1.Identification of Mequindox and Its Metabolites by High Performance Liquid Chromatography Combined with Ion Trap-Time of Flight-Mass Spectrometry
Yingchun LIU ; Hongbin SI ; Limin HE ; Huanzhong DING ; Xianhui HUANG ; Jianxin CHEN ; Zhangliu CHEN ; Zhenling ZENG
Chinese Journal of Analytical Chemistry 2010;38(1):82-86
The chemical structures of mequindox related metabolites in chicken plasma had been investigated using high performance liquid chromatography combined with linear ion trap quadrupole(LC-ESI/LTQ) and high performance liquid chromatography combined with ion trap-time of flight-mass spectrometry (LC-ESI/IT-TOF).Samples were separated by Hypersil BDS C_(18) and symmetry Shield columns, respectively, and 0.01% formic acid aqueous(A) and methanol(B) were used as mobile phase with gradient elution.Electros pray ionization mass spectrometric(ESI) source was used and operated in positive ion mode.When chickens were orally administered with mequindox at dosage of 20 mg/kg, blood samples were collected from the brachi al vein.Mequindox and its metabolites were extracted by the mixture of acetonitrile and acetoacetate (3:2, V/V).After solvent evaporated, the residue was dissolved in 30% methanol aqueous and the solution was detected by LC/IT-TOF MS and LC-ESI/LTQ.The molecule weight from LC-ESI/IT-TOF was analyzed by software Shimadzu's Composition and the mass chromatogram from LC-ESI/LTQ was analyzed by software Xcalibur 2.0.7.According to the molecular weight and MS~n data, referring the metabolic reaction rules, five chemical structures of mequindox related metabolites in chicken plasma were identified.Metabolites (M1-M4) were synthesized to verify the structure of metabolites.The metabolites are 3-methyl-2-(1-hydroxy) ethyl-qui-noxaline-N~1,N~4-dioxide(Ml), 3-methyl-2-(1-hydroxy) ethyl-quinoxaline-N~4-oxide(M2), 3-methyl-2-acetyl-quinoxaline-N~4-oxide, 3-methyl-2-acetyl-quinoxaline (M4), 3-hydroxymethyl-2-(1-hydroxy) ethyl-quinoxa-line-N~1,N~4-dioxide (M5).
2.Effect of BML-111 on acute lung injury induced by hemorrhagic shock and resuscitation in rats
Guangzhi WANG ; Hongbin LI ; Si GUO ; Jie GONG ; You SHANG ; Shanglong YAO
Chinese Journal of Anesthesiology 2014;34(1):90-93
Objective To evaluate the effect of lipoxin A4 receptor agonist BML-111 on acute lung injury induced by hemorrhagic shock and resuscitation in rats.Methods Thirty-two healthy male Sprague-Dawley rats,aged 6-8 weeks,weighing 200-250 g,were randomized into 4 groups (n =8 each) using a random number table:sham operation group (S group),hemorrhagic shock/resuscitation group (HSR group),BML-111 group,and BML-111 plus BOC-2 (lipoxin A4 receptor antagonist) group (BOC-2 group).The animals were anesthetized with 2% pentobarbital sodium 80 mg/kg,tracheostomized and mechanically ventilated.Left common carotid artery was cannulated for blood-letting and fluid infusion.Hemorrhagic shock was induced according to the method described by Kochanek et al.MAP was reduced to 35-45 mmHg and maintained at this level for 30 min.The animals were then resuscitated for 30 min with infusion of the blood withdrawn and lactated Ringer' s solution 2 times the volume of blood withdrawn.In BML-111 and BOC-2 groups,BML-111 (1 mg/kg) was injected intraperitoneally at the beginning of resuscitation.In BOC-2 group,BOC-2 (50 μg/kg) was injected intraperitoneally before blood-letting.The rats were sacrificed at 2 h after completion of resuscitation.Bronchoalveolar lavage fluid (BALF) was collected for determination of neutrophil count.Lungs were excised for microscopic examination of the pathological changes and for determination of wet/dry lung weight ratio (W/D ratio),contents of interleukin-1 (IL-1β) and IL-6,and phosphorylation of mitogen-activated protein kinase (MAPK).Results Compared with group S,the neutrophil count in BALF,W/D ratio,contents of IL-1β and IL-6,and phosphorylation of MAPK were significantly increased in HSR group (P < 0.05).The neutrophil count in BALF,W/D ratio,contents of IL-1β and IL-6,and phosphorylation of MAPK were significantly lower in BML-111 group than in HSR group,and higher in BOC-2 group than in BML-111 group (P < 0.05).Conclusion BML-111 can attenuate acute lung injury induced by hemorrhagic shock and resuscitation in rats and inhibition of activation of MAPK pathways and reduction of inflammatory responses in lung tissues are involved in the mechanism.
