cells.

Acute myeloid leukemia (AML) is an aggressive cancer with complex issues of drug resistance and a poor prognosis; thus, effective therapeutics is urgently needed for AML. In this study, we designed and synthesized dual cyclooxygenase-2 (COX-2) and histone deacetylase (HDAC) inhibitors, IMC-HA and IMC-OPD, and applied them for the treatment of AML. IMC-HA comprised a COX-2 inhibitor skeleton of indomethacin (IMC) and an HDAC inhibitor moiety of the hydroxamic group and was found to exhibit potent antiproliferative activity against AML cells (THP-1 and U937) and low cytotoxicity toward normal cells. Molecular docking simulations suggested that IMC-HA had a high binding affinity for HDAC and COX-2, with binding energies of -6.8 and -9.0kcal/mol, respectively. Mechanistic studies revealed that IMC-HA induced apoptosis and G0/G1 phase arrest in AML cells, which were characterized by alterations in the expression of apoptotic and cell cycle-related proteins. Further study demonstrated that IMC-HA also inhibited the MEK/ERK signaling pathway in AML cells. Overall, we believe that IMC-HA could serve as a potent COX-2/HDAC dual inhibitor and improve the treatment of AML.

Comparison and Assessment of Anti-Inflammatory and Antioxidant Capacity Between EGCG and Phosphatidylcholine-Encapsulated EGCG.

To compare and evaluate the differences between EGCG and phosphatidylcholine-encapsulated EGCG in terms of their anti-inflammatory and antioxidant capacities. In this study, transdermal absorption experiments were conducted to compare the absorption capacity of EGCG and phosphatidylcholine-encapsulated EGCG. Subsequently, the disparity in anti-inflammatory and antioxidant efficacy between EGCG and phosphatidylcholine-encapsulated EGCG were evaluated through cytotoxicity experiments, as well as the determination of cellular inflammatory factors and the measurement of ROS content under different treatment conditions. The concentration of EGCG, encapsulated in phosphatidylcholine, in porcine skin is 40.76 ± 1.29 μg/cm2, which is significantly higher than the concentration of EGCG alone (31.62 ± 2.01 μg/cm2). Also, the ability of phosphatidylcholine-encapsulated EGCG to suppress inflammatory factors such as tumor necrosis factor-α (TNF-α), cyclooxygenase-2 (COX-2), and prostaglandin E2 (PGE2) was notably superior to that of EGCG alone. Both phosphatidylcholine-encapsulated EGCG and EGCG showed excellent ROS scavenging ability in terms of antioxidant capacity. The percutaneous absorption and anti-inflammatory impact of EGCG encapsulated within phosphatidylcholine were substantially enhanced when compared to EGCG by itself. Additionally, both formulations exhibited enhanced ROS scavenging capacities, albeit the variance between them was not pronounced. These insights furnish a vital theoretical underpinning for the utilization of phosphatidylcholine-encapsulated EGCG in cosmetic applications, specifically for fostering products with anti-inflammatory and antioxidant benefits.

Efficacy and Mechanism of Schisandra chinensis Fructus Water Extract in Alzheimer's Disease: Insights from Network Pharmacology and Validation in an Amyloid-β Infused Animal Model.

Schisandra chinensis Fructus (SCF) is a traditional medicinal herb containing lignans that improves glucose metabolism by mitigating insulin resistance. We aimed to investigate the therapeutic potential and action mechanism of SCF for Alzheimer's disease (AD) using a network pharmacology analysis, followed by experimental validation in an AD rat model. The biological activities of SCF's bioactive compounds were assessed through a network pharmacology analysis. An AD rat model was generated by infusing amyloid-β peptide (Aβ) (25-35) into the hippocampus to induce Aβ accumulation. The AD rats were fed either 0.5% dextrin (AD-Con) or 0.5% SCF (AD-SCF group) in a high-fat diet for seven weeks. The rats in the normal/control group received an Aβ (35-25) infusion (no Aβ deposition) and were fed a control diet (Normal-C). Aβ deposition, memory function, inflammation, and glucose/lipid metabolism were evaluated. The network analysis revealed significant intersections between AD-related targets and bioactive SCF compounds, like gomisin A, schisandrin, and longikaurin A. Key AD genes prostaglandin-endoperoxide synthase-2 (PTGS2, cyclooxygenase-2) and acetylcholinesterase (AChE) were linked to SCF compounds. In the rats with AD induced by bilaterally infusing amyloid-β (25-35) into the hippocampus, the 0.5% SCF intake mitigated hippocampal amyloid-β deposition, neuroinflammation, memory deficits, and dysregulated glucose and lipid metabolism versus the AD controls. SCF reduced hippocampal AChE activity, inflammatory cytokine expression related to PTGS2, and malondialdehyde contents and preserved neuronal cell survival-related factors such as brain-derived neurotrophic factor and ciliary neurotrophic factor similar to normal rats. The neuroprotective effects validated the network analysis findings. SCF could be a potential AD therapeutic agent by activating the parasympathetic nervous system to reduce hippocampal oxidative stress and inflammation, warranting further clinical investigations of its efficacy.

