Ge-gen decoction alleviates primary dysmenorrhoea symptoms in a rat model.

BACKGROUND: Existing treatments for primary dysmenorrhoea (PD), such as NSAIDs, impart side effects. Ge-Gen decoction (GGD), a traditional Chinese medicine, has shown promise in treating PD, but its exact mechanisms remain unclear. Here, we aimed to investigate the efficiency of GGD in alleviating PD using a rat model to understand its precise mechanism of action. METHODS: We established a rat model of dysmenorrhoea induced by oestradiol and oxytocin. The PD rats were administered GGD or Ibuprofen (positive control) intragastrically once daily for seven consecutive days. Serum levels of prostaglandin E2 (PGE2), prostaglandin F2 alpha (PGF2α), ß-endorphin (ß-EP), thromboxane B2 (TXB2), 6-keto-prostaglandin F1α (6-keto-PGF1α) were determined using an enzyme-linked immunosorbent assay (ELISA). The expression levels of oestrogen receptor alpha (ERα) and cyclooxygenase-2 (COX-2) in uterine tissue were measured using immunohistochemical assays, and those of phosphorylated and total extracellular signal-regulated protein kinases 1 and 2 (ERK1/2) were assessed using western blot analysis. RESULTS: Treatment with GGD significantly reduced writhing behaviour, histopathological scores, and levels of COX-2, PGE2, and PGF2α in the serum of PD rats. Additionally, GGD increased ß-EP content and inhibited ERK1/2 activation and ERα expression in uterine tissues. CONCLUSIONS: The results of this study suggest that GGD alleviates PD in rats by suppressing the COX-2-mediated release of PGE2 and PGF2α, modulating the ERα/ERK1/2/COX-2 pathway, and increasing ß-EP content. These results provide insights into the potential mechanisms of GGD in treating PD and support its further investigation as an alternative therapy for this condition.

Ge-Gen decoction is commonly used to alleviate primary dysmenorrhoea. However, its anti-dysmenorrhoea mechanism remains elusive. In this study, using a rat model of primary dysmenorrhoea, we demonstrate that Ge-Gen decoction reduced the levels of cyclooxygenase-2, prostaglandin E2, and prostaglandin F2 alpha in serum and phosphorylated extracellular signal-regulated protein kinases 1 and 2 in the uterus. These results suggest that Ge-Gen decoction alleviates primary dysmenorrhoea via inactivation of the oestrogen receptor alpha/extracellular signal-regulated protein kinases 1 and 2/cyclooxygenase-2 pathway. This study enhances our understanding of the pathogenesis of primary dysmenorrhoea and may potentially inform the development of novel treatment approaches.

Comparative Analysis of Phytochemical Profiles and Selected Biological Activities of Various Morphological Parts of Ligustrum vulgare.

Ligustrum vulgare (LV), widely cultivated in Europe and often used in hedges, has been historically recognized in folk medicine for its potential health benefits. This study focused on exploring the untargeted identification of secondary metabolites in ethanol extracts (70% v/v) from different morphological parts (young shoots, leaves, flowers and fruits) of LV at various stages of plant development, using ultra-high-performance liquid chromatography with high-resolution mass spectrometry (UHPLC-HRMS). Additionally, the selected biological activities (antioxidant activity, cyclooxygenase-2 inhibition (COX-2), α-amylase inhibition and cytotoxicity) of the tested extracts were determined. Untargeted metabolomics showed that LV extracts were a rich source of phenylethanoid compounds, flavonoids, iridoids and their derivatives. The flowers of LV had the highest content of oleuropein (33.43 ± 2.48 mg/g d.w.). The lowest antioxidant activity was obtained for ripe and post-seasonal fruits, while in the case of other samples, the activity was at a similar level. All tested extracts showed α-amylase and COX-2 inhibitory activity. In addition, LV extracts showed strong antiproliferative properties in colorectal (HT29) and liver (HepG2) cancer cell lines. The obtained results show the difference in the content of bioactive compounds in various morphological parts of Ligustrum vulgare. These differences may influence the multifaceted medicinal potential of this plant.

Target-guided isolation and purification of cyclooxygenase-2 inhibitors from Meconopsis integrifolia (Maxim.) Franch. by high-speed counter-current chromatography combined with ultrafiltration liquid chromatography.

