Proteomic and metabolomic profiling of plasma predicts immune-related adverse events in older patients with advanced non-small cell lung cancer.

The clinical success of immune checkpoint inhibitors is compromised by the fact of immune-related adverse events (irAEs), especially for older patients. To identify predictive biomarkers for older patients with irAEs, we used multiplex immunoassay and flow cytometry and liquid chromatography-tandem mass spectrometry to test immune factors and plasma protein and metabolites levels in non-small cell lung cancer (NSCLC) patients. The results showed that older patients with irAEs displayed lower CD28, CD4+ T cell, and B cell and higher interleukin (IL)-10 and CCL2 levels at baseline. Besides, lower aldolase, fructose-bisphosphate B (ALDOB), higher ST6GAL1, and lower lactate/pyruvate ratio at baseline were found in older patients with irAEs. Based on metabolomic markers, predictive models were developed to distinguish patients with grade 2-4 irAEs from grade 0-1 (Area under curve, AUC = 0.831) and to distinguish patients with grade 3-4 irAEs from grade 2 (AUC = 1). Our results confirmed the predictive value of plasma metabolites for irAEs in older patients with NSCLC.

Downregulation of MYBL1 in endothelial cells contributes to atherosclerosis by repressing PLEKHM1-inducing autophagy.

MYBL1 is a strong transcriptional activator involved in the cell signaling. However, there is no systematic study on the role of MYBL1 in atherosclerosis. The aim of this study is to elucidate the role and mechanism of MYBL1 in atherosclerosis. GSE28829, GSE43292 and GSE41571 were downloaded from NCBI for differentially expressed analysis. The expression levels of MYBL1 in atherosclerotic plaque tissue and normal vessels were detected by qRT-PCR, Western blot and Immunohistochemistry. Transwell and CCK-8 were used to detect the migration and proliferation of HUVECs after silencing MYBL1. RNA-seq, Western blot, qRT-PCR, Luciferase reporter system, Immunofluorescence, Flow cytometry, ChIP and CO-IP were used to study the role and mechanism of MYBL1 in atherosclerosis. The microarray data of GSE28829, GSE43292, and GSE41571 were analyzed and intersected, and then MYBL1 were verified. MYBL1 was down-regulated in atherosclerotic plaque tissue. After silencing of MYBL1, HUVECs were damaged, and their migration and proliferation abilities were weakened. Overexpression of MYBL1 significantly enhanced the migration and proliferation of HUVECs. MYBL1 knockdown induced abnormal autophagy in HUVEC cells, suggesting that MYBL1 was involved in the regulation of HUVECs through autophagy. Mechanistic studies showed that MYBL1 knockdown inhibited autophagosome and lysosomal fusion in HUVECs by inhibiting PLEKHM1, thereby exacerbating atherosclerosis. Furthermore, MYBL1 was found to repress lipid accumulation in HUVECs after oxLDL treatment. MYBL1 knockdown in HUVECs was involved in atherosclerosis by inhibiting PLEKHM1-induced autophagy, which provided a novel target of therapy for atherosclerosis.

Effects of Supercritical CO2 Treatment on Color, Lipid Oxidation, Heme Iron, Non-Heme Iron and Metmyoglobin Contents in Ground Pork.

The color, lipid oxidation, heme iron (HI) and non-heme iron (NHI) contents, metmyoglobin content and Soret band of myoglobin of ground pork subjected to supercritical CO2 treatment under different conditions, or to heat treatment (40°C, 2 h) and subsequent storage at 4°C were evaluated during 9-day period. Supercritical CO2 treatment significantly increased CIE L* and CIE b* values of ground pork during subsequent storage, while the HI content was slightly affected. In general, CIE a* value and metmyoglobin content were decreased. Supercritical CO2 treatment for 2 h could increase the thiobarbituric acid-reactive substances (TBARS) value, while treatment for 1 h or less had no effect. The NHI content could be increased only after treatment at above 40°C or 17.2 MPa for 2 h. The Soret band of myoglobin was shifted to longer wavelength. Increasing treatment temperature from 35°C to 45°C could increase CIE L*, CIE a*, CIE b* and TBARS values, HI and NHI contents of the ground pork, while decreasing metmyoglobin content. As the treatment pressure increased from 13.8 MPa to 20.7 MPa, CIE b* and TBARS values were decreased, while the NHI and metmyoglobin contents were increased. However, the other parameters were unchanged. Extending exposure time from 0.5 h to 2 h could increase CIE L*, CIE b* and TBARS values, HI contents, while decreasing CIE a* value and metmyoglobin content. Correlation analysis showed that the TBARS value was significantly and negatively correlated with the HI content or metmyoglobin content in samples treated at 40°C or above for 2 h.

