Cyclooxygenase-2 Gene Expression Research Articles

Investigation of the Relationship between COX-2 Expression, Clinicopathological Symptoms, and Prognosis of the Patients with Gastric Cancer

Introduction: The present study was designed to investigate the relationship between COX-2 gene expression, clinicopathological symptoms, and prognosis of patients with gastric cancer.
 Methods: The present cross-sectional study was designed to achieve the scientific aims among patients referred to the Imam Reza Hospital, Mashhad, Iran from 2008 to 2018. In this study, patients' demographic characteristics, clinical signs and symptoms, tumor findings, type of surgery, and COX-2 gene expression level (using Real Time PCR technique) were recorded. The data were analyzed by SPSS software (version 16), statistical tests, including Chi-square, Fisher's exact, Gamma coefficient, one-way ANOVA, or Kruskal-Wallis test at a significance level of 0.05.
 Results: Out of 110 patients included in the study, 55 patients were evaluated according to the criteria. Among them 21 (38.2%) patients had decreased levels of gene expression, in 18 (32.7%) patients gene expression was in the normal range, and in 16 (29.1%) patients increased gene expression was seen. The average survival months of patients after surgery were 23.41 ± 23.07 months. No statistically significant relationship was found between increased COX-2 gene expression and clinical signs, age, sex, survival (by month), tumor progression, type of surgery, histological features, family history of cancer, and other cases (P> 0.05); However, considering the annual survival in the annual classification showed a significant relationship with the expression of COX-2 gene (P=0.028).
 Conclusion: Despite no significant relationship between COX-2 gene expression and studied parameters in the patients with gastric cancer in this study, the existence of a significant relationship between gene expression and annual survival of patients seems so important.

Ceratonia siliqua pods (Carob) methanol extract alleviates doxorubicin-induced nephrotoxicity via antioxidant, anti-inflammatory and anti-apoptotic pathways in rats

Doxorubicin (DOX) is an anti-neoplastic therapy, but its use is limited by its deleterious toxic effects including nephrotoxicity and cardiotoxicity. This work aimed at assessing the potential protective effect of Ceratonia siliqua methanol extract (CME) on DOX-induced nephrotoxicity in 5 groups of Wistar rats. Nephrotoxicity was induced experimentally by intraperitoneal (IP) injection of DOX (15 mg/kg). DOX increased serum creatinine, urea, sodium, and potassium levels. It elevated MDA levels in the renal tissue but decreased the concentration of GSH and the activity of GST, CAT, and SOD. Meanwhile, it decreased the level of immunomodulatory anti-inflammatory mediators: IL-10 and TGF-β, as well as the activity of MPO but increased the level of IL-6, TNF-α, and caspase-3 in the renal tissue. DOX has upregulated COX-2, caspase-9, and Bax gene expression and downregulated the Bcl-2 gene expression. Immunolabeling of renal tubular epithelium in DOX-intoxicated rats was moderate to strong against Bax, COX-2, and NF-kβ and weak against Bcl-2. Treatment with CME significantly restored the levels of kidney function parameters and the levels of oxidative stress markers. It stimulated the production of IL-10 and TGF-β and decreased the level of IL-6 and TNF-α. CME reverted the gene expression of COX-2, caspase-9, and Bax. Microscopically, CME alleviated the DOX-induced renal damage. Phytochemical analysis revealed the presence of 26 compounds in the CME. No signs of acute toxicity were recorded by CME up to 4000 mg/kg b. wt. orally into mice. Finally, CME could effectively alleviate the deleterious effects of DOX on the kidney. The safety of carob extract encourages its use in the preparation of valuable therapeutic agents.

Alpha-pinene modulates inflammatory response and protects against brain ischemia via inducible nitric oxide synthase-nuclear factor-kappa B-cyclooxygenase-2 pathway.

