[Sequence analysis of Paragonimus internal transcribed spacer 2 and cyclooxygenase 1 genes in freshwater crabs in Henan Province].

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.

15-lipoxygenase and cyclooxygenase expression profile and their related modulators in COVID-19 infection.

BACKGROUND: The role of the lipoxygenase (LOX) and cyclooxygenase (COX) enzymes in maintaining cellular homeostasis and regulating immune responses promoted us in this study to analyze the pattern of changes in 15-lipoxygenase and cyclooxygenase isoforms and their related cytokines in SARS-CoV-2 infection. METHODS: 15-LOX-1, 15-LOX-2, COX-1 and COX-2 gene expression levels were determined using qRT-PCR in nasopharynx specimens from patients with severe [N = 40] and non-severe [N = 40] confirmed SARS-CoV-2 infections and healthy controls. Circulating levels of lL-6, lL-10, PGE2, and IFN-γ were measured in patients and healthy controls using ELISA assay. The associations between the measured variables and the patient's clinic-pathological characteristics were assessed for all groups. RESULTS: The expression level of 15-LOX-1 was elevated significantly in male patients with severe infection; although female patients showed a different expression profile. 15-LOX-2 expression level was considerably increased in male patients with severe infection; while changes in its expression remained inconclusive in female patients. The relationship between 15-LOX expression and the male gender was prominent. Both COX isoforms expression showed elevation in male and female patients that were correlated with disease severity. The simultaneous increase in lL-6, PGE2 and IFN-γ levels also decrease in lL-10 in patients with severe infection indicating the possible regulatory network related to the COX and 15-LOX enzymes in the output of the SARS-CoV-2 infection. CONCLUSION: The results of this study determined the pattern of possible changes in key enzymes of prostaglandin and eicosanoids synthesis pathway and their mediators, which can be helpful in mapping the SARS-CoV-2 pathogenicity and pharmaceutical approaches.

The PTGS1 (rs1330344) CC Genotype Contributes to Susceptibility to Kawasaki Disease in Southern Chinese Children.

Kawasaki disease (KD) is an acute systemic vascular disease complicated by coronary artery injury. Although polymorphisms in prostaglandin-endoperoxide synthase 1 (PTGS1) are being increasingly explored in cardiovascular diseases, little is known regarding the connection between PTGS1 polymorphisms and KD risk. We evaluated 834 KD patients and 1474 healthy controls to explore the relationship between PTGS1 polymorphisms (rs1330344 and rs5788) and KD risk. Our results showed that the rs1330344 CC genotype was significantly associated with KD risk and coronary artery injury in children with KD. In combined analysis, individuals with 1-2 unfavorable genotypes had an increased risk of KD, compared with those with no risk genotype. Stratified analysis indicated that the rs1330344 CC genotype is strongly associated with increased risk of KD in children aged ≤60 months and females. Moreover, carrying 1-2 of these SNP genotypes had a higher risk of KD than those who harbored none of them in children ≤60 months of age and females; the risk of coronary artery dilatations/small aneurysms and medium/giant aneurysms was also significantly increased in KD patients. In summary, the PTGS1 rs1330344 CC genotype is associated with increased susceptibility to KD, which may contribute to KD pathogenesis and serve as a genetic biomarker.

The specific deletion of cyclooxygenase-1 in megakaryocytes/platelets reduces intestinal polyposis in ApcMin/+ mice.

