Toxoplasma gondii suppresses proliferation and migration of breast cancer cells by regulating their transcriptome.

BACKGROUND: Breast cancer is the most common cancer in women worldwide. Toxoplasma gondii (T. gondii) has shown anticancer activity in breast cancer mouse models, and exerted beneficial effect on the survival of breast cancer patients, but the mechanism was unclear. METHODS: The effect of tachyzoites of T. gondii (RH and ME49 strains) on human breast cancer cells (MCF-7 and MDA-MB-231 cells) proliferation and migration was assessed using cell growth curve and wound healing assays. Dual RNA-seq was performed for T. gondii-infected and non-infected cells to determine the differentially expressed genes (DEGs). Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and Protein-Protein Interaction Networks analysis (PPI) were performed to explore the related signaling pathway and hub genes. Hub genes were validated using the Kaplan-Meier plotter database, and Pathogen Host Interaction (PHI-base) database. The results were verified by qRT-PCR. RESULTS: The tachyzoites of T. gondii decreased the expression of Ki67 and increased the expression of E-cadherin, resulting in suppressing the proliferation and migration of infected human breast cancer cells. The inhibitory effect of T. gondii on breast cancer cells showed a significant dose-response relationship. Compared with the control group, 2321 genes were transcriptionally regulated in MCF-7 cells infected with T. gondii, while 169 genes were transcriptionally regulated in infected MDA-MB-231 cells. Among these genes, 698 genes in infected MCF-7 cells and 67 genes in infected MDA-MB-231 cells were validated by the publicly available database. GO and KEGG analyses suggested that several pathways were involved in anticancer function of T. gondii, such as ribosome, interleukin-17 signaling, coronavirus disease pathway, and breast cancer pathway. BRCA1, MYC and IL-6 were identified as the top three hub genes in infected-breast cancer cells based on the connectivity of PPI analysis. In addition, after interacting with breast cancer cells, the expression of ROP16 and ROP18 in T. gondii increased, while the expression of crt, TgIST, GRA15, GRA24 and MIC13 decreased. CONCLUSIONS: T. gondii transcriptionally regulates several signaling pathways by altering the hub genes such as BRCA1, MYC and IL-6, which can inhibit the breast tumor growth and migration, hinting at a potential therapeutic strategy.

Restraint Stress, Foot Shock and Corticosterone Differentially Alter Autophagy in the Rat Hippocampus, Basolateral Amygdala and Prefrontal Cortex.

Autophagy is a conserved lysosomal degradation process that has recently been found to be associated with stress-related psychological diseases. However, previous studies have yielded inconsistent results regarding the effects of various stress patterns on autophagy in different brain regions. This discrepancy may arise from differences in autophagy flux across nuclei, the type of stress experienced, and the timing of autophagy assessment after stress exposure. In this study, we assessed autophagy flux in the rat hippocampus (HPC), medial prefrontal cortex (mPFC), and basal lateral amygdala (BLA) by quantifying protein levels of p-ULK1, LC3-I, LC3-II, and p62 via Western blot analysis at 15 min, 30 min, and 60 min following various stress paradigms: restraint stress, foot shock, single corticosterone injection, and chronic corticosterone treatment. We found that: (1) hippocampal autophagy decreased within 1 h of restraint stress, foot shock, and corticosterone injection, except for a transient increase at 30 min after restraint stress; (2) autophagy increased 1 h after restraint stress and corticosterone injection but decreased 1 h after foot shock in mPFC; (3) In BLA, autophagy increased 1 h after foot shock and corticosterone injection but decreased 1 h after restraint stress; (4) Chronic corticosterone increased autophagy in mPFC and BLA but had no effects in HPC. These findings suggest that stress regulates autophagy in a brain region- and stressor-specific manner within 1 h after stress exposure, which may contribute to the development of stress-related psychological disorders.

Identification of Cofilin-1 as a novel biomarker of atopic dermatitis using iTRAQ quantitative proteomics.