Cerebral ischemia-reperfusion leads to brain tissue injury. Inflammation and apoptosis play pivotal roles in the pathology. α-Pinene is an organic compound of many aromatic plants and is known as a potent agent to possess antioxidant, and anti-inflammatory properties. Here, we sought to identify the anti-inflammatory and anti-apoptosis mechanism by which α-Pinene improves brain ischemia injury. Male Wistar rats underwent MCAO surgery for 1h and different doses of alpha-pinene (25, 50, and 100mg/kg) were intraperitoneally injected immediately after reperfusion to test this hypothesis. IV, NDS, gene and protein expression of inducible nitric oxide synthase (iNOS), cyclogenase-2 (COX-2), nuclear factor kappa B (NF-κB) p65, and caspase-3 were assessed 24h after reperfusion. Results demonstrated that NF-κB p65, iNOS, and COX-2 gene and protein expression increased in the hippocampus, cortex, and striatum after 24h of reperfusion, and alpha-pinene significantly inhibited NF-kB p65, iNOS, and COX-2 expression. Also, alpha-pinene significantly reduced the ischemia/reperfusion-induced caspase-3 activation in CA1 area of hippocampus. Results showed that alpha-pinene protects the cerebral against ischemic damage caused by MCAO, and this effect may be through the regulating iNOS -NF-kappa B- COX-2 and caspase-3 inflammatory and apoptotic pathways.

Effect of antioxidant and anti-inflammatory on bioactive components of carrot (Daucus carota L.) leaves from Jeju Island

The present study evaluated the potential of carrot (Daucus carota L.) leaf, a non-edible part of carrots, which are among the most consumed vegetables worldwide. The antioxidant activities of 70% ethanol extract (EEC) and hot water extract (HEC) of carrot leaves were compared. The results revealed that the total polyphenol content, total flavonoid content, and DPPH and ABTS radical scavenging activities were higher in EEC than in HEC. Both extracts protected the cells against H2O2-induced toxicity and markedly reduced the levels of reactive oxygen species in RAW 264.7 cells. Moreover, pretreatment of RAW 264.7 cells with EEC and HEC prior to H2O2 (500 μM) exposure increased superoxide dismutase and glutathione peroxidase activities in these cells. Notably, EEC and HEC increased intracellular catalase activity by 36.77 and 6.39 times, respectively. Compared to HEC, EEC remarkably inhibited the production of lipopolysaccharide-induced nitric oxide and reduced the gene expression of IL-6, IL-1β, iNOS, COX-2, and TNF-α. Comparative analysis of the composition of the extracts using HPLC–UV suggested notably higher contents of catechin, chlorogenic acid, caffeic acid, rutin, quercetin, and cynaroside in EEC than in HEC. Collectively, these results imply that carrot leaves are a potentially beneficial natural source of antioxidants and anti-inflammatory compounds in functional foods.

Study on mechanism of Zishen Pill treating benign prostatic hyperplasia based on serum pharmacochemistry and network pharmacology

Zishen Pill (ZSP) is a traditional Chinese medicine that is frequently used to treat Benign Prostatic Hyperplasia (BPH), however its specific mechanism of action and active ingredients are yet unknown. We used a combination of serum pharmacochemistry and network pharmacology and a series of biochemical assays to explore the action mechanism of ZSP in the treatment of BPH. The BPH rat model was created using testosterone propionate, and following oral ZSP administration, the components of ZSP in rat serum were detected by UPLC-Q-Exactive Orbitrap/MS method. A "component-target-disease" network and PPI networks were constructed on this foundation. The primary mechanism of ZSP decreasing BPH in rats was studied by KEGG pathway and GO analysis. Finally, the potential pathways and key targets were further verified in vivo by molecular biology and immunological methods. 46 substances were charactered from rat serum, and 164 anti-BPH targets were screened from the database. According to network pharmacology, the primary targets were CASP3, STAT3, JUN, and PTGS2/COX2. Three related pathways (PI3K/Akt signaling pathway, AGE-RAGE signaling pathway and EGFR tyrosine kinase inhibitor resistance) were closely related to the therapeutic effects of ZSP. The findings of molecular biology demonstrated that ZSP may bring Bcl-2, BAX, CASP3, COX2, and 5LOX protein and gene expression in BPH rats appreciably closer to that of normal rats. Additionally, ZSP can lessen the expression of inflammatory cytokines in BPH rats, including VEGF, TNF-α, CCL5, and interleukin. ConclusionThe above results suggest that ZSP may reduce BPH through inflammation/immunity and apoptosis/proliferation-related pathways. This study offers a fresh approach to investigate the basic pharmacological effects and mechanism of ZSP in the treatment of BPH.

