Isolation, Structural Characterization, and Biological Activity of the Two Acidic Polysaccharides from the Fruits of the Elaeagnus angustifolia Linnaeus.

Elaeagnus angustifolia Linnaeus is a medicinal plant and its fruit has pharmacological activity such as antiinflammatory, antiedema, antinociceptive, and muscle relaxant functions, etc. Two acidic homogeneous polysaccharides (EAP-H-a1 and EAP-H-a2) were isolated from the fruits of Elaeagnus angustifolia L. through DEAE-52 and Sephadex G-75 column chromatography, and the physicochemical, structural properties, and biological activities of the polysaccharides were investigated. Both EAP-H-a1 and EAP-H-a2 were composed of Rha, Ara, Xyl, Glc, and Gal with the molar ratios of 13.7:20.5:23.3:8.8:33.4 and 24.8:19.7:8.2:8.4:38.6, respectively, and with the molecular weights of 705.796 kDa and 439.852 kDa, respectively. The results obtained from Fourier transform infrared spectroscopy (FTIR) confirmed the polysaccharide nature of the isolated substances. Congo red assay confirmed the existence of a triple-helix structure. Scanning electron microscopy (SEM) and X-ray diffraction (XRD) analysis revealed that EAP-H-a1 and EAP-H-a2 had irregular fibrous, filament-like surfaces; and both had crystalline and amorphous structures. Bioactivity analysis showed that the crude polysaccharide, EAP-H-a1, and EAP-H-a2 had clear DPPH and ABTS free radical scavenging activity, and could promote the secretion of NO and the phagocytic activities of RAW 264.7 and THP cells, which showed clear antioxidant and immuno-regulatory activity. These results indicated that Elaeagnus angustifolia L fruit acidic polysaccharides may have potential value in the pharmaceutical and functional food industries.

A network pharmacology strategy combined with in vitro experiments to investigate the potential anti-inflammatory mechanism of Prunus cerasifera Ehrhart.

This study aimed to investigate the anti-inflammatory activity of Prunus cerasifera Ehrhart (EHP). LC-MS/MS, network pharmacology, enzyme-linked immunosorbent assay (ELISA), and Western blot analysis methods were used to investigate the chemical composition and the anti-inflammatory mechanism of EHP. The LC-MS/MS results showed that flavonoids and phenolic acids were the major compounds in EHP. The network pharmacology analysis results indicated that EHP was related to TNF, inflammatory cytokine, and MAPK signaling pathway. ELISA and Western blot results showed that EHP impeded the increase in inflammatory factors, inducible nitric oxide synthase (iNOS), cyclooxygenase 2 (COX-2), nuclear transcription factors κB (p65), MAPK pathway, pyrolytic relevant proteins nod-like receptor family pyrin domain-containing 3 (NLRP3), and interleukin-1ß (IL-1ß) induced by lipopolysaccharide (LPS) and activated the nuclear factor erythroid 2-related factor 2 (Nrf2)/hemeoxygenase-1 (HO-1) pathway. Therefore, this research highlighted the potential application of P. cerasifera in the development of anti-inflammatory foods that prevented inflammatory diseases. PRACTICAL APPLICATIONS: In recent years, many synthetic drugs with anti-inflammatory effect have the disadvantages of high price and side effects. Thus, the development of anti-inflammatory drugs from natural resources has its application value. In this study, LPS-stimulated RAW264.7 cells were used to establish inflammatory model to verify the anti-inflammatory effect of Prunus cerasifera (EHP). The results showed that P. cerasifera possessed anti-inflammatory activity through inhibiting pro-inflammatory cytokines secretion, NF-κB, MAPK pathway, and NLRP3 inflammasome activation. Therefore, P. cerasifera has the potential to develop into functional food to prevent the progress of various inflammatory-related diseases.

Chemical component and in vitro protective effects of Matricaria chamomilla (L.) against lipopolysaccharide insult.