Heat-inducible CAR-T overcomes adverse mechanical tumor microenvironment in a 3D bioprinted glioblastoma model.

Glioblastoma (GBM) presents a significant therapeutic challenge due to the limited efficacy of existing treatments. Chimeric antigen receptor (CAR) T-cell therapy offers promise, but its potential in solid tumors like GBM is undermined by the physical barrier posed by the extracellular matrix (ECM). To address the inadequacies of traditional 2D cell culture, animal models, and Matrigel-based 3D culture in mimicking the mechanical characteristics of tumor tissues, we employed biomaterials and digital light processing-based 3D bioprinting to fabricate biomimetic tumor models with finely tunable ECM stiffness independent of ECM composition. Our results demonstrated that increased material stiffness markedly impeded CAR-T cell penetration and tumor cell cytotoxicity in GBM models. The 3D bioprinted models enabled us to examine the influence of ECM stiffness on CAR-T cell therapy effectiveness, providing a clinically pertinent evaluation tool for CAR-T cell development in stiff solid tumors. Furthermore, we developed an innovative heat-inducible CAR-T cell therapy, effectively overcoming the challenges posed by the stiff tumor microenvironment.

Multi-omics data-based analysis characterizes molecular alterations of the vesicle genes in human colorectal cancer.

The role of vesicular genes in the development of colorectal cancer (CRC) is crucial. Analyzing alterations in these genes at multi-omics can aid in understanding the molecular pathways behind colorectal carcinogenesis and identifying potential treatment targets. However, studies on the overall alteration of vesicular genes in CRC are still lacking. In this study, we aimed to investigate the relationship between vesicle genetic alterations and CRC progression. To achieve this, we analyzed molecular alterations in CRC vesicle genes at eight levels, including mRNA, protein, and epigenetic levels. Additionally, we examined CRC overall survival-related genes that were obtained from a public database. Our analysis of chromatin structural variants, DNA methylation, chromatin accessibility, and proteins (including phosphorylation, ubiquitination, and malonylation), along with RNA-seq data from the TCGA database, revealed multiple levels of alterations in CRC vesicle genes in the collected tissue samples. We progressively examined the alterations of vesicle genes in mRNA and protein levels in CRC and discovered the hub genes. Further investigation identified the probable essential transcription factors. This study contributes to a thorough knowledge of the connection between vesicle gene alterations at multiple levels and the development of CRC and offers a theoretical framework for the identification of novel treatment targets.

Integration of 3D bioprinting and multi-algorithm machine learning identified glioma susceptibilities and microenvironment characteristics.

Glioma, with its heterogeneous microenvironments and genetic subtypes, presents substantial challenges for treatment prediction and development. We integrated 3D bioprinting and multi-algorithm machine learning as a novel approach to enhance the assessment and understanding of glioma treatment responses and microenvironment characteristics. The bioprinted patient-derived glioma tissues successfully recapitulated molecular properties and drug responses of native tumors. We then developed GlioML, a machine learning workflow incorporating nine distinct algorithms and a weighted ensemble model that generated robust gene expression-based predictors, each reflecting the diverse action mechanisms of various compounds and drugs. The ensemble model superseded the performance of all individual algorithms across diverse in vitro systems, including sphere cultures, complex 3D bioprinted multicellular models, and 3D patient-derived tissues. By integrating bioprinting, the evaluative scope of the treatment expanded to T cell-related therapy and anti-angiogenesis targeted therapy. We identified promising compounds and drugs for glioma treatment and revealed distinct immunosuppressive or angiogenic myeloid-infiltrated tumor microenvironments. These insights pave the way for enhanced therapeutic development for glioma and potentially for other cancers, highlighting the broad application potential of this integrative and translational approach.

