1.β-Elemene improves endothelial cells dysfunction, and abnormal proliferation and migration of vascular smooth muscle cells
Lanlan DUAN ; Jing DONG ; Xiangcheng FAN ; Junyi ZHU ; Yifan ZHANG ; Jichun HAN ; Jing SHANG
Journal of China Pharmaceutical University 2020;51(3):333-339
This study aimed to investigate whether β-elemene could improve the dysfunction of vascular endothelial cells induced by low shear force (LSS), and the proliferation and migration of vascular smooth muscle cells induced by oxidized low-density lipoprotein (ox-LDL). Parallel plate flow chambers and ox-LDL were used to establish vascular endothelial cells (ECs) dysfunction model and vascular smooth muscle cell (VSMCs) proliferation and migration model, respectively, and the effects of β-elemene on ECs dysfunction and VSMCs proliferation and migration were examined. The activity of ROS in ECs was measured by DHE and the activity of NO in ECs was tested by DAF-FM DA. The protein phosphorylation of Akt and ERK in ECs were detected by Western blot. The proliferation of VSMCs was measured by MTT. The migration of VSMCs was examined by cell scratch test and Transwell assay. The gene expression of MMP-2 and MMP-9 in VSMCs was measured by RT-qPCR. In ECs, β-elemene could significantly reduce the LSS-induced increase in ROS, significantly increase the LSS-induced decrease in NO, decrease the phosphorylation of ERK, and increase the phosphorylation of Akt. In VSMCs, β-elemene could significantly reduce the proliferation and migration of VSMCs induced by ox-LDL, and reduce the gene expression of MMP-2 and MMP-9. To conclude, β-elemene can improve the LSS-induced ECs dysfunction and ox-LDL-induced VSMCs proliferation and migration.
2.Drug resistance induction and analysis of differential expression protein on adult Schistosoma j aponicum induced by ED50 PZQ
Lanlan DONG ; Jing XU ; Bo ZHAO ; Song LIANG ; Yanyan WANG ; Zhixun GUAN ; Yun CAO ; Chaoming XIA
Chinese Journal of Zoonoses 2014;(12):1171-1180
ABSTRACT:In the present study ,we aimed to identify differentially expressed proteins between induced worms (the infec‐ted mice were treated intragastrically with ED50 PZQ) and uninduced worms (control group) for clarifying the mechanism of PZQ .ED50 PZQ was used to administrate mice that were infected with S .japonicum via intragastric incubation for consecutive‐ly 30 days .Twenty‐one days later ,mice were sacrificed after treatment with 200 mg/kg PZQ for continuously five days ,and the male worms were obtained and some of them were subjected in DMEM medium with different concentrations of PZQ in vitro for 16 hours .Then the worms were washed twice and incubated in PZQ‐free medium for 72 hours .Compared with control group ,the induced worms had lesser sensitivity to PZQ .The survival rate of induced worms was 75 .6% in vitro when the con‐centration of PZQ was 112 mol/L (the concentration was 8 times of uninduced worms Lethal Concentration ) ,significantly higher than that in the uninduced worms (11 .1% ,P<0 .05) ,showing obviously tolerance .The other induced and uninduced worms were acquired and collected for 2D‐DIGE and MALDI‐TOF‐MS ,and combined with bioinformatics to analyse the func‐tion of the identified protein .Thirty differential expression proteins were confirmed between induced and uninduced worms ,in‐cluding 12 proteins up‐regulated and 18 proteins down‐regulated .These proteins respectively ascribed to cytoskeleton‐associat‐ed protein ,glucose and energy metabolism enzymes ,stress proteins ,thioredoxin peroxidase enzymes ,and other protease .Up‐or down‐regulation of these differential proteins indicated that PZQ promote or inhibit the expression of some specific genes . These findings may help to clarify the mechanism of PZQ ,simultaneously ,providing a scientific basis for exploring new vaccine candidate antigens and targets for drug therapy .
