Comparation between Guidance for Judicial Expertise of Medical Malpractice and Medical Association Identification Rules of Medical Damage.

On the basis of retaining the technical identification system of medical negligence, the Medical Association Identification Rules of Medical Damage mainly provides technical services for various types of conciliation work about doctor-patient dispute. Its identification work is still influenced by the thinking of medical negligence technical identification and has certain administrative color. Guidance for Judicial Expertise of Medical Malpractice is mainly reflected that the trial of civil cases and pre-trial mediation of courts need service. Its procedures and evidence review are strictly required by the litigation rules and has the characteristics of public legal services provided as a third-party neutral institution. Technical identification of medical damage, whether organized by the Medical Association or the forensic identification institutions, is carried out under the background of the current Regulations on the Prevention and Treatment of Medical Disputes and the Civil Code of the People's Republic of China; both have a corresponding positive role in regulating the medical damage identification activities, and have also laid a certain foundation for the establishment of a unified identification system in the future in China. To understand the different characteristics of the medical damage identification rules issued by the Chinese Medical Association and the Ministry of Justice, and to improve the understanding of the standardization of the forensic identification of medical damage, a comparative study was conducted on Medical Association Identification Rules of Medical Damage and Guidance for Judicial Expertise of Medical Malpractice from seven aspects: Concept and legal status, entrust of identification, identification acceptance, identification procedures, identification presentation meeting, theory of medical malpractice evaluation, consequences and causality of medical damage. The subject of evaluation, the function of evidence review, the role of consulting experts, the technical standard system of malpractice evaluation and other contents were emphatically analyzed.

The Association between the C5263T Mutation in the Mitochondrial ND2 Gene and Coronary Heart Disease among Young Chinese Han People.

OBJECTIVE: This study aimed to investigate the genetic background of mitochondrial genes in young patients with Coronary heart disease (CHD) to provide a foundation for the early prevention of young patients with CHD. METHODS: 115 cases of young (⋜ 45 years) CHD Chinese Han patients (case group), 100 cases of older (> 45 years) Chinese Han CHD patients (experimental group) hospitalized and 100 cases of healthy people through physical examination (control group) at the General Hospital of PLA between January 2014 and December 2015 were selected. General information, clinical assessment, pedigree analysis, and mitochondrial full sequence scanning were performed. The pedigrees of one patient harbouring the C5263T mutation were recruited. Mitochondrial functional analysis including cellular reactive oxygen species (ROS) levels and mitochondrial membrane potential (MMP) were performed on pedigrees with the C5263T mutation (mutation group) and without the mutation (non-mutation group). RESULTS: The differences in biochemical tests (P > 0.05) between the case group and experimental group were not significant. The C5263T single-nucleotide mutation of the mitochondrial ND2 gene was observed in 2 young CHD patients in the case group. The premature CHD of these 2 patients followed a pattern of maternal inheritance. The mutation group (I1, II2) had higher ROS levels (4750.82 ± 1045.55 vs. 3888.58 ± 487.60, P = 0.022) and lower MMP levels (P = 0.045) than the non-mutation group (II1, III1, III2). CONCLUSION: We speculated that the mitochondrial C5263T mutation might be associated with the occurrence CHD in Chinese Han young people.

Surface-Dependent Hydrogen Evolution Activity of Copper Foil.

Single-crystal planes are ideal platforms for catalytic research. In this work, rolled copper foils with predominantly (220) planes were used as the starting material. By using temperature gradient annealing, which caused grain recrystallization in the foils, they were transformed to those with (200) planes. In acidic solution, the overpotential of such a foil (10 mA cm-2) was found to be 136 mV lower than that of a similar rolled copper foil. The calculation results show that hollow sites formed on the (200) plane have the highest hydrogen adsorption energy and are active centers for hydrogen evolution. Thus, this work clarifies the catalytic activity of specific sites on the copper surface and demonstrates the critical role of surface engineering in designing catalytic properties.

Ag-doped Cu nanosheet arrays for efficient hydrogen evolution reaction.

