BACKGROUND:With the deepen understanding on the biological function of Rho/ROCK pathway, new ROCK inhibitors continue to be discovered, and ROCK inhibitors show good promoting effects on the survival, proliferation and migration of keratocytes. Research on ROCK inhibitors wil provide more donor materials or seed cels for regenerative medicine and clinical cel transplantation. OBJECTIVE:To summarize and explore the progress in the treatment and application of corneal disease using the ROCK inhibitors Y-27632 and Y-39983. METHODS:The PubMed database and CNKI database were retrieved by computer to search the relevant literature published between 2008 to 2015 using the key words of “corneal endothelial cel, corneal epithelial cel, ROCK inhibitor, Y-39983, Y-27632” in English and Chinese, respectively. Relevant articles in line with the theme were screened and analyzed. RESULTS AND CONCLUSION:Totaly 264 papers were initialy searched. At last, 45 papers were selected. Currently there are two main ROCK inhibitors: Y-27632 and Y-39983, but both of which are stil in basic research stage and clinical testing stage. Y-27632 promotes the proliferation and activity of corneal epithelial stem cel after resuscitation; Y-39983 as a novel ROCK inhibitor can be better to inhibit Rho kinases activity than Y-27632, thereby more effectively promoting the healing of the corneal endothelium. There are many studies on the application of ROCK inhibitors in corneal treatment, but not a stable method established to obtain seed cels. Each method has its own advantages and disadvantages, and how to overcome these disadvantages and to find fast and stable access to seed cels is the future direction of development.

Traumatic brain injury (TBI)has been classified as mild,moderate,or severe,on the basis of the Glasgow coma scale (GCS)score.Mild TBI is estimated to account for 90% of all cases of TBI, and it has become a serious public health problem,with morbidity increasing year by year.At present,there is a lack of accepted uniform definition of mild TBI.Clinically,mild TBI and concussion are interchangeable terms.In recent years,advances in brain imaging,biomarkers determination,and neuropathology have encouraged people to revise and update their knowledge about mild TBI.In view of the high prevalence of mild TBI in the emergency and community,and the absence of the data concerning the long-term effects of mild TBI, further research is needed about how to reduce morbidity and costs, alleviate delayed consequences,and develop evidence-based interventions to improve outcomes.

The occurrence and development of type 2 diabetes mellitus(T2DM) are related to abnormal energy metabolism.L-carnitine,as one of the most important drugs to improve energy metabolism,has gradually been in a hot research in recent years.It can prevent cell lipid peroxidation,prevent oxidation by free radical scavenging,promote long-chain fatty acids into mitochondria for β-oxidation and increase lipid metabolism and improving diabetic complications.It can also decrease insulin resistance (IR),and lower fasting blood glucose (FBG).L-carnitine may have certain inhibitory effect on the development of T2DM.Early application of L-carnitine may improve the complication of T2DM diabetes,and provide a new way for the treatment of T2DM.

Cardiac valve replacement is the most common surgical procedure in cardiac surgery,which often results in non-surgical bleeding.There are many factors that cause non-surgical bleeding after cardiac surgery,which can be summed up as the disease itself,the artificial device of cardiopulmonary bypass and the special requirements during the operation,including the abnormality of liver function,extracorporeal circulation conduits,oxygenators and blood pumps,hypothermia,hemodilution,neutralization between protamine and heparin.At present,most hospitals still use the results of blood loss and coagulation test as the reference for blood transfusion.Only a few hospitals have used thromboelastogram to guide blood transfusion,which is still in the exploratory stage,so far there is no unified reference standard.This review will focus on the related factors leading to non-surgical bleeding and the concept of accurate blood transfusion and reducing blood transfusion after heart valve replacement in order to prevent the occurrence of rethoracotomy to hemostasis.

