A Facile Strategy for Multiplex Protein Detection by a Fluorescent Microsphere-Based Digital Immunoassay.

The digital immunoassay is a highly sensitive detection technique based on single-molecule counting and is widely used in the ultrasensitive detection of biomarkers. Herein, we developed a fluorescent microsphere-based digital immunoassay (FMDIA) by employing fluorescent microspheres as both the carriers for immunoreaction and fluorescent reports for imaging. In this approach, the target protein in the sample was captured by fluorescent microspheres to form a biotin-labeled sandwich immunocomplex, and then, the fluorescent microspheres containing the target protein molecules were captured by adding streptavidin-coated magnetic beads (SA-MBs). By counting the proportion of fluorescence-positive magnetic beads, the concentration of the target protein can be precisely quantified. As a proof of concept, α fetoprotein (AFP) and human interleukin-6 (IL-6) were used to assess the analytical performance of the proposed FMDIA, and limit of detection (LOD) values of 21 pg/mL (0.30 pM) and 0.19 pg/mL (7.3 fM) were achieved, respectively. The results of AFP detection in serum samples of patients and healthy people were consistent with the reference values given by the hospital. Furthermore, by adding fluorescent microspheres of various colors for encoding, the proposed FMDIA can easily realize the simultaneous detection of multiple proteins without the need to introduce multiple modified magnetic beads. This multiplex protein detection strategy, in which the reactions are first carried out on the fluorescent microspheres and then magnetic beads are used to capture the fluorescent reporters containing the target molecules, provides a new idea for digital assays.

Rare case of septic shock combined with meningitis caused by Pasteurella multocida without a history of cat and dog bites.

BACKGROUND: Pasteurella multocida is a zoonotic pathogen that mainly causes local skin and soft tissue infections in the human body through cat and dog bites. It rarely causes bacteraemia (or sepsis) and meningitis. We reported a case of septic shock and meningitis caused by P. multocida in a patient without a history of cat and dog bites. CASE PRESENTATION: An 84-year-old male patient was urgently sent to the emergency department after he was found with unclear consciousness for 8 h, accompanied by limb tremors and urinary incontinence. In the subsequent examination, P. multocida was detected in the blood culture and wound secretion samples of the patient. However, it was not detected in the cerebrospinal fluid culture, but its DNA sequence was detected. Therefore, the patient was clearly diagnosed with septic shock and meningitis caused by P. multocida. The patient had no history of cat or dog contact or bite. The patient was subsequently treated with a combination of penicillin G, doxycycline, and ceftriaxone, and he was discharged after 35 days of hospitalisation. CONCLUSION: This report presented a rare case of septic shock and meningitis caused by P. multocida, which was not related to a cat or dog bite. Clinical doctors should consider P. multocida as a possible cause of sepsis or meningitis and should be aware of its potential seriousness even in the absence of animal bites.

Amplification-free detection of Mpox virus DNA using Cas12a and multiple crRNAs.

Mpox virus (MPXV) is a zoonotic DNA virus that caused human Mpox, leading to the 2022 global outbreak. MPXV infections can cause a number of clinical syndromes, which increases public health threats. Therefore, it is necessary to develop an effective and reliable method for infection prevention and control of epidemic. Here, a Cas12a-based direct detection assay for MPXV DNA is established without the need for amplification. By targeting the envelope protein gene (B6R) of MPXV, four CRISPR RNAs (crRNAs) are designed. When MPXV DNA is introduced, every Cas12a/crRNA complex can target a different site of the same MPXV gene. Concomitantly, the trans-cleavage activity of Cas12a is triggered to cleave the DNA reporter probes, releasing a fluorescence signal. Due to the application of multiple crRNAs, the amount of active Cas12a increases. Thus, more DNA reporter probes are cleaved. As a consequence, the detection signals are accumulated, which improves the limit of detection (LOD) and the detection speed. The LOD of the multiple crRNA system can be improved to ~ 0.16 pM, which is a decrease of the LOD by approximately ~ 27 times compared with the individual crRNA reactions. Furthermore, using multiple crRNAs increases the specificity of the assay. Given the outstanding performance, this assay has great potential for Mpox diagnosis.