Protective Effects of Aquilaria agallocha and Aquilaria malaccensis Edible Plant Extracts against Lung Cancer, Inflammation, and Oxidative Stress—In Silico and In Vitro Study

The family Thymelaeaceae, which includes huge evergreen trees that are sparsely distributed in tropical rainforests, includes the genus Aquilaria. Numerous medical conditions, including inflammation, cancer, and oxidative stress have been traditionally treated using Aquilaria agallocha and Aquilaria malaccensis. In this study, we evaluated in silico and biological activity with A. agallocha and A. malaccensis sample for more conformation. Raw 264.7 macrophage cells and HacaT cells were used, together with the MTT, ROS, NO, and wound healing assays, to investigate the possible cytotoxicity in A549 lung cancer. Thus, A. agallocha and A. malaccensis showed significant cytotoxicity against A549 cancer cells at 1000 µg/mL. Furthermore, we observed an elevated ROS level in cancer cells. The wound healing assay showed cancer cell inhibition activity. While BCL-2 decreased in the intrinsic route, p53, Bax, Caspase 3, and Caspase 9 were elevated by A.A and A.M. Additionally, we have also conducted an in silico evaluation followed by molecular dynamics (MD) simulations, along with ADMET and biological activity prediction to further validate the experimental results. In normal cells, both samples showed less toxicity at 1000 µg/mL and suppressed the LPS-treated NO and ROS levels against the inflammation. Additionally, A.A and A.M suppressed the pro-inflammatory gene expression of COX-2, iNOS, TNF-α, IL-6, and IL-8 in RAW 264.7 cells. On the other hand, A.A and A.M extract effectively suppressed oxidative stress by increasing the antioxidative gene expression in H2O2-induced HaCat cells at 50 μg/mL. This study revealed that the plant extracts from A. agallocha and A. malaccensis could exert a cytotoxic effect on lung adenocarcinoma cells through the activation of an intrinsic signaling pathway. Moreover, it could be a potential source of anti-inflammatory, antioxidant, and anti-cancer agents after consideration of in vivo and clinical studies.

Tetrandrine Represses Inflammation and Attenuates Osteoarthritis by Selective Inhibition of COX-2.

There is a lack of effective and long-term safe drugs for the treatment of osteoarthritis (OA). Tetrandrine (Tet) has been approved and used to treat rheumatoid arthritis for several decades, but its effect on OA has not been investigated. Herein, we explored the effect of Tet on OA and its underlying mechanism. OA was induced using destabilization of the medial meniscus (DMM) in C57BL/6J mice. The animals were randomly divided into sham, DMM, Tet, celecoxib (CXB), and indomethacin (INDO) groups. Each group was given solvent or corresponding drugs by gavage for 7 weeks after convalescence. Pathological staining, OARSI scores, micro-computed tomography and behavior tests were performed to evaluate the effects of Tet. Tet remarkably alleviated cartilage injury in the knee joint, limited bone remodeling in the subchondral bone, and delayed progression of OA. Tet also significantly relieved joint pain and maintained function. Further mechanistic studies revealed that Tet lowered inflammatory cytokine levels and selectively suppressed gene and protein expression of cyclooxygenase (COX)-2 but not COX-1 (P<0.01). Tet also reduced the production of prostaglandin E2 without damaging the gastric mucosa. We found that Tet could selectively inhibit COX-2 gene expression and decrease cytokine levels in mice, thus reducing inflammation and improving OA without obvious gastric adverse events. These results provide a scientific basis for the clinical application of Tet in the treatment of OA.

Galectins are critical regulators of cytokine signalling at feto-maternal interface in infection-associated spontaneous preterm birth

