OBJECTIVE: To investigate the sequences of internal transcribed spacer 2 (ITS2) and cyclooxygenase 1 (COX1) genes of Paragonimus metacercariae in freshwater crabs in Henan Province, identify the species of Paragonimus and evaluate its genetic relationships with Paragonimus isolates from other provinces in China. METHODS: Freshwater crabs were collected from 8 survey sites in Zhengzhou, Luoyang, Pingdingshan, Nanyang and Jiyuan cities of Henan Province from 2016 to 2021, and Paragonimus metacercariae were detected in freshwater crabs. Genomic DNA was extracted from Paragonimus metacercariae, and the ITS2 and COX1 genes were amplified using PCR assay, followed by sequencing of PCR amplification products. The gene sequences were spliced and aligned using the software DNASTAR, and aligned with the sequences of Paragonimus genes in the GenBank. Phylogenetic trees were created using the MEGA6 software with the Neighbor-Joining method based on ITS2 and COX1 gene sequences, with Fasciola hepatica as the outgroup. RESULTS: The detection rates of Paragonimus metacercariae were 6.83% (11/161), 50.82% (31/61), 18.52% (5/26), 8.76% (12/137), 14.29% (9/63), 17.76% (19/105), 18.50% (32/173) and 42.71% (41/96) in freshwater crabs from 8 survey sites in Zhengzhou, Luoyang, Pingdingshan, Nanyang and Jiyuan cities of Henan Province, with a mean detection rate of 19.46% (160/822), and a mean infection intensity of 0.57 metacercariae/g. The amplified ITS2 and COX1 gene fragments of Paragonimus were approximately 500 bp and 450 bp in lengths, respectively. The ITS2 gene sequences of Paragonimus metacercariae from 8 survey sites of Henan Province showed the highest homology (99.8% to 100.0%) with the gene sequence of P. skrjabini (GenBank accession number: MW960209.1), and phylogenetic analysis showed that the Paragonimus in this study was clustered into the same clade with P. skrjabini from Sichuan Province (GenBank accession number: AY618747.1), Guangxi Zhuang Autonomous Region (GenBank accession number: AY618729.1) and Hubei Province (GenBank accession number: AY618751.1), and P. miyazaki from Fujian Province (GenBank accession number: AY618741.1) and Japan (GenBank accession number: AB713405.1). The COX1 gene sequences of Paragonimus metacercariae from 8 survey sites of Henan Province showed the highest homology (90.0% to 100.0%) with the gene sequence of P. skrjabini (GenBank accession number: AY618798.1), and phylogenetic analysis showed that the Paragonimus in this study was clustered into the same clade with all P. skrjabini and clustered into the same sub-clade with P. skrjabini from Hubei Province (GenBank accession numbers: AY618782.1 and AY618764.1). CONCLUSIONS Paragonimus species from freshwater crabs in Henan Province were all characterized as P. skrjabini, and the ITS2 and COX1 gene sequences had the highest homology to those of P. skrjabini from Hubei Province. The results provide insights into study of Paragonimus in Henan Province and China.
Animals ; Paragonimus/genetics* ; Brachyura/genetics* ; Cyclooxygenase 1/genetics* ; Phylogeny ; China/epidemiology* ; Sequence Analysis, DNA ; Paragonimiasis