BACKGROUND: Atopic dermatitis (AD) is a chronic relapsing inflammatory skin condition; however, little is known about the pathogenesis and serum biomarker of this disease. METHODS: Isobaric tagging for relative and absolute quantitation (iTRAQ) proteomic assay was adopted to identify and quantify the differentially expressed proteins (DEPs) in the serum of AD patients. Bioinformatic analysis, including GO, Reactome, GSEA, PPI, and ssGSEA analysis, were used to identified the enriched pathways, hub proteins and immune cells. The expression level and distribution of hub proteins were confirmed by ELISA and IHC. RESULTS: Sixty-six DEPs were identified with iTRAQ proteomic assay by analyzing serum from AD patients and normal subjects. GO and Reactome analysis shown the alternated pathway were mainly involved in immunity, oxidative stress, and actin cytoskeleton. The GSEA and PPI network analysis among the DEPs were carried out and identified Cofilin-1 and profilin-1 as the core components of this network. Additionally, the disruption of Th1/Th2/Th17 cell balance and the significantly reducing of Treg, MDSC, and γδT cells was also found in AD patients using the ssGSEA analysis. Further ELISA and IHC assay validated the significantly elevated expression of Cofilin-1 in AD patients. CONCLUSION: Our results suggested that Cofilin-1 may serve as a novel biomarker for AD diagnosis.

Renal protective effects of astragaloside IV, in diabetes mellitus kidney damage animal models: A systematic review, meta-analysis.

Astragaloside IV (ASIV) is the essential active component of astragalus that has diverse biological activities. Previous research has suggested its potentially beneficial effects on diabetic nephropathies. However, its effects and protective mechanism remain unclear. In this study, we conducted a preclinical systematic review to evaluate the efficacy and potential mechanisms of ASIV in reducing kidney damage in diabetes mellitus (DM) models. Studies were searched from nine databases until January 2020. A random-effects model was used to calculate combined standardised mean difference estimates and 95 % confidence intervals. Risk of bias of studies was assessed using the Systematic Review Center for Laboratory Animal Experimentation risk of bias tool 10-item checklist. RevMan 5.3 software was used for statistical analysis. Twenty-three studies involving 562 animals were included in the meta-analysis. Studies quality scores ranged from 2 to 5. The ASIV group induced a marked decrease in serum creatinine (P < 0.00001), blood urea nitrogen (P < 0.00001), 24-h urine protein (P < 0.00001) and pathological score (P < 0.001) compared with the control group. The determined potential mechanisms of ASIV action were relieving oxidative stress, delaying renal fibrosis, anti-apoptosis and anti-inflammatory action. We conclude that ASIV exerts renal protective effects in animals with DM through multiple signalling pathways.

LncRNA MEG3 contributes to adenosine-induced cytotoxicity in hepatoma HepG2 cells by downregulated ILF3 and autophagy inhibition via regulation PI3K-AKT-mTOR and beclin-1 signaling pathway.

Adenosine is a promising cytotoxic reagent for tumors, long noncoding RNA (lncRNA) maternally expressed gene 3 (MEG3) has been indicated to play critical roles in tumorigenesis, ILF3 has been recognized as a MEG3-binding protein, however, the roles of adenosine and MEG3 on hepatoma are still ambiguous. To clarify the effects of MEG3 on the adenosine-induced cytotoxicity in hepatoma, MEG3 and ILF3 lentivirus were transduced into human hepatoma HepG2 cells to stimulate overexpression of MEG3 (OE MEG3) and overexpression of ILF3 (OE ILF3), furthermore, ILF3 small interfering RNA (siRNA) was also applied to downregulate the expression of ILF3. In this study, autophagy was markedly inhibited by low concentration of adenosine, which present by not only inhibited transformation from LC3-I to LC3-II and autophagosomes formation, but also the elevation of mTOR and reduction of beclin-1 proteins. Furthermore, low concentration of adenosine also exerted marked cytotoxicity representing induced cell apoptosis together with reductions of cell viability and migration, which were also markedly enhanced by OE MEG3. Novelly and excitingly, adenosine markedly stimulated MEG3 expression, OE MEG3 markedly decreased the ILF3 expression in HepG2 cells, and the adenosine-induced autophagy inhibition, together with the ratio of p-PI3K/PI3K, p-AKT/AKT, and p-mTOR/mTOR were also boosted by OE MEG3. More interestingly, OE ILF3 increased autophagy, whereas downregulated ILF3, especially in the case of adenosine, led to marked autophagy inhibition by decreasing beclin-1. The present study demonstrates autophagy inhibition is involved in the adenosine-induced cytotoxicity in HepG2 cells, the cytotoxicity can be synergized by OE MEG3 via downregulated ILF3 to activate PI3K/Akt/mTOR and inactivate the beclin-1 signaling pathway. In conclusion, MEG3 and inhibition of autophagy might be potential targets for augmenting adenosine-induced cytotoxicity in hepatoma.