SR-16234, a Unique Selective Estrogen Receptor Modulator, Suppressed Proliferation and Pain-Related Factor Expression by Inhibition of the Nuclear Factor-kappa B Pathway in Endometriotic Stromal Cells.

What is the effect of SR-16234 (SR), a selective estrogen receptor (ER) modulator, on human endometriotic stromal cells (ESCs)? Endometriotic tissues were obtained from 21 patients undergoing laparoscopic surgery for ovarian endometriomas (OEs). Normal eutopic endometrium during the luteal phase was obtained from 18 patients without endometriosis. ESCs isolated from OEs and normal eutopic endometrial stromal cells (NESCs) were cultured with SR and subsequently exposed to tumor necrosis factor (TNF)-α. After 48 h of incubation, the effect of SR on cell proliferation was evaluated by the WST-8 assay. The gene expressions of inflammatory and pain-related factors, including interleukin (IL)-6, IL-8, cyclooxygenase (COX)-2, transient receptor potential vanilloid (TRPV)1, ESR1, and ESR2, were evaluated by real-time RT-PCR. The phosphorylation of Inhibitor κBα (IκBα), extracellular signal-regulated kinase (ERK)1/2, and Protein Kinase B (AKT) were evaluated by western blot analysis. ILs, prostaglandin (PG) E2, and intranuclear p65 syntheses were assessed by ELISA. SR treatment repressed TNF-α-induced proliferation by 20% in ESCs but not NESCs. SR also reduced IL-6, IL-8, COX-2, TRPV1, ESR1, and ESR2 mRNA expressions and ILs protein, and PGE2 synthesis in ESCs, whereas in NESCs, only TRPV1 mRNA expression was decreased. SR suppressed TNF-α-induced phosphorylated IκBα levels by approximately 50%, and intranuclear p65 protein was reduced by 30% compared to addition of only TNF-α in ESCs. However, SR did not affect the phosphorylation of AKT and ERK1/2. SR appears to be a potential therapeutic agent for endometriosis by suppressing inflammatory and pain-related factor expressions by inhibiting the nuclear factor-kappa B pathway.

P2Y12 receptor-mediated cyclooxygenase 2 (COX-2) expression enhances tumor cell progression in a mouse model of lymphoma.

The pro-inflammatory enzyme cyclooxygenase 2 (COX-2) has been known to impart metastatic property to cancer cells. However, blocking of COX-2 with nonsteroidal anti-inflammatory drugs or COX-2-specific inhibitors has failed in clinical trials due to adverse effects associated with their prolonged use. We have previously shown that extracellular ATP (eATP), a major component of the tumor microenvironment, enhances COX-2 expression several-fold, both in macrophages and in various cancer cells, by acting on purinergic (P2) receptors. In this study, we show that blocking of P2 receptors significantly reduced tumor growth in a mouse model of lymphoma. Tumors were induced in mice through subcutaneous injection of syngeneic EL4 lymphoma cells. Various P2 receptor antagonists were injected within the tumors after they were palpable. The broad-spectrum P2 receptor antagonist, suramin, P2X7 receptor-specific antagonist, oATP, P2Y6 receptor-specific antagonist, MRS 2578, and P2Y12 receptor-specific antagonist, AR-C 69931, all showed significant arrest in tumor growth. Both suramin and AR-C 69931-treated tumors showed strong reduction in COX-2 expression and modulation of various metastatic markers. Disaggregated cells from AR-C 69931-treated tumors, when injected intravenously in naïve mice, did not exhibit metastasis in various tissues which was observed in mice injected with cells from saline-treated tumors. Our results show that blocking of P2 receptors is a therapeutic alternative to inhibit COX-2 expression, and thereby, arrest tumor progression and metastasis.