3.Interferon Gamma Release Assays from T Lymphocytes in Patients with Tuberculosis Infection
Peili ZHANG ; Yiqing LIU ; Jing SHAO ; Lanlan CHEN ; Dengran NIU ; Wenbing DUAN ; Dong ZHANG
Journal of Modern Laboratory Medicine 2017;32(1):22-25,29
Objective To explore the application value of interferon gamma release assay (IGRAs)in the clinical detection of tuberculosis infected T lymphocytes.Methods Used IGRAs method to detect the 11 968 outpatients and hospitalized pa-tients from 2013 to 2016 with tuberculosis screening.According to the distribution department analysis,also of positive case detection according to age and gender were analysis and comparison and analysis on the uncertainty of results,different methods were compared.Results Among the 11 968 cases,2 048 cases were positive,the positive rate was 17.11%,and the uncertain result was 107 cases,which accounted for 0.89% of the total number.The positive rates from 2013 to 2016 were 19.65%,21.35%,15.82% and 13.56%,respectively.In the detection and screening of pulmonary and pulmonary tuberculo-sis,the positive rates of the department of respiration,the digestive department,the oncology department,the department of neurology and the department of gynecology were 22.07%,20.27%,23.38%,12.84% and 11.86%,respectively.In the positive screening,men accounted for 62.11%,women accounted for 37.89%,men were significantly higher than women.By age group,was less than or equal to 15,16~25,26~45,46~65,was more than orequal to 66 years old,positive rate were 1.96%,18.51%,16.54%,21.25% and 25.73%,respectively.Analysis of uncertain outcome data,department of respira-tion,rheumatism,department of hematology,accounted for 1.99% and 2.35%,respectively.Compared with other laboratory methods,the IGRAs method had obvious advantages.Conclusion Tuberculosis occurs in various body organs,there were differences in gender and age of Mycobacterium tuberculosis infection.IGRAs is a sensitive and specific method for rapid de-tection of Mycobacterium tuberculosis infection,although it can not be used as a diagnostic indicator,but in patients with suspected tuberculosis IGRAs has a larger clinical application value for the further diagnosis of disease.
4.Simultaneous Contents Determination of Chloramphenicol and Diphenhydramine Hydrochloride in Chloro-benzene Alcohol Solution by HPLC
Shan LIU ; Linlan JIANG ; Xinmin XIE ; Ming YAN ; Dong LI ; Tengxia LI ; Hui LI ; Lanlan LIN ; Liping HAN
China Pharmacy 2016;27(27):3847-3849
OBJECTIVE:To establish a method for the simultaneous contents determination of chloramphenicol and diphenhydr-amine hydrochloride in Chlorobenzene alcohol solution. METHODS:HPLC was performed on the column of Diamonsil C with mo-bile phase of phosphate buffer-acetonitrile(66.5∶33.5,V/V)containing 0.01 mol/L sodium heptane,column temperature was 35 ℃, flow rate was 1 ml/min,detection wavelength was 258 nm for diphenhydramine hydrochloride and 277 nm for chloramphenicol, the injection volume was 10μl. RESULTS:The linear range was 34.8-139.4μg/ml for chloramphenicol(r=0.999 8)and 93.0-930.5μg/ml for diphenhydramine hydrochloride (r=0.999 8);RSDs of precision,stability and reproducibility tests were lower than 2.0%;recoveries were 97.70%-104.02%(RSD=2.2%,n=9)and 96.72%-101.72%(RSD=1.6%,n=9). CONCLUSIONS:The method is specific,accurate with good precision,and suitable for the quality control of Chlorobenzene alcohol solution.