IntroductionSpontaneous preterm birth (sPTB) is a global health issue. Studies suggest infections are chiefly associated with sPTB and galectins (gals) play a role in regulation of innate and adaptive maternal immune response against pathogens during sPTB. The aim of this study was to describe the gene expression of gal -1, -3, -8, -9, -13 in relation to gene expression of cyclooxygenase-2 (COX-2) and the cytokines IL-8, IL-10, TNF-α, IFN-ϒ in the setting of sPTB and confirmed infection with Chlamydia trachomatis, Mycoplasma hominis, and Ureaplasma urealyticum. MethodsPlacental samples were collected from 120 term control and 120 sPTB pregnancies. PCR was used to detect specific pathogens. Gene expression of galectins, cytokines, and COX-2 was performed using real time qPCR. ResultsFold-change expression of gal -1, -3, -8, -9, -13 was 5.13, 6.11, 1.14, 5.23 and 7.16 (p<0.001), respectively; while IL-10, IL-8, TNF-α, IFN-ϒ and COX-2 was 6.29, 6.55, 6.35, 6.36 and 2.73-fold upregulated (p<0.05), respectively in infected sPTB. Gal-1 was positively correlated with IL-10 (r=0.49, p=0.003) while gal-3 showed significant correlation with IL-8 (r=0.42, p=0.0113), TNF-α (r=0.65, p=< 0.001) and COX-2 (r=0.72, p=0.001). However, gal-8 was not significantly correlated with any cytokine. Gal-9, -13 were negatively correlated with IFN-ϒ (r=-0.45, p=0.006) and IL-8 (r=-0.39, p=0.018). DiscussionGal-1, -9, -13 are anti-inflammatory and might play role in immune-tolerance while gal-3 is pro-inflammatory and possibly responsible for immunogenic response, having potential to anticipate the clinical beginning of preterm labour during infection.

Anti-Inflammatory and Vasorelaxant Effects Induced by an Aqueous Aged Black Garlic Extract Supplemented with Vitamins D, C, and B12 on Cardiovascular System.

Multiple studies demonstrated biological activities of aged black garlic, including anti-inflammatory, antioxidant, and cardioprotective effects. We aimed to investigate the protective effects of an aged black garlic water extract (ABGE) alone or in association with multivitamins consisting of combined Vitamins D, C, and B12, on mouse heart specimens exposed to E. coli lipopolysaccharide (LPS). Moreover, we studied the hydrogen sulphide (H2S) releasing properties and the membrane hyperpolarization effect of the Formulation composed by ABGE and multivitamins, using Human Aortic Smooth Muscle Cells (HASMCs). ABGE, vitamins D and C, and the Formulation suppressed LPS-induced gene expression of cyclooxygenase (COX)-2, tumor necrosis factor (TNF)-α, interleukin (IL)-6, nuclear factor-kB (NF-kB), and inducible nitric oxide synthase (iNOS) on mouse heart specimens. The beneficial effects induced by the extract could be related to the pattern of polyphenolic composition, with particular regard to gallic acid and catechin. The Formulation also increased fluorescence values compared to the vehicle, and it caused a significant membrane hyperpolarization of HASMCs compared to ABGE. To conclude, our present findings showed that ABGE, alone and in association with multivitamins, exhibited protective effects on mouse heart. Moreover, the Formulation increased intracellular H2S formation, further suggesting its potential use on cardiovascular disease.

ARGININE SUPPLEMENTATION HAS A DOSE-DEPENDENT EFFECT ON CHONDROGENIC DIFFERENTIATION

Nitric oxide is a free radical which in vivo is solely produced during the conversion of the amino acid arginine into citrulline by nitric oxide synthase enzymes. Recently, the importance of nitric oxide on inflammation and bone metabolism has been investigated. However, the knowledge regarding possible in vitro effects of arginine supplementation on chondrogenic differentiation is limited.ATDC5, a cell line which is derived from mouse teratocarcinoma cells and which is characterized as chondrogenic cell line, were proliferated in Dulbecco's Modified Eagle Medium (DMEM)/F12 and subsequently differentiated in proliferation medium supplemented with insulin, transferrin and sodium-selenite and where arginine was added in four different concentrations (0, 7.5, 15 and 30 mM). Samples were harvested after 7 or 10 days and were stored at −80 °C for subsequent RNA isolation for qPCR analysis. To determine chondrogenic differentiation, Alcian Blue staining was performed to stain the proteoglycan aggrecan, which is secreted by differentiated ATDC5 cells. All measurements were performed in triplo.Alcian Blue staining showed a qualitative increase of proteoglycan aggrecan secretion in differentiated ATDC5 cells after treatment with 7 and 15 mM arginine, with additional increased expression of ColII, ColX, Bmp4 and Bmp6. Treatment with 30 mM arginine inhibited chondrogenic differentiation and expression of aforementioned genes, however, Cox-2 and Vegfa gene expression were increased in these samples. Bmp7 was not significantly expressed in any experimental condition.The obtained results are suggestive for a dose-dependent effect of arginine supplementation on chondrogenic differentiation and associated gene expression, with 7.5 and 15 mM as most optimal concentrations and implications for apoptosis after incubation with 30 mM arginine. A future recommendation would be to investigate the effects of citrulline in a similar experiment, as this shows even more promising results to enhance the nitric oxide metabolism in sepsis and bone healing.