Clinical and experimental evidence sustain the role of cyclooxygenase (COX)-1 in intestinal tumorigenesis. However, the cell type expressing the enzyme involved and molecular mechanism(s) have not been clarified yet. We aimed to elucidate the role of platelet COX-1 (the target of low-dose aspirin in humans) in intestinal tumorigenesis of ApcMin/+ mice, considered a clinically relevant model. To realize this objective, we generated an ApcMin/+ mouse with a specific deletion of Ptgs1(COX-1 gene name) in megakaryocytes/platelets (ApcMin/+;pPtgs1-/-mice) characterized by profound inhibition of thromboxane(TX)A2 biosynthesis ex vivo (serum TXB2; by 99%) and in vivo [urinary 2,3-dinor-TXB2(TXM), by 79%]. ApcMin/+ mice with the deletion of platelet COX-1 showed a significantly reduced number (67%) and size (32%) of tumors in the small intestine. The intestinal adenomas of these mice had decreased proliferative index associated with reduced COX-2 expression and systemic prostaglandin(PG)E2 biosynthesis (urinary PGEM) vs. ApcMin/+mice. Extravasated platelets were detected in the intestine of ApcMin/+mice. Thus, we explored their contribution to COX-2 induction in fibroblasts, considered the primary polyp cell type expressing the protein. In the coculture of human platelets and myofibroblasts, platelet-derived TXA2 was involved in the induction of COX-2-dependent PGE2 in myofibroblasts since it was prevented by the selective inhibition of platelet COX-1 by aspirin or by a specific antagonist of TXA2 receptors. In conclusion, our results support the platelet hypothesis of intestinal tumorigenesis and provide experimental evidence that selective inhibition of platelet COX-1 can mitigate early events of intestinal tumorigenesis by restraining COX-2 induction.

Human mtRF1 terminates COX1 translation and its ablation induces mitochondrial ribosome-associated quality control.

Translation termination requires release factors that read a STOP codon in the decoding center and subsequently facilitate the hydrolysis of the nascent peptide chain from the peptidyl tRNA within the ribosome. In human mitochondria eleven open reading frames terminate in the standard UAA or UAG STOP codon, which can be recognized by mtRF1a, the proposed major mitochondrial release factor. However, two transcripts encoding for COX1 and ND6 terminate in the non-conventional AGA or AGG codon, respectively. How translation termination is achieved in these two cases is not known. We address this long-standing open question by showing that the non-canonical release factor mtRF1 is a specialized release factor that triggers COX1 translation termination, while mtRF1a terminates the majority of other mitochondrial translation events including the non-canonical ND6. Loss of mtRF1 leads to isolated COX deficiency and activates the mitochondrial ribosome-associated quality control accompanied by the degradation of COX1 mRNA to prevent an overload of the ribosome rescue system. Taken together, these results establish the role of mtRF1 in mitochondrial translation, which had been a mystery for decades, and lead to a comprehensive picture of translation termination in human mitochondria.

Morphological Identification and Phylogenetic Analysis of Laelapin Mite Species (Acari: Mesostigmata: Laelapidae) from China.

Laelapinae mites are involved in transmission of microbial diseases between wildlife and humans, with an impact on public health. In this study, 5 mite members in the subfamily Laelapinae (laelapin mites; LM) were morphologically identified by light microscopy, and the phylogenetic relationship of LM was analyzed in combination with the sequence information of part of the LM cytochrome oxidase subunit I (cox1) gene. The morphological identification revealed that 5 mites belonged to the genera Laelaps and Haemolaelaps, respectively. Sequence analysis showed that the ratio of non-synonymous mutation rate to synonymous mutation rate of LM was less than 1, indicating that the LM cox1 gene had undergone purifying selection. Phylogenetic analysis showed that the Laelapinae is a monophyletic group. The genera Haemolaelaps and Hyperlaelaps did not separated into distinct clades but clustered together with species of the genus Laelaps. Our morphological and molecular analyses to describe the phylogenetic relationships among different genera and species of Laelapinae provide a reference for the improvement and revision of the LM taxonomy system.

The influence of a nanoparticle gel loaded with siRNA-cyclooxygenase on flexor tendon healing: an in vivo animal study.

We investigated the influence of cyclooxygenase (COX)-1 and COX-2 siRNAs delivered through a nanoparticle-gel system on the strength of flexor tendon repairs. Sixteen flexor digitorum profundus (FDP) tendons of chicken toes were transected, repaired and wrapped with gels to evaluate gel adherence. We found that the gel adhered to the tendon surface firmly. Next, 56 tendons were used in a first set of in vivo experiments to compare the therapeutic effects of different doses of COX siRNAs. Another 15 tendons were added in a second set to further assess the effects of a dosage of 12 µg. After 4 weeks, the mean strength of the repaired tendons increased most notably in the toes treated with 12 µg COX siRNAs, and the number of samples with low strength (<35 N) was significantly smaller than in the group without molecular treatment. We conclude that COX-1 and COX-2 siRNAs delivered through a nanoparticle-gel system increased the healing strength of the repaired tendons.

