Table of Content

28 February 2022, Volume 37 Issue 2

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Dynamic changes and clinical significance of CXCR5,sPD-1 and Hcy in AIDS patients during therapy

SHI Penghui, FAN Weiguang, ZHANG Zhen, SU Miaomiao, MENG Juan, LU Xinli

2022, 37(2):  103-107.  DOI: 10.3969/j.issn.1673-8640.2022.02.001

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Objective To investigate the changes and clinical significance of serum C-X-C chemokine receptor 5（CXCR5）,soluble programmed cell death receptor-1（sPD-1） and homocysteine（Hcy） in acquired immune deficiency syndrome（AIDS） patients during antiretroviral therapy（ART）. Methods Totally,120 AIDS patients were enrolled in AIDS group,and 45 healthy subjects were enrolled as control group. The human immunodeficiency virus（HIV）-1 load,the absolute number of CD4⁺T cells,CXCR5,sPD-1,Hcy,blood lipids [triglyceride（TG）,total cholesterol（TC）,low-density lipoprotein cholesterol（LDL-C） and high-density lipoprotein cholesterol（HDL-C）] were determined in AIDS patients on 1 d before therapy and 3,6 and 12 months after therapy,respectively. Pearson correlation analysis was used to evaluate the correlations between each index and the absolute number of CD4⁺ T cells and HIV-1 load. Results Before therapy,the levels of sPD-1 and Hcy in AIDS group were higher than those in control group（P<0.01）,and CXCR5 and the absolute number of CD4⁺T cells were lower than those in control group（P<0.01）. Compared with those before therapy,with the prolongation of ART time,the absolute numbers of CD4⁺ T cells and CXCR5 levels in AIDS patients were increased（P<0.05）. The peak was reached at 6 months after therapy. Hcy,TC,TG and LDL-C were increased after 12 months of therapy（P<0.05）. HDL-C did not change. The HIV-1 load began to decrease after 3 months of therapy（P<0.05）,and the decrease was more significant after 6 months of therapy（P<0.05）. The level of serum sPD-1 decreased after 12 months of therapy（P<0.05）. Pearson correlation analysis showed that the absolute number of CD4⁺ T cells was positively correlated with CXCR5 and Hcy（r=0.192 and 0.401,P<0.05）,and there was a negative correlation with sPD-1（r=-0.708,P<0.05）. The HIV-1 load was negatively correlated with CXCR5 and Hcy（r=-0.682 and -0.318,P<0.05）,and it was positively correlated with sPD-1（r=0.368,P<0.05）. Conclusions The dynamic monitoring of the absolute number of CD4⁺ T cells,HIV-1 load,CXCR5,sPD-1,Hcy and blood lipid changes of AIDS patients during ART can help monitor the progress of HIV infection in time.

Evaluation of the interference of serum index in the determination of prealbumin by immunoscattering turbidimetry according to CLSI EP07-A3 document

HAN Guang, XU Qiongfeng, DUAN Shumin, LUO Jiahuan, WANG Yunxiu, ZHANG Xiujuan, WAN Zemin, KE Peifeng, HUANG Xianzhang

2022, 37(2):  108-111.  DOI: 10.3969/j.issn.1673-8640.2022.02.002

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Objective To investigate the interference effect of hemolysis,icterus and lipemia on the determination of prealbumin（PA） by immunoscattering turbidimetry. Methods A fresh serum sample was used as a basic sample,interfering substances [hemoglobin（Hb）,bilirubin and lipid] were added,and the PA level was determined by immunoscattering turbidimetry. According to the EP07-A3 document of the Clinical and Laboratory Standards Institute（CLSI）,the paired difference experiment and dose-effect experiment were performed. Results PA had no interference with 10 g/L Hb（hemolysis index of 1 000）and 200 mg/L（340 μmol/L） bilirubin（icterus index of 20）. The lipid turbidity interfering substance with a lipid turbidity index of 987 had a negative interference on the determination of PA. The degree of interference had a linear relationship with the lipid turbidity index. For samples with low values of PA,when the lipid turbidity index≤120,there was no interference to PA determination. The linear equation between lipid turbidity index and interference rate was Y=-0.077X+0.979（r ²=0.999）. For samples with high values of PA,when the lipid turbidity index≤203,there was no interference to PA determination. The linear equation between lipid turbidity index and interference rate was Y=-0.040X-0.168（r²=0.992）. Conclusions Generally clinical hemolytic and icteric samples will not interfere with the PA determination results by immunoscattering turbidimetry. The samples with lipid turbidity index>120 will cause negative interference to PA determination results.

