Objective:To establish a rapid detection method for human astrovirus based on TaqMan-probe real-time fluorescent quantitative reverse transcription-polymerase chain reaction (RT-PCR).Methods:According to the conservative sequence of human astrovirus ORF1 b gene, we designed the amplification primers and specific fluorescent probe to establish the human astrovirus TaqMan real-time fluorescent quantitative RT-PCR rapid detection method. The specificity, sensitivity and stability of the method were evaluated. We also used this method to detect human astrovirus in clinical samples. Results:The established human astrovirus TaqMan real-time fluorescent quantitative RT-PCR detection method has good specificity and repeatability for human astrovirus, and the sensitivity can reach 10 2 copies/μl. After testing the clinical samples, the detection rate of human astrovirus by our method was 100%. Conclusions:The human astrovirus TaqMan real-time fluorescent quantitative RT-PCR detection method established in this study is simple, rapid, sensitive, specific and stable. It can be used for clinical human astrovirus detection and epidemiological investigation.

Objective To study application of Chinese general practice assessment questionnaire (GPAQ)consultation version 2.0 in evaluating patients' satisfaction with community health-care service (CHS).Methods First,origihal GPAQ consultation version 2.0 in English was translated into Chinese language by three experts in general practice/family medicine field,then translated it back into English,finally a Chinese version of GPAQ was formed.Ten medical doctors who were on duty on 25 April,2008 at each out-patient clinic from CHS of Daxing,Nancaiyuan,Yongning and Hancunhe were selected randomly as targets for evaluation of service satisfaction.A sample of the first 30 patients(1200 in total)aged more than 16 years who visited the clinic on that day were selected to fill out the Chinese GPAQ consultation version 2.0 to express their satisfaction with the medical doctors they visited,and reliability and validity of the questionnaire was then evaluated.Results Cronbach's alpha ranged 0.734 to 0.813 for each domain of the Chinese version of GPAQ2.0.and cumulated contribution of three common factors with the eigen-value more than one reached 58.722%.Scores of patients' satisfaction withreceptionists,access,continuity,doctor's communication skills andpatient enablement were significantly lower,as compared to those of national bench-mark in the United Kingdom,with t-values of -6.397,-11.729,-6.328,-59.871 and -2.210,and all P＜0.05.respectively.Conclusions The Chinese version of GPAO 2.0 achieves good reliability and modcrate validity,and can be used as an instrument for evaluating patient's satisfaction with quality of CHS in China as further revision in the future studies.

Objective To learn the quality of life (QOL) and mental health status of the physically disabled people in Zhejiang Province,in order to provide evidence for improving the quality of life and mental health of the physically disabled people.Methods Using questionnaires and interviews to make household surveys of 950 physically disabled people.The questionnaire included the brief quality of life scale (SF-36) and the general health questionnaire (GHQ-28) in the anxiety and depression scale,physical and mental health status.The basic information and other living conditions of physically disabled people were collected through interviews.Results A total of 907 valid questionnaires were recovered,and the effective response rate was 95.47％.Scores of QOL of 907 physically disabled people was 60.89,and anxiety and depression scores were more than 4 points and there was a high score on anxious and depression.The scores of quality of life,anxiety and depression were not statistically significant between males and females(P ＞0.05).There were significant differences in all dimensions of SF-36 and anxiety and depression in different levels of physically disabled people(P ＜ 0.01).There was a significant negative correlation between the quality of life and anxiety and depression in physically disabled people(rs =-0.626、-0.639,P ＜ 0.01).The multiple linear regression showed that physiological function,role physical,physical pain and general health were the influencing factors of anxiety of the physically disabled people,and role physical,physical pain and general health were the influencing factors of depression of the physically disabled people.Condusion The quality of life of physically disabled people were comparably poor,and presented anxiety,depression and other psychological problems.Physical function status directly affects the quality of life and mental health status.