Nucleic Acid Amplification-Free Digital Detection Method for SARS-CoV-2 RNA Based on Droplet Microfluidics and CRISPR-Cas13a.

Most of the methods currently developed for RNA detection based on CRISPR were combined with nucleic acid amplification. As a result, such methods inevitably led to certain disadvantages such as multiple operations, expensive reagents, and amplification bias. To solve the above problems, we developed a highly sensitive and specific nucleic acid amplification-free digital detection method for SARS-CoV-2 RNA based on droplet microfluidics and CRISPR-Cas13a. In this assay, thousands of monodisperse droplets with a size of 30 µm were generated within 2 min by a negative pressure-driven microfluidic chip. By confining a single target RNA recognition event to an independent droplet, the collateral cleavage products of activated Cas13a could be accumulated in one droplet. By combining the droplet microfluidics and CRISPR-Cas13a, SARS-CoV-2 RNA could be easily detected within 30 min with a detection limit of 470 aM. The performance of this assay was verified by specificity experiments and spiking and recovery experiments with human saliva. Compared with many developed methods for SARS-CoV-2 RNA detection, our method is time- and reagent-saving and easy to operate. Taken together, this digital detection method based on droplet microfluidics and CRISPR-Cas13a provides a promising approach for RNA detection in clinical diagnostics.

CRISPR/Cas13a Trans-Cleavage-Triggered Catalytic Hairpin Assembly Assay for Specific and Ultrasensitive SARS-CoV-2 RNA Detection.

New coronavirus (SARS-CoV-2), which has caused the coronavirus disease 2019 (COVID-19) pandemic, has brought about a huge burden on global healthcare systems. Rapid and early detection is important to prevent the spread of the pandemic. Here, an assay based on CRISPR/Cas13a and catalytic hairpin assembly (CHA), termed as Cas-CHA, was developed for ultrasensitive and specific detection of SARS-CoV-2 RNA. Upon specific recognition of the target, the CRISPR/Cas13a collaterally cleaved a well-designed hairpin reporter and triggered the CHA reaction. Under optimized conditions, the assay detected the SARS-CoV-2 RNA with a wide range of 100 aM to 100 nM and realized a low detection limit of 84 aM. At the same time, the whole detecting process could be completed within 35 min. More importantly, the assay was able to distinguish SARS-CoV-2 RNA from common human coronaviruses and analyze in saliva samples. By the flexible design of crRNA, the assay was expanded to detect other viruses. The clinical sample analysis verified that the proposed assay held a great potential for practical testing.

[Relationship between vitamin D and parathyroid hormone in Chinese elderly people].

OBJECTIVE: To explore the relationship between serum 25-hydroxyvitamin D(25(OH)D) and serum parathyroid hormone(PTH) level in Chinese people aged 50 years and above, and to probe the optimum threshold for vitamin D sufficiency preliminarily, and apply this threshold to predict the risk of metabolic syndrome(Mets) in this population. METHODS: A total of 750 serum samples were selected from the biological samples' bank of Chinese Chronic Diseases and Nutritional Survey(CCDNS, 2015-2017) by stratified sampling, basic information(including age, gender, season, etc. ) were collected from questionnaire and physical measurement of the subjects were conducted unified. Serum 25(OH)D concentration was determined by high performance liquid chromatography tandem mass spectrometer, and PTH and interleukin-6(IL-6) were detected by electrochemiluminescence method. Phosphorus, albumin(Alb), creatinine(Cr) in blood were detected by automatic biochemical analyzer. Factors affecting the concentration of 25(OH)D and PTH were found by multiple linear regression and adjusted by generalized additive model separately, threshold was predicted by locally weighted regression and smoothing scatterplot, abbreviated as LOESS, and the exact threshold of 25(OH)D was found when PTH reached the plateau by nonlinear least squares estimation and segmented regression. Relationship between 25(OH)D and Mets was analyzed by multivariate logistic regression using the different cut-off points in Chinese elderly people. RESULTS: Reference threshold for vitamin D deficiency in Chinese elderly people can be preliminarily discovered as serum total 25(OH)D was 19.62 ng/mL, and 28.44 ng/mL can be used as reference threshold for sufficient vitamin D. Sufficient 25(OH)D(≥28.44 ng/mL) could reduce the risk of Mets significantly(OR=0.617(0.439-0.869)) after adjusting for confounding factors such as sex, age, region, season, ect. A plateau in PTH was observed at a 25(OH)D concentration of 20.03-28.43 ng/mL for male whereas 13.12-26.33 ng/mL for female by gender stratification analysis, but no cut-off point was obtained statistically. CONCLUSION: Reference threshold for vitamin D sufficiency in Chinese elderly people was preliminarily observed in the range of 19.62-28.44 ng/mL when PTH was maximally inhibited, and the threshold may vary with gender. Applying the threshold we also found that more sufficient levels of vitamin D were protective against Mets in this population.