Objective To investigate the effect and mechanism of isorhapontigenin (ISO) in the protection of mice from the lipopolysaccharide (LPS)-induced acute lung injury (ALI). Methods RAW264.7 cells were cultured in vitro with different concentrations of ISO and the viability of the cells was measured by CCK-8 assay.Further,RAW264.7 cells were induced with 200 ng/ml LPS and then treated with ISO and the autophagy inhibitor 3-methyladenine (3-MA).Western blotting was employed to determine the expression of inflammatory cytokines [interleukin (IL)-1β,IL-6,tumor necrosis factor-α (TNF-α),P65,phospho-P56 (p-P65),IκB,phospho-IκB (p-IκB),inducible nitric oxide synthase (iNOS),cyclooxygenase-2 (COX-2),and high mobility group box-1 (HMGB1)] and autophagy markers (LC3Ⅱ/Ⅰ,Beclin1,and P62).The reactive oxygen species (ROS) production of the cells was measured with the DCFH-DA probe.The mouse model of ALI was established by intraperitoneal injection of LPS (15 mg/kg).The pathological changes of the lung tissue were observed via HE staining.The expression of inflammatory cytokines and autophagy markers in the lung tissue was determined by Western blotting and the content of ROS in bronchoalveolar lavage fluid (BALF) by flow cytometry. Results ISO down-regulated the expression of IL-1β,IL-6,TNF-α,iNOS,COX-2,and HMGB1 and inhibited the ROS production in the LPS-induced RAW264.7 cells (all P<0.05).Furthermore,it promoted the expression of LC3Ⅱ/Ⅰ and Beclin1 and inhibited the expression of P62,thereby activating autophagy (all P<0.05).However,the addition of 3-MA up-regulated the expression of p-P65/P65,p-IκB,iNOS,COX-2,and HMGB1,down-regulated that of IκB (all P<0.001),and promote the production of ROS.ISO mitigated the pathological changes in the lung tissue of ALI mice.It down-regulated the expression of p-P65/P65,p-IκB,iNOS,COX-2,and HMGB1 and up-regulated that of IκB in the lung tissue (all P<0.001) and decreased the ROS production in BALF.However,such protective effect was reversed by 3-MA. Conclusion ISO may induce autophagy of macrophages to protect mice from LPS-induced ALI.
Animals ; Mice ; Acute Lung Injury/pathology* ; Beclin-1/pharmacology* ; Cyclooxygenase 2/metabolism* ; Cytokines ; HMGB1 Protein/metabolism* ; Interleukin-6 ; Lipopolysaccharides ; Lung/pathology* ; NF-kappa B/metabolism* ; Reactive Oxygen Species/metabolism* ; Tumor Necrosis Factor-alpha/metabolism*

OBJECTIVE: To elucidate the underlying mechanism of Panax notoginseng saponin (PNS) on gastric epithelial cell injury and barrier dysfunction induced by dual antiplatelet (DA). METHODS: Human gastric mucosal epithelial cell (GES-1) was cultured and divided into 4 groups: a control, a DA, a PNS+DA and a LY294002+PNS+DA group. GES-1 apoptosis was detected by flow cytometry, cell permeability were detected using Transwell, level of prostaglandins E2 (PGE2), 6-keto-prostaglandin F1α (6-keto-PGF1α) and vascular endothelial growth factor (VEGF) in supernatant were measured by enzyme linked immunosorbent assay (ELISA), expression of phosphatidylinositide 3-kinase (PI3K), phosphorylated-PI3K (p-PI3K), Akt, phosphorylated-Akt (p-Akt), cyclooxygenase-1 (COX-1), cyclooxygenase-2 (COX-2), glycogen synthase kinase-3β (GSK-3β) and Ras homolog gene family member A (RhoA) were measured by Western-blot. RESULTS: DA induced apoptosis and hyper-permeability in GES-1, reduced supernatant level of PGE2, 6-keto-PGF1α and VEGF (P<0.05). Addition of PNS reduced the apoptosis of GES-1 caused by DA, restored the concentration of PGE2, 6-keto-PGF1α and VEGF (P<0.05). In addition, PNS attenuated the alteration of COX-1 and COX-2 expression induced by DA, up-regulated p-PI3K/p-Akt, down-regulated RhoA and GSK-3β. LY294002 mitigated the effects of PNS on cell apoptosis, cell permeability, VEGF concentration, and expression of RhoA and GSK-3β significantly. CONCLUSIONS PNS attenuates the suppression on COX/PG pathway from DA, alleviates DA-induced GES-1 apoptosis and barrier dysfunction through PI3K/Akt/ VEGF-GSK-3β-RhoA network pathway.
Cyclooxygenase 1 ; Epithelial Cells/metabolism* ; Glycogen Synthase Kinase 3 beta ; Humans ; Panax notoginseng ; Phosphatidylinositol 3-Kinases/metabolism* ; Platelet Aggregation Inhibitors ; Proto-Oncogene Proteins c-akt/metabolism* ; Saponins/pharmacology* ; Vascular Endothelial Growth Factor A ; rhoA GTP-Binding Protein

