Quality markers research on Chinese herbal pieces: Quality evaluation and quality standards of steamed and vinegar-processed pieces of Curcumae Rhizoma / 中草药

A demonstration research on Chinese herbal decoction pieces of Curcumae Rhizoma was performed based on the concept of quality markers (Q-markers), standard establishment, and research modes. The chemical constituents of both steamed processed pieces of Curcumae Rhizoma and vinegar-processed pieces of Curcumae Rhizoma decoction pieces were identified by ultra- performance liquid chromatography/quadrupole-time of flight mass spectrometry (UPLC-Q/TOF-MS). The major effective components were analyzed through pharmacodynamics, drug property, pharmacokinetics studies, and correlation analysis of chemical constituents. The Q-markers were determined by all the results. At last, five compounds including curdione, curcumol, germacrone, furanodiene and β-elemene were selected as Q-markers. The quality control methods of multi-component assaying and fingerprint were also established. Overall, this study provides a demonstration for the study of quality markers of Chinese herbal decoction pieces.

Research and development thought on biopharmaceutics classification system of Chinese materia medica / 中国中药杂志

The biopharmaceutics classification system( BCS) is a scientific framework or method for classifying drugs based on drug solubility and permeability,which can be used to provide drug bioavailability-absorption correlation analysis. Based on the characteristics of multi-component and multi-target of traditional Chinese medicine( TCM) as well as the concept,method and technology of BCS,the research group proposed biopharmaceutics classification system of Chinese materia medica( CMMBCS) and carried out research and data accumulation of classical prescriptions. Based on the previous research results,further development ideas under the CMMBCS concept and framework were further proposed in this study. In the course of research,the influence of the intermediate links of the complex interactions of the multi-component environment was omitted,and the component absorption studies on the main clinical effects of prescription ingredients were directly concerned,or the components and data were reversely extracted from the aspects of metabolism,pharmacodynamic pathways and absorption principles. Studies were conducted from two aspects( single component and compound prescription) to comprehensively evaluate the absorption properties of TCM compound. In the research path,the different ways in which Chinese medicine could exert its efficacy were fully considered,and CMMBCS classification and establishment rules were clarified mainly by focusing on the absorption pathway into the blood. Specifically,the network pharmacology and molecular docking technology were used to screen the compound index components of TCM; the absorption rules were studied by the physiologically based pharmacokinetic models and the absorption parameters of CMMBCS were calculated by reverse reasoning. Then the CMMBCS classification of TCM prescription was corrected by studying the efficacy or absorption pathway. In this paper,the theoretical framework and research methodology of CMMBCS were systematically improved based on the establishment of CMMBCS basic theory,the supplementary of drug-oriented research ideas and the application of modern mature Chinese medicine methodology.

Knowledge of the consequences and use of drugs for CostaRica niversity students / Conhecimento das consequências e do uso de drogas em estudantes universitários costarriquenses / Conocimiento de las consecuencias y uso de drogas en estudiantes universitarios costarricenses

ABSTRACT Objective: to determine the relationship between knowledge of consequences and drug use in undergraduate students of a university in San José, Costa Rica. Method: the cross-sectional study examines the demographic profile of the sample and the relationship between knowledge of consequences, drug use and academic performance. The study focuses on three types of drugs: alcohol, marijuana and cocaine. Three variables will be analyzed: demographic data, knowledge of consequences and use of drugs. Results: the relationship between knowledge of consequences and use of drugs was made using of the T-test. The sample had 272 students, 28.2% (n=77) of them were men and 71.4% were women (n=195). They were selected from the areas of social sciences (n=137, 50.2%), and from the area of health sciences (n=136; 49.8%). Alcohol was the most used drug (n=217, 79.8%), followed by marijuana (n=72, 26.6%) and finally cocaine (n=3, 1.1%) in the last 12 months. Conclusion: the results shown indicate that there is no significant relationship between such variables. The findings are important at the level of drug policies to support the development of new preventive strategies for drug use.

RESUMO Objetivo: determinar a relação entre o conhecimento das consequências e o uso de drogas em estudantes de graduação de uma universidade em San José, Costa Rica. Método: este estudo, de corte transversal, examina o perfil demográfico da amostra e a relação entre o conhecimento das consequências, o uso de drogas e o desempenho acadêmico. A pesquisa centra-se em três tipos de drogas: álcool, maconha e cocaína. Três variáveis foram analisadas: os dados demográficos, o conhecimento das consequências e o uso de drogas. Resultados: a relação entre o conhecimento das consequências e o uso de drogas foi realizada através do uso do T-test. A amostra foi composta por 272 estudantes, sendo 28,2% (n=77) homens e 71,4% mulheres (n=195); selecionados da área de ciências sociais (n=137; 50,2%), e da área de ciências da saúde (n=136; 49,8%). O álcool foi a droga mais utilizada (n=217; 79,8%), seguida da maconha (n=72; 26,6%) e, finalmente, a cocaína (n=3; 1.1%) nos últimos 12 meses. Conclusão: os resultados demostram que não existe relação significativa entre tais variáveis, e são importantes no nível das políticas de drogas para apoiar o desenvolvimento de novas estratégias preventivas para o uso de drogas.

