Development of a sandwich ELISA for detecting 3AB non-structural protein of foot-and-mouth disease virus / 生物工程学报

Antigenic purity is important for quality control of the foot-and-mouth (FMD) whole virus inactivated vaccine. The recommended method for evaluation the antigenic purity of FMD vaccine is to check the serum conversion to non-structural protein (NSP) 3AB antibody after 2 to 3 times inoculation of animals with inactivated vaccine. In this study, we developed a quantitative ELISA to detect the amount of residual 3AB in vaccine antigen, to provide a reference to evaluate the antigenic purity of FMD vaccine. Monoclonal antibody (Mab) of NSP 3A and HRP-conjugated Mab of NSP 3B were used to establish a sandwich ELISA to quantify the NSP 3AB in vaccine antigen of FMD. Purified NSP 3AB expressed in Escherichia coli was serially diluted and detected to draw the standard curve. The detectable limit was determined to be the lowest concentration of standard where the ratio of its OD value to OD blank well was not less than 2.0. Results: The OD value was linearly corelated with the concentration of 3AB protein within the range between 4.7 and 600 ng/mL. The correlation coefficient R² is greater than 0.99, and the lowest detectable limit is 4.7 ng/mL. The amount of 3AB protein in non-purified inactivated virus antigen was detected between 9.3 and 200 ng/mL depending on the 12 different virus strains, whereas the amount of 3AB in purified virus antigen was below the lowest detectable limit. The amount of 3AB in 9 batches of commercial FMD vaccine antigens was between 9.0 and 74 ng/mL, whereas it was below the detectable limit in other 24 batches of commercial vaccine antigens. Conclusion: the sandwich ELISA established in this study is specific and sensitive to detect the content of 3AB protein in vaccine antigen of FMD, which will be a useful method for evaluation of the antigenic purity and quality control of FMD inactivated vaccine.

Expressions of TLR4 and Dectin-1 in cornea tissue of rats with Aspergillus fungal keratitis and their effects in pathogenesis of keratitis / 吉林大学学报(医学版)

Objective To explore the expressions of Dectin-1 and Toll-like receptor 4 (TLR4)and the secretory charateristics of inflammatory factors of the Aspergillus fumigatus infected cornea of the rats,and to clarify theirs role in the pathogenesis of rat keratitis.Methods 40 SD rats were separated into control group (n = 10)and aspergillus keratitis group (n=30).The rats in aspergillus keratitis group were infected by Aspergillus fumigatus in two eyes for 12,24 and 48 h,and the cornea tissue was took and ELISA kit was used to detect the level of inflammatory cytokines IL-1,TNF-a,IL- 6,IL-10,and NF-κB;immunohistochemistry,Western blotting and RT-PCR methods were used to determine the Dectin-1 and TLR4 protein and mRNA expressions.Results The rat models were successfully established.The eyes of the rats in control group were limpid tested by corneal slit lamp, however,there existed a clear boundary film on the surface of eyes of the rats in aspergillus keratitis group infected for 12 h,white infiltrates and bloodshot in the eyes in 24 h group and the white infiltrates were further strengthened and almost covered the entire eyes in 48 h group. The order of inflammatory cytokines IL-1,TNF-α,IL-6,IL-10, NF-κB was:48 h group> 24 h group> 12 h group> control group (P < 0.05),so did the expression levels of Dectin-1 and TLR4 protein and mRNA in various groups tested by Western blotting and RT-PCR methods.The expression levels of Dectin-1 and TLR4 were increased with the prolongation of time investigated by immunohistochemical staining.Conclusion Dectin-1 and TLR4 are secreted in excess after the eyes are infected with fumigatus,which promotes the expressions of related inflammatory cytokines and results in the occurrence of keratitis.