Interaction of immune cells with renal cancer development: Mendelian randomization (MR) study.

BACKGROUND: Renal cell carcinoma (RCC) is a prevalent and extensively immune-infiltrated malignancy of the urinary system. Immune cells play a crucial role in both the progression and therapeutic interventions targeting RCC. Nevertheless, the interplay between RCC and immune cells remains understudied, lacking substantial evidence supporting their causal relationship. METHODS: For the purpose of investigating the causal connection between RCC and immune cell characteristics, a two-way two-sample Mendelian randomization (MR) analysis was carried out in this study. The aim was to determine whether specific immune cell traits have a causal impact on the risk of RCC. In order to achieve this, publicly accessible genetic data was utilized to examine and establish the potential relationship between 731 immune cell characteristics and the likelihood of developing RCC. Additionally, various techniques were applied to verify the reliability, variability, and presence of horizontal pleiotropy in the outcomes. RESULTS: We found a bidirectional causal relationship between RCC and immune cells according to the MR analysis results. It should be noted that CD4-CD8-T cells (OR = 1.61, 95%CI = 1.02-2.55, P = 4.07 × 10-2) pose a risk for RCC, whereas BAFF-R (OR = 0.69, 95%CI = 0.53-0.89, P = 5.74 × 10-3) and CD19 (OR = 0.59, 95%CI = 1.02-2.55, P = 4.07 × 10-2) on B cells act as protective factors. Furthermore, the presence of RCC reduces the levels of B cells (OR = 1.05, 95%CI = 1.01-1.09, P = 1.19 × 10-2) and CD8 + T cells (OR = 1.04, 95%CI = 1.00-1.08, P = 2.83 × 10-2). CONCLUSIONS: Our research illustrates the intricate correlation between immune cells and RCC, presenting novel insights for the prospective safeguarding against RCC risk and the exploration of fresh therapeutic targets.

Epstein-Barr virus-driven metabolic alterations contribute to the viral lytic reactivation and tumor progression in nasopharyngeal carcinoma.

Metabolic reprogramming induced by Epstein-Barr virus (EBV) often mirrors metabolic changes observed in cancer cells. Accumulating evidence suggests that lytic reactivation is crucial in EBV-associated oncogenesis. The aim of this study was to explore the role of metabolite changes in EBV-associated malignancies and viral life cycle control. We first revealed that EBV (LMP1) accelerates the secretion of the oncometabolite D-2HG, and serum D-2HG level is a potential diagnostic biomarker for NPC. EBV (LMP1)-driven metabolite changes disrupts the homeostasis of global DNA methylation and demethylation, which have a significantly inhibitory effect on active DNA demethylation and 5hmC content. We found that loss of 5hmC indicates a poor prognosis for NPC patients, and that 5hmC modification is a restriction factor of EBV reactivation. We confirmed a novel EBV reactivation inhibitor, α-KG, which inhibits the expression of EBV lytic genes with CpG-containing ZREs and the latent-lytic switch by enhancing 5hmC modification. Our results demonstrate a novel mechanism of which metabolite abnormality driven by EBV controls the viral lytic reactivation through epigenetic modification. This study presents a potential strategy for blocking EBV reactivation, and provides potential targets for the diagnosis and therapy of NPC.

Meta-analysis of dynamic contrast enhancement and diffusion-weighted MRI for differentiation of benign from malignant non-mass enhancement breast lesions.