RESUMEN Objetivo: determinar la relación entre conocimiento de consecuencias y uso de drogas en estudiantes de pregrado de una universidad en San José, Costa Rica. Método: el estudio, de corte transversal, examina el perfil demográfico de la muestra y la relación entre conocimiento de consecuencias, uso de drogas y rendimiento académico. El estudio se enfoca en tres tipos de droga: alcohol, marihuana y cocaína. Se analizarán tres variables: datos demográficos, conocimiento de consecuencias y uso de drogas. Resultados: la relación entre conocimiento de consecuencias y uso de drogas se realizó mediante el uso de la prueba T-test. La muestra fue de 272 estudiantes, con un 28.2% (n=77) de hombres y 71.4% de mujeres (n=195); seleccionados de las áreas de ciencias sociales (n=137; 50.2%), y del área de ciencias de la salud (n=136; 49.8%). El alcohol fue la droga más utilizada (n=217; 79.8%), seguida por marihuana (n=72; 26.6%) y finalmente la cocaína (n=3; 1.1%) en los últimos 12 meses. Conclusion: los resultados mostrados indican que no hay una relación significativa entre tales variables. Los hallazgos son importantes a nivel de políticas de drogas para apoyar el desarrollar de nuevas estrategias preventivas de uso de drogas.

Estudio Transversal: Consumo de Sustancias Psicotrópicas en Pacientes que Acuden al Servicio de Psicología del Centro de Salud No 1 de Azogues - Ecuador, 2017 / Cross ­ Sectional Research: Consumption of psychotropic substances in Psychology service patients´ of the Health Center No. 1 of Azogues, 2017

INTRODUCCIÓN: El consumo de sustancias psicotrópicas de manera continua actúan sobre el sistema nervioso central, estimulando o deprimiéndolo. Los objetivos del estudio fueron determinar la prevalencia del consumo de sustancias psicotrópicas en pacientes que acuden al servicio de Psicología, establecer cuáles son los psicotrópicos que se consumen, su frecuencia, su asociación con edad y sexo e identificar riesgo. MÉTODOS: Se trata de un estudio transversal, con un universo de 1182 pacientes, obtenidos del registro diario automatizado de consultas y atenciones médicas ambulatorias del Ministerio de Salud Pública, que midió la prevalencia del consumo de psicotrópicos en el área de Psicología del centro de salud No 1 de Azogues ­ Ecuador, durante el año 2017. Se utilizó la clasificación internacional de enfermedades 10 para los respectivos diagnósticos y se tabuló en los programas estadísticos SPSS versión 22 y EPIDAT 3.1. RESULTADOS: El 52.2 % eran hombres, la media de edad fue de 26 años. Se estableció una prevalencia de 9 % para el consumo de sustancias psicotrópicas. Los psicotrópicos más consumidos fueron: alcohol (3.6 %), cannabinoides (2.9 %) y múltiples drogas (1.8 %), teniendo más frecuencia de consumo, el sexo masculino (15.4 % frente a 1.9 % del femenino) y en el grupo etario de 10 - 19 años con un 13.9 %, seguido de los de 20 ­ 29 años con 11.1%. El uso nocivo tuvo una prevalencia de 57.55 %, la dependencia 34.9 % y la abstinencia un 7.55 %. CONCLUSIONES: Existe asociación estadística entre el consumo y las variables sexo y edad (ambas P: 0.000). El sexo masculino y ser adolescente/joven (10 ­ 29 años) constituyen factores de riesgo para el consumo de sustancias psicotrópicas (RP 7.9; LC: 4.2 ­ 14.6 y RP 2.5; LC: 1.6 ­ 3.7 respectivamente).

BACKGROUND: The consumption of psychotropic substances acts continuously on the central nervous system, stimulating or depressing it. The aim of the study they should determine the prevalence of use of psychotropic substances in patients who attended the psychology service, establish which psychotropic drugs are consumed, their frequency, their association with age and sex, and identify the risk. METHODS:It is a cross-sectional study, with a universe of 1182 patients, obtained of the automated daily record of outpatient consultations and medical attentions of the Ministry of Public Health, which measured the prevalence of psychotropic use in the area of Psychology of the health center Nº 1 of Azogues - Ecuador, during the year 2017. It was used the international classification of diseases 10 for the respective diagnoses and it was tabulated in the statistical programs SPSS version 22 and EPIDAT 3.1. RESULTS: The 52.2 % were men, the average age was 26 years. A prevalence of 9 % was established for the consumption of psychotropic substances. The psychotropics most consumed were: alcohol (3.6 %), cannabinoids (2.9 %) and multiple drugs (1.8 %), having more frequency of consumption, the male sex (15.4 % versus 1.9 % of the female) and in the age group of 10 -19 years with 13.9 %, followed by those aged 20 - 29 with 11.1 %. Harmful use had a prevalence of 57.55 %, dependence 34.9 % and abstinence a 7.55 %. CONCLUSIONS: There was a statistical association between consumption and sex and age variables (both P: < 0.001). Male and adolescent / young (10 - 29 years) are risk factors for the consumption of psychotropic substances (RP 7.9, LC: 4.2 - 14.6 and RP 2.5; LC: 1.6 - 3.7 respectively).

Drug-target protein interaction prediction based on AdaBoost algorithm / 生物医学工程学杂志

The drug-target protein interaction prediction can be used for the discovery of new drug effects. Recent studies often focus on the prediction of an independent matrix filling algorithm, which apply a single algorithm to predict the drug-target protein interaction. The single-model matrix-filling algorithms have low accuracy, so it is difficult to obtain satisfactory results in the prediction of drug-target protein interaction. AdaBoost algorithm is a strong multiple classifier combination framework, which is proved by the past researches in classification applications. The drug-target interaction prediction is a matrix filling problem. Therefore, we need to adjust the matrix filling problem to a classification problem before predicting the interaction among drug-target protein. We make full use of the AdaBoost algorithm framework to integrate several weak classifiers to improve performance and make accurate prediction of drug-target protein interaction. Experimental results based on the metric datasets show that our algorithm outperforms the other state-of-the-art approaches and classical methods in accuracy. Our algorithm can overcome the limitations of the single algorithm based on machine learning method, exploit the hidden factors better and improve the accuracy of prediction effectively.

