Nicotinamide Riboside Augments Human Macrophage Migration via SIRT3-Mediated Prostaglandin E2 Signaling.

NAD+ boosting via nicotinamide riboside (NR) confers anti-inflammatory effects. However, its underlying mechanisms and therapeutic potential remain incompletely defined. Here, we showed that NR increased the expression of CC-chemokine receptor 7 (CCR7) in human M1 macrophages by flow cytometric analysis of cell surface receptors. Consequently, chemokine ligand 19 (CCL19, ligand for CCR7)-induced macrophage migration was enhanced following NR administration. Metabolomics analysis revealed that prostaglandin E2 (PGE2) was increased by NR in human monocytes and in human serum following in vivo NR supplementation. Furthermore, NR-mediated upregulation of macrophage migration through CCL19/CCR7 was dependent on PGE2 synthesis. We also demonstrated that NR upregulated PGE2 synthesis through SIRT3-dependent post-transcriptional regulation of cyclooxygenase 2 (COX-2). The NR/SIRT3/migration axis was further validated using the scratch-test model where NR and SIRT3 promoted more robust migration across a uniformly disrupted macrophage monolayer. Thus, NR-mediated metabolic regulation of macrophage migration and wound healing may have therapeutic potential for the topical management of chronic wound healing.

PGE2 Potentiates Orai1-Mediated Calcium Entry Contributing to Peripheral Sensitization.

Peripheral sensitization is one of the primary mechanisms underlying the pathogenesis of chronic pain. However, candidate molecules involved in peripheral sensitization remain incompletely understood. We have shown that store-operated calcium channels (SOCs) are expressed in the dorsal root ganglion (DRG) neurons. Whether SOCs contribute to peripheral sensitization associated with chronic inflammatory pain is elusive. Here we report that global or conditional deletion of Orai1 attenuates Complete Freund's adjuvant (CFA)-induced pain hypersensitivity in both male and female mice. To further establish the role of Orai1 in inflammatory pain, we performed calcium imaging and patch-clamp recordings in wild-type (WT) and Orai1 knockout (KO) DRG neurons. We found that SOC function was significantly enhanced in WT but not in Orai1 KO DRG neurons from CFA- and carrageenan-injected mice. Interestingly, the Orai1 protein level in L3/4 DRGs was not altered under inflammatory conditions. To understand how Orai1 is modulated under inflammatory pain conditions, prostaglandin E2 (PGE2) was used to sensitize DRG neurons. PGE2-induced increase in neuronal excitability and pain hypersensitivity was significantly reduced in Orai1 KO mice. PGE2-induced potentiation of SOC entry (SOCE) was observed in WT, but not in Orai1 KO DRG neurons. This effect was attenuated by a PGE2 receptor 1 (EP1) antagonist and mimicked by an EP1 agonist. Inhibition of Gq/11, PKC, or ERK abolished PGE2-induced SOCE increase, indicating PGE2-induced SOCE enhancement is mediated by EP1-mediated downstream cascade. These findings demonstrate that Orai1 plays an important role in peripheral sensitization. Our study also provides new insight into molecular mechanisms underlying PGE2-induced modulation of inflammatory pain.Significance Statement Store-operated calcium channel (SOC) Orai1 is expressed and functional in dorsal root ganglion (DRG) neurons. Whether Orai1 contributes to peripheral sensitization is unclear. The present study demonstrates that Orai1-mediated SOC function is enhanced in DRG neurons under inflammatory conditions. Global and conditional deletion of Orai1 attenuates complete Freund's adjuvant (CFA)-induced pain hypersensitivity. We also demonstrate that prostaglandin E2 (PGE2) potentiates SOC function in DRG neurons through EP1-mediated signaling pathway. Importantly, we have found that Orai1 deficiency diminishes PGE2-induced SOC function increase and reduces PGE2-induced increase in neuronal excitability and pain hypersensitivity. These findings suggest that Orai1 plays an important role in peripheral sensitization associated with inflammatory pain. Our study reveals a novel mechanism underlying PGE2/EP1-induced peripheral sensitization. Orai1 may serve as a potential target for pathological pain.

Protective effect of Amauroderma rugosum ethanol extract and its primary bioactive compound, ergosterol, against acute gastric ulcers based on LXR-mediated gastric mucus secretions.