Snapin interacts with G-protein coupled receptor PKR2.

Mutations in Prokineticin receptor 2 (PKR2), a G-protein-coupled receptor, have been identified in patients with Kallmann syndrome and/or idiopathic hypogonadotropic hypogonadism, characterized by delayed puberty and infertility. In this study, we performed yeast two-hybrid screening by using PKR2 C-terminus (amino acids 333-384) as a bait, and identified Snapin as a novel interaction partner for PKR2. The interaction of Snapin and PKR2 was confirmed in GST pull-down and co-immunoprecipitation studies. We further demonstrated that two α-helix domains in Snapin are required for the interaction. And the interactive motifs of PKR2 were mapped to YFK (343-345) and HWR (351-353), which shared a similar sequence of two aromatic amino acids followed by a basic amino acid. Disruption of Snapin-PKR2 interaction did not affect PKR2 signaling, but increased the ligand-induced degradation, implying a role of Snapin in the trafficking of PKR2.

Effects of Ellagic Acid on Glucose and Lipid Metabolism: A Systematic Review and Meta-Analysis.

Background: Abnormal glucose and lipid metabolism (GALM) serve as both a cause and an inducer for the development of the disease. Improvement and treatment of GALM are an important stage to prevent the occurrence and development of the disease. However, current clinical treatment for GALM is limited. Ellagic acid (EA), a common polyphenol present in foods, has been shown to improve abnormalities in GALM observed in patients suffering from metabolic diseases. Objective: This study used a meta-analysis method to systematically assess the effects of EA on GALM. Method: As of November 8, 2023, a comprehensive search was conducted across 5 databases, namely, PubMed, Embase, Web of Science, Cochrane Library, and Google Scholar to identify randomized controlled trials (RCTs) in which EA served as the primary intervention for diseases related to GALM. The risk of bias within the included studies was assessed according to the Cochrane Handbook. All statistical analyzes were performed using RevMan 5.4 software. Results: In this study, a total of 482 articles were retrieved, resulting in the inclusion of 10 RCTs in the meta-analysis. The results showed that EA could reduce fasting blood glucose (FBG) (p = 0.008), increase insulin secretion (p = 0.01), improve insulin resistance index (HOMA-IR) (p = 0.003), decrease triglyceride (TG) (p = 0.004), and reduce cholesterol (Chol) (p = 0.04) and low-density lipoprotein (LDL-c) (p = 0.0004). EA had no significant effect on waist circumference (WC), body weight (BW), body mass index (BMI), 2 hours after prandial blood glucose (2 h-PG), total cholesterol (TC), and high-density lipoprotein (HDL-c). Conclusions: The effect of improvement in glucose and lipids of EA was closely related to the dose and the intervention time. EA can improve GALM caused by diseases. To corroborate the findings of this study and improve the reliability of the results, EA is imperative to refine the research methodology and increase the sample size in future investigations.

Predicting the potential distribution of Dendrolimus punctatus and its host Pinus massoniana in China under climate change conditions.

Introduction: Dendrolimus punctatus, a major pest endemic to the native Pinus massoniana forests in China, displays major outbreak characteristics and causes severe destructiveness. In the context of global climate change, this study aims to investigate the effects of climatic variations on the distribution of D. punctatus and its host, P. massoniana. Methods: We predict their potential suitable distribution areas in the future, thereby offering a theoretical basis for monitoring and controlling D. punctatus, as well as conserving P. massoniana forest resources. By utilizing existing distribution data on D. punctatus and P. massoniana, coupled with relevant climatic variables, this study employs an optimized maximum entropy (MaxEnt) model for predictions. With feature combinations set as linear and product (LP) and the regularization multiplier at 0.1, the model strikes an optimal balance between complexity and accuracy. Results: The results indicate that the primary climatic factors influencing the distribution of D. punctatus and P. massoniana include the minimum temperature of the coldest month, annual temperature range, and annual precipitation. Under the influence of climate change, the distribution areas of P. massoniana and its pests exhibit a high degree of similarity, primarily concentrated in the region south of the Qinling-Huaihe line in China. In various climate scenarios, the suitable habitat areas for these two species may expand to varying degrees, exhibiting a tendency to shift toward higher latitude regions. Particularly under the high emission scenario (SSP5-8.5), D. punctatus is projected to expand northwards at the fastest rate. Discussion: By 2050, its migration direction is expected to closely align with that of P. massoniana, indicating that the pine forests will continue to be affected by the pest. These findings provide crucial empirical references for region-specific prevention of D. punctatus infestations and for the rational utilization and management of P. massoniana resources.