Prediction analysis of ametropia in children aged from 5 to 13 years old / 重庆医学

Objective To investigate the characteristics of children ametropia in the development process ,predict and analyze the refractive state with the measurement data of refractive factors on the children aged from 5 to 13 years old .Methods 461 children (922 eyes) ,aged from 5 to 13 years old ,were selected from outpatients who went to ophthalmology clinic from January 2011 to September 2012 .The static diopter was measured by the optometry with the continuous mydriasis of 5 days using the ophthalmic gel of 1% atropine .And the biological measured values of refractive factors were obtained by the A type ultrasonic and keratome-ter .Meanwhile ,the parameters including the height (H) ,weight and the dominant eye were recorded to analyze the characteristics of children ametropia in the development process ,and then the equation between the diopter and the correlative factors including the measurement data of refractive factors and growth factors was established by the stepwise multiple regression analysis .Results (1) The myopia degree of the right eye static diopter was significantly deeper than the left (P0 .05) .In the group in which the right eye was dominant ,the difference of the left eye and right eye refractive factors was not statistically significant (P>0 .05) except the instance in which K1 of the right eye was bigger than the left eye(P<0 .05) .(2) In the period of 5 to 13 years old ,the myopia developing trend of girls was earlier than boys .(3) The ocular axis length(AL) and H development had a close relationship ,and the correlation equation of them was AL =16 .135+0 .052H(r=0 .993) .(4) The multiple linear regression equation for the relationship between the static diopter (D) ,ocular AL and mean value of the corneal curvature (K) was D=84 .271-0 .937K -1 .878AL(r=0 .871) .Conclusion The ocular AL and H development of children have a close relationship .The static diopter(D) of the children aged from 5 to 13 years old can be predicted by the ocular axis length(AL) and corneal curvature(K) , w hich can be used to prevent myopia reasonably .

Enhanced immune response of a novel T-cell immunogen in vaccine for foot-and-mouth disease / 生物工程学报

We investigated the enhanced immune response of a recombinant T cell immunogen as an effective cellular immune adjuvant. The T cell immunogen named TI contained several T cell epitopes from the VP1, VP4, 3A and 3D proteins of foot-and-mouth disease virus (FMDV) and two pan-T helper (T(H)) cell sites to broaden the immunogenicity of the protein. Meanwhile, another fusion protein named OA-VP1 was expressed in bacteria, which contained two VP1 proteins of O and Asia1 type FMDV. Mice were vaccinated with commercially inactivated vaccine or OA-VP1 protein with or without the TI immunogen. The results show that mice inoculated with inactivated vaccine or OA-VP1 protein supplemented with TI immunogen produced significantly higher level of neutralizing antibodies (P < 0.01 or P < 0.05) than the mice only inoculated with inactivated vaccine or OA-VP1 protein by microneutralization assay. An obvious increase in T cell number by flow cytometric analysis and significantly higher concentration of IFN-gamma secreted in culture media of spleen lymphocytes were observed in groups supplemented with TI immunogen (P < 0.01). TI immunogen was an effective stimulator for humoral and cellular immunity and could help improve the immunogenicity of inactivated vaccine or protein subunit vaccine.

Regulating effects of Wnt signaling pathway on differentiation of bone marrow mesenchymal stem cells into osteoblasts and adipocytes / 中国组织工程研究

BACKGROUND:Disequilibrium of proportion of adipogenesis and osteogenesis from bone marrow mesenchymal stem cells (BMSCs)is associated with many bone diseases.However,it has been demonstrated that Wnt signaling pathway could play an important role in regulation of BMSC differentiation.OBJECTIVE:To investigate the different gene expression profiles and to find the target gene on Wnt signaling pathway of the BMSCs,after being induced to osteoblasts and adipocytes respectively using Wnt signaling pathway PCR array.METHODS:The third-passage BMSCs,after being induced to osteoblasts and adipocytes respectively for 7 days.The total mRNA of MSCs was extracted by Trizol.BMSC morphology was observed following osteogenic and adipogenic induction under an inverted microscope.Gene array was detected by rat Wnt signaling pathway PCR array.Non-induction group served as controls.The ratio of increase/reduction gene of osteoblasts and adipocytes was calculated.RESULTS AND CONCLUSION:Under an inverted microscope,BMSCs with high homogenicity were obtained following passage 3.BMSCs differentiated into osteoblasts following osteogenic induction,and into adipocytes following adipogenic induction.Compared with non-induction group,fifteen genes(Dkk1,kremen,FZD1,FZD7,et al.)were expressed up-regulated(ratio ＞ 2)and 16(sFrp 5,β-catenin,Dvl3,Tcf7,et al.)genes down-regulated(ratio ＜ 0.5)when the third-passage BMSCs were induced to adipocytes.Six genes(Dkk1,kremen,β-catenin,Wnt11,et al.)were expressed up-regulated and 15 genes(sFrp5,sFRP4,Fzd1,et al.)down-regulated when BMSCs being induced to osteoblasts.Above-mentioned results suggested that Wnt signaling pathway plays an important role in the osteoblast and adipocyte differentiation from BMSCs.