Association between Vitamin Intake and Chronic Kidney Disease According to a Variant Located Upstream of the PTGS1 Gene: A Cross-Sectional Analysis of Shika Study.

Chronic kidney disease (CKD) patients have been advised to take vitamins; however, the effects have been controversial. The individual differences in developing CKD might involve genetic variants of inflammation, including variant rs883484 located upstream of the prostaglandin-endoperoxide synthase 1 (PTGS1) gene. We aimed to identify whether the 12 dietary vitamin intake interacts with genotypes of the rs883484 on developing CKD. The population-based, cross-sectional study had 684 Japanese participants (≥40 years old). The study used a validated, brief, self-administered diet history questionnaire to estimate the intake of the dietary vitamins. CKD was defined as estimated glomerular filtration < 60 mL/min/1.73 m2. The study participants had an average age of 62.1 ± 10.8 years with 15.4% minor homozygotes of rs883484, and 114 subjects had CKD. In the fully adjusted model, the higher intake of vitamins, namely niacin (odds ratio (OR) = 0.74, 95% confidence interval (CI): 0.57−0.96, p = 0.024), α-tocopherol (OR = 0.49, 95% CI: 0.26−0.95, p = 0.034), and vitamin C (OR = 0.97, 95% CI: 0.95−1.00, p = 0.037), was independently associated with lower CKD tendency in the minor homozygotes of rs883484. The results suggested the importance of dietary vitamin intake in the prevention of CKD in middle-aged to older-aged Japanese with minor homozygous of rs883484 gene variant.

Infection of the Asian gray shrew Crocidura attenuata (Insectivora: Soricidae) with Sarcocystis attenuati n. sp. (Apicomplexa: Sarcocystidae) in China.

BACKGROUND: Data on the genus Sarcocystis in insectivores are limited. The Asian gray shrew Crocidura attenuata is one of the most common species of the insectivore family Soricidae in South Asia and Southeast Asia. To our knowledge, species of Sarcocystis have never been recorded previously in this host. METHODS: Tissues were obtained from 42 Asian gray shrews caught in 2017 and 2018 in China. Sarcocysts were observed using light microscopy (LM) and transmission electron microscopy (TEM). To describe the parasite life cycle, muscle tissues of the host infected with sarcocysts were force-fed to two beauty rat snakes Elaphe taeniura. Individual sarcocysts from different Asian gray shrews, and oocysts/sporocysts isolated from the small intestines and feces of the experimental snakes, were selected for DNA extraction, and seven genetic markers, namely, two nuclear loci [18S ribosomal DNA (18S rDNA) and internal transcribed spacer region 1 (ITS1)], three mitochondrial genes [cytochrome oxidase subunit 1 (cox1), cox3 and cytochrome b], and two apicoplast genes (RNA polymerase beta subunit and caseinolytic protease C), were amplified, sequenced and analyzed. RESULTS: Sarcocysts were found in 17 of the 42 (40.5%) Asian gray shrews. Under LM, the microscopic sarcocysts showed saw- or tooth-like protrusions measuring 3.3-4.5 µm. Ultrastructurally, the sarcocyst wall contained numerous lancet- or leaf-like villous protrusions, similar to those described for type 9h of the common cyst wall classification. The experimental beauty rat snakes shed oocysts/sporocysts measuring 11.9-16.7 × 9.2-10.6 µm with a prepatent period of 10-11 days. Comparison of the newly obtained sequences with those previously deposited in GenBank revealed that those of 18S rDNA and cox1 were most similar to those of Sarcocystis scandentiborneensis recorded in the tree shrews Tupaia minor and Tupaia tana (i.e., 97.6-98.3% and 100% identity, respectively). Phylogenetic analysis based on 18S rDNA or ITS1 sequences placed this parasite close to Sarcocystis spp. that utilize small animals as intermediate hosts and snakes as the known or presumed definitive host. On the basis of morphological and molecular characteristics and host specificity, the parasite was proposed as a new species, named Sarcocystis attenuati. CONCLUSIONS: Sarcocysts were recorded in Asian gray shrews, to our knowledge for the first time. Based on morphological and molecular characterization, a new species of parasite is proposed: Sarcocystis attenuati. According to the LM and TEM results, S. attenuati sarcocysts are distinct from those of Sarcocystis spp. in other insectivores and those of S. scandentiborneensis in tree shrews. The 18S rDNA or cox1 sequences of Sarcocystis attenuati shared high similarity with those of Sarcocystis scandentiborneensis, Sarcocystis zuoi, Sarcocystis cf. zuoi in the Malayan field rat, and Sarcocystis sp. in the greater white-toothed shrew. Therefore, we suggest that more research on the relationships of these closely related taxa should be undertaken in the future.

