Preparation and In Vitro Characterization of Gelatin Methacrylate for Corneal Tissue Engineering

BACKGROUND@#Corneal disease is second only to cataract considered as the leading cause of blindness in the world, with high morbidity. Construction of corneal substitutes In Vitro by tissue engineering technology to achieve corneal regeneration has become a research hotspot in recent years. We conducted in-depth research on the biocompatibility, physicochemical and mechanical properties of rat bone marrow mesenchymal stem cells (rBM-MSCs)-seeded gelatin methacrylate (GelMA) as a bioengineered cornea. @*METHODS@#Four kinds of GelMA with different concentrations (7, 10, 15 and 30%) were prepared, and their physicchemical, optical properties, and biocompatibility with rBM-MSCs were characterized. MTT, live/dead staining, cell morphology, immunofluorescence staining and gene expression of keratocyte markers were performed. @*RESULTS@#7%GelMA hydrogel had higher equilibrium water content and porosity, better optical properties and hydrophilicity. In addition, it is more beneficial to the growth and proliferation of rBM-MSCs. However, the 30%GelMA hydrogel had the best mechanical properties, and could be more conducive to promote the differentiation of rBM-MSCs into keratocyte-like cells. @*CONCLUSION@#As a natural biological scaffold, GelMA hydrogel has good biocompatibility. And it has the ability to promote the differentiation of rBM-MSCs into keratocyte-like cells, which laid a theoretical and experimental foundation for further tissue-engineered corneal stromal transplantation, and provided a new idea for the source of seeded cells in corneal tissue engineering.

A review of advances in intraocular fluid detection for high myopia and its relevant fundus diseases / 中华眼底病杂志

The fundus lesions caused by high myopia (HM) often lead to irreversible visual impairment or even blindness. However, the pathogenesis of HM and its fundus lesions is still unclear, the intraocular fluid detection technology of micro samples has brought new prospects for the early diagnosis, monitoring and intervention of the fundus lesions. The molecules associated with HM are various and functionally diverse, intermolecular interactions are staggered and the specific mechanism is complex. With the development of intraocular fluid detection technology, while gradually revealing the role of each molecule in the pathogenesis of HM, it is expected to successfully assist clinical work in the future, providing outpost markers for the progress of myopia and targets for early intervention, or providing a new therapy choice for HM fundus lesions at the molecular level targeting pathogenesis, which is expected to provide more accurate and effective treatment for HM patients in the future.

A review of advances in intraocular fluid detection for high myopia and its relevant fundus diseases / 中华眼底病杂志

The fundus lesions caused by high myopia (HM) often lead to irreversible visual impairment or even blindness. However, the pathogenesis of HM and its fundus lesions is still unclear, the intraocular fluid detection technology of micro samples has brought new prospects for the early diagnosis, monitoring and intervention of the fundus lesions. The molecules associated with HM are various and functionally diverse, intermolecular interactions are staggered and the specific mechanism is complex. With the development of intraocular fluid detection technology, while gradually revealing the role of each molecule in the pathogenesis of HM, it is expected to successfully assist clinical work in the future, providing outpost markers for the progress of myopia and targets for early intervention, or providing a new therapy choice for HM fundus lesions at the molecular level targeting pathogenesis, which is expected to provide more accurate and effective treatment for HM patients in the future.

Inhibitory effects of Genistein on hyperoxia-induced retinal neovascularization / 中华实验眼科杂志