Pharmacodynamic principles and the time course of delayed and cumulative drug effects

This tutorial reviews the principles of the concentration — effect relationship for the usual case when drug effects are delayed relative to changes in circulating concentrations. The key processes determining delay are distribution from the circulation to the receptor, binding to the receptor to produce a stimulus and translation of the receptor stimulus into an effect through turnover of physiological mediators. Some clinical outcomes are dependent on the accumulation of drug action which is predictable in terms of basic pharmacokinetic and pharmacodynamic concepts.

Protection effects of metformin on biological behaviour of human vascular endothelial cells under inflammatory conditions / 中华实验眼科杂志

Background Studies showed that inflammatory process participates in the pathogenesis anddevelopment of diabetic retinopathy targeting retinal vascular endothelial cells (RVECs).A growing body of evidence revealed that metformin reduces the risk of micro-and macro-vascular complications by protecting blood-brain barrier,however,whether it plays a protective effect on human retinal vascular by similar mechanism is still unelucidated.Objective This study was to investigate the effects of metformin on the proliferation,migration and secreting monocyte chemotactic protein-1 (MCP-1) and interleukin-8 (IL-8) of human retinal vascular endothelial cells (RVECs) under the stimulation of tumor necrosis factor-alpha (TNF-α).Methods RVECs were cultured and divided into normal control group,metformin (5 mmol/L) group,TNF-α 2.5 ng/ml group,and TNF-α+metformin (5,10,20 and 40 mmol/L,respectively) groups.Corresponding drugs were added into medium according to grouping for 24 hours.Cell numbers were calculated before and after treatment.The metabolic activity (absorbancy) of RVECs was measured with MTS assay.Cell migration of RVECs was assessed with transwell migration assay.The MCP-1 and IL-8 concentrations in the cell supernatant were detected by ELISA assay.Results The number of the cells was significantly different among the normal control group,metformin group,TNF-α group,and TNF-α+metformin (5,10,20 and 40 mmol/L,respectively) groups (F =189.31,P ＜ 0.01).The metabolic activities of RVECs were 0.32 + 0.02,0.32±0.03,0.97 ± 0.02,0.90 ± 0.05,0.76 ± 0.15,0.74 ± 0.05 and 0.41 ± 0.03;migrated cell numbers were (1 214±49),(1 200±45),(1 648±43),(1 309±48),(1 279±73),(961±60) and (942±106)/field;the concentrations of MCP-1 were (0.385 ±0.050),(0.362±0.060),(2.285 ±0.200),(1.131 ±0.180),(0.622 ± 0.120),(0.537±0.090) and (0.492±0.130) μg/ml,and those of IL-8 were (0.385±0.080),(0.390±0.120),(1.123±0.130),(0.899±0.180),(0.680±0.060),(0.417±0.090) and (0.335±0.100) μg/ml in the normal control group,metformin group,TNF-α group,and TNF-α + metformin (5,10,20 and 40 mmol/L,respectively) groups,showing significant differences among the groups (F =73.31,103.89,150.92,268.32,all at P＜ 0.01).The cell number,cell metabolic activity,migrated cell number,and MCP-1 and IL-8 levels in the cell supernatant were evidently increased in the TNF-α group compared with the normal control group,and those in the TNF-α+10 mmol/L metformin group,TNF-e +20 mmol/L metformin group and TNF-α+40 mmol/L metformin group were significantly decreased in comparison with the TNF-α group (all at P＜0.05).Conclusions Metformin can inhibit TNF-α-induced proliferation,migration and MCP-1 and IL-8 secretion of the cells,and therefore plays a protective role on RVECs in the inflammatory environment.

Pharmacodynamic principles and the time course of immediate drug effects

This tutorial defines the principles of the concentration - effect relationship which are the basis of pharmacodynamics. The two key parameters of pharmacodynamics are the maximum response (Emax) and the concentration producing 50% of Emax (C₅₀). The time course of effect is illustrated under the assumption that drug effects are immediately related to concentration in the central compartment e.g. plasma. The related idea of duration of drug action and its relationship to dose is shown to have a simple relationship with drug half-life.

Response of Leptomeningeal Dissemination of Anaplastic Glioma to Temozolomide: Experience of Two Cases

The incidence of leptomeningeal dissemination (LMD) of anaplastic glioma has been increasing. LMD can be observed at the time of initial presentation or the time of recurrence. As a result of both rarity and unusual presentation, a standard therapy has not yet been suggested. In contrast to leptomeningeal carcinomatosis for systemic solid cancers, a relatively prolonged survival is observed in some patients with LMD of anaplastic gliomas. Treatment modalities include whole craniospinal irradiation, intra-cerebrospinal fluid (CSF) chemotherapy, and systemic chemotherapy. In some cases, response to temozolomide (TMZ), with or without combined radiation has been reported. Here, we report two cases of LMD of an anaplastic glioma. In one case LMD presented at the time of diagnosis, and in the other at the time of recurrence after radiation. CSF cytology was positive in both cases, and persisted in spite of intrathecal methotrexate chemotherapy. Later, TMZ was prescribed for progressing brain parenchymal lesions, and both radiological and cytological responses were obtained after oral TMZ treatment.