A zinc-infiltration process was adopted to prepare silver-doped copper nanosheet arrays. The larger atomic radius of Ag introduces tensile stress, which lowers the electron density at the s-orbitals of Cu atoms and improves the adsorption capability for hydrogen atoms. As a catalyst for hydrogen evolution, these silver doped copper nanosheet arrays achieved a low overpotential of 103 mV at 10 mA cm-2 in 1 M KOH, which is 604 mV lower than that of pure copper foil.

An electrochemical sensor for rapid determination of ractopamine based on a molecularly imprinted electrosynthesized o-aminothiophenol film.

A simple electrochemical sensor based on a molecularly imprinted polymer film as the recognition element was developed for ractopamine (RAC) detection. This is the first report of a RAC-imprinted film on a gold electrode surface, synthesized through an electrochemical method using o-aminothiophenol as the functional monomer. The imprinting mechanism and experimental parameters affecting the capability of the imprinted film are discussed here. The sensor was successfully applied with constant potential amperometry for RAC detection in an indirect process with potassium ferricyanide as an electrochemical probe. The sensor had a rapid equilibrium time (120 s), high binding affinity and selectivity towards RAC, and with good reproducibility and stability. Under the experimental conditions applied, a linear relationship between the relative amperometric response and RAC ranged from 2.0 × 10(-7) to 1.4 × 10(-6) mol L(-1), with a lower limit of detection (LOD) of 2.38 × 10(-8) mol L(-1) (signal to noise ratio = 3). The sensor was tested with feed samples spiked with trace amounts of RAC, with good recoveries between 87.4 and 90.5 %.

Endophytic Fungus Phomopsis liquidambaris Enhances Fe Absorption in Peanuts by Reducing Hydrogen Peroxide.

Iron (Fe) deficiency in alkaline calcium soil is a problem that needs to be solved urgently as Fe is an essential and commonly limiting nutrient for plants. Endophytic fungus, Phomopsis liquidambaris (P. liquidambaris), has been reported to promote Fe absorption in peanuts (Arachis hypogaea L.), however, the mechanisms remain unclear. Under prolonged Fe deficiency, an increase in hydrogen peroxide (H2O2) often triggers a series of signaling events and leads to the inhibition of Fe acquisition. The main purpose of this study was to explore whether and how the endophytic fungus P. liquidambaris promote Fe absorption in peanut through regulating H2O2 and assisting in resisting oxidative stress. In this study, we detected the Fe deficiency-induced transcription factor (FIT), Fe2+ transporter (IRT1), and ferric reduction oxidase 2 (FRO2) of peanuts, and confirmed that they were negatively related to Fe concentration. Similarly, FIT, IRT1, and FRO2 were also inhibited by H2O2. The addition of P. liquidambaris reduces H2O2 under Fe-deficiency with an increase in Fe content, while the exogenous addition of H2O2 further decreases it, and the addition of catalase (CAT) under Fe-deficiency reverses this phenomenon. Through transcriptome analysis, we proved that the expression of FIT, IRT1, FRO2 and CAT are consistent with our hypothesis, and P. liquidambaris has a stress-mitigating effect on peanuts mainly via CAT, glutathione peroxidase, and malondialdehyde. Our study proved the Fe-absorption promoting effect and stress mitigation effect of P. liquidambaris under Fe-deficiency in peanuts, and their combined usage may help peanuts grow better.

A system for site-specific integration of transgenes in mammalian cells.

Mammalian cell expression systems are the most commonly used platforms for producing biotherapeutic proteins. However, development of recombinant mammalian cell lines is often hindered by the unstable and variable transgene expression associated with random integration. We have developed an efficient strategy for site-specific integration of genes of interest (GOIs). This method enables rapid and precise insertion of a gene expression cassette at defined loci in mammalian cells, resulting in homogeneous transgene expression. We identified the Hipp11 (H11) gene as a "safe harbor" locus for gene knock-in in CHO-S cells. Using clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 mediated homologous recombination, we knocked in a DNA cassette (the landing pad) that includes a pair of PhiC31 bacteriophage attP sites and genes facilitating integrase-based GOI integration. A master cell line, with the landing pad inserted correctly in the H11 locus, was established. This master cell line was used for site-specific, irreversible recombination, catalyzed by PhiC31 integrase. Using this system, an integration efficiency of 97.7% was achieved with green fluorescent protein (GFP) after selection. The system was then further validated in HEK293T cells, using an analogous protocol to insert the GFP gene at the ROSA26 locus, resulting in 90.7% GFP-positive cells after selection. In comparison, random insertion yielded 0.68% and 1.32% GFP-positive cells in the CHO-S and HEK293T cells, respectively. Taken together, these findings demonstrated an accurate and effective protocol for generating recombinant cell lines to provide consistent protein production. Its likely broad applicability was illustrated here in two cell lines, CHO-S and HEK293T, using two different genomic loci as integration sites. Thus, the system is potentially valuable for biomanufacturing therapeutic proteins.