ETHNOPHARMACOLOGICAL RELEVANCE: Chamomile (Matricaria chamomilla L.) is a popular herbal tea for the treatment of hepatitis and cholecystitis in traditional Uygur medicines. AIM OF THE STUDY: To investigate the anti-inflammatory activity and chemical composition of M. chamomilla, and clarify its molecular mechanism. MATERIALS AND METHODS: M. chamomilla was extracted with 75% ethanol and then extracted with different solvents to obtain five fractions, namely petroleum ether fraction (EOPE), dichloromethane fraction (EOD), ethyl acetate fraction (EOEA), n-butanol fraction (EOB), and water fraction (EOW). Cytotoxicity and the effect on the nitric oxide (NO) production of RAW264.7 cells induced by LPS of the five fractions were screened, and the most active one (EOD) was selected for further investigations. The components of EOD were identified by LC-MS/MS analysis in combination with comparison of retention time and UV absorption with authentic compounds by HPLC. In addition, five most abundant compounds of EOD were isolation by column chromatography and semi-preparative HPLC and their structures were further confirmed by HRMS and NMR data analysis and comparison with data in literatures. Then the underlying anti-inflammatory mechanism of EOD were predicted through Network pharmacology using the identified compounds from EOD, and further verified by Western Blot and ELISA experiments. RESULTS: EOD showed the most significant inhibition ratio against NO in RAW264.7 cells without toxicity among the tested five fractions. Thirty-seven compounds including flavonoid-O-glycoside, flavonoid aglycone, methylated flavonoid aglycone, phenolic acid, coumarin, sesquiterpene, and triterpene were identified from EOD by LC-MS/MS and comparison with authentic compounds. The five most abundant compounds in EOD were isolated and determined to be axillarin (26), tricin (30), chrysoeriol (31), centaureidin (33) and chrysosplenetin (35). IL-6, NF-κB, ERK1 and ERK2 cascade, TNF were the most important anti-inflammatory targets of EOD predicted by Network pharmacology. Western Blot and ELISA experiments revealed that EOD significantly decreased the protein expression levels of inflammatory factors (PGE2, MCP-1, IL-6, TNF-α), iNOS, COX-2, NF-κB (p-P65 and p-IκBα), MAPKs (p-p38, p-ERK and p-JNK), and increased the protein expression levels of Nrf2, HO-1 and CYP2E1. In addition, EOD blocked the p65 protein into the nucleus and promoted the nuclear translocation of Nrf2 in RAW264.7 cells induced by LPS. CONCLUSION: M. chamomilla exerted anti-inflammatory effect via NF-κB, MAPK and Nrf2/HO-1 pathways. It could be further applied as a safe anti-inflammatory agent from natural source.

Anti-inflammatory activity of Anchusa italica Retz. in LPS-stimulated RAW264.7 cells mediated by the Nrf2/HO-1, MAPK and NF-κB signaling pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Anchusa italica Retz. (Boraginaceae) is an important medicinal plant for the treatment of meningitis and pneumonia in traditional Uygur medicines. AIM OF THE STUDY: To clarify the anti-inflammatory activity of A. italica, to reveal its molecular mechanisms, and to discover the anti-inflammatory active ingredients. MATERIALS AND METHODS: Dried and crushed aerial parts of A. italica were extracted with 75% ethanol to yield crude extract (AICE) and AICE was fractionated to obtain petroleum ether extract (AIPE), dichloromethane extract (AIDE), ethyl acetate extract (AIEE), n-butanol extract (AIBE) and residues (AIW). By measuring the effects of AIPE, AIDE, AIEE, AIBE and AIW on cell viability and nitric oxide (NO) in Lipopolysaccharide (LPS) stimulated RAW264.7 cell lines, AIDE with the lowest cytotoxicity and NO contents was finally selected for further chemical and anti-inflammatory investigations. LC-MS/MS experiment was applied to analyze the chemical composition of AIDE. MTT and Griess methods were used to detect the cell viability and to quantify the nitrite levels in culture supernatants, respectively. Prostaglandin E2 (PGE2), interleukin-6 (IL-6), interleukin-1ß (IL-1ß) and tumor necrosis factor α (TNF-α) production was examined by ELISA assays. Real-time quantitative PCR was used to detect the expression of hemeoxygenase-1 (HO-1), Nrf2-mediated quinone oxidoreductase 1 (NQO-1), glutathione S-transferase A 1 (GSTA1) and glutathione S-transferase M 1 (GSTM1) mRNA. Western blot analysis was employed to examine the protein expression and enzymatic activities. RESULTS: In preliminary anti-inflammatory screening, AIDE showed the lowest cytotoxicity and the most significant inhibitory effect on the production of NO (the inhibitory is 89%) induced by LPS among the tested five extracts. Thirty-three compounds including twenty-five triterpenoids were identified by LC-MS/MS analysis. AIDE could inhibit LPS-induced the over-expression of NO, IL-6, PGE2, IL-1ß and TNF-α and down-regulate the levels of extracellular signal-regulated protein kinase (ERK), c-Jun N-terminal kinase (JNK), P38-MAPK (P38) and nuclear transcription factors κB-P65 (P65) phosphorylation. It promoted the mRNA expression level of HO-1, NQO-1, GSTA1 and GSTM1 and the protein expression level of nuclear factor erythroid 2-related factor 2 (Nrf2) and HO-1. After the treatment of AIDE, P65 nuclear translocation was inhibited and Nrf2 nuclear translocation was increased. In addition, the protein expression of pyrolytic relevant protein nod-like receptor family pyrin domain-containing 3 (NLRP3) and IL-1ß were decreased after the AIDE treatment. CONCLUSIONS: Anchusa italica Retz. exerted its anti-inflammatory activity by inhibiting the mitogen-activated protein kinase (MAPK), nuclear transcription factors κB (NF-κB) and pyrolytic relevant proteins, down-regulating inflammatory factor levels, and activating the Nrf2/HO-1 pathway. Triterpenoids might be its major active anti-inflammatory ingredients.