3.Effect of BML-111 on NF-κB pathway during acute lung injury induced by hemorrhagic shock and resuscitation in rats
Hong QI ; Jie GONG ; Si GUO ; Hongbin LI ; Shiying YUAN ; You SHANG ; Shanglong YAO
Chinese Journal of Anesthesiology 2014;34(7):856-858
Objective To evaluate the effect of BML-111 on NF-κB pathway during acute lung injury induced by hemorrhagic shock and resuscitation in rats.Methods Thirty-two adult male Sprague-Dawley rats,weighing 200-250 g,were randomly divided into 4 groups (n =8 each) using a random number table:sham operation group (group S),hemorrhagic shock and resuscitation group (group HSR),BML-111 group,and BML-111 + BOC-2 (lipoxin A4 receptor antagonist) group (group BOC-2).The animals were anesthetized with intraperitoneal pentobarbital sodium.Hemorrhagic shock was induced by blood letting and maintained for 30 min.The animals were then resuscitated for 30 min by infusion of the shed blood and lactated Ringer's solution.In group BOC-2,BOC-2 (50 μg/kg) was injected intraperitoneally before blood letting.In BML-111 and BOC-2 groups,BML-111 (1 mg/kg) was injected intraperitoneally at the beginning of resuscitation.The rats were sacrificed at 2 h after the end of resuscitation and lungs were removed for determination of pathological changes,myeloperoxidase (MPO) activity,intercellular adhesion molecule-1 (ICAM-1) expression (by immunohistochemistry),tumor necrosis factor-alpha (TNF-α) content (by ELISA),and NF-κB p65 and IκB-α expression (by Western blot).Results Compared with group S,the MPO activity,ICAM-1 expression,and TNF-α content were significantly increased,NF-κB p65 expression was up-regulated,and IκB-α expression was down-regulated in group HSR.Compared with group.HSR,the MPO activity,ICAM-1 expression,and TNF-α content were significantly decreased,NF-κB p65 expression was down-regulated,IκB-α expression was up-regulated,and pathological changes of lung were attenuated in group BML-111.Compared with group BML-111,the MPO activity,ICAM-1 expression,and TNF-α content were significantly increased,NF-κB p65 expression was up-regulated,and lκ:B-α expression was down-regulated,and pathological changes of lung were aggravated in group BOC-2.Conclusion BML-1 11 inhibits activation of NF-κB pathway and inflammatory responses,thus mitigating acute lung injury induced by hemorrhagic shock and resuscitation in rats.