Dual COX-2/TNF-α Inhibitors as Promising Anti-inflammatory and Cancer Chemopreventive Agents: A Review

: Cyclooxygenases (COX) play a pivotal role in inflammation and are responsible for the production of prostaglandins (PGs). Two types of COXs have been identified as key biological targets for drug design: Constitutive COX-1 and inducible COX-2. Nonsteroidal anti-inflammatory drugs (NSAIDs) target COX-1, while selective COX-2 inhibitors are designed for COX-2. These COX isoforms are involved in multiple physiological and pathological pathways throughout the body. Overproduction of tumor necrosis factor-alpha (TNF-α) plays a role in COX-2's inflammatory activity. Tumor necrosis factor-alpha can contribute to cardiac fibrosis, heart failure, and various cancers by upregulating the COX-2/PGE2 axis. Therefore, suppressing COX activity has emerged as a potentially effective treatment for chronic inflammatory disorders and cancer. This review explores the mechanisms of TNF-α-induced COX-2/PGE2 expression, a significant pathophysiological feature of cancer development. Furthermore, we summarize chemical compounds with dual COX-2/TNF-α inhibitory actions, providing an overview of their structure-activity relationship. These insights may contribute to the development of new generations of dual-acting COX-2/TNF-α inhibitors with enhanced efficacy.

Melissa officinalis Regulates Lipopolysaccharide-Induced BV2 Microglial Activation via MAPK and Nrf2 Signaling.

Neuroinflammation and microglial activation play critical roles in neurodegenerative diseases such as Alzheimer's and Parkinson's disease. Modulating microglial activation may help prevent the progression of these disorders. This study aimed to investigate the effects and mechanisms of Melissa officinalis ethanol extract on lipopolysaccharide (LPS)-induced microglial activation in BV2 cells. Cell viability and nitric oxide (NO) production were assessed using MTT assay and Griess reagent, while inflammatory cytokine levels were measured by qPCR. Key inflammatory pathways, including MAPK, TLR4, and antioxidant biomarkers, were analyzed through western blot and immunofluorescence. Rosmarinic acid content in M. officinalis was determined using high-performance liquid chromatography (HPLC). The results demonstrated that M. officinalis ethanol extract significantly inhibited LPS-induced NO production and reduced inflammatory cytokine expression. Additionally, it downregulated inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), TLR4, NF-κB, and MAPK signaling pathways (p38, JNK, ERK), while increasing the expression of antioxidant markers, including Nrf2, HO-1, catalase, and SOD2. In conclusion, M. officinalis ethanol extract exerts neuroprotective effects by modulating inflammation and enhancing antioxidant defenses, suggesting its potential in the prevention and treatment of inflammation-related neurodegenerative diseases.

Ferroptosis contributes to the development of cardiac dysfunction in iron overload cardiomyopathy