To explore the therapeutic effect of Kangfuxiaomi shuan II on cervicitis in rats.

PURPOSE: The rat cervicitis model was established with 20% phenol glue to explore the therapeutic effect of Kangfuxiaomi shuan II on rat cervicitis and its mechanism. METHODS: After modeling, the rats were treated with Shuangzuotai suppository (37.84 mg/kg), Kangfuxiaoyan shuan (205.6 mg/kg) and Kangfuxiaomi shuan II (40, 80, 160 mg/kg). The histopathological changes and injury degree of cervix in rats were evaluated by vulvar inflammation score and organ index. The therapeutic effect of Kangfuxiaomi shuan II on cervicitis was evaluated by detecting the levels of copper-protein (CP), C-reactive protein (CRP), Rat interleukin 6 (IL-6), superoxide dismutase (SOD) and malondialdehyde (MDA) in serum and epidermal growth factor (EGF), cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) in cervical tissue. RESULTS: Compared with the model group, the vulvar inflammation score and cervical index of rats in other groups decreased significantly (P<0.01). Kangfuxiaomi shuan II could significantly reduce the levels of CP, CRP, and MDA in serum of rats with cervicitis, and significantly increase the activity of SOD in serum of rats with cervicitis (P<0.01). The levels of EGF and iNOS in cervical tissue of rats also increased in different degrees, while the level of COX-2 decreased significantly (P<0.01), which significantly improved the pathological degree of vulvar inflammation in rats with cervicitis. CONCLUSIONS: Kangfuxiaomi shuan II has a certain therapeutic effect on cervicitis in rats, and its mechanism may be related to the regulation of inflammatory cytokine network and immunity.

Network Pharmacology Reveals That Resveratrol Can Alleviate COVID-19-Related Hyperinflammation.

Hyperinflammation is related to the development of COVID-19. Resveratrol is considered an anti-inflammatory and antiviral agent. Herein, we used a network pharmacological approach and bioinformatic gene analysis to explore the pharmacological mechanism of Resveratrol in COVID-19 therapy. Potential targets of Resveratrol were obtained from public databases. SARS-CoV-2 differentially expressed genes (DEGs) were screened out via bioinformatic analysis Gene Expression Omnibus (GEO) datasets GSE147507, followed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis; then, protein-protein interaction network was constructed. The common targets, GO terms, and KEGG pathways of Resveratrol targets and SARS-CoV-2 DEGs were confirmed. KEGG Mapper queried the location of common targets in the key pathways. A notable overlap of the GO terms and KEGG pathways between Resveratrol targets and SARS-CoV-2 DEGs was revealed. The shared targets between Resveratrol targets and SARS-CoV-2 mainly involved the IL-17 signaling pathway, NF-kappa B signaling pathway, and TNF signaling pathway. Our study uncovered that Resveratrol is a promising therapeutic candidate for COVID-19 and we also revealed the probable key targets and pathways involved. Ultimately, we bring forward new insights and encourage more studies on Resveratol to benefit COVID-19 patients.