5.MR signal and spectroscopy analysis of lumbar spine bone marrow in Tibetan
Yousen WU ; Haihua BAO ; Hongqian ZHANG ; Weixia LI ; Lanlan DONG ; Xiaofei YANG
Journal of Practical Radiology 2017;33(12):1910-1912,1916
Objective To investigate the MR signal intensity and spectroscopy characteristics of lumbar bone marrow in normal adult of Tibetan and Han nationality in high altitude.Methods According to the inclusion criteria,lumbar MRI examinations in Tibetan and Chinese volunteers(each 50 cases)were obtained.For each inspector,the lumbar 3 vertebra was selected,TFE sequence was used to measure the signal strength,T2map was used to measured the T2time,and 1H-MRS was used to measure the lumbar bone marrow spectrum signal.The measured data were statistically analyzed.Results Compared with the Han nationality in Xining area,T1-TFE sequence of Tibetan showed obvious low signal in the lumbar vertebra.The relative signal intensity were significantly different(P=0.001).For measurement of T2time,there was no significant differences(P=0.061).The spectrum analysis showed a line of low fat high water for Tibetan,and a line of low water high fat for Han nationality in Xining area.There were significant differences for water peak intensity,Width, Height and peak Area,but no significant differences in Lipid peak.There were significant differences(P<0.05)on Lipid water absorption ratio, fat water ratio,fat fraction between the two groups.Conclusion The Tibetan shows a low signal on T1WI sequence of lumbar spine which is considered in hypoxia condition for a long time,and the bone marrow red pulp associated with water content increased sig-nificantly.
6.Effect of down-regulation of lncRNA LINC00263 targeting miR-4458 on regulating radiosensitivity of breast cancer SK-BR-3 cells
Lanlan WEN ; Dongjuan WANG ; Hui DONG ; Jiwei ZHAO ; Cuimin ZHU ; Pingping LIN ; Lanfang LIU ; Qingshan LI
Chinese Journal of Radiation Oncology 2021;30(11):1195-1201
Objective:To evaluate the effect of down-regulating lncRNA LINC00263 targeting miR-4458 on the proliferation, migration, invasion and radiosensitivity of breast cancer SK-BR-3 cells.Methods:The expression differences of LINC00263 in breast cancer tissues, adjacent tissues, normal breast epithelial cells and breast cancer cells were determined by qRT-PCR. Transfection of LINC00263 shRNA in breast cancer SK-BR-3 cells down-regulated the expression of LINC00263, and the cloning experiment was used to detect the radiosensitivity. Breast cancer SK-BR-3 cells were treated with 6 Gy irradiation. CCK-8 assay was employed to detect cell proliferation. Flow cytometry was adopted to detect cell apoptosis. Transwell chamber test was performed to detect cell migration and invasion. Western blot was used to detect the expression levels of C-Caspase-3 and C-Caspase-9, MMP-2 and MMP-9 proteins. Bioinformatics software predicted that LINC00263 and miR-4458 had complementary binding sites, and the luciferase reporter system was utilized determine the targeting relationship between LINC00263 and miR-4458. LINC00263 shRNA and miR-4458 inhibitor were co-transfected into breast cancer SK-BR-3 cells, and 6 Gy irradiation was given to detect the changes in cell proliferation, apoptosis, invasion and migration.Results:The expression level of LINC00263 in breast cancer tissues was higher than that in adjacent tissues. The expression level of LINC00263 in breast cancer cells was higher compared with that in normal breast epithelial cells. The radiosensitivity of breast cancer SK-BR-3 cells was increased after transfection of LINC00263 shRNA. Transfection of LINC00263 shRNA and radiation exerted a synergistic effect, jointly inhibited breast cancer cell proliferation, migration and invasion, promoted cell apoptosis, up-regulated the expression levels of C-Caspase-3 and C-Caspase-9 proteins in cells, and down-regulated those of MMP-2 and MMP-9 proteins. Down-regulation of LINC00263 targetedly up-regulated miR-4458 expression. miR-4458 inhibitor reversed the inhibitory effect of LINC00263 shRNA combined with radiation on the proliferation, migration, invasion and apoptosis promotion of breast cancer SK-BR-3 cells.Conclusion:Down-regulating lncRNA LINC00263 targeting miR-4458 inhibits the proliferation, migration and invasion of breast cancer SK-BR-3 cells, and improves cell radiosensitivity.