Hemophilia A ameliorated in mice by CRISPR-based in vivo genome editing of human Factor VIII.

Hemophilia A is a monogenic disease with a blood clotting factor VIII (FVIII) deficiency caused by mutation in the factor VIII (F8) gene. Current and emerging treatments such as FVIII protein injection and gene therapies via AAV-delivered F8 transgene in an episome are costly and nonpermanent. Here, we describe a CRISPR/Cas9-based in vivo genome editing method, combined with non-homologous end joining, enabling permanent chromosomal integration of a modified human B domain deleted-F8 (BDD-F8) at the albumin (Alb) locus in liver cells. To test the approach in mice, C57BL/6 mice received tail vein injections of two vectors, AAV8-SaCas9-gRNA, targeting Alb intron 13, and AAV8-BDD-F8. This resulted in BDD-F8 insertion at the Alb locus and FVIII protein expression in the liver of vector-, but not vehicle-, treated mice. Using this approach in hemophilic mice, BDD-F8 was expressed in liver cells as functional human FVIII, leading to increased plasma levels of FVIII and restoration of blood clotting properties in a dose-dependent manor for at least 7 months, with no detectable liver toxicity or meaningful off-target effects. Based on these findings, our BDD-F8 genome editing approach may offer an efficacious, long-term and safe treatment for patients with hemophilia A.

Hemodynamics associated with atrial fibrillation directly alters thrombotic potential of endothelial cells.

An experimental in vitro model of the hemodynamics that occur in atrial fibrillation (AFib) in the left atrial appendage (LAA) was developed to study changes in human endothelial cell thrombotic potential. We applied human-derived sinus rhythm and AFib hemodynamic shear stress patterns to primary human endothelial cells (ECs) in culture. We found that ECs exposed to AFib hemodynamics have increased thrombotic potential as measured by increased expression of pro-thrombotic gene markers and fibrin deposition on the endothelium. Treatment with the factor Xa inhibitor, apixaban, attenuated fibrin deposition thickness while increasing fibrin density at the endothelial cell surface. This study suggests that altered hemodynamics associated with AFib play a key role in driving the thrombotic potential of the LAA endothelium.

Molecularly imprinted quartz crystal microbalance sensor based on poly(o-aminothiophenol) membrane and Au nanoparticles for ractopamine determination.

A molecularly imprinted quartz crystal microbalance (QCM) sensor for ractopamine (RAC) detection was developed by electrodepositing a poly-o-aminothiophenol membrane on an Au electrode surface modified by self-assembled Au nanoparticles (AuNPs). The modified electrodes were characterized by cyclic voltammetry, electrochemical impedance spectroscopy and scanning electron microscopy. This molecularly imprinted QCM sensor showed good frequency response in RAC binding measurements and the introduction of AuNPs demonstrated performance improvements. Frequency shifts were found to be proportional to concentration of RAC in the range of 2.5×10(-6) to 1.5×10(-4) mol L(-1) with a detection limit of 1.17×10(-6) mol L(-1) (S/N=3). The sensor showed a good selective affinity for RAC (selectivity coefficient >3) compared with similar molecules and good reproducibility and long-term stability. This research has combined the advantages of high specific surface area of AuNPs, high selectivity from molecularly imprinted electrodeposited membrane and high sensitivity from quartz crystal microgravimetry. In addition, the modified electrode sensor was successfully applied to determine RAC residues in spiked swine feed samples with satisfactory recoveries ranging from 87.7 to 95.2%.

FGF21 suppresses hepatic glucose production through the activation of atypical protein kinase Cι/λ.