Low vitamin D status is associated with inflammatory response in older patients with cerebral small vessel disease.

This study aimed to determine the association of the NF-κB inflammatory signaling pathway with vitamin D status in older cerebral small vessel disease (SVD) patients. We measured serum 25(OH)D, pro-and anti-inflammatory cytokines, and mRNA levels of the vitamin D-activating enzyme, CYP27B1, as well as NF-kB, COX-2, the chemokine-CCL2, IL-1β, IL-6, TNF-α, TGF-β, and IL-10, in cerebral SVD patients aged ≥60years presenting with vascular dementia and age and gender-matched healthy controls. Low vitamin D status (insufficiency: serum 25(OH)D 12-20ng/ml; deficiency: ≤12ng/ml) was more prevalent among patients compared to controls. The mRNA levels of NF-kB, COX-2, CCL2, IL-1β, and IL-6, and serum levels of pro-inflammatory cytokines (IL-1α, IL-1β, IL-6, and TNF-α) were significantly higher in cases compared to controls. There was a significant correlation between CYP27B1 and NF-kB, COX-2, CCL2, and IL-1β gene expression. Serum IL-1α, IL-1β, and IL-6 concentrations and the expression of CCL-2, NF-kB2, and NF-kB3 genes were higher in vitamin D-deficient subjects compared to vitamin D-sufficient subjects. There was a significant negative correlation between serum 25(OH)D and IL-1α, IL-6, and TNF-α, and a positive correlation between 25(OH)D and IL-10. Low vitamin D is associated with an inflammatory response via NF-kB signaling, which could play a role in the etio-pathogenesis of SVD. Further large-scale studies are required to validate our findings.

Terminal Complement Activation Is Induced by Factors Released from Endplate Tissue of Disc Degeneration Patients and Stimulates Expression of Catabolic Enzymes in Annulus Fibrosus Cells.

Terminal complement complex (TCC) deposition was identified in human degenerated discs. To clarify the role of terminal complement activation in disc degeneration (DD), we investigated respective activating mechanisms and cellular effects in annulus fibrosus (AF) cells. Isolated cells from human AF, nucleus pulposus (NP), and endplate (EP) were stimulated with human serum alone or with zymosan and treated with either the C3 inhibitor Cp40 or the C5 antibody eculizumab. Complement activation was determined via anaphylatoxin generation and TCC deposition detection. Thereby, induced catabolic effects were evaluated in cultured AF cells. Moreover, C5 cleavage under degenerative conditions in the presence of AF cells was assessed. Zymosan-induced anaphylatoxin generation and TCC deposition was significantly suppressed by both complement inhibitors. Zymosan induced gene expression of ADAMTS4, MMP1, and COX2. Whereas the C3 blockade attenuated the expression of ADAMTS4, the C5 blockade reduced the expression of ADAMTS4, MMP1, and COX2. Direct C5 cleavage was significantly enhanced by EP conditioned medium from DD patients and CTSD. These results indicate that terminal complement activation might be functionally involved in the progression of DD. Moreover, we found evidence that soluble factors secreted by degenerated EP tissue can mediate direct C5 cleavage, thereby contributing to complement activation in degenerated discs.

