A simple and cost-effective indirect competitive enzyme-linked immune sorbent assay (ic-ELISA) was developed to rapidly screen the content of aflatoxin B1 (AFB1) in lotus seeds, and the results were confirmed by ultra-fast liquid chromatography-tandem mass spectrometry( UFLC-MS/MS). Matrix-matched calibration expressed a good linearity ranging from 0. 171 to 7. 25 µg · L(-1) for AFB, with R2 > 0.978. The medium inhibitory concentration( IC50 ) for AFB1 was 1.29 µg · L(-1), the recovery for AFB1 was 74.73% to 126.9% with RSD < 5%, and the limit of detection (IC10) was 0.128 µg · L(-1). The developed ic-ELSIA method was applied to rapid analysis of AFB, in 20 lotus seeds samples and the results indicated that the contents of AFB, in samples 1-15 were in the range of 1. 19- 115. 3 µg · kg(-1) and in 40% of the samples exceeded the legal limit(5 µg · kg(-1)), while the contamination rate of AFB, in samples 16-20 was 40%. Pearson correlation coefficient(r) reached 0.997 for AFB1 content in the samples detected by ic-ELSIA and UFLC-MS/MS methods. The results proved that the developed ic-ELISA method is simple, sensitive and reliable, and can be used for rapid and high-throughput screening of AFB1 in lotus seeds
Aflatoxin B1 ; analysis ; Drug Contamination ; Enzyme-Linked Immunosorbent Assay ; methods ; Loteae ; chemistry ; Seeds ; chemistry

For diabetes mellitus, little research has been done on the tissue-based or cell-based drug screening model, which has advantages over traditional animal diabetic model in high specificity, high screening volume, low cost and simple manipulation. Considering that the maintenance of complete islet tissue structure is the prerequisite for islet cells to perform their functions normally, an in vitro islet-based drug screening model for diabetes mellitus was established and evaluated. Pancreatic islets were isolated from 3 weeks old mice of either sex by collagenase digestion and density gradient centrifugation as prescribed by Ramanadham S. The volume of 0.1% (W/V) collagenase IV, 0.1% (W/V) Hyaluroridase and 0.1% (W/V) DNase I were 4 times, 2 times and 1 times that of the islets to be digested. And a 2 hours' cold digestion at 4 degrees C was followed by a 10 minutes' warm digestion at 37 degrees C. Under the optimized digestion condition, the islet recovery could be increased by 10%. The isolated islets could survive 6 weeks in vitro and show stable insulin secretion in the first 10 days after inoculation. The obtained islets were cultured in RPMI-1640 medium at 37 degrees C with 5% CO2. Then a diabetic model was established by selecting streptozotocin (STZ) as the evocator and nitric oxide (NO) as the responding index. After 1 day's inoculation, islets culture was treated with STZ, whose concentration ranged from 0 to 5.0 mmol/L. NO was measured by a colorimetric assay at 540nm based on the Griess reaction for 10 min with 0.1 mL Griess reagent and 0.1 mL culture supernatants. Insulin secretion was assayed by RIA methods. Due to the islets-related inoculation variations, NO release and insulin content were both expressed as a percentage of the value recorded in basal experiment which was in the only presence of Krebs culture medium. It was testified that the amount of NO released from islet itself remained steady at 30-35 mmol/L regardless of the changes of STZ concentration from 0 to 5.0 mmol/L. However the NO content in the supernatants of islets culture had close relationship with STZ concentration. This indicated that in this STZ-induced islet diabetic model, NO mainly comes from STZ when it dissolves in water. On the other hand, when STZ changed from 0 to 5.0 mmol/L, the dose-dependent relationship between NO content and insulin secretion showed that the increase of NO came along with the decrease of insulin secretion, which is an important symbol of islet function. As a kind of oxidative free radical, NO is capable of impair islet cells. Thus, NO is a reliable responding index of the model. The optimal STZ concentration in the model is finally determined to be 5.0 mmol/L, under which condition the NO content and insulin secretion is 10.81 times and 0.43 times that in the medium before STZ is added. So if anything is effective in lowering the NO content in the culture, it could protect islets cells from the oxidative attacks of NO. Finally, as an application of the model, the scavenging effect of KOSCr on NO was studied. In a series of KOSCr with different chromium content, all had shown better NO scavenging effects than KOS itself, which could give us an enlightenment of the influence of chromium ion on oligosaccharide. And 1 g/mL KOSCr with 3.519% chromium content can significantly inhibit the NO formation. This has lain a theoretic basis for the research of KOSCr bioactivity and quality control. These results suggested that the STZ-induced diabetic islet model which is impaired by NO free radical can be used effectively, fast and conveniently when screening potential diabetes drugs.
Animals ; Chromium ; pharmacology ; Diabetes Mellitus, Experimental ; metabolism ; Disease Models, Animal ; Female ; In Vitro Techniques ; Insulin ; metabolism ; Islets of Langerhans ; drug effects ; metabolism ; Male ; Mice ; Mice, Inbred ICR ; Nitric Oxide ; metabolism ; Streptozocin ; pharmacology