Meconopsis integrifolia (Maxim.) Franch. is used extensively in traditional Tibetan medicine for its potent anti-inflammatory properties. In this study, six cyclooxygenase-2 (COX-2) inhibitors were purified from M. integrifolia using high-speed counter-current chromatography guided by ultrafiltration liquid chromatography (ultrafiltration-LC). First, ultrafiltration-LC was performed to profile the COX-2 inhibitors in M. integrifolia. The reflux extraction conditions were further optimized using response surface methodology, and the results showed that the targeted COX-2 inhibitors could be well enriched under the optimized extraction conditions. Then the six target COX-2 inhibitors were separated by high-speed countercurrent chromatography with a solvent system composed of ethyl acetate/n-butanol/water (4:1:4, v/v/v. Finally, the six COX-2 inhibitors, including 21.2 mg of 8-hydroxyluteolin 7-sophoroside, 29.6 mg of 8-hydroxyluteolin 7-[6'''-acetylallosyl-(1→2)-glucoside], 42.5 mg of Sinocrassoside D3, 54.1 mg of Hypolaetin 7-[6'''-acetylallosyll-(l→2)-3''-acetylglucoside, 30.6 mg of Hypolaetin 7-[6'''-acetylallosyll-(l→2)-6''-acetylglucoside and 17.8 mg of Hypolaetin were obtained from 500 mg of sample. Their structures were elucidated by 1 H-NMR spectroscopy. This study reveals that ultrafiltration-LC combined with high-speed counter-current chromatography is a robust and efficient strategy for target-guided isolation and purification of bioactive molecules. It also enhances the scientific understanding of the anti-inflammatory properties of M. integrifolia but also paves the way for its further medicinal applications.

Comparative study of the antioxidant and anti-inflammatory effects of the natural coumarins 1,2-benzopyrone, umbelliferone and esculetin: in silico, in vitro and in vivo analyses.

The aim of this work was to compare the anti-inflammatory and antioxidant effects of three natural coumarins: 1,2-benzopyrone, umbelliferone and esculetin. The antioxidant capacity of coumarins was evaluated using both chemical and biological in vitro assays. Chemical assays included DPPH and ABTS∙+ radical scavenging as well as ferric ion reducing ability power (FRAP) assay. Inhibition of mitochondrial ROS generation and lipid peroxidation in brain homogenates were used as biological in vitro assays. The experimental method of carrageenan-induced pleurisy in rats was used for the in vivo investigation of the anti-inflammatory activity. In silico molecular docking analysis was undertaken to predict the affinity of COX-2 to the coumarins. Considering the antioxidant capacity, esculetin was the most efficient one as revealed by all employed assays. Particularly, the mitochondrial ROS generation was totally abolished by the compound at low concentrations (IC50 = 0.57 µM). As for the anti-inflammatory effects, the COX-2 enzyme presented good affinities to the three coumarins, as revealed by the molecular docking analyses. However, considering the in vivo anti-inflammatory effects, 1,2-benzopyrone was the most efficient one in counteracting pleural inflammation and it potentiated the anti-inflammatory actions of dexamethasone. Umbelliferone and esculetin treatments failed to reduce the volume of pleural exudate. Overall, therefore, our results support the notion that this class of plant secondary metabolites displays promising effects in the prevention and/or treatment of inflammation and other diseases associated with oxidative stress, although the singularities regarding the type of the inflammatory process and pharmacokinetics must be taken into account.

2-Hydroxybenzohydrazide as a novel potential candidate against nociception, inflammation, and pyrexia: in vitro, in vivo, and computational approaches.

The current study was designed to evaluate the 2-hydroxybenzohydrazide (HBH) as a drug having efficacy against pyrexia, inflammation, and nociception. Besides, the therapeutic effects of HBH on oxidative stress and C-reactive proteins were also evaluated. The pharmacological studies on HBH (20-60 mg/kg) were conducted using nociception, inflammation, and pyrexia standard models. Naloxone antagonism was performed to assess the possible involvement of opioidergic mechanisms. The antioxidant study was conducted on ABTS and DPPH assays using gallic acid as a standard. Moreover, the binding capability of HBH with enzymes cyclooxygenase-I/II (COX-I/II) was determined using molecular modeling analysis. The findings indicated that the HBH dose-dependently inhibited pain, inflammation, and pyrexia. The HBH has significant anti-nociceptive and anti-inflammatory activities at 60 mg/kg (***p < 0.001), similar to the lower doses of diclofenac sodium (50 mg/kg) and tramadol (30 mg/kg). The HBH at 60 mg/kg reduced pyrexia as paracetamol (150 mg/kg). The HBH at 20-60 mg/kg doses declined the plasma C-reactive protein concentration. The mechanistic studies showed that the anti-nociceptive effect of HBH was antagonized by naloxone, indicating that the opioidergic mechanisms are involved. Furthermore, computational studies showed that the HBH exhibited an affinity for COX-I/II target receptors. The HBH significantly inhibited ABTS and DPPH radicals (IC50 = 33.81 and 26.74 µg/ml). These results proposed that the HBH has significant antipyretic, anti-inflammatory, and anti-nociceptive activities involving opioidergic mechanism.

In Vitro Analysis of Selected Antioxidant and Biological Properties of the Extract from Large-Fruited Cranberry Fruits.