Chemical Composition of the Hazelnut Kernel (Corylus avellana L.) and Its Anti-inflammatory, Antimicrobial, and Antioxidant Activities.

The chemical composition of hazelnut kernels (Corylus avellana L.) and their COX-2 inhibitory, antimicrobial, and 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical-scavenging activities were investigated. Six previously undescribed indoleacetic acid glycosides, hazelnutins A-F (1-6), and five known compounds (7-11) were isolated from the hazelnut kernels. The structures of compounds 1-6 were successfully identified by high-resolution-electrospray ionization-mass spectrometry and NMR data, and their absolute configurations were established by electron-capture detector spectroscopy analyses in corporation with quantum chemical calculations. Furthermore, the absolute configurations of compounds 7 and 8 were unambiguously confirmed for the first time. Compounds 8-11 were discovered in hazelnut kernels for the first time. Compounds 1-5 inhibited COX-2 expression with inhibition rates ranging from 36.10 to 64.08%. Compounds 3, 4, and 8 could inhibit the proliferation of Candida albicans. Compound 11 exhibited potent antioxidant activity against ABTS and DPPH with IC50 values of 11.22 and 13.21 µmol/L, respectively. Compounds 8 and 10 exhibited moderate antioxidant activity against ABTS.

Lignanamides from the roots of Limonium gmelinii (Willd.) Kuntze and their anti-diabetic, cytotoxic and anti-inflammatory activities.

Nine undescribed lignanamides, limoniumins A-I, together with ten known lignanamides and two known phenolics were isolated from ethyl acetate extract of the roots of Limonium gmelinii (Plumbaginaceae). Their structures were determined by spectroscopic analysis including 1D and 2D NMR and HRESIMS experiments. Limoniumin A is the first hybrid lignanamide of phenylpropanoid and coumarin. All tested lignanamides showed significant inhibitory activity against α-glucosidase stronger than positive control and remarkable inhibitory effect to PTP1B with IC50 values less than 10 µM. In addition, some lignanamides exhibited moderate cytotoxic activity against HeLa and MCF-7 cells and anti-inflammatory activity against COX-2 in a dose-dependent way.

To explore the therapeutic effect of Kangfuxiaomi shuan II on cervicitis in rats

ABSTRACT Purpose: The rat cervicitis model was established with 20% phenol glue to explore the therapeutic effect of Kangfuxiaomi shuan II on rat cervicitis and its mechanism. Methods: After modeling, the rats were treated with Shuangzuotai suppository (37.84 mg/kg), Kangfuxiaoyan shuan (205.6 mg/kg) and Kangfuxiaomi shuan II (40, 80, 160 mg/kg). The histopathological changes and injury degree of cervix in rats were evaluated by vulvar inflammation score and organ index. The therapeutic effect of Kangfuxiaomi shuan II on cervicitis was evaluated by detecting the levels of copper-protein (CP), C-reactive protein (CRP), Rat interleukin 6 (IL-6), superoxide dismutase (SOD) and malondialdehyde (MDA) in serum and epidermal growth factor (EGF), cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) in cervical tissue. Results: Compared with the model group, the vulvar inflammation score and cervical index of rats in other groups decreased significantly (P<0.01). Kangfuxiaomi shuan II could significantly reduce the levels of CP, CRP, and MDA in serum of rats with cervicitis, and significantly increase the activity of SOD in serum of rats with cervicitis (P<0.01). The levels of EGF and iNOS in cervical tissue of rats also increased in different degrees, while the level of COX-2 decreased significantly (P<0.01), which significantly improved the pathological degree of vulvar inflammation in rats with cervicitis. Conclusions: Kangfuxiaomi shuan II has a certain therapeutic effect on cervicitis in rats, and its mechanism may be related to the regulation of inflammatory cytokine network and immunity.

Wasp Venom Possesses Potential Therapeutic Effect in Experimental Models of Rheumatoid Arthritis.