Objective To examine HIV prevalence and related risk factors among men who have sex with men(MSM) in Chongqing, and to explore the feasibility of using respondent-driven sampling (RDS) in the survey. Methods Based on results from formative research, a RDS survey was designed and conducted to collect demographic, behavioral and serologic data. RDSAT was used to calculate point estimation and confidence intervals. SPSS was used for bi-variate analysis using RDSAT exported weighed data. NETDRAW was used to draw network diagram. Results Among 617 subjects recruited, the adjusted HIV and syphilis prevalence rates were 16.8% and 10.9%, respectively. 73.0% of the subjects were 20 to 29 years old and 72.9% were officially registered residents of Chongqing. 83.4% were single, with the proportion of students the highest, accounting for 24.6%. During the last six months, 83.4% of them reported ever having anal sex, and 54.0% reported having unprotected anal sex. Conclusion This survey confirmed that Chongqing had a higher reported HIV prevalence among MSM than from other Chinese cities. Comprehensive intervention services were required to address this alarmingly high prevalence, with focus on intervention through internet and those having syphilis infection. RDS seemed one of the effective ways of recruiting hidden MSM populations in Chongqing which had a large population of active MSM who did not frequently visit MSM venues as compared with snowball method.

The quantitative identification and enrichment of viable regulatory T cells (Treg) requires reliable surface markers that are selectively expressed on Treg. Foxp3 is the accepted marker of natural Treg, but it cannot be used to isolate cells for functional studies. CD127 is a new surface marker expressed in Treg cells. In this study, two populations of Treg, including CD4(+)CD25(+)CD127(low/-) and CD4(+)CD25(+)Foxp3(+)T cells, and profiles of the Foxp3 expression in CD4(+)CD25(+)CD127(low/-) cells were compared to evaluate which population is better. The peripheral blood cells were collected and spleen suspension of BALB/C mice were prepared, and using triple staining CD4, CD25, CD127 and CD4, CD25, Foxp3. The profiles of Treg, including CD4(+)CD25(+)CD127(low/-) and CD4(+)CD25(+)Foxp3(+) were detected by flow cytometry. The quadruple staining CD4, CD25, Foxp3 and CD127 were used to determine the CD127 expression in CD4(+)CD25(+)Foxp3(+) cells. The results showed that on T cell subset the median expression levels of CD4(+), CD4(+)CD25(+) were 39.02%, 5.35% in peripheral blood and 23.49%, 3.86% in spleen. On CD4(+) T cell subset, the median expression level of CD4(+)CD25(+)CD127(low/-) and CD4(+)CD25(+)Foxp3(+)T cells were 7.13%, 3.97% in peripheral blood and 12.8%, 8.23% in spleen. The ratio of CD4(+)CD25(+)CD127(low/-) T cells was higher than that of CD4(+)CD25(+)Foxp3(+) cells in both peripheral blood and spleen cells (P < 0.01). The CD4(+)CD25(+)CD127(low/-) cells highly expressed Foxp3, while the CD4(+)CD25(+)Foxp3(+)T cells lowly expressed CD127. It is concluded that compared with the CD4(+)CD25(+)Foxp3(+) populations, CD4(+)CD25(+)CD127(low/-) T cells better fit the definition of naturally occurring regulatory T cells in peripheral blood cells and spleen of BALB/C mice. CD127(low/-) is a characteristic marker on surface of CD4(+)CD25(+) Treg cells, and has been confirmed to be more specific marker for quantitatively sorting Treg cells.
Animals ; Biomarkers ; blood ; Female ; Interleukin-7 Receptor alpha Subunit ; analysis ; Mice ; Mice, Inbred BALB C ; Spleen ; cytology ; T-Lymphocytes, Regulatory ; metabolism

OBJECTIVEDevelopment and application of a real time fluorescent quantitative PCR (FQ-PCR) assay for detecting WU polyomavirus in children with low respiratory tract infections.

METHODSThe VP2 gene of WU polyomavirus was selected as the detection target, from which the real time primers and probes were designed. The standard curve was established by using recombinant plasmid as template. And the FQ-PCR assay for specific detection of WU polyomavirus was established. The specificity, sensitivity and reproducibility of the method were evaluated. Furthermore, the clinical specimens from children with respiratory tract infections collected in Wenling First People's Hospital were quantitatively detected using this method.