Purpose: The objective of this study was to conduct a meta-analysis comparing the diagnostic efficacy of models based on diffusion-weighted imaging (DWI)-MRI, dynamic contrast enhancement (DCE)-MRI, and combination models (DCE and DWI) in distinguishing benign from malignant non-mass enhancement (NME) breast lesions. Materials and methods: PubMed, Embase, and Cochrane Library were searched, from inception to January 30, 2023, for studies that used DCE or DWI-MRI for the prediction of NME breast cancer patients. A bivariate random-effects model was used to calculate the meta-analytic sensitivity, specificity, and area under the curve (AUC) of the DCE, DWI, and combination models. Subgroup analysis and meta-regression analysis were performed to find the source of heterogeneity. Results: Of the 838 articles screened, 18 were eligible for analysis (13 on DCE, five on DWI, and four studies reporting the diagnostic accuracy of both DCE and DWI). The funnel plot showed no publication bias (p > 0.5). The pooled sensitivity and specificity and the AUC of the DCE, DWI, and combination models were 0.58, 0.72, and 0.70, respectively; 0.84, 0.69, and 0.84, respectively; and 0.88, 0.79, 0.90, respectively. The meta-analysis found no evidence of a threshold effect and significant heterogeneity among trials in terms of DCE sensitivity and specificity, as well as DWI specificity alone (I2 > 75%). The meta-regression revealed that different diagnostic criteria contributed to the DCE study's heterogeneity (p < 0.05). Different reference criteria significantly influenced the heterogeneity of the DWI model (p < 0.05). Subgroup analysis revealed that clustered ring enhancement (CRE) had the highest pooled specificity (0.92) among other DCE features. The apparent diffusion coefficient (ADC) with a mean threshold <1.3 × 10-3 mm2/s had a slightly higher sensitivity of 0.86 compared to 0.82 with an ADC of ≥1.3 × 10-3 mm2/s. Conclusion: The combination model (DCE and DWI) outperformed DCE or DWI alone in identifying benign and malignant NME lesions. The DCE-CRE feature was the most specific test for ruling in NME cancers.

Clinical profile, treatment and quality of life of patients with psoriatic arthritis in Malaysia: A population-based cross-sectional study.

Psoriatic arthritis (PsA) is a major comorbidity of psoriasis and may lead to irreversible joint damage and disability. This study aims to describe the clinical profile, treatment and quality of life (QoL) of patients with PsA in Malaysia. This is a multicentre retrospective cross-sectional study of psoriasis patients who were notified to the Malaysian Psoriasis Registry (MPR) from January 2007 to December 2018. Of 21 735 psoriasis patients, 2756 (12.7%) had PsA. The male to female ratio was 1:1. The mean age of psoriasis onset for PsA patients was 34.73 ± 14.44 years. They had a higher rate of family history of psoriasis (26% vs. 22.4%, p < 0.001), scalp (82.7% vs. 81.0%, p = 0.04) and nail involvement (73.3% vs. 53.3%, p < 0.001), obesity (62.6% vs. 54.4%, p < 0.001), dyslipidaemia (23.8% vs. 15.4%, p < 0.001), hypertension (31.1% vs. 22.7%, p < 0.001) and diabetes mellitus (20.9% vs. 15.2%, p < 0.001) compared to non-PsA patients. More than half (54.3%) had severe psoriasis [(body surface area >10% and/or Dermatology Life Quality Index (DLQI) >10)]. Most had oligo-/monoarthropathy (40.3%), followed by distal interphalangeal arthropathy (31.3%), symmetrical polyarthropathy (28.3%), spondylitis/sacroiliitis (8.2%) and arthritis mutilans (3.2%). Nearly 40% of PsA patients received systemic treatment, but only 1.6% received biologic agents. QoL was more significantly affected in PsA than in non-PsA patients (mean DLQI 10.12 ± 7.16 vs. 9.52 ± 6.67, p < 0.001). One in eight patients with psoriasis in Malaysia had PsA. They had a higher incidence of comorbidities, severe disease, impaired QoL and were more likely to receive systemic and biological treatment compared to non PsA patients.

Prediction of histopathologic grades of bladder cancer with radiomics based on MRI: Comparison with traditional MRI.