BACKGROUND: Amauroderma rugosum (Blume & T. Nees) Torrend (Ganodermataceae) is an edible mushroom with a wide range of medicinal values. Our previous publication demonstrated the therapeutic effects of the water extract of A. rugosum (WEA) against gastric ulcers. However, the protective effects of the ethanol extract of A. rugosum (EEA) on gastric mucosa and its major active constituents have not yet been elucidated. PURPOSE: This study aims to evaluate the gastroprotective effects and underlying mechanisms of EEA and its fat-soluble constituent, ergosterol, in acute gastric ulcers. STUDY DESIGN AND METHOD: SD rats were pre-treated with EEA (50, 100, and 200 mg/kg) or ergosterol (5, 10, and 20 mg/kg), and acute gastric ulcer models were constructed using ethanol, gastric mucus secretion inhibitor (indomethacin) or pyloric-ligation. The gastric ulcer area, histological structure alterations (H&E staining), and mucus secretion (AB-PAS staining) were recorded. Additionally, Q-PCR, western blotting, immunohistochemistry, ELISA, molecular docking, molecular dynamics simulations, MM-GBSA analysis, and surface plasmon resonance assay (SPR) were used to investigate the underlying mechanisms of the gastroprotective effect. RESULT: Compared with WEA, which primarily exerts its anti-ulcer effects by inhibiting inflammation, EEA containing fat-soluble molecules showed more potent gastroprotective effect through the promotion of gastric mucus secretion, as the anti-ulcer activity was partly blocked by indomethacin. Meanwhile, EEA exhibited anti-inflammatory effects by suppressing the production of IL-6, IL-1ß, TNF-α, and NO, thereby inhibiting the MAPK pathway. Significantly, ergosterol (20 mg/kg), the bioactive water-insoluble compound in EEA, exhibited a gastroprotective effect comparable to that of lansoprazole (30 mg/kg). The promotion of gastric mucus secretion contributed to the effects of ergosterol, as indomethacin can completely block it. The upregulations of COX1-PGE2 and C-fos, an activator protein 1 (AP-1) transcription factor, were observed after the ergosterol treatment. Ergosterol acted as an LXRß agonist via van der Waals binding and stabilizing the LXRß protein without compromising its flexibility, thereby inducing the upregulation of AP-1 and COX-1. CONCLUSION: EEA and its primary bioactive compound, ergosterol, exert anti-ulcer effects by promoting gastric mucus secretion through the LXRß/C-fos/COX-1/PGE2 pathway.

Enzyme-treated red rice (Oryza sativa L.) bran extracts mitigate inflammatory markers in RAW 264.7 macrophage cells and exhibit anti-inflammatory efficacy greater/comparable to ferulic acid, catechin, γ-tocopherol, and γ-oryzanol.

ETHNOPHARMACOLOGICAL RELEVANCE: Rice (Oryza sativa L.), a staple food for a significant portion of the global population, has been recognized for its traditional medicinal properties for centuries. Rice bran, a by-product of rice milling, contains many bioactive compounds with potential pharmaceutical and therapeutic benefits. In recent years, research has highlighted the anti-inflammatory potential of rice bran, contributed by the bioactive components concentrated in their bran but, unfortunately, entrapped in the bran matrix, with limited bioavailability. Previous studies have reported that the enzymatic treatment of rice bran improves the bran's bioactive compound profile but did not investigate its impact on chronic conditions such as inflammation. AIM OF THE STUDY: This study investigates the anti-inflammatory effects of endo-1,4-ß-xylanase (ERB) and Viscozyme (VRB) treated red rice bran extracts against lipopolysaccharide-induced inflammation in RAW264.7 macrophages in comparison with non-enzyme-treated bran (CRB). Further established their efficacy with known anti-inflammatory compounds-ferulic acid (FA), catechin (CAT), γ-tocopherol (GTP), and γ-oryzanol (ORZ). MATERIALS AND METHODS: The RAW 264.7 macrophage cells were pre-treated with non-toxic concentrations (10-200 µg/mL) of FA, CAT, GTP, ORZ, CRB, ERB, and VRB, followed by inflammatory stimulation with LPS for 24 h. Further, the cell supernatant and pellets were harvested to study the anti-inflammatory effects by evaluating and measuring their efficacy in inhibiting pro-inflammatory cytokines (TNF-α, IL-6, IL-10, IL-1ß) and mediators (ROS, NO, PGE2, COX2, iNOS) through biochemical, ELISA, and mRNA expression studies. RESULTS: The findings showed that both ERB and VRB effectively inhibited the production of pro-inflammatory markers (TNF-α, IL-6) and mediators (ROS, NO, PGE2) by downregulating mRNA expressions of inflammatory genes (TNF-α, IL-1ß, IL-6, IL-10, COX2, iNOS) and demonstrated anti-inflammatory efficacy higher than CRB. On comparison, ERB demonstrated exceptional efficacy by causing a reduction of 48% in ROS, 20% in TNF-α, and 23% in PGE2 at 10 µg/mL, surpassing the anti-inflammatory capabilities of all the bioactive compounds, FA and ORZ, respectively. At the same time, VRB exhibited remarkable efficacy by reducing NO production by 52% at 200 µg/mL and IL-6 by 66% at 10 µg/mL, surpassing FA, CAT, ORZ, and GTP. Further, ERB downregulated the mRNA expression of IL-10 and iNOS, while VRB downregulated TNF-α, IL-1ß, and COX2 expression. Both extracts equally downregulated IL-6 expression at 10 µg/mL, demonstrating the efficacy more remarkable/on par with established anti-inflammatory compounds. CONCLUSIONS: Overall, enzyme-treated rice bran/extract, particularly ERB, possesses excellent anti-inflammatory properties, making them promising agents for alternatives to contemporary nutraceuticals/functional food against inflammatory diseases.

Peony Pollen Protects against Primary Dysmenorrhea in Mice by Inhibiting Inflammatory Response and Regulating the COX2/PGE2 Pathway.