Anti-aging effect of simvastatin and telmisartan on retinas and its mechanism in rats / 中华实验眼科杂志

Background Statins has prominent roles in regulating lipids,anti-inflammation,autoxidation and protecting vascular endothelial cells.Sartans can promote cell growth and the expression of cytokines.Since the pleiotropic effects of statins and sartans on a variety of cell types,it is inferred that the two medicines can delay retinal aging.Objective This study was to explore the anti-aging effect of simvastatin and telmisartan on the physiological aging of retina.Methods Sixty-six three-month-old healthy SD rats were selected in this study,and 6 of them served as the youth group and the right eyeballs were immediately enucleated.The other rats were raised until 9-month-old in the same conditions and then randomly divided into the simvastatin group,telmisartan group and the control group with 20 rats for each group.The simvastatin of 5 mg/kg and telmisartan of 8 mg/kg were given by intragastric administration once a day in the simvastatin group and the telmisartan group until 17-month-old,and the equal amount of normal saline was used in the control group in the same way.The number of survival rats was 12 in the simvastatin group,10 in the telmisartan group and 8 in the control group.The right eyes were enucleated after heart perfusion of 4％ paraformaldehyde solution for the preparation of retinal paraffin sections.Retinal thickness was measured by pathological examination,and the expressions of the retinal neuron markers,including Thy-1,protein kinase C-α (PKC-ot),opsin and rhodopsin,were detected by immunofluorescence technique to evaluate the morphology of retinal ganglion cells (RGCs),bipolar cells as well as the thickness of the outer segment of photoreceptors.Results The retinal structure was clear in the rats of the youth group.However,the RGCs arrangement and inner segment (IS) and outer segment (OS) structure were abnormal in the simvastatin group,the telmisartan group and the control group.Compared with the rats of the youth group,the thickness of outer nuclear layer (ONL),outer plexiform layer (OPL),inner nuclear layer (INL),inner plexiform layer (IPL) and the total thickness of the aging rats were decreased,and the IS/OS thickness was increased in the simvastatin group and the telmisartan group (all at P＜ 0.01).Thy-1 stain showed that the number of RGCs was reduced in the simvastatin group,telmisartan group and the control group compared with the youth group,and that in the simvastatin group was increased in comparison with the control group (all at P＜0.01).PKC-αt stain exhibited that the density of bipolar cells was increased but the axon terminal bouton was declined in the simvastatin group,telmisartan group and the control group compared with the youth group,and the axon terminal bouton was declined in the simvastatin group compared with the youth group and the control group (all at P=0.000).Opsin and rhodopsin stains displayed that the OS thickness was increased in the simvastatin group,telmisartan group and the control group compared with the youth group,and that in the telmisartan group was reduced in comparison with the control group (all at P＜0.01).Conclusions As SD rat aging,retinal thickness is gradually attenuated and the number of RGCs is gradually declined.Although the density of bipolar cells seem to be unchanged,their synaptic connections are decreased and the OS is thicken.Simvastatin and telmisartan can delay retinal senescence by protecting retinal neurons against aging and thinning thickened OS.

Quality markers research on Chinese materia medica: Quality evaluation and quality standards of Corydalis Rhizoma / 中草药

Corydalis Rhizoma was taken as an example for demonstration research based on the concept of quality markers (Q-markers), confirmed standards and research modes were proposed by the author. The chemical constituents of Corydalis Rhizoma were identified using high performance liquid chromatography/electrospray ionization quadruple time-of-flight mass spectrometry (HPLC/ESI-Q-TOF MS). The source and specificity of chemical constituents were confirmed by analyzing biosynthetic pathway and component specificity. The major effective components were clarified through efficacy, drug property, pharmacokinetic studies, and correlation analysis of chemical constituents. The Q-markers were determined by integrating all the results of the studies. At last, seven alkaloidal compounds including tetrahydropalmatine, corydaline, coptisine, palmatine, dehydrocorydaline, tetrahydrojatrorrhizine, and protopine have been selected as Q-markers and quality control methods of multi-component assaying and fingerprint have been established.

Relation of "property-response-component" and action mechanism of Yuanhu Zhitong Dropping Pills based on quality marker (Q-Marker) / 中草药

The research of effective components of Chinese materia medica (CMM) is one of the key points of modernization of traditional Chinese medicine (TCM), is the core of interpreting overall efficacy and mechanism of TCM, and is the important foundation of realizing the safety, efficacy, and quality control of CMM. Previous studies mostly focused on the binary research mode of "component-effect". However, in this paper, the triarchic theory of "property-effect-component" was proposed and the research strategy and mode of exploring the mechanism, effective components, and quality markers (Q-Marker) of TCM were established based on the basic properties of TCM, basic concepts and terms of TCM theory and also related with the core contents of TCM theory including syndrome differentiation and treatment, rules of treatment, method of treatment, compatibility regularity, and drug property. The Yuanhu Zhitong Dropping Pills (YZDP) were taken as an example for demonstration research and corydaline, tetrahydropalmatine, protopine, imperatorin, and isoimperatorin were determined to be the Q-Marker of YZDP.

