Aim: A new derivative LC-MS method was developed for the simultaneous determination of zofenopril and its active metabolite zofenoprilat to investigate the pharmacokinetic characteristics of zofenopril and zofenopri-lat in healthy Chinese volunteers after single and multiple oral doses of zofenopril calcium tablets. Methods: Ten Chinese healthy volunteers were given three single oral doses of 15,30, and 60 mg, respectively, and consecutively the multiple doses of 30 mg. The concentration and pharmacokinetic parameters of both the parent drug and the active metabolite were simultaneously determined by derivative LC-MS method using p-bromophenacyl bromide (p-BPB) as the derivative reagent. Results: After the single oral administrations of 15, 30, and 60 mg of zofeno-pril calcium, there was no significant difference in the t_(1/2) of both zofenopril and zofenoprilat among the three do-ses. The values of AUC_(0-24h) and c_(max) for both zofenopril and zofenoprilat showed the good linearities to the dosage over the dose range from 15 mg to 60 mg. There were no significant differences in AUC_(0-24h) and c_(max) for both com-pounds between female and male volunteers. After multiple oral administration( 30 mg once daily for 6 days ), the average steady state plasma concentration( c_(av)) for zofenopril was (5. 07 ±1. 06) ng/mL with the degree of fluctu-ation (DF) of 14. 26 ± 2. 94. The c_(av) for zofenoprilat was (6. 28 ± 1. 87) ng/mL with the DF of 11. 61 ±4. 68. The accumulation index values for zofenopril and zofenoprilat were 0. 94 ± 0. 31 and 0. 83±0. 13, respec-tively. Conclusion: Both zofenopril and zofenoprilat were demonstrated of linear kinetics after single administra-tion and showed no accumulation after multiple administration of the test zofenopril calcium tablets. There was significant difference in the pharmacokinetic characteristics for zofenopril calcium between healthy Chinese and European volunteers.

The 1.23 kb DNA fragment encoding the early protein EP0 of pseudorabies virus (PRV) Ea strain was amplified by PCR technique and cloned into pBluescriptII sk+.Three sequencing plasmids containing the partial fragment of the EP0 gene were constructed and the sequences were obtained by Sanger's sequencing technique. Compared with PRV InFh strain, there were multipile site-mutations and a deleted-mutation in the EP0 gene of PRV strain Ea，and the diversity of amino acid residues also existed.Then, the EP0 gene was inserted into an expression vector, pET-28a, fused into the downstream of the 6ΧHis-Tag in frame, to yield the expression plasmid pETEP0. After induction by IPTG, a high expression of fusion protein was obtained, SDS-PAGE analysis and Western blotting showed that the fusion protein was 62kD and the protein was specific to antisera against PRV Ea strain. This indicated that the EP0 gene be expressed in BL21(DE3) and the expression products have immuno-genicity.

Objective This study is aimed at determining whether anti-von Willebrand factor (VWF) autoantibodies are present in the plasma of idiopathic thrombotic thrombocytopenic purpura (TTP) patients with normal ADAMTS13 activity and undetectable anti-ADAMTS13 antibodies,and at examining whether murine monoclonal antibodies (mAbs) against human VWF decrease the susceptibility of VWF to ADAMTS13 in vitro.Methods Anti-VWF autoantibodies and ultralarge VWF (UL-VWF) multimers were measured in plasma samples of 53 adult patients with idiopathic TTP by enzyme-linked immunosorbent assay and sodium dodecylsulphate-agarose gel electrophoresis,respectively.Moreover,the effects of eight murine mAbs to different human VWF domains on VWF cleavage by ADAMTS13 were evaluated under fluid shear stress and static/denaturing conditions,respectively.Results Anti-VWF antibodies and UL-VWF multimers were detected in two TTP patients with normal ADAMTS13 activity and undetectable anti-ADAMTS13antibodies.The SZ34,an anti-VWF mAb,inhibited VWF proteolysis mediated by ADAMTS13 under flow,but not static conditions.Conclusion Anti-VWF antibody may be one of the causes of idiopathic TTP with normal ADAMTS13 activity and undetectable anti-ADAMTS13 antibodies.