COX-1, COX-2 and CYP2C19 variations may be related to cardiovascular events due to acetylsalicylic acid resistance.

BACKGROUND: In some stent implanted patients, cardiovascular events (CE) may occur. Acetylsalicylic acid (ASA) is routinely administered to these patients in order to prevent the occurrence of CE. CE may be related to gene variations which cause ASA resistance (AR). Therefore, it was aimed to investigate the relationship between COX-1, COX-2, CYP2C9 and CYP2C19 variations with CE due to AR. MATERIALS AND RESULTS: Seventy-four stent implanted patients, using 100 mg of ASA per day during five years were enrolled into the study. Following stent implantation, thirty-eight patients who had a CE within five years due to AR and 36 patients without CE were enrolled in patient and control group, respectively. AR was confirmed by platelet aggregation testing. After DNA isolation from blood; COX-1, COX-2, CYP2C19 and CYP2C9 variations were investigated with real-time polymerase chain reaction. At the end of this study, heterozygous genotype of COX-1 was found statistically high in patients whereas heterozygous genotype of CYP2C19*17 was found statistically high in controls. The presence of C and G allele in COX-1 and COX-2 were found statistically high in patients, respectively. The presence of T allele in CYP2C19*17 was found statistically high in controls. Heterozygous genotype of COX-1 variation was found statistically high in patients who have AR. Additionally heterozygous genotype of CYP2C19*17 was found statistically high in patients who have low thrombosis risk. CONCLUSIONS: COX-1 and COX-2 gene mutations may increase the risk of CE due to AR whereas CYP2C19*17 may have a protective effect in this process.

Comparative subcellular localization of NRF2 and KEAP1 during the hepatocellular carcinoma development in vivo.

The activation of Nuclear Factor, Erythroid 2 Like 2 - Kelch Like ECH Associated Protein 1 (NRF2-KEAP1) signaling pathway plays a critical dual role by either protecting or promoting the carcinogenesis process. However, its activation or nuclear translocation during hepatocellular carcinoma (HCC) progression has not been addressed yet. This study characterizes the subcellular localization of both NRF2 and KEAP1 during diethylnitrosamine-induced hepatocarcinogenesis in the rat. NRF2-KEAP1 pathway was continuously activated along with the increased expression of its target genes, namely Nqo1, Hmox1, Gclc, and Ptgr1. Similarly, the nuclear translocation of NRF2, MAF, and KEAP1 increased in HCC cells from weeks 12 to 22 during HCC progression. Likewise, colocalization of NRF2 with KEAP1 was higher in the cell nuclei of HCC neoplastic nodules than in surrounding cells. Moreover, immunofluorescence analyses revealed that the interaction of KEAP1 with filamentous Actin was disrupted in HCC cells. This disruption may be contributing to the release and nuclear translocation of NRF2 since the cortical actin cytoskeleton serves as anchoring of KEAP1. In conclusion, this evidence indicates that NRF2 is progressively activated and promotes the progression of experimental HCC.

Musa sp. Leaves Extract Ameliorates the Hepato-Renal Toxicities Induced by Cadmium in Mice.

Heavy metals intoxication causes several health problems that necessitate finding new protective and therapeutic approaches. This study aimed to evaluate the impact of Musa sp. leaves extract (MLE) on hepato-renal toxicities induced by cadmium (Cd) in male mice. The phytochemical screening, metal chelating activity (MCA), and the median lethal dose (LD50) of MLE were determined. Fifty CD-1 male mice were used and intraperitoneally (i.p.) injected with MLE (1000 to 5000 mg/kg b.wt) for MLE LD50 determination. Another 50 mice were used for evaluating the effect of MLE on Cd toxicity. Blood samples were collected for hematological, liver, and kidney functions assessments. Liver tissue homogenates were used for determination of oxidant/antioxidant parameters. Liver and kidney tissues were harvested for histopathological and molecular investigations. MLE showed potent in vitro antioxidant activities. The MCA and LD50 of the MLE were 75 µg/mL and 3000 mg/kg b.wt, respectively. MLE showed beneficial therapeutic activity against hepato-renal toxicities in Cd-intoxicated mice, evidenced by improving the hematological, biochemical, histopathological, and molecular alterations.