[Association of vitamin D gene polymorphisms and serum 25-hydroxyvitamin D in Chinese women of childbearing age].

OBJECTIVE: To analyze the relationship between vitamin D(VitD)-related single nucleotide polymorphism(SNP) and 25-hydroxyvitamin D(25(OH) D) levels and VitD nutritional status. METHODS: A total of 1507 women of childbearing age aged 18-45 were selected from the sample bank of "2015 Chinese adult chronic disease and nutrition monitoring". Basic information(including region, season, age, height, weight, etc. ) of the subjects was collected. The SNPs related to VitD metabolism were screened, and the improved multiple ligase detection reaction was used for SNP testing. Liquid chromatography tandem mass spectrometry was used to determine the serum 25(OH)D concentration. The effects of genotypes on 25(OH)D level and VitD deficiency were analyzed by generalized linear model and binary logistic regression model, respectively. RESULTS: After adjusting for latitude, region, region type, season and age, CYP2R1 rs12794714, GC rs2282679, GC rs7041 and VDR rs2228570 were associated with serum 25(OH)D levels in women of childbearing age. The risk of VitD deficiency in individuals carrying GG genotype at rs2282679 was significantly higher than that in individuals carrying TT genotype(OR=2.466, 95%CI 1.690-3.598, P<0.001), and the risk of VitD deficiency in individuals carrying A allele at rs2228570 was lower than that in individuals carrying G allele(OR_(AA)=0.625, 95%CI 0.446-0.876, P_(AA)=0.006;OR_(GA)=0.661, 95%CI 0.502-0.869, P_(GA)=0.003). CONCLUSION: The genotype distribution of CYP2R1 rs12794714, GC rs2282679, GC rs7041 and VDR rs2228570 may be related to serum 25(OH)D level or VitD nutritional status of Chinese women of childbearing age.

Significant association of urinary alpha-1-microglobulin compared to urinary neutrophil gelatinase-associated lipocalin with renal insufficiency in patients with type 2 diabetes.

AIM: Various studies have reported that urinary neutrophil gelatinase-associated lipocalin (NGAL), an indicator of tubular damage, may be an effective biomarker of renal impairment in patients with diabetes. This study aimed to compare the ability of urinary alpha-1-microglobulin (a traditional tubular damage marker) with NGAL for evaluating renal insufficiency in patients with type-2 diabetes. METHODS: Urinary albumin-to-creatinine ratio (ACR) and estimated glomerular filtration rate (eGFR) were used to determine whether 513 participants with type-2 diabetes had renal dysfunction. Urinary alpha-1-microglobulin-to-creatinine ratio (A1MCR) and NGAL-to-creatinine ratio (NCR) were calculated. RESULTS: Although both A1MCR and NCR were significantly higher among participants with renal insufficiency than among participants without renal damage, the difference in A1MCR values between participants with and without renal insufficiency was relatively greater than the difference in NCR values, especially among the male subjects. The correlation of ACR or eGFR with A1MCR was stronger than that of ACR or eGFR with NCR. A1MCR showed a good capability for detecting renal dysfunction (area under the curve = 0.80), its cut-off value was 14.82 mg/g, corresponding to 71.4% sensitivity and 73.1% specificity. The diagnostic efficiency of A1MCR was significantly higher than that of NCR. CONCLUSION: The results indicated that the traditional tubular damage marker A1MCR was more significantly associated with renal insufficiency defined by ACR and/or eGFR and may have a higher diagnostic efficiency compared with the efficiency of NCR in patients with type-2 diabetes.

