Overexpression Of Cyclooxygenase-2 Research Articles

(-)-Methyl-Oleocanthal, a New Oleocanthal Metabolite Reduces LPS-Induced Inflammatory and Oxidative Response: Molecular Signaling Pathways and Histones Epigenetic Modulation.

The antioxidant and anti-inflammatory responses of (−)-methyl-oleocanthal (met-OLE), a new metabolite of the extra virgin olive oil (EVOO) phenolic oleocanthal (OLE), were explored in lipopolysaccharide (LPS)-induced murine peritoneal macrophages. Possible signaling pathways and epigenetic modulation of histones were studied. Met-OLE inhibited LPS-induced intracellular reactive oxygen species (ROS) and nitrite (NO) production and decreased the overexpression of the pro-inflammatory enzymes COX-2, mPGES-1 and iNOS in murine macrophages. In addition, met-OLE was able to significantly decrease the activation of p38, JNK, and ERK mitogen-activated protein kinases (MAPKs) and blocked canonical and non-canonical inflammasome signaling pathways. On the contrary, met-OLE upregulated haem oxigenase 1 (HO-1) and nuclear factor (erythroid-derived 2)-like 2 (Nrf-2) expression in treated cells. Finally, met-OLE pretreated spleen cells counteracted LPS induction, preventing H3K18 acetylation or H3K9 and H3K27 demethylation. Overall, these results provide novel mechanistic insights into the beneficial effects of met-OLE regarding the regulation of the immune–inflammatory response through epigenetic changes in histone markers. This revealing evidence suggests that the methylated metabolite of OLE may contribute significantly to the beneficial effects that are associated with the secoiridoid-related compound and the usual consumption of EVOO.

Evaluation of Cyclooxygenase-2 and p53 Expression in Pterygium Tissue Following Preoperative Intralesional Ranibizumab Injection.

Introduction: Overexpression of vascular endothelial growth factor (VEGF), cyclooxygenase-2 (COX-2), and p53 are the postulated aetiopathogenesis in pterygium. VEGF is responsible for the induction of COX-2 expression, whereas p53 plays an important role in the regulation of VEGF. This study aimed to evaluate the immunohistochemistry of COX-2 and p53 expressions from excised pterygium tissue from patients who received intralesional ranibizumab (anti-VEGF) injection 2 weeks prior to pterygium surgery.Materials and Methods: An interventional comparative study involving patients presenting with primary pterygium was conducted between September 2015 and November 2017. The patients were randomized into either the intervention or control group. Patients in the intervention group were injected with intralesional ranibizumab (0.5 mg/0.05 ml) 2 weeks prior to surgery. Both groups underwent pterygium excision followed by conjunctival autograft. Immunohistochemistry staining was performed to evaluate COX-2 and p53 expressions in the excised pterygium tissue.Results: A total of 50 patients (25 in both the intervention and control groups) were recruited. There were 34 (68%) patients with grade III pterygium and 16 (32%) patients with grade IV pterygium. There was statistically significant difference in reduction of COX-2 expression in the epithelial layer [84.0% (95% CI: 63.9, 95.5)] (p = 0.007) and stromal layer [84.0% (95% CI: 63.9, 95.5)] (p < 0.001) between intervention and control groups. There was no significant difference in the reduction of p53 expression between the two groups.Conclusion: This study demonstrated the possible use of intralesional anti-VEGF treatment prior to pterygium excision as a potential future modality of adjunctive therapy for pterygium surgery.

Exploration of the quantitative Structure-Activity relationships for predicting Cyclooxygenase-2 inhibition bioactivity by Machine learning approaches