PURPOSE: To compare biparametric magnetic resonance imaging (bp-MRI) radiomics signatures and traditional MRI model for the preoperative prediction of bladder cancer (BCa) grade. MATERIALS AND METHODS: This retrospective study included 255 consecutive patients with pathologically confirmed 113 low-grade and 142 high-grade BCa. The traditional MRI nomogram model was developed using univariate and multivariate logistic regression by the mean apparent diffusion coefficient (ADC), vesical imaging reporting and data system, tumor size, and the number of tumors. Volumes of interest were manually drawn on T2-weighted imaging (T2WI) and ADC maps by 2 radiologists. Using one-way analysis of variance, correlation, and least absolute shrinkage and selection operator methods to select features. Then, a logistic regression classifier was used to develop the radiomics signatures. Receiver operating characteristic (ROC) analysis was used to compare the diagnostic abilities of the radiomics and traditional MRI models by the DeLong test. Finally, decision curve analysis was performed by estimating the clinical usefulness of the 2 models. RESULTS: The area under the ROC curves (AUCs) of the traditional MRI model were 0.841 in the training cohort and 0.806 in the validation cohort. The AUCs of the 3 groups of radiomics model [ADC, T2WI, bp-MRI (ADC and T2WI)] were 0.888, 0.875, and 0.899 in the training cohort and 0.863, 0.805, and 0.867 in the validation cohort, respectively. The combined radiomics model achieved higher AUCs than the traditional MRI model. decision curve analysis indicated that the radiomics model had higher net benefits than the traditional MRI model. CONCLUSION: The bp-MRI radiomics model may help distinguish high-grade and low-grade BCa and outperforming the traditional MRI model. Multicenter validation is needed to acquire high-level evidence for its clinical application.

Interferon-γ in the tumor microenvironment promotes the expression of B7H4 in colorectal cancer cells, thereby inhibiting cytotoxic T cells.

The bioactivity of interferon-γ (IFN-γ) in cancer cells in the tumor microenvironment (TME) is not well understood in the current immunotherapy era. We found that IFN-γ has an immunosuppressive effect on colorectal cancer (CRC) cells. The tumor volume in immunocompetent mice was significantly increased after subcutaneous implantation of murine CRC cells followed by IFN-γ stimulation, and RNA sequencing showed high expression of B7 homologous protein 4 (B7H4) in these tumors. B7H4 promotes CRC cell growth by inhibiting the release of granzyme B (GzmB) from CD8+ T cells and accelerating apoptosis in CD8+ T cells. Furthermore, interferon regulatory factor 1 (IRF1), which binds to the B7H4 promoter, is positively associated with IFN-γ stimulation-induced expression of B7H4. The clinical outcome of patients with CRC was negatively related to the high expression of B7H4 in cancer cells or low expression of CD8 in the microenvironment. Therefore, B7H4 is a biomarker of poor prognosis in CRC patients, and interference with the IFN-γ/IRF1/B7H4 axis might be a novel immunotherapeutic method to restore the cytotoxic killing of CRC cells.

Arsenic exposure and pulmonary function decline: Potential mediating role of TRAIL in chronic obstructive pulmonary disease patients.

BACKGROUND: Environmental arsenic (As) exposure is strongly related to the progression of chronic obstructive pulmonary disease (COPD). Pulmonary epithelial cells apoptosis is implicated in the pathophysiological mechanisms of COPD. However, the role of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), one biomarker of apoptosis, remains unclear in As-mediated pulmonary function alternations in COPD patients. METHODS: This study included 239 COPD patients. The serum level of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) was measured by enzyme-linked immunosorbent assay (ELISA). The blood As level was determined through inductively coupled plasma mass spectrometry (ICP-MS). RESULTS: Blood As levels exhibited a negative and dose-dependent correlation with pulmonary function. Per unit elevation of blood arsenic concentrations was related to reductions of 0.339 L in FEV1, 0.311 L in FVC, 1.171% in FEV1/FVC%, and 7.999% in FEV1% in COPD subjects. Additionally, a positive dose-response correlation of blood As with serum TRAIL was found in COPD subjects. Additionally, the level of serum TRAIL was negatively linked to lung function. Elevated TRAIL significantly mediated As-induced decreases of 11.05%, 13.35%, and 31.78% in FVC, FEV1, and FEV1%, respectively among the COPD patients. CONCLUSION: Blood As level is positively correlated with pulmonary function decline and serum TRAIL increase in individuals with COPD. Our findings suggest that elevated TRAIL levels may serve as a mediating mechanism through which As contributes to declining lung function in COPD patients.

