Search

Objective: Osteoclast (OC) over-activation is an important cause of bone loss that is strongly correlated with inflammation. Although the CD163/TWEAK/Fn14 axis has been implicated in several inflammatory pathologies, its contributions to inflammatory bone loss remain poorly understood. This study aimed to evaluate the interaction of the CD163/TWEAK/Fn14 axis with OC in inflammatory bone loss. Methods: To assess the role of CD163 in bone homeostasis, we characterized the bone phenotypes of CD163-deficient mice and their wild-type littermates. CD163 and TWEAK levels were evaluated in the bone marrow of mice with LPS-induced bone loss and individuals with rheumatoid arthritis (RA). Bone mass changes were assessed using uCT and histology following supplementation with recombinant mouse CD163 protein (rCD163) or blockade of TWEAK/Fn14 signaling in CD163-deficient mice and mice with LPS-induced bone loss. The impact of CD163/TWEAK on OC differentiation and bone resorption capacity was analyzed in vitro. Results: CD163 deficiency caused decreased bone mass and increased OC abundance. Lower CD163 expression and higher TWEAK expression were observed in the bone marrow of mice with LPS-induced bone loss and individuals with RA. TWEAK, mainly derived from CD68+ macrophages, was responsible for bone loss, and supplementing rCD163 or blocking TWEAK/Fn14 signaling contributed to rescue bone loss. TWEAK/Fn14 synergistically promoted RANKL-dependent OC differentiation and bone resorption capability through downstream mitogen-activated protein kinases (MAPK) signaling, while the pro-osteoclastic effect of TWEAK was suppressed by CD163. Conclusion: Our findings suggest that the CD163/TWEAK/Fn14 axis is a potential therapeutic target for inflammatory bone loss by regulating osteoclastogenesis.

Cyprinus acutidorsalis (Wang, 1979) is an endemic fish in China that is sparsely distributed in the Hainan provinces and Guangxi Zhuang Autonomous Region (GZAR). In this study, the complete mitochondrial genome of C. acutidorsalis from the Hainan population from the Wanquan River was sequenced, and its phylogenetic relationship was analyzed. The circular mtDNA was 16,581 bp in length, and the overall base composition was A (32.0%), C (27.5%), T (24.8%), and G (15.70%), with a slight bias toward A + T. The complete mitogenome encoded 13 protein-coding genes (PCGs), 22 tRNA genes, two rRNA genes, and a control region. Phylogenetic analysis indicated that the most closely related fish to C. acutidorsalis from the Hainan population was C. acutidorsalis from the Guangxi population. These findings offer basic molecular data and a better understanding of the phylogenetic relationships among the Cyprinus species.

BackgroundDiabetes mellitus (DM), a chronic metabolic disease characterized by elevated blood sugar, leads to delayed or non-healing wounds, increasing amputation risks, and placing a significant burden on patients and society. While extensive research has been conducted on adipose-derived stem cells (ADSCs) for promoting wound healing, there is a scarcity of studies focusing on diabetic wounds, particularly those employing proteomics and bioinformatics approaches.ObjectiveThis study aimed to investigate the mechanisms by which ADSCs promote diabetic wound healing using proteomics and bioinformatics techniques.MethodsHealthy rat fat tissue was used to isolate ADSCs. A T2DM rat model with back wounds was established. The experimental group received ADSC injections around the wound, while the control group received PBS injections. Wound healing rates were documented and photographed on days 0, 3, 7, 10, and 14. On day 7, wound tissues were excised for HE and Masson’s staining. Additionally, on day 7, tissues were analyzed for protein quantification using 4D-DIA, with subsequent GO and KEGG analyses for differentially expressed proteins (DEPs) and protein-protein interaction (PPI) network analysis using STRING database (String v11.5). Finally, Western blot experiments were performed on day 7 wounds to verify target proteins.Results and ConclusionsIn all measured days postoperatively, the wound healing rate was significantly higher in the ADSC group than in the PBS group (day 7: p < 0.001, day 10: p = 0.001, day 14: p < 0.01), except on day 3 (p > 0.05). Proteomic analysis identified 474 differentially expressed proteins, with 224 key proteins after PPI analysis (78 upregulated and 146 downregulated in the ADSC group). The main cellular locations of these proteins were “cellular anatomical entity” and “protein-containing complex”, while the biological processes were “cellular processes” and “biological regulation”. The primary molecular functions were “binding” and “catalytic activity”, with GO enrichment focused on “Wnt-protein binding”, “neural development”, and “collagen-containing extracellular matrix”. Further analysis of PPI network nodes using LASSO regression identified Thy1 and Wls proteins, significantly upregulated in the ADSC group, as potentially crucial targets for ADSC application in diabetic wound treatment.

Abstract Background Suture knotting is the basis of surgical skills. In the process of surgical skills learning, the surrounding environment, especially the light, will affect the efficiency of learning. This study investigated the effect of optical environment on the learning of stitching and knotting skills. Methods A total of 44 medical students were randomly divided into four groups and participated in the study of suture knotting in four different optical environments. During the process, we assess objective pressure level by testing salivary amylase activity Likert scale and objective structured clinical examination (OSCE) was used to estimate the subjective psychological state and overall skill mastery in surgical suturing respectively. Results Under high illumination conditions (700 lx), the salivary amylase activity of the high color temperature group (6000 K) was significantly higher than that of the low color temperature group (4000 K) (p

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease caused by the SFTS virus (SFTSV). Predicting the incidence of this disease in advance is crucial for policymakers to develop prevention and control strategies. In this study, we utilized historical incidence data of SFTS (2013–2020) in Shandong Province, China to establish three univariate prediction models based on two time-series forecasting algorithms Autoregressive Integrated Moving Average (ARIMA) and Prophet, as well as a special type of recurrent neural network Long Short-Term Memory (LSTM) algorithm. We then evaluated and compared the performance of these models. All three models demonstrated good predictive capabilities for SFTS cases, with the predicted results closely aligning with the actual cases. Among the models, the LSTM model exhibited the best fitting and prediction performance. It achieved the lowest values for mean absolute error (MAE), mean square error (MSE), and root mean square error (RMSE). The number of SFTS cases in the subsequent 5 years in this area were also generated using this model. The LSTM model, being simple and practical, provides valuable information and data for assessing the potential risk of SFTS in advance. This information is crucial for the development of early warning systems and the formulation of effective prevention and control measures for SFTS.

Microbial natural products and their derivatives have been developed as a considerable part of clinical drugs and agricultural chemicals. Marine microbial natural products exhibit diverse chemical structures and bioactivities with substantial potential for the development of novel pharmaceuticals. However, discovering compounds with new skeletons from marine microbes remains challenging. In recent decades, multiple approaches have been developed to discover novel marine microbial natural products, among which heterologous expression has proven to be an effective method. Facilitated by large DNA cloning and comparative metabolomic technologies, a few novel bioactive natural products from marine microorganisms have been identified by the expression of their biosynthetic gene clusters (BGCs) in heterologous hosts. Heterologous expression is advantageous for characterizing gene functions and elucidating the biosynthetic mechanisms of natural products. This review provides an overview of recent progress in heterologous expression-guided discovery, biosynthetic mechanism elucidation, and yield optimization of natural products from marine microorganisms and discusses the future directions of the heterologous expression strategy in facilitating novel natural product exploitation.

The known universal primers for environmental DNA (eDNA) metabarcoding of fish are mainly located in the mitochondrial ribosome gene regions. Not only are the reference sequences insufficient but also some relative fish species could not be identified using these 12S and 16S primers. In this study, the freshwater fishes of Hainan Island were selected to meet six investigation targets. (1) Six conserved regions were selected through the cytochrome c oxidase subunit I (COⅠ) sequences of 8 orders, 26 families, 101 genera, and 150 species; (2) Based on the base variation, species identification, eDNA degradation, and read length requirements of high-throughput sequencing, 26 primers were designed from four flanking conserved regions; however, six primers did not meet the Premier score requirements. (3) The first-round PCR results of 72 freshwater fish species on Hainan Island showed that 11 primers had high universality. Among them, there were 5 primers with more than 70 species successfully amplified and brighter PCR bands than marker. The second-round of PCR results showed that the amplification success rate of 3×2 (forward and reverse, respectively) primer combinations generated by the five primers was 100%, and the optimal primer combinations after PCR band length and brightness screening were "HN-A-F4, HN-D-R3" (hereinafter referred to as HN-COⅠ). (4) High-throughput sequencing results of 30 water samples showed that the total number of clean reads, fish sequences, operational taxonomic units (OTUs), and fish OTUs generated by HN-COⅠ were 0.77, 1.22, 0.85, and 1.32 times those of MiFish-U, respectively. (5) The proportion of fish OTUs annotated to species, genus, and family was 81.40%, 11.63%, and 6.98% for HN-COⅠ and 81.54%, 4.62%, and 13.85% for MiFish-U, respectively. The non-metric multidimensional scaling (NMDS) clustering pattern of "primer + water sample" formed two distinct groups with an obvious boundary (stress=0.15). The minimum and average values of pairwise genetic distances of species amplicons within genera of HN-COⅠ were 1.23 and 1.57 times those of MiFish-U, respectively. (6) The correlation coefficient of the linear regression equation of common carp biomass and sequence copies of the six indoor density groups was low. HN-COⅠ and MiFish-U could not accurately reflect the biomass of common carp. In this study, the advantages and disadvantages of HN-COⅠ were screened and compared with those of MiFish-U. The results showed that HN-COⅠ primers have a high targeting ability for freshwater fishes on Hainan Island. HN-COⅠ are advantageous in preventing eDNA contamination from non-target organisms, such as microorganisms and mammals, and is also conducive to the detection and accurate identification of freshwater fishes.