Research advance in the effect of potassium channel on pathogenesis of idiopathic optic neuritis / 眼科研究

Idiopathic optic neuritis (ION) is a common disorder in neuro-ophthalmology. It harms vision function seriously. However, there are a lots of controversy and confusion about its etiology, etiopathogenesis and outcome of treatments. Recently, potassium channel, such as stichodactyla helianthus peptide (ShK) and TRAM-34, was found to be involved in the immunopathogenesis of ION, and regulation of potassium channel provide a novel immunomodulatory therapy for ION. This paper reviewed the research advance of potassium channel in ION. It is expected to further clarify the pathogenesis and search for effective treatment.

Application of biomaterials to aqueous drainaging devices combined with non-penetrating trabecular surgery / 中国组织工程研究

The implantation of aqueous drainaging devices combined with non-penetrating trabecular surgery(NPTS) has been an effective way to treat open-angle glaucoma.Studies concerning characteristics and outcome of various implants have obtained great progresses,and a large number of implants have been used in clinic.This study was designed to summarize present progress of aqueous drainaging devices combined with NPTS by analyzing aqueous drainaging devices combined with NPTS.

Optimization of the Forming Technology of Zuojin Dropping Pill by Orthogonal Experiment / 中国药房

OBJECTIVE:To optimize the forming technology for Zuojin dropping pill.METHODS:The base material and cooling agent were optimized by orthogonal experiment with the type of base material,the ratio of drug to base material,the temperature of physic liquor,the type of the cooling agent as factors taking the mixing state,forming condition and dissolution time as indexes; the dripping condition was optimized with the temperature of the cooling agent,the caliber of the dripping mouth,the dripping distance and dripping speed as factors taking the variation coefficient of the pill weight difference as index. RESULTS:The optimal forming technology was as follows:with polyglycol 4000 as base and paraffin oil as cooling agent; the cooling temperature was 12 ℃;the caliber of the dripping mouth was 3.5/4.9 mm;the dripping distance was 10 cm and the dripping speed was 25 drop?min-1.CONCLUSION:The forming technology is stable,reproducible and the preparation was well-shaped,and the forming technology serves as a reference for the production of Zuojin dropping pill.

Extraction of the Volatile Oil from Huoxiang Zhengqi Prescription Process with Supercritical CO_2 Fluid:Optimization of the Technical Condition by Orthogonal Test / 中国药房

OBJECTIVE:To establish the optimal technical condition for the extraction of volatile oil from the volatile herbs of Huoxiang zhengqi prescription with supercritical CO2 fluid(SFE-CO2).METHODS:The technical condition for the extraction of volatile oil from the volatile herbs of Huoxiang zhengqi prescription were optimized using L9(34)orthogonal design table with extraction pressure,the extraction temperature,the pressure and temperature of separator Ⅰ as factors and with the yield of the volatile oil as index.RESULTS:A highest extraction yield of volatile oil from the volatile herbs of Huoxiang zhengqi prescription was achieved(2.057%)under an optimal technical condition as follows:with extraction pressure at 25 MPa,the extraction temperature at 30 ℃,the pressure for separator Ⅰ at 9 MPa and the temperature of separator Ⅰ at 45 ℃.CONCLUSION:Extraction of volatile oil from the volatile herbs of Huoxiang zhengqi prescription by SFE-CO2 is characterized by high extraction yield,stable,pure operation,safe,and by which the biological activity of the volatile oil can be maintained,thus it is a potential extraction and separation method.