Effect of oridonin on oxylipins in the livers of mice with acute liver injury induced by D-galactosamine and lipopolysaccharide.

BACKGROUND AND PURPOSE: Oridonin (Ori) has been shown to protect against acute liver injury (ALI) induced by D-galactosamine (D-GalN) and lipopolysaccharide (LPS). Oxylipins are oxidation products of polyunsaturated fatty acids (PUFAs) and are key proinflammatory mediators. This study aimed to investigate the changes in oxylipins in the livers of mice with D-GalN/LPS-induced ALI and the effects of Ori on these changes. RESULTS: 54 oxylipins in liver tissues were identified and qualitatively and quantitatively analyzed by ultra-performance liquid chromatography-electrospray ionization triple quadrupole mass spectrometry (UPLC-QTRAP/MS/MS). The levels of 12-HETE, 12-HEPE, 14(S)-HDHA, PGE2, dihomo-γ-linolenic acid and 13-HOTrE in the liver were significantly increased in the D-GalN/LPS-induced ALI group compared with the control group, and the levels of EPA and 7-HDHA were significantly decreased. However, pretreatment with Ori dramatically decreased the levels of 12-HETE, 12-HEPE, 14(S)-HDHA, PGE2 and 13-HOTrE compared with those of the ALI group and induced 7-HDHA and 15-oxoETE. Moreover, Ori reduced the protein levels of COX-1, COX-2, ALOX5, ALOX12 and ALOX15 induced by D-GalN/LPS, indicating that Ori altered oxylipins through the COX and LOX pathways. CONCLUSIONS: These results suggest that the protective effect of Ori on ALI is partly mediated by affecting the oxylipin pathway.

Genetic diversity and structure of Oncomelania hupensis hupensis in two eco-epidemiological settings as revealed by the mitochondrial COX1 gene sequences.

BACKGROUND: Oncomelania hupensis hupensis is the only intermediate host of Schistosoma japonicum, the causative agent of schistosomiasis in China and is therefore of significant medical and veterinary health importance. Although tremendous progress has been achieved, there remains an understudied area of approximately 2.06 billion m2 of potential snail habitats. This area could be further increased by annual flooding. Therefore, an understanding of population genetics of snails in these areas may be useful for future monitoring and control activities. METHODS AND RESULTS: We sampled snails from Hexian (HX), Zongyang (ZY) and Shitai (ST) in Anhui (schistosomiasis transmission control), and from Hengtang (HT), Taicang (TC), Dongsan (DS) and Xisan (XS) in Jiangsu (schistosomiasis transmission interrupted), downstream of Anhui. ST, DS and XS are classified as hilly and mountainous areas, and HX, ZY, TC and HT as lake and marshland areas. The mitochondrial cytochrome c oxidase subunit I gene were sequenced. Out of 115 snails analyzed, 29 haplotypes were identified. We observed 56 (8.72%) polymorphic sites consisting of 51 transitions, four transversions and one multiple mutational change. The overall haplotype and nucleotide diversity were 0.899 and 0.01569, respectively. Snail populations in Anhui had higher genetic diversity than in Jiangsu. 73.32% of total variation was distributed among sites and 26.68% within sites. Snails were significantly separated according to eco-epidemiological settings in both network and phylogenetic analyses. CONCLUSION: Our results could provide important guidance towards assessing coevolutionary interactions of snails with S. japonicum, as well as for future molluscan host monitoring and control activities.

High-throughput sequencing for species authentication and contamination detection of 63 cell lines.