A231 THE FLAVANOID CYANIDIN REDUCES INTESTINAL INFLAMMATORY DAMAGE BY MAINTAINING INTESTINAL PERMEABILITY

Abstract Background Since the discovery and the use of proton pump inhibitors (PPI), the incidence of gastric ulcers has reduced, however, there has been an increase in intestinal lesions since PPIs do not guarantee protection to the intestine. To protect the deeper layers of the intestinal wall, the intestinal barrier tightly regulates the passage of pro-inflammatory molecules, microorganisms, toxins, and antigens. The epithelium is damaged during intestinal inflammation due to the action of inflammatory cytokines such as TNF resulting in ulceration. Cyanidin is a flavonoid of the anthocyanin class, found in several red fruits, with anti-inflammatory and antioxidant activity. Purpose We hypothesized that the administration of cyanidin can decrease the deleterious effects caused by polypharmacy (PPI + NSAID) in the small intestine by reducing intestinal permeability. Method In vivo: Male mice (8 weeks old) were treated to induce intestinal ulcers caused by polypharmacy (combination of PPI - lansoprazole 20 mg/kg daily, non-steroidal anti-inflammatory drug (NSAID) - acetylsalicylic acid 10 mg/kg daily and COX-2 selective NSAID, celecoxib 20 mg/kg daily). On the ninth day, oral treatment with cyanidin (5 mg/kg) or vehicle was started until the fourteenth day, then the animals were killed for parameter analysis lesion. ELISA was performed to quantify interleukins (IL-10, IL-6, IL-1β) and cytokine (TNF), and we assessed the antioxidant profile (SOD, CAT, and GSH) and measured gene expression of TNF, IL-10, IL-6 TRL-4, HMOX-1, MMP 2 and 9, COX-1, MUC-3, ZO-1, CL-1. In vitro: Monolayers of colonic epithelial cell lines (Caco-2 at 21 days of confluence) were mounted in Ussing chambers to assess barrier function and to determine transepithelial resistance (TER). To analyze the permeability response to injury, we utilized TNF and IFN (25 ng/mL) with cyanidin (10 or 100 uM) for 48 h in transwell plates, with measurement of total intestinal permeability using 4 kD FITC-dextran. Result(s) Analysis of mouse intestine indicated that cyanidin (5 mg/kg) significantly reduced expression of IL-6 and TNF, TLR4, and HMOX-1 (p&amp;lt;0.05), and increased gene expression of MUC-3, CL-1, occludin, COX-1, and IL-10 (p&amp;lt;0.05) without altering antioxidant parameters. Cyanidin (100 mM) maintained barrier function as shown by transepithelial electrical resistance (TER), and also significantly reversed the detrimental effects of the inflammatory cytokine on FITC-dextran flux in Caco-2 cells (p&amp;lt;0.05). This may be related to the increased expression of occludin and ZO-1 in the intestinal epithelium of mice. Conclusion(s) These in vivo and in vitro results suggest that cyanidin decreases the polypharmacy-induced intestinal inflammatory response while maintaining the integrity of the intestinal epithelium. Other trials are underway to elucidate these mechanisms. Please acknowledge all funding agencies by checking the applicable boxes below Other Please indicate your source of funding; NSERC; FAPESP Disclosure of Interest None Declared

Adding New Scientific Evidences on the Pharmaceutical Properties of Pelargonium quercetorum Agnew Extracts by Using In Vitro and In Silico Approaches

Pelargonium quercetorum is a medicinal plant traditionally used for treating intestinal worms. In the present study, the chemical composition and bio-pharmacological properties of P. quercetorum extracts were investigated. Enzyme inhibition and scavenging/reducing properties of water, methanol, and ethyl acetate extracts were assayed. The extracts were also studied in an ex vivo experimental model of colon inflammation, and in this context the gene expression of cyclooxygenase-2 (COX-2) and tumor necrosis factor α (TNFα) were assayed. Additionally, in colon cancer HCT116 cells, the gene expression of transient receptor potential cation channel subfamily M (melastatin) member 8 (TRPM8), possibly involved in colon carcinogenesis, was conducted as well. The extracts showed a different qualitative and quantitative content of phytochemicals, with water and methanol extracts being richer in total phenols and flavonoids, among which are flavonol glycosides and hydroxycinnamic acids. This could explain, at least in part, the higher antioxidant effects shown by methanol and water extracts, compared with ethyl acetate extract. By contrast, the ethyl acetate was more effective as cytotoxic agent against colon cancer cells, and this could be related, albeit partially, to the content of thymol and to its putative ability to downregulate TRPM8 gene expression. Additionally, the ethyl acetate extract was effective in inhibiting the gene expression of COX-2 and TNFα in isolated colon tissue exposed to LPS. Overall, the present results support future studies for investigating protective effects against gut inflammatory diseases.