This study investigated the in vitro antioxidant and biological properties of ethanol extracts obtained from the fruits of the highbush cranberry. The produced extracts exhibited a high content of polyphenols (1041.9 mg 100 g d.m.-1) and a high antioxidant activity (2271.2 mg TE g 100 d.m.-1 using the DPPH method, 1781.5 mg TE g 100 d.m.-1 using the ABTS method), as well as a substantial amount of vitamin C (418.2 mg 100 g d.m.-1). These extracts also demonstrated significant in vitro biological activity. Studies conducted on the Saccharomyces cerevisiae cellular model revealed the strong antioxidant effects of the extract, attributed to a significant reduction in the levels of reactive oxygen species (ROS) within the cells, confirming the utility of the extracts in mitigating oxidative stress. Moreover, inhibitory properties were demonstrated against factors activating metabolic processes characteristic of inflammatory conditions. It was observed that the cranberry extract inhibits the activity of cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) non-selectively. Additionally, the extract was found to be a highly active inhibitor of acetylcholinesterase (AChE), potentially suggesting the applicability of this extract in the prevention of neurodegenerative diseases, including Alzheimer's disease.

Effect of digoxin, sodium valproate, and celecoxib on the cerebral cyclooxygenase pathway and neuron-specific enolase under the pentylenetetrazole-induced kindling in mice.

Neuroinflammation plays an important role in the pathogenesis of epilepsy, so it is necessary to clarify the influence of standard antiepileptic drugs as well as adjuvant agents (e.g., cardiac glycoside digoxin, which previously showed a clear anticonvulsant potential) on cyclooxygenase pathway and neuron-specific enolase under the conditions of chronic epileptogenesis. The aim of the article is to determine the effect of digoxin, sodium valproate, and celecoxib per se, as well as the combination of digoxin with sodium valproate on the content of cyclooxygenase 1 and 2 types, prostaglandins E2, F2α, I2, thromboxane B2, 8-isoprostane and neuron-specific enolase in the brain of mice in the pentylenetetrazole-induced kindling model. It was found that only the combination of sodium valproate with digoxin provides a complete protective effect (absence of seizures) and shows the clearest influence on neuroinflammation markers and neuronal damage than monotherapy with each of these drugs and celecoxib, which appeared to be an ineffective anticonvulsant. The obtained results indicate that digoxin is a promising adjuvant drug to classical antiepileptic drugs (mostly sodium valproate) in epilepsy treatment.c.

Sirtuin 6 ameliorates arthritis through modulating cyclic AMP-responsive element binding protein/CCN1/cyclooxygenase 2 pathway in osteoblasts.

INTRODUCTION: CCN1 is an immediate-early gene product pivotal for arthritis progression. We have previously shown that sirtuin 6 (SIRT6) inhibited hypoxia-induced CCN1 expression in osteoblasts. Herein we examined the contribution of cyclic AMP-responsive element binding protein (CREB)/CRE to this suppressive action and the influence of CCN1 on cyclooxygenase (COX) 2 synthesis. MATERIALS AND METHODS: MC3T3-E1 murine osteoblasts were cultured under normoxia (21% oxygen) or hypoxia (2% oxygen). Expressions of CCN1, phospho-CREB (Ser133), COX2 and relevant kinases were assessed by Western blot. SIRT6 was overexpressed in cultured osteoblasts and arthritic joints by a lentiviral-based technique. Activities of CCN1 gene promoter constructs were examined by luciferase reporter assay. Interaction between CREB and CCN1 promoter was assessed by chromatin immunoprecipitation (ChIP). Collagen-induced arthritis (CIA) was established in 20 rats to evaluate the effects of SIRT6 therapy on osteoblastic expressions of phospho-CREB, CCN1 and COX2. RESULTS: SIRT6 suppressed hypoxia-enhanced CCN1 expression and CREB phosphorylation. Attenuation of calcium/calmodulin-dependent protein kinase II (CaMKII) may be responsible for SIRT6-induced CREB inhibition. CRE at - 286 bp upstream of the ATG start codon was essential for CCN1 expression under hypoxia and SIRT6 reduced hypoxia-stimulated CREB/CRE interaction. Forced expression of CREB rescued SIRT6-suppressed CCN1 synthesis. CCN1 induced COX2 expression in osteoblasts. In rat CIA, the therapeutic effect of SIRT6 was accompanied by decreases in osteoblastic expressions of phospho-CREB, CCN1 and COX2. CONCLUSION: Our study indicated that the benefits of SIRT6 to inflammatory arthritis and bone resorption are at least partially derived from its modulation of CREB/CCN1/COX2 pathway in osteoblasts.

Trilobatin rescues fulminant hepatic failure by targeting COX2: Involvement of ROS/TLR4/NLRP3 signaling.