On the Supercritical Fluid Extraction Technology of Olibanum / 中国药房

OBJECTIVE:To study the supercritical CO2 fluid extraction(SFE-CO2)technology of Olibanum.METHODS:The optimum extraction technology was obtained through orthogonal test with the content of acetic octylester determined by GC and taken as index for the evaluation of the technology.RESULTS:The optimal conditions were as follows:pressure 25 MPa,temperature 60℃ and extraction time 3h.CONCLUSION:The process of SFE-CO2 is suitable for Olibanum extraction.

High glucose concentration incseases the MAPK and TGF?_2 expression in cultured P38 human umbilical vein endothelial cells / 基础医学与临床

Objective To examine the effects of glucose on the expression of P38 MAPK and TGF?2 in cultured human umbilical vein endothelial cells.Methods Human umbilical vein endothelial cells(HUVECs)were incubated in a culture medium with 11.2 mmol/L,16.8 mmol/L and 33.6 mmol/L glucose concentrations for 24 h,48 h and 72 h respectively.The expression P38 MAPK and TGF?2 was measured by RT-PCR and Western-blot.Results When D-glucose concentrations rose,HUVECs exposed to high glucose concentration(11.2 mmol/L,16.8 mmol/L and 33.6 mmol/L)showd increase in cell expression of P38 MAPK and TGF?2 after 48 h and 72 h exposure as compared with those cultured in medium of low glucose concentration(5.6 mmol/L).ConclusionHigh concentration of glucose can arrest the proliferative response.Even a short-term exposure of endothelial cells(ECs)to high glucose concentration may lead to their activation associated with increased expression of P38 MAPK and TGF?2.High Glucose Concentration on P38 MAPK and TGF?2 expression may participate the pathogenesy of diabetic retinopathy.

Effect of curcumin eye ointment on scar formation in filtering blebs after trabeculectomy in rabbits / 第三军医大学学报

Objective To study the effect of curcumin ointment on scar formation in filtering blebs after trabeculectomy in rabbits. Methods Twenty-seven rabbits( 27 eyes) were randomly divided into control group,and matrix( placebo) group,and curcumin group,9 in each group. Trabeculectomy for right eyes was performed on all rabbits. Eyes in control group were not treated with any drug,eyes in matrix group were smeared with 0. 1 g common eye ointment ( 3 times a day) ,and eyes in curcumin group were smeared with 0. 1 g curcumin eye ointment ( 3 times a day). Then,intraocular pressure ( IOP) ,filtering blebs,inflammatory cells,collagen,fibroblasts and myofibroblasts were detected by immunohistochemical staining. Results No significant difference was observed in IOP among the 3 groups before and 1,3,28 d after operation. However,a significant difference was found in IOP among the 3 groups in 7,14 and 21 d after operation ( P

Compound anisodine and physical therapy treat child amblyopia:report of 300 cases / 第三军医大学学报

Objective To investigate the combination of compound anisodine and physical therapy in the treatment of amblyopia,and its possible mechanism.Methods Totally 300 outpatients with amblyopia(3 to 14 years old) were randomly and equally divided into treatment group and control group.In the treatment group,subcutaneous injection of compound anisodine(2 ml,once per day) to the superficial temporal artery of eye was given for a course of 14 d and followed by another course after 5 days'interval.Cover treatment was carried out at the same time.The control group was only treated with physical therapy.Vision,central retinal artery peak systolic velocity(PSV) and diastolic resistance index(RI) were measured during the 3 months' follow-up.Results The 3 to 6-year-old efficiency was 75.0%,7 to 9-year-old efficiency was 69.6%,and 10 to 14-year-old efficiency was 61.1% ;The difference of efficiency between therapy group and control group was very significant(P0.05).Conclusion Compound anisodine plus physical therapy for amblyopia at different ages and varying degrees are effective and safe.The mechanism may be due to enhanced retinal blood supply.