Inhibition of curcumin on the expression of IL-1 β-induced nuclear factor-κB-dependent inflammatory gene in rabbit RPE cells / 中华实验眼科杂志

Background Interleukin-1β (IL-1β) is an important inflammation-related factor in the initial stage of proliferative vitreoretinopathy (PVR).The previous research showed that curcumin can inhibit IL-1 β-induced proliferation of rabbit retinal pigment epithelium (RPE) cells,but the anti-inflammatory mechanism and effect of curcumin are still undefined.Objective This study was to observe the migration of IL-1β-induced rabbit RPE cells,and evaluate the function and mechanism of inhibition of curcumin on IL-1β-induced inflammation of RPE cells.Methods Cultured rabbit RPE cells of generation 4 were used in this experiment.The cells were cultured in serum-free DMEM and 0,0.1,1.0 and 10.0 μg/L IL-1β were separately added in the medium for 24 hours.The expressions of cyclooxygenase-2 (COX-2) protein and mRNA in the cells were detected by Western blot and reverse transcription PCR to determine the optimal concentration of IL-1β.The cells were divided into IL-1β group and curcumin+IL-1β group,and 1.0 μg/L IL-1 or 1.0 μμg/L IL-1 β combined with 10 μg/ml curcumin was respectively added into the medium for 24,48 and 72 hours.The cells cultured by only serum-free medium served as the control group.Hematoxylin and eosin staining was conducted for the cells to count the number of cells migrating into the injured area under the optical microscope.The relative expression levels of COX-2 protein and mRNA in the cells were detected by Western blot and reverse transcription PCR,and the relative expression levels of nuclear factor (NF)-κBp65 and inhibitor of NF-κB-α (IκB-α) protein were also detected by Western blot assay.The expression intensity and location of NF-κBp65,IκB-α and COX-2 in the cells were detected by immunochemistry.Results RPE cells just isolated from the rabbit eyes were in round shape and abundant in melanin.The melanin significantly decreased in the fourth generations of RPE cells.The shape of cells became long and narrow,and net shaped distribution.Immunochemistry demonstrated the strong positive response of RPE cells for keratin (AE1/AE3).There were (31.93 ±1.21),(36.27±2.50) and (38.33±2.40) migratory cells in the control group after 24,48 and 72 hours respectively.The number of migratory cells increased to 45.73 ± 2.30,71.13 ± 1.92 and 80.60 ± 1.71 in the IL-13 group,but obviously decreased to 13.13 ± 2.20,14.93 ± 1.10 and 12.60 ± 1.51 in the curcumin + IL-1β group.A Significant increase in the migrating cell number was found in the IL-1 β group compared with the control group and the curcumin+IL-1β group in various time points (all at P＜0.05).The relative expression levels of COX-2 protein and mRNA peaked in the 1.0 μg/L IL-1β group,so 1.0 μg/L of IL-1β was determined as the optimal concentration in the experiment.In 24,48 and 72 hours after culture,the expression levels of COX-2 protein and mRNA in the cells were significantly lower in the curcumin + IL-1β group than those in the control group (all at P＜0.05).The relative expression level reached peak in NF-κBp65 protein and lowed bottom in IκB-α proteins at 48 hours after cultured in the IL-1β group,and the reverse trend was seen in the curcumin+IL-1β group,with the significant differences between the two groups (both at P＜0.05).Immunochemistry showed that NF-κBp65 was expressed strongly in the cell nuclei and cytoplasm in the IL-1 β group and presented the weaker expression in the control group and the curcumin+IL-1 β group.Compared with the control group,the expression was weaker in IκB-α and stronger in COX-2 in the IL-1β group.In addition,the expression of IκB-α was enhanced and that of COX-2 was attenuated in the curcumin+IL-1β group in comparison with the IL-1β group.Conclusions Curcumin inhibits the movement of rabbit RPE cells induced by IL-1β.IL-1β up-regulates the expression of COX-2 by activating NF-κB signal pathway,and curcumin plays an anti-inflammatory role by blocking this pathway.

Rapid differentiation of human ESCs into RPE cells by defined xeno-free culture system / 中华实验眼科杂志

Background Subretinal transplantation of retinal pigment epithelium (RPE) cells for the treatment of age-related macular degeneration (AMD) have accelerated the drive to develop xeno-free cultivation system that support the rapid differentiation of human embryonic stem cells (hESCs) into ES-RPE cells.Objective This study was to report a modified xeno-free culture system and method for accelerating derivation of hESCs to differentiate into RPE cells.Methods This study was approved by Ethic Committee of Zhongshan Ophthalmic Center.HESC H1 line was cloned and cuhured in Vitronectin XFTM-coated 6-well dish with xenogenetic-free medium.Cells were cultured in 50 ng/ml noggin,10 ng/ml DKK-1 and 10 ng/ml insulin like growth factor-1 (IGF-1) medium for 2 days,and then the concentration of noggin was decreased to 10 ng/ml and 5 ng/ml basic fibroblast growth factor (bFGF) and cultured for the following 2 days.Sequentially,noggin and bFGF were removed and cultured for 2 days.Finally,1 μmol/L CHIR99021 was added in medium for 6 days.Morphological changes in the progress of ESCs differentiation into RPE were observed by Living Cell Imaging System.The expression of Mitf and RPE65,RPE cellsspecific markers,in the cells were detected by immunofluorescence technique,and the relative expression levels of RPE cells-specific marker mRNA were assayed using real time fluorescent quantitation PCR.Results Polygonalshape monolayer cells which contained pigments were initially observed at day 14 after cultured with the cobblestonelike arrangement.Mitf and RPE65 were strongly expressed in the hES-derived RPE cells 35 days after induced,showing red fluorescence,and the cells presented hexagonal shape at cultured day 60 with numerous pigment granules in cytoplasm.Compared with before differentiation,the expression levels of Mitf mRNA in hES-RPE cells increased by (3.43±2.77) folds and (8.91 ± 2.83) folds,and the expression levels of RPE65 mRNA increased by (14.60 ± 3.94) folds and (87.16 ±9.32) folds at day 7 and day 14 after differentiation,respectively (all at P＜0.05).Conclusions A defined xeno-free culture system is successfully established by adding niacinamide,DKK-l,noggin,IGF-1 and CHIR99021 in xeno-free medium,and this system can accelerate the derivation and differentiation of hESCs into RPE-like cells.