Abstract Background Ferroptosis is a novel form of cell death, and its role in iron overload cardiomyopathy (IOC) has not been elucidated. Purpose To explore whether ferroptosis of cardiomyocytes contributes to cardiac dysfunction in IOC by cardiac MR (CMR) and molecular biology techniques in vivo. Methods A total of 14 Sprague-Dawley (SD) rats were randomly divided into two groups: the control group (n=6) and the IOC group (n=8). The latter was induced by intraperitoneal injection of iron dextran on alternate days for 9 weeks and underwent a 7T CMR for assessment of cardiac function, including left ventricular ejection fraction (LVEF) and global longitude strain (GLS), along with the assessment of iron overload severity and fibrosis using Cine, T2 mapping, and late gadolinium enhancement (LGE), respectively. Circulating iron overload was determined via blood biochemistry analysis, measuring serum iron, ferritin, and transferrin levels. Furthermore, Prussian blue and Masson staining were employed to identify iron and fibrosis deposition within the myocardium, respectively. Cardiac ultrastructure, especially mitochondria, was examined via transmission electron microscopy (TEM). A sensitive fluorescent probe, dihydroethidium (DHE), was employed for the detection of reactive oxygen species (ROS) levels. Malondialdehyde (MDA), prostaglandin-endoperoxide synthase 2 (PTGS2), and glutathione peroxidase 4 (GPX4) were utilized to identify and quantify levels of lipid peroxidation, thereby providing comprehensive insights into cardiac cellular ferroptosis. Results 6 rats survived in the control group and 5 in the IOC group. Compared with the control group, the IOC rats had reduced T2 values of the cardiac septum (P<0.001), with normal LVEF (P>0.05) but decreased GLS (P<0.01), seen in Fig 1A. Based on blood biochemistry and Prussian blue staining, they were evident that IOC rats exhibited both circulating and cardiac iron overload, as seen in Figure 1B, C. Masson staining showed no obvious myocardial fibrosis, which was consistent with negative CMR LGE results, seen in Fig 1A, B. TEM further elucidated cardiac mitochondrial injuries in IOC rats, including focal vacuolization, swelling, crest disruption, and even disappearance, seen in Fig 2A. The fluorescent probe results revealed a striking increase in red fluorescence within the myocardium of IOC rats, seen in Fig 2B. Additionally, IOC rats exhibited significantly elevated levels of MDA and PTGS2mRNA (P<0.001), accompanied by notable downregulation of GPX4mRNA expression (P<0.001), seen in Fig 2C. These findings suggested the occurrence of ferroptosis in the myocardium of IOC rats. Conclusions In IOC, ferroptosis rather than necrosis is the primary driver of cardiac dysfunction, which is characterized by maintaining normal LVEF but experiencing a reduction in GLS without accompanying myocardial fibrosis. Our study sheds light on the potential involvement of ferroptosis in the pathogenesis of IOC.

Aspirin inhibits atherosclerosis through macrophage pyroptosis by interacting with Lactobacillus johnsonii and its metabolite butyrate

Abstract Introduction Atherosclerotic cardiovascular disease (ASCVD) is a chronic progressive disease closely related to metabolism and inflammation. Aspirin inhibits cyclooxygenase (COX) and has antiplatelet aggregation effects, considered as the cornerstone of ASCVD treatment. Numerous studies have shown that gut microbiota (GM) and its metabolites affects host metabolism. Studies have also shown that people taking aspirin exhibit specific changes in the microbiota profile. However, whether the interaction of aspirin with GM and its metabolites affects the therapeutic efficacy of aspirin in treating atherosclerosis (AS) have not been reported. Purpose This study aimed to explore the interaction of aspirin with the GM and its metabolites in the treatment of AS, to reveal the specific mechanism of the GM-metabolite axis and provide potential new targets for the combined treatment of AS. Methods To establish the mouse model of atherosclerosis by high-fat diet, and the atherosclerotic plaque burden and composition were analyzed. The role of GM in aspirin anti-atherosclerosis was explored by antibiotics and fecal microbiota transplantation (FMT). The third generation 16S RNA sequencing was used to explore the effects of aspirin on species diversity and abundance of GM, and to screen differential microorganisms for intervention of AS mice. UHPLC-MS and GC-MS were used to explore the changes of intestinal metabolites induced by aspirin. The correlation between gut microbiota and metabolites was also investigated. Finally, in vitro and in vivo experiments were performed to confirm the effects and mechanisms of metabolites on macrophage pyroptosis and atherosclerosis. Results Aspirin has anti-atherosclerosis effect. After removing gut microbiota, the therapeutic effect of aspirin on atherosclerosis was decreased. Three generations of 16S rRNA gene sequencing results showed that aspirin supplementation alters the diversity of gut microbiota, and increased the relative abundance of Lactobacillus johnsonii(L.johnsonii) and Ligilactobacillus murinus(L.murinus). After gavage of L.johnsonii and L.murinus, the plaque burden and macrophage pyroptosis were significantly reduced in AS mice. Colonization with L.johnsonii and L.murinus significantly increased the abundance of probiotics and butyrate level in the feces of AS mice and L.johnsonii and L.murinus were positively correlated with butyrate. Supplementation of butyrate inhibited macrophage pyroptosis and atherosclerosis in vivo. In vitro, butyrate could inhibit the expression of NLRP3/Caspase-1/IL-1β and pyroptosome production, and this process mainly by activating butyrate receptors GPR41, GPR43 and GPR109A. Conclusions The gut microbiota may mediate the anti-AS effect of aspirin by promoting an increase in the abundance of L.johnsonii and L.murinus. Butyrate produced by Lactobacillus may mediate the anti-AS effect of aspirin, and inhibiting macrophage pyroptosis and the progression of AS.