ABSTRACT: Transport stress (TS) not only weakens poultry performance but also affects animal welfare. Additionally, TS can evoke cardiac damage by triggering sterile inflammation in chicks, but the underlying mechanism is not fully understood. Here, we aimed to elucidate how TS induces sterile inflammation and heart injury and to clarify the antagonism effect of astragalus polysaccharides (APS). We randomly divided 60 chicks (one-day-old female) into 5 groups (n = 12): Control_0h (Con_0h) group (chicks were slaughtered at initiation), Control group (stress-free control), TS group (simulated TS exposure for 8 h), TS plus water (TS+W) group, and TS plus APS (TS+APS) group. Before simulation transport, the chicks of TS+W and TS+APS groups were, respectively, dietary with 100 μL of water or APS (250 μg/mL). H&E staining was employed for cardiac histopathological observation. ELISA assay was used to measure oxidative stress marker levels (GSH, GPX, GST, and MDA). A commercial kit was used to isolate the mitochondrial portion, and qRT-PCR was employed to measure the mitochondrial DNA (mtDNA) levels. Furthermore, we evaluated the activity of mtDNA-mediated NF-κB, NLRP3 inflammasome, and cGAS-STING inflammatory pathways and the expression of downstream inflammatory factors by Western Blotting or qRT-PCR. Our findings revealed that APS notably relieved TS-induced myocardial histopathological lesions and infiltrations. Likewise, the decrease in proinflammatory factors (TNF-α, IL-1β, and IL-6) and IFN-β by APS further supported this result. Meanwhile, TS caused severe oxidative stress in the chick heart, as evidenced by decreased antioxidant enzymes and increased MDA. Importantly, APS prevented mtDNA stress and leakage by reducing oxidative stress. Interestingly, TS-induced mtDNA leakage caused a series of inflammation events via mtDNA-PRRs pathways, including TLR21-NF-κB, NLRP3 inflammasome, and cGAS-STING signaling. Encouragingly, all these adverse changes related to inflammation events induced by mtDNA-PRRs activation were all relieved by APS treatment. In summary, our findings provide the first evidence that inhibition of mtDNA-PRRs pathway-mediated sterile inflammation by APS could protect against TS-induced cardiac damage in chicks.

Tick-borne Apicomplexan parasites pose a significant threat to both public health and animal husbandry. Identifying potential pathogenic parasites and gathering their epidemiological data are essential for prospectively preventing and controlling infections. In the present study, genomic DNA of ticks collected from two goat flocks (Goatflock1 and Goatflock2) and one dog group (Doggroup) were extracted and the 18S rRNA gene of Babesia/Theileria/Colpodella spp. was amplified by PCR and sequenced. Phylogenetic analysis was conducted based on the obtained sequences. The differences in pathogen positive rates between ticks of different groups were statistically analyzed using the Chi-square or continuity-adjusted Chi-square test. As a result, two pathogenic Theileria (T.) luwenshuni genotypes, one novel pathogenic Colpodella sp. HLJ genotype, and two potential novel Colpodella spp. (referred to as Colpodella sp. struthionis and Colpodella sp. yiyuansis in this study) were identified in the Haemaphysalis (H.) longicornis ticks. Ticks of Goatflock2 had a significantly higher positive rate of Colpodella spp. than those from Goatflock1 (χ2=92.10; P = 8.2 × 10−22) and Doggroup (χ2=42.34; P = 7.7 × 10−11), and a significantly higher positive rate of T. luwenshuni than Doggroup (χ2=5.38; P = 0.02). However, the positive rates of T. luwenshuni between Goatflock1 and Goatflock2 were not significantly different (χ2=2.02; P = 0.16), and so as the positive rates of both pathogens between Goatflock1 and Doggroup groups (P > 0.05). For either Colpodella spp. or T. luwenshuni, no significant difference was found in prevalence between male and female ticks. These findings underscore the potential importance of Colpodella spp. in domestic animal-attached ticks, as our study revealed two novel Colpodella spp. and identified Colpodella spp. in H. longicornis for the first time. The study also sheds light on goats' potential roles in the transmission of Colpodella spp. to ticks and provides crucial epidemiological data of pathogenic Theileria and Colpodella. These data may help physicians, veterinarians, and public health officers prepare suitable detection and treatment methods and develop prevention and control strategies.

Abstract Background Retifanlimab is a humanized monoclonal antibody targeting programmed death protein‐1, and INCB001158 is an oral arginase inhibitor. This phase Ib study investigated retifanlimab, INCB001158, and their combination in Japanese patients with advanced solid tumors. Methods Patients received retifanlimab (500 mg every 4 weeks [Q4W] i.v.) or escalating doses of INCB001158 (75 or 100 mg twice daily [BID]) monotherapy in Part 1 and combination of retifanlimab (500 mg Q4W) and INCB001158 (100 mg BID) in Part 2. Primary endpoints were safety, tolerability, dose‐limiting toxicities (DLTs), and determination of recommended phase II doses in Japanese patients. Results Eighteen patients (retifanlimab or INCB001158 monotherapy and combination; n = 6 each) were enrolled at 2 sites in Japan. There were no DLTs, fatal adverse events (AEs), or discontinuations due to AEs. Rash (all grade 1) was the most common treatment‐emergent AE with retifanlimab (n = 6). Treatment‐related AEs were reported with retifanlimab (n = 4) or INCB001158 (n = 2) monotherapy and with combination (n = 4); an immune‐related AE (thyroid disorder, grade 2) was reported with combination. Two responses were observed with retifanlimab monotherapy (1 complete, 1 partial) and 1 stable disease (SD), for an overall response rate of 33.3% (95% confidence interval [CI], 4.3–77.7) and disease control rate (DCR) of 50% (95% CI, 11.8–88.2). Three patients had SD with INCB001158 monotherapy (DCR 50%; 95% CI, 11.8–88.2). No responses or SD were observed with combination therapy. Conclusion Retifanlimab, INCB001158, and their combination had acceptable safety profiles. Promising retifanlimab antitumor activity warrants further investigation in Japanese patients.

The main objective of the present study was to investigate factors related to seafarers’ mental health. A sample of seafarers from 12 countries participated in the study. A list of stressors was used to assess both perception of exposure to these stressors and their subjective significance. The Symptom Checklist (SCL-90) was used to assess seafarers’ mental health on 5 of 9 subscales: Depression, Anxiety, Hostility, Interpersonal Sensitivity, and Somatisation. Three significant findings emerged from the analyses. The first was that 3 types of stressors contribute significantly to mental health problems: (1) environmental factors (eg, vibration), (2) social problems (eg, bullying, homesickness, working alone), and (3) health problems (eg, physical injuries, viruses, and the illnesses). The second finding was that both stress and mental health issues determine seafarers’ motivation for their work and their consideration regarding leaving the maritime industry. The third finding was that factors contributing to seafarers’ consideration of leaving the industry were mainly related to social stressors such as isolation from family and friends, cultural differences at work, demands from supervisors, and bullying. Factors such as bad weather, working shifts, length of employment contract or a ban on disembarkment in ports were found to be relatively less important for seafarers as factors toward considering leaving the industry. The implications of these findings are discussed.