Cyclophosphamide induced physiological and biochemical changes in mice with an emphasis on sensitivity analysis.

The widespread use of cyclophosphamide (CP) in medical treatment had caused ubiquitous contamination in the environment. To data, many studies have been carried out on the toxic effect of CP. However, among these toxic effects of CP, which are the most sensitive remains unclear. Present study aimed to investigate the toxicity of CP on mice and evaluate the sensitivity of physiological-biochemical parameters upon exposure of mice to CP. Results showed that as compared with the control group, CP caused significant reduction in body weight (p < 0.01), spleen coefficient (p < 0.01), leukocyte density (p < 0.01) and alanine transaminase (ALT) in kidney (p < 0.01); However superoxide dismutase (SOD), malondialdehyde (MDA), ALT in liver and creatinine (Cr) in kidney significantly (p < 0.05) increased. Among the suppressed physiological and biochemical parameters, the sensitivity to CP toxicity was generally ranked as body weight > leukocyte density > ALT in kidney > spleen coefficient; while among the stimulated parameters, the sensitivity was ranked as MDA (liver) > Cr (kidney) > ALT (liver). Overall, the most sensitive parameters to CP toxicity may be associated with growth, immune system and the normal function of liver and kidney.

[Relationship between rs7041 polymorphism of GC gene and serum vitamin D status in Chinese women of childbearing age].

OBJECTIVE: To investigate the association of rs7041 polymorphism of GC gene that encodes the vitamin D-binding protein with serum vitamin D status in Chinese women of childbearing age. METHODS: A total of 1812 plasma samples of women childbearing aged 18-44 years old were selected by stratified random sampling technology from the established biological samples bank of Chinese Chronic Diseases and Nutrition Survey(CCDNS, 2015-2018). The serum 25(OH)D status was detected by enzyme-linked immunosorbent assay. The genotypes of rs7041 in the GC gene were analyzed by improved multiple ligase detection reaction method. RESULTS: A total of 1812 childbearing women aged 18-49 years were included in this study. The frequency of rs7041 genotypes in the study were distributed according to the Hardy-Weinberg equilibrium, indicating sufficient representativeness of our sample. The median serum 25(OH)D status was 16. 69(12. 04, 21. 69)ng/mL. The higher 25(OH)D levels was detected in the overall sample, southern women or women with normal vitamin D status with the CC genotype than the AA genotype(P<0. 05). Before and after correction, the risk of vitamin D insufficiency in the women carrying the CC genotype was decreased significantly compared with the women carrying the AA genotype(OR=0. 571, 95%CI 0. 373-0. 873). And the CC genotype of rs7041 was associated with a significant decrease in risk of 25(OH)D deficiency(in the subgroup of southern childbearing women, OR=0. 284, 95%CI 0. 144-0. 560 and in the subgroup of northern childbearing women, OR=0. 109, 95%CI 0. 015-0. 798). CONCLUSION: The GC rs7041 with A/C polymorphism are significantly correlated with 25(OH)D status in Chinese childbearing women, mutant CC genotype is a protective factor for vitamin D non-normal status risks.

Urinary orosomucoid 1 protein to creatinine ratio as a potential biomarker for early screening of kidney impairment in type-2 diabetes patients.