Cyclooxygenase-2 (COX-2) overexpression in many humans plays a key role in carcinogenic and inflammation-associated diseases. Several therapeutic and pharmaceutic drugs have been developed but some of them either lack the potency to control the COX-2 related diseases or promote unwanted side effects. The prospect and possibility for the development of drugs with novel therapeutic and pharmacological properties using the quantitative structure–activity relationship (QSAR) model were investigated. Themodel involves the application of chemical descriptors and supervised machine learning to predict the bioactivity classes of molecules for COX-2 inhibition using real multidimensional COX-2 inhibitors obtained from a curated database. The PubChem Fingerprints is the class of descriptor used in developing the model. A model performance check is carried out on 22 SciKit-Learn models and a comparative analysis of their predictive performance in classifying bioactivity of compounds is given in terms of validation accuracy. Unsupervised learning using principal component analysis (PCA) and t-distributed stochastic neighbor embedding (t-SNE) algorithms are applied as techniques to explore the effect of dimensionality in the training data. The PCA algorithm is a tool used to reduce the number of variables contained in the dataset, though preservingas much information as possible. The t-SNE algorithm is mainly used for data exploration and visualization of the multi-dimensional data set. Despite their high predictive performances, the eXtreme Gradient Boosting Classifier (XGB Classifier) algorithm is the ultimate performer. More so, hyperparameter tuning, and regularization account for excellent model statistics with higher predictive power at 10-fold cross-validation. Model metrics including log-loss probability (0.1208), accuracy score (0.9484), Matthew’s correlation coefficient (0.8741), among others proved adequately significant. Furthermore, the developed model is validated using recommended OECD metrics, such as Precision, Recall, and Balanced Accuracy for classification. The results in this study offer important pharmacological insight, and this insight can lead to designing novel bioactive drugs with undesired side effects. The proposed QSAR approach in this study achieves a futuristic performance when applied in drug development schemes different from conventional methods.

Cox-2 Antagonizes the Protective Effect of Sevoflurane on Hypoxia/Reoxygenation-Induced Cardiomyocyte Apoptosis through Inhibiting the Akt Pathway.

Objective To uncover the protective role of sevoflurane on hypoxia/reoxygenation-induced cardiomyocyte apoptosis through the protein kinase B (Akt) pathway. Methods An in vitro hypoxia/reoxygenation (H/R) model was established in cardiomyocyte cell line H9c2. Sevoflurane (SEV) was administrated in H9c2 cells during the reoxygenation period. Viability, layered double hydroxide (LDH) release, and apoptosis in H9c2 cells were determined to assess H/R-induced cell damage. Relative levels of apoptosis-associated genes were examined. Moreover, phosphorylation of Akt was determined. Results H/R injury declined viability and enhanced LDH release and apoptotic rate in H9c2 cells. Cyclooxygenase-2 (Cox-2) was upregulated following H/R injury, which was partially reversed by SEV treatment. In addition, SEV treatment reversed changes in viability and LDH release owing to H/R injury in H9c2 cells, which were further aggravated by overexpression of Cox-2. The Akt pathway was inhibited in H9c2 cells overexpressing Cox-2. Conclusions Sevoflurane protects cardiomyocyte damage following H/R via the Akt pathway, and its protective effect was abolished by overexpression of Cox-2.

Fisetin suppresses cigarette smoke extract-induced epithelial to mesenchymal transition of airway epithelial cells through regulating COX-2/MMPs/β-catenin pathway

Cigarette smoke exposure leads to upregulation of cyclooxygenase‐2 (COX‐2), an inducible enzyme that synthesizes prostaglandin E2 (PGE2) and promotes airway inflammation. COX-2 overexpression is frequently implicated in inflammation, invasion, metastasis, and epithelial-mesenchymal transition (EMT). However, its detailed molecular mechanism in cigarette smoke induced EMT is not clear. Further, fisetin, a bioflavonoid, exhibits antioxidant and anti-inflammatory properties, but its effect in modulating COX-2-mediated inflammation and downstream sequelae remains unexplored. Therefore, we have investigated the mechanism of cigarette smoke-induced COX-2-mediated EMT in airway epithelial cells and examined the role of fisetin in controlling this aberration. MTT, trypan blue staining, gelatin zymography, Western blotting, invasion, wound healing, and tumor sphere formation assays in cigarette smoke extract (CSE) and/or fisetin treated airway epithelial cells, and in-silico molecular docking studies were performed. Results revealed that CSE exposure increased the expression and activity of COX-2, MMP-2/9, and β-catenin and also enhanced expression of EMT markers leading to higher migration and invasion potential of airway epithelial cells. A specific COX-2 inhibitor NS-398 as well as fisetin treatment reversed the expression of EMT biomarkers, reduced the activity of MMP-2/9, and blocked the migration and invasion potential induced by CSE. Further, PGE2 also increased MMPs activity, invasion, and migration potential similar to CSE, which were significantly reversed by fisetin. In-silico studies showed a high binding affinity of fisetin to key EMT associated proteins, validating its anti-EMT potential. Thus, our study firstly unearths the mechanism of CSE-induced EMT in airway epithelial cells via COX-2/MMP/β-catenin pathway, and secondly, it reveals that fisetin could significantly reverse CSE-induced EMT by inhibiting COX-2, indicating that fisetin could be an effective drug candidate for cigarette smoke-induced lung dysfunction.