Objective:To observe the efficacy and safety of dexzopiclone plus auricular acupressure in intervening primary insomnia.Methods:A total of 72 participants who met the inclusion criteria were enrolled in a randomized controlled trial,with 36 cases allocated to a treatment group and 36 cases allocated to a control group.Both groups were given dexzopiclone as the routine treatment.Patients in the treatment group were given auricular acupressure with Wang Bu Liu Xing (Semen Vaccariae) seeds at the auricular acupoints related to sleep and emotion based on meridian theory,whereas for patients in the control group,the medical plasters with Wang Bu Liu Xing (Semen Vaccariae) seeds were only gently stuck to acupoints unrelated to sleep without stimulation.Patients in both groups were required to visit the hospital once a week for replacing the seeds and plasters.The course of intervention lasted for 8 weeks and the patients were followed up for another 4 weeks.Pittsburgh sleep quality index (PSQI) and Karolinska sleep diary (KSD) were used to evaluate the outcomes.Meanwhile,adverse effects were monitored and recorded.Results:In the enrolled 72 cases,4 patients (one in the treatment group and three in the control group) reported thirst and a bitter taste,and one case in the control group reported nausea and vomiting.At last,3 cases in the control group dropped out for adverse reactions,and 69 cases completed the clinical trial.After 8 weeks of treatment,the global scores of PSQI in both treatment and control groups decreased significantly compared with the baseline (both P＜0.001).Furthermore,the global score of PSQI in the treatment group was lower than that in the control group (P＜0.01).The global scores of PSQI in both groups at the follow-up were significantly different from the baseline (both P＜0.001),but insignificantly different compared with the post-treatment results (both P＞0.05).According to KSD,both treatment protocols could prolong the total sleep time,shorten sleep-onset latency,improve sleep efficacy and sleep quality significantly,and the changes in the treatment group were more significant.The total effective rate was 88.9％ in the treatment group,higher than 81.8％ in the control group,though the difference was statistically insignificant (P＞0.05).Conclusion:Dexzopiclone plus auricular acupressure is effective and safe for patients with primary insomnia both in short and long terms,and it is more effective than monotherapy of dexzopiclone.

Objective This study aimed to evaluate the effects of in-utero exposure to HIV and ART on pregnancy outcome and early growth of children.Methods This cohort study enrolled 802 HIV-infected pregnant women between October 2009 and May 2018 in Guangzhou,China The women were assigned to receive combination ART (cART) or mono/dual ART or no treatment.The primary outcomes were the combined endpoints of any adverse pregnancy outcome [including ectopic pregnancy,spontaneous abortion,stillbirth,preterm birth,small for gestational age (SGA)] and adverse early growth outcome (including infant death,HIV infection of mother-to-child transmission,and underweight,wasting and stunting of infants at 4 weeks of age).Results Adverse pregnancy outcomes occurred in 202 (35.1％) of all enrolled HIV-infected women,and 121 (31.3％) of all infants exhibited adverse effects on early growth at 4 weeks of age.The rates of adverse pregnancy outcomes,spontaneous abortion,ectopic pregnancy,stillbirth,infant death and perinatal HIV infection were higher among women not receiving ART,compared to those treated with cART or mono/dual ART (P ＜ 0.05).However,women treated with cART had a higher rate of SGA,compared to untreated women (P ＜ 0.05).No differences in early infant growth were observed among the different treatment regimens.Conclusion Our findings underscore the essentiality of prioritizing HIV-positive pregnant women for ART,as even mono/dual ART available in resource-limited countries could improve pregnancy outcomes and infant survival