Serum HDL-C subfractions as predictors of cardiovascular calcification in hemodialysis patients: novel insights and clinical implications.

Objective: This study aims to explore the relationship between cardiovascular calcification (CVC) and serum levels of high-density lipoprotein cholesterol (HDL-C) and its subfractions in hemodialysis (HD) patients. Methods: HD patients and healthy participants were recruited based on specific inclusion and exclusion criteria. Various blood indicators were measured, and demographic information was recorded. HDL-C particle levels were quantified using lipophilic fluorescent dye staining and capillary electrophoresis (microfluidic platform). Coronary artery calcium scores and valve calcification were used to classify HD patients into calcification and non-calcification groups. Results: Compared to healthy participants, HD patients showed a significant increase in HDL-C, high-density lipoprotein 2 cholesterol (HDL2-C), and high-density lipoprotein 3 cholesterol (HDL3-C) levels (p < 0.001). Further division of HD patients into calcification and non-calcification groups revealed higher serum HDL3-C concentrations (p = 0.002) and a higher HDL3-C/HDL-C ratio (p = 0.04) in the calcification group. Additionally, elevated HDL3-C levels were found to be an independent risk factor for CVC in HD patients (p = 0.040). The ROC curve analysis showed an AUC value of 0.706 for HDL3-C (p = 0.002). Conclusion: Our study indicates that elevated serum HDL3-C levels in HD patients are an independent risk factor for CVC and can serve as a potential predictor for CVC events. However, more studies need to verify its potential as a predictive indicator..

DoS attacks resilience of heterogeneous complex networks via dynamic event-triggered impulsive scheme for secure quasi-synchronization.

This paper addresses the secure quasi-synchronization issue of heterogeneous complex networks (HCNs) under aperiodic denial-of-service (DoS) attacks with dynamic event-triggered impulsive scheme (ETIS). The heterogeneity of networks and the aperiodic DoS attacks, which hinder communication channels and synchronization goals, present challenges to the analysis of secure quasi-synchronization. The ETIS leverages impulsive control and dynamic event-triggered scheme (ETS) to handle the network heterogeneity and the DoS attacks. We give specific bounds on the attack duration and frequency that the network can endure, and obtain synchronization criteria that relate to event parameters, attack duration, attack frequency, and impulsive gain by the variation of parameter formula and recursive methods. Moreover, we prove that the dynamic ETS significantly reduces the controller updates, saves energy without sacrificing the system decay rate, and prevents the Zeno phenomenon. Finally, we validate our control scheme with a numerical example.

Oridonin Attenuates Diabetes­induced Renal Fibrosis via the Inhibition of TXNIP/NLRP3 and NF­κB Pathways by Activating PPARγ in Rats.

INTRODUCTION: Oridonin possesses remarkable anti-inflammatory, immunoregulatory properties. However, the renoprotective effects of oridonin and the underlying molecular mechanisms in diabetic nephropathy (DN). We hypothesized that oridonin could ameliorate diabetes­induced renal fibrosis. METHODS: Streptozocin (STZ)-induced diabetic rats were provided with a high-fat diet to establish a type 2 diabetes mellitus (T2DM) animal model, and then treated with Oridonin (10, 20 mg/kg/day) for two weeks. Kidney function and renal fibrosis were assessed. High glucose-induced human renal proximal tubule epithelial cells (HK-2) were also treated with oridonin. The expression of inflammatory factors and fibrotic markers were analyzed. RESULTS: Oridonin treatment preserved kidney function and markedly limited the renal fibrosis size in diabetic rats. The renal fibrotic markers were inhibited in the oridonin 10 mg/kg/day and 20 mg/kg/day groups compared to the T2DM group. The expression of thioredoxin-interacting proteins/ nod-like receptor protein-3 (TXNIP/NLRP3) and nuclear factor (NF)­κB pathway decreased, while that of peroxisome proliferator-activated receptor-gamma (PPARγ) increased in the oridonin treatment group compared to the non-treated group. In vitro, PPARγ intervention could significantly regulate the effect of oridonin on the high glucose-induced inflammatory changes in HK-2 cells. CONCLUSION: Oridonin reduces renal fibrosis and preserves kidney function via the inhibition of TXNIP/NLRP3 and NF­κB pathways by activating PPARγ in rat T2DM model, which indicates potential effect of oridonin in the treatment of DN.