Is intestinal metaplasia the point of no return in the progression of gastric carcinogenesis?

Is intestinal metaplasia the point of no return in the progression of gastric carcinogenesis?

Effects of dietary zinc deficiency on esophageal squamous cell proliferation and the mechanisms involved.

BACKGROUNDDietary zinc deficiency has been shown to be associated with the development of esophageal cancer in humans, but the exact mechanism of action is not knownAIMTo observe the effects of dietary zinc deficiency on esophageal squamous cell proliferation. METHODSThirty C57BL/6 mice were randomly divided into three groups: A zinc-sufficient (ZS) group, zinc-deficient (ZD) group, and zinc-replenished (ZR) group. For weeks 1–10, zinc levels in the mice diets were 30.66–30.89 mg/kg in the ZS group and 0.66–0.89 mg/kg in the ZD and ZR groups. During weeks 10–12, the ZR group was switched to the ZS diet; the other two groups had no changes in their diets. Changes in body weight, serum, and esophageal tissue zinc concentrations were assessed as well as differences in the expression of proliferating cell nuclear antigen (PCNA), mitogen-activated protein kinase p38 (p38MAPK), nuclear factor kappa B (NF-κB) p105, NF-κB p65, and cyclooxygenase (COX)-2 proteins in the esophageal mucosa.RESULTSThe body weight and zinc concentration in the serum and esophageal mucosa were significantly lower in the ZD and ZR groups than in the ZS group (P < 0.05). In ZD mice, there was a marked proliferation of basal cells in the esophageal mucosa, resulting in a disturbance in the arrangement of basal cells in layers 2–4, a thickening of the squamous layer, and a significant increase in the expression of the above-mentioned five proteins involved in proliferation and inflammation in the esophageal mucosa. Two weeks after switching to the ZS diet, the serum zinc concentration in the ZR group increased, and the expression of PCNA, NF-κB p105, and COX-2 decreased, but the concentration of zinc in the esophageal mucosa and the structure of the esophageal mucosa did not display any significant changesCONCLUSIONThe ZD diet decreased the growth rate and promoted the proliferation of esophageal squamous cells in mice. The mechanism of proliferation was related to the induced overexpression of COX-2, P38, PCNA, and NF-κB (p105 and p65), and the ZR diet reduced the expression of PCNA, NF-κB p105, and COX-2, thereby reversing this process.​

Discovery of Phenolic Glycoside from Hyssopus cuspidatus Attenuates LPS-Induced Inflammatory Responses by Inhibition of iNOS and COX-2 Expression through Suppression of NF-κB Activation.

It seems quite necessary to obtain effective substances from natural products against inflammatory response (IR) as there are presently clinical problems regarding accompanying side effects and lowered quality of life. This work aimed to investigate the abilities of hyssopuside (HY), a novel phenolic glycoside isolated from Hyssopus cuspidatus (H. cuspidatus), against IR in lipopolysaccharide (LPS)-induced RAW 264.7 cells and mouse peritoneal macrophages. The results indicated that HY could reduce nitric oxide (NO) production and inhibit the production and secretion of pro-inflammatory mediators including tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-1β (IL-1β) in LPS-stimulated macrophages. Moreover, data from the immunofluorescence study showed that HY suppressed nuclear translocation of nuclear factor-kappa B (NF-κB) upon LPS induction. The Western blot results suggested that HY reversed the LPS-induced degradation of IκB (inhibitor of NF-κB), which is normally required for the activation of NF-κB. Meanwhile, the overexpression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) diminished significantly with the presence of HY in response to LPS stimulation. On the other hand, HY had a negligible impact on the activation of mitogen-activated protein kinase (MAPK) pathways. Moreover, an in silico study of HY against four essential proteins/enzymes revealed that COX-2 was the most efficient enzyme for the interaction, and binding of residues Phe179, Asn351, and Ser424 with HY played crucial roles in the observed activity. The structure analysis indicated the typical characterizations with phenylethanoid glycoside contributed to the anti-inflammatory effects of HY. These results indicated that HY manipulated its anti-inflammatory effects mainly through blocking the NF-κB signal transduction pathways. Collectively, we believe that HY could be a potential alternative phenolic agent for alleviating excessive inflammation in many inflammation-associated diseases.