BACKGROUND: Early screening of diabetic kidney disease (DKD) remains a major challenge. Our aim was to evaluate the value of urinary orosomucoid 1 protein (UORM1) in early renal impairment screening in type-2 diabetes patients. METHODS: The concentration of UORM1, the UORM1-to-creatinine ratio (UORM1CR), the urinary albumin-to-creatinine ratio (ACR), the alpha-1-microglobulin-to-creatinine ratio (A1MCR) and estimated glomerular filtration rate (eGFR) were measured in 406 type-2 diabetes patients. Any positive values for ACR, A1MCR and/or eGFR were considered as indicative of renal impairment. RESULTS: On average, the levels of UORM1 and UORM1CR were about seven times higher in subjects with renal injury than in those without. Both UORM1 and UORM1CR, when adjusted via logarithm-transformation, were significantly related to ACR, A1MCR and eGFR levels. The highest correlation was observed between UORM1CR and A1MCR (r = 0.85, P < .001). The cut-off values for UORM1 (2.53 mg/L) and UORM1CR (3.69 mg/g) for the early diagnosis of kidney impairment were obtained from receiver operating characteristic curves. UORM1CR obviously had higher diagnostic efficiency corresponding to 83.26% sensitivity and 90.32% specificity than UORM1. Likewise, its sensitivity was higher than those of ACR, A1MCR and eGFR. Bad glycaemic control had the highest risk of increased UORM1CR (odds ratio [OR] = 2.81, P < .001), while high HDL-C (high-density lipoprotein cholesterol) decreased the risk of increased UORM1CR (OR = 0.38, P = .017). CONCLUSION: The UORM1CR (>3.69 mg/g) has the high diagnostic efficiency for the early screening of renal impairment in type-2 diabetes patients. Furthermore, good glycaemic control and high HDL-C might be protective factors against UORM1CR increase.

Rapid identification of the Candida glabrata species complex by high-resolution melting curve analysis.

BACKGROUND: Candida glabrata is a common pathogen that causes invasive candidiasis. Among non-albicans Candida infections, C glabrata infections are associated with the highest fatality rates. Candida glabrata sensu stricto, Candida nivariensis, and Candida bracarensis have been identified and together form the C glabrata species complex. It is difficult to detect the two rare species by traditional laboratory methods. This study established a method for the rapid identification of members of the C glabrata species complex based on high-resolution melting curve (HRM) analysis and evaluated its practical application. METHODS: The internal transcribed spacer (ITS) region was used as target gene region to design specific primers. HRM analysis was performed with three subspecies of the C glabrata species complex and negative controls to test its specificity and sensitivity. To evaluate its practical application, the HRM technique was tested with clinical isolates, and the results were compared with the DNA sequencing results. RESULTS: Differences were detected among the melting profiles of the members of the C glabrata species complex. The negative controls were not amplified, indicating the high specificity of the method. The minimum detection limits of C glabrata sensu stricto, C nivariensis, and C bracarensis were approximately 1 × 101  copies/µL or less. The results of the HRM analysis of the clinical isolates were consistent with the DNA sequencing results. CONCLUSIONS: The HRM method is sensitive and can be used to rapidly identify the members of the C glabrata species complex. The method can allow early and targeted treatment of patients with invasive candidiasis.

Performance Verification of Alternative Quality Control Materials for Urine Albumin Assessment.

BACKGROUND: Accurate detection of urine albumin is important for evaluating the progression of diabetic kidney disease. However, two levels of daily quality control may not be practically feasible in some small clinical laboratories owing to a small number of patient samples and high costs. We aimed to prepare homemade quality control material (HQM) to measure urine albumin and then verify its performance. METHODS: Normal saline solution and fresh mixed urine samples from five donors with serious kidney disease were used to prepare two levels of HQM (HQM1 and HQM2). Anhydrous ethylene glycol and sodium azide were used as antifreeze and as a preservative, respectively. RESULTS: Before being separated into Eppendorf tubes, 20 tests for HQM1 and HQM2 were performed, resulting in mean ± SD of 19.52 ± 0.91 mg/L and 105.28 ± 3.71 mg/L, respectively. After having been divided, the vial-to-vial variations of HQM1 and HQM2 were small (4.93% and 3.70%, respectively). The stability of HQM1 and HQM2 stored at 2 - 8°C was about 2 months and 80 days, respectively, and when stored at -20°C, remained stable for more than 8 months. After 1 - 8 months of cryopreservation at -20°C, once opened, the HQM in every Eppendorf tube could be kept for at least five days (CV < 6.1%). CONCLUSIONS: Our HQM stored at -20°C remained stable for a long time, and so could be considered as an alternative to standard QMs in the clinical laboratory.