Peony pollen contains multiple nutrients and components and has been used as a traditional Chinese medicine with a long history, but the effect of the treatment of primary dysmenorrhea remains to be clarified. The aim of this study is to investigate the therapeutic effect of peony pollen on primary dysmenorrhea mice and the potential mechanism. A uterus contraction model in vitro and primary dysmenorrhea mice were used to evaluate the treatment effect of peony pollen on primary dysmenorrhea. The primary dysmenorrhea mice were treated with 62.5 mg/kg, 125 mg/kg, or 250 mg/kg of peony pollen, and the writhing response, latency period, histopathological changes in the uterus, prostaglandin E2 (PGE2) and prostaglandin F2α (PGF2α) levels, and infiltration of neutrophils and macrophages were investigated. Protein expression of interleukin 1 ß (IL-1ß), interleukin 6 (IL-6), NOD-like receptor thermal protein domain associated protein 3 (NLRP3), cyclooxygenase-2 (COX-2), microsomal prostaglandin-E synthase 1 (mPGEs-1), BCL2-Associated X (Bax), B-cell lymphoma-2 (BCL-2), caspase-3, and cleaved caspase-3 were detected by Western blot, and the oxidative stress related marker malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and reactive oxygen species (ROS) were evaluated. Peony pollen could attenuate spontaneous or oxytocin-induced uterus contractions in vitro. Moreover, peony pollen decreased the writhing times, prolonged the writhing latency, and reduced the pathological damage of uterine tissues. Furthermore, the inflammatory cell infiltration and the protein expression of IL-1ß, IL-6, and NLRP3 were decreased. The COX-2/PGE2 pathway was inhibited; oxidative stress and apoptosis in the uterus also improved in the uterus of primary dysmenorrhea mice. Peony pollen exerts a positive effect on primary dysmenorrhea by inhibiting the inflammatory response and modulating oxidative stress and apoptosis by regulating the COX-2/PGE2 pathway.

Neutralizing tumor-related inflammation and reprogramming of cancer-associated fibroblasts by Curcumin in breast cancer therapy.

Tumor-associated inflammation plays a vital role in cancer progression. Among the various stromal cells, cancer-associated fibroblasts are promising targets for cancer therapy. Several reports have indicated potent anti-inflammatory effects attributed to Curcumin. This study aimed to investigate whether inhibiting the inflammatory function of cancer-associated fibroblasts (CAFs) with Curcumin can restore anticancer immune responses. CAFs were isolated from breast cancer tissues, treated with Curcumin, and co-cultured with patients' PBMCs to evaluate gene expression and cytokine production alterations. Blood and breast tumor tissue samples were obtained from 12 breast cancer patients with stage II/III invasive ductal carcinoma. Fibroblast Activation Protein (FAP) + CAFs were extracted from tumor tissue, treated with 10 µM Curcumin, and co-cultured with corresponding PBMCs. The expression of smooth muscle actin-alpha (α-SMA), Cyclooxygenase-2(COX-2), production of PGE2, and immune cell cytokines were evaluated using Real-Time PCR and ELISA, respectively. Analyzes showed that treatment with Curcumin decreased the expression of genes α-SMA and COX-2 and the production of PGE2 in CAFs. In PBMCs co-cultured with Curcumin-treated CAFs, the expression of FoxP3 decreased along with the production of TGF-ß, IL-10, and IL-4. An increase in IFN-γ production was observed that followed by increased T-bet expression. According to our results, Curcumin could reprogram the pro-tumor phenotype of CAFs and increase the anti-tumor phenotype in PBMCs. Thus, CAFs, as a component of the tumor microenvironment, are a suitable target for combination immunotherapies of breast cancer.

Ginsenoside Rh2 and its octyl ester derivative inhibited invasion and metastasis of hepatocellular carcinoma via the c-Jun/COX2/PGE2 pathway.

BACKGROUND: Liver cancer is a topical global health issue. The treatment of liver cancer meets significant challenges in the high recurrence rate and invasive incidence. Therefore, the treatment strategies that target epithelial-mesenchymal transition (EMT) induced by cyclooxygenase 2 (COX2)/ prostaglandin E2 (PGE2) pathway have become epidemic. Ginsenoside Rh2 has been proved to inhibit the EMT. However, the underlying mechanisms remain unclear. Moreover, the octyl ester derivative of Rh2 (Rh2-O) exhibited superior anti-proliferative and immunomodulatory effects than Rh2 in our previous researches, which indicated that Rh2-O might also exert inhibitory effects on invasion and metastasis. PURPOSE: The aim of current study is to explore the inhibitory effects of Rh2 and Rh2-O on invasion and metastasis of hepatocellular carcinoma, and to investigate whether these effects are dependent on the c-Jun/COX2/PGE2 pathway. STUDY DESIGN: The Huh-7 liver cancer cells and the H22 tumor-bearing mice were treated with Rh2 and Rh2-O. METHOD: In this paper, the inhibitory effects of Rh2 and Rh2-O on invasion and metastasis were tested by wound healing, trans-well assay and tumor-bearing mice, and the involvement of c-Jun/COX2/PGE2 pathway were verified by exogenous PGE2, activation of COX2 and overexpression of c-Jun. RESULTS: The results showed that Rh2 and Rh2-O could efficiently inhibit the invasion and metastasis in a dose-dependent manner (p < 0.05). And the Rh2-O showed stronger effects than Rh2. Moreover, the exogenous PGE2, activation of COX2 by exogenous LPS and the overexpression of c-Jun by transfection all reversed the inhibitory effects of Rh2 and Rh2-O on metastasis or EMT (p < 0.05). CONCLUSION: Rh2 and Rh2-O could inhibit the invasion and metastasis of hepatocellular carcinoma via restraining the EMT, which was mediated by c-Jun/COX2/PGE2 pathway.