Anti-proliferation and anti-inflammation effects of corilagin in rheumatoid arthritis by downregulating NF-κB and MAPK signaling pathways

Ethnopharmacological relevanceThe dried aboveground part of Geranium Wilfordii Maxim. (G. Wilfordii) is a traditional Chinese herbal medicine named lao-guan-cao. It has long been used for dispelling wind-dampness, unblocking meridians, and stopping diarrhea and dysentery. Previous investigations have revealed that 50% ethanolic extract of G. Wilfordii has anti-inflammatory and anti-proliferation activities on TNF-α induced murine fibrosarcoma L929 cells. Corilagin (COR) is a main compound in G. Wilfordii with the content up to 1.69 mg/g. Pharmacology study showed that COR has anti-inflammatory, anti-tumor, anti-microorganism, anti-oxidant, and hepatoprotective effects. However, there is no any investigation on its anti-proliferation and anti-inflammation effects in rheumatoid arthritis (RA). Aim of the studyThe present study aimed to evaluate the potential pharmacological mechanisms of anti-proliferation and anti-inflammation effects of COR in RA. Materials and methodsIn vitro, MH7A cells model induced by IL-1β was used. The anti-proliferation activity of COR was assessed by Cell Counting Kit-8 (CCK-8) assay, and the anti-migration and anti-invasion activity of COR was determined by wound healing assay and transwell assay, respectively. Furthermore, apoptosis assay by flow cytometer was used to measure the pro-apoptotic effect of COR. The mRNA expressions of Bax, Bcl-2, IL-6, IL-8, MMP-1, MMP-2, MMP-3, MMP-9, COX-2, and iNOS were measured by qRT-PCR, and related protein were further verified by ELISA kits or Western blot. Moreover, protein levels associated with NF-κB and MAPK signaling pathways of p65, P-p65, IκBα, P-IκBα, ERK1/2, P-ERK1/2, JNK, P-JNK1/2/3, p38, and P-p38 were determined by Western blot. The nuclear translocation of NF-κB-p65 was detected by immunofluorescent staining. In vivo, adjuvant-induced arthritis (AIA) rat model was used, and the body weight, paw swelling, and arthritis score during the entire period were measured. Histopathological analysis of joints of synovial tissues was also determined. The expression of pro-inflammatory cytokines in serum including IL-6, TNF-α, IL-1β, and IL-17 were measured. ResultsThe in vitro results showed that COR could dose-dependently inhibit the proliferation, migration, and invasion of IL-1β-induced MH7A cells, as well as promote its apoptosis. Moreover, it also suppressed the over-expression of Bcl-2, IL-6, IL-8, MMP-1, MMP-2, MMP-3, MMP-9, COX-2, and iNOS while up-regulated the level of Bax. Besides, the ratios of P-p65/p65, P-IκBα/IκBα, P-ERK/ERK, P-JNK/JNK, and P-p38/p38 were decreased, and the nuclear translocation of p65 induced by IL-1β was blocked by COR. In vivo results indicated that COR significantly reduced the paw swelling and arthritis score in AIA rats, and inhibited synovial tissue hyperplasia and erosion, as well as inflammatory cells infiltration. It also decreased the serum pro-inflammatory cytokines (IL-6, TNF-α, IL-1β, and IL-17) production. ConclusionThese results revealed that COR exerted anti-rheumatoid arthritis effect, and its underlying mechanisms may be related to inhibiting the proliferation, migration, and invasion of synovial fibroblasts, enhancing cell apoptosis, and suppressing inflammatory responses via downregulating NF-κB and MAPK signaling pathways.

LncRNA XIST Prevents Tendon Adhesion and Promotes Tendon Repair Through the miR-26a-5p/COX2 Pathway.

Tendon adhesion is the biggest obstacle to repair of tendon injury. Long-chain non-coding RNA X-inactive specific transcript (lncRNA XIST) is highly expressed in populations at high risk of tendon injury. However, whether XIST participates in tendon injury and the specific mechanism remain unknown. Here, we aimed to explore the effects and underlying mechanism of XIST in tendon injury. A mouse model of tendon injury was constructed by the transection method in vivo. XIST and COX2 were highly expressed in tendon tissues of mice with tendon injury, while miR-26a-5p was lowly expressed. Fibroblasts were isolated from tendon injury mice. Overexpression of XIST promoted fibroblast proliferation and upregulated α-SMA and Collagen I protein expression, while silencing XIST indicated the opposite effects. Further dual-luciferase reporter gene assay and RIP assay verified a targeting relationship between XIST and miR-26a-5p, as well as miR-26a-5p and COX2, and XIST targeted miR-26a-5p to act on COX2 expression. miR-26a-5p inhibition and COX2 overexpression reversed the decrease in fibroblast proliferation and the downregulation of α-SMA and Collagen I expression caused by XIST silencing, while interference with si-COX2 eliminated the effects of miR-26a-5p inhibitor. This study revealed that XIST promoted fibroblast proliferation and the formation of tendon adhesion through miR-26a-5p/COX2 pathway, suggesting that XIST/miR-26a-5p/COX2 may be a potential target for the treatment of tendon injury.