Associations of social determinants of health with life expectancy and future health risks among individuals with type 2 diabetes: two nationwide cohort studies in the UK and USA.

BACKGROUND: Social determinants of health (SDHs) are the primary drivers of preventable health inequities, and the associations between SDHs and health outcomes among individuals with type 2 diabetes remain unclear. This study aimed to estimate the associations of combined SDHs with life expectancy and future health risks among adults with type 2 diabetes from the UK and USA. METHODS: In an analysis of two nationwide cohort studies, adults with type 2 diabetes were identified from the UK Biobank from March 13, 2006, to Oct 1, 2010 (adults aged 37-73 years) and the US National Health and Nutrition Examination Survey (NHANES) from 1999 to 2018 (adults aged ≥20 years). Participants with type 2 diabetes at baseline were included in our analysis. Participants without information on SDHs or follow-up were excluded. The UK Biobank assessed 17 SDHs and the US NHANES assessed ten SDHs, with each SDH dichotomised into advantaged and disadvantaged levels. The combined score of SDHs were calculated as the sum of the weighted scores for each SDH. Participants were then categorised into tertiles (favourable, medium, and unfavourable SDH groups). Primary outcomes were life expectancy and mortality in both cohorts, and incidences of cardiovascular disease, diabetes-related microvascular disease, dementia, and cancer in the UK Biobank. Outcomes were obtained from disease registries up until Dec 31, 2021, in the UK Biobank and Dec 31, 2019, in the US NHANES cohorts. FINDINGS: We included 17 321 participants from the UK Biobank cohort (median age 61·0 years [IQR 56·0-65·0]; 6028 [34·8%] women and 11 293 [65·2%] men) and 7885 participants from the NHANES cohort (mean age 59·2 years [95% CI 58·7-59·6]; 3835 [49·1%, weighted] women and 4050 [50·9%, weighted] men) in our analysis. In the UK Biobank, 3235 deaths (median follow-up 12·3 years [IQR 11·5-13·2]), 3010 incident cardiovascular disease (12·1 years [10·8-13·0]), 1997 diabetes-related microvascular disease (8·0 years [7·1-8·9]), 773 dementia (12·6 years [11·8-13·5]), and 2259 cancer cases (11·3 years [10·4-12·2]) were documented; and the US NHANES documented 2278 deaths during a median follow-up of 7·0 years (3·7-11·2). After multivariable adjustment, compared with the favourable SDH group, the hazard ratio was 1·33 (95% CI 1·21-1·46) in the medium SDH group and 1·89 (1·72-2·07) in the unfavourable SDH group in the UK Biobank cohort; 1·51 (1·34-1·70) in the medium SDH group and 2·02 (1·75-2·33) in the unfavourable SDH group in the US NHANES cohort for all-cause mortality; 1·13 (1·04-1·24) in the medium SDH group and 1·40 (1·27-1·53) in the unfavourable SDH group for incident cardiovascular disease; 1·13 (1·01-1·27) in the medium SDH group and 1·41 (1·26-1·59) in the unfavourable SDH group for incident diabetes-related microvascular disease; 1·35 (1·11-1·64) in the medium SDH group and 1·76 (1·46-2·13) in the unfavourable SDH group for incident dementia; and 1·02 (0·92-1·13) in the medium SDH group and 1·17 (1·05-1·30) in the unfavourable SDH group for incident cancer in the UK Biobank cohort (ptrend<0·010 for each category). At the age of 45 years, the mean life expectancy of participants was 1·6 years (0·6-2·3) shorter in the medium SDH group and 4·4 years (3·3-5·4) shorter in the unfavourable SDH group than in the favourable SDH group in the UK Biobank. In the US NHAHES cohort, the life expectancy was 1·7 years (0·6-2·7) shorter in the medium SDH group and 3·0 years (1·8-4·3) shorter in the unfavourable SDH group, compared with the favourable group. INTERPRETATION: Combined unfavourable SDHs were associated with a greater loss of life expectancy and higher risks of developing future adverse health outcomes among adults with type 2 diabetes. These associations were similar across two nationwide cohorts from varied social contexts, and were largely consistent across populations with different demographic, lifestyle, and clinical features. Thus, assessing the combined SDHs of individuals with type 2 diabetes might be a promising approach to incorporate into diabetes care to identify socially vulnerable groups and reduce disease burden. FUNDING: The National Natural Science Foundation of China, the National Key R&D Program of China, and the Fundamental Research Funds for the Central Universities.