Suppressing effects of MMPs inhibitor GM6001, MMP-2/9 inhibitor Ⅰ ,MMP-2/9 inhibitor Ⅱ on migration of human lens epithelial cells / 中华实验眼科杂志

Background The primary pathologic mechanism of posterior capsular opacification (PCO) is proliferation,migration and epithelial-mesenchymal transformation of residuary lens epithelial cells (LECs) following cataract surgery.Matrix metalloproteinases (MMPs) play a role during the migration of LECs.Researches showed that GM6001,a broad inhibitor of MMPs,can arrest the migration of LECs,but as specific inhibitors of MMPs,the efficacy and safety of MMP-2/9 inhibitor Ⅰ and Ⅱ on LECs migration remain unclear.Objective This study was to determine and compare the inhibitory efficacy among GM6001,MMP-2/9 inhibitor Ⅰ and Ⅱ on human LECs and search the clinical medication to prevent PCO.Methods Human LECs were cultured and passaged in vitro,and the cells of 3-4 generation were incubated in 6-well plates.Then the cells of 70％ confluent monolayer were cultured in DMEM without fetal bovine serum for 12 hours.GM6001,MMP-2/9 inhibitor Ⅰ and Ⅱ at different concentrations (0.25,0.50,1.00,2.00,4.00,8.00,16.00,32.00,64.00,128.00 μmol/L) were added into the culture medium for 24 hours separately,and regularly cultured cells served as the control group.A bare area was made by a 200 μl sterile spear on the cell layer,and the migrated distance and inhibitory rate were calculated.The second or third generation of cells were incubated in 96-well plates at a density of 5×105/ml (200 μl/well).GM6001 (128.00 μmol/L),MMP-2/9 inhibitor Ⅰ (64.00 μmol/L) and Ⅱ (32.00 μmol/L) were added into the culture medium for 24 hours,and the cell viability was assayed by using MTT assay.Results Cultured cells grew well with irregular arrangement and presented the polygon in shape.The migrated distance was gradually reduced as the increase of concentrations of GM6001,MMP-2/9 inhibitor Ⅰ and Ⅱ,showing significant differences among the various concentration groups (GM6001:F=248.647,P＜0.05;MMP-2/9 inhibitor Ⅰ:F=357.125,P＜0.05;MP2/9 inhibitor Ⅱ:F=396.374,P＜ 0.05).The cell migrated distance in the control group was set to 1,the relative migrated distances were 0.478 ± 0.091,0.294±0.088 and 0.191 ±0.081 in the GM6001 group,MMP-2/9 inhibitor Ⅰ group and MMP-2/9 inhibitor Ⅱ group at the concentrations of 32.00 μmol/L,respectively,showing a significant difference among the groups (F =116.031,P＜0.01),and cell migrated distance was obviously shorter in the MMP-2/9 inhibitor Ⅱ group than that in the GM6001 group or MMP-2/9 inhibitor Ⅰ group (all at P＜0.01).The A values were 0.607±0.016,0.567±0.015,0.583±0.010 and 0.595 ±0.0138 in the control group,GM6001 group (128.00 μmol/L),MMP-2/9 inhibitor Ⅰ group (64.00 μmol/L) and MMP-2/9 inhibitor Ⅱ group (32.00 μmol/L),respectively,without significant difference among the groups (F=1.403,P＞0.05).Conclusions GM6001,MMP-2/9 inhibitor Ⅰ and Ⅱ reduce the mobility of human LECs effectively but do not affect the viability of the cells in vitro.MMP-2/9 inhibitor Ⅱ appears to be most dominant in inhibiting migration of human LECs.

Promoting effect of curcumin on induced differentiation of human embryonic stem cells into retinal pigment epithelial-like cells / 中华实验眼科杂志