Objective To explore the effects of disease adaptation intervention in young and middle-aged patients undergoing early stage of peritoneal dialysis.Methods Using convenience sampling method,a total of 88 patients undergoing early stage of peritoneal dialysis were assigned into the experimental group and the control group from the inpatient ward of Nephrology Department of a tertiary hospital in Zhengzhou City.The experimental group received the comprehensive adaptation intervention and routine care,and the control group only received routine care.Before and after the intervention,the Psychosocial Adjustment to Illness Scale(PAIS-SR) and Short Form of 36 health survey questionnaire (SF-36) were used to assess the disease adaptability and quality of life of two groups,and the degree of edema and serum albumin index of two groups were recorded.Results Before and after the intervention,there were significant differences in the scores of psychosocial adaptation and each dimension,the scores of physical health and mental health in the experimental group,and the differences were statistically significant (P＜ 0.05);there were significant differences in the scores of psychosocial adaptation and each dimension in control group,and the differences were statistically significant(P＜0.05),but the differences of the scores of physical health and mental health were not statistically significant(P＞0.05).After the intervention,there were significant differences in the scores of psychosocial adaptation and each dimension,the scores of physical health and mental health between the two groups(P＜0.05);but the differences of serum albumin and edema index of two groups were not statistically significant(P＞0.05).Conclusion Comprehensive adaptation intervention can improve the ability of disease adaptation and the quality of life in young and middle-aged patients undergoing early stage of peritoneal dialysis.

Objective:To analyze the availability ofpediatricessential medicines in China, and to provide reference and suggestions for the further responses to the demand for pediatric medication in China. Methods :Using survey data collected from 19 provinces in China through the cooperation of the National Health Commission and our research group, the availability of pediatricessential medicines in China was analyzed from the perspectives of drug use and a-vailability with its influencing factors. Results :The survey results showed that the availability of pediatricessential medicines in China is generally low. The shortage in such medicines does not occur in individual regions, individual medical institutions or individual treatment areas, but it is still general in the surveyed parts of the country. The two fundamental reasons for the noticed shortage are :the research on special varieties for Children is expensive and the pricing mechanism of suitable varieties is inappropriate, which results in insufficient motivation for R&D Enterprises. Conclusions :This article suggests that the existing national pediatric medication support policies should be put in place as soon as possible, andbe further implemented for nationwide pediatric consumption. It is also recommended that health, drug supervision, bidding, pricing, medical insurance and other departments should work together to make more preferential policies to help enterprises develop special pediatric drugs to further promote the supply and respond to the demand of medicine for children in China.

Lysine decarboxylase is a key enzyme involved in the upstream biosynthesis of lycopodium alkaloids (LAs) such as huperzine A, contributing to the decarboxylation of lysine to 1,5-pentanediamine (cadaverine). Three lysine decarboxylase genes (HsLDC-L1, HsLDC-L2, HsLDC-L3) were successfully cloned from Huperzia serrata using transcriptomic sequence data mining strategy combined with reverse transcription PCR. The physicochemical properties, secondary and tertiary structures, amino acid identities, and evolutionary relationship of the three LDCs were analyzed by online bioinformatics analysis platforms and DNAMAN, MEGA 7.0 software, revealing that all of these proteins had the conserved PLP binding domain and active site residues were completely conserved in LDCs. Phylogenetic analysis showed that these LDCs were located in the same branch as other known LDCs from LA-producing plants. Accordingly, the ORFs of these three HsLDCs were inserted into different expression plasmids for further expression in E. coli. However, only HsLDC-L1 was successfully expressed in E. coli BL21 (DE3) by inserting into a pCold TF vector. The recombinant protein was purified by Ni²⁺ affinity chromatography purification. HsLDC-L1 contains 469 amino acid residues, with a calculated molecular weight of 50.50 kDa. HsLDC-L1 expectedly catalyzed the decarboxylation of lysine to produce cadaverine. In addition, HsLDC-L1 can also catalyze the generation of putrescine from ornithine. However, it cannot catalyze the decarboxylation of tyrosine, phenylalanine, tryptophan and histidine. The results not only provide insight into the biosynthesis of LAs including huperzine A, but also provide a critical genetic element for the overproduction of Δ¹-piperideine and pelletierine, the essential biosynthetic precursors of LAs, using synthetic biology strategies.

