Objective:To establish an identical UPLC-MS/MS method for determination of related substances of 15 drugs(bezafibrate,acipimox,gliquidone,glimepiride,glipizide,glibenclamide,enalapril,captopril,irbesartan,amlodipine,felodipine,nimodipine,nitrendipine,nisoldipine,nifedipine).Methods:The chromatography was performed on Waters ACQUITY UPLCHSS T3(2.1 mm × 100 mm,1.8 μm)column,mobile phase was 0.01 mmol·L-1 Ammonium formate solution containing 0.1％formic acid and methanol with gradient elution.The flow rate was 0.3 mL·min-1,and the injection volume was 2 μL.The detection was carried out by electrospray ioniza-tion(ESI).Multiple-reaction monitoring(MRM)was employed with switching electrospray ion source polarity in positive and negative mode.Results:The 21 related substances showed good linearity(r ≥0.998 35)in the tested ranges.The precision(RS D≤3.45％)and stability(RSD≤3.48％)of the method were good for the related sub-stances.The average recoveries of the related substances were in the range of 91.81％-101.84％with relative standard deviations RSD≤4.66％.Conclusion:The method is rapid,simple,accurate and highly sensitive,and can be used as a auxiliary test method and confirmatory method for current drug standards.

Objective:To establish an identical UPLC-MS/MS method for determination of related substances of 15 drugs(bezafibrate,acipimox,gliquidone,glimepiride,glipizide,glibenclamide,enalapril,captopril,irbesartan,amlodipine,felodipine,nimodipine,nitrendipine,nisoldipine,nifedipine).Methods:The chromatography was performed on Waters ACQUITY UPLCHSS T3(2.1 mm × 100 mm,1.8 μm)column,mobile phase was 0.01 mmol·L-1 Ammonium formate solution containing 0.1％formic acid and methanol with gradient elution.The flow rate was 0.3 mL·min-1,and the injection volume was 2 μL.The detection was carried out by electrospray ioniza-tion(ESI).Multiple-reaction monitoring(MRM)was employed with switching electrospray ion source polarity in positive and negative mode.Results:The 21 related substances showed good linearity(r ≥0.998 35)in the tested ranges.The precision(RS D≤3.45％)and stability(RSD≤3.48％)of the method were good for the related sub-stances.The average recoveries of the related substances were in the range of 91.81％-101.84％with relative standard deviations RSD≤4.66％.Conclusion:The method is rapid,simple,accurate and highly sensitive,and can be used as a auxiliary test method and confirmatory method for current drug standards.

Guillain-Barré syndrome (GBS) is an autoimmune-mediated peripheral neuropathy. The main therapeutic strategies for GBS are intravenous immunoglobulin and plasma exchange; however, a part of patients respond not well to current strategies. The definite mechanism of GBS remains not clear and animal models are powerful tools to investigate the pathogenesis of GBS and explore new treatments. Till now, a series of animal models have been established. This paper is aimed to review these models with focus on their characteristics, advantages and disadvantages, which will be helpful for the further experimental studies of GBS.