Embryo Microinjection for Transgenesis in Drosophila.

Transgenesis in Drosophila is an essential approach to studying gene function at the organism level. Embryo microinjection is a crucial step for the construction of transgenic flies. Microinjection requires some types of equipment, including a microinjector, a micromanipulator, an inverted microscope, and a stereo microscope. Plasmids isolated with a plasmid miniprep kit are qualified for microinjection. Embryos at the pre-blastoderm or syncytial blastoderm stage, where nuclei share a common cytoplasm, are subjected to microinjection. A cell strainer eases the process of dechorionating embryos. The optimal time for dechorionation and desiccation of embryos needs to be determined experimentally. To increase the efficiency of embryo microinjection, needles prepared by a puller need to be beveled by a needle grinder. In the process of grinding needles, we utilize a foot air pump with a pressure gauge to avoid the capillary effect of the needle tip. We routinely inject 120-140 embryos for each plasmid and obtain at least one transgenic line for around 85% of plasmids. This article takes the phiC31 integrase-mediated transgenesis in Drosophila as an example and presents a detailed protocol for embryo microinjection for transgenesis in Drosophila.

Association of ultraprocessed food consumption with risk of microvascular complications among individuals with type 2 diabetes in the UK Biobank: a prospective cohort study.

BACKGROUND: The poor nutritional characteristics and potentially harmful molecules in ultraprocessed foods (UPFs) are risk factors for diabetic microvascular complications. However, the evidence regarding UPFs and diabetic microvascular complications remains limited. OBJECTIVES: We aimed to evaluate the associations between UPF consumption and risk of diabetic microvascular complications, to examine the underlying biological pathways (e.g., inflammation and lipid profile), and to identify whether the associations differ by type of UPF dietary patterns. METHODS: We included a prospective cohort of UK Biobank participants with type 2 diabetes (T2D) having at least one 24-h dietary recall (N = 5685). UPFs were defined using the Nova classification. Principal component analysis was used to derive UPF consumption patterns. Associations of UPFs and their consumption patterns with microvascular complications were assessed using Cox proportional hazards regression models. Mediation analyses were used to estimate the mediating effects of 22 biomarkers. RESULTS: During a median of 12.7 y of follow-up, 1243 composite microvascular complications events occurred (599 diabetic retinopathy, 237 diabetic neuropathy, and 662 diabetic kidney disease events). Five consumption patterns were identified (spread and bread, cereal prepared with liquids, dairy-based products, sugary beverage and snack, and mixed beverage and savory snack patterns). A 10% increment in the proportion of UPF was associated with higher hazards of the composite microvascular complications (hazard ratio [HR]: 1.08; 95% confidence interval [CI]: 1.03, 1.13) and diabetic kidney disease (HR: 1.13; 95% CI: 1.06, 1.20). Triglycerides, C-reactive protein, and body mass index collectively explained 22.0% (9.6%-43.0%) of the association between UPF intake and composite microvascular complications. Pattern high in mixed beverage and savory snack was associated with a higher risk of composite microvascular complications. CONCLUSIONS: Higher UPF consumption was associated with higher risks of diabetic microvascular complications, and the association was partly mediated through multiple potential ways.

Serum 25-hydroxyvitamin D concentrations, vitamin D receptor polymorphisms, and risk of infections among individuals with type 2 diabetes: a prospective cohort study.