Association between dietary carbohydrate intake, glycemic index and glycemic load, and risk of gastric cancer.

PURPOSE: The association between dietary carbohydrate intake, glycemic index (GI) and glycemic load (GL), and risk of gastric cancer (GC) has been investigated by many studies. However, the results of these studies were controversial. The aim of our study was to systematically assess this issue. METHODS: PUBMED and EMBASE were searched up to March 2015, and either a fixed- or a random-effects model was adopted to estimate overall relative risks (RRs). Dose-response, meta-regression, subgroup, and publication bias analyses were applied. RESULTS: Twenty-six studies with approximately 540,000 participants were finally included in this meta-analysis. High level of dietary carbohydrate intake (pooled RR 1.17, 95 % CI 0.91-1.50), GI (pooled RR 1.17, 95 % CI 0.80-1.69), and GL (pooled RR 1.06, 95 % CI 0.90-1.26) were all nonsignificantly associated with incidence of GC. In addition, no significant dose-response relationship was observed between carbohydrate intake, GI and GL, and the risk of GC. However, further subgroup analyses based on gender and geographic region suggested a significant association between higher carbohydrate intake (pooled RR 1.52, 95 % CI 1.10-2.08), GL (pooled RR 1.41, 95 % CI 1.04-1.92), and GC risk in males subgroup, and between higher carbohydrate intake (pooled RR 1.69, 95 % CI 1.36-2.09) and GC risk in Asian studies. CONCLUSIONS: No significant association was found between dietary carbohydrate intake, GI and GL, and risk of GC. However, significantly positive association was observed in the males subgroup and Asian studies.

Alanine aminotransferase is more sensitive to the decrease in hepatitis B virus-DNA load than other liver markers in chronic hepatitis B patients.

BACKGROUND: A direct correlation between hepatitis B virus DNA (HBV-DNA) and liver markers has not been identified in chronic hepatitis B (CHB) patients. However, the effect of HBV-DNA changes on liver markers remains unclear. We explored the association between decreased HBV-DNA and liver makers in CHB patients. METHODS: Chronic hepatitis B patients who visited Jinhua Central Hospital twice were selected for analysis. Finally, 171 participants with a 1-log reduction in HBV-DNA between the two visits were enrolled as the case group, and 158 participants with no significant changes in HBV-DNA were enrolled as the control group. RESULTS: There was no significant correlation between HBV-DNA and liver markers (P>.05). However, in longitudinal analysis, alanine aminotransferase (ALT), aspartate aminotransferase (AST), and gamma-glutamyl transpeptidase (GGT) were significantly different between the two tests (P<.05) in the case group. Conversely, there was no significant difference in the control group. When HBV-DNA decreased >26 times, ALT was reduced by half or more. A similar trend was observed with a decrease of >63 times for AST and a decrease of >76 times for GGT. CONCLUSIONS: A large change in HBV-DNA can lead to a significant variation in liver markers. In particular, ALT was more sensitive than other liver markers to a reduction in HBV-DNA.

Interleukin-1B 31 C>T polymorphism combined with Helicobacter pylori-modified gastric cancer susceptibility: evidence from 37 studies.