BACKGROUND: The area of myocardial infarction is the determinative factor of acute myocardial infarction prognosis. Amelioration of blood transportation and replacement therapy can reduce infarction area. Bone marrow mesenchymal stem cells can differentiate into cardiovascular tissue and are easy to obtain. After cultured and expanded in vitro, they can become the ideal cells for cardiovascular replacement therapy.OBJECTIVE: To evaluate the therapeutic effect of intracoronary transplantation of bone marrow mesenchymal stem cells in the treatment of myocardial infarction. DESIGN: Self-control observation taking the patients as subjects.SETTING: Department of Cardiology, Department of Nuclear Medicine,Echocardiogram Room, Nanjing First Hospital Affiliated to Nanjing Medical University.PARTICIPANTS: Totally 20 patients with acute myocardial infarction who received the therapy of bone marrow mesenchymal stem cells transplantation in the Department of Cardiology, Nanjing First Hospital Affiliated to Nanjing Medical University during March 2003 to March 2004 were recurited. Informed consents were obtained from the patients, and the complete postoperative follow up was over 3 months. The patients include 15 male and 5 female, and they were aged (64±10) years.METHODS: All the patients underwent percutaneous coronary intervention (PCI) to treat infarction-related blood vessel. Autologous bone marrow was taken from the patients, then stem cells were extracted to be performed in vitro induction, differentiation and proliferation, and transplanted infarction-related blood vessel through coronary artery at the mean number of (21.7±30.14)× 107 within 2 weeks. Before and 3 months after transplantation of stem cells, patients underwent gated dual-isotopic myocardial perfusion/metabolic imaging (18-fluoro-2-deoxy-glucose, 18F-FDG) examination. Survived and necrotic myocardia were predicted and infarction area was obtained. At the same time, wall motion and heart function index were evaluated with ultrasound cardiography (UCG)examination, and they were re-checked 3 months after operation to evaluate the amelioration of wall motion and heart function index. A 5-point scale was used in the evaluation of gated dual-isotopic myocardial perfusion/metabolic imaging (18F-FDG) examination: point 0: normal, 1: sparse, 2:obviously sparse, 3: defected. Evaluative standard of UCG: point 1: normal,2: reduced, 3: obviously reduced, 4: no ventricular wall motion or paradoxical motion; Wall motion with 2 points or more than 2 points suggests it is improved.MAIN OUTCOME MEASURES: ① Results of gated dual-isotopic myocardial perfusion/ metabolic imaging (18F-FDG-SPECT); ②Infarctionrelated myocardial segment score and heart function index before and after stem cell transplantation of patients in ECG follow-up observation.RESULTS: All the 20 patients participated in the result analysis.Results of gated dual-isotopic myocardial perrusion/metabolic imaging (18F-FDG-SPECT): The myocardial perfusion defect area of 20 patients was significantly reduced after therapy than before therapy [(33±15)%,-(44±18)% ,P ＜ 0.05]; Metabolie defect area was significantly reduced after therapy than before therapy [(33±17)%, (43±21)% ,P ＜ 0.05];Before therapy, there were 199 segments, in which blood flow reperfusion was matched to glycometabolism defect, and they were determined as necrotic myocardium. After therapy, blood flow perfusion metabolism was improved in 79 segments, but blood flow perfusion and glycometabolism were not improved significantly in 120 segments (P ＜ 0.05). Results of UCG: ejection fraction of patients was significantly larger after therapy than before therapy [(53±8)%, (42±7)% ,P ＜ 0.05].CONCLUSION: Intracoronary transplantation of human bone marrow mesenchymal stem cells for treating myocardial infarction is simple to operate. After therapy, the infarction area is obviously reduced, myocardial blood flow perfusion and metabolism of necrotic area improve, myocardial segments without survival determined before operation reduce sigrificantly and the heart function of patients improve.

Adolescent ; Amino Acid Metabolism, Inborn Errors ; complications ; Child ; Female ; Humans ; Male ; Mental Disorders ; etiology

This study was aimed to investigate the effect of CD44 gene silence on the drug resistance and biologic activity of human multidrug resistant leukemia cell line K562/A02. The oligonucleotides of CD44 gene were designed according to related data of GenBank, double-stranded DNA was produced by annealing, and was inserted into pGCsilencerU6/Neo/GFP vector. The resultant recombinant plasmid pGCsiRNA-CD44 was transfected into K562/A02 cell line. Expressions of CD44, mdr-1 and blc-2 mRNA were assayed by real time RT-PCR. The 50% inhibitory concentration (IC50) of doxorubicin (ADM) for K562/A02 cell line was determined by MTT method. Cell cycle was determined by flow cytometry. The morphology of apoptotic cells was examined by Hochst 33258 staining. The results indicated that the siRNA eukaryotic plasmid directing at CD44 gene could effectively silence the CD44 gene of K562/A02 cells; as compared with control group, the CD44 expression in K562/A02 cells transfected with 4 pGCsiRNA-CD44 plasmids was obviously inhibited, while the inhibition of CD44 expression in cells transfected with siCD44-1 was strongest. After being transfected with pGCsiRNA-CD44, the expression of CD44 mRNA in K562/A02 cells reduced by 64.1% (p<0.05), at the same time the expression of mdr-1 and bcl-2 mRNA in pGCsiRNA-CD44-transfected K562/A02 cells reduced by 25.6% and 50.8% respectively. IC50 of K562/A02 cells after transfection decreased to (8.77+/-1.63) microg/ml and was obviously lower than that of control (17.97+/-1.61) microg/ml (p<0.01). After transfection for 48 hours, the ratio of K562/A02 cells in G0/G1 increased by 10.7%, and the cells displayed karyopyknosis, nuclear margination and apoptotic bodies. It is concluded that the siRNA plasmid specifically targeting CD44 gene can remarkably down-regulate the expression of CD44 gene, inhibit K562/A02 cell proliferation, induce its apoptosis and effectively reverse the multidrug resistance of K562/A02 cells.
Apoptosis ; Cell Line, Tumor ; Cell Proliferation ; Drug Resistance, Multiple ; genetics ; Drug Resistance, Neoplasm ; genetics ; Gene Silencing ; Genetic Vectors ; Humans ; Hyaluronan Receptors ; genetics ; K562 Cells ; RNA, Small Interfering