Cell lines are widely used in research and for diagnostic tests and are often shared between laboratories. Lack of cell line authentication can result in the use of contaminated or misidentified cell lines, potentially affecting the results from research and diagnostic activities. Cell line authentication and contamination detection based on metagenomic high-throughput sequencing (HTS) was tested on DNA and RNA from 63 cell lines available at the Canadian Food Inspection Agency's National Centre for Foreign Animal Disease. Through sequence comparison of the cytochrome c oxidase subunit 1 (COX1) gene, the species identity of 53 cell lines was confirmed, and eight cell lines were found to show a greater pairwise nucleotide identity in the COX1 sequence of a different species within the same expected genus. Two cell lines, LFBK-αvß6 and SCP-HS, were determined to be composed of cells from a different species and genus. Mycoplasma contamination was not detected in any cell lines. However, several expected and unexpected viral sequences were detected, including part of the classical swine fever virus genome in the IB-RS-2 Clone D10 cell line. Metagenomics-based HTS is a useful laboratory QA tool for cell line authentication and contamination detection that should be conducted regularly.

Malvidin Protects against and Repairs Peptic Ulcers in Mice by Alleviating Oxidative Stress and Inflammation.

Peptic ulcer episodes cause damage to the stomach and intestine, with inflammatory cell infiltration and oxidative stress as the main players. In this study, we investigated the potential of anthocyanidin malvidin for preventive and curative peptic ulcer treatment. The anthocyanidin effects were examined in gastric ulcer mouse models induced by ethanol, non-steroidal anti-inflammatory drugs (NSAIDs), ischemia-reperfusion (IR), acetic acid and duodenal ulcer induced by polypharmacy. Expression levels of oxidative and inflammatory genes were measured to investigate the mechanism of anthocyanin activity. At a dose of 5 mg·kg-1, Malvidin prevented gastric ulcer induction by ethanol, NSAID and repaired the tissue after 6 days of IR. Moreover, the anthocyanidin accelerated the healing of acetic acid-induced ulcer, increased the gene expression of EGF and COX-1, and downregulated MMP-9. Anthocyanin treatment mitigated the effect of polypharmacy on inflammation and oxidative stress observed in the intestine. Additionally, the compound downregulated cytokine expression and TLR4 and upregulated HMOX-1 and IL-10, exhibiting protective activity in the mouse gut. Malvidin thus prevented gastric and duodenal ulcers due to prominent anti-inflammatory and antioxidative effects on the gastrointestinal tract that were related to gene expression modulation and an increase in endogenous defense mechanisms.

Automatic barcode gap discovery reveals diverse clades of Rhipicephalus spp. and Haemaphysalis spp. ticks from small mammals in 'Asir, Saudi Arabia.

BACKGROUND: The ixodid tick genera Rhipicephalus and Haemaphysalis contain several species of medical and/or veterinary importance, but their diversity in some regions of the world remains under-explored. For instance, very few modern studies have been performed on the taxonomy of these genera on the Arabian Peninsula. METHODS: In this study, we trapped small mammals in the 'Asir Mountains of south-western Saudi Arabia and collected tick specimens for morphological examination and molecular barcoding, targeting three mitochondrial loci: cox1, 16S rRNA and 12S rRNA. RESULTS: We obtained a total of 733 ticks (608 Haemaphysalis spp. and 125 Rhipicephalus spp.) from 75 small mammal hosts belonging to six species. All tick specimens were immature except for nine adults recovered from a hedgehog (Paraechinus aethiopicus). Morphologically, the Rhipicephalus ticks resembled R. camicasi, but the Haemaphysalis ticks showed differences in palp morphology compared with species previously described from Saudi Arabia. Phylogenetic analysis and automatic barcode gap discovery identified a novel clade of Rhipicephalus sp. representing most of the nymphs. This was most closely related to R. leporis, R. guilhoni and R. linnaei. The adult ticks and a small proportion of nymphs clustered with R. camicasi sequences from a previous study. Finally, the Haemaphysalis nymphs formed two distinct clades that were clearly separated from all reference sequences but closest to some African species. CONCLUSIONS: This apparent high level of tick diversity observed in a single study site of only ~ 170 km2, on a relatively small number of hosts, highlights the potential for the discovery of new tick species on the Arabian Peninsula.

Prostaglandin E2 increases the expression of cyclooxygenase-2 in cultured rat microglia.