The hygiene hypothesis proposes that decreased exposure to infectious agents in developed countries may contribute to the development of allergic and autoimmune diseases. Trichinella spiralis, a parasitic roundworm, causes trichinellosis, also known as trichinosis, in humans. T. spiralis had many hosts, and almost any mammal could become infected. Adult worms lived in the small intestine, while the larvae lived in muscle cells of the same mammal. T. spiralis was a significant public health threat because it could cause severe illness and even death in humans who eat undercooked or raw meat containing the parasite. The complex interactions between gastrointestinal helminths, gut microbiota, and the host immune system present a challenge for researchers. Two groups of mice were infected with T. spiralis vs uninfected control, and the experiment was conducted over 60 days. The 16S rRNA gene sequences and untargeted LC/MS-based metabolomics of fecal and serum samples, respectively, from different stages of development of the Trichinella spiralis-mouse model, were examined in this study. Gut microbiota alterations and metabolic activity accompanied by parasite-induced immunomodulation were detected. The inflammation parameters of the duodenum (villus/crypt ratio, goblet cell number and size, and histological score) were involved in active inflammation and oxidative metabolite profiles. These profiles included increased biosynthesis of phenylalanine, tyrosine, and tryptophan while decreasing cholesterol metabolism and primary and secondary bile acid biosynthesis. These disrupted metabolisms adapted to infection stress during the enteral and parenteral phases and then return to homeostasis during the encapsulated phase. There was a shift from an abundance of Bacteroides in the parenteral phase to an abundance of probiotic Lactobacillus and Treg-associated-Clostridia in the encapsulated phase. Th2 immune response (IL-4/IL-5/IL-13), lamina propria Treg, and immune hyporesponsiveness metabolic pathways (decreased tropane, piperidine and pyridine alkaloid biosynthesis and biosynthesis of alkaloids derived from ornithine, lysine, and nicotinic acid) were all altered. These findings enhanced our understanding of gut microbiota and metabolic profiles of Trichinella -infected mice, which could be a driving force in parasite-shaping immune system maintenance.

The tumor microenvironment stimulates the angiogenic activity of endothelial cells (ECs) to facilitate tumor vascularization, growth, and metastasis. The involvement of microRNA-186-5p (miR-186) in regulating the aberrant activity of tumor-associated ECs has so far not been clarified. In the present study, we demonstrated that miR-186 is significantly downregulated in ECs microdissected from human non-small cell lung cancer (NSCLC) tissues compared with matched non-malignant lung tissues. In vitro analyses of primary human dermal microvascular ECs (HDMECs) exposed to different stimuli indicated that this miR-186 downregulation is triggered by hypoxia via activation of hypoxia-inducible factor 1 alpha (HIF1α). Transfection of HDMECs with miR-186 mimic (miR-186m) significantly inhibited their proliferation, migration, tube formation, and spheroid sprouting. In contrast, miR-186 inhibitor (miR-186i) exerted pro-angiogenic effects. In vivo, endothelial miR-186 overexpression inhibited the vascularization of Matrigel plugs and the initial growth of tumors composed of NSCLC cells (NCI-H460) and HDMECs. Mechanistic analyses revealed that the gene encoding for protein kinase C alpha (PKCα) is a bona fide target of miR-186. Activation of this kinase significantly reversed the miR-186m-repressed angiogenic activity of HDMECs. These findings indicate that downregulation of miR-186 in ECs mediates hypoxia-stimulated NSCLC angiogenesis by upregulating PKCα.

Type H vessels are specialized blood vessels found in the bone marrow that are closely associated with osteogenic activity. They are characterized by high expression of endomucin and CD31. Type H vessels form in the cancellous bone area during long bone development to provide adequate nutritional support for cells near the growth plate. They also influence the proliferation and differentiation of osteoprogenitors and osteoclasts in a paracrine manner, thereby creating a suitable microenvironment to facilitate new bone formation. Because of the close relationship between type H vessels and osteogenic activity, it has been found that type H vessels play a role in the physiological and pathological processes of bone diseases such as fracture healing, osteoporosis, osteoarthritis, osteonecrosis, and tumor bone metastasis. Moreover, experimental treatments targeting type H vessels can improve the outcomes of these diseases. Here, we reviewed the molecular mechanisms related to type H vessels and their associated osteogenic activities, which are helpful in further understanding the role of type H vessels in bone metabolism and will provide a theoretical basis and ideas for comprehending bone diseases from the vascular perspective.

Abstract Background Left-to-right shunt congenital heart disease is more likely to induce recurrent respiratory infections in the patients which exacerbate pulmonary hypertension and thereby impairs cardiac function. It is urgent to explore a non-invasive and accurate diagnostic method that can show the cardiac anatomy and associated malformations in clinical research. Objective To determine the diagnostic value of peripheral mucin domain protein-3 (Tim-3), N-terminal pro-brain natriuretic peptide (NT proBNP), sestrin2 testing in patients with the left-to-right shunt congenital heart disease and heart failure. Methods Fifty-two neonates with with left to right shunt congenital heart disease and 30 healthy neonates were enrolled. Blood samples were collected within 24 h of admission from newborns for determining the content of TiM-3, NT proBNP, and Sestrin2. Analyzing the ROC curve provided insight into the diagnostic accuracy. Both a Spearman’s rank correlation test and a logistic regression analysis were carried out. Results TiM-3, NT proBNP, and Sestrin2 levels in peripheral blood were statistically different in the three groups (P

Abstract Purpose To systematically evaluate the curative efficacy and safety of platelet-rich plasma (PRP) combined with hyaluronic acid (HA) in the treatment of knee osteoarthritis (KOA), comparing with platelet-rich plasma alone. Methods Cochrane Library, PubMed, China National Knowledge Infrastructure (CNKI) and Embase were searched for randomized controlled trials (RCTs) and cohort studies regarding the efficacy and safety of platelet-rich plasma (PRP) combined with hyaluronic acid (HA) in the treatment of knee osteoarthritis (KOA) comparing with platelet-rich plasma alone before January 15, 2022. The methodological quality of the ultimately included studies was assessed comprehensively, and meta-analysis was implemented using RevMan 5.3 software. Results Thirteen articles (9 RCTs, 4 cohort studies), including 1118 patients, were covered. There was no significant difference between the PRP + HA therapy and PRP-alone therapy in VAS scores at 3 months, 6 months and 12 months, WOMAC total scores at 3 months and KOOS at 1 month and 6 months. Compared with PRP-alone therapy, PRP + HA therapy was associated with significantly better improvement in VAS scores at 1 month, WOMAC total scores at 6 months, KOOS at 3 months, IKDC scores at 6 months and Lequesne index scores at 3 and 6 months. However, the smallest treatment effect of VAS scores, WOMAC total scores, KOOS and IKDC scores did not exceed the minimum clinically important difference (MCID). However, PRP + HA therapy got a greater reduction in the rate of adverse events, compared with PRP-alone therapy. Conclusion The results of this meta-analysis indicated that PRP + HA therapy was not found to be superior to PRP-alone therapy in pain relief and function improvement for patients with KOA. However, combined PRP with HA injections was generally safer than PRP injections alone, by assessing the incidence of adverse events.

High-fat diet induces bone marrow inflammation and osteoarthritis phenotype in knee joint, but the underlying mechanisms is unknown. Here, we report that high-fat diet induces aberrant bone formation and cartilage degeneration in knee joint. Mechanistically, a high-fat diet increases the number of macrophages and the secretion of prostaglandins in subchondral bone, promoting bone formation. Metformin treatment is able to decrease the number of macrophages and also the level of prostaglandins induced by high-fat diet in subchondral bone. Importantly, metformin rescues aberrant bone formation and cartilage lesions by decreasing the number of osteoprogenitors and type-H vessels, which also results in relief of osteoarthritis pain response. Thus, we demonstrate prostaglandins secreted by macrophages may be a key reason for high-fat diet induced aberrant bone formation and metformin is a promising therapy for high-fat diet induced osteoarthritis.

Rotundic acid (RA), an ursane-type pentacyclic triterpene acid isolated from the dried barks of Ilex rotunda Thunb. (Aquifoliaceae), possesses diverse bioactivities. To further study its pharmacokinetics, a simple and sensitive liquid chromatography with triple quadrupole mass spectrometry (LC-QqQ-MS/MS) method was developed and validated to quantify RA concentration in rat plasma and tissue using etofesalamide as an internal standard (IS). Plasma and tissue samples were subjected to one-step protein precipitation. Chromatographic separation was achieved on a ZORBAX Eclipse XDB-C18 column (4.6 mm × 50 mm, 5 μm) under gradient conditions with eluents of methanol:acetonitrile (1:1, V/V) and 5 mM ammonium formate:methanol (9:1, V/V) at 0.5 mL/min. Multiple reaction monitoring transitions were performed at m/z 487.30 → 437.30 for RA and m/z 256.10 → 227.10 for IS in the negative mode. The developed LC-QqQ-MS/MS method exhibited good linearity (2–500 ng/mL) and was fully validated in accordance with U.S. Food and Drug Administration bioanalytical guidelines. Dose proportionality and bioavailability in rats were determined by comparing pharmacokinetic data after single oral (10, 20, and 40 mg/kg) and intravenous (10 mg/kg) administration of RA. Tissue distribution was studied following oral administration at 20 mg/kg. The results showed that the absolute bioavailability of RA after administration at different doses ranged from 16.1% to 19.4%. RA showed good dose proportionality over a dose range of 10–40 mg/kg. RA was rapidly absorbed in a dose-dependent manner and highly distributed in the liver. In conclusion, this study is the first to systematically elucidate the absorption and distribution characteristics of RA in rats, which can provide additional information for further development and evaluation of RA in drug metabolism and pharmacokinetic studies.