The prognostic value of the stress hyperglycemia ratio for all-cause and cardiovascular mortality in patients with diabetes or prediabetes: insights from NHANES 2005-2018.

BACKGROUND: The Stress hyperglycemia ratio (SHR) is a novel marker reflecting the true acute hyperglycemia status and is associated with clinical adverse events. The relationship between SHR and mortality in patients with diabetes or prediabetes is still unclear. This study aimed to investigate the predictive value of the SHR for all-cause and cardiovascular mortality in patients with diabetes or prediabetes. METHODS: This study included 11,160 patients diagnosed with diabetes or prediabetes from the National Health and Nutrition Examination Survey (2005-2018). The study endpoints were all-cause and cardiovascular mortality, and morality data were extracted from the National Death Index (NDI) up to December 31, 2019. Patients were divided into SHR quartiles. Cox proportion hazards regression was applied to determine the prognostic value of SHR. Model 1 was not adjusted for any covariates. Model 2 was adjusted for age, sex, and race. Model 3 was adjusted for age, sex, race, BMI, smoking status, alcohol use, hypertension, CHD, CKD, anemia, and TG. RESULTS: During a mean follow-up of 84.9 months, a total of 1538 all-cause deaths and 410 cardiovascular deaths were recorded. Kaplan-Meier survival analysis showed the lowest all-cause mortality incidence was in quartile 3 (P < 0.001). Multivariate Cox regression analyses indicated that, compared to the 1st quartile, the 4th quartile was associated with higher all-cause mortality (model 1: HR = 0.89, 95% CI 0.74-10.7, P = 0.226; model 2: HR = 1.24, 95% CI 1.03-1.49, P = 0.026; model 3: HR = 1.30, 95% CI 1.08-1.57, P = 0.006). The 3rd quartile was associated with lower cardiovascular mortality than quartile 1 (model 1: HR = 0.47, 95% CI 0.32-0.69, P < 0.001; model 2: HR = 0.66, 95% CI 0.45-0.96, P = 0.032; model 3: HR = 0.68, 95% CI 0.46-0.99, P = 0.049). There was a U-shaped association between SHR and all-cause mortality and an L-shaped association between SHR and cardiovascular mortality, with inflection points of SHR for poor prognosis of 0.87 and 0.93, respectively. CONCLUSION: SHR is related to all-cause and cardiovascular mortality in patients with diabetes or prediabetes. SHR may have predictive value in those patients.

Association of blood cadmium concentration with chronic obstructive pulmonary disease progression: a prospective cohort study.

BACKGROUND: Prior studies in patients with chronic obstructive pulmonary disease (COPD) had indicated a potential correlation between cadmium (Cd) exposure and reduction in lung function. Nevertheless, the influence of Cd exposure on the progression of COPD remained unknown. Exploring the relationship between Cd exposure and the progression of COPD was the aim of this investigation. METHODS: Stable COPD patients were enrolled. Blood samples were collected and lung function was evaluated. Regular professional follow-ups were conducted through telephone communications, outpatient services, and patients' hospitalization records. RESULTS: Each additional unit of blood Cd was associated with upward trend in acute exacerbation, hospitalization, longer hospital stay, and death within 2 years. Even after adjusting for potential confounding factors, each 1 unit rise in blood Cd still correlated with a rise in the frequencies of acute exacerbation, longer hospital stay, and death. Moreover, COPD patients with less smoking amount, lower lung function and without comorbidities were more vulnerable to Cd-induced disease deterioration. CONCLUSION: Patients with COPD who have higher blood Cd concentration are susceptible to worse disease progression.

