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

Objective To investigated the effects of combined arsenic trioxide(ATO) and resveratrol(Res)on the viability of NB4 human leukemia cells. Methods NB4 human leukemia cell was used in this experiment.Cells were cultured in ATO (0,0.1875,0.3750,0.7500, 1.1250, 1.5000,2.2500,3.0000,5.0000 μmol/L) and Res (0, 1.5625,3.1250,6.2500, 12.5000, 18.7500,25.0000,37.5000,50.0000 μmol/L). Cell viabilities were measured by MTT in different treatment groups. Half inhibitory concentration(IC50) was calculated. The ratio of concentration of ATO and Res 1.5∶ 18,1.5∶ 25,1.5∶ 35 was added to cells, and the combination index(CI) was calculated. The level of ROS in control, ATO( 1.5000 μmol/L), Res(25.0000 μmol/L) and ATO(0.9000 μmol/L) + Res( 12.5000μmol/L) groups was measured by chemiluminescence assay. Results ①ATO( ≥0.7500 μmol/L) reduced the viability of NB4 cells in a concentration-dependent manner(P ＜ 0.05 ), and IC50 was (1.78 ± 0.11 )μmol/L. ②)Res (≥18.7500 μ mol/L) dose-dependently decreased the viability of NB4 cells (P ＜ 0.05 ), and IC50 was ( 18.71 ±0.18)μ mol/L. ③Combination of ATO and Res showed an antagonistic effect on NB4 cells viability. ④The ROS in Res group( 1670.55 ± 13.97) was significantly lower than that in control group(2345.88 ± 14.48,P ＜ 0.05). The ROS in ATO group (3092.42 ± 94.84) was significantly higher than that in control group(P ＜ 0.05). The ROS in ATO + Res group (1860.27 ± 15.99) was significantly lower than that in ATO group(P ＜ 0.05). Conclusions NB4 cell survival rate can be decreased by ATO and Res. The combination of arsenic trioxide and Res presents an antagonistic effect on NB4 cell viability, in part by reducing intracellular ROS formation.

A 24-year-old male suffered from acute nephritic syndrome, liver dysfunction, and mesenteric mass. Laboratory examination showed a variety of autoantibodies (ANA, SM, and A-β2-GP1) were positive. The biopsies of the kidney and the mesenteric mass were performed. The diagnosis was typeⅤ + Ⅲ lupus nephritis accompanied with Castleman's disease. Then the patient was given induction therapy of glucocorticoids and cyclophosphamide for the first 3 months, followed by rituximab as maintenance therapy. The patient was followed up after 0, 3, and 9 months. After 3-month treatment, lupus nephritis was partially remitted, and systemic lupus erythematosus disease activity index (SLEDAI) decreased to 4 scores in an inactivity phase from 20 scores in a serious activity phase at baseline. Nine months later, lupus nephritis was completely remitted and 50％ mesenteric mass was regressed through CT scanning. Lupus nephritis can accompany with multicentric Castleman's disease. Due to lack of clinical specificity and effective therapy, patients may have a high misdiagnosis rate and poor prognosis. The most reliable way to establish a definitive diagnosis relays on histopathologic confirmation. The management of induction therapy of glucocorticoids and cyclophosphamide, followed maintenance therapy of rituximab may become a beneficial treatment.

OBJECTIVE To investigate the neuroprotective effect and possible mechanisms of lute-olin-7-O-β-D-glucuronide (LGU) against focalcerebral ischemic injury. METHODS The focal cerebral ischemic injury model was established by middle cerebral artery occlusion (MCAO). Male Sprague Dawley rats were randomly divided into sham group,model group(MCAO),LGU group(0.24,0.72 and 2.16 mg·kg-1)and positive control group(Edaravone at 5 mg·kg-1).LGU was injected intravenously 30 min after MCAO.Neurological severity score,infarct volume and brain water content were detected 24 h after MCAO and the levels of Na+-K+ATPase,Ca2+ATPase,TNF-α and IL-1β were detected to explore the possible mechanisms.For the therapeutic time window test,LGU(0.72 mg·kg-1)was injected intrave-nously 0.5, 2, 4, 6, 8, 10 and 12 h respectively after MCAO. To evaluate motion behavior, LGU were injected intravenously 30 min after MCAO and once per day during detection period. The changes of motor coordination were detected by rotating rod method and grip strength analysis, and the changes of gaits were detected using DigiGait Imaging System. RESULTS LGU improved the neurological severity score, infarct volume ratio and brain water content. The therapeutic time window of LGU for cerebral infarction and brain edema was at least 6 h and for neurological dysfunction was 12 h.LGU also prolonged the latency on rotarod, increased the forelimb tension and improved 8 gait parameters, including stance duration,stride length,stance width,paw area,paw area variability,gait symmetry,ataxia coefficient and tau propulsion.Furthermore,LGU increased Na+-K+-ATPase and Ca2+-ATPase levels in the cortex and hippocampus in the ischemic side,reduced the levels of TNF-α and IL-1β in the serum. CONCLUSION LGU has a significant neuroprotective effect against cerebral ischemic injury via improving energy metabolism and reducing inflammation.