Objective To understand microcystins (MCs) pollution in important waters of Shaoxing, so as to provide evidence for risk assessment and supervision of water quality safety. Methods Water samples were collected from 15 points set in three Shaoxing waters during September 2016 to September 2017. Six kinds of MCs (MC-LR, MC-RR, MC-YR, MC-LA, MC-LY, MC-LF) in samples were detected by ultra performance liquid chromatography tandem mass spectrometry, and the results were compared according to different time and place. Results MCs were detected positive in 22 of the 135 samples , the total detection rate was 16.3％.The detection rates of Xiaoshao River, Puyang River and Cao′e River were 7.9％, 18.5％和26.7％ respectively; the detection rates in July, August and September were 13.3％, 40％ and 46.7％ respectively, and the other months were not detected. In 22 positive samples, the detection rates of 6 MCs from high to low as follows: MC-LR (100％), MC-RR (100％), MC-YR (22.7％), MC-LY (18.2％), MC-LF (13.6％) and MC-LA (0％) . Conclusion In the three important waters of Shaoxing, microcystins were polluted in summer and autumn, which were mainly MC-LR and MC-RR.

OBJECTIVETo study the influence of anastomoses sequence on the hemodynamics in off-pump coronary artery bypass grafting (CABG), and to investigate the strategy to stabilize the hemodynamics.

METHODSFrom March 2005 to March 2007, 67 patients with serious triple-vessel coronary artery lesions (male 45, female 22) with a age range from 44 to 81-years-old were enrolled for off-pump CABG. All the patients underwent left internal mammary artery-left anterior descending branch (LIMA-LAD) anastomose firstly, followed by the foreword anastomose of the other two vessels. According to the anastomose sequence of posterior descending branch (PDA) and obtuse marginal branch (OM), the patients were divided into three groups. Group I (n = 22) did the sequence of PDA-OM-aortic root (Ao). Group II (n = 14) did the sequence of Ao-PDA-OM. Group III (n = 31) did the sequence of Ao-OM-PDA. The hemodynamics markers, including heart rate (HR), central venous pressure (CVP), mean artery pressure (MAP), cardiac index (CI), left ventricular-stroke work index (LVSWI) and right ventricular stroke work index (RVSWI), were analyzed before LIMA-LAD anastomose to serve as baseline. And these markers were re-measured at LIMA-LAD anastomose, immediately after LIMA-LAD anastomose, at PDA anastomose and at OM anastomose.

RESULTSAll the patients survived after off-pump CABG. Compared with the baseline, there were no significant hemodynamic changes when doing LIMA-LAD anastomose. When the anastomose finished, the hemodynamic indices improved significantly. When performing OM anastomose, HR and CVP increased significantly, and MAP, CI, LVSWI and RVSWI decreased remarkably in group I and group III, while there were no significant changes of MAP, CI and RVSWI in group II . When performing PDA anastomose, HR and CVP increased significantly, and MAP, CI, LVSWI and RVSWI decreased remarkably in group I and group II, while there were no significant hemodynamic changes in group III except HR.

CONCLUSIONProximal anastomose first when performing off-pump CABG, followed by distal anastomoses of target vessels is beneficial to those who have hard exposure and difficult anastomose of OM and PDA.