Prostaglandin E2 (PGE2) plays pivotal roles in controlling microglial activation with the EP2 receptor, a PGE2 receptor subtype. Activated microglia are often reported to increase cyclooxygenase (COX)-2 expression, followed by PGE2 production, but it is unclear whether extracellular PGE2 is involved in microglial PGE2 synthesis. In the present study, we report that PGE2 increases COX-2 protein in microglia. In a culture system, PGE2 at 10-6 M for 3 h increased COX-2 and microsomal PGE synthase (mPGES)-1 mRNA levels, and reduced mPGES-2, but did not affect COX-1 or cytosolic PGE synthase (cPGES) in microglia. PGE2 at 10-6 M for 3 h also increased the COX-2 protein level, but did not affect COX-1, mPGES-1, mPGES-2, or cPGES. An EP2 agonist, ONO-AE1-259-01, also increased COX-2 and mPGES-1 mRNA levels, and reduced mPGES-2, but did not affect COX-1 or cPGES, whereas an EP1 agonist, ONO-DI-004, an EP3 agonist, ONO-AE-248, and an EP4 agonist, ONO-AE1-329, had no effect. Similar to PGE2, ONO-AE1-259-01 increased the COX-2 protein level, but did not affect COX-1, mPGES-1, mPGES-2, or cPGES. In addition, the effects of PGE2 were inhibited by an EP2 antagonist, PF-04418948, but not by an EP1 antagonist, ONO-8713, an EP3 antagonist, ONO-AE3-240, or an EP4 antagonist, ONO-AE3-208, at 10-6 M. On the other hand, lipopolysaccharide (LPS) increased PGE2 production, but the LPS-induced PGE2 production was not affected by ONO-8713, PF-04418948, ONO-AE3-240, or ONO-AE3-208. These results indicate that PGE2 increases COX-2 protein in microglia through the EP2 receptor supporting the idea that extracellular PGE2 has a triggering aspect for microglial activation.

In situ imaging reveals disparity between prostaglandin localization and abundance of prostaglandin synthases.

Prostaglandins are important lipids involved in mediating many physiological processes, such as allergic responses, inflammation, and pregnancy. However, technical limitations of in-situ prostaglandin detection in tissue have led researchers to infer prostaglandin tissue distributions from localization of regulatory synthases, such as COX1 and COX2. Herein, we apply a novel mass spectrometry imaging method for direct in situ tissue localization of prostaglandins, and combine it with techniques for protein expression and RNA localization. We report that prostaglandin D2, its precursors, and downstream synthases co-localize with the highest expression of COX1, and not COX2. Further, we study tissue with a conditional deletion of transformation-related protein 53 where pregnancy success is low and confirm that PG levels are altered, although localization is conserved. Our studies reveal that the abundance of COX and prostaglandin D2 synthases in cellular regions does not mirror the regional abundance of prostaglandins. Thus, we deduce that prostaglandins tissue localization and abundance may not be inferred by COX or prostaglandin synthases in uterine tissue, and must be resolved by an in situ prostaglandin imaging.

Molecular identification of Sarcocystis species in diaphragm muscle tissue of European mouflon (Ovis gmelini musimon) from Austria.

Previous morphological studies suggested that mouflon may have sarcocysts similar to those of sheep. However, to date, no molecular-based studies of the species of Sarcocystis infecting mouflon have been done. The present study identified Sarcocystis species in diaphragm muscle samples from 20 European mouflon (Ovis gmelini musimon). Molecular identification using the cox1 sequence analysis was performed on sarcocysts excised from muscle tissue and on DNA from digested muscle samples. Both frequency and intensity of infection in mouflon were high with 19 of 20 animals testing Sarcocystis positive and > 50 cysts per gram of tissue recovered from 10 of the 19 Sarcocystis positive animals. Molecular analysis revealed dominant Sarcocystis tenella (18/19 animals) and Sarcocystis arieticanis (1/19 animals), whose known intermediate hosts are sheep. In addition, Sarcocystis capracanis, which is known to form sarcocysts in goats, was detected in two animals. The results of this study demonstrated the digestion method to be superior over the direct isolation of sarcocysts for the molecular identification of Sarcocystis species in a certain host. Future research of Sarcocystis diversity in wild ovine and caprine species is needed.