Cell Proliferation and Tumor Induction by Ochratoxin A in Mouse Skin and Evaluation of Cyclin D1 and Cyclooxygenase-2 Expressions.

Motivation. Skin tumor is one of the frequent occurring forms of cancer where 2-3 million instances are reported worldwide. The ultraviolet rays along with the environmental pollutants and other contaminants can be the potential factors of skin cancer. Cyclin D1 is a serious gene included in controlling the development through the G1 phase of the cell cycle. Ochratoxin A (OTA) is a naturally existing mycotoxin which majorly occurs in food like grains. It is responsible for producing the splitting of single-strand DNA and is identified to be cancer-causing. It is established as a critical risk factor towards reproductive health in both males and females. Methodology. A single dose of ochratoxin A was used for topical application for assessment of skin tumor promotion activity, hyperplasia, ornithine decarboxylase activity, and expression of cyclin D1 and COX-2 in mouse skin. Enhancement in the synthesis of DNA, activation of the epidermal growth factor receptor, and overexpression of cyclin D1 and COX-2 were noted. Primary murine keratinocyte cell culture was cultured with Waymouth's medium. Western blot analysis and real-time polymerase chain reaction (RT-PCR) were used to detect the expression of cyclin D1 and COX-2. Chromatin immunoprecipitation (ChIP) assays were used to the association between AP-1 transcription and nuclear factor-kappaB (NF-κB) with COX-2 and cyclin D1 promoters. Results. The results found that cyclin D1 and COX-2 were responsible for stimulating OTA-induced PMK proliferation and hyperplasia. Implications. EGFR-mediated pathways were also responsible for tumor promotion due to OTA.

Celecoxib reduces hepatic vascular resistance in portal hypertension by amelioration of endothelial oxidative stress

The balance between endothelial nitric oxide (NO) synthase (eNOS) activation and production of reactive oxygen species (ROS) is very important for NO homeostasis in liver sinusoidal endothelial cells (LSECs). Overexpression of cyclooxygenase‐2 (COX‐2), a major intravascular source of ROS production, has been observed in LSECs of cirrhotic liver. However, the links between low NO bioavailability and COX‐2 overexpression in LSECs are unknown. This study has confirmed the link between low NO bioavailability and COX‐2 overexpression by COX‐2‐dependent PGE2‐EP2‐ERK1/2‐NOX1/NOX4 signalling pathway in LSECs in vivo and in vitro. In addition, the regulation of COX‐2‐independent LKB1‐AMPK‐NRF2‐HO‐1 signalling pathway on NO homeostasis in LSECs was also elucidated. The combinative effects of celecoxib on diminishment of ROS via COX‐2‐dependent and COX‐2‐independent signalling pathways greatly decreased NO scavenging. As a result, LSECs capillarisation was reduced, and endothelial dysfunction was corrected. Furthermore, portal hypertension of cirrhotic liver was ameliorated with substantial decreasing hepatic vascular resistance and great increase of portal blood flow. With the advance understanding of the mechanisms of LSECs protection, celecoxib may serve as a potential therapeutic candidate for patients with cirrhotic portal hypertension.

Scutellarein relieves the death and inflammation of tubular epithelial cells in ischemic kidney injury by degradation of COX-2 protein

