Objective To investigate the effect of dissociation of the human retinal pigment epithelium (RPE) cells by ficoll hypaque gradient centrifugation. Methods The primary human RPE cells and subcultured human RPE cells were dissociated with ficoll gradient centrifugation solution (d= 1.077 g/ml) and the same divided cells as the control were dissociated with routine normal culture medium centrifugation. The Trypan blue (0.4%) rejection staining was used, and the mouse anti-human monoclonal antibody and fluorescein isothiocyanate (FITC) labeled rabbit anti-mouse IgG were utilized for indirect immunoreactivity for the test of human cytokeratin (CK) in active RPE cells cytoplasm. Flow cytometry assay was used to analyzed the percentages of CK positive staining RPE cells. simultaneously, the cells configuration, growth condition, the rate of clone formation, and the purifying result were observed under the fluorescent and confocal microscope. Results The survival rate and positive rate of CK of RPE cells in experimental group were higher than those in the control ( P

Objective　To investigate the expression and role of inflammatory cytokines like interleukin-1β(IL-1β)、interleukin-6(IL-6)、interleukin-8(IL-8) and tumor necrosis factor-alpha(TNF-α)in epiretinal membranes(ERM)of eyes with proliferative vitreoretinopathy(PVR).Methods　Nineteen epiretinal membranes were obtained from eyes undergoing vitrectomy for retinal detachment complicated with PVR and observed by immunohistochemical methods.Results　Expression of IL-1β、IL-6、IL-8 and TNF-α were observed in 9、12、11and 15 membranes respectively, with positive staining mostly in the extracellular matrix of epiretinal membranes. Only one membrane showed positive to IL-6 intracellularly, expression for all the cytokines simultaneously in 4 membranes.Conclusion　The findings indicate that cytokine-mediated pathways of inflammation are involved in the pathogenesis of PVR.

AIM: To investigate changes of Ca 2+ activated potassium channels (K Ca) in autogenous vein grafts. METHODS: The contraction of venous ring was measured by means of perfusion in vitro. The intimal proliferation and proliferation of cultured smooth muscle cells (vascular smooth muscle cells, VSMCs) were observed by the means of computerised image analysis and MTT method, respectively. Furthermore, whole cell mode of patch clamp was used to record K Ca of VSMCs isolated from autogenous vein grafts. RESULTS: 1 week after transplantation there were no significant differences of contraction and intimal relative thickness between autogenous vein grafts and control. Contraction and intimal relative thickness of autogenous vein graft were significantly increased 2 weeks after transplantation (P

Transient receptor potential A1 (TRPAl) is a cold sensitive cation channel, which could also be activated by various pungent compounds. As a transduction channel in a number of sensory modalities, TRPAl expressing in heterogonous systems serves to provide great convenience in pharmacological analysis and functional investigation. Due to cellular toxicity, establishment of stable TRPAl cell line has always been challenging. Nevertheless, the first stable human embryonic kidney (HEK-293) cell line with un-controlled expression of TRPAl was successfully established. It was also confirmed that this stable cell line retained TRPAl expression for more than 25 passages in culture. The functional analysis of the cell response verified the stability and specificity of this novel recombinant TRPAl cell line. Altogether, the data indicated this TRPAl-HEK cell line would be a useful tool for functional analysis of TRPAl and for the development of high throughput screening (HTS) compatible assay in the effort to identify TRPAl modulators.

Objective:To study influence of intravenous injection urapidil and nitroglycerin micro pump on blood pressure and heart rate (HR) of hypertension patients undergoing tooth extraction.Methods:116 hypertension patients underwent electrocardiographic monitoring tooth extraction in our hospital from January 2015 to October 2016 were enrolled in this study.According to the random number table method,the patients were divided into observation group and control group with 58 cases,the control group was given nitroglycerin plus pump static point to maintain,the observation group was given intravenous urapidil and pump maintenance,compared the changes of systolic blood pressure (SBP),diastolic blood pressure (DBP) and HR in two groups before operation,at anesthesia,10min after anesthesia,in operation,10 min after operation,and compared the adverse reactions condition of the two groups.Results:The levels of SBP and DBP in two groups in operation and 10 min after operation were significantly lower than that before operation,and the SBP and DBP levels in the observation group in operation were significantly lower than control group,the differences were statistically significant (all P＜0.05).The HR in control group in operation,10 min after operation were significantly higher than before operation,while the observation group were significantly lower control group,the differences were statistically significant (P＜0.05).The total incidence of adverse reactions in the observation group was 6.90％(4/58),which has no significant difference than 10.34％ (6/58) in control group (P＞0.05).Conclusion:Intravenous injection urapidil has little effect on blood pressure and HR in hypertension patients undergoing tooth extraction,and with good safety,which is worthy of promotion.