BACKGROUND: Evidence on the association between serum 25-hydroxyvitamin D [25(OH)D] and infections among patients with type 2 diabetes (T2D), a group susceptible to vitamin D deficiency and infections, is limited. OBJECTIVES: We aimed to examine this association in individuals with T2D, and to evaluate whether genetic variants in vitamin D receptor (VDR) would modify this association. METHODS: This study included 19,851 participants with T2D from United Kingdom Biobank. Infections were identified by linkage to hospital inpatient and death registers. Negative binomial regression models were used to estimate incidence rate ratios (IRRs) and 95% confidence intervals (CIs), with adjustment of potential confounders. RESULTS: In patients with T2D, the incidence rate of infections was 29.3/1000 person-y. Compared with those with 25(OH)D of 50.0-74.9 nmol/L, the multivariable-adjusted IRRs and 95% CIs of total infections, pneumonia, gastrointestinal infections, and sepsis were 1.44 (1.31, 1.59), 1.49 (1.27, 1.75), 1.47 (1.22, 1.78), and 1.41 (1.14, 1.73), respectively, in patients with 25(OH)D <25.0 nmol/L. Nonlinear inverse associations between 25(OH)D concentrations and the risks of total infections (P-overall < 0.001; P-nonlinear = 0.002) and gastrointestinal infections (P-overall < 0.001; P-nonlinear = 0.040) were observed, with a threshold effect at ∼50.0 nmol/L. The vitamin D-infection association was not modified by genetic variants in VDR (all P-interaction > 0.050). CONCLUSIONS: In patients with T2D, lower serum 25(OH)D concentration (<50 nmol/L) was associated with higher risks of infections, regardless of genetic variants in VDR. Notably, nonlinear inverse associations between 25(OH)D concentrations and the risks of infections were found, with a threshold effect at ∼50.0 nmol/L. These findings highlighted the importance of maintaining adequate vitamin D in reducing the risk of infections in patients with T2D.

Functional analysis of the distal region of the third intracellular loop of PROKR2.

Mutations in the G-protein-coupled receptor PROKR2 have been identified in patients with idiopathic hypogonadotropic hypogonadism (IHH) and Kallmann syndrome (KS) manifesting with delayed puberty and infertility. Recently, the homozygous mutation V274D was identified in a man displaying KS with an apparent reversal of hypogonadism. The affected amino acid, valine 274, is located at the junction region of the third intracellular loop (IL3) and the sixth transmembrane domain (TM6). In this study, we first studied the effect of V274D and related mutations (V274A, V274T, and V274R) on the signaling activity and cell surface expression of PROKR2. Our data indicate that a charged amino acid substitution at residue 274 of PROKR2 results in low cell surface expression and loss-of-function. Furthermore, we studied the effects of two clusters of basic amino acids located at the proximal region of Val274 on the cell surface expression and function of PROKR2. The deletion of RRK (270-272) resulted in undetectable cell surface expression, whereas RKR (264-266)-deleted PROKR2 was expressed normally on the cell surface but showed loss-of-function due to a deficiency in G-protein coupling. Our data indicate that the distal region of the IL3 of PROKR2 may differentially influence receptor trafficking and G-protein coupling.

An outbreak of norovirus gastroenteritis associated with a secondary water supply system in a factory in south China.

BACKGROUND: Between September 17 and October 3, 2009, hundreds of workers employed in a manufacturing factory in Shenzhen, a city in south China developed a sudden onset of acute gastroenteritis. A retrospective cohort study is designed to identify the risk factors and control this outbreak. METHODS: Information on demographic characteristics, working place, the history of contact with a person having diarrhea and/or vomiting, drink water preference and frequency, eating in the company cafeteria or outside the company, hand-washing habits and eating habits is included. Furthermore, in order to find the contamination source, we investigated the environment around the underground reservoir and collected water samples from the junction between municipal supply water system and underground reservoir to test potential bacteria and virus, examine the seepage tracks on the wall of the underground reservoir from the side of septic tank, and check the integrity and attitude of this lid. Relative risk was presented and Chi-square test was performed. All the analyses were performed with OpenEpi software version 2.3.1 online. RESULTS: The cohort study demonstrated that the workers who had direct drink water were 3.0 fold more likely to suffer from acute gastroenteritis than those who consumed commercial bottled water. The direct drinking water, water of the tank of buildings, and the underground reservoir were positive only for norovirus. Norovirus was also detected from stool and rectal swab samples from patients with acute gastroenteritis. The underground reservoir was found to be the primary contamination source. Further environmental investigation showed that the norovirus contaminated substance entered into the underground reservoir via access holes in lid covering this underground reservoir. CONCLUSION: This acute gastroenteritis outbreak was caused by the secondary supply system contaminated by norovirus in this factory. The outbreak of gastroenteritis cases caused by norovirus frequently occurred in China due to a lack of surveillance and supervision, and due to faults in the construction of such water systems. Therefore, more attentions should pay to the secondary supply water system in China.