BACKGROUND: Fulminant hepatic failure (FHF) lacks efficient therapies notwithstanding increased comprehending of the inflammatory response and oxidative stress play crucial roles in the pathogenesis of this type of hepatic damage. Trilobatin (TLB), a naturally occurring food additive, is endowed with anti-inflammation and antioxidant properties. PURPOSE: In current study, we evaluated the effect of TLB on FHF with a mouse model with d-galactosamine/lipopolysaccharide (GalN/LPS)-induced FHF and LPS-stimulated Kupffer cells (KCs) injury. METHODS: Mice were randomly divided into seven groups: control group, TLB 40 mg/kg + control group, GalN/LPS group, TLB 10 mg/kg + GalN/LPS group, TLB 20 mg/kg + GalN/LPS group, TLB 40 mg/kg + GalN/LPS group, bifendate 150 mg/kg + GalN/LPS group. The mice were administered intragastrically TLB (10, 20 and 40 mg/kg) for 7 days (twice a day) prior to injection of GalN (700 mg/kg)/LPS (100 µg/kg). The KCs were pretreated with TLB (2.5, 5, 10 µM) for 2 h or its analogue (10 µM) or COX2 inhibitor (10 µM), and thereafter challenged by LPS (1 µg/ml) for 24 h. RESULTS: TLB effectively rescued GalN/LPS-induced FHF. Furthermore, TLB inhibited TLR 4/NLRP3/pyroptosis pathway, and caspase 3-dependent apoptosis pathway, along with reducing excessive cellular and mitochondrial ROS generation and enhancing mitochondrial biogenesis. Intriguingly, TLB directly bound to COX2 as reflected by transcriptomics, molecular docking technique and surface plasmon resonance assay. Furthermore, TLB failed to attenuate LPS-induced inflammation and oxidative stress in KCs in the absence of COX2. CONCLUSION: Our findings discover a novel pharmacological effect of TLB: protecting against FHF-induced pyroptosis and apoptosis through mediating ROS/TLR4/NLRP3 signaling pathway and reducing inflammation and oxidative stress. TLB may be a promising agent with outstanding safety profile to treat FHF.

The Interaction of Dietary Pectin, Inulin, and Psyllium with Copper Nanoparticle Induced Changes to the Cardiovascular System.

We aimed to analyze how supplementation with a standard (recommended, 6.5 mg/kg) or enhanced (two-times higher, 13 mg/kg) dose of copper (Cu), in the form of nanoparticles (NPs) along with dietary intervention via the implementation of diverse types of fiber, affects the cardiovascular system in rats. Nine-week-old male Wistar Han rats (n/group = 10) received, for an additional 6 weeks, a controlled diet with cellulose as dietary fiber and ionic Cu (in the form of carbonate salt). The experimental groups received cellulose, pectin, inulin, and psyllium as dietary fiber, together with CuNPs (6.5 or 13 mg/kg diet). After the experimental feeding, samples of blood, hearts, and thoracic arteries were collected for further analysis. Compared to pectin, and under a standard dose of CuNPs, inulin and psyllium beneficially increased the antioxidant capacity of lipid- and water-soluble compounds in the blood, and decreased heart malondialdehyde. Moreover, pectin decreased heart catalase (CAT) and cyclooxygenase (COX)-2 in the aortic rings compared to inulin and psyllium under standard and enhanced doses of copper. When the dose of CuNPs was enhanced, inulin and psyllium potentiated vasodilation to acetylcholine by up-regulation of COX-2-derived vasodilator prostanoids compared to both cellulose and pectin, and this was modulated with selective inducible nitric oxide synthase (iNOS) inhibitor for psyllium only. Moreover, inulin decreased heart CAT compared to psyllium. Our results suggest that supplementation with dietary fiber may protect the vascular system against potentially harmful metal NPs by modulating the antioxidant mechanisms.

Triterpenoid and Coumarin Isolated from Astilbe grandis with Anti-Inflammatory Effects through Inhibiting the NF-κB Pathway in LPS-Induced RAW264.7 Cells.