Unloading-Induced Skeletal Interoception Alters Hypothalamic Signaling to Promote Bone Loss and Fat Metabolism.

Microgravity is the primary factor that affects human physiology in spaceflight, particularly bone loss and disturbances of the central nervous system. However, little is known about the cellular and molecular mechanisms of these effects. Here, it is reported that in mice hindlimb unloading stimulates expression of neuropeptide Y (NPY) and tyrosine hydroxylase (TH) in the hypothalamus, resulting in bone loss and altered fat metabolism. Enhanced expression of TH and NPY in the hypothalamus occurs downstream of a reduced prostaglandin E2 (PGE2)-mediated ascending interoceptive signaling of the skeletal interoception. Sympathetic antagonist propranolol or deletion of Adrb2 in osteocytes rescue bone loss in the unloading model. Moreover, depletion of TH+ sympathetic nerves or inhibition of norepinephrine release ameliorated bone resorption. Stereotactic inhibition of NPY expression in the hypothalamic neurons reduces the food intake with altered energy expenditure with a limited effect on bone, indicating hypothalamic neuroendocrine factor NPY in the facilitation of bone formation by sympathetic TH activity. These findings suggest that reduced PGE2-mediated interoceptive signaling in response to microgravity or unloading has impacts on the skeletal and central nervous systems that are reciprocally regulated.

Histological, Physiological and Biomechanical Effects of Low-Level Laser Therapy on Tendon Healing in Animals and Humans: A Systematic Review.

Low-level Laser Therapy (LLLT) was widely used in clinical practice for tendon disorders. However, the underlying mechanisms and effectiveness of LLLT in treating tendon injury remain unclear. Therefore, the present study was conducted aiming to summarize the evidence regarding the histological, physiological, and biomechanical effects of LLLT on tendon healing in animal and human models. Four databases were searched for relevant literature. Four independent reviewers screened abstracts and full-text articles, extracted relevant data, evaluated the risk of bias, and quantified the quality of evidence. Database searches yielded 1400 non-duplicated citations. Fifty-five studies were included (50 animal and five human studies). Animal studies revealed that LT had stimulating effects on collagen organization, collagen I and collagen II formation, matrix metalloproteinase (MMP)-8, transforming growth factor ß1, vascular endothelial growth factor, hydroxyproline, maximum load, maximum elongation before breaking, and tendon stiffness. However, LLLT had inhibitory effects on the number of inflammatory cells, histological scores, relative amount of collagen III, cyclooxygenase-2, prostaglandin E2 (PGE2), interleukin-6, tumor necrosis factor-α, MMP-1, and MMP-3. Although one human study found that LLLT reduced the concentration of PGE2 in peritendinous tissue of the Achilles tendon, other human studies revealed that the effects of LLLT on the physiology and biomechanics of human tendons remained uncertain. LLLT facilitates tendon healing through various histological, physiological, and biomechanical effects in animal models. Only post-LLLT anti-inflammatory effects were found in human studies.

Inhibition of Microsomal Prostaglandin E2 Synthase Reduces Collagen Deposition in Melanoma Tumors and May Improve Immunotherapy Efficacy by Reducing T-cell Exhaustion.

The arachidonic acid pathway participates in immunosuppression in various types of cancer. Our previous observation detailed that microsomal prostaglandin E2 synthase 1 (mPGES-1), an enzyme downstream of cyclooxygenase 2 (COX-2), limited antitumor immunity in melanoma; in addition, genetic depletion of mPGES-1 specifically enhanced immune checkpoint blockade therapy. The current study set out to distinguish the roles of mPGES-1 from those of COX-2 in tumor immunity and determine the potential of mPGES-1 inhibitors for reinforcing immunotherapy in melanoma. Genetic deletion of mPGES-1 showed different profiles of prostaglandin metabolites from that of COX-2 deletion. In our syngeneic mouse model, mPGES-1-deficient cells exhibited similar tumorigenicity to that of COX-2-deficient cells, despite a lower ability to suppress PGE2 synthesis by mPGES-1 depletion, indicating the presence of factors other than PGE2 that are likely to regulate tumor immunity. RNA-sequencing analysis revealed that mPGES-1 depletion reduced the expressions of collagen-related genes, which have been found to be associated with immunosuppressive signatures. In our mouse model, collagen was reduced in mPGES-1-deficient tumors, and phenotypic analysis of tumor-infiltrating lymphocytes indicated that mPGES-1-deficient tumors had fewer TIM3+ exhausted CD8+ T cells compared with COX-2-deficient tumors. CAY10678, an mPGES-1 inhibitor, was equivalent to celecoxib, a selective COX-2 inhibitor, in reinforcing anti-PD-1 treatment. Our study indicates that mPGES-1 inhibitors represent a promising adjuvant for immunotherapies in melanoma by reducing collagen deposition and T-cell exhaustion. Significance: Collagen is a predominant component of the extracellular matrix that may influence the tumor immune microenvironment for cancer progression. We present here that mPGES-1 has specific roles in regulating tumor immunity, associated with several collagen-related genes and propose that pharmacologic inhibition of mPGES-1 may hold therapeutic promise for improving immune checkpoint-based therapies.