Nonsteroidal anti-inflammatory drug (NSAID)-exacerbated respiratory disease (NERD) has attracted a great deal of attention because of its association with severe asthma. However, it remains widely underdiagnosed in asthmatics as well as the general population. Upon pharmacological inhibition of cyclooxygenase 1 by NSAIDs, production of anti-inflammatory prostaglandin E2 and lipoxins ceases, while release of proinflammatory cysteinyl leukotrienes increases. To determine the underlying mechanisms, many studies have attempted to elucidate the genetic variants, such as single nucleotide polymorphisms, responsible for alterations of prostaglandins and leukotrienes, but the results of these genetic studies could not explain the whole genetic pathogenesis of NERD. Accordingly, the field of epigenetics has been introduced as an additional contributor to genomic alteration underlying the development of NERD. Recently, changes in CpG methylation, as one of the epigenetic components, have been identified in target tissues of NERD. This review discusses in silico analyses of both genetic and epigenetic components to gain a better understanding of their complementary roles in the development of NERD. Although the molecular mechanisms underlying NERD pathogenesis remain poorly understood, genetic and epigenetic variations play significant roles. Our results enhance the understanding of the genetic and epigenetic mechanisms involved in the development of NERD and suggest new approaches toward better diagnosis and management.
Anti-Inflammatory Agents, Non-Steroidal ; Asthma ; Computer Simulation ; Cyclooxygenase 1 ; Diagnosis ; Dinoprostone ; Epigenomics ; Genetics ; Leukotrienes ; Lipoxins ; Methylation ; Polymorphism, Single Nucleotide ; Prostaglandins

BACKGROUND: Abnormal upregulation of prostaglandin E₂ (PGE₂) is considered to be a key oncogenic event in the development and progression of inflammation-associated human colon cancer. It has been reported that 15-hydroxyprostaglandin dehydrogenase (15-PGDH), an enzyme catabolizing PGE₂, is ubiquitously downregulated in human colon cancer. 15-Deoxy-Δ(12,14)-prostaglandin J₂ (15d-PGJ₂), a peroxisome proliferator-activated receptor γ ligand, has been shown to have anticarcinogenic activities. In this study, we investigate the effect of 15d-PGJ₂ on expression of 15-PGDH in human colon cancer HCT116 cells. METHODS: HCT116 cells were treated with 15d-PGJ₂ analysis. The expression of 15-PGDH in the treated cells was measured by Western blot analysis and RT-PCR. In addition, the cells were subjected to a 15-PGDH activity assay. To determine which transcription factor(s) and signaling pathway(s) are involved in 15d-PGJ₂-induced 15-PGDH expression, we performed a cDNA microarray analysis of 15d-PGJ₂-treated cells. The DNA binding activity of AP-1 was measured by an electrophoretic mobility shift assay. To determine whether the AP-1 plays an important role in the 15d-PGJ₂-induced 15-PGDH expression, the cells were transfected with siRNA of c-Jun, a major subunit of AP-1. To elucidate the upstream signaling pathways involved in AP-1 activation by 15d-PGJ₂, we examined its effect on phosphorylation of Akt by Western blot analysis in the presence or absence of kinase inhibitor. RESULTS: 15d-PGJ₂ (10 μM) significantly upregulated 15-PGDH expression at the mRNA and protein levels in HCT-116 cells. 15-PGDH activity was also elevated by 15d-PGJ₂. We observed that genes encoding C/EBP delta, FOS-like antigen 1, c-Jun, and heme oxygenase-1 (HO-1) were most highly induced in the HCT116 cells following 15d-PGJ₂ treatment. 15d-PGJ₂ increased the DNA binding activity of AP-1. Moreover, transfection with specific siRNA against c-Jun significantly reduced 15-PGDH expression induced by 15d-PGJ₂. 15d-PGJ₂ activates Akt and a pharmacological inhibitor of Akt, LY294002, abrogated 15d-PGJ₂-induced 15-PGDH expression. We also observed that an inhibitor of HO-1, zinc protoporphyrin IX, also abrogated upregulation of 15-PGDH and down-regulation of cyclooxygenase-2 expression induced by 15d-PGJ₂. CONCLUSIONS: These finding suggest that 15d-PGJ₂ upregulates the expression of 15-PGDH through AP-1 activation in colon cancer HCT116 cells.
Blotting, Western ; Colon ; Colonic Neoplasms ; Cyclooxygenase 2 ; DNA ; Down-Regulation ; Electrophoretic Mobility Shift Assay ; HCT116 Cells ; Heme Oxygenase-1 ; Heme ; Humans ; Oligonucleotide Array Sequence Analysis ; Oxidoreductases ; Peroxisomes ; Phosphorylation ; Phosphotransferases ; RNA, Messenger ; RNA, Small Interfering ; Transcription Factor AP-1 ; Transfection ; Up-Regulation ; Zinc