The roots of Astilbe grandis, known as "Ma sang gou bang", are used as a Miao traditional medicine with anti-inflammatory and analgesic properties. However, the active components and mechanism of action of this plant remain mostly uncharacterized. The aim of this study was to identify its active components and verify their pharmacological activity. The extract of A. grandis root was separated using various chromatographic methods. As a result, we obtained one novel triterpenoid, named astigranlactone (1), which has an unusual lactone moiety formed between C-7 and C-27. Additionally, a known coumarin compound, 11-O-galloyl bergenin (2) was isolated from this plant. The structures of these two compounds were elucidated by extensive NMR experiments in conjunction with HR-ESI-MS data. To the best of our knowledge, both compounds were isolated from this species for the first time. Moreover, we tested the anti-inflammation effect of the two compounds by establishing a cellular inflammation model induced by LPS in RAW264.7 cells. The effect of different concentrations of these compounds on the activity of RAW264.7 cells was assessed using a CCK8 assay. The levels of nitric oxide (NO), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1ß (IL-1ß) in the supernatant of each group were evaluated using the Griess method and an enzyme-linked immunosorbent assay (ELISA). Western blot and quantitative real-time PCR (qRT-RCR) were used to measure the levels of cyclooxygenase 2 (COX-2) and nitric oxide synthase (iNOS) gene expression. Our findings revealed that these two compounds inhibited the high levels of NO, TNF-α, IL-6, IL-1ß, COX-2, and iNOS (induced by LPS). Mechanistic studies demonstrated that these two compounds reduced the activation of the nuclear transcription factor-B (NF-κB) signaling pathway by inhibiting the phosphorylation of p65. Therefore, our study indicates that compounds 1 and 2 can exert a definite anti-inflammatory effect by inhibiting the NF-κB signaling pathway.

Ameliorating effect of polyherbal formulation (Febrojith) on acute and chronic inflammatory model via inhibiting inflammation and oxidative stress.

ETHNOPHARMACOLOGICAL RELEVANCE: Inflammation is a complex biological response of the tissue to noxious stimuli, which causes several debilitating inflammatory disorders. Currently, various conventional medicines are available, but their consumption causes adverse effects, hence researchers focused on alternatives like medical herbs from natural sources, as one of the most promising sources of therapeutic agents for inflammation. Febrojith is a well-known traditional Ayurvedic formulation obtained from the treasures of Ayurveda with a unique blend of herbs that are used effectively in preventing and combating a broad spectrum of infections, fevers, and also enhancing immunity for many years. However, its anti-inflammatory, efficacy and underlying mechanism remained unexplored. AIM OF THE STUDY: In the present study, we investigated the chemical characterization and in vivo anti-inflammatory efficacy of Febrojith (FB) on acute and chronic inflammatory models via inhibiting inflammation and oxidative stress. MATERIALS AND METHODS: FB was analyzed for chemical characterization & its phytoconstituents by UV-Vis spectrum, FT-IR, and GC-MS analysis. The anti-inflammatory activity of FB was studied on carrageenan-induced acute and adjuvant-induced chronic experimental models. The inflammatory cytokines and mediators were measured using the ELISA & Colorimetry techniques. Histopathology and cytology of paw tissue and synovium were analyzed by H&E and Papanicolau's (PAP)-staining methods. RESULTS: 100 mg/kg bwt was found to be a potent dose from the carrageenan model and evaluated its effect in the adjuvant-induced chronic arthritic model. In the chronic model, 84% of edema inhibition was observed at the dose of 100 mg/kg bwt. Moreover, the supplementation of FB was shown to significantly (p ≤ 0.05) decrease the TBARS level and activity of myeloperoxidase in the paw tissue. In addition, adjuvant-induced production of various pro-inflammatory cytokines like TNF-α, IL-1ß, IL-6, PGE2, NO and COX-2 were suppressed in inflamed rats subjected to FB supplementation. It also revealed that FB supplementation significantly (p ≤ 0.05) reduced the haematological markers. From the histopathology and cytological analysis, we found a reduction in the edema formation, and infiltration of inflammatory cells after the supplementation of FB. CONCLUSION: In conclusion, FB might be used as an effective and potent drug against inflammation.

In silico molecular docking analysis of three molecules isolated from Litsea guatemalensis Mez on anti-inflammatory receptors.

BACKGROUND: The Litsea genus has four native species from Mesoamerica. Litsea guatemalensis Mez. is a native tree, traditionally used as a condiment and herbal medicine in the region. It has demonstrated antimicrobial, aromatic, anti-inflammatory and antioxidant activity. Bioactive fractionation attributed the anti-inflammatory and anti-hyperalgesic activities to pinocembrin, scopoletin, and 5,7,3´4´-tetrahydroxy-isoflavone. In silico analysis, these molecules were analyzed on receptors involved in the anti-inflammatory process to determine which pathways they interact. OBJECTIVE: To analyze and evaluate 5,7,3',4'-tetrahydroxyisoflavone, pinocembrin, and scopoletin using the in silico analysis against selected receptors involved in the inflammatory pathway. METHOD: Known receptors involved in the anti-inflammatory process found as protein-ligand complex in the Protein Data Bank (PDB) were used as references for each receptor and compared with the molecules of interest. The GOLD-ChemScore function, provided by the software, was used to rank the complexes and visually inspect the overlap between the reference ligand and the poses of the studied metabolites. RESULTS: 53 proteins were evaluated, each one in five conformations minimized by molecular dynamics. The scores obtained for dihydroorotate dehydrogenase were greater than 80 for the three molecules of interest, scores for cyclooxygenase 1 and glucocorticoid receptor were greater than 50, and identified residues with interaction in binding sites overlap with the reference ligands in these receptors. CONCLUSION: The three molecules involved in the anti-inflammatory process of L. guatemalensis show in silico high affinity to the enzyme dihydroorotate dehydrogenase, glucocorticoid receptors and cyclooxygenase-1.