Protective mechanisms of Tuina therapy against lipopolysaccharide-induced fever in young rabbits based on untargeted metabolomics analysis.

OBJECTIVE: To investigate the effect of Tuina on the plasma metabolites of lipopolysaccharide-induced febrile in infant rabbits. METHODS: Twenty-four infant New Zealand rabbits were selected and randomly divided into three groups: saline, model, and Tuina. The fever model was established by injecting LPS intravenously through the ear margin vein in the model group and Tuina group, respectively. The modeling was considered successful when the anal temperature increased by 0.5℃ or above within 1 h. In the Tuina group, six Tuina techniques (i.e., opening Tianmen / the heaven gate, pushing Kangong / the superciliary arch, kneading Taiyang and the prominent bone behind the ears, clearing Tianheshui, spine pinching) that alleviate fever were performed on the young rabbits 1 h after the modeling, whereas the model and saline groups were not given Tuina treatment, with the real-time anal temperature monitored during the experiment. The plasma was taken 3 h after the modeling for liquid chromatography-mass spectrometry (LC-MS) untargeted metabolomics study. RESULTS: Our results showed a fever-reducing effects of Tuina therapy on lipopolysaccharide-induced fever in young rabbits, as indicated by a significantly lower anal temperature, maximum rise in body temperature, and body response index at 2 and 3 h after modeling in the Tuina group compared to the model group, with reductions in the PGE2 expression observed in the blood and hypothalamus. The differential metabolites including riboflavin, nicotinamide N-oxide, porphobilinogen, 5-hydroxyindoleacetic acid, gamma-aminobutyric acid, and lysoPC (16:1 (9Z)/0:0) were found following the Tuina intervention. Tuina primarily involves glycine-serine-threonine, arginine-proline, porphyrin-chlorophyll, pyrimidine, primary bile acid biosynthesis, and cyanoamino acid metabolic pathways. CONCLUSION: Tuina therapy has proven to be effective in reducing body temperature and down-regulating PGE2 expression in LPS-induced febrile young rabbits, with its mechanism of fever-reducing action possibly associated with the changes in plasma metabolites and metabolic pathways.

An in vitro anti-inflammatory effect of Thai propolis in human dental pulp cells.

OBJECTIVE: To explore the potential for development of Thai propolis extract as a pulp capping agent to suppress pulpal inflammation from dental pulp infections. This study aimed to examine the anti-inflammatory effect of the propolis extract on the arachidonic acid pathway, activated by interleukin (IL)-1ß, in cultured human dental pulp cells. METHODOLOGY: Dental pulp cells, isolated from three freshly extracted third molars, were first characterized for their mesenchymal origin and treated with 10 ng/ml of IL-1ß in the presence or absence of non-toxic concentrations of the extract from 0.08 to 1.25 mg/ml, as determined by the PrestoBlue cytotoxic assay. Total RNA was harvested and analyzed for mRNA expressions of 5-lipoxygenase (5-LOX) and cyclooxygenase-2 (COX-2). Western blot hybridization was performed to investigate COX-2 protein expression. Culture supernatants were assayed for released prostaglandin E2 levels. Immunofluorescence was conducted to determine involvement of nuclear factor-kappaB (NF-kB) in the inhibitory effect of the extract. RESULTS: Stimulation of the pulp cells with IL-1ß resulted in the activation of arachidonic acid metabolism via COX-2, but not 5-LOX. Incubation with various non-toxic concentrations of the propolis extract significantly inhibited upregulated COX-2 mRNA and protein expressions upon treatment with IL-1ß (p<0.05), resulting in a significant decrease in elevated PGE2 levels (p<0.05). Nuclear translocation of the p50 and the p65 subunits of NF-kB upon treatment with IL-1ß was also blocked by incubation with the extract. CONCLUSIONS: Upregulated COX-2 expression and enhanced PGE2 synthesis upon treatment with IL-1ß in human dental pulp cells were suppressed by incubation with non-toxic doses of Thai propolis extract via involvement of the NF-kB activation. This extract could be therapeutically used as a pulp capping material due to its anti-inflammatory properties.

Targeting PTGES/PGE2 axis enhances sensitivity of colorectal cancer cells to 5-fluorouracil.

Insensitivity and resistance to 5-fluorouracil (5FU) remain as major hurdles for effective and durable 5FU-based chemotherapy in colorectal cancer (CRC) patients. In this study, we identified prostaglandin E synthase (PTGES)/prostaglandin E2 (PGE2) axis as an important regulator for 5FU sensitivity in CRC cells. We found that PTGES expression and PGE2 production are elevated in CRC cells in comparison to normal colorectal epithelial cells. Depletion of PTGES significantly enhanced the inhibitory effect of 5FU on CRC cell viability that was fully reverted by exogenous supplement of PGE2. Inhibition of PTGES enzymatic function, by either inducing loss-of-function mutant or treatment with selective inhibitors, phenocopied the PTGES depletion in terms of 5FU sensitization. Mechanistically, PTGES/PGE2 axis modulates glycolysis in CRC cells, thereby regulating the 5FU sensitivity. Importantly, high PTGES expression is correlated with poor prognosis in 5FU-treated CRC patients. Thus, our study defines PTGES/PGE2 axis as a novel therapeutic target for enhancing the efficacy of 5FU-based chemotherapy in CRC.