OBJECTIVE To investigate the protective effect and mechanisms of luteolin-7-O-β-d-glucuronide (LGU) on oxygen glucose deprivation (OGD)-induced H9C2 cardiomyocytes injury. METH-ODS The protective effect of LGU on OGD-induced H9C2 cardiomyocytes death were investigated by MTT assay. The microfilament change of H9C2 cardiomyocytes was detected by phalloidin staining and the lactate dehydrogenase (LDH) leakage rate was also detected by LDH kit. In order to explore the possible mechanisms of LGU, ATP content, intracellular Ca2+fluorescent intensity and concentra-tion, mitochondrial membrane potential (MMP)and the expressions of apoptosis-related proteins were detected by ATP kit,CLSM(Fluo-3/AM probe),Ca2+kit,CLSM(JC-1 probe)and western blotting meth-od, respectively. RESULTS The inhibition of H9C2 cardiomyocyte survival rate inducedby OGD was improvedby pretreated with LGU in a concentrationdependent manner. The microfilaments injury as well as the increase of LDH leakage rate were also improvedby pretreated with LGU.The ATP content was significantly decreased,intracellular Ca2+fluorescent intensity and concentration were significantly increased and the MMP was significantly decreased 4 hafter OGD. LGU significantly reversed the de-crease of intracellular ATP content,the increase of Ca2+fluorescent intensity and concentration and the decrease of MMP.The release of cytochrome C,the expressionsof caspase-9 and caspase-3 in H9C2 cardiomyocytes were increased 16 h after OGD.LGUsignificantly inhibited the changes of these apop-tosis-related proteins. CONCLUSION LGU has a significant protective effect against OGD-induced H9C2 cardiomyocytes injury through inhibiting calcium overload,increasing ATP content,improving mi-tochondrial function and inhibiting apoptosis.

Objective To explore the relationship between plasticity ofhippocampus neuronal morphology and pathogenesis of epilepsy by observing the changes of mossy fiber sprouting (MFS) and P-glycoprotein (P-gp) expression in the hippoeampus of rat models of amygdala-kindling epilepsy.Methods Ninety male Wistar rats were randomly divided into epilepsy model group (n=40),drug treatment group (n=40) and sham-operated group (n=10).Models of chronic epilepsy were established by stimulating the amygdale; rats in the drug treatment group were perfused antiepileptic drug levetiracetam into stomach [100 mg/(kg·d),twice daily].At different observation time points (1,2,4 and 8 weeks after the treatment),Timm staining was employed to observe the changes of MFS; immunohistochemical method was used to detect the dynamic changes of P-gp.Results (1) After the success of model making,abnormal MFS levels in the hippocampal CA3 transparent layer were noted; lowest MFS scores were noted in S1 subgroup,which showed no significant difference as compared with those in sham-operated group (P＞0.05); the MFS scores in S2,S4 and S8 subgroups increased gradually with a peak level at 8 weeks after inducement,which showed significant difference as compared with those in sham-operated group (P＜0.05).No significant differences on MFS grading scores were noted in the drug treatment group between each two time points (P＞0.05).(2) The P-gp expression in the epilepsy model group showed significant difference at different time points (P＜0.05); highest expression level was noted at one week after the surgery,and returned to normal at four weeks.However,the P-gp expression showed no significant difference in the drug treatment group between each two time points (P＞0.05),except for 1 week after treatment.Conclusion MFS is the important mechanism of chronic epilepsy,and P-gp is the product of epilepsy,which is the main reason of epilepsy drug resistance.

BACKGROUNDLack of fluoroscopic landmarks can make valve deployment more difficult in patients with absent aortic valve (AV) calcification. The goal of this article was to evaluate the feasibility and effectiveness of transcatheter implantation of a valved stent into the AV position of a goat, assisted with a microcatheter which provides accurate positioning of coronary artery ostia to help valved stent deployment.

METHODSThe subjects were 10 healthy goats in this study. A microcatheter was introduced into the distal site of right coronary artery (RCA) through femoral artery sheath. A minimal thoracic surgery approach was used to access the apex of the heart. The apex of the left ventricle was punctured; a delivery catheter equipped with the valved stent was introduced over a stiff guidewire into the aorta arch. We could accurately locate the RCA ostia through the microcatheter placed in the RCA under fluoroscopy. After correct valve position was confirmed, the valved stent was implanted after rapid inflation of the balloon. The immediate outcome of the function of the valved stents was evaluated after implantation.

RESULTSAll ten devices were successfully implanted into the AV position of the goats. Immediate observation after the procedure showed that the valved stents were in the desired position after implantation by angiography, echocardiogram. No obstruction of coronary artery ostia occurred, and no moderate to severe aortic regurgitation was observed.