Rheumatoid arthritis (RA) is an autoimmune disease. Wasp venom (WV), which is considered as a traditional folk medicine in Jingpo nationality in Yunnan, China, relieves rheumatoid arthritis. The current study aimed to investigate the effect of wasp venom ameliorating rheumatoid arthritis symptoms in experimental rats. We established a model of type II collagen- (CII-) induced arthritis (CIA) in SD rats and examined the inhibition of inflammation and autoimmune response. The antiarthritic effects of WV were evaluated through the paw swelling, and histopathological score and histopathology changes of the affected paw were assessed. The anti-inflammation effects were assayed by the level of IL-6, TNF-α, IL-1ß, and the number of inflammatory cells in peripheral blood. The alteration of the T cell subset ratio in the spleen of rats was detected by flow cytometry, and at the same time, the viscera index and immune serum globulin levels were evaluated. The results suggested that various doses of WV (0.125, 0.25, and 0.5 mg/kg) significantly alleviated paw swelling and arthritis score in CIA rats with the untreated control (P < 0.05). WV (0.25 and 0.5 mg/kg) relieved synovial tissue lesions of ankle joints and histopathology scores of synoviocyte hyperplasia and inflammatory cell infiltration with vehicle group (P < 0.05). Regarding immunological regulation, 0.5 mg/kg WV lowered the immune serum globulin levels (P < 0.05), and we further found that WV (0.5 mg/kg) suppressed the immune response of Th cells, while enhancing the functions of Tc cells and Treg cells in spleen cells markedly (P < 0.05). The immunosuppressive action of WV displayed was analogous to its inhibitory effect on IL-1ß, TNF-α, IL-8, IL-6, COX-2, and PGE2 levels in rat serum. In conclusion, these findings demonstrated that WV exhibited antiarthritic activity, which might be associated with their inhibitory effects on immunoregulation and anti-inflammatory action.

Homologous Gold Nanoparticles and Nanoclusters Composites with Enhanced Surface Raman Scattering and Metal Fluorescence for Cancer Imaging.

A large number of deaths from cancer can be attributed to the lack of effective early-stage diagnostic techniques. Thus, accurate and effective early diagnosis is a major research goal worldwide. With the unique phenomenon of localized surface plasmon resonance (LSPR), plasmonic nanomaterials have attracted considerable attention for applications in surface-enhanced Raman scattering (SERS) and metal-enhanced fluorescence (MEF). Both SERS and MEF are ultra-sensitive methods for the detection and identification of early tumor at molecular level. To combine the merits of the fast and accurate imaging of MEF and the stable and clear imaging of SERS, we propose a novel dual functional imaging nanoprobe based on gold nanoparticles and gold nanocluster composites (denoted AuNPC-RGD). The gold nanoparticles are used as LSPR substrates to realized enhancement of Raman or fluorescence signal, while the gold nanoclusters serve as a fluorophore for MEF imaging, and exhibit better biocompatibility and stability. Furthermore, target molecule of cyclic Arg-Gly-Asp (cRGD) is incorporated into the composite to improve delivery efficiency, selectivity and imaging accuracy. These integrated properties endow AuNPC-RGD composites with outstanding biocompatibility and excellent imaging abilities, which could be used to achieve accurate and effective diagnosis for early cancer.

A novel tick-borne phlebovirus, closely related to severe fever with thrombocytopenia syndrome virus and Heartland virus, is a potential pathogen.

Tick-borne viral diseases have attracted much attention in recent years because of their increasing incidence and threat to human health. Severe fever with thrombocytopenia syndrome phlebovirus (SFTSV) and Heartland virus (HRTV) were recently identified as tick-borne phleboviruses (TBPVs) in Asia and the United States, respectively, and are associated with severe human diseases with similar clinical manifestations. In this study, we report the first identification and isolation of a novel TBPV named Guertu virus (GTV) from Dermacentor nuttalli ticks in Xinjiang Province, China, where TBPVs had not been previously discovered. Genome sequence and phylogenetic analyses showed that GTV is closely related to SFTSV and HRTV and was classified as a member of the genus Phlebovirus, family Phenuiviridae, order Bunyavirales. In vitro and in vivo investigations of the properties of GTV demonstrated that it was able to infect animal and human cell lines and can suppress type I interferon signaling, similar to SFTSV, that GTV nucleoprotein (NP) can rescue SFTSV replication by replacing SFTSV NP, and that GTV infection can cause pathological lesions in mice. Moreover, a serological survey identified antibodies against GTV from serum samples of individuals living in Guertu County, three of which contained neutralizing antibodies, suggesting that GTV can infect humans. Our findings suggested that this virus is a potential pathogen that poses a threat to animals and humans. Further studies and surveillance of GTV are recommended to be carried out in Xinjiang Province as well as in other locations.