Hu'po Anshen decoction promotes fracture healing in mice with traumatic brain injury through BMP2-COX2-ATF4 signaling pathway.

Hu'po Anshen decoction (HPASD), a traditional Chinese medicine used to treat concussion and fracture, could regulate the expression of bone morphogenetic protein 2 (BMP2). However, whether HPASD affects the fracture healing of traumatic brain injury (TBI) combined with a fracture through BMP2 and its downstream signals remains obscure. The chondrocyte-specific BMP2 conditional knockout mice and chondrocyte-specific cyclooxygenase-2 (COX2) overexpression mice were generated. BMP2 conditional knockout mice were treated with fracture surgery, fracture combined with TBI, or fracture combined with TBI followed by different doses of HPASD (2.4, 4.8, and 9.6 g/kg), respectively. TBI was induced by Feeney's weight-drop technique. The fracture callus formation and fracture sites were determined by X-ray, micro-CT, and histological analyses. The expressions of chondrocyte-, osteoblast-, and BMP2/COX2 signal-related targets were determined by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and western blot assays. The specific absence of BMP2 in chondrocytes led to the prolonged formation of cartilage callus, a delay in the osteogenesis initiation and the downregulation of RUNX2, Smad1/5/9, EP4, ERK1/2, RSK2, ATF4. Overexpression of COX2 partially reverses the effects of chondrocyte-specific BMP2 knockout mice. HPASD promoted cartilage callus formation and osteogenesis initiation, as accompanied by upregulated expression levels of RUNX2, Smad1/5/9, EP4, ERK1/2, RSK2, and ATF4 in a time-dependent and concentration-dependent manner in chondrocyte-specific BMP2 knockout mice. Overall, our findings demonstrated that HPASD induced COX2 transcription through the BMP2-Smad1/5/9-RUNX2 axis, and then affected fracture healing through the COX2-mediated EP4-ERK1/2-RSK2-ATF4 axis.

Lafoensia pacari alleviates intestinal damage by modulating cyclooxygenase-2: In silico and in vivo evaluation in a colitis model.

BACKGROUND: Inflammatory bowel diseases (IBD) are a worldwide health problem and mainly affect young people, consequently affecting the workforce. Available treatments are often associated with side effects, and new therapeutic options are needed. For centuries, plants have represented important substrates in the field of drug development. Lafoensia pacari (L. pacari) is a plant whose pharmaceutical potential has been described, and may have biological activity relevant to the treatment of IBD symptoms. AIM: To investigate the activity of keto-alcoholic extracts of L. pacari with respect to ameliorating the inflammatory and nociceptive symptoms of acute experimental colitis in mice. METHODS: Keto-alcoholic extracts of L. pacari leaves and bark were administered to male and female Swiss mice weighing 25 g to 30 g (n = 8 male mice and n = 8 female mice). The effect of these extracts was observed in an acetic acid-induced acute experimental model of colitis with regard to antinociception/analgesia and inflammatory tissue damage. Recorded macroscopic indices included the Wallace score and the colon weight obtained using a precision scale. Mechanical hyperalgesia was determined using an electronic analgesimeter. Behavior related to overt pain was determined by quantifying the number of writhing instances within 20 min of administration of acetic acid. Molecular docking was performed using human and murine cyclooxygenase-2 (COX-2) with 3 flavonoids (ellagic acid, kaempferol, and quercetin) on the AutoDock Vina software. Analysis of variance followed by Tukey's posttest was used with P < 0.05 indicating significance. RESULTS: In this murine model of colitis, administration of extracts from L. pacari ameliorated acetic acid-induced writhing and colitis-associated inflammatory pain. These improvements may be attributable to the reduction in edema, inflammation (e.g., ulcers, hyperemia, and bowel wall damage), and the intensity of abdominal hyperalgesia. The keto-alcoholic extracts of L. pacari leaves and bark administered at a dose of either 100 mg/kg or 300 mg/kg significantly reduced the number of writhing events when compared to the negative control (P < 0.05). Additionally, extracts of L. pacari bark also performed better than Dipyrone. Leaf extracts administered at 10 mg/kg, 30 mg/kg, and 100 mg/kg and bark extracts administered at 30 mg/kg significantly reduced or prevented the development of edema in the colon of treated mice, while mesalazine did not. Moreover, using molecular docking, we observed that the flavonoids present in L. pacari extracts bind to COX-2, an event not unique to ellagic acid. CONCLUSION: The results of this study demonstrate a potential novel application of L. pacari extracts for the reduction of inflammation and promotion of antinociception/analgesia as demonstrated by our findings in a murine model of colitis. These findings were also corroborated by in silico analyses, and suggest that L. pacari extracts may be a promising therapeutic agent in the treatment of IBD.