Syringa pinnatifolia Hemsl.( SP) is a representative Mongolian folk medicine with the effects of inhibiting Heyi related diseases,clearing heat and relieving pain. It has been used for the treatment of Heyi-induced heart tingling,heart palpitations,upset,insomnia and other symptoms. Total ethanol extract( T) and major fraction( M) of SP have been evaluated its anti-ischemic effects,and the mechanism was related to the regulation of cyclooxygenase( COX)-mediated inflammatory pathway and p53-mediated apoptosis pathway in our previous studies. This study reports the chemical fractionation on M by which to obtain subfractions( I and M_3),and the pharmacological evaluation of M,I,and M_3 against myocardial ischemia in mice. The result showed that I and M reduced the values of LVEDd and LVEDs,significantly increased EF and FS values,increased serum CK-MB and LDH levels in mice,and reduced in inflammatory cells infiltration and collagen deposition in the infarcted myocardial tissue,suggesting that M and I possess the same degree anti-myocardial is chemia equally whereas M_3 has no this effect. Related mechanism studies suggested that I can reduce the expression of COX-1,COX-2 and p53 protein in myocardial tissue in a dose-dependent manner. This study lays the foundation for further chemical segmentation and clarification of pharmacological substance groups,paving the way for the full use and benefits to be use of systematic biological methods to analyze the pharmacological basis of SP against myocardial ischemia.
Animals ; Cyclooxygenase 1/metabolism* ; Cyclooxygenase 2/metabolism* ; Heart/drug effects* ; Medicine, Mongolian Traditional ; Membrane Proteins/metabolism* ; Mice ; Myocardial Ischemia/drug therapy* ; Myocardium/metabolism* ; Plant Extracts/therapeutic use* ; Syringa/chemistry* ; Tumor Suppressor Protein p53/metabolism*

OBJECTIVETo detect the expression level of cyclooxygenase-1（COX-1） and cyclooxygenase-2（COX-2） in the platelet of iron deficiency anemia（IDA）women at childbearing age and to explore its correlation with the different indexes of anemia and platelets.

METHODSForty female IDA patients at childbearing age and 35 healthy controls were enrolled in this study. The Flow cytometry was used to detect the expression of platelet COX-1 and COX-2，the platelet aggregation function as examined by turbidimetric method，and the levels of serum ferritin were analyzed by electrochemical luminescence method，the leval of serum iron was determined by ELISA，and the correlation of different indexes was analyzed.

RESULTSCompared with healthy controls，the levels of platelet COX-1 and COX-2 were significantly lower in female IDA patients at Childbearing age（P<0.05），but platelet count（Plt），mean platelet volume（MPV） and platelet aggregation rate（PAgT）were not statistically different between the 2 groups（P > 0.05）. The expression level of platelet COX-1 positively correlated with those of Hb（r =0.623，P<0.01），serum iron（r =0.321，P<0.05） and HCT（r=0.305，P<0.05）. but the platelet COX-2 expression did not corelate with these indexs.

CONCLUSIONThe expression of platelet COX-1 and COX-2 in female IDA patients at Childbearing age markedly decrease，and the expression level of platelet COX-1 closely relates with the severity of anemia，that possesses reference value for clinical diagnosis of female IDA patients at Childbearing age..

In the present study, we carried out a phytochemical investigation of the ethanol extract of the aerial parts of Baeckea frutescens, which resulted in the isolation of two new flavonoid glycosides, myricetin 3-O-(5″-O-galloyl)-α-L-arabinofuranoside (1), 6-methylquercetin 7-O-β-D-glucopyranoside (2), one new methylchromone glycoside, 7-O-(4', 6'-digalloyl)-β-D-glucopyranosyl-5-hydroxy-2-methylchromone (3), together with three known compounds (4-6). The structures of these isolated compounds were established on the basis of 1D and 2D NMR techniques and chemical methods. The anti-inflammatory activities of the compounds 1-6 were evaluated for their inhibitory effects against cyclooxygenases-1 and -2 in vitro. Compounds 1-6 showed potent COX-1 and COX-2 inhibiting activities in vitro with IC values ranging from 1.95 to 5.54 μmol·L and ranging from 1.01 to 2.27 μmol·L, respectively.
Anti-Inflammatory Agents ; chemistry ; isolation & purification ; Cyclooxygenase 1 ; chemistry ; Cyclooxygenase 2 ; chemistry ; Cyclooxygenase Inhibitors ; chemistry ; isolation & purification ; Flavonoids ; chemistry ; isolation & purification ; Molecular Structure ; Myrtaceae ; chemistry ; Plant Components, Aerial ; chemistry ; Plant Extracts ; chemistry ; isolation & purification