Conjugated linoleic acid supplementation changes prostaglandin concentration ratio and alters the expression of genes involved in maternal-fetal recognition from bovine trophoblast cells in vitro.

Early embryonic mortality caused by maternal-fetal recognition failure in the three weeks after fertilization represents a major cause of reproductive inefficiency in the cattle industry. Modifying the amounts and ratios of prostaglandin (PG) F2α and PGE2 can benefit the establishment of pregnancy in cattle. Adding conjugated linoleic acid (CLA) to endometrial and fetal cells culture affects PG synthesis, but its effect on bovine trophoblast cells (CT-1) is unknown. The aim of this study was to determine the effects of CLA (a mixture of cis- and trans-9, 11- and -10,12-octadecadienoic acids) on PGE2 and PGF2α synthesis and the expression of transcripts involved with maternal-fetal recognition of bovine trophectoderm. Cultures of CT-1 were exposed to CLA for 24, 48 and 72 h. Transcript abundance was determined by qRT-PCR and hormone profiles were quantified by ELISA. The PGE2 and PGF2α concentrations were reduced in the culture medium of CLA-exposed CT-1 compared to that of unexposed cells. Furthermore, CLA supplementation increased the PGE2:PGF2α ratio in CT-1 and had a quadratic effect (P < 0.05) on the relative expression of MMP9, PTGES2, and PTGER4. The relative expression levels of PTGER4 were reduced (P < 0.05) in CT-1 cultured with 100 µM CLA than in the unsupplemented and 10 µM-CLA groups. Treatment of CT-1 with CLA decreased PGE2 and PGF2α synthesis but a biphasic effect of CLA was observed on the PGE2:PGF2α ratio and relative abundance of transcripts with 10 µM CLA providing maximal improvements in each endpoint. Our data suggest that CLA may influence eicosanoid metabolic process and extracellular matrix remodeling.

Molecular docking and anti-ulcerative potential of Cucumis (L. Inodorous) on ibuprofen induced gastric ulceration in male wistar animals.

BACKGROUND: The use of NSAIDs have caused stomach injury by inhibiting endogenous mucosal prostaglandin production. Cucumis melo is reported to possess antiulcer potential. This study investigates the mechanism underlying the antiulcer potentials of Cucumis melo (CUM). METHODS: Thirty-five male Wistar rat were randomly assigned to each of seven groups; A(control given water and rat pellets), B(gastric ulcer induced with ibuprofen 400 mg/kg), C (Misoprotol 200 µg/kg), D to G (pretreated with different variation of CUM extract; 25 %, 50 %, 75 % and 100 % at a dose of 1 ml/kg for 3 weeks prior to gastric ulcer induction). Ulcer score, ulcer index and percentage inhibition, total gastric acidity was measured. Antioxidant activities, Malondialdehyde, H+/K+ ATPase, PGE2, TNF-α was done by spectrophotometry. Molecular docking investigation of Cucumis melo compounds against Prostaglandin E2 was carried out. Level of significance was tested at P ≤ 0.05 using Tukey post hoc. RESULT: Total gastric acidity, ulcer score, ulcer index, MDA, TNF-α significantly decreased after CUM treatment when compared to group B. The percentage inhibition, antioxidant activities, PGE2 concentration was significantly increased in all treatment groups compared to group B. Interactions of selected compounds of CUM with Prostaglandin E2 at various docking pockets showed folic acid has highest binding affinity followed by delta7-avenasterol and codisterol to PGE2 receptor. this study shows that one of the mechanisms by which CUM exhibits its antiulcer potential by enhancing Prostaglandin synthesis and antioxidant capacity. Therefore, Cucumis melo can therefore be explored as novel antiulcer agents.

AAV-glycine receptor α3 alleviates CFA-induced inflammatory pain by downregulating ERK phosphorylation and proinflammatory cytokine expression in SD rats.