RESULTSIn this study, the FQ-PCR method was established to detect a specific fragment in VP2gene of WU polyomavirus. The standard curve coefficient R2 was 0.998. And this method can detect as low as 50 copies recombinant plasmid. The clinical specimens of sputum and throat swab from children with respiratory tract infections were quantitatively detected using this method. 7 sputum specimens were detected as WU polyomavirus positive in 700 sputum specimens, the positive ratio was 1.00%. No positive specimens were detected in 146 specimens of throat swabs and 846 blood samples from same patient population.

CONCLUSIONThe results indicated that the FQ-PCR assay method established in this study was specific, rapid and sensitive for detecting WU polyomavirus in children with lower respiratory tract infections. The sputum specimen is more suitable to be used for gene detection of WU polyomavirus than throat swab or blood.

Child ; Child, Preschool ; Female ; Humans ; Infant ; Male ; Polyomavirus ; isolation & purification ; Real-Time Polymerase Chain Reaction ; methods ; Respiratory Tract Infections ; virology ; Sputum ; virology

n increasing trend.

OBJECTIVETo establish a three-dimentional liver function evaluation system using 99mTc-diethyl iminodiacetic acid (99mTc-EHIDA) scintigraphy based on single photon emission computed tomography (SPECT).

METHODSTotally 16 patients with liver lesions were divided into cirrhosis group and non-cirrhosis group. SPECT was performed 2 days before operation and 5 days after operation. Serum liver functions were examined on the same day of scintigraphy. SPECT images of areas of interest of heart and liver were aquired. Time of the peak of EHIDA density in liver (Tpeak), five-minutes heart liver index (HLI5), blood clearance index (HH15), receptor index (LHL15), and the predictive values were calculated.

RESULTSTpeak was not significantly different between two groups, while HLI5, HH15, and LHL15 were significantly different (P = 0.033, P = 0.001, and P = 0.005). HLI, and LHL15 were significantly correlated with preoperative total protein and prealbumin levels (P = 0.003, P = 0.015, P = 0.022, P = 0.038) and post-operative prealbumin (P = 0.037, P = 0.042). The predictive values of HLI5 and LHL15 correlated well with postoperative HLI5 and LHL15 (r = 0.675, P = 0.016; r = 0.629, P = 0.028).

CONCLUSIONThe three-dimentional liver function evaluation system using 99mTc-EHIDA based on liver SPECT may facilitate the further studies of risks of liver surgery.

Adult ; Aged ; Animals ; Female ; Humans ; Liver Diseases ; diagnosis ; diagnostic imaging ; physiopathology ; Liver Function Tests ; Male ; Middle Aged ; Postoperative Period ; Preoperative Period ; Radiopharmaceuticals ; administration & dosage ; Technetium Tc 99m Diethyl-iminodiacetic Acid ; administration & dosage ; Tomography, Emission-Computed, Single-Photon

Entinostat plus exemestane in hormone receptor-positive (HR+) advanced breast cancer (ABC) previously showed encouraging outcomes. This multicenter phase 3 trial evaluated the efficacy and safety of entinostat plus exemestane in Chinese patients with HR + ABC that relapsed/progressed after ≥1 endocrine therapy. Patients were randomized (2:1) to oral exemestane 25 mg/day plus entinostat (n = 235) or placebo (n = 119) 5 mg/week in 28-day cycles. The primary endpoint was the independent radiographic committee (IRC)-assessed progression-free survival (PFS). The median age was 52 (range, 28-75) years and 222 (62.7%) patients were postmenopausal. CDK4/6 inhibitors and fulvestrant were previously used in 23 (6.5%) and 92 (26.0%) patients, respectively. The baseline characteristics were comparable between the entinostat and placebo groups. The median PFS was 6.32 (95% CI, 5.30-9.11) and 3.72 (95% CI, 1.91-5.49) months in the entinostat and placebo groups (HR, 0.76; 95% CI, 0.58-0.98; P = 0.046), respectively. Grade ≥3 adverse events (AEs) occurred in 154 (65.5%) patients in the entinostat group versus 23 (19.3%) in the placebo group, and the most common grade ≥3 treatment-related AEs were neutropenia [103 (43.8%)], thrombocytopenia [20 (8.5%)], and leucopenia [15 (6.4%)]. Entinostat plus exemestane significantly improved PFS compared with exemestane, with generally manageable toxicities in HR + ABC (ClinicalTrials.gov #NCT03538171).