ObjectivesAcute kidney injury (AKI) is a clinical syndrome that usually caused by ischemia/reperfusion (I/R). Previous studies have revealed the protection of scutellarein against ischemia in nervous system. This study aimed to demonstrate the potential of scutellarein in ischemic AKI. MethodsAnimal model of ischemic AKI was established by clamping bilateral kidney pedicles in Sprague-Dawley rats. HK-2 cells were exposed to oxygen glucose deprivation/reoxygenation (OGD/R) to induce a cell model of AKI. The effects of scutellarein pre-treatment were detected by H&E staining, TUNEL, ELISA, CCK-8, LDH activity assay, ROS generation, flow cytometry, qRT-PCR and western blotting. Bioinformatic analysis was performed to probe the targets of scutellarein. ResultsThe blood urea nitrogen (BUN) and serum creatinine (SCr) levels in rats treated with scutellarein were lower than that in model groups. Scutellarein suppressed the pathological injury of kidney, and dose-dependently inhibited the apoptosis and pro-inflammatory cytokines release (IL-1β, IL-6 and IL-18). Scutellarein prevented OGD/R-induced HK-2 cell loss and cytotoxicity. ROS generation, apoptosis, and inflammation induced by OGD/R were all inhibited by scutellarein. By searching on the TCMSP and Symmap databases, COX-2 was screened out as a target of scutellarein. Scutellarein has no significant impacts on COX-2 mRNA expression, but could inhibit its protein level. Scutellarein promoted COX-2 protein degradation via enhancing autophagy. Furthermore, overexpression of COX-2 partly eliminated the renal protection of scutellarein in HK-2 cells. ConclusionsScutellarein was suggested as a renal protective agent against ischemia-induced damage in AKI. The protective properties of scutellarein may be through inhibition of COX-2.

A Systematic Review of the Cyclooxygenase-2 (COX-2) Expression in Rectal Cancer Patients Treated with Preoperative Radiotherapy or Radiochemotherapy.

The main objective of this systematic review is to investigate the expression level of the cyclooxygenase-2 (COX-2) in rectal cancer treated with either preoperative radiotherapy or radiochemotherapy. In addition, we have summarized the effects of preoperative treatment of rectal cancer with regards to the expression levels of COX-2. A systematic literature review was performed in The Cochrane Library, PubMed, Web of Science, and Scopus databases on 1 January 2021 with the usage of the following search string—(cyclooxygenase-2) OR (COX-2) AND (rectal cancer) AND (preoperative radiochemotherapy) OR (preoperative radiotherapy). Among the 176 included in the analysis, only 13 studies were included for data extraction with a total number of 2095 patients. The results of the analysis are based on the articles concerning the expression of COX-2 in rectal cancer among patients treated with preoperative radiotherapy or radiochemotherapy. A COX-2 expression is an early event involved in rectal cancer development. In cases of negative COX-2 expression, radiotherapy and radiochemotherapy might contribute to the reduction of a local recurrence. Therefore, COX-2 may be considered as a biologic factor while selecting patients for more effective, less time-consuming and less expensive preoperative treatment. However, the utility of the administration of COX-2 inhibitors to patients with COX-2 overexpression, in an attempt to improve the patients’ response rate to the neoadjuvant treatment, needs an assessment in further clinical trials.

Missense mutations involvement in COX-2 structure, and protein-substrate binding affinity: in-silico study

Cyclooxygenase-2 (COX-2) is an inducible inflammatory enzyme, which produces prostanoids from arachidonic acid. COX-2 overexpression and over-activity can cause inflammation, tumorigenesis, and angiogenesis. Prostanoids are the main reason for the inflammation, and increase of mitogenesis by COX-2. So, any change such as mutations that can lead to COX-2 over-activity could ignite the tumor situations with increase of prostanoids production is one of its ways. The aim of this study was to check the effect of 166 missense mutations of COX-2 on protein features that can affect the COX-2 activity such as protein stability, fluctuation, 2D structure, and its binding affinity with the substrate by in silico methods, network modeling, and docking calculations, by which 44 of them shown to be deleterious. Among them, the S124I and S474F mutations can increase the stability of the protein. 11.36% of deleterious nsSNPs were part of the substrate-binding region among which the M508T, H337R, and V511G have the potential to affect the protein by 2D structure alteration. V511G can improve binding affinity and H337R showed a small decrease in the deformation overall energy that can represent a decrease in the stability of COX-2. Also, L517S showed a significant decrease in the binding power of COX-2/substrate but based on the anisotropic network modeling this mutation has a dual effect on COX-2 stability. These nsSNPs/mutations have the potential causing an increase or decrease of tumorigenesis because increasing of COX-2 stability and its binding affinity can lead to altering its activity.