Reduced glutathione protects retina and optic nerve of rats after optic neuritis / 第三军医大学学报

Objective To investigate the effect of reduced glutathione(rGSH)on the retina and optic nerve in rats with experimental optic neuritis,and the underlying mechanism.Methods Animal model of experimental allergic encephalomyelitis(EAE),which was as used for acute demyelinating optic neuritis were established in 48 female SD rats,and then the rats were randomly divided into EAE group(n=18)and GSH group(n=30).Another 6 female SD rats served as normal control.Rats of GSH group received an intraperitoneal injection of 100 mg?kg-1?d-1 GSH for 14 d from the beginning of immunization,while the rats of EAE group were given saline alternatively.To observe pathological changes in the optic nerve,retinal ganglion cells(RGCs)apoptosis,and measure malondialdehyde(MDA),glutathione peroxidase(GSH-PX),nitric oxide(NO)levels in the retina were observed and measured in the early period,peak period and recovery period(12 to 14 d,16 to 18 d,or 26 to 28 d after the immunization)of the neuritis.Results There were 18 rats(60%)from GSH group having the disease with a symptom score of 1 to 3.Compared with those of EAE group,their optic nerve fibers were in a comparatively good order,and more glial cells and not obvious demyelination were observed in morphology;Significant lesser apoptotic RGCs were observed in GSH group than in EAE group at the early and peak period(P

ffect of basic fibroblast growth factor on rabbit lens epithelial cell proliferation / 眼科新进展

Objective　 To study the effect of basic fibroblast growth factor (bFGF) on proliferating of rabbit lens epithelial cells(RLECs).Methods　The second and three generations of RLECs were exposed to different concentrations of bFGF, and the proliferation characteristics of the cells were measured with methyl thiazolyl tetrazolium (MTT) assay. The ultrastructure of cell were observed with transmission electron microscope (TEM).The changs of cells cycles were observed with flow cytometer (FCM).Results　 After the treatment of bFGF, RLECs showed the marked proliferation , especially in 10μg*L-1 of bFGF . TEM result showed that the cells were more active with bFGF, FCM result showed that the S phase cell obviously increased.Conclusion　bFGF is an important factor that can promote proliferation of RLECs.

Protective Effect of Zingiber officinale Extract on Retinal Ischemia-reperfusion Injury in Rats / 中国药房

OBJECTIVE: To investigate the protective effect of Zingiber officinale extract on retinal ischemia-reperfusion (RIR) injury in rats. METHODS: 45 SD rats were randomly divided into normal control group, model group and Z.officinale extract group (400 mg?kg-1).RIR models were induced by infusing normal saline into anterior chambers for l hour. During five days before modeling,Z.officinale extract group were oral administration of Z.officinale extract two times per day by i.g. and model group distilled water. The medication regime remained after modeling. The activity of SOD and the content of malonyldialdehyde(MDA) and NO in retina were measured at 12 h,24 h,48 h and 72 h after modeling. The retinal ultrastructural changes were detected under the electron microscope. RESULTS: Compared with model group, in high-dose of Z.officinale extract group Z.officinale extract could strengthen the activity of SOD, reduce the levels of MDA and NO(P

Study on Preparation Process for Yegan Drop Pills / 中国药房

OBJECTIVE: To establish an optimized preparation process for Yegan Drop Pills. METHODS: The orthogonal test was applied to the selection of basic substance ( polyglycol) , temperature of cooling agent ( usoline) , and ratio in quantity of the extract to the basic substance. RESULTS: The optimal preparation process is under the condition of using polyglycol 4000, and cooling temperature at 10～ 15℃ . CONCLUSION: The Yegan Drop Pills prepared by using the technique are good in shape, stable in quality and easy to replicate. The technique can provide reference for the manufacture of Yegan Drop Pills.