Objective: To introduce the method of galvanic vestibular stimulation-vestibular evoked myogenic potentials (GVS-VEMP) as well as to observe and analyze the parameters and elicited rate of GVS-cVEMP and GVS-oVEMP in healthy young people in China. Methods: Twenty six normal young subjects were recruited for conventional examinations of GVS-VEMP. The subjects were 21-37 years old, average age was (25.8±3.7) years old, including 13 males and 13 females. The galvanic stimulation intensity of 3 mA/1 ms was used to evoke cVEMP and oVEMP on the sternocleidomastoid and inferior extraocular muscles respectively, and the intensity of stimulus was decreased until the response disappeared, the threshold, latency, amplitude, interval phase and interaural amplitude ratio(IAR) were calculated. SPSS18.0 software was used for statistical analysis. Results: All subjects were elicited normal GVS-cVEMP and GVS-oVEMP under 3 mA/1 ms, the elicited rate was 100%. The threshold of GVS-cVEMP was (1.18±0.47) mA, p1 latency was (10.43±1.54) ms, n1 latency was (17.91±1.20) ms, the amplitude was (102.47±56.77) uV and IAR was (0.26±0.20). The threshold of GVS-oVEMP was (1.12±0.50) mA, n1 latency was (8.46±1.05) ms, p1 latency was (11.83±1.27) ms, the amplitude was (9.12±6.82) uV and IAR was (0.25±0.20). In terms of gender and lateral comparison, only the GVS-oVEMP amplitude was higher for male than for female, which had significant statistical difference (P<0.05), and there was no statistical difference in the other parameters between GVS-cVEMP and GVS-oVEMP. Conclusion GVS-cVEMP and GVS-oVEMP could be elicited in healthy youth population, and the parameters could provide reference for subsequent vestibular function evaluation.

Objective: To observe the parameters of the results of suppression head impulse paradigm (SHIMP) in healthy adults, and to provide reference for evaluating vestibular oculomotor reflex function in patients with peripheral vertigo. Methods: Fifty healthy adults, 22 males and 28 females, aged from 23-65 years, with an average age of (38.5±11.6) years, were recruited from January to March 2018. Parameters provided by the video head pulse software included the gains, the latency and the peak velocity of saccades, and comparison was made with head impulse paradigm (HIMP). Results: All subjects were elicited anti-compensatory saccades in SHIMP. The normal values of left and right gains were 1.02 and 1.10 in HIMP, and 0.93 and 1.01 in SHIMP respectively. The left and right saccades latency were (201.1± 50.8)ms and (187.0± 42.9)ms, and the peak saccadic velocity were (302.7±58.5)°/s and (291.5±46.5)°/s in SHIMP; there were small but significant difference between two sides about gains in HIMP and SHIMP, as well as latency in SHIMP(P<0.05); there were small but significant difference between HIMP and SHIMP about gains in ipsilateral(P<0.01); there were no significant difference between two sides about peak saccadic velocity in SHIMP(P>0.05). Conclusions SHIMP can be used for the examination of vestibular oculomotor reflex function in adult population. It is easy to be operated and is convenient for clinical application. Combined with head pulse test, the function of the semicircular canal can be evaluated together.

Objective: To estabilsh animal methods of bone-conducted vibration elicited cervical vestibular-evoked myogenic potentials (BCV-cVEMP) and ocular vestibular-evoked myogenic potentials (BCV-oVEMP) in healthy guinea pigs. Methods: Eleven healthy (250-350 g) and awake guinea pigs were selected and undertake conventional BCV-cVEMP and BCV-oVEMP examination in prone position. Parameters of waveforms were cauculated. Results: The BCV-cVEMP and BCV-oVEMP both could be elicited in 100% (22/22) in guinea pigs respectively, threshold was (85.5±10.8)dB SPL and (90.7±10.6)dB SPL for cVEMP and oVEMP; n1 latency was (4.5±1.3)ms and (4.3±1.5)ms for cVEMP and oVEMP; p1 latency was (5.8±1.4)ms and (5.6±1.7)ms respectively; n1-p1 interwave latency was (1.2±0.4)ms for cVEMP and (1.4±0.6)ms for oVEMP, amplitude was (21.5±17.3)μV and (24.0±16.3)μV respectively. Conclusion Both BCV-cVEMP and BCV-oVEMP can be successfully elicited in healthy guinea pigs.