Chitosan are abundant polysaccharides in nature, which play many important roles in food, medicine, agriculture, environmental protection and other fields. The deacetylation degree of chitosan is also one of the most important indexes to evaluate their performance. Therefore, the convenience and accuracy of deacetylation degree detection method are highly required in application. In this paper, the chemical analysis methods, spectral analysis methods, destructive analysis methods and electrical characteristics analysis methods are summarized, and the advantages and disadvantages of various methods and their improvement schemes are discussed. It is found that chemical analysis and spectroscopy have not only good detection results, but also a wider range of applications, while electrical characteristics analysis methods have broad prospects for development. The antibacterial and fresh-keeping ability of chitosan with a high degree of deacetylation is valued, while chitosan with a low degree of deacetylation plays an important role in medicine and environmental protection. The functions of chitosan of antibacterial, preservation and packaging in the food industry and application in environmental protection and tissue engineering were introduced to provide reference for the determination of the deacetylation degree of chitosan.

Abstract Background Radioresistance inducing by hypoxic microenvironment of hepatocellular carcinoma is a major obstacle to clinical radiotherapy. Advanced nanomedicine provides an alternative to alleviate the hypoxia extent of solid tumor, even to achieve effective synergistic treatment when combined with chemotherapy or radiotherapy. Results Herein, we developed a self-assembled nanoparticle based on hemoglobin and curcumin for photoacoustic imaging and radiotherapy of hypoxic hepatocellular carcinoma. The fabricated nanoparticles inhibited hepatoma migration and vascular mimics, and enhanced the radiosensitivity of hypoxic hepatoma cells in vitro via repressing cell proliferation and DNA damage repair, as well as inducing apoptosis. Benefit from oxygen-carrying hemoglobin combined with polyphenolic curcumin, the nanoparticles also effectively enhanced the photoacoustic contrast and the efficacy of radiotherapy for hepatocellular carcinoma in vivo. Conclusions Together, the current study offered a radiosensitization platform for optimizing the efficacy of nanomedicines on hypoxic radioresistant tumor. Graphical Abstract

MicroRNAs (miRNAs) expressed in endothelial cells (ECs) are powerful regulators of angiogenesis, which is essential for tumor growth and metastasis. Here, we demonstrated that miR-22 is preferentially and highly expressed in ECs, while its endothelial level is significantly downregulated in human non-small cell lung cancer (NSCLC) tissues when compared to matched nontumor lung tissues. This reduction of endothelial miR-22 is possibly induced by NSCLC cell-secreted interleukin-1β and subsequently activated transcription factor nuclear factor-κB. Endothelial miR-22 functions as a potent angiogenesis inhibitor that inhibits all of the key angiogenic activities of ECs and consequently NSCLC growth through directly targeting sirtuin 1 and fibroblast growth factor receptor 1 in ECs, leading to inactivation of AKT/mammalian target of rapamycin signaling. These findings provide insight into the molecular mechanisms of NSCLC angiogenesis and indicate that endothelial miR-22 represents a potential target for the future antiangiogenic treatment of NSCLC.

Abstract Objectives The COVID-19 pandemic has impacted the mental health of people worldwide. Psychological resilience has been shown to buffer against the threat of the pandemic (i.e., COVID-19 fear) and sustain mental health. The extent to which psychological resilience factors impact mental health maintenance, however, is unclear, given broad differences in infection rates, prevention approaches, government interventions across different cultures and contexts. Our study examines resilience factors and how they protect individuals from COVID-19-related fear and sustain their mental health. Data description Data were collected from 1583 (Mage = 32.22, SD = 12.90, Range = 19–82) respondents from Japan, China, the United States, and Malaysia between October to November 2020. We collected data across age and sex, marital status, number of children, and occupations. We also accounted for stay-at-home measures, change in income, COVID-19 infection status, place of residence, and subjective social status in the study. Our variables included mental health-related and resilience constructs, namely (i) fear of COVID-19, (ii) depression, anxiety, and stress; (iii) present, past, and future life satisfaction, (iv) sense of control, (v) positive emotions, (vi) ego-resilience, (vii) grit, (viii) self-compassion, (ix) passion, and (x) relational mobility. All questionnaires were assessed for their suitability across the four countries with the necessary translation checks. Results from this study can be instrumental in examining the impact of multiple resilience factors and their interaction with demographic variables in shaping mental health outcomes.

Tumor endothelial cells (TECs) are key stromal components of the tumor microenvironment, and are essential for tumor angiogenesis, growth and metastasis. Accumulating evidence has shown that small single-stranded non-coding microRNAs (miRNAs) act as powerful endogenous regulators of TEC function and blood vessel formation. This systematic review provides an up-to-date overview of these endothelial miRNAs. Their expression is mainly regulated by hypoxia, pro-angiogenic factors, gap junctions and extracellular vesicles, as well as long non-coding RNAs and circular RNAs. In preclinical studies, they have been shown to modulate diverse fundamental angiogenesis-related signaling pathways and proteins, including the vascular endothelial growth factor (VEGF)/VEGF receptor (VEGFR) pathway; the rat sarcoma virus (Ras)/rapidly accelerated fibrosarcoma (Raf)/mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) pathway; the phosphoinositide 3-kinase (PI3K)/AKT pathway; and the transforming growth factor (TGF)-β/TGF-β receptor (TGFBR) pathway, as well as krüppel-like factors (KLFs), suppressor of cytokine signaling (SOCS) and metalloproteinases (MMPs). Accordingly, endothelial miRNAs represent promising targets for future anti-angiogenic cancer therapy. To achieve this, it will be necessary to further unravel the regulatory and functional networks of endothelial miRNAs and to develop safe and efficient TEC-specific miRNA delivery technologies.

An electrochemical sensor based on AuPtPd trimetallic nanoparticles functionalized multi-walled carbon nanotubes coupled with chitosan modified glassy carbon electrode (GCE/CS/MWCNTs-AuPtPd) was proposed for the rapid detection of bisphenol A (BPA). AuPtPd trimetallic nanoparticles were first assembled on MWCNTs to obtain MWCNTs-AuPtPd nanocomposite. Then, the MWCNTs-AuPtPd was further dispersed on the chitosan-modified electrode surface to fabricate the GCE/CS/MWCNTs-AuPtPd sensor. Due to the superior catalytic properties of MWCNTs-AuPtPd and the good film formation of chitosan, the constructed sensor displayed good performances for BPA detection. The structural morphology of CS/MWCNTs-AuPtPd was characterized in many ways, such as SEM, TEM and UV-vis. The designed sensor showed a linear relationship in concentration range from 0.05 to 100 µM for BPA detecting, and the detection limit was 1.4 nM. The GCE/CS/MWCNTs-AuPtPd was further used to realize the detection of BPA in food samples, and the recovery was between 94.4% and 103.6%. Those results reflected that the electrochemical sensor designed by CS/MWCNTs-AuPtPd nanocomposites was available, which could be used for the monitoring of food safety.

Angiogenesis, the development of new blood vessels from pre-existing ones, is an essential process determining numerous physiological and pathological conditions. Accordingly, there is a high demand for research approaches allowing the investigation of angiogenic mechanisms and the assessment of pro- and anti-angiogenic therapeutics. The present review provides a selective overview and critical discussion of such approaches, which, in line with the 3R principle, all share the common feature that they are not based on animal experiments. They include in vitro assays to study the viability, proliferation, migration, tube formation and sprouting activity of endothelial cells in two- and three-dimensional environments, the degradation of extracellular matrix compounds as well as the impact of hemodynamic forces on blood vessel formation. These assays can be complemented by in vivo analyses of microvascular network formation in the chorioallantoic membrane assay and early stages of zebrafish larvae. In addition, the combination of experimental data and physical laws enables the mathematical modeling of tissue-specific vascularization, blood flow patterns, interstitial fluid flow as well as oxygen, nutrient and drug distribution. All these animal-free approaches markedly contribute to an improved understanding of fundamental biological mechanisms underlying angiogenesis. Hence, they do not only represent essential tools in basic science but also in early stages of drug development. Moreover, their advancement bears the great potential to analyze angiogenesis in all its complexity and, thus, to make animal experiments superfluous in the future.

Context Rotundic acid (RA), a plant-derived pentacyclic triterpene acid, has been reported to possess extensive pharmacological activities. The poor bioavailability limits its further development and potential clinic application. Objective To clarify the potential mechanism for poor oral bioavailability. Materials and methods The single-dose pharmacokinetics of orally administered RA (10 mg/kg) in Sprague–Dawley rats without or with verapamil (25 or 50 mg/kg) were investigated. Additionally, MDCKII-MDR1 and Caco-2 cell monolayers, five recombinant human cytochrome P450 (rhCYP) enzymes (1A2, 2C8, 2C9, 2D6 and 3A4), and rat liver microsomes were also conducted to investigate its potential mechanism. Results Verapamil could significantly affect the plasma concentration of RA. Co-administered verapamil at 25 and 50 mg/kg, the AUC0–∞ increased from 432 ± 64.2 to 539 ± 53.6 and 836 ± 116 ng × h/mL, respectively, and the oral clearance decreased from 23.6 ± 3.50 to 18.7 ± 1.85 and 12.2 ± 1.85 L/h/kg, respectively. The MDCKII-MDR1 cell assay showed that RA might be a P-gp substrate. The rhCYPs experiments indicated that RA was mainly metabolized by CYP3A4. Additionally, verapamil could increase the absorption of RA by inhibiting the activity of P-gp, and slow down the intrinsic clearance of RA from 48.5 ± 3.18 to 12.0 ± 1.06 µL/min/mg protein. Discussion and conclusions These findings indicated that verapamil could significantly affect the pharmacokinetic profiles of RA in rats. It was demonstrated that P-gp and CYP3A were involved in the transport and metabolism of RA, which might contribute to the low oral bioavailability of RA.