Gastric cancer is one of the most common malignancies worldwide. Interleukin-1-beta (IL-1ß) is a pro-inflammatory cytokine and potent inhibitor of gastric acid secretion. Some studies provided evidence of the association between IL-1B 31 polymorphism and gastric cancer risk while other studies did not. Therefore, we conducted a comprehensive meta-analysis to reassess the association. A systematic literature search of the PubMed and EMBASE databases identified 37 studies with 6108 cases and 8980 controls for this meta-analysis. The crude odd ratios (ORs) and the 95% confidence intervals (CIs) were calculated to evaluate the strength of the association. Meta-regression was used to determine the major source of heterogeneity across the studies. The pooled analysis did not suggest the significant association of IL-1B 31 C>T polymorphism with gastric cancer risk. Stratified analysis was performed by ethnicity, source of control, genotype method, and indicated a significantly increased gastric cancer risk associated with IL-1B 31T variant in the population-based subgroup (heterozygous model: OR = 1.22, 95% CI = 1.03-1.45). Moreover, stratified analysis by Helicobacter pylori infection status indicated that IL-1B 31 polymorphism increased gastric cancer risk in infection-positive subgroup (homozygous model: OR = 1.35, 95% CI = 1.02-1.78; heterozygous model: OR = 1.31, 95% CI = 1.04-1.66; recessive model: OR = 1.29, 95% CI = 1.04-1.61). The study suggested that IL-1B 31 polymorphism might confer susceptibility to gastric cancer in the presence of H. pylori infection, indicating a gene-environment interaction in gastric carcinogenesis.

Mechanism of Dose-Dependent Regulation of UBE1L by Polyphenols in Human Bronchial Epithelial Cells.

Ubiquitin activating enzyme E1-like (UBE1L) is the activating enzyme for ISG15ylation (ISG15, interferon stimulated gene 15). UBE1L is thought to be a candidate tumor suppressor gene and has positive activity against stress responses such as viral infections. Both type I interferon and retinoic acid are known to induce UBE1L expression. However, the molecular mechanism of UBE1L regulation is unclear. Here, the effect of several chemopreventive polyphenols on UBE1L expression in human bronchial epithelial cells (Beas-2B) was investigated. Lower concentrations of curcumin, (-)-epigallocatechin-3-gallate (EGCG) and resveratrol upregulated UBE1L, while high concentrations of curcumin, EGCG and resveratrol downregulated UBE1L levels. Interestingly, curcumin, EGCG and resveratrol diminished intracellular reactive oxygen species (ROS) at lower concentrations but generated ROS at higher concentrations. The antioxidant N-acetylcysteine (NAC) increased UBE1L protein levels, while pro-oxidants such as hydrogen peroxide and tert-butyl hydroperoxide (tBHP) decreased UBE1L protein levels, indicating that the intracellular redox status is associated with UBE1L expression. Kinase inhibitors were used to examine the contribution of mitogen-activated protein kinase (MAPK) activity to the polyphenol-regulated UBE1L. Only the inhibition of c-Jun N-terminal kinase (JNK) significantly reduced UBE1L expression. Knockdown of nuclear factor erythroid-2 related factor-2 (Nrf2) caused a concomitant decrease in UBE1L protein levels. It is concluded from the above mentioned results that JNK/Nrf2 signal pathway is involved in the regulation of UBE1L via intracellular ROS status when cells came in contact with polyphenols.

Evaluation of the diagnostic accuracy of nonstructural protein 1 Ag-based tests for dengue virus in Asian population: a meta-analysis.