Background Pluripotent stem cell-derived retinal pigment epithelial (RPE) cells holds great promise for the treatment of age-related macular degeneration (AMD) and retinitis pigmentosa (RP),but the poor induction efficiency and the according high cost of RPE differentiation hindere its clinical applications.Curcumin is proved to have a promoting effect on the induced differentiation of embryonic stem cells (ESCs).However,the mechanism of curcumin on differentiation of human ESCs into RPE-like cells remains unclear.Objective This study aimed to explore the underlying molecular mechanism of curcumin on directed differentiation of human ESCs into RPE-like cells.Methods Human ESCs strains were cultured in the Matrigel-coated 6-well plate with mTeSRTM 1 medium until over-confluence,and basic fibroblast growth factor was withdrawn there after to induce automatic differentiation.Curcumin at the final concentration 1 μmol/L was added in the first day of differentiation for 24 hours,and the cells without curcumin in the medium served as the control group.Total RNA and protein were extracted at 3 weeks and 5 weeks after induction.RT-PCR,Western blot and immunofluorescence were performed to examine the expressions of the biomarks of stem cells and RPE cells as well as Wnt/β-catenin signaling pathway components.The endocytosis of polystyrene microsphere by induced RPE (iRPE) cells was investigated to verify their function of phagocytosis which features RPE cells.Results Pigmented cells were found from 3 weeks through 5 weeks after induction in the curcumin group,but only less pigmented cells were seen in the fifth week after induction in the control group.In the third and fifth week after induction,the relative expression levels of NANOG mRNA in the iRPE cells were significantly lower than those in the control group (t =13.086,P =0.022;t =34.186,P =0.004),and the relative expression levels of Pax6,RX,CRALBP and RPE65 mRNA were higher in the curcumin group than those of the control group (all at P＜0.01).Western blot assay showed that the expressing bands for CRALBP,RPE65 and MITF enhanced in iRPE cells with a similar appearance in human RPE cells.However,these expressions were all absent in human ESCs.Immunofluorescence staining showed the positive expressions of Pax6,MITF and ZO-1 in cytoplasm of iRPE cells in the curcumin group with a purified efficacy 100％.The fluorescence dye-doped polystyrene microspheres in cytoplasm were obvious in the iRPE cells like positive controls,but the polystyrene microsphere was absent in the negative controls.From 3 weeks through 5 weeks after induced,the relative expression levels of Lef1,MYC and TCF7 mRNA (the dwnstream target genes of Wnt signaling pathway),FZD3 mRNA (Wnt receptor),Wnt2B mRNA (Wnt ligand) and Wnt7B mRNA were significantly reduced in the curcumin group compared with the control group (all at P＜0.01).Conclusions Curcumin promotes the differentiation of human ESCs into RPE-like cells by stimulating the activation of Wnt signaling pathway,and therefore accelerate the differentiation and mature of iRPE cells.

Development of Syllabuses for Biological Nursing Science Subjects based on Learning Outcomes: Structure and Function of Human Body, Pathogenic Microbiology, Pathophysiology, and Mechanisms and Effects of Drugs

PURPOSE: The purpose of this study was to develop syllabuses of basic nursing science subjects (Structure and Function of Human Body, Pathogenic Microbiology, Pathophysiology, and Mechanisms and Effects of Drug) based on learning outcomes. METHODS: We developed a 3-phase plan to develop the syllabuses. In the first phase, The Korean Society of Biological Nursing Science (KSBNS) held a workshop in May, 2013 in which professors who are in charge of basic nursing science subjects shared opinions about learning outcomes. As a result, initial prototype syllabuses came out. In the second phase, revised syllabuses based on learning outcomes were presented and discussed in the conference held by KSBNS in November, 2013. In the last phase, a research team who taught basic nursing science subjects finalized the syllabuses during the meetings. RESULTS: Syllabuses of 4 basic nursing science subjects were developed. CONCLUSION: These syllabuses of 4 subjects need to be disseminated throughout nursing colleges. Further revision needs to be made according to the circumstances and context of each school.

Neuroprotective effects of minocycline on retinal ganglion cells in early stage of optic nerve crush injury / 中华实验眼科杂志

Background Minocycline possesses neuroprotective effect in a variety of animal models and clinical trials of central nervous system,but whether it works on optic nerve injury remains unclear.Objective This study aimed to observe the protective effects of minocycline on retinal ganglion cells (RGCs) in the early stage of optic nerve crush and explore its mechanism.Methods One hundred and thirty-six clean C57BL/6J mice were randomly divided into normal control group,normal saline solution group and minocycline group.The optic nerve crush injury models were induced in the left eyes of the mice in the normal saline solution group and minocycline group by a cross-action forceps for 3 seconds.Minocycline was injected intraperitoneally in the minocycline group firstly 45 mg/kg(0.4 ml) and followed by 22.5 mg/kg per day after 24 hours until sacrifice of the animals,and the equivalent volume of normal saline solution was injected in the same way in the normal saline solution group.The mice were euthanized at 4,7,11,14 days postoperatively and the left eyeballs were collected.Retinal flat mounts and DAPI staining was used to observe and compare the change of RGCs density among different groups and various time points.Apoptosis of mice RGCs were assessed by TdT-mediated dUTP nick end labeling (TUNEL).Real-time polymerase chain reaction (real-time PCR) was used to detect the expression of CD11b mRNA in retinal microglials.Results DAPI staining in retinal flat mounts showed that the average RGCs density was (77.50±2.38)/0.01 mm2 and (70.00±2.94) /0.01 mm2 in the 4th and 7th day after modeling in the normal saline solution group,and those in the minocycline group were (88.75 ± 2.36) /0.01 mm2 and (81.00 ± 3.92)/0.01 mm2,with significant differences between the two groups (t4d =-6.708,P＜0.01 ;t7d =--4.491,P＜0.01).The apoptotic RGCs were (12±1)/mm and (4±1)/mm in the normal saline solution,which were significantly more than (4±1)/mm and (1±0)/mm in the minocycline group (t4 d =12.832,P＜0.01 ; t7d =3.455,P =0.026).However,no significant difference was found in apoptotic RGCs in postoperative 11 days and 14 days between the normal saline solution group and the minocycline group (P =0.708,0.777).The expressing levels of CD11 b mRNA in the retinal microglials were significantly higher in the 4th and 7th day in the normal saline solution group than those in the minocycline group (t4 d =8.312,P＜0.01 ;t7d=5.407,P＜0.01),but were not significantly different in the 11st and 14th day after modeling between the two groups (P=0.055,0.170).Conclusions Minocycline can play a neuroprotective effect on RGCs in the early stage of optic nerve crush in mice by inhibiting microglia activation and decreasing RGCs apoptosis.