Role of miR-383 and miR-146b in different propensities to obesity in male mice.

The study was designed to investigate the possible mechanisms of hepatic microRNAs (miRs) in regulating local thyroid hormone (TH) action and ultimately different propensities to high-fat diet (HFD)-induced obesity. When obesity-prone (OP) and obesity-resistant (OR) mice were fed HFD for 7 weeks, OP mice showed apparent hepatic steatosis, with significantly higher body weight and lower hepatic TH receptor b (TRb) expression and type 1 deiodinase (DIO1) activity than OR mice. Next-generation sequencing technology revealed that 13 miRs in liver were dysregulated between the two phenotypes, of which 8 miRs were predicted to target on Dio1 or TRb When mice were fed for 17 weeks, OR mice had mild hepatic steatosis and increased Dio1 and TRb expression than OP mice, with downregulation of T3 target genes (including Srebp1c, Acc1, Scd1 and Fasn) and upregulation of Cpt1α, Atp5c1, Cox7c and Cyp7a1 A stem-loop qRT-PCR analysis confirmed that the levels of miR-383, miR-34a and miR-146b were inversely correlated with those of DIO1 or TRb. Down-regulated expression of miR-383 or miR-146b by miR-383 inhibitor (anti-miR-383) or miR-146b inhibitor (anti-miR-146b) in free fatty acid-treated primary mouse hepatocytes led to increased DIO1 and TRb expressions, respectively, and subsequently decreased cellular lipid accumulation, while miR-34a inhibitor (anti-miR-34a) transfection had on effects on TRb expression. Luciferase reporter assay illustrated that miR-146b could directly target TRb 3'untranslated region (3'UTR). These findings suggested that miR-383 and miR-146b might play critical roles in different propensities to diet-induced obesity via targeting on Dio1 and TRb, respectively.

The Expression of microRNA-223 and FAM5C in Cerebral Infarction Patients with Diabetes Mellitus.

The aim of the study was to investigate the expression of miR-223 and FAM5C in the peripheral blood mononuclear cells (PBMCs) of cerebral infarction patients with or without diabetes. Sixteen cases with diabetes mellitus (DM), 14 cases with cerebral infarction (CI), 12 cases with cerebral infarction and diabetes mellitus (CIDM), and 18 healthy subjects were included in this study. Real-time PCR was used to quantify mRNA expression. Western blot was used to detect FAM5C protein level. Recombinant plasmids expressing miR-223-3p and 3' UTR of FAM5C were constructed. Dual-luciferase reporter system was used to analyze the binding of miR-223-3p to FAM5C 3' UTR. FAM5C mRNA and protein level were significantly higher in the PBMCs of CIDM patients compared with healthy controls (P < 0.05). miR-223-3p expression in PBMCs was significantly lower in DM patients than in healthy controls (P < 0.05). The expression of miR-223-3p was negatively correlated with FAM5C mRNA in all patients and healthy controls. Co-transfection of miR-223-3p plasmid with FAM5C 3'UTR dual-luciferase plasmid significantly inhibited the luciferase activity (P < 0.01). FAM5C, but not miR-223, is a risk factor for CI in type 2 DM patients.

Role of thyroid hormone homeostasis in obesity-prone and obesity-resistant mice fed a high-fat diet.

BACKGROUND: The exact mechanism for different propensities to obesity when consuming a high-fat diet (HFD) is largely unknown. Thyroid hormone (TH) is an important modulator of energy homeostasis and body weight. OBJECTIVE: The present study aimed to find the potential mechanisms of TH in the development of obesity-prone (OP) and obesity-resistant (OR) mice after short-term and long-term HFD feeding. METHODS: C57Bl/6 male mice were randomly divided into two groups: a low-fat diet (LFD) group and an HFD group. In the 7th week, HFD-fed mice were classified as OP or OR according to upper and lower tertiles of body weight. Half of the mice were sacrificed at this time point and the remaining mice were kept on feeding and sacrificed in the 27th week. Indirect calorimetry was performed. At harvest, serum was used for ELISA assays and oxidative stress biomarkers determination. Tissues were dissected for deiodinases activity and relative mRNA expression determination, as well as antioxidant capacity evaluation. RESULTS: In the 7th week, OP mice showed a significant body weight gain, decreased energy expenditure (EE), normal circulating TH levels, and activated HPT axis, whereas OR mice had normal body weight and maintained T(3) levels only through enhancing hepatic D1 activity. In the 27th week, OR mice gained more body weight than LFD mice accompanied by an activation of HPT axis and decreased hepatic deiodination. Genes involved in TH production were down-regulated in OP mice and up-regulated in OR mice. Changes in deiodinases activity and thyroid function were related with redox status in specific tissues. Furthermore, OP mice had more serious hepatic steatosis than OR mice, with up-regulation of T(3) target genes (e.g. Srebp1c, Acc1, Fasn) involved in lipid synthesis and down-regulation of Pgc1α, Cyp7a1 and Cpt1α. CONCLUSIONS: HPT axis function and deiodinases activity might be involved in different propensities to obesity and the ability of OR mice to resist obesity was limited.