7.Deletion of D8L region reducing the immunogenicity of recombinant vaccinia virus vector
Ziling ZHANG ; Kangli CAO ; Shimeng BAI ; Lanlan DONG ; Tianhan YANG ; Chen ZHAO ; Jianqing XU ; Xiaoyan ZHANG
Chinese Journal of Microbiology and Immunology 2023;43(11):836-842
Objective:To reduce the immunogenicity of vaccinia virus vector by replacing the D8L region, which is a neutralizing antibody epitope in vaccinia virus, with an exogenous gene.Methods:A gene fragment encoding influenza virus hemagglutinin (HA) was inserted into the D8L region to replace it using homologous recombination technique. Then, a recombinant vaccinia virus influenza vaccine was constricted. A recombinant vaccinia virus vaccine with the TK region expressing HA was used as a control. The expression of HA was validated by Western blot. BALB/c mice were immunized with the vaccines and the serum antibody titers two weeks after each immunization were evaluated by ELISA and hemagglutination inhibition assay. The protective efficacy of the recombinant vaccinia virus was assessed through a challenge experiment.Results:Western blot confirmed the successful expression of HAD8L protein in the constructed recombinant vaccines. ELISA and hemagglutination inhibition assay showed that after the primary immunization, the anti-HA antibody titer induced by the recombinant vaccinia virus with D8L region mutation was slightly higher than that induced by the vaccine with TK region mutation, and the difference was statistically significant with the increase of immunization times ( P<0.05). The recombinant vaccinia virus with D8L region mutation showed significantly lower immunogenicity than the recombinant virus with TK region mutation after the primary immunization, but there was no significant difference between them with the increase of immunization times ( P>0.05). After H1N1pdm challenge, no virus was detected in the mice immunized with the recombinant vaccinia virus with D8L region mutation and the mice showed mild lung inflammation and less tissue damage. Conclusions:This study indicated that inserting exogenous genes into the D8L region of the neutralizing antibody epitope in the vaccinia virus vector could help to reduce the immunogenicity of the vector itself and enhance the immunogenicity of the exogenous genes. This provided a reference for the use of the vaccinia virus vector as a delivery tool in the field of vaccines or gene therapy.
8.Remote programming after vagus nerve stimulation in refractory epilepsy
Yinbao QI ; Dong ZHANG ; Lanlan WANG ; Xianming FU ; Ruobing QIAN
Chinese Journal of Neuromedicine 2021;20(9):932-935
Objective:To explore the efficacy of remote programming after vagus nerve stimulation (VNS) in patients with refractory epilepsy.Methods:Thirty-four patients who received VNS in our hospital from October 2019 to October 2020 were chosen in our study. Among them, 19 patients accepted remote programming (remote programming group) and 15 patients regularly came to the outpatient clinic for regulation (outpatient regulation group). The seizure frequency, response rate (seizure frequency decreased by≥50%), McHugh grading, and incidence of postoperative complications between the 2 groups were compared 6 months after VNS.Results:The seizure frequency in the remote programming group and outpatient regulation group was 2 (0, 4) times/month and 4(1, 24) times/month, without significant difference ( Z=-1.602, P=0.105). The proportion of patients enjoying effective treatment in the two groups was 13/19 and 8/15, without significant difference ( P=0.781). Results of McHugh grading showed no significant difference between the two groups ( Z=-0.728, P=0.467). The proportion of patients with postoperative complications in the two groups was 3/19 and 2/15, without significant difference ( P=0.625). Conclusion:The remote programming after VNS is safe and effective, which can become an important complementary approach for outpatient regulation.