Synthesis and Biological Evaluation of Gentiopicroside Derivatives as Novel Cyclooxygenase-2 Inhibitors with Anti-Inflammatory Activity.

Purpose: Nonsteroidal anti-inflammatory drugs cause a series of adverse reactions. Thus, the search for new cyclooxygenase-2 selective inhibitors have become the main direction of research on anti-inflammatory drugs. Gentiopicroside is a novel selective inhibitor of cyclooxygenase-2 from Chinese herbal medicine. However, it is highly hydrophilic owing to the presence of the sugar fragment in its structure that reduces its oral bioavailability and limits efficacy. This study aimed to design and synthesize novel cyclooxygenase-2 inhibitors by modifying gentiopicroside structure and reducing its polarity. Materials and Methods: We introduced hydrophobic acyl chloride into the gentiopicroside structure to reduce its hydrophilicity and obtained some new derivatives. Their in vitro anti-inflammatory activities were evaluated against NO, TNF-α, PGE2, and IL-6 production in the mouse macrophage cell line RAW264.7 stimulated by lipopolysaccharide. The in vivo inhibitory activities were further tested against xylene-induced mouse ear swelling. Molecular docking predicted that whether new compounds could effectively bind to target protein cyclooxygenase-2. The inhibitory activity of new compounds to cyclooxygenase-2 enzyme were verified by the in vitro experiment. Results: A total of 21 novel derivatives were synthesized, and exhibit lower polarities than the gentiopicroside. Most compounds have good in vitro anti-inflammatory activity. The in vivo activity results demonstrated that 8 compounds were more active than gentiopicroside. The inhibition rate of some compounds was higher than celecoxib. Molecular docking predicted that 6 compounds could bind to cyclooxygenase-2 and had high docking scores in accordance with their potency of the anti-inflammatory activity. The confirmatory experiment proved that these 6 compounds had significant inhibitory effect against cyclooxygenase-2 enzyme. Structure-activity relationship analysis presumed that the para-substitution with the electron-withdrawing groups may benefit the anti-inflammatory activity. Conclusion: These gentiopicroside derivatives especially PL-2, PL-7 and PL-8 may represent a novel class of cyclooxygenase-2 inhibitors and could thus be developed as new anti-inflammatory agents.

Preparation and Evaluation of 6-Gingerol Derivatives as Novel Antioxidants and Antiplatelet Agents.

Ginger (Zingiber officinale) is widely used as a spice and a traditional medicine. Many bioactivities have been reported for its extracts and the isolated compounds, including cardiovascular protective effects. Different pathways were suggested to contribute to these effects, like the inhibition of platelet aggregation. In this study, we synthesised fourteen 6-gingerol derivatives, including eight new compounds, and studied their antiplatelet, COX-1 inhibitor, and antioxidant activities. In silico docking of selected compounds to h-COX-1 enzyme revealed favourable interactions. The investigated 6-gingerol derivatives were also characterised by in silico and experimental physicochemical and blood-brain barrier-related parameters for lead and preclinical candidate selection. 6-Shogaol (2) was identified as the best overall antiplatelet lead, along with compounds 3 and 11 and the new compound 17, which require formulation to optimize their water solubility. Compound 5 was identified as the most potent antioxidant that is also promising for use in the central nervous system (CNS).

Vitamin D3 supplementation could ameliorate the inflammatory and redox status in the muscular phase of trichinellosis.

Nutritional supplements, particularly vitamin D, have been widely used worldwide in the treatment of various infections, including parasites. This study aimed to evaluate the potential effects of vitamin D3 supplementation on the muscular phase of trichinellosis in experimental animals. Mice were divided as follows: (group I): infected untreated, (group IIa) infected and treated with vitamin D3 for 12 doses beginning 2 weeks before infection and continuing after infection, (group IIb) infected and treated with vitamin D3 for 8 doses beginning on the same day of infection, (group III) normal control, (group IVa) which received vitamin D3 for 12 doses and (group IVb) which received vitamin D3 for 8 doses. Mice were sacrificed 35 days after infection and total muscle larval count, and histopathological examination of muscle samples with immunohistochemical staining of cyclooxygenase 2 (COX2) and inducible nitric oxide synthase (iNOS) were performed. Muscle relative cathelicidin mRNA expression was assessed, as well as serum levels of muscle enzymes CK and LDH, interleukin-4 (IL-4), IL-10, IL-17 and interferon-gamma (INF-γ). Vitamin D3 supplementation significantly reduced muscle larval count, inflammatory cellular infiltration, COX2 and iNOS expression. Furthermore, it increased cathelicidin gene expression, decreased serum levels of CK and LDH and affected serum cytokine levels, increasing serum IL-4 and IL10 levels while decreasing serum INF γ and IL-17. In conclusion, vitamin D3 supplementation has favorable outcomes on the muscle phase of trichinellosis, including anti-inflammatory, antioxidant, and immunomodulatory effects.