Influence of triamcinolone acetonide on peribulbus tissues via periocular injection / 中华实验眼科杂志

Background Triamcinolone acetonide (TA) is often used in the treatment of uveitis and fundus disease,but whether it has toxic effect to normal tissue around eyes is unclear.Objective The aim of this study was to explore the adverse and toxic effects of TA on the periorbitally normal tissue following the periocular injection.Methods Twenty-seven New Zealand albino rabbits were randomly divided into experimental group,normal saline solution group and blank control group.TA was periocularly injected twice (20 mg for each) at 1-month interval in the rabbits of the experimental group,and 0.5 ml normal saline solution was used in the same way in the rabbits of the normal saline solution group.Not any drug was used in the blank group.The rabbits were sacrificed 1 month and 2 months,3 months after the secondary injection.For the preparation of the specimens of extraocular muscles,peribulbus adipose tissue,lacrimal gland and optic nerve.Hematoxylin and eosin staining of the specimens was performed to examine the the pathological change under the optical microscope,and the ultrastructural of peribulbus tissues was observed under the transmission electron microscope.Results No statistically significant change was found in IOP among various time points and groups (Fgroup =0.952,P =0.881 ; Ftime =7.297,P =0.411).The hyphological structures and ultrastructure of extraocular muscles,peribulbus adipose tissue,lacrimal gland and optic nerve were normal in the rabbits of the normal saline solution group and the blank control group.However,the optical microscopy showed varying degrees of atrophy or dissolving in the extraocular muscle fibers,the increase of glandular epithelial cells and enlargement of nuclei in the lacrimal gland as well as visible disorder arrangement of nerve fibers,vacuoles degeneration and the decrease of glial cells in optial nerve tissue in the rabbits of the experimental group from 1 month though 3 months after the secondary injection.Under the transmission electron microscope,the muscle stripes,bright band and dark band were incompletely disappeared,and expansionary sarcoplasmic reticulum and oval nucleus at the inferior to sarolemma were seen in the extraocular muscles.In addition,swelling lacrimal gland epithelial cells,thickening nuclear membrane and more organelles were exhibited in the lacrimal gland.In the optic never tissue,uneven myelin,space between axons and myelin,lamellar separation and degeneration also were revealed in the rabbits of the experimental group after injection of TA.Peribulbus adipose tissue was near normal both by optical microscope and transmission electron microscope in the experimental group.Conclusions The periocular injection of TA dose not elevated the IOP.However,it will result in the histogical and ultrastructural damages of extraocular muscles,lacrimal gland and optic nerve.

The randomized controlled study on timolol preventing refractive regression after LASIK in high myopic eyes / 中华实验眼科杂志

Background Refractive regression is a common complication of high myopia after laser in situ keratomileusis (LASIK),and it affects the stability of surgery.Objective This study was to observe the preventive effect of timolol on refractive regression in high myopia after LASIK.Methods A perspective randomized controlled trial was performed under the approval of Affiliated Hospital of Zunyi Medical College and the informed consent of the patients.Sixty eyes of 60 patients with high myopia (-7.16±0.95) D for LASIK were randomized into experimental group and control group.Regular eye drops were topically administered in the patients after LASIK in both groups,and timolol 0.5 ％ was added topically from 1 day after LASIK.The uncorrected visual acuity (UCVA),best corrected visual acuity (BCVA),spherical equivalent (SE),corneal anterior surface curvature,intraocular pressure (IOP),central corneal thickness (CCT) were measured before LASIK,1 week,1 month,3 months and 6 months after LASIK,respectively.Residual stromal bed thickness was calculated before LASIK.The differences of above-mentioned indexes were analyzed and compared between the two groups and among the various time points using repetitive measurements analysis of variance,independent simple t test and Bonferroni test.Results No significant differences were found in the demography between the experimental group and the control group,including age,UCVA,BCVA,SE,IOP,corneal anterior surface curvature,CCT and residual stromal bed thickness (all at P＞0.05).UCVA was significantly different between the two groups among various time points (Fgroup =3.91,P＜0.05 ; Ftime =3.80,P＜0.05),and the UCVA was significantly higher in the experimental group than the control group 6 months after LASIK (t=2.97,P＜0.05),and UCVA was gradually increased as the lapse of postoperative time with the significant difference between 7 days and 6 months after LASIK in the experimental group (P＜0.05).No significant difference was seen in the BCVA between the two groups at various postoperative time points (Fgroup =2.44,P＞0.05;Ftime =2.31,P＞0.05).SE refraction in both groups were significantly reduced from 7 days through 6 months after LASIK,showing significant difference between the groups at various time points after LASIK(Fgroup =11.52,P＜0.05;Ftime =22.06,P＜0.05).The SE refraction was higher in the experimental group than that in the control group 6 months after LASIK (t =2.47,P＜0.05).Corrected IOP in the experimental group was lower than that of the control group at 7 days,1 month and 3 months after LASIK,respectively (Fgroup =14.83,P＜0.05).The change of CCT was not statistically different between the control group and the experimental group (Fgroup =0.04,P＞0.05).The anterior corneal surface curvature was steady in the experimental group during the following-up duration after LASIK,while the control group was growing gradually (Ftime =18.73,P＜0.05).Conclusions The study data show that topical administration of 0.5％ timolol early in high myopia following LASIK is effective for the reduction of the refractive regression.It is suggested that 0.5％ timolol can prevent cornea ectasia by lowering IOP.Reduction of the IOP may contribute to improving the regression after keratorefractive surgery in high myopic eyes.