Objective To investigate the effect of Genistein,a tyrosinase inhibitor,on retinal neovascularization in mice.Methods Thirty-six 7-day-old C57BL/6J mice were randomly assigned into hyperoxiainduced group,Genistein group,DMSO group and normal control group.The mice in the hyperoxia-induced group,Genistein group and DMSO group were fed in a static chamber with the oxygen volume fraction (75±2)％ for 5 days and then sent back to natural environment for 5 days to establish retinal neovascular models,and 1 μl Genistein diluted by 5％ dimethyl sulfoxide (DMSO) (400 mg/L) and 1 μl DMSO were intravitreally injected in the 12-dayold mice of Genistein group and DMSO group,respectively.The mice in the normal control group were bred in natural environment.The fluorescence angiography was carried out in 17-day-old mice (2 mice in each group) to prepare the whole retinal flatmounts,and the morphology of retinal vessels was observed under the fluorescence microscope.The other mice in various groups were sacrificed and the retinas were collected.The expressions of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) in mRNA and protein levels were detected by realtime fluorescence quantitative PCR and Western blot,respectively.The use and care of the mice complied with regulations for the management of laboratory animals.Results Retinal vessels were normal in the mice of normal control group.In the mice of the hyperoxia-induced group and DMSO group,retinal vessels were tortuous,and neovacularization and non-perfusion areas were visible.In the Genistein group,retinal vessels were clearly visible,but non-perfusion areas were exhibited.The relative expression levels of VEGF mRNA in retinas were 0.64±0.25,0.37±0.23,0.03±0.02 and 0.04±0.02,and the relative expression levels of bFGF mRNA in retinas were 21.40±3.07,17.22±2.63,0.52±0.25 and 0.67±0.23,in the hyperoxia-induced group,DMSO group,Genistein group and normal control group,and the expressions of VEGF and bFGF in mRNA level in the hyperoxia-induced group and DMSO group were significantly higher than those in the Genistein group and normal control group (all at P＜0.05).The protein expression levels of VEGF and bFGF were 1.01 ±0.05 and 0.97±0.06 in the hyperoxia-induced group,1.06±0.07 and 1.03±0.08 in the DMSO group,0.73±0.05 and 0.76±0.07 in the Genistein group,0.52±0.05 and 0.56± 0.05 in the normal control group.The expressions of VEGF and bFGF in protein level in the hyperoxia-induced group and DMSO group were significantly higher than those in the Genistein group and normal control group (all at P＜0.05).Conclusions Genistein can inhibit hyperoxia-induced retinal neovascularization may be by downregulating the expressions of VEGF and bFGF in retinas.

Effect of Cathepsin B and vascular endothelial growth factor on the retinal neovascularization induced by hyperoxia / 中华实验眼科杂志

Background Vascular endothelial growth factor (VEGF) plays a key role in the vascular development and neovascularization.Studies have proved that Cathepsin B is related to the formation of neovascularization,but its mechanism is unclear.Objective This study was to investigate the expression of Cathepsin B and VEGF in the retinal neovascularization induced by hyperoxia and the relationship between them.Methods Forty-four 7-day-old C57BL/6J mice were randomly assigned into 4 groups:normal control group,hyperoxia-induced group,normal control (NC)-green fluorescent protein (GFP)-lentivirus (Lv) group and Cathepsin B-RNA interference (RNAi)-Lv group,with 11 mice 22 eyes for each group.The mice in normal control group were survival in natural environment.The other three groups of 7-day-old mice were put in a sealed box of oxygen volume fraction (75±2)％ for 5 days and then sent back to normal environment.The mice of hyperoxia-induced group did not have any drug intervention,while the 12-day-old mice of NC-GFP-Lv group and Cathepsin B-RNAi-Lv group received intravitreal injection of NC-GFP-Lv 1 μl or Cathepsin B-RNAi-Lv 1 μl.All 17-day-old mice were sacrificed and the retinas were collected.The mRNA expression levels of Cathepsin B and VEGF were performed by real-time PCR;the protein expression levels of Cathepsin B and VEGF were detected by Western blot.Results Fluorescence microscope results showed that the layer and branch of retinal neovascularization were less in the Cathepsin B-RNAi-Lv group than those in the NC-GFP-Lv group and hyperoxia-induced group.The relative expression levels of Cathepsin B and VEGF mRNA in each group were significantly different (F =444.89,P =0.00;F =519.78,P =0.00).The relative expression levels of Cathepsin B and VEGF mRNA in hyperoxia-induced group,NC-GFP-Lv group and Cathepsin B-RNAi-Lv group were higher than those in normal control group,and the relative expression levels of Cathepsin B and VEGF mRNA in Cathepsin B-RNAi-Lv group were lower than those in hyperoxia-induced group and NC-GFP-Lv group,with significant differences between them (all at P ＜ 0.05).The relative expression levels of Cathepsin B and VEGF protein in each group were significantly different (F =54.37,P =0.00;F =79.65,P =0.00).The relative expression levels of Cathepsin B and VEGF protein in hyperoxia-induced group,NC-GFP-Lv group and Cathepsin B-RNAi-Lv group were higher than those in normal control group,and the relative expression levels of Cathepsin B and VEGF protein in Cathepsin B-RNAi-Lv group were lower than those in hyperoxia-induced group and NC-GFP-Lv group,with significant differences between them (all at P＜0.05).Conclusions The expression levels of Cathepsin B and VEGF in retinal neovascularization of hyperoxia-induced group were significantly higher than those in normal control group,Cathepsin B-RNAi-Lv can inhibit the mRNA and protein expression of Cathepsin B and VEGF,the expression of VEGF may be influenced by the expression of Cathepsin B.