Abstract Cracking caused by shrinkage deformation of cement-based materials at early age is a major problem leading to material failure in restrained conditions. Carbon nanotubes (CNTs) are incorporated into cement-based materials, and the autogenous shrinkage and crack resistance of the new composite materials obtained by linear shrinkage and ring tests are studied to solve the destruction of the materials caused by the shrinkage of cement-based materials. The results showed that addition of CNTs significantly inhibited the autogenous shrinkage of cement-based materials with maximum reduction rate above 40%. CNTs also significantly improved the cracking resistance of cement-based materials. The optimal effect was noticed at CNTs content of 0.1 wt%. The incorporation of CNTs not only inhibits the autogenous shrinkage of cement-based materials, but also inhibits the drying shrinkage of cement-based materials to some extent. Therefore, carbon nanotubes have the potential to solve the destruction of materials caused by shrinkage of cement-based materials.

Body size is sensitive to environmental changes and one of the fundamental traits linking ecological functions. Size structure has been suggested as a useful indicator for environmental monitoring and assessment in aquatic ecosystems. However, the organisms’ size structure and the relationship with environmental factors remain seldom addressed in reservoir ecosystems. In this study, we investigated the size spectrum, size diversity of the zooplankton and their relationships with environmental conditions across nitrogen and phosphorus gradients in the Xiangxi Bay of Three Gorges Reservoir, China. We further tested the hypotheses that how nutrient and water temperature affect zooplankton size structure: nutrients indirectly affect zooplankton size spectrum and size diversity via phytoplankton (H1); increasing water temperature will reduce size diversity and result in a steeper size spectrum (H2); size diversity is a more robust metric indicating environment changes than the size spectrum in high dynamic ecosystems (H3). We found that both the size spectrum and size diversity showed high spatiotemporal dynamics. The size spectrum ranged from −3.373 to −0.984. The size diversity ranged from 0.631 to 3.291. Spatially, the lowest values of the size spectrum and size diversity were observed in the upstream areas of Xiangxi Bay, where nitrogen and phosphorus concentrations are high and low, respectively. And in temporal dynamics, lower values of the size spectrum and size diversity were generally observed in March and April. Further analyses based on the structural equation model (SEM) found a clear pathway revealing that nutrient variables affect the zooplankton abundance and size structure, supporting hypothesis H1. That is, dissolved inorganic nitrogen had an indirect effect on the zooplankton abundance, size spectrum, and size diversity by influencing the concentration of phytoplankton chlorophyll a. In addition, results of SEM suggested that increased water temperature had a significant negative effect on the size diversity but had non-significant effects on zooplankton abundance and size spectrum. This finding suggests that size diversity is a reliable and useful index in measuring the zooplankton size structure in reservoir ecosystems with high dynamics, which may have a wide application in environmental monitoring and assessment, especially for complex and dynamic aquatic ecosystems.

Attractin (ATRN) is a widely expressed glycoprotein that is involved in energy homeostasis, neurodevelopment, and immune response. It is encoded by a gene spanning 180 kb on chromosome 20p13, a region previously implicated in schizophrenia by linkage studies. To address a possible role of ATRN in disorders of the central nervous system, we created an atrn knockout zebrafish line and performed behavioral tests. Adult atrn–/– zebrafish exhibited more pronounced attack behavior relative to wild-type control zebrafish in a tracking analysis. Biochemical analysis revealed elevated testosterone levels in atrn–/– zebrafish. At the gene expression level, we noted an upregulation of cyp51 and hsd17b7, key proteins in testosterone synthesis in the brains of both adult and larvae of atrn–/– zebrafish. In order to further elucidate the relationship between testosterone and behavioral syndromes, we then compared testosterone levels of 9,008 psychiatric patients and 247 healthy controls from the same catchment area. Of all subjects examined, male subjects with schizophrenia exhibited lower testosterone levels compared with controls. In contrast, female subjects with a diagnosis of schizophrenia or bipolar disorder featured higher testosterone levels than did same sex controls. Purposeful sampling of extreme groups showed reduced ATRN expression in a subset of these subjects. Finally, we identified 14 subjects with ATRN mutations. All of whom displayed abnormal testosterone levels. In summary, the interplay of ATRN and testosterone may help to explain sexual dimorphisms in selected behavioral phenotypes.

Summary: Germline alterations of the NF2 gene cause neurofibromatosis type 2, a syndrome manifested with benign tumors, and Nf2 deletion in mice also results in slow tumorigenesis. As a regulator of the Hippo signaling pathway, NF2 induces LATS1/2 kinases and consequently represses YAP/TAZ. YAP/TAZ oncoproteins are also inhibited by motin family proteins (Motins). Here, we show that the Hippo signaling is fine-tuned by Motins in a NF2-dependent manner, in which NF2 recruits E3 ligase RNF146 to facilitate ubiquitination and subsequent degradation of Motins. In the absence of NF2, Motins robustly accumulate to restrict full activation of YAP/TAZ and prevent rapid tumorigenesis. Hence, NF2 deficiency not only activates YAP/TAZ by inhibiting LATS1/2 but also stabilizes Motins to keep YAP/TAZ activity in check. The upregulation of Motins upon NF2 deletion serves as a strategy for avoiding uncontrolled perturbation of the Hippo signaling and may contribute to the benign nature of most NF2-mutated tumors.

BackgroundOsteoporosis is a common complication of acute fracture, which can lead to fracture delayed union or other complications and resulting in poor fracture healing. Bisphosphate is a common anti-osteoporosis drug, but its application in fracture patients is still controversial because of its inhibitory effect on bone resorption.MethodStudies were acquired from literature databases in accordance with established inclusion criteria. Standard mean difference (SMD) and 95% confidence intervals (Cls) were calculated to evaluate the effectiveness of the bisphosphonates treatment in fracture patients. Data analysis was conducted with the Review Manager 5.4.1 software.ResultsA total of 16 studies involving 5022 patients obtained from selected databases were examined. As expected, bisphosphate had no significant effect on fracture healing time, but it could significantly increase BMD and prevent osteoporosis. Meanwhile, bisphosphate can inhibit both bone resorption and bone formation markers, resulting in low bone turnover state.ConclusionThis meta-analysis showed that bisphosphonate have no significant effect on fracture healing time but they do increase the changes in BMD and reduce bone synthesis and resorption markers. Early application of bisphosphonates after injury in the appropriate patient population should be considered.

Abstract Objectives We aimed to evaluate the global scientific output of research of acupuncture on insomnia and explore the hotspots and frontiers from 1999 to 2018, by using bibliometric methods. Methods Articles about acupuncture therapy on insomnia were retrieved from the Web of Science Core Collection (WoSCC). We used Citespace V to analyze publication years, journals, countries, institutions, authors and cited authors. We plotted the reference co-citation network and key words to analyze the research hotspots and trends. Results Until August 31, 2018 31, 2018, a total of 292 records in acupuncture therapy on insomnia research were identified from 1999 to 2018. The number and rate of the annual publication gradually increased. Respectively, SLEEP and J NEUROPSYCH CLIN N (journal of neuropsychiatry and clinical neurosciences) ranked the first in the frequency and centality of cited joural. Among countries, China ranked highest in the number of publications and the top 3 institutes were in Hong Kong. Chung KF and Yeung WF were the most productive authors and YEUNG WF ranked the first in the cited authors. In the ranking of frequency and in cited reference, the first was the article published in by CAO HJ and KALAVAPALLI R. ‘Randomized controlled trial’ had a high frequncy and centrality in keyword. Conclusions A higher degree of acceptance acupuncture was obtained in the Asian. Recently, systematic reviews and clinic trials most focused on electrocacupuncture and acupressure among the acupuncture therapy.