4.Evaluation of ventricular systolic function in patients with rheumatoid arthritis using two-dimensional speckle tracking echocardiography
Xiashan ZHANG ; Guojun ZHAO ; Zhian LI ; Shasha DUAN ; Yilu SHI ; Xiaolei WANG ; Si HA ; Yong WANG ; Lijie BAI ; Hongbin LI
Chinese Journal of Ultrasonography 2016;(1):19-23
Objective To evaluate the left ventricular and right ventricular systolic function by two‐dimensional speckle tracking echocardiography ( 2D‐STE) in patients with rheumatoid arthritis ( RA ) . Methods Fifty five patients with RA and 50 healthy subjects were received echocardiography . 2D‐STE were applied for all the subjects to obtain left ventricular global longitudinal strain ( LVGLS) and right ventricular global longitudinal strain ( RVGLS) .Tricuspid annular plane systolic excursion ( TAPSE) and the change ratio of right ventricular area ( RVACR) were measured by echocardiography . The anti‐cyclic citrullinated peptide antibodies ( anti‐CCP‐II) ,rheumatoid factor ( RF) ,C‐reactive protein ( CRP) and erythrocyte sedimentation rate ( ESR) were detected in both group . The LVGLS and RVGLS in RA group were used to conduce correlation analysis with the level of anti‐CCP‐II ,RF ,CRP , ESR and the duration of disease . Results There was a significant decrease in RVGLS and LVGLS in RA group compared with control group( P <0 .05) . However there was no statistical differenc in TAPSE and RVACR between RA and control group( P > 0 .05) . The anti‐CCP‐II ,RF ,CRP and ESR in RA group increased significantly compared with control group ( P < 0 .001 ) . The result of correlation analysis showed there was no correlation between RVGLS ,LVGLS and anti‐CCP‐II ,RF ,CRP ,ESR in RA group . However ,RVGLS and LVGLS were negatively correlated with the duration of disease . Conclusions LVGLS and RVGLS in RA patients were lower than those in healthy people ,strain decreases with the extension of disease duration ,2D‐STE may be an efficacious assessment to assess left ventricular and right ventricular systolic function in patients w ith RA .
5.Development of a multiplex qRT-PCR assay for detection of African swine fever virus, classical swine fever virus and porcine reproductive and respiratory syndrome virus
Yating CHEN ; Kaichuang SHI ; Huixin LIU ; Yanwen YIN ; Jing ZHAO ; Feng LONG ; Wenjun LU ; Hongbin SI
Journal of Veterinary Science 2021;22(6):e87-
Background:
African swine fever virus (ASFV), classical swine fever virus (CSFV), and porcine reproductive and respiratory syndrome virus (PRRSV) are still prevalent in many regions of China. Co-infections make it difficult to distinguish their clinical symptoms and pathological changes. Therefore, a rapid and specific method is needed for the differential detection of these pathogens.
Objectives:
The aim of this study was to develop a multiplex real-time quantitative reverse transcription polymerase chain reaction (multiplex qRT-PCR) for the simultaneous differential detection of ASFV, CSFV, and PRRSV.
Methods:
Three pairs of primers and TaqMan probes targeting the ASFV p72 gene, CSFV 5′untranslated region, and PRRSV ORF7 gene were designed. After optimizing the reaction conditions, including the annealing temperature, primer concentration, and probe concentration, multiplex qRT-PCR for simultaneous and differential detection of ASFV, CSFV, and PRRSV was developed. Subsequently, 1,143 clinical samples were detected to verify the practicality of the assay.
Results:
The multiplex qRT-PCR assay could specifically and simultaneously detect the ASFV, CSFV, and PRRSV with a detection limit of 1.78 × 10 0 copies for the ASFV, CSFV, and PRRSV, but could not amplify the other major porcine viruses, such as pseudorabies virus, porcine circovirus type 1 (PCV1), PCV2, PCV3, foot-and-mouth disease virus, porcine parvovirus, atypical porcine pestivirus, and Senecavirus A. The assay had good repeatability with coefficients of variation of intra- and inter-assay of less than 1.2%. Finally, the assay was used to detect 1,143 clinical samples to evaluate its practicality in the field. The positive rates of ASFV, CSFV, and PRRSV were 25.63%, 9.36%, and 17.50%, respectively. The co-infection rates of ASFV+CSFV, ASFV+PRRSV, CSFV+PRRSV, and ASFV+CSFV+PRRSV were 2.45%, 2.36%, 1.57%, and 0.17%, respectively.
Conclusions
The multiplex qRT-PCR developed in this study could provide a rapid, sensitive, specific diagnostic tool for the simultaneous and differential detection of ASFV, CSFV, and PRRSV.