Objective To discuss the alterations of functional brain properties in patients with idiopathic trigeminal neuralgia (ITN) based on graph theoretical approaches.Methods A total of 20 patients with pain in the distribution of left trigeminal nerve and 22 matched healthy controls were recruited in our hospital from May 2013 to January 2015.Resting-state functional MRI data in both groups were collected and analyzed with GRETNA software to obtain the matrices of brain network.From these connectivity matrices,a number of key characteristics were computed,including network efficiency,weighted shortest path length,weighted clustering coefficient and small-wordless.A two-tailed t test was used to appraise the significance of any inter-group differences in complex network measures investigated.Network-based statistic (NBS) approach was used to localize specific pairs of brain regions in which functional connectivity strength were altered in patients.Pearson's correlation analyses were performed to identify the correlations between clinical data and altered network properties in patients.Results ITN patients had significantly increased weighted clustering coefficient in the right calcarine (CAL) and bilateral thalamus,increased total efficiency in the left superior frontal gyrus (SFG),and increased regional efficiency in the right SFG,right CAL,bilateral cuneus and left pallidum,and significantly reduced weighted shortest path length for regional nodal characteristics in the right superior parietalis lobules (P＜0.05).The total efficiency in the left SFG and regional efficiency in the right SFG in ITN patients were negatively correlated with course of disease (r=-0.500,P=0.035;r=-0.501,P=0.034).As compared with the healthy controls,the ITN patients had enhanced functional loop connection between left SFG and right supplementary motor area,between left SFG and right SFG,between left SFG and right middle frontal gyrus,between left SFG and right orbital middle frontal gyrus,between left SFG and partes opercularis of right gyrus frontalis inferior (IFGoperc),between left SFG and bilateral putamen (PUT),between left SFG and right PAL,between right PUT and left interior superior frontal gyrus,and between left PUT and right caudate nucleus.The connectivity strength between left SFG and right IFGoperc was negatively correlated with disease duration (r=-0.487,P=-0.041).Conclusion Alterations in regional nodal characteristics may suggest central sensitization and modulation in ITN patients.

Transient expression is the major method to express foot-and-mouth disease virus (FMDV) capsid proteins in mammalian cells. To achieve stable expression of FMDV capsid proteins and efficient assembly of virus like particles (VLPs) in cells, the plasmids of piggyBac (PB) transposon-constitutive expression and PB transposon-tetracycline (Tet) inducible expression vectors were constructed. The function of the plasmids was tested by fluorescent proteins. By adding antibiotics, the constitutive cell pools (C-WT, C-L127P) expressing P12A3C (WT/L127P) genes and the inducible cell pools (I-WT, I-L127P) expressing P12A3C (WT/L127P) genes were generated. The genes of green fluorescent protein, 3C protease and reverse tetracycline transactivator (rtTA) were integrated into chromosome, which was confirmed by fluorescence observation and PCR testing. The cell pool I-L127P has a stronger production capacity of capsid proteins and VLPs, which was confirmed by Western blotting and enzyme linked immunosorbent assay (ELISA), respectively. In conclusion, inducing the chromosomal expression of FMDV capsid proteins was firstly reported, which may facilitate the technical process of mammalian production of FMDV VLPs vaccine and the construction of mammalian inducible expression systems for other proteins.
Animals ; Foot-and-Mouth Disease Virus/genetics* ; Capsid Proteins ; Viral Proteins/metabolism* ; Foot-and-Mouth Disease/prevention & control* ; Tetracyclines/metabolism* ; Viral Vaccines ; Antibodies, Viral ; Mammals/metabolism*

To improve the specific recognition and presentation of virus-like particle (VLPs), and to develop immune-targeted VLPs vaccine, the gene fragment encoding OVA₂₅₇₋₂₆₄ peptide was inserted into the VP3 gene of foot-and-mouth disease virus (FMDV) between the 171th and 172th amino acids (aa) or 173th and 174th aa by reverse PCR. The recombinant proteins were expressed by using Escherichia coli and assembled into chimeric VLP (VLP(OVA)) in vitro after purification. The VLP(OVA) was measured by dynamic light scattering and transmission electron microscopy. The recombinant protein and the assembled VLPs were evaluated by Western blotting, enzyme-linked immunosorbent assay and laser scanning confocal microscopy to confirm the insertion of OVA₂₅₇₋₂₆₄ peptide into VP3 and its location. The results show that insertion of OVA₂₅₇₋₂₆₄ into the 173th and 174th aa of FMDV VP3 did not affect the assembly of VLPs. The VLP(OVA) in size was larger than VLPs, and the OVA₂₅₇₋₂₆₄ peptide was located on the surface of VLP(OVA).
Animals ; Escherichia coli ; genetics ; Foot-and-Mouth Disease ; virology ; Foot-and-Mouth Disease Virus ; genetics ; Recombinant Proteins ; genetics ; metabolism ; Vaccines, Virus-Like Particle