Inflammation and Hypoxia Negatively Impact the Survival and Immunosuppressive Properties of Mesenchymal Stromal Cells In Vitro

Abstract Mesenchymal stromal cells (MSC) are nonhematopoietic cells with fibroblast-like morphology and multipotent capacity that are widely used in pre-clinical and clinical investigations. Unfortunately, the efficiency of MSC treatment is hindered by the poor survival rate after transplantation at the damaged tissue. The goal of this study was to investigate the fate of MSC exposed to various stimuli mimicking the in vivo microenvironment post transplantation. To this aim, murine bone marrow–derived MSC were stimulated with IFNγ and TNFα under low oxygen (hypoxia) or atmospheric (normoxia) conditions for 24 to 72 hours, in order to better mimic an ischemic injury. The results showed that MSC pre-stimulation with TNFα and IFNγ enhanced immunosuppressive pathways by over-expression of NOS2, IDO, COX2 and production of NO. However, MSC viability was affected by these two cytokines in dose-dependent and time-dependent manners. Besides, priming with TNFα and/or IFNγ under low oxygen concentrations revealed that significantly increased cell mortality rate and decreased NO production. Our data suggest that both hypoxia and inflammation could impact the cell survival after transplantation and reinforces the necessity of further investigations to better understand MSC behavior after transplantation in order to identify the MSC-based strategies with the highest therapeutic potential.

Diclofenac down-regulates COX-2 induced expression of CD44 and ICAM-1 in human HT29 colorectal cancer cells.

Cyclooxygenase-2 (COX-2) is expressed in a variety of human colorectal cancer cells and can contribute to carcinogenesis. This study aimed to investigate the effect of diclofenac (DCF), a selective COX-2 inhibitor, on cell adhesion molecules and apoptosis in human colon adenocarcinoma cells. Levels of homing cell adhesion molecule (H-CAM, CD44), intercellular adhesion molecule-1 (ICAM-1, CD54), vascular cell adhesion molecule-1 (VCAM-1, CD106), and epithelial cell adhesion molecule (EpCAM, CD326) were evaluated in cancer cells overexpressing (HT29) or not expressing (HCT116) COX-2. Cell viability was determined by MTT assay, COX-2 protein levels and activity were assessed by immunofluorescence and fluorometric analysis, respectively. Endogenous levels of polyunsaturated fatty acids (PUFAs) were measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS) while expression of cell adhesion molecules was analyzed by flow cytometry. Annexin V-FITC/propidium iodide-labelling and fluorometric caspase-3 activity measurements were carried out to determine apoptosis. Flow cytometry analysis revealed that the percentage of CD44 and ICAM-1 staining in HCT116 cells was significantly lower compared to HT29 cells. In HT29 cells, phorbol 12-myristate 13-acetate (PMA) induced COX-2 expression and increased CD44 and ICAM-1 levels were down-regulated by diclofenac. Stimulation of COX-2 activity in HT29 cells via PMA significantly decreased diclofenac associated increase in PUFA levels. Treatment with both diclofenac and PMA significantly increased the number of apoptotic cells and caspase-3 activity in colon adenocarcinoma cells compared to control groups. In conclusion, diclofenac's effect to retard colorectal tumor growth and metastasis occurs in COX-2 overexpressing colon cancer cells by increased apoptosis and decreased expression of CD44 and ICAM-1.

DNMT3B decreases extracellular matrix degradation and alleviates intervertebral disc degeneration through TRPA1 methylation to inhibit the COX2/YAP axis.

Intervertebral disc degeneration (IVDD) is a main cause of low back pain that is associated with extracellular matrix (ECM) degradation and inflammation. This study aims to investigate the role of DNMT3B and its regulatory mechanisms in IVDD. IVDD rat models were constructed followed by transfections with oe-DNMT3B or oe-YAP in order to explore the role of DNMT3B in the development of IVDD. After that transfection, nucleus pulposus (NP) cells were isolated and transfected with oe-DNMT3B, oe-TRPA1, si-YAP, oe-YAP or oe-COX2 in order to investigate the functions of DNMT3B in NP cells. DNMT3B was poorly expressed in IVDD tissues and NP cells whereas TRPA1, COX2, and YAP were highly expressed. The proliferation or apoptosis of NP cells was detected through CCK-8 assay or flow cytometry, respectively. Overexpression of DNMT3B promoted the proliferation of NP cells, inhibited their apoptosis, as well as increasing the expression of collagen II and aggrecan and decreasing expression of MMP3 and MMP9. Besides, DNMT3B suppressed inflammation and alleviated IVDD. Mechanistically, DNMT3B modified the TRPA1 promoter by methylation to inhibit the expression of COX2. Overexpression of COX2 promoted the apoptosis of NP cells and decreased the expression of YAP, which was reversed by upregulating DNMT3B. DNMT3B may promote the proliferation of NP cells and prevent their ECM degradation through the TRPA1/COX2/YAP axis, thereby alleviating IVDD in rats.