9.Performance verification of LIAISON chemiluminescence immunity analyzer
Lanlan CHEN ; Jing SHAO ; Yiqing LIU ; Peili ZHANG ; Dengran NIU ; Wenbing DUAN ; Wanhui ZHAO ; Dong ZHANG
International Journal of Laboratory Medicine 2018;39(2):149-152
Objective To verify the performance of LIAISON chemiluminescence immunoassay analyzer in the prenatal screening for TORCH .Methods Reference to the US Institute of Clinical and Laboratory Stand-ards(NCCLS) series of documents and literature and combining with actual work ,we designed the verification program ,and tested and evaluated the LIAISON chemiluminescent immunoassay systems for the measurement precision ,accuracy ,linearity analysis ,clinical reportable range and biological reference intervals of Tox IgG , Tox IgM ,Rub IgG ,Rub IgM ,CMV IgG ,CMV IgM ,HSV IgG ,HSV IgM .We also compared the results with analysis performance provided by manufacturers (Italy LIAISON ) or recognized quality indicators .Results Intra-assay imprecision CV values were between 3 .58% -7 .03% ,which were less than the predetermined range;inter-assay imprecision CV values were between 3 .13% -10 .73% .Linear range validation regression coefficients a values were between 0 .97 -1 .03 and r2 >0 .95 .The linear relationship met the requirements . Both biological reference interval and reportable range meet the requirements .Conclusion The performance of LIAISON chemiluminescence immunoassay detection system satisfied the clinical requirements ,and the meas-urement results had advantages of high sensitivity ,specificity ,stability ,wide detection range ,good accuracy and repeatability ,which was suitable for clinical application .
10.Establishment of mouse models of lymphoma with dual-labeled EBV-immortalized lymphoblastoid cell lines by intravenous versus subcutaneous injection
Lanlan FANG ; Ting DONG ; Ying ZHOU ; Yulu SUN ; Yang GAO ; Yunqing XIONG ; Chaojiang GU
Chinese Journal of Clinical Oncology 2023;50(24):1243-1247
Objective:To establish a green fluorescent protein(GFP)and firefly luciferase(Luc)double-labeled Epstein-Barr virus(EBV)infec-ted B lymphoblastoid cell lines(B-LCL)and apply them to mouse models,then compare the advantages and disadvantages of models inocu-lated by intravenous(IV)or subcutaneous(SC).Methods:B lymphoblastoid cell lines double-tagged with GFP/Luc(B-LCL-GL)were con-structed through lentivirus transduction,puromycin intervention.Subcutaneous xenograft and hematogenous metastasis models were re-spectively established by subcutaneous or intravenous injection of B-LCL-GL cells at three concentrations in(NOD)/Prkdcscid/IL-2Rγnull(NPG)mice for in vivo bioluminescence imaging.Results:In the B-LCL-GL group,the ratio of the GFP-positive cell population was 92.5%,and the average luminescence intensity was as high as 4.80E+08 Photons/s,which was considerably higher than that of untreated B-LCLs.In the hematogenous metastasis models,tumor bioluminescence was initially located in the peritoneal area and then spread throughout the en-tire body between 7 and 28 days.In the subcutaneous xenograft models,strong central and weak peripheral tumor-related biolumines-cence signal was detected on day 7 in the three groups,which then spread throughout the body on day 28 in the high-dose group.Taken to-gether,there was no significant difference in tumor progression between the two routes of administration when using the same dose of B-LCL-GL cells.However,the survival analysis indicated that the IV injection group,in which all the mice ultimately died,had a shorter time frame for testing than that of the SC injection group,in which the mice survived until day 100 in the low-dose and medium-dose groups,thus allowing for long-term testing.Conclusions:GFP and Luc dual-positive B-LCLs were successfully established to generate hematogenous metastasis and subcutaneous xenograft models,which allow the monitoring of the location and size of lymphomas in vivo.It provide plat-form for the study of tumor characteristics and selecting anti-tumor drugs.