Helminth-derived molecules have the ability to modulate the host immune system. Our previous study identified a tetradecapeptide derived from Trichinella spiralis paramyosin (Ts-pmy) that could bind to human complement component C9 to inhibit its polymerization, making the peptide a candidate therapeutic agent for complement-related immune disorders. Here, the peptide underwent an N-terminal modification with a membrane-targeting signal (a unique myristoylated peptide) to improve its therapeutic efficacy. We found that the modified peptide had a binding affinity to human C9 that was similar to that of the original peptide, as confirmed by microscale thermophoresis assays. The binding of the modified peptide to human C9 resulted in the inhibition of C9-related complement activation, as reflected by the decreased Zn2+-induced C9 polymerization and the decreased C9-dependent lysis of rabbit erythrocytes. In addition, the original and modified peptides could both bind to recombinant mouse C9 and inhibit the C9-dependent lysis of rabbit erythrocytes in normal mouse serum (NMS), which meant that the peptides could cross the species barrier to inhibit complement activity in mice. Further in vitro and in vivo analyses confirmed that the peptide modification increased the retention time of the peptide. Furthermore, intraarticular injection of the modified peptide markedly ameliorated knee swelling and joint damage in mice with antigen-induced arthritis (AIA), as assessed histologically. These results suggested that the Ts-pmy-derived peptide modified with a membrane-targeting signal was a reasonable candidate therapeutic agent for membrane attack complex (MAC)-related diseases [such as rheumatoid arthritis (RA)] and the study presented a new modification method to improve the potential therapeutic effects of the peptide.

Mutations in the enzyme isocitrate dehydrogenase 1/2 (IDH1/2) are the most common somatic mutations in low-grade glioma (LGG). The Hippo signaling pathway is known to play a key role in organ size control, and its dysregulation is involved in the development of diverse cancers. Large tumor suppressor 1/2 (LATS1/2) are core Hippo pathway components that phosphorylate and inactivate Yes-associated protein (YAP), a transcriptional co-activator that regulates expression of genes involved in tumorigenesis. A recent report from The Cancer Genome Atlas (TCGA) has highlighted a frequent hypermethylation of LATS2 in IDH-mutant LGG. However, it is unclear if LATS2 hypermethylation is associated with YAP activation and prognosis of LGG patients. Here, we performed a network analysis of the status of the Hippo pathway in IDH-mutant LGG samples and determined its association with cancer prognosis. Combining TCGA data with our biochemical assays, we found hypermethylation of LATS2 promoter in IDH-mutant LGG. LATS2 hypermethylation, however, did not translate into YAP activation but highly correlated with IDH mutation. LATS2 hypermethylation may thus serve as an alternative for IDH mutation in diagnosis and a favorable prognostic factor for LGG patients.

Background: Schizophrenia, regarded as a neurodevelopmental disorder, is characterized by positive symptoms, negative symptoms, and cognitive dysfunction. Investigating the spontaneous brain activity in patients with schizophrenia can help us understand the underlying pathophysiologic mechanism of schizophrenia. However, results concerning abnormal neural activities and their correlations with cognitive dysfunction/psychopathology of patients with schizophrenia were inconsistent.Methods: We recruited 57 first-diagnosed and drug-naive patients with schizophrenia and 50 matched healthy controls underwent magnetic resonance imaging. The Positive and Negative Syndrome Scale (PANSS) and the MATRICS Consensus Cognitive Battery were used to assess the psychopathology/cognitive dysfunction. Regional homogeneity (ReHo) was used to explore neural activities. Correlation analyses were calculated between abnormal ReHo values and PANSS scores/standardized cognitive scores. Lastly, support vector machine analyses were conducted to evaluate the accuracy of abnormal ReHo values in distinguishing patients with schizophrenia from healthy controls.Results: Patients with schizophrenia showed cognitive dysfunction, and increased ReHo values in the right gyrus rectus, right inferior frontal gyrus/insula and left inferior frontal gyrus/insula compared with those of healthy controls. The ReHo values in the right inferior frontal gyrus/insula were positively correlated with negative symptom scores and negatively correlated with Hopkins verbal learning test-revised/verbal learning. Our results showed that the combination of increased ReHo values in the left inferior frontal gyrus/insula and right gyrus rectus had 78.5% (84/107) accuracy, 85.96% (49/57) sensitivity, and 70.00% specificity, which were higher than other combinations.Conclusions: Hyperactivities were primarily located in the prefrontal regions, and increased ReHo values in the right inferior frontal gyrus/insula might reflect the severity of negative symptoms and verbal learning abilities. The combined increases of ReHo values in these regions might be an underlying biomarker in differentiating patients with schizophrenia from healthy controls.

Multiepitope peptide vaccine has some advantages over traditional recombinant protein vaccine due to its easy and fast production and possible inclusion of multiple protective epitopes of pathogens. However, it is usually poorly immunogenic and needs to conjugate to a large carrier protein. Peptides conjugated to a central lysine core to form multiple antigen peptides (MAPs) will increase the immunogenicity of peptide vaccine. In this study, we constructed a MAP consisting of CD4+ T cell and B cell epitopes of paramyosin (Pmy) of Trichinella spiralis (Ts-Pmy), which has been proved to be a good vaccine candidate in our previous work. The immunogenicity and induced protective immunity of MAP against Trichinella spiralis (T. spiralis) infection were evaluated in mice. We demonstrated that mice immunized with MAP containing CD4+ T cell and B cell epitopes (MAP-TB) induced significantly higher protection against the challenge of T. spiralis larvae (35.5% muscle larva reduction) compared to the MAP containing B cell epitope alone (MAP-B) with a 12.4% muscle larva reduction. The better protection induced by immunization of MAP-TB was correlated with boosted antibody titers (both IgG1 and IgG2a) and mixed Th1/Th2 cytokine production secreted by the splenocytes of immunized mice. Further flow cytometry analysis of lymphocytes in spleens and draining lymph nodes demonstrated that mice immunized with MAP-TB specifically enhanced the generation of T follicular helper (Tfh) cells and germinal center (GC) B cells, while inhibiting follicular regulatory CD4+ T (Tfr) cells and regulatory T (Treg) cells. Immunofluorescence staining of spleen sections also confirmed that MAP-TB vaccination enhanced the formation of GCs. Our results suggest that CD4+ T cell epitope of Ts-Pmy is crucial in vaccine component for inducing better protection against T. spiralis infection.

Abstract Background To assess the association of different gay-related stressful events (GRSEs) with depressive symptoms in Chinese men who have sex with men (MSM). Method A total of 807 MSM were recruited using respondent-driven sampling from four cities in northeastern China. GRSEs were measured using the Gay Related Stressful Life Events Scale, and depressive symptoms were assessed using the Self-Rating Depression Scale (SDS). Results A total of 26.0% of study participants experienced GRSEs in the past three months, and the average SDS score was lower than the previously reported national average for China. The study participants had significantly elevated risks of depression (SDS score ≥ 53) due to recent troubles with a boss (OR = 4.92, 95% CI = 1.87–12.97) or a workmate (OR = 3.68, 95% CI = 1.52–8.88), loss of a close friend (OR = 2.41, 95% CI = 1.39–4.18), argument with a close friend (OR = 2.07, 95% CI = 1.33–3.22), and being physically assaulted (OR = 2.08, 95% CI = 0.98–4.43). Arguments with family members or classmates had no significant effect on depression. Multiple logistic regression analysis showed that the number of GRSEs, a lower level of education, more advanced age, and HIV infection significantly increased the risk of depression. Conclusions There are large differences in the associations of different types of GRSEs with depressive symptoms. Reducing the stigmatization and discrimination toward MSM in all social environments and improving the capability of MSM to cope with different types of GRSEs may improve their emotional wellbeing.

Abstract Although human disturbance indirectly influences lotic assemblages through modifying physical and chemical conditions, identifying thresholds of human disturbance would provide direct evidence for preventing anthropogenic degradation of biological conditions. In the present study, we used data obtained from tributaries of the Three Gorges Reservoir in China to detect effects of human disturbance on streams and to identify disturbance thresholds for benthic diatoms. Diatom species composition was significantly affected by three in-stream stressors including TP, TN and pH. Diatoms were also influenced by watershed % farmland and natural environmental variables. Considering three in-stream stressors, TP was positively influenced by % farmland and % impervious surface area (ISA). In contrast, TN and pH were principally affected by natural environmental variables. Among measured natural environmental variables, average annual air temperature, average annual precipitation, and topsoil % CaCO3, % gravel, and total exchangeable bases had significant effects on study streams. When effects of natural variables were accounted for, substantial compositional changes in diatoms occurred when farmland or ISA land use exceeded 25% or 0.3%, respectively. Our study demonstrated the rationale for identifying thresholds of human disturbance for lotic assemblages and addressed the importance of accounting for effects of natural factors for accurate disturbance thresholds.