BACKGROUND: Nonstructural protein 1 (NS1) Ag-based tests are useful for detecting dengue virus (DENV), but there is lack of evidence on the diagnostic accuracy of NS1 Ag-based tests in Asian population. Thus, we conducted this meta-analysis to obtain the overall estimated and summarized performance of the NS1 Ag-based tests in the detection of DENV in Asia. METHODS: PubMed, Embase and Medline were searched for studies that evaluated the diagnostic validity of NS1 Ag-based tests between January 1990 and November 2014. Data were analyzed by Meta-Disc and STATA software. RESULTS: A total of 18 studies including 3342 dengue cases and 1904 control cases which fulfilled the inclusion criteria were considered for analysis. The pooled sensitivity and specificity for NS1 Ag-based tests was 66 % (95 % CI 64.5-67.5) and 97.9 % (95 % CI 97.3-100), respectively. STRIP has the overall highest sensitivity (72.9 %, 95 % CI 70.1-75.5). According to viral serotype, the test with the highest sensitivity for DENV1, DENV2 and DENV3 were Platelia (83.7 %, 95 % CI 79.7-87.1), Panbio (71.8 %, 95 % CI 65.5-80.9) and STRIP (81.9 %, 95 % CI 75.5-87.2) respectively. The highest sensitivity for primary infection was Platelia (95.1 %, 95 % CI 92.6-96.9) and for secondary infection was STRIP (64 %, 95 % CI 53.2-73.9). CONCLUSION: Our meta-analysis suggests that NS1 Ag-based test is a good diagnostic method for DENV with a high specificity. However, viral serotype, serological status, clinical severity and the duration of illness are the main factors influencing the diagnostic accuracy.

Curcumin Reactivates Silenced Tumor Suppressor Gene RARß by Reducing DNA Methylation.

Reactivation of tumor suppressor genes by nontoxic bioactive food component represents a promising strategy for cancer chemoprevention. Retinoic acid receptor ß (RARß), one member of the RAR receptor family, is considered as a tumor suppressor. Reduced expression of RARß has been reported in lung cancer and other solid tumors. DNA hypermethylation of the promoter region of RARß is a major mechanism for its silencing in tumors. Recently, curcumin has been considered as a potential DNA methyltransferase inhibitor. Herein, we demonstrated that curcumin significantly elevate RARß expression at the mRNA and protein levels in tested cancer cells. Additionally, curcumin decreased RARß promoter methylation in lung cancer A549 and H460 cells. Mechanistic study demonstrated that curcumin was able to downregulate the mRNA levels of DNMT3b. In a lung cancer xenograft node mice model, curcumin exhibited protective effect against weight loss because of tumor burden. Tumor growth was strongly repressed by curcumin treatment. As the results from in vitro, RARß mRNA were increased and DNMT3b mRNA were decreased by curcumin treatment compared with the mice in control group. Altogether, this study reveals a novel molecular mechanism of curcumin as a chemo-preventive agent for lung cancer through reactivation of RARß.

Study on the Dose-Response Relationship between Magnesium and Type 2 Diabetes of Childbearing Women in the China Adult Chronic Disease and Nutrition Surveillance 2015.

Background: Magnesium (Mg) is an essential element and participates in many metabolic pathways. Many studies have found a certain negative correlation between magnesium and blood glucose parameters, but the dose-response relationship between them is still a relatively narrow research field. We aim to explore the dose-response relationship between plasma and dietary Mg and type 2 diabetes (T2DM) among childbearing women in a nationally representative sample. And we will also initially explore the threshold of dietary and plasma magnesium in the prevention of T2DM and their consistency. Methods: A total of 2912 18-44 year-old childbearing women were recruited from the China Adult Chronic Disease and Nutrition Surveillance (2015). Multivariate logistic regression was used to explore the dose-response relationship between plasma and dietary Mg and glucose parameters. The threshold effect between Mg and T2DM was explored by a restricted cubic spline regression. Results: It was found that when plasma Mg was increased by 0.041 mmol/L, the risk of T2DM, impaired fasting glucose (IFG), and HbA1c-hyperglycemia was reduced by 18%, 19%, and 18%, respectively. The possible threshold value for plasma Mg to prevent the risk of T2DM was 0.87 mmol/L. Through the quality control of the sample dietary survey data, 2469 cases were finally included for dietary analysis. And the possible threshold value for dietary Mg to prevent the risk of T2DM was 408 mg/d. Taking the recommended dietary Mg intake of 330 mg/d as the reference group, when the Mg intake reached 408 mg/d, the risk of T2DM was significantly reduced. And the average plasma Mg level of the people whose dietary intake reached 408 mg/d was 0.87 mmol/L. Conclusions: These results indicate that dietary Mg and plasma Mg have good consistency on the threshold effect of glucose parameters in women of childbearing age.