Adult ; Aged ; Aged, 80 and over ; Anastomosis, Surgical ; methods ; Coronary Artery Bypass, Off-Pump ; methods ; Coronary Artery Disease ; physiopathology ; surgery ; Coronary Vessels ; surgery ; Female ; Hemodynamics ; Humans ; Male ; Middle Aged ; Monitoring, Intraoperative ; Retrospective Studies

BACKGROUNDThe production of neural stem cells (NSCs) derived from embryonic stem (ES) cells was usually very low according to previous studies, which was a major obstacle for meeting the needs of clinical application. This study aimed at investigating whether astrocytes could promote production of NSCs derived from ES cells in vitro.

METHODSMouse ES cells line-D3 was used to differentiate into NSCs with astrocytes as inducing stromal cells by means of three-stage differentiation procedure. Another group without astrocytes served as control. The totipotency of ES cells was identified by observation of cells' morphology and formation of teratoma in severe combined immunodeficiency disease (SCID) mice. The quantity and purity of NSCs derived from ES cells were analyzed using clonogenic assay, immunohistochemical staining and flow cytometry assay. The plasticity of NSCs was detected by differentiating test. Octamer-binding transcription factor 4 (Oct-4) and nestin, the specific marker genes of ES cells and NSCs respectively, were detected continuously using reverse transcription-polymerase chain reaction (RT-PCR) method to monitor the process of cell differentiation.

RESULTSThe ES cells of D3 line could maintain the ability of differentiating into cellular derivations of all three primary germ layers after continuous passage culture. At the end of two-stage of inducing process, 23.2 +/- 3.5 neurospheres per plate formed in astrocyte-induced group and only 0.8 +/- 0.3 per plate in the control group (clonogenic assay, P < 0.01), and the ratio of nestin positive cells was (50.2 +/- 2.8)% in astrocyte-induced group and only (1.4 +/- 0.5)% in the control group (flow cytometry, P < 0.01). With the induction undergoing, the expression of Oct-4 gradually decreased and then disappeared, while the expression of nestin was increased step by step, and the ratio of nestin positive cells was up to 91.4% by the three-stage differentiation. The nestin positive cells could be further induced into neurons, astrocytes, and oligodendrocytes in differentiating medium supplemented with fetal calf serum. The results of differentiating test showed that the ratio of NF-200 and NSE positive cells was (42.7 +/- 2.6)% in astrocyte-induced group and only (11.2 +/- 1.8)% in the control group (P < 0.01).

CONCLUSIONSAstrocytes can not only increase the production of NSCs derived from ES cells but also promote the differentiation of NSCs toward neuronal lineage.

Animals ; Astrocytes ; physiology ; Cell Differentiation ; Cell Lineage ; Cells, Cultured ; Embryo, Mammalian ; cytology ; Mice ; Neurons ; cytology ; Stem Cells ; cytology

Herein, solid lipid nanoparticles (SLN) were proposed as a new drug delivery system for adefovir dipivoxil (ADV). The octadecylamine-fluorescein isothiocynate (ODA-FITC) was synthesized and used as a fluorescence maker to be incorporated into SLN to investigate the time-dependent cellular uptake of SLN by HepG2.2.15. The SLN of monostearin with ODA-FITC or ADV were prepared by solvent diffusion method in an aqueous system. About 15 wt% drug entrapment efficiency (EE) and 3 wt% drug loading (DL) could be reached in SLN loading ADV. Comparing with free ADV, the inhibitory effects of ADV loaded in SLN on hepatitis B surface antigen (HBsAg), hepatitis B e antigen (HBeAg) and hepatitis B virus (HBV) DNA levels in vitro were significantly enhanced.
Adenine ; administration & dosage ; analogs & derivatives ; pharmacokinetics ; Amines ; Antiviral Agents ; administration & dosage ; pharmacokinetics ; Cell Line ; Drug Delivery Systems ; Fluorescein-5-isothiocyanate ; Fluorescent Dyes ; Glycerides ; Hepatitis B virus ; drug effects ; Humans ; Nanoparticles ; Nanotechnology ; Organophosphonates ; administration & dosage ; pharmacokinetics