Background: Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal human cancers. Aberrant expression of genes plays important role in the procession of PDAC. The analysis of gene expression profile will contribute to the research of carcinoma mechanism. Objective: This present study is focused to investigate the differentially expressed genes (DEGs) from 3 PDAC microarray datasets, which would provide candidate genes for putative biomarkers to understand the mechanism of PDAC and potential targets of treatment. Method: Based on the overlap genes obtained from 3 GEO datasets, the hub genes were identified using STRING and Cytoscape plugin MCODE. The enrichment and function analysis were applied using DAVID. The protein-protein interaction network was performed using cBioPortal and UCSC Xena. The Oncomine was finally used to determine the candidate gene by analyzing their expression between pancreas sample and PDAC sample. Results: 25 hub genes were selected from a total of 1006 DEGs from 3 GEO datasets, consisting of 14 upregulated genes and 11 downregulated genes. The overall decline of hub gene expression enriched in G1 phase of cell cycle in other subtypes of pancreatic cancer. Oncomine database was ultimately performed to determine the 8 candidate genes, including CXCL5, CCL20, NMU, F2R, ANXA1, EDNRA, LPAR6, and GNA15. Conclusions: Conclusively, 8 candidate genes would become the potential PDAC combined biomarkers for diagnosis and therapeutic strategies.

Talaromyces leycettanus JCM12802 is a great producer of thermophilic glycoside hydrolases (GHs). In this study, two cellulases (TlCel5A and TlCel6A) belonging to GH5 and GH6 respectively were expressed in Pichia pastoris and functionally characterized. The enzymes had acidic and thermophilic properties, showing optimal activities at pH 3.5-4.5 and 75-80°C, and retained stable at temperatures up to 60°C and over a broad pH range of 2.0-8.0. TlCel5A and TlCel6A acted against several cellulose substrates with varied activities (3,101.1 vs. 92.9 U/mg to barley β-glucan, 3,905.6 U/mg vs. 109.0 U/mg to lichenan, and 840.3 and 0.09 U/mg to CMC-Na). When using Avicel, phosphoric acid swollen cellulose (PASC) or steam-exploded corn straw (SECS) as the substrate, combination of TlCel5A and TlCel6A showed significant synergistic action, releasing more reduced sugars (1.08-2.87 mM) than the individual enzymes. These two cellulases may represent potential enzyme additives for the efficient biomass conversion and bioethanol production.

Abstract Background Dendritic cells (DCs) are antigen-presenting cells that regulate T cell responses for many infectious diseases. The tissue-dwelling nematode Trichinella spiralis expresses paramyosin (TsPmy) not only as a structural protein but also as an immunomodulator to alleviate complement attack by binding to some host complement components. Whether TsPmy is involved in other immunomodulatory pathway and how TsPmy interacts with host DCs is still unknown. Methods Mouse bone marrow-derived DCs were incubated with recombinant TsPmy (rTsPmy) for activation. Maturation of DC was determined by the expression of surface markers CD40, CD80, CD86 and MHCII. The rTsPmy-pulsed DCs were co-incubated with T. spiralis-sensitized or naïve mouse CD4+ T cells to observe their activation on T cells and polarizing regulatory T cells using flow cytometry. Cytokines were measured by enzyme-linked immunosorbent assays (ELISA). Results TsPmy was able to activate mouse bone marrow-derived DCs to semi-mature status characterized by expressing surface CD40 and CD86, but not CD80 and MHCII. The semi-mature TsPmy-pulsed DCs were able to stimulate T. spiralis-sensitized CD4+ T cells to proliferate. Incubation of TsPmy-pulsed DCs with naïve CD4+ splenocytes polarized the latter to CD4+CD25+Foxp3+ regulatory T cells. However, mice immunized with rTsPmy only induce the CD4+CD25−Foxp3+ T cell population, associated with high level of IL-10, TGF-β and IL-17A. Conclusions During T. spiralis infection, TsPmy plays an important role in modulating the host immune system by stimulating DCs to differentiate the CD4+ T cells to regulatory T cells, in addition to binding to components of the host complement cascade, as survival strategies to live in host.

Deoxyglycychloxazol (TY501) is a glycyrrhetinic acid derivative which exhibits high anti-inflammatory activity and reduced pseudoaldosteronism compared to glycyrrhetinic acid. In this study, a sensitive and rapid liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was established for the quantitation of TY501 in rat plasma. Plasma samples were treated by precipitating protein with methanol and supernatants were separated by a Symmetry C8 column with the mobile phase consisting of methanol and 10 mM ammonium formate (containing 0.1% of formic acid) (90:10, v/v). The selected reaction monitoring (SRM) transitions were performed at m/z 647.4→191.2 for TY501 and m/z 473.3→143.3 for astragaloside aglycone (IS) in the positive ion mode with atmospheric pressure chemical ionization (APCI) source. Calibration curve was linear over the concentration range of 5–5000 ng/mL. The lower limit of quantification was 5 ng/mL. The mean recovery was over 88%. The intra- and inter-day precisions were lower than 6.0% and 12.8%, respectively, and the accuracy was within ±1.3%. TY501 was stable under usual storage conditions and handling procedure. The validated method has been successfully applied to a pharmacokinetic study after oral administration of TY501 to rats at a dosage of 10 mg/kg.

Phenazines are secondary metabolites with broad spectrum antibiotic activity and thus show high potential in biological control of pathogens. In this study, we identified phenazine biosynthesis (phz) genes in two genome-completed plant pathogenic bacteria Pseudomonas syringae pv. tomato (Pst) DC3000 and Xanthomonas oryzae pv. oryzae (Xoo) PXO99A. Unlike the phz genes in typical phenazine-producing pseudomonads, phz homologs in Pst DC3000 and Xoo PXO99A consisted of phzC/D/E/F/G and phzC/E1/E2/F/G, respectively, and the both were not organized into an operon. Detection experiments demonstrated that phenazine-1-carboxylic acid (PCA) of Pst DC3000 accumulated to 13.4 μg L−1, while that of Xoo PXO99A was almost undetectable. Moreover, Pst DC3000 was resistant to 1 mg mL−1 PCA, while Xoo PXO99A was sensitive to 50 μg mL−1 PCA. Furthermore, mutation of phzF blocked the PCA production and significantly reduced the pathogenicity of Pst DC3000 in tomato, while the complementary strains restored these phenotypes. These results revealed that Pst DC3000 produces low level of and is resistant to phenazines and thus is unable to be biologically controlled by phenazines. Additionally, phz-mediated PCA production is required for full pathogenicity of Pst DC3000. To our knowledge, this is the first report of PCA production and its function in pathogenicity of a plant pathogenic P. syringae strain.

Protein kinase CK2 is a crucial regulator of endothelial cell proliferation, migration and sprouting during angiogenesis. However, it is still unknown whether this kinase additionally affects the angiogenic activity of other vessel-associated cells. In this study, we investigated the effect of CK2 inhibition on primary human pericytes. We found that CK2 inhibition reduces the expression of nerve/glial antigen (NG)2, a crucial factor which is involved in angiogenic processes. Reporter gene assays revealed a 114 bp transcriptional active region of the human NG2 promoter, whose activity was decreased after CK2 inhibition. Functional analyses demonstrated that the pharmacological inhibition of CK2 by CX-4945 suppresses pericyte proliferation, migration, spheroid sprouting and the stabilization of endothelial tubes. Moreover, aortic rings of NG2−/− mice showed a significantly reduced vascular sprouting when compared to rings of NG2+/+ mice, indicating that NG2 is an important regulator of the angiogenic activity of pericytes. In vivo, implanted Matrigel plugs containing CX-4945-treated pericytes exhibited a lower microvessel density when compared to controls. These findings demonstrate that CK2 regulates the angiogenic activity of pericytes through NG2 gene expression. Hence, the inhibition of CK2 represents a promising anti-angiogenic strategy, because it does not only target endothelial cells, but also vessel-associated pericytes.

Trichinellosis is a serious food-borne parasitic zoonosis worldwide. In the effort to develop vaccine against Trichinella infection, we have identified Trichinella spiralis Heat shock protein 70 (Ts-Hsp70) elicits partial protective immunity against T. spiralis infection via activating dendritic cells (DCs) in our previous study. This study aims to investigate whether DCs were activated by Ts-Hsp70 through TLR2 and/or TLR4 pathways.After blocking with anti-TLR2 and TLR4 antibodies, the binding of Ts-Hsp70 to DCs was significantly reduced. The reduced binding effects were also found in TLR2 and TLR4 knockout (TLR2-/- and TLR4-/-) DCs. The expression of TLR2 and TLR4 on DCs was upregulated after treatment with Ts-Hsp70 in vitro. These results suggest that Ts-Hsp70 is able to directly bind to TLR2 and TLR4 on the surface of mouse bone morrow-derived DCs. In addition, the expression of the co-stimulatory molecules (CD80, CD83) on Ts-Hsp70-induced DCs was reduced in TLR2-/- and TLR4-/- mice. More evidence showed that Ts-Hsp70 reduced its activation on TLR2/4 knockout DCs to subsequently activate the naïve T-cells. Furthermore, Ts-Hsp70 elicited protective immunity against T. spiralis infection was reduced in TLR2-/- and TLR4-/- mice correlating with the reduced humoral and cellular immune responses.This study demonstrates that Ts-Hsp70 activates DCs through TLR2 and TLR4, and TLR2 and TLR4 play important roles in Ts-Hsp70-induced DCs activation and immune responses.