BACKGROUND: Glycine receptors (GlyRs) play key roles in the processing of inflammatory pain. The use of adeno-associated virus (AAV) vectors for gene therapy in human clinical trials has shown promise, as AAV generally causes a very mild immune response and long-term gene transfer, and there have been no reports of disease. Therefore, we used AAV for GlyRα1/3 gene transfer in F11 neuron cells and into Sprague-Dawley (SD) rats to investigate the effects and roles of AAV-GlyRα1/3 on cell cytotoxicity and inflammatory response. METHODS: In vitro experiments were performed using plasmid adeno-associated virus (pAAV)-GlyRα1/3-transfected F11 neurons to investigate the effects of pAAV-GlyRα1/3 on cell cytotoxicity and the prostaglandin E2 (PGE2)-mediated inflammatory response. In vivo experiment, the association between GlyRα3 and inflammatory pain was analyzed in normal rats after AAV-GlyRα3 intrathecal injection and after complete Freund's adjuvant (CFA) intraplantar administration. Intrathecal AAV-GlyRα3 delivery into SD rats was evaluated in terms of its potential for alleviating CFA-induced inflammatory pain. RESULTS: The activation of mitogen-activated protein kinase (MAPK) inflammatory signaling and neuronal injury marker activating transcription factor 3 (ATF-3) were evaluated by western blotting and immunofluorescence; the level of cytokine expression was measured by ELISA. The results showed that pAAV/pAAV-GlyRα1/3 transfection into F11 cells did not significantly reduce cell viability or induce extracellular signal-regulated kinase (ERK) phosphorylation or ATF-3 activation. PGE2-induced ERK phosphorylation in F11 cells was repressed by the expression of pAAV-GlyRα3 and administration of an EP2 inhibitor, GlyRαs antagonist (strychnine), and a protein kinase C inhibitor. Additionally, intrathecal AAV-GlyRα3 administration to SD rats significantly decreased CFA-induced inflammatory pain and suppressed CFA-induced ERK phosphorylation, did not induce obvious histopathological injury but increased ATF-3 activation in dorsal root ganglion (DRGs). CONCLUSIONS: Antagonists of the prostaglandin EP2 receptor, PKC, and glycine receptor can inhibit PGE2-induced ERK phosphorylation. Intrathecal AAV-GlyRα3 administration to SD rats significantly decreased CFA-induced inflammatory pain and suppressed CFA-induced ERK phosphorylation, did not significantly induce gross histopathological injury but elicited ATF-3 activation. We suggest that PGE2-induced ERK phosphorylation can be modulated by GlyRα3, and AAV-GlyRα3 significantly downregulated CFA-induced cytokine activation.

Exploring the anti-inflammatory potential of Colocasia esculenta root extract in in-vitro and in-vivo models of inflammation.

ETHNOPHARMACOLOGICAL RELEVANCE: Colocasia esculenta (CE) (L.) Schott is an annual herbaceous tropical plant from the family of Araceae which has been traditionally used for the healing of various ailments such as asthma, arthritis, internal hemorrhage, diarrhea, and neurological disorders. The plant is reported to have potential anti-microbial, anti-fungal, antimetastatic, anti-hepatotoxic, and anti-lipid peroxidative activities. AIM OF THE STUDY: The present study is designed to explore the potential anti-inflammatory property of Colocasia esculenta methanolic root extract (CEMRE) on carrageenan-induced rat paw edema and lipopolysaccharide (LPS) stimulated RAW264.7 cells. MATERIALS AND METHODS: Carrageenan-induced rat paw edema model was used to investigate the in vivo anti-inflammatory action of CEMRE. Adult male Wistar rats (180-220 g; n = 6) were pre-treated with CEMRE (100, 200, and 400 mg/kg BW) orally before 1 h of injection of 1% carrageenan. Indomethacin (10 mg/kg BW) was given orally as the standard drug. Serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), nitric oxide (NO), prostaglandinE2 (PGE2), and cytokines levels were measured. Liquid chromatography-mass spectrometry (LC-MS) was done to identify the phytoconstituents present in CEMRE. The inhibitory activity of CEMRE was investigated against cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) in in vitro assessment of LPS-stimulated RAW264.7 cells. The RAW 264.7 cells were pre-treated with Indomethacin (5 µM and 10 µM) and CEMRE (17 µg/ml and 34 µg/ml) followed by induction of LPS (1 µg/ml) for 24 h. Docking analyses were also performed to explore the interaction of important phytoconstituents (Sinapic acid, Acetylsalicylic acid, L-fucose, Salicylic acid, Quinic acid, Zingerone, and Gingerol) of CEMRE with COX-2 and iNOS. RESULTS: Pre-treatment with CEMRE (400 mg/kg) could inhibit the paw inflammation significantly which was elevated due to carrageenan induction. The inhibition is comparable to that of the standard drug Indomethacin. The concentration of serum AST, ALT, ALP, NO, PGE2 and cytokines were also considerably lowered in the CEMRE-treated group as compared to the carrageenan-induced group. CEMRE (34 µg/ml) inhibited the LPS-stimulated relative expression of mRNA of COX-2 and iNOS and significantly reduced the expression of nitric oxide and prostaglandin E2. Docking analyses revealed promising interaction with low binding energies between Sinapic acid with both the target proteins COX-2 and iNOS. CONCLUSION: Collectively, our results suggested that CEMRE exhibited effective anti-inflammatory actions on carrageenan-induced rat paw edema and LPS-treated RAW 264.7 cells by reducing the in vivo paw edema inhibition, inhibiting the serum NO, PGE2, cytokines and also reduced the in vitro production of NO, PGE2 along with expressions of mRNA COX-2 and iNOS. Molecular docking demonstrated good binding affinities among the target proteins and ligand Sinapic acid. Thus the bioactive compound from CE need to be isolated and purified.