Diabetes: a potential mediator of associations between polycyclic aromatic hydrocarbon exposure and stroke.

Epidemiological evidence suggests associations between exposure to polycyclic aromatic hydrocarbons (PAHs) and cardiovascular diseases (CVD), while diabetes is a common risk factor for CVD. The present study aims to clarify the effect of high PAH exposure on diabetes and stroke in general population. A total of 7849 individuals aged 20 years or older from the National Health and Nutrition Examination Survey 2007-2016 were included in the study. The logistic regression analysis modeled the association between PAH exposure and diabetes as well as stroke. The analysis yielded odds ratios (ORs) and 95% confidence intervals (CIs). The study also evaluated the potential mediating role of diabetes in the relation between PAH exposure and stroke via mediating effect analyses. Of the 7849 eligible participants, 1424 cases of diabetes and 243 cases of stroke were recorded. After adjusting for covariates including age, gender, smoking status, drinking status, education level, marital status, physical activity, hypertension, low-density lipoprotein cholesterol, and BMI, the ORs for stroke in the highest quartile (Q4) of total urinary PAHs were 1.97 (95% CI 1.11-3.52, P = 0.022) as compared to the lowest quartile (Q1) of total urinary PAHs. The ORs for diabetes in the Q4 of total urinary PAHs were 1.56 (95% CI 1.15-2.12, P = 0.005), while the ORs between Q4 and Q1 for stroke and diabetes concerning exposure to 2-hydroxynaphthalene were 2.23 (95% CI 1.17-4.25, P = 0.016) and 1.40 (95% CI 1.07-1.82, P = 0.015), respectively. The mediation analysis found that diabetes accounted for 5.00% of the associations between urinary PAHs and the prevalence of stroke. Urinary metabolites of PAH have been linked to stroke and diabetes. Increasing the risk of diabetes may play a significant role in mediating the association between exposure to PAHs and increased risk of stroke. Monitoring and improving glucose metabolism in individuals with high exposure to PAHs may aid in reducing the prevalence of stroke.

Prokaryotic Expression and Affinity Purification of DDX3 Protein.

BACKGROUND: DDX3 is a protein with RNA helicase activity that is involved in a variety of biological processes, and it is an important protein target for the development of broad-spectrum antiviral drugs, multiple cancers and chronic inflammation. OBJECTIVE: The objective of this study is to establish a simple and efficient method to express and purify DDX3 protein in E. coli, and the recombinant DDX3 should maintain helicase activity for further tailor-made screening and biochemical function validation. METHODS: DDX3 cDNA was simultaneously cloned into pET28a-TEV and pNIC28-Bsa4 vectors and transfected into E. coli BL21 (DE3) to compare one suitable prokaryotic expression system. The 6×His-tag was fused to the C-terminus of DDX3 to form a His-tagging DDX3 fusion protein for subsequent purification. Protein dissolution buffer and purification washing conditions were optimized. The His-tagged DDX3 protein would bind with the Ni-NTA agarose by chelation and collected by affinity purification. The 6×His-tag fused with N-terminal DDX3 was eliminated from DDX3 by TEV digestion. A fine purification of DDX3 was performed by gel filtration chromatography. RESULTS: The recombinant plasmid pNIC28-DDX3, which contained a 6×His-tag and one TEV cleavage site at the N terminal of DDX3 sequence, was constructed for DDX3 prokaryotic expression and affinity purification based on considering the good solubility of the recombinant His-tagging DDX3, especially under 0.5 mM IPTG incubation at 18 °C for 18 h to obtain more soluble DDX3 protein. Finally, the exogenous recombinant DDX3 protein was obtained with more than 95% purity by affinity purification on the Ni-NTA column and removal of miscellaneous through gel filtration chromatography. The finely-purified DDX3 still retained its ATPase activity. CONCLUSION: A prokaryotic expression pNIC28-DDX3 system is constructed for efficient expression and affinity purification of bioactive DDX3 protein in E. coli BL21(DE3), which provides an important high-throughput screening and validation of drugs targeting DDX3.