Fine epitope mapping of the central immunodominant region of nucleoprotein from Crimean-Congo hemorrhagic fever virus (CCHFV).

Crimean-Congo hemorrhagic fever (CCHF), a severe viral disease known to have occurred in over 30 countries and distinct regions, is caused by the tick-borne CCHF virus (CCHFV). Nucleocapsid protein (NP), which is encoded by the S gene, is the primary antigen detectable in infected cells. The goal of the present study was to map the minimal motifs of B-cell epitopes (BCEs) on NP. Five precise BCEs (E1, 247FDEAKK252; E2a, 254VEAL257; E2b, 258NGYLNKH264; E3, 267EVDKA271; and E4, 274DSMITN279) identified through the use of rabbit antiserum, and one BCE (E5, 258NGYL261) recognized using a mouse monoclonal antibody, were confirmed to be within the central region of NP and were partially represented among the predicted epitopes. Notably, the five BCEs identified using the rabbit sera were able to react with positive serum mixtures from five sheep which had been infected naturally with CCHFV. The multiple sequence alignment (MSA) revealed high conservation of the identified BCEs among ten CCHFV strains from different areas. Interestingly, the identified BCEs with only one residue variation can apparently be recognized by the positive sera of sheep naturally infected with CCHFV. Computer-generated three-dimensional structural models indicated that all the antigenic motifs are located on the surface of the NP stalk domain. This report represents the first identification and mapping of the minimal BCEs of CCHFV-NP along with an analysis of their primary and structural properties. Our identification of the minimal linear BCEs of CCHFV-NP may provide fundamental data for developing rapid diagnostic reagents and illuminating the pathogenic mechanism of CCHFV.

Detection of platelet microRNA expression in patients with diabetes mellitus with or without ischemic stroke.

AIMS: The objective of this study was to investigate the role of plasma and platelet microRNAs in the occurrence of ischemic stroke in patients with diabetes mellitus. METHODS: miR-223, miR-146a, miR-495, and miR-107 expression in the plasma and platelets, blood glucose concentration, and platelet activation rate were measured in patients with diabetes mellitus and ischemic stroke, diabetes mellitus only, ischemic stroke only, and healthy controls. Platelet activity was measured by flow cytometric measurement of P-selectin expression, while miRNA was measured by real-time PCR. RESULTS: The expressions of platelet and plasma miR-223 and miR-146a were significantly downregulated in patients with ischemic stroke and diabetes mellitus or diabetes mellitus only, but not in patients with ischemic stroke only compared to healthy controls. The expressions of platelet and plasma miR-495 and miR-107 showed no significant differences among these four groups. The expression of platelet miR-223 and miR-146a significantly correlated with plasma miR-223 and miR-146a levels, blood glucose concentration, and platelet activation rate. CONCLUSIONS: Hyperglycemia may downregulate the expressions of miR-223 and miR-146a, leading to subsequent platelet activation in patients with diabetes mellitus. Low platelet and plasma miR-223 and miR-146a expression is a risk factor for ischemic stroke in Chinese diabetes mellitus patients.

Association of BSG genetic polymorphisms with atherosclerotic cerebral infarction in the Han Chinese population.