Hedychium flavum flower essential oil: Chemical composition, anti-inflammatory activities and related mechanisms in vitro and in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: Hedychium flavum, an ornamental, edible, and medicinal plant, is extensively cultivated as a source of aromatic essential oils (EO). Its flower is a traditional Chinese medicine for treating inflammation-related diseases like indigestion, diarrhea, and stomach pain. In particular, H. flavum flower EO has been used in cosmetics and as an aromatic stomachic to treat chronic gastritis in China. AIM OF THE STUDY: This research aimed to analyze H. flavum flower EO's chemical composition and explore its anti-inflammatory activities and related mechanisms in vitro and in vivo. MATERIALS AND METHODS: EO's chemical composition was determined by GC-FID/MS analysis. For in vitro test, the anti-inflammatory activity of EO was demonstrated by measuring the LPS-induced release of NO, PGE2, IL-1ß, TNF-α, and IL-6 in RAW264.7 macrophages, and then its related mechanisms were explored using qRT-PCR, western blot, and immunofluorescent staining analysis. Next, EO's in vivo anti-inflammatory potential was further evaluated using a xylene-induced ear edema model, in which ear swelling and TNF-α, IL-6, and IL-1ß levels in serum and tissue were examined. RESULTS: The main components of EO were ß-pinene (20.2%), α-pinene (9.3%), α-phellandrene (8.3%), 1,8-cineole (7.1%), E-nerolidol (5.4%), limonene (4.4%), borneol (4.1%), and ß-caryophyllene (3.7%). For the anti-inflammatory activities in vitro, EO dramatically reduced the LPS-stimulated NO and PGE2 release by suppressing the mRNA and protein expression of iNOS and COX-2. Meanwhile, it remarkably decreased IL-6, TNF-α, and IL-1ß production by inhibiting their mRNA levels. Related mechanism studies indicated that it not only inhibited IκBα phosphorylation and degradation, leading to blockade of NF-κB nuclear transfer but also suppressed MAPKs (ERK, p38, and JNK) phosphorylation in LPS-stimulated RAW264.7 cells. Further in vivo assay showed that EO ameliorated xylene-induced ear edema in mice and reduced TNF-α, IL-6, and IL-1ß levels in serum and tissue. CONCLUSIONS: H. flavum EO exerted significant anti-inflammatory activity in vivo and in vitro, and its mechanism of action is related to the inhibition of MAPK and NF-κB activation. Thus, H. flavum EO could be considered a novel and promising anti-inflammatory agent and possess high potential for utilization in the pharmaceutical field.

Transgenic Rice Seed Extracts Exert Immunomodulatory Effects by Modulating Immune-Related Biomarkers in RAW264.7 Macrophage Cells.

Protopanaxadiol (PPD), a native active triterpenoid present in Panax ginseng, has been reported to exert immune-related effects. We previously created PPD-producing transgenic rice by introducing the P. ginseng protopanaxadiol synthase and dammarenediol-II synthase genes into Dongjin rice. In the present study, the seeds of the T4 generation of this transgenic rice were tested for their immunomodulatory effects in RAW264.7 macrophage cells. Treatment with transgenic rice seed extract in RAW264.7 cells (i) significantly enhanced nitric oxide (NO) production in a dose-dependent manner without any cytotoxicity (up to 100 µg/mL), (ii) upregulated the expression of immune-related genes and increased production of the inflammation mediator prostaglandin E2 (PGE2), and (iii) activated nuclear factor-κB (NF-κB) and mitogen-activated protein kinase (MAPK) by promoting the phosphorylation of NF-κB p65, p38 MAPK, and c-Jun N-terminal protein kinase (JNK). In lipopolysaccharide (LPS)-treated RAW264.7 cells used to mimic the inflammation condition, treatment with transgenic rice seed extract significantly reduced NO production, proinflammatory cytokine expression, and PGE2 production, all of which are LPS-induced inflammation biomarkers, by inhibiting the phosphorylation of NF-κB p65, p38 MAPK, and JNK. Collectively, these results indicate that PPD-producing transgenic rice has immunomodulatory effects.

Akebia Saponin D Inhibits the Inflammatory Reaction by Inhibiting the IL-6-STAT3-DNMT3b Axis and Activating the Nrf2 Pathway.

Akebia saponin D (ASD) is derived from the Dipsacus asper Wall. ex Henry, which is a traditional Chinese medicine commonly used to treat rheumatic arthritis (RA). However, the in-depth mechanism of the anti-inflammatory effect of ASD is still unclear. This study aimed to preliminarily explore the anti-inflammatory effect of ASD and the underlying mechanisms from the perspective of DNA methylation and inflammation-related pathways. We found that ASD significantly reduced the production of multiple inflammatory mediators, including nitric oxide (NO) and prostaglandin E2 (PGE2), in LPS-induced RAW264.7 cells. The expression of DNA methyltransferase (DNMT) 3b and inducible nitric oxide synthase (iNOS) was also obviously inhibited by the ASD treatment. The protein and mRNA levels of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) were also significantly inhibited by ASD. ASD inhibited the macrophage M1 phenotype, inhibited the high level of DNMT3b, and downregulated the signal transducer and activator of the transcription 3 (STAT3) pathway to exert its anti-inflammatory activity. Furthermore, DNMT3b siRNA and Nrf2 siRNA significantly promoted the anti-inflammatory effect of ASD. Our study demonstrates for the first time that ASD inhibits the IL-6-STAT3-DNMT3b axis and activates the nuclear factor-E2-related factor 2 (Nrf2) signaling pathway to achieve its inhibitory effect on inflammatory reactions.