Fibroblast growth factor 21 (FGF21) has been identified as a potent and robust metabolic regulator. Administration of recombinant FGF21 protein to rodents and rhesus monkeys exerts strong anti-diabetic effects. Previous studies have demonstrated that FGF21 inhibits glucose output in the rat H4IIE hepatoma cell line. We performed pharmacological studies to investigate the mechanisms by which FGF21 regulates glucose production in these cells. We found that both insulin and FGF21 suppressed gene expression of G6Pase and PEPCK. Accordingly, glucose production was inhibited. The FGF21 effects were phosphoinositide 3-kinase (PI3K)-dependent, and, unlike insulin, Akt-independent. Additionally, we found that FGF21 induced PKCι/λ phosphorylation in a PI3K-dependent manner; and that a non-isoform selective PKC inhibitor blocked FGF21 inhibition of glucose production, while an inhibitor of classical and novel PKC isoforms had no effect on FGF21 inhibitory activity. Furthermore, hepatic PKCι/λ phosphorylation was upregulated in FGF21-treated diabetic db/db mice.These data support the proposition that FGF21 inhibits hepatic glucose production by the PI3K-dependent activation of PKCι/λ.

The Rb-related p130 protein controls telomere lengthening through an interaction with a Rad50-interacting protein, RINT-1.

The oncogenic process often leads to a loss of normal telomere length control, usually as a result of activation of telomerase. Nevertheless, there are also telomerase-independent events that involve a Rad50-dependent recombination mechanism to maintain telomere length. Previous work has implicated the Rb family of proteins in the control of telomere length, and we now demonstrate that the p130 member of the Rb family is critical for telomere length control. p130 interacts specifically with the RINT-1 protein, previously identified as a Rad50-interacting protein. We further show that RINT-1 is essential for telomere length control. We propose that p130, forming a complex with Rad50 through RINT-1, blocks telomerase-independent telomere lengthening in normal cells. Given previous work implicating E2F in the control of telomerase gene expression, these results thus point to complementary roles for the Rb/E2F pathway in the control of telomere length.

A geminivirus replication protein interacts with a protein kinase and a motor protein that display different expression patterns during plant development and infection.

The geminivirus protein AL1 initiates viral DNA replication, regulates its own expression, and induces plant gene transcription. To better understand how AL1 interacts with host proteins during these processes, we used yeast two-hybrid library screening and a baculovirus protein interaction system to identify plant proteins that interact with AL1. These studies identified a Ser/Thr kinase, a kinesin, and histone H3 as AL1 partners. The kinase is autophosphorylated and can phosphorylate common kinase substrates in vitro. The kinesin is phosphorylated in insect cells by a cyclin-dependent kinase. Immunostaining of Nicotiana benthamiana and Arabidopsis showed that kinase protein levels and subcellular location are regulated during plant development and geminivirus infection. By contrast, the kinesin is ubiquitous even though it is associated with the spindle apparatus in mitotic cells. Together, our results establish that AL1 interacts with host proteins involved in plant cell division and development. Possible functions of these host factors in healthy and geminivirus-infected plants are discussed.

A novel motif in geminivirus replication proteins interacts with the plant retinoblastoma-related protein.

The geminivirus replication factor AL1 interacts with the plant retinoblastoma-related protein (pRBR) to modulate host gene expression. The AL1 protein of tomato golden mosaic virus (TGMV) binds to pRBR through an 80-amino-acid region that contains two highly predicted alpha-helices designated 3 and 4. Earlier studies suggested that the helix 4 motif, whose amino acid sequence is strongly conserved across geminivirus replication proteins, plays a role in pRBR binding. We generated a series of alanine substitutions across helix 4 of TGMV AL1 and examined their impact on pRBR binding using yeast two-hybrid assays. These experiments showed that several helix 4 residues are essential for efficient pRBR binding, with a critical residue being a leucine at position 148 in the middle of the motif. Various amino acid substitutions at leucine-148 indicated that both structural and side chain components contribute to pRBR binding. The replication proteins of the geminiviruses tomato yellow leaf curl virus and cabbage leaf curl virus (CaLCuV) also bound to pRBR in yeast dihybrid assays. Mutation of the leucine residue in helix 4 of CaLCuV AL1 reduced binding. Together, these results suggest that helix 4 and the conserved leucine residue are part of a pRBR-binding interface in begomovirus replication proteins.