Objective To explore the diagnostic and score value of ultrasound on hemophiliac arthropathy referring to MRI on the diagnosis and score of hemophiliac arthropathy Methods The ultrasound and MRI examinations were performed on 42 joints of 42 hemophilia patients 14 knees 14 ankles and 14 elbows The consistency of ultrasound and magnetic resonance imaging in the detection and score of joint diseases was compared Finally inter-and intra-observer agreement of ultrasound scoring system were tested Results The consistency of ultrasound and magnetic resonance imaging was excellent κ=0 763-0 896 P < 0 001 in the detection of early soft tissue lesions effusion or hemarthrosis synovial hypertrophy hemosiderin excellent κ=0 793 P <0 001 in the detection of cartilage loss poor κ=0 133 P = 0 132 in the detection of erosions and poor κ= 0 100 P = 0 137 in the detection of subchondral cysts The consistency of ultrasound and magnetic resonance imaging was good to excellentκ=0 684-0 833 P < 0 001 in the score of early soft tissue lesions effusion or hemarthrosis synovial hypertrophy and hemosiderin and poor to good κ=0 145 -0 635 P <0 001 in the score of advanced osteochondral lesions cartilage loss and bone erosions The inter-observer agreement was good to excellent κ=0 676-0 870 P <0 001 for early soft tissue lesions and moderate to excellent κ=0 421- 0 75 1 P < 0 001 for advanced osteochondral lesions The intra-observer agreement was good to excellent κ=0 705-0 885 P <0 001 for early soft tissue lesions and moderate to good κ=0 532 -0 732 P <0 001 for advanced osteochondral lesions Conclusions Ultrasound plays an important role in detecting early soft tissue changes effusion or hemarthrosis synovial hypertrophy hemosiderin and cartilage loss which helps follow-up and guide clinical treatment.

OBJECTIVETo investigate the effect of DNA methylation in combination with histone deacetylase inhibitor on transcription regulation of Ras associated domain family gene 1(RASSF1A) tumor suppressor gene and the molecular biological behaviors in U266 cells.

METHODSThe U266 cells were treated with different doses of 5-Aza-2'-deoxycytidine (5-Aza-CdR) and Valproate (VPA) each alone or in combination. Methylation-specific PCR (MSP) was used to detect CpG island methylation in RASSF1A promoter. Quantitative real-time reverse transcription polymerase chain reaction (RQ-PCR) was used to examine the expression of RASSF1A gene in U266 cells. MTT was used for cell proliferation. Cell apoptosis and cell cycle were analyzed by flow cytometry.

RESULTSThe methylation of RASSF1A gene promoter was detected in U266 cells, while there was little RASSF1A gene expressing in the control group. The demethylation effect could be detected in the 5-Aza-CdR treated and combined treatment groups but no in the VPA group. The expression level of RASSF1A was induced by 5-Aza-CdR in a concentration-dependent manner while VPA had no such effect. The expression level of RASSF1A mRNA was increased significantly in the combined treatment group. Higher growth inhibition and apoptosis effects were found in 5-Aza-CdR and VPA combination group than that in 5-Aza-CdR or VPA alone group (P < 0.05). After treatment with 5-Aza-CdR or VPA alone for 72 h, more cells were arrested in G(0)/G(1) phase as conpared with control group (P < 0.05), and even more cells were so arrested in combined treatment group (P < 0.05).

CONCLUSIONDNA methylation and histone deacetylase inhibitor can synergistically induce demethylation of the RASSF1A gene, re-express RASSF1A gene silenced in U266 cells, inhibit the proliferation of U266 cells and induce cell apoptosis.