Optimization of transfect conditions for cathepsin B-RNAi-lentivirus to the brain micro vascular endothelial cells / 中国综合临床

ObjectiveTo Research on the best condition of transfecting RNAi plasmid into the mice brain micro vascular endothelial cells to interfere the tissue enzymes and protease through the cathepsin B-RNAi-lentivirus.MethodsThe brain micro vascular endothelial cells ( bEND.3 ) were divided into three groups at random:culture medium completely group (NP group),culture medium with 5 mg/L polybrene group ( P1 group) and culture medium with 8 mg/L polybrene group ( P2 group),each group has five different dimensional MOI (1,5,10,15,20),after transfected,48 h,72 h,96 h,the fluorescence rate of GFP protein were counted by a fluorescence microscope to obtain the transfected rate,the MOI of the best optimization condition were observed;After one week,the effect of the most appropriate of MOI to cytoactive and survival rate calculated through MTT.ResultsA small amount of green fluorescent protein(GFP) in each group were observed after transfected for 48 h,the transfection rate of each group were 1％ -9％ ;In the same MOI groups,there was no significant difference between the 72 h and 96 h ( P ＞ 0.01 ) ; In groups of MOI ( 1,5,15 ),the prevalence transfection rate of each group were lower than 20％ and in MOI(10,20) group,the transfection rate were between 71％ -97％ and 60％ - 84％,and the transfection rate in group 10 was significantly higher than group 20 ( P ＜ 0.01 ) ; P1 group showed significantly higher survival rate than that in P2 group by MTT assay [ ( 81.38 ± 6.69 ) ％ vs (62.03 ± 5.13)％,t =15.795,P ＜ 0.001 ].ConclusionThe most optimal conditions of CathepsinB-RNAi-Lentivirus infection bEND.3 depend on MOI(10),polybrene(5 mg/L) and the transfection time of 72 h.

An improved method for primary culture of mouse pulmonary microvascular endothelial cells / 南方医科大学学报

<p><b>OBJECTIVE</b>To establish an improved method for culturing primary mouse pulmonary microvascular endothelial cells (PMVECs).</p><p><b>METHODS</b>An improved tissue block adherent culture method was used to isolate and culture the PMVECs from C57 mice. The cultured cells were identified by factor VIII-related antigen and CD31 antigen, and the growth of cells cultured using the improved method and the conventional method was compared.</p><p><b>RESULTS</b>The cultured primary pulmonary microvascular endothelial cells showed a short fusiform or round morphology, and the cell monolayer displayed a cobble stone-like appearance. The cultured cells were positive for VIII-related antigen and CD31 antigen. The cell growth was accelerated in the cell cultures with the improved method compared with that in conventional cell cultures.</p><p><b>CONCLUSION</b>The improved culture method allows more efficient acquisition of primary mouse PMVECs of a greater purity.</p>

Quality Analysis of Finished Water of Municipal Centralized Water Supply System in Shenzhen / 环境与健康杂志

Objective To investigate the water quality of finished water of the centralized water supply system in Shenzhen according to 106 indexes of water quality standards of China.Methods The collection and preservation of water samples was according to the standard examination methods for drinking water-collection and preservation of water samples (GB/T 5750.2-2006).The sanitary quality of the finished water samples collected from 35 centralized water supply systems in cities were determined and evaluated according to the Standards for Drinking Water Quality (GB 5749-2006) in May of 2008.Results The average qualified rate of drinking water was 82.9% (29/35) in Shenzhen.Among 106 indexes,4 indexes (such as turbidity,aluminum,manganese and free chlorine residue) exceeded the standard limits in degrees in some centralized water supply system.Conclusion According to the results of the present paper,it is considered that the water quality of the product water from the centralized water supply system in Shenzhen is good.

Statistical Results of the Cochlear Implant Case Study Using a Database Application Software / 대한의료정보학회지

OBJECTIVE: Using the CI database application software and SPSS to process cochlear implanted patients' data, in order to know what factors influence the cochlear implant effects. METHODS: Design a cochlear implant questionnaire; get answers from young patients' parents through telephone calls. Input the patients' data to the database with our software, analyze the data using SPSS. RESULTS: 120 cochlear implant recipients were investigated. Those patients are all between 9 and 20 years old. 106 parents answered the inquisition. Some beneficial statistical results like Communion ability and method, comprehension ability, and other factors that influence CI effect were elicited. CONCLUSION: Even for Pre-lingual deaf adolescent, cochlear implantation is still a good choice to acquire better communion ability and to improve the life quality. The statistical analysis results of our follow up investigation indicated that the earlier the speech training intervention, and adopting lip-reading, combining with scientific language training after CI operation, the better CI effect could be acquired. And also the speech communion ability could be improved quickly. Besides, the CI database application software provided great advantages to this study.