Objective: To observe the features of air-conducted sound elicited ocular vestibular-evoked myogenic potential(ACS-oVEMP) and cervical vestibular-evoked myogenic potential(ACS-cVEMP) in patients with Meniere disease (MD). To analyze the relationship between air-conducted sound elicited vestibular-evoked myogenic potentials (ACS-VEMP) responses and clinical stages of disease, as well as its clinical application of cervical and ocular vestibular evoked myogenic potentials in MD. Method: Fifty six patients with MD and 50 normal subjects (100 ears) were recruited for conventional cVEMP and oVEMP examinations. Grades of vestibular function were also collected for patients with MD. The relationship between VEMPs abnormity, grades of vestibular function and clinical stages of MD were analyzed. Results: The abnormal rates of cVEMP and oVEMP in MD patients were 57.1% (32/56) and 64.3% (36/56), which were significantly higher than those in normal subjects respectively (χ²=22.286, P=0.000; χ²=15.217, P=0.000). The abnormal rates of cVEMP and oVEMP in MD patients of stage Ⅰ to stage Ⅳ were 20.0% (1/5) and 40.0% (2/5), 50.0% (9/18) and 50.0% (9/18), 59.3% (16/27) and 70.4% (19/27), and 100.0% (6/6) and 100.0% (6/6) respectively. There was a significant difference in cVEMP abnormity between four stages of MD patients (P=0.046). Significant correlation was found between clinical stages and the grades of vestibular dysfunction (r_s=0.417, P=0.001). Conclusions Dysfunction of vestibular otolithic organs and their input pathways in patients with MD can be detected by cVEMP and oVEMP tests. The abnormal rates of VEMP could show an gradually increasing trend with the development of MD stages. And the extent of vestibular lesions could be detected by cVEMP and oVEMP tests, which may provide a reference for clinical staging of MD.

Objective To investigate the functions of triple point-mutants of hypoxia-inducible factor 1α(HIF1α)in angiogenesis in bone defect region under normoxic conditions.Methods Triple point-mutations (the 402,564 and 803 amino acids)in HIF1α coding sequence (CDS)were induced.The triple mutant of HIF1α(402/564/803)was inserted into the adenovirus pAdEasy-1 system to complete viral packaging and titer measurements. The wild-type HIF1α gene and the empty adenovirus vector were packaged in the same way.For the in vitro experiment,the rabbit bone marrow mesenchymal stem cells (MSCs)were divided into four experimental groups:A,MSCs infected with viral solution containing mutant HIF1α;B,MSCs infected with viral solution containing wild-type HIF1α;C,MSCs infected with viral solution without any HIF1α;and D,MSCs without viral infection. The efficiency of infection was observed by the expression of human renilla reniformis green fluorescent protein (hrGFP).The expression levels of HIF1α mRNA and protein in infected cells in each experimental group were measured.For the in vivo experiment,the MSCs were divided into the same four groups and infected with the virus solutions from each group and cultured under normoxic conditions.At 72h after the infection,the MSCs were used as seed cells and transplanted into Apatite-wollastonite magnetic bioactive glass-ceramic (AW MGC)vector to construct artificial tissue-engineering scaffolds,and then the scaffolds were implanted into the in vivo rabbit radial bone defect model.The animals from each group were sacrificed 8 weeks after the surgery and the tissues from the implantation region were harvested for evaluation of angiogenesis.Results The 402,564 and 803 amino acids in CDS area were point mutated into alanine;three types of recombinant adenovirus were successfully constructed, packaged and characterized.The expression levels of HIF1αmRNA were significantly higher in Group A and Group B than in Group C and Group D (P <0.05 ).There was no significant difference between Group A and Group B group or between Group C and Group D (P >0.05 ).The HIF1α protein expression in Group A was significantly higher than that in the other three groups (P <0.05);however,there was no significant difference among groups B,C and D (P >0.05).There was prominent angiogenesis in bone defect region in Group A,but not in the other three groups.Conclusion ① Triple point-mutants of HIF1αefficiently expressed functional proteins under normoxic conditions.② Triple point-mutants of HIF1αeffectively promoted in vivo angiogenesis in bone defect region.