Objective To investigate the possible damaging effect of infrasound on ultrastructure and permeability of rats′ blood retinal barrier (BRB). Methods Twenty mature male rats, averagely divided into 5 groups according to the exposure duration, were exposed to infrasound at a 16 Hz frequency and 130 dB sound pressure level in a pressure chamber for 2 hours per day. After exposed for 0, 1 day, 7, 14, and 21 days respectively, ultrastructural changes of rats′ BRB were observed through injection of lanthanum (La) nitrate solution, which was used as a tracer to demonstrate the breakdown of the BRB. Results With prolonging the duration of infrasound exposure, BRB structure lesion, chondriosome tumefaction, endoplasmic reticulum expandedness, membrane disc damage, retinal pigment epithelial cells distortion and putrescence, karyotheca expandedness, and La leakage on each level of retina aggravated gradually. Conclusion Infrasound may cause the breakdown of BRB, and the lesions aggravated with prolonged infrasound exposure time.

Objective Our previous study showed that cycloxygenase-2 (COX-2) and vascular endothelial growth factor (VEGF) are expressed in choroidal neovascularization (CNV) and the expression of COX-2 is prior to VEGF, indicating that COX-2 is probably one of upstream regulatory factors of VEGF. The aim of this study was to observe inhibition and mechanism of intravitreous injection of celecoxib, a selective COX-2 inhibitor, on experimental choroidal neovascularization in a laser-induced rat model. Methods Retinal photocoagulation was performed in 36 right eyes of 36 male Brown Norway rats to establish CNV models with the laser parameter as follows:wavelength 532 nm, power 80 mW, spot diameter 100 p, m and time shutter 100 ms. Eight or ten spots were irradiated in the position of 1.5 - 2. 0 PD to optic disc. Celecoxib or normal saline solution was intravitreously injected via scleral incision in 18 right eyes of 18 rats, respectively. The thickness and area of CNV were qualified by HE staining(n =3) and by choroidal flatmount (n =3) at day 14 after photocoagulation under the light microscope. The expressions of VEGF and COX-2 in RPE-choroid-sclera complex were examined by Western blot(n =6) and RT-PCR(n =6) at day 7 after photocoagulation. The experimental procedure followed the Standard of Association for Research in Vision and Ophthalmology. The license for animal administration was obtained. Results After intravitreous injection with celecoxib, the thickness and area of CNV were significantly smaller in celecoxib group than normal saline group on 14 days (69. 75 μm ± 7. 50 μm vs 45. 84 μm ± 5. 59 μm in thickness and 87 854 pixel~2 ± 6 735 pixel~2 vs 61 101 pixel~2 ± 6 314 pixel~2 in area, P =0.00). The expressions of VEGF protein and mRNA were obviously lower in celecoxib group compared with normal saline group(t = 3. 755, P = 0.02; t =3. 155, P =0. 03) . No significant difference was found in expression of COX-2 mRNA between the two groups (t = 0. 581, P = 0. 59). Conclusion Intravitreous injection of celecoxib can effectively inhibit CNV by downregulating VEGF level, which is a new approach for the treatment of CNV.

Clinical practice is the critical stage of training medical students in basic clinical quality.For the current major issues,such as lack of clinical case resources,lack of teaching conditions,and less chances of practice for students,cases library combined with medical human patient simulator is used in extern's teaching.Through this teaching,the students can touch numerous and comprehensive clinical cases,grasp clinical techniques proficiently,predominate diagnosis and treatment of common diseases in Orthopaedics efficiently by using medical human patient simulator.The effect of clinical practice teaching is improved greatly.The practice of related teaching means and methods provides a new way for the reform of the practice teaching.

OBJECTIVETo investigate the effect of cartilage-derived growth factor (CDGF) on cultured rabbit chondrocytes, and the relation between CDGF, insulin-like growth factor-I (IGF-I), and proliferation and metabolism of chondrocytes.

METHODSCDGF was extracted from chicken xiphoid in our laboratory. Chondrocytes were isolated from 3-week-old New Zealand male rabbits, and cultured in Ham's F-12 medium containing 10% fetal calf serum. The chondrocyts of second generation were treated with CDGF and/or IGF-I of different concentrations. With chloramine T method and MTT method, we compared the content of hydroxyproline in Ham's F-12 medium and observed proliferation and energy synthesis of chondrocytes.

RESULTSCDGF and IGF-I both stimulated the proliferation and synthesis of hydroxyproline of chondrocytes dose-dependently. The optimal concentration of CDGF was 16 ng/ml and 32 ng/ml respectively, and that of IGF-I was 30 ng/ml. There was obvious synergic effect between CDGF and IGF-I.

CONCLUSIONCDGF can stimulate the proliferation and collagen synthesis of chondrocytes and has synergistic effect with IGF-I.

Animals ; Cartilage ; metabolism ; Cell Division ; drug effects ; Chondrocytes ; drug effects ; physiology ; Dose-Response Relationship, Drug ; Drug Synergism ; Hydroxyproline ; biosynthesis ; Insulin-Like Growth Factor I ; pharmacology ; Male ; Proteins ; pharmacology ; Rabbits