Cyclooxygenase-2 regulates PTHrP transcription in human articular chondrocytes and is involved in the pathophysiology of osteoarthritis in rats.

BackgroundCyclooxygenase-2 (COX-2) inhibitors are prescribed for the management of osteoarthritis (OA)-associated pain and inflammation. However, the role of COX-2 in normal and osteoarthritic articular chondrocytes has not been well investigated. We hypothesize that COX-2 plays a role in articular chondrocytes under normal conditions and during OA progression.MethodsIn vivo COX-2 levels in articular cartilage of normal and papain-induced osteoarthritic rats were compared. The role of COX-2 in human articular chondrocytes (HACs) was tested in vitro by COX-2 overexpression or activity inhibition. The levels of COX-2 and marker gene for normal function or articular cartilage degeneration were evaluated: mRNA by qRT-PCR; proteins by western blotting or immunohistochemistry; and glycosaminoglycan (GAG) by Safranin O–fast green staining. Parathyroid hormone-related protein (PTHrP) promoter activity was detected with luciferase reporter assays.ResultsIn the OA rat study, COX-2 and PTHrP were simultaneously increased in osteoarthritic rat chondrocytes, while increased PTHrP levels were reduced by celecoxib, a COX-2 selective inhibitor. The levels of normal cartilage matrices, GAG and type II collagen decreased, while markers of degeneration, collagen type X and MMP13 were elevated in osteoarthritic articular chondrocytes. Celecoxib rescued the loss of GAG and the increased collagen type X and MMP13 levels. In vitro, COX-2 overexpression in HACs significantly increased Col2a1, Col10a1, PTHrP and MMP13 mRNA expression, which was decreased when COX-2 activity was suppressed. More importantly, COX-2 overexpression upregulated the PTHrP transcription, mRNA expression and protein levels.ConclusionCOX-2 plays a pathophysiological role by preventing terminal differentiation of articular chondrocytes by upregulating PTHrP expression at the early stage of OA progression.The Translational potential of this articleCOX2 up-regulates PTHrP expression in normal and OA articular chondrocytes.

CURCUMIN NANOPARTICLES VERSUS CURCUMIN IN AMELIORATION OF INFLAMMATORY AND PATHOLOGICAL CHANGES DURING THE MIGRATORY PHASE OF MURINE TRICHINELLOSIS

Trichinella spiralis can cause systemic inflammatory manifestations all over the body beforehabituating their destination in the striated muscles. Its new borne larvae (NBL) most dangerousphase go via bloodstream of different organs during migration.This study explored the anti-inflammatory, antioxidant and anti-apoptotic effects of Curcumin(Cur) and Curcumin nanoparticles (Cur-Nano) on inflammatory and pathological changes occurredin different organs in murine trichinellosis during larval migratory phase.Forty (40) male Swiss albino mice were divided into four groups of ten mice each. GI: miceinfected and treated with Cur, GII: mice infected and treated with Cur-Nano, GIII: infected nontreatedmice (positive control) and GIV: non-infected non-treated (negative control). Mice wereinfected orally with 200 T. spiralis larvae per mouse. GI & GII received treatment on 13th daypost infection (dpi) for 5 successive days. All mice were sacrificed on the 30th dpi. Effects of Cur& Cur-Nano were evaluated by counting T. spiralis encysted larvae in muscle by light microscope.Frozen sera (-20°C) were used for quantitative estimation of ALT, AST, TNFα, transforminggrowth factor-β (TGF-β) and Troponin (Trop). Specimens of lung, liver, brain and heartwere fixed in neutral buffered formalin for H&E and immunohistochemical staining.The results showed that number of encysted T. spiralis larvae in GI & GII was significantlyreduced compared to GIII. In both treated groups (GI & GII), there was a significant reduction inmean levels of AST, ALT, TNFα, TGF-β, & Trop. There was significant improvement of bileduct injury and proliferation, alveolar and pleural inflammation, and bronchial epithelial proliferation,as well as improvement of degenerative changes in brain and heart. The infected nontreatedgroup (GIII) showed significant overexpression of Cyclooxygenase-2 (COX2), caspase 3,glial fibrillary acidic protein (GFAP), arginase and granzyme b, with a significant low expressionof peroxisome proliferator-activated receptors (PPARs) as compared to treated groups.