The Basigin (BSG, also known as CD147/extracellular matrix metalloproteinase inducer) belongs to the immunoglobulin superfamily (IgSF). It is a cellular receptor for cyclophilin A (CypA), and is originally known as tumor cell collagenase stimulatory factor (TCSF), which could abundantly expressed on the surface of tumor cells, haematopoietic, monocytes, epithelial endothelial cells and smooth muscle cells. Accumulating evidence showed that BSG played an important role in stimulating the secretion of matrix metalloproteinases (MMPs), which has been reported to be involved in the development of atherosclerosis. Since atherosclerosis is an important risk factor for atherosclerotic cerebral infarction (ACI), we speculate that BSG genetic polymorphisms may influence formation of atherosclerosis and then development of ACI. This study aimed to detect the potential association of the single nucleotide polymorphisms (SNP, -631 G > T, -318 G > C, 10141 G > A and 10826 G > A) of BSG gene in Hunan Han Chinese population with ACI. We genotyped 199 ACI patients and 188 matched healthy controls for the four BSG SNP by method of matrix-assisted laser desorption/ionization-time-offlight mass spectrometry (MALDI-TOF MS). Our results suggested that all the polymorphisms were observed in the subjects from Changsha area of Hunan Province. However, no significant difference was observed between the distribution of these SNP in cases and controls. Therefore, we speculate that BSG genetic polymorphisms might not be an important factor in the development of ACI in our Chinese Han population.

A functional polymorphism at miR-491-5p binding site in the 3'-UTR of MMP-9 gene confers increased risk for atherosclerotic cerebral infarction in a Chinese population.

BACKGROUND: and purpose: Matrix metalloproteinases 9 (MMP-9) has been reported to play a critical role in the pathophysiology of atherosclerotic cerebral infarction (ACI). Here we assessed association of MMP-9 polymorphisms with ACI susceptibility and the function of SNPs through microRNA mediated regulation. METHODS: Genotyping was performed using MALDI-TOF mass spectrometry. Reporter gene plasmids with the MMP-9 3'UTR carrying either the mutant or the wild-type MMP-9 allele were constructed. Also, we constructed pcDNA-3.1-miR-491-5p recombinant plasmid, which transiently co-transfected human umbilical vein endothelial cells (HUVEC) with the reporter plasmids. Reporter plasmids, miR-491-5p mimics and inhibitor were transfected into HUVE cells line by lipofectamine. MMP-9 mRNA expression in HUVEC was detected by RT-PCR and protein level by ELISA. RESULTS: The rs1802908 and rs2664517 polymorphisms were not observed in all subjects from Hunan Han Chinese. No significant difference in genotype distribution of rs20544 and rs9509 between cases and controls were observed (p＞0.05). The rs1056628CC genotype had a significantly increased risk for ACI as compared with carries of the rs1056628 A allele (total χ(2) = 12.041, P = 0.002). Reporter gene assay revealed that the rs1056628 A allele showed lower reporter activity than the rs1056628C allele. Hsa-miR-491-5p had effect on modulation of MMP-9 gene in vitro. The rs1056628 A→C variant in the 3'-UTR of the MMP-9 increased MMP-9 protein expression in cultured HUVECs. CONCLUSIONS: Our data suggested that the rs1056629A→C variation contributes to an increased risk of ACI by increasing MMP-9 expression through affecting binding of miR-491 to the polymorphic site in the 3'-UTR of MMP-9.

Association of PTEN genetic polymorphisms with atherosclerotic cerebral infarction in the Han Chinese population.

The phosphatase and tensin homologue (PTEN) gene is critical to the pathological development of atherosclerosis, a major risk factor for atherosclerotic cerebral infarction (ACI). However, it remains unclear whether genetic polymorphism in the PTEN gene can affect the risk of developing ACI. In the present haplotype-based case-control study, we investigated whether polymorphisms of the PTEN gene were associated with ACI in a Han Chinese population. Using matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS), we genotyped 300 patients with ACI and 204 matched healthy controls for five single nuclear polymorphisms (SNP) of the PTEN gene (rs2299939, rs17431184, rs555895, rs12357281 and rs2673836). Linkage disequilibrium and haplotype construction were analyzed using SHEsis software. We found that the rs2299939 PTEN intronic polymorphism was significantly associated with a risk of ACI in our Han Chinese population. The rs555895 and rs17431184C/T variants were not associated with risk of ACI. Two other polymorphisms were not detected in any subject in this study. The rs2299939C/rs17431184T/rs555895T PTEN haplotype was also significantly associated with risk of ACI. Our study provides the first evidence of an association between a PTEN intronic variant and the risk of ACI. It also provides new a new line of evidence for the critical role of PTEN in the development of stroke.