CONCLUSIONSWhen the procedure of transcatheter implantation of a balloon-expandable valved stent into the AV position of goats is assisted with microcatheter positioning coronary artery ostia, the success rate of operation can be increased in those with noncalcified AV.

Animals ; Aortic Valve ; surgery ; Female ; Goats ; Heart Valve Prosthesis Implantation ; methods ; Male ; Transcatheter Aortic Valve Replacement ; methods

OBJECTIVETo study the distribution of human immunodeficiency virus-1 (HIV-1) genotypes in Hubei province.

METHODSEpidemiological survey was carried out to HIV-1 carriers who were identified in Hubei province. HIV-1 env V3-V4, gag P17/24 and the first exon of tat region were amplified by nested-polymerase chain reaction(nPCR) .The sequences were determined, and phylogenetic analyses were then performed.

RESULTS4 HIV-1 strains or circulating recombinant forms (CRFs) were identified in Hubei province with subtype B' the predominant which covered 5 kinds of populations including former blood donors, blood receivers, spouses of the infected people, sex workers and their clients, homosexuals, mainly distributed in the areas with many former blood donors. CRF08-BC and CRF01-AE were found distributed in economically more developed cities or southern area of the province, and the major transmission routes was through sexual contact. Only 1 patient, an injecting drug user, was identified having subtype C.

CONCLUSIONSubtype B' was the main epidemic subtypes in Hubei province while CRF08-BC, CRF01-AE and subtype C were also circulating in the province, indicating the transmission of the disease might to become more complex.

China ; epidemiology ; HIV Infections ; epidemiology ; HIV-1 ; classification ; Humans ; Molecular Epidemiology ; Phylogeny ; RNA, Viral ; genetics ; Sequence Analysis, RNA

To investigate the expression of telomerase in cord blood hematopoietic stem/progenitor cells during their committed differentiation in vitro and provide an index of monitoring the proliferating potential of the hematopoietic stem/progenitor cells and security for clinical application. Human CD34 positive cells were isolated from umbilical cord blood by using magnetic cell sorting system (MACS), and were induced to differentiation with hematopoietic growth factors (SCF + IL3 + IL6 + GCSF and SCF + IL3 + IL6 + EPO) in a liquid culture system. The telomerase activity and the cytalytic subunit of telomerase (hTERT) of the cells were analysed during different periods of culture by using TRAP-PCR, TRAP-ELISA, Western blot and RT-PCR techniques, respectively. The results showed that a peak of cell growth was achieved on day 14 - 21 during induction of differentiation in vitro. Total cell number could increase 1006.4 +/- 103.2 times and could not increase there after. Telomerase activity and hTERT expression were low in freshly isolated cord blood CD34(+) cells and increased after about 7 days of culture in addition of cytokine combinations of SCF + IL3 + IL6 + GCSF and SCF + IL3 + IL6 + EPO, respectively. The telomerase activity and hTERT decreased after 14 days of culture and were not detected after 28 days of culture. It was concluded that the hematopoietic stem/progenitor cells can be expanded in large number in vitro and do not have the character of immortality and the telomerase activity could be a useful index in hematopoiesis regulation.
Antigens, CD34 ; blood ; Blotting, Western ; Cell Differentiation ; DNA-Binding Proteins ; Enzyme-Linked Immunosorbent Assay ; Fetal Blood ; cytology ; Hematopoietic Stem Cells ; cytology ; enzymology ; Humans ; Polymerase Chain Reaction ; RNA, Messenger ; analysis ; Telomerase ; genetics ; metabolism

To evaluated the feasibility of preventing infection after high dose chemotherapy and radiotherapy using the granulocytes derived from differentiated from hematopoietic stem/progenitor cells ex vivo, human CD34-positive cells were isolated from umbilical cord blood by using a high-gradient magnetic cell sorting system (MACS), and the cells committedly differentiated with hematopoietic cytokines (SCF + IL-3 + IL-6 + G-CSF) in a liquid culture system. The expanded cell number, ratio of the viable cells, chromosome and phenotype of the differentiated cells and safety analysis of expanded cells were detected by using cell count, trypan blue exclusion test, karyotype analysis, flow cytometry and tumorigenic model of nude mice, respectively. The results showed that the combination of cytokines increased cell number by (1006.4 +/- 103.2) folds and flow cytometric analysis showed myeloid marker CD11b expressed in the about 60% cells. The growth peak of differentiated cells was at 14 days of culture and decreased at about 33 days. No abnormality was found in the karyotype analysis of expanded cells. No tumor was found in the nude mice injected with expanded cells after 35 days and the expanded cells had the ability of phagocytizing bacteria. It is concluded that the cells, differentiated from CD34(+) cells, expanded ex vivo possess the function of granulocyte and it was safe for clinical trial.
Animals ; Antigens, CD34 ; analysis ; Cell Differentiation ; Fetal Blood ; cytology ; Granulocytes ; cytology ; immunology ; Hematopoietic Stem Cells ; cytology ; Humans ; Karyotyping ; Mice ; Mice, Inbred BALB C ; Phagocytosis