Glucagon like peptide-1 (GLP-1) stimulates glucose-dependent insulin secretion. Dipeptidyl peptidase-4 (DPP-4) inhibitors, which block inactivation of GLP-1, are currently in clinical use for type 2 diabetes mellitus. Recently, GLP-1 has also been reported to have neuroprotective effects in cases of cerebral ischemia. We therefore investigated the neuroprotective effects of GLP-1 receptor (GLP-1R) agonist, exendin-4 (ex-4), after cerebral ischemia-reperfusion injury. Transient middle cerebral artery occlusion (tMCAO) was induced in rats by intracerebroventricular (i.c.v.) administration of ex-4 or ex9-39. Oxygen-glucose deprivation was also induced in primary neurons, bEnd.3 cells, and BV-2. Ischemia-reperfusion injury reduced expression of GLP-1R. Additionally, higher oxidative stress in SOD2 KO mice decreased expression of GLP-1R. Downregulation of GLP-1R by ischemic injury was 70% restored by GLP-1R agonist, ex-4, which resulted in significant reduction of infarct volume. Levels of intracellular cyclic AMP, a second messenger of GLP-1R, were also increased by 2.7-fold as a result of high GLP-1R expression. Moreover, our results showed that ex-4 attenuated pro-inflammatory cyclooxygenase-2 (COX-2) and prostaglandin E₂ after MCAO. C-Jun NH₂ terminal kinase (JNK) signaling, which stimulates activation of COX-2, was 36% inhibited by i.c.v. injection of ex-4 at 24 h. Islet-brain 1 (IB1), a scaffold regulator of JNK, was 1.7-fold increased by ex-4. GLP-1R activation by ex-4 resulted in reduction of COX-2 through increasing IB1 expression, resulting in anti-inflammatory neuroprotection during stroke. Our study suggests that the anti-inflammatory action of GLP-1 could be used as a new strategy for the treatment of neuroinflammation after stroke accompanied by hyperglycemia.
Animals ; Brain Ischemia ; Cyclic AMP ; Cyclooxygenase 2 ; Diabetes Mellitus, Type 2 ; Down-Regulation ; Glucagon* ; Glucagon-Like Peptide 1 ; Glucagon-Like Peptide-1 Receptor ; Hyperglycemia ; Infarction, Middle Cerebral Artery ; Insulin ; Mice ; Neurons ; Neuroprotection ; Neuroprotective Agents ; Oxidative Stress ; Phosphotransferases ; Rats ; Reperfusion Injury ; Second Messenger Systems ; Stroke*

Since previous studies have reported that hydroquinone (HQ) exerted immunosuppressive and anti-inflammatory activity, various HQ derivatives have been synthesized and their biological activities investigated. In this study, we explored the anti-inflammatory activity of JS-III-49, a novel HQ derivative, in macrophage-mediated inflammatory responses. JS-III-49 suppressed the production of the inflammatory mediators nitric oxide (NO) and prostaglandin E2 (PGE2) and down-regulated the mRNA expression of the inflammatory enzymes cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) as well as the expression of the pro-inflammatory cytokines interleukin-6 (IL-6) and IL-1b without cytotoxicity in LPS-stimulated RAW264.7 cells. JS-III-49 inhibited nuclear translocation of the NF-kB transcription factors p65 and p50 by directly targeting Akt, an upstream kinase of the NF-kB pathway, in LPS-stimulated RAW264.7 cells. However, JS-III-49 did not directly inhibit the kinase activities of Src and Syk, which are upstream kinases of Akt, in LPS-stimulated RAW264.7 cells. Moreover, JS-III-49 suppressed the nuclear translocation of c-Fos, one of the components of AP-1, by specifically targeting p38, an upstream mitogen-activated protein kinase (MAPK) in the AP-1 pathway in LPS-stimulated RAW264.7 cells. These results suggest that JS-III-49 plays an anti-inflammatory role in LPS-stimulated macrophages by targeting Akt and p38 in the NF-kB and AP-1 pathways, respectively.
Cyclooxygenase 2 ; Cytokines ; Dinoprostone ; Interleukin-6 ; Macrophages ; NF-kappa B ; Nitric Oxide ; Nitric Oxide Synthase Type II ; Phosphotransferases ; Protein Kinases ; RNA, Messenger ; Transcription Factor AP-1 ; Transcription Factors