BACKGROUND:Lipopexia induced by glucocorticoid is fat precipitation in marrow due to abnormal lipomatabolism,or differentiation of cells resulting from hormone-affected bone marrow mesenchymal cells.The precise generating procedure remains unclear.OBJECTIVE:To Jnvastigate effects of vadous concentrations of dexamethasone on adipogenic differentiation of bone marrow mesenchymal cells.DESIGN,TIME AND SETTING:The observational study was performed at the Department of Biochemistry,China Medical University from January 2007 to January 2009.MATERIALS:A total of 80 Sprague-Dawley rats aged 3 or 4 weeks,weighing (110±10) g,of both genders,were used in this study.METHODS:Rat bone marrow mesenchymal cells were isolated and subcultured.Bone marrow masenchymal cells at the third passage were used as samples.MAIN OUTCOME MEASURES:Cellular morphologic changes after treatment of 10~(-7) mol/L dexamethasone were observed under an inverted microscope,as well as at 3,7,14 and 21 days under normal culture condition.Morphological changes of bone marrow mesenchymal cells were observed following alkaline phosphatase and Sudan Ⅲ staining.Alkaline phosphatase activity was measured using phenol reagent.Effects of dexamethasone on cell proliferation were measured using MTT assay.RESULTS:Following 24 hours of incubation,a few adherent cells were found in the bottom of the culture flask,showing spindle shape.With prolonged time,adherent cells became more,presenting radiated shape.Following passage,cells distributed uniformly,showing typical fibroblast-shape.Following dexamethasone stimulation,cells changed from spindle-shape into polygonal or irregular shape.In the late phase,with increased concentration and prolonged time,cell colonies disappeared;cells adhered,and died.In normal cultured cells,no or a few orange particles were found.In dexamethasone-cultured cells,with increased hormone concentration and prolonged time,Sundan Ⅲ stained particles increased.At 21 days,alkaline phosphatase activity under normal culture was separately 1.57-,4.49-and 5.0-fold of 10~(-8),10~(-7),10~(-6) mol/L dexamethasone group.At 7,14 and 21 days,alkaline phosphatase activity under normal culture was separately 2.93-,3.80-and 4.39-fold of 10~(-7) mol/L dexamethasone group (P ＜ 0.05).High concentration of dexamethasone had significant inhibitory effects on cell proliferation activity.Significant difference was detected as compared 10~(-7) mol/L and 10~(-6) mol/L to other groups (P ＜ 0.05).CONCLUSION:High-dose dexamethasone enhances adipogenic and suppresses osteoblastic differentiation of bone marrow mesenchymal cells.The effect will increase along with the increasing content and prolonged duration of demamethasone stimulation.

AIM: To develop a simple and sensitive high-performance liquid chromatography (HPLC) for the quantification of zidovudine and to study the pharmacokinetics of two kinds of zidovudine capsules in Chinese healthy volunteers. METHODS :The concentrations of zidovudine in plasma were determined by a validated HPLC method with UV detection. A randomized two-way crossover study was conducted in 18 fasting volunteers to compare plasma pharmacokinetic profile and single-dose tolerability of a new zidovudine capsules. RESULTS: The main pharmacokinetic parameters of two formulations, reference and test capsules, were as follows: cmax were (2 252±s 837) μg·L-1 and (2 300±1 099) μg·L-1; tmax were (0.49±0.19) h and (0.5±0.3) h;t1/2 ke were (0.93±0.19) h and (0.99±0.24) h; AUC0-t were (2 530±452) μg·h·L-1 and (2 467±605) μg·h·L-1;AUC0-∞ were(2 689 ± 414) μg·h·L-1 and (2 583±575) μg·h·L-1. The results of ANOVA and two one-side t test statistical analysis for lg AUC0-t, lg AUC0-∞ and lg cmax showed that two formulations were bioequivalent. CONCLUSION:The method is convenient, sensitive, accurate and reproducible, and could be applied to determining the plasma zidovudine concentration and studying on pharmacokinetics. Two zidovudine capsules are bioequivalent in Chinese healthy volunteers.