Schizochytrium spp. Dietary Supplementation Modulates Immune-Oxidative Transcriptional Signatures in Monocytes and Neutrophils of Dairy Goats.

The high propensity of dietary polyunsaturated fatty acids (PUFA) to oxidation can induce a cascade of cellular immune-oxidative imbalances. On the other hand, PUFA, namely docosapentaenoic acid (ω6-DPA) and docosahexaenoic acid (DHA) can exert immunomodulatory effects by suppressing a pro-inflammatory response. Thus, the objective of this study was to investigate the effect of dietary supplementation with Schizochytrium spp. levels, rich in both ω6-DPA and DHA on the transcriptional profiling of genes involved in oxidative homeostasis and innate immunity of dairy goats' monocytes and neutrophils. Twenty-four dairy goats were divided into four homogeneous sub-groups; the diet of the control group (CON) had no Schizochytrium spp. while those of the treated groups were supplemented daily with 20 (ALG20), 40 (ALG40), and 60 (ALG60) g/goat/day. The mRNA levels of MGST1 in neutrophils were downregulated (p = 0.037), while in monocytes, SOD2 and SOD3 were downregulated (p = 0.010 and p = 0.044, respectively) in ALG60 compared to the CON group. GPX2 mRNA levels were downregulated (p = 0.036) in ALG20 and ALG60 compared to the CON group in neutrophils. NOX1 was upregulated (p = 0.043) in the neutrophiles of ALG60-goats. NOX2 was upregulated (p = 0.042) in the monocytes of ALG40-fed goats, while higher (p = 0.045) levels were also found in the ALG60 group in neutrophils. The mRNA levels of COX2 were downregulated (p = 0.035) in monocytes of the ALG40 and ALG60 groups. The mRNA levels of PTGER2 were also downregulated (p = 0.004) in monocytes of Schizochytrium-fed goats, while in neutrophils, significant downregulation (p = 0.024) was only found for ALG60 compared to the CON group. ALOX5AP mRNA levels were significantly decreased (p = 0.033) in ALG60 compared to the CON group in monocytes. LTA4H mRNA levels were increased (p = 0.015) in ALG60 compared to ALG20 and ALG40 groups in monocytes, while in neutrophils, a significant downregulation (p = 0.028) was observed in ALG20 compared to the CON group. The inclusion of more than 20 g Schizochytrium spp./day in goats' diet induced imbalances in mechanisms that regulate the antioxidant system, while downregulated the expression of pro-inflammatory pathways in monocytes and neutrophils.

Rheum rhaponticum and Rheum rhabarbarum Extracts as Modulators of Endothelial Cell Inflammatory Response.

BACKGROUND: Inflammation, endothelial dysfunction, and alterations in blood physiology are key factors contributing to atherosclerosis and other cardiovascular disorders. Hence, modulation of endothelial function and reducing its pro-inflammatory and pro-thrombotic activity is considered one of the most important cardioprotective strategies. This study aimed to evaluate the anti-inflammatory potential of rhubarb extracts isolated from petioles and underground organs of Rheum rhabarbarum L. (garden rhubarb) and R. rhaponticum L. (rhapontic rhubarb) as well as two stilbenoids, typically found in these plants, i.e., rhapontigenin (RHPG) and its glycoside, rhaponticin (RHPT). METHODS: Analysis of the anti-inflammatory effects of the indicated rhubarb-derived substances involved different aspects of the endothelial cells' (HUVECs) response: release of the inflammatory mediators; cyclooxygenase (COX-2) and 5-lipoxygenase (5-LOX) expression as well as the recruitment of leukocytes to the activated HUVECs. The ability of the rhubarb-derived extracts to inhibit COX-2 and 5-LOX activities was examined as well. The study was supplemented with the in silico analysis of major components of the analyzed extracts' interactions with COX-2 and 5-LOX. RESULTS: The obtained results indicated that the examined plant extracts and stilbenes possess anti-inflammatory properties and influence the inflammatory response of endothelial cells. Biochemical and in silico tests revealed significant inhibition of COX-2, with special importance of rhaponticin, as a compound abundant in both plant species. In addition to the reduction in COX-2 gene expression and enzyme activity, a decrease in the cytokine level and leukocyte influx was observed. Biochemical tests and computational analyses indicate that some components of rhubarb extracts may act as COX-2 inhibitors, with marginal inhibitory effect on 5-LOX.