Antioxidant and Anti-Inflammatory Properties of Conceivable Compounds from Glehnia littoralis Leaf Extract on RAW264.7 Cells.

Glehnia littoralis is a medicinal plant, but the scientific basis is still unclear. This study thoroughly investigated phenols from Glehnia littoralis extract (GLE) to determine their potential as anti-inflammatory and antioxidant agents. High-performance liquid chromatography (HPLC) and mass spectrometry (MS) were used to analyze the compounds in GLE. In addition, we performed GLE in vitro in macrophages after lipopolysaccharide (LPS)-induced inflammation. The extract contained eight peaks representing phenolic compounds and one peak representing riboflavin, with the corresponding mass spectrometry data documented. These biologically active compounds were purified by ultrafiltration using LC to determine their ability to target cyclooxygenase-2 (COX-2) and 2,2-diphenyl-1-picrylhydrazyl (DPPH). The results showed that significant compounds were identified, demonstrating a binding affinity for both COX-2 and DPPH. This suggests that the compounds showing excellent binding affinity for COX-2 and DPPH may be the main active ingredients. Vital inflammatory cytokines, including COX-2, inducible nitric oxide synthase (iNOS), mitogen-activated protein kinase (MAPK), and nuclear factor kappa B (NF-κB), were found to be down-regulated during the treatment. In addition, we revealed that the selected drugs exhibited potent binding capacity to inflammatory factors through molecular docking studies. In addition, we confirmed the presence of phenolic components in GLE extract and verified their possible anti-inflammatory and antioxidant properties. This study provided evidence for an efficient strategy to identify critical active ingredients from various medicinal plants. These data may serve as a baseline for further investigations of applying GLE in the pharmaceutical industry.

Uterine stromal but not epithelial PTGS2 is critical for murine pregnancy success.

Use of non-steroidal anti-inflammatory drugs that target prostaglandin synthase (PTGS) enzymes have been implicated in miscarriage. Further, PTGS2-derived prostaglandins are reduced in the endometrium of patients with a history of implantation failure. However, in the mouse model of pregnancy, peri-implantation PTGS2 function is controversial. Some studies suggest that Ptgs2 -/- mice display deficits in ovulation, fertilization, and implantation, while other studies suggest a role for PTGS2 only in ovulation but not implantation. Further, the uterine cell type responsible for PTGS2 function and role of PTGS2 in regulating implantation chamber formation is not known. To address this we generated tissue-specific deletion models of Ptgs2 . We observed that PTGS2 ablation from the epithelium alone in Ltf cre/+ ; Ptgs2 f/f mice and in both the epithelium and endothelium of the Pax2 cre/+ ; Ptgs2 f/f mice does not affect embryo implantation. Further, deletion of PTGS2 in the ovary, oviduct, and the uterus using Pgr cre/+ ; Ptgs2 f/f does not disrupt pre-implantation events but instead interferes with post-implantation chamber formation, vascular remodeling and decidualization. While all embryos initiate chamber formation, more than half of the embryos fail to transition from blastocyst to epiblast stage, resulting in embryo death and resorbing decidual sites at mid-gestation. Thus, our results suggest no role for uterine epithelial PTGS2 in early pregnancy but instead highlight a role for uterine stromal PTGS2 in modulating post-implantation embryo and implantation chamber growth. Overall, our study provides clarity on the compartment-specific role of PTGS2 and provides a valuable model for further investigating the role of stromal PTGS2 in post-implantation embryo development.