Deleted in Liver Cancer-1 (DLC1), a member of the RhoGAP family of proteins, functions as a tumor suppressor in several cancers including breast cancer. However, its clinical relevance is unclear in breast cancer. In this study, expression of DLC1 was correlated with prognosis using publicly available breast cancer gene expression datasets and quantitative Reverse Transcription PCR in cohorts of Estrogen Receptor-positive (ER+) breast cancer. Low expression of DLC1 correlates with poor prognosis in patients with ER+ breast cancer with further decrease in metastatic lesions. The Cancer Genome Atlas (TCGA) data showed that down regulation of DLC1 is not due to methylation or mutations. To seek further insights in understanding the role of DLC1 in ER+ breast cancer, we stably overexpressed DLC1-full-length (DLC1-FL) in T-47D breast cancer cells; this inhibited cell colony formation significantly in vitro compared to its control counterpart. Label-free global proteomic and TiO2 phosphopeptide enrichment assays (ProteomeXchange identifier PXD008220) showed that 205 and 122 phosphopeptides were unique to DLC1-FL cells and T-47D-control cells, respectively, whereas 6,726 were quantified by phosphoproteomics analysis in both conditions. The top three significant clusters of differentially phosphopeptides identified by DAVID pathway analysis represent cell-cell adhesion, mRNA processing and splicing, and transcription regulation. Phosphoproteomics analysis documented an inverse relation between DLC1 expression and several phosphopeptides including epithelial cell transforming sequence 2 (ECT2). Decreased phosphorylation of ECT2 at the residue T359, critical for its active conformational change, was validated by western blot. In addition, the ECT2 T359-containing phosphopeptide was detected in both basal and luminal patient-derived breast cancers breast cancer phosphoproteomics data on the Clinical Proteomic Tumor Analysis Consortium (CPTAC) Assay portal. Together, for the first time, this implicates ECT2 phosphorylation in breast cancer, which has been proposed as a therapeutic target in lung cancer. In conclusion, this data suggests that low expression of DLC1 is associated with poor prognosis. Targeting ECT2 phosphopeptides could provide a promising mechanism for controlling poor prognosis seen in DLC1low ER+ breast cancer.

Trichinellosis is a worldwide zoonosis and remains a serious public health problem. Interrupting parasite transmission via vaccination of livestocks with a potent vaccine is a practical approach to prevent human Trichinellosis. Our previous studies have identified that paramyosin of Trichinella spiralis (Ts-Pmy) is a good vaccine candidate against Trichinellosis. In this study, a novel multi-epitope vaccine (MEP) was constructed by using four CD4+ T cell epitopes (P2, P3, P4, and P5) and one B cell epitope (YX1) from Ts-Pmy and expressed as a soluble recombinant protein (rMEP) in Escherichia coli. Mice immunized with rMEP vaccine produced significant higher muscle larval reduction (55.4%) than that induced by immunization of parental rTs-Pmy (34.4%) against T. spiralis infection. The better protection is associated with rMEP induced high levels of anti-rMEP specific IgG and subclass IgG1/IgG2a, elevated T cell proliferation of splenocytes and secretion of IFN-γ, IL-4 and IL-5. The cellular response to individual T cell epitope also showed that splenocytes from mice immunized with rMEP strongly response to the stimulation of synthetic epitope peptide P2, P3, and P4, but not to P5, suggesting that most of T cell epitopes are exposed and processed well during immunization that may contribute to the high protection induced by the immunization of rMEP. This study implies that epitope vaccine is a promising approach for the development of vaccines against Trichinellosis.

Abstract Background Trichinella spiralis expresses paramyosin (Ts-Pmy) not only as a structural protein but also as an immunomodulator that inhibits host complement as a survival strategy. Previous studies demonstrated that Ts-Pmy bound to complement components C8 and C9 and inhibited the polymerization of C9 during the formation of the membrane attack complex (MAC). The C9 binding domain of Ts-Pmy was identified within 14 amino acid residues at the C-terminus of Ts-Pmy. The production of a monoclonal antibody that specifically targets the C9 binding site is necessary for further studies of Ts-Pmy function and may be used as a therapeutic agent for T. spiralis infection. Methods In this study, a monoclonal antibody against the complement C9 binding domain of Ts-Pmy (mAb 9G3) was produced using hybridoma technology. The binding activity of the mAb produced for recombinant or native Ts-Pmy and the blockade of Ts-Pmy binding to C9 by the mAb were assessed by Western blot analysis. The effect of the mAb on the viability of T. spiralis was observed by co-incubation of T. spiralis with mAb 9G3 in the presence of complement in vitro and by passive transfer of the mAb into naive mice following T. spiralis larval challenge. Results mAb 9G3 was successfully produced against the C9 binding domain of Ts-Pmy and bound specifically not only to recombinant Ts-Pmy but also to native Ts-Pmy expressed in different stages of T. spiralis, including adult worms, newborn larvae and muscle larvae. The binding of mAb 9G3 to Ts-Pmy efficiently blocked the binding of Ts-Pmy to human complement C9, resulting in a significant increase in the complement-mediated killing of newborn larvae in vitro and reduced infectivity of T. spiralis larvae in mice passively transferred with the mAb. Conclusions mAb 9G3 is a specific antibody that binds to the C9 binding domain of Ts-Pmy and interferes with Ts-Pmy’s complement-binding activity. Therefore, this mAb is a protective antibody that has potential as a preventive and therapeutic agent for T. spiralis infection.

Abstract Background Trichinellosis is an important foodborne zoonosis that is distributed worldwide. Trichinella spiralis may evade host complement-mediated attack by expressing complement inhibitory proteins, such as paramyosin (Pmy). Previous studies have shown that Trichinella spiralis paramyosin (Ts-Pmy) is able to bind to the human complement component C9 to inhibit the complement activation and protect the parasite from complement-mediated attack. Further determination of the complement-binding domain on Ts-pmy will enable us to better understand the Ts-Pmy’s biofunction in the immune evasion and provide feasible approach to develop epitope-based subunit vaccine against trichinellosis. Methods The complement C9 binding region on Ts-Pmy was determined by expression of overlapped fragments of Ts-Pmy and their binding activities to C9. The exact binding site was further narrowed-down to a 14-amino acid peptide at C-terminus using synthesized peptides with different size of amino acid sequence. The C9 complement-binding of the 14-amino acid peptide and its interference in the C9 polymerization and the complement-mediated lysis of rabbit erythrocytes was investigated. Results The protein interaction between human C9 and native Ts-Pmy was further confirmed by immunoprecipitation with T. spiralis lysates. The fragmental expression and C9 binding assays identified that the binding region of Ts-Pmy to C9 is located within 831–885 of Ts-Pmy C-terminus. The exact binding site on Ts-Pmy to C9 was narrowed down to 14 amino acid residues (866Val-879Met) by using different sizes of synthesized peptides. In the presence of the synthesized 14-amino acid peptide, human C9 polymerization and the hemolytic activity of the human complement was inhibited. Conclusions Our results revealed the precise molecular basis for T. spiralis to produce Ts-Pmy as an immunomodulator to evade the attack of the host complement system as a survival mechanism.

BACKGROUND:Our previous studies showed that Trichinella spiralis paramyosin (TsPmy) is an immunomodulatory protein that inhibits complement C1q and C8/C9 to evade host complement attack. Vaccination with recombinant TsPmy protein induced protective immunity against T. spiralis larval challenge. Due to the difficulty in producing TsPmy as a soluble recombinant protein, we prepared a DNA vaccine as an alternative approach in order to elicit a robust immunity against Trichinella infection. METHODS AND FINDINGS:The full-length TsPmy coding DNA was cloned into the eukaryotic expression plasmid pVAX1, and the recombinant pVAX1/TsPmy was transformed into attenuated Salmonella typhimurium strain SL7207. Oral vaccination of mice with this attenuated Salmonella-delivered TsPmy DNA vaccine elicited a significant mucosal sIgA response in the intestine and a systemic IgG antibody response with IgG2a as the predominant subclass. Cytokine analysis also showed a significant increase in the Th1 (IFN-γ, IL-2) and Th2 (IL-4, 5, 6, 10) responses in lymphocytes from the spleen and MLNs of immunized mice upon stimulation with TsPmy protein. The expression of the homing receptors CCR9/CCR10 on antibody secreting B cells may be related to the translocation of IgA-secreted B cells to local intestinal mucosa. The mice immunized with Salmonella-delivered TsPmy DNA vaccine produced a significant 44.8% reduction in adult worm and a 46.6% reduction in muscle larvae after challenge with T. spiralis larvae. CONCLUSION:Our results demonstrated that oral vaccination with TsPmy DNA delivered by live attenuated S. typhimurium elicited a significant local IgA response and a mixed Th1/Th2 immune response that elicited a significant protection against T. spiralis infection in mice.