[Construction and expression of the Echinococcus granulosus recombinant BCG-EgG1Y162].

OBJECTIVE: To construct and express Echinococcus granulosus recombinant bacille Calmette-Guerin (BCG) strain rBCG-EgG1Y162. METHODS: The encoding gene of the antigen EgG1Y162 of E. granulosus was recombined with E. coli-Mycobacterium shuttle expression plasmid vector pMV361 by genetic engineering technique, and transformed into E. coli for amplification. The recombinant plasmid rpMV-EgG1Y162 was identified by PCR, double digestion with restriction enzymes, and sequence analysis. The confirmed rpMV-EgG1Y162 was transformed into BCG strain via electroporation technique to construct the recombinant rBCG-EgG1Y162. After identification by PCR and double digestion with restriction enzymes, the recombinant strain was cultured for about 2 weeks. In order to induce the expression of target protein, the rBCG was placed in 45 degrees C for 30 min. SDS-PAGE and Western blotting were used to analyze the expressive protein. RESULTS: The product of recombinant plasmid rpMV-EgG1Y162 was approximately 360 bp by PCR amplification and double digestion with restriction enzymes, consistent with the expected fragment length. Sequencing results showed that the inserted sequence was correct. The rBCG-EgG1Y162 grew well and the identification of PCR and enzyme digestion revealed accuracy. The results of SDS-PAGE and Western blotting showed that the relative molecular weight (M(r)) of the protein was about 71 000. CONCLUSION: The E. granulosus rBCG-EgG1Y162 strain is constructed and expressed.

Multi-frequency ultrasound-assisted cellulase extraction of protein from mulberry leaf: Kinetic, thermodynamic, and structural properties.

The effects of different extraction methods (traditional extraction, ultrasound extraction, cellulase extraction, and ultrasound-assisted cellulase extraction) on the yield of mulberry leaf protein (MLP) were investigated, and the results revealed that multi-frequency ultrasound-assisted cellulase extraction was the most efficient extraction method. The mechanism of the synergistic extraction method used to efficiently extract protein from mulberry leave was investigated, focusing on the kinetics and thermodynamics of the enzymatic process. The results revealed that kinetic parameters KM decreased by 14.07% and kA increased by 5.02%, and the thermodynamic parameters Ea, ΔH, and ΔS decreased by 44.81%, 48.41%, and 21.12 %, respectively, following the process of multi-frequency ultrasound (MFU) pretreatment. The spectral analysis with fluorescence spectra manifested that ultrasound exposed hydrophobic groups and induced molecular unfolding of MLP. Atomic force microscope showed that ultrasound decreased particle size while increasing flexibility of MLP. The effect of ultrasound increases the binding frequency of cellulase and substrates, resulting in greater affinity between the two and promoting the solubilization of MLP. This study provides a theoretical basis to improve the application prospects of MLP.

Design and functional preliminary investigation of recombinant antigen EgG1Y162-EgG1Y162 against Echinococcus granulosus.

In the early stage, our research group cloned Echinococcus granulosus-specific antigen, EgG1Y162, from protoscolex and adult worms of E. granulosus. In order to enhance the immunogenicity of the vaccine, we prepared a recombinant vaccine by tandemly linking EgG1Y162, splicing the protein and linker at the gene level. This approach is expected to improve the immunogenicity of the vaccine by enhancing the molecular weight of the protein and increasing the antigenic epitopes. Bioinformatics was used to predict the physicochemical properties, transmembrane domain, protein structure, and T-/B-cell antigenic epitope of different recombinant proteins, EgG1Y162-linker-EgG1Y162. Finally, the linker sequence, "GGGGSGGG," which had the least influence on the migration of recombinant protein T/B epitope and can fold normally in series with EgG1Y162, was selected to design the recombinant vaccine. The plasmid was produced using genetic engineering techniques, and the recombinant protein, EGG1Y162-GGGGSGGG-EgG1Y162, was induced to be expressed and purified. EgG1Y162-GGGGSGGG-EgG1Y162 was identified to be correctly expressed with 100% specificity. Compared with EgG1Y162, EgG1Y162-GGGGSGGG-EgG1Y162 was more likely to promote dendritic cell maturation. EgG1Y162-GGGGSGGG-EgG1Y162 was speculated to have the potential to improve antigen immunogenicity by increasing the molecular weight and antigenic epitope.