The special nucleic acid fragments, 5' untranslated region (5' UTR) and internal ribosome entry site (IRES) of foot-and-mouth disease virus (FMDV), which interact with the capsid proteins, were selected as scaffolds to investigate the assembly efficiency of foot-and-mouth disease (FMD) virus-like particles (VLPs). The assembled product was characterized by evaluation of particle size, surface potential, gel retardation assay, nuclease digestion experiments, size-exclusion chromatography, transmission electron microscopy and circular dichroism analysis. The results confirmed that the 5' UTR and IRES of FMDV co-assembled with the FMD VLPs and facilitated the assembly efficiency of FMD-VLPs. It demonstrates that the assembly efficiency of 75S particles of VLPs-5'UTR was significantly higher than those of the VLPs (P<0.001) and VLPs-IRES group (P<0.01). Comparatively the assembly efficiency of 12S particles of VLPs-IRES was significantly higher than those of the VLPs (P<0.000 1) and VLPs-5'UTR (P<0.000 1). It showed that the 5' UTR represented more effective in facilitating the assembly of VLPs. This study proposes an optimized strategy for improving the assembly efficiency of VLPs for the development of VLPs vaccine.
5' Untranslated Regions ; Capsid Proteins/metabolism* ; Foot-and-Mouth Disease Virus/physiology* ; Internal Ribosome Entry Sites ; Nucleic Acids/metabolism* ; Virus Assembly

The stability of virus-like particles (VLPs) is currently the main factor affecting the quality of foot-and-mouth disease VLPs vaccines. In order to further improve the quality of the VLPs vaccine of foot-and-mouth disease (FMD), three amino acid modification sites were designed and screened through kinetic analysis software, based on the three-dimensional structure of FMDV. The three mutant recombinant plasmids were successfully prepared by the point mutation kit, transformed into Escherichia coli strain BL21 and expressed in vitro. After purification by Ni ion chromatography column, SDS-PAGE proved that the three amino acid mutations did not affect the expression of the target protein. The results of the stability study of three FMD mutant VLPs obtained by in vitro assembly show that the introduction of internal hydrophobic side chain amino acids made the morphology of VLPs more uniform (N4017W), and their stability was significantly improved compared to the other two VLPs. The internal hydrophobic force of the capsid contributes to the formation of VLPs and helps to maintain the stability of the capsid, providing new experimental ideas for improving the quality of VLPs vaccines, and helping to promote the development of VLPs vaccines.
Amino Acids ; Animals ; Capsid Proteins/genetics* ; Foot-and-Mouth Disease/prevention & control* ; Foot-and-Mouth Disease Virus/genetics* ; Kinetics ; Vaccines, Virus-Like Particle/genetics* ; Viral Vaccines/genetics*

With the continuous progress of research methodology in the real world and the growing maturity of artificial intelligence technology， a method for conducting “quantitative” research to guide clinical practice based on traditional Chinese medicine （TCM） diagnosis and treatment data was gradually developed. However， there is still a need for further improvements in the overall design of studies and the transformation of findings into clinical practice. Based on this， we put forward a comprehensive overall design concept and application approach for real-world study and artificial intelligence research based on clinical diagnosis and treatment data of TCM. This approach consists of five steps： Constructing a research-based database with a large sample size and high data quality； Mining and classification of core prescriptions； Conducting cohort studies to evaluate the effectiveness of core prescriptions； Utilizing case-control studies to clarify the dominant population； Establishing predictive models to achieve precision medicine. Additionally， it is imperative for researchers to establish a standardized system for collecting TCM variables and processing data， optimize the determination and measurement methods of confounding factors， further improve and promote methodologies， and strengthen the training of interdisciplinary talents. By following this research method， we anticipate that the clinical translation of research findings will be facilitated， leading to advancements in TCM precision medicine. Real-world study and artificial intelligence research share similar research foundations， and clinical applications complement each other. In the future， the two will merge together.