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

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). 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. 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.

COX-2/PGE2 upregulation contributes to the chromosome 17p-deleted lymphoma

Deletions of chromosome 17p, where TP53 gene locates, are the most frequent chromosome alterations in human cancers and associated with poor outcomes in patients. Our previous work suggested that there were p53–independent mechanisms involved in chromosome 17p deletions-driven cancers. Here, we report that altered arachidonate metabolism, due to the deficiency of mouse Alox8 on chromosome 11B3 (homologous to human ALOX15B on chromosome 17p), contributes to the B cell malignancy. While the metabolites produced from lipoxygenase pathway reduced, chromosome 11B3 deletions or Alox8 loss, lead to upregulating its paralleling cyclooxygenase pathway, indicated by the increased levels of oncometabolite prostaglandin E2. Ectopic PGE2 prevented the apoptosis and differentiation of pre-B cells. Further studies revealed that Alox8 deficiency dramatically and specifically induced Cox-2(Ptgs2) gene expression. Repressing Cox-2 by its shRNAs impaired the tumorigenesis driven by Alox8 loss. And, in turn, tumor cells with Alox8 or 11B3 loss were sensitive to the COX-2 inhibitor celecoxib. This correlation between COX-2 upregulation and chromosome 17p deletions was consistent in human B-cell lymphomas. Hence, our studies reveal that the arachidonate metabolism abnormality with unbalanced ALOX and COX pathways underlies human cancers with 17p deletions and suggest new susceptibility for this disease.

Depigmenting topical therapy based on a synergistic combination of compounds targeting the key pathways involved in melasma pathophysiology.

Melasma has a complex pathophysiology with different cell types and signalling pathways involved. Paracrine factors secreted by keratinocytes, fibroblasts and endothelial cells act on melanocytes and stimulate melanogenesis. These paracrine factors are involved in the oxidative stress, inflammatory, vascular and hormonal pathways, among others. Damage of the dermoepidermal barrier also occurs and facilitates melanin deposition in the dermis, also known as dermal or mixed melasma. We used artificial intelligence tools to define the best combination of compounds for skin pigmentation inhibition. Mathematical models suggested the combination of retinol, diosmin and ferulic acid to be the most effective one. In vitro cellular tyrosinase activity assay proved that this combination had a synergistic depigmenting effect. Further assays proved that the combination could inhibit key pathways involved in melasma by downregulating ET-1 and COX-2 gene expression and IBMX-induced dendricity in human melanocytes, and upregulated the gene expression of IL-1b, TIMP3 and several endogenous antioxidant enzymes. The combination also reduced melanin levels in a phototype VI 3D epidermis model. These results indicate that the combination of retinol, diosmin and ferulic acid is an effective synergistic complex for the treatment of melasma by regulating the key molecular pathways involved in skin hyperpigmentation pathophysiology.

Olive leaves extract alleviates inflammation and modifies the intrinsic apoptotic signal in the leukemic bone marrow.

Current anti-leukemic chemotherapies with multiple targets suffer from side effects. Synthetic drugs with huge off-target effects are detrimental to leukemic patients. Therefore, natural plant-based products are being increasingly tested for new anti-leukemic therapy with fewer or no side effects. Herein, we report the effect of ethanolic olive leaves extract (EOLE) on the K562 cell line and on the bone marrow (BM) of N-ethyl-N-nitrosourea (ENU)-induced leukemic mice. Using standard methodologies, we assessed viability, chromatin condensation, and induction of apoptosis in EOLE-treated K562 cells in-vitro. The anti-leukemic activity of EOLE was assayed by measuring ROS, levels of various cytokines, expression of iNOS and COX-2 gene, and changes in the level of important apoptosis regulatory and cell signaling proteins in-vivo. K562 cells underwent apoptotic induction after exposure to EOLE. In the BM of leukemic mice, EOLE therapy decreased the number of blast cells, ROS generation, and expression of NF-κB and ERK1/2. IL-6, IL-1β, TNF-α, iNOS, and COX-2 were among the inflammatory molecules that were down-regulated by EOLE therapy. Additionally, it decreased the expression of anti-apoptotic proteins BCL2A1, BCL-xL, and MCL-1 in the BM of leukemic mice. Chronic inflammation and anomalous apoptotic mechanism both critically contribute to the malignant transformation of cells. Inflammation in the tumor microenvironment promotes the growth, survival, and migration of cancer cells, accelerating the disease. The current investigation showed that EOLE treatment reduces inflammation and alters the expression of apoptosis regulatory protein in the BM of leukemic mice, which may halt the progression of the disease.

Probiotic Potential of the Marine Isolate Enterococcus faecium EA9 and In Vivo Evaluation of Its Antisepsis Action in Rats.

This study aims to obtain a novel probiotic strain adapted to marine habitats and to assess its antisepsis properties using a cecal ligation and puncture (CLP) model in rodents. The marine Enterococcus faecium EA9 was isolated from marine shrimp samples and evaluated for probiotic potential after phenotypical and molecular identification. In septic animals, hepatic and renal tissues were histologically and biochemically evaluated for inflammation and oxidative stress following the probiotic treatment. Moreover, gene expressions of multiple signaling cascades were determined using RT-PCR. EA9 was identified and genotyped as Enterococcus faecium with a 99.88% identity. EA9 did not exhibit any signs of hemolysis and survived at low pH and elevated concentrations of bile salts. Moreover, EA9 isolate had antibacterial activity against different pathogenic bacteria and could thrive in 6.5% NaCl. Septic animals treated with EA9 had improved liver and kidney functions, lower inflammatory and lipid peroxidation biomarkers, and enhanced antioxidant enzymes. The CLP-induced necrotic histological changes and altered gene expressions of IL-10, IL-1β, INF-γ, COX-2, SOD-1, SOD-2, HO-1, AKT, mTOR, iNOS, and STAT-3 were abolished by the EA9 probiotic in septic animals. The isolate Enterococcus faecium EA9 represents a promising marine probiotic. The in vivo antisepsis testing of EA9 highlighted its potential and effective therapeutic approach.

Evaluation of You-Gui-Wan critical compounds inhibiting ALOX-5 and HDC gene expression in RBL-2H3 cells using a fractional factorial design

Ethnopharmacological relevanceThe traditional Chinese medicine (TCM) You-Gui-Wan (YGW) has been used to treat asthma for hundreds of years. Aim of the studyYGW is composed of 10 types of medicinal materials. However, the immune mechanism of YGW in asthma treatment has not been elucidated. Therefore, this study investigated asthma symptoms attenuated by YGW and the underlying immune regulatory mechanism. Materials and methodsIntratracheal (i.t.) stimulation of BALB/c mice with Dermatophagoides pteronyssinus (Der p) was performed once per week (40 μL, 2.5 μg/μL). For six consecutive weeks, different doses of YGW (0.2 g/kg and 0.5 g/kg) were orally administered 30 min before stimulation with Der p. After the last stimulation, airway hyperreactivity, lung gene expression, and total immunoglobulin E (IgE) in blood were evaluated using a whole-body plethysmograph system, real-time PCR, and ELISA, respectively. In addition, DNP-IgE/DNP-BSA was added to stimulate mast cells (RBL-2H3), and YGW or various compound compositions (Trial) were added to RBL-2H3 cells for 30 min to evaluate the effects of the drug on mast cell degranulation and on gene expression. JMP 5.1 software was used to design and analyze YGW's critical compounds by which it inhibited ALOX-5 and HDC gene expression in RBL-2H3 cells. ResultsYGW significantly decreased serum total IgE levels and airway hyperresponsiveness in asthmatic mice. YGW also reduced the gene expression of IL-6, TNF-α, IL-4, IL-13, and COX-2 in the lungs of asthmatic mice and RBL-2H3 cells. YGW and the compound (Trial 21) present in YGW inhibited the gene expression of ALOX-5 and HDC in RBL-2H3 cells. ConclusionThe experimental results indicate that YGW exhibits anti-airway hyperresponsiveness and specific immunomodulatory effects. In addition, YGW synergistically inhibits ALOX-5 and HDC gene expression in mast cells through a combination of 21 compounds, including luteolin, quercetin, and β-carotene.