Feasibility of indirect method to establish the cut-off value of neonatal G6PD screening in Shanghai

JI Wei, TIAN Guoli, ZHU Zhixing, ZHOU Zhuo, WANG Yanmin, ZHANG Xiaofen

2022, 37(2):  112-116.  DOI: 10.3969/j.issn.1673-8640.2022.02.003

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Objective To investigate the feasibility of establishing the cut-off value of glucose-6-phosphate dehydrogenase（G6PD） screening for newborns in Shanghai by indirect method. Methods The G6PD determination data of 134 747 newborns were collected,and the Kolmogorov-Smirnov test was used to analyze the datum distribution. The skewed distribution data were converted into an approximately normal distribution using Box-Cox. The interquartile range（Turkey） method was used to eliminate outliers and establish a reference interval,with the lower limit of the reference interval [single-side 5th percentile value（P₅）] as the screening cut-off value. The established cut-off value with the lower limit of the reference interval declared by the manufacturer and the cut-off value established by a small sample in the clinical verification phase of the Neonatal Screening Laboratory of Shanghai Children's Hospital were compared,the relative deviation was calculated,and it was compared with the reference change value（RCV）. Results Among 134 747 newborns,233 cases of G6PD deficiency were diagnosed,and the incidence of G6PD deficiency was 1∶579. The indirect method was used to establish the G6PD screening cut-off value in Shanghai,which was 317.8 U/L for male infants and 325.7 U/L for female infants. The relative deviation of this cut-off value from the lower limit of the reference interval declared by the manufacturer and the cut-off value established by the Neonatal Screening Laboratory of Shanghai Children's Hospital based on a small sample was higher than RCV. The G6PD levels of 233 cases of G6PD deficiency were lower than the established screening cut-off value. Based on this screening cut-off value,the coincidence rate of both negative and positive for 70 external quality assessment samples reached 100%. Conclusions Using indirect method to establish the reference interval and the cut-off value of G6PD can improve the screening efficiency of neonatal G6PD deficiency.

Correlation between the expression levels of MMP mRNA and ADAMTS mRNA with the degree of intervertebral disc degeneration

XU Mingyuan, XU Guisen, YANG Xiaokun, LUO Yong, GUAN Sishu, FAN Yunfei

2022, 37(2):  117-121.  DOI: 10.3969/j.issn.1673-8640.2022.02.004

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Objective To investigate the correlation between the expression levels of matrix metalloproteinase（MMP） mRNA and a disintegrin and metalloproteinase with thrombospondin motifs（ADAMTS） mRNA with the degree of intervertebral disc degeneration. Methods A total of 160 disc degenerative disease patients were enrolled,and according to the results of magnetic resonance imaging（MRI） and Pfirrmann classification method,they were classified into Ⅰ level（30 cases）,Ⅱ level（32 cases）,Ⅲ level（35 cases）,Ⅳ level（33 cases） and Ⅴ level（30 cases）. The expressions of MMP-1 mRNA,MMP-3 mRNA,MMP-9 mRNA,MMP-13 mRNA,ADAMTS-1 mRNA,ADAMTS-4 mRNA,ADAMTS-5 mRNA,interleukin（IL）-1α mRNA,IL-1β mRNA,IL-6 mRNA and IL-8 mRNA in the nucleus pulposus were determined. Spearman correlation analysis was used to evaluate the correlation between each indicator and Pfirrmann classification,and Pearson correlation analysis was used to evaluate the correlation between each indicator. Results The relative expressions of MMP-3 mRNA,MMP-9 mRNA,IL-1α mRNA and IL-1β mRNA in Ⅳ and Ⅴ level groups were higher than those in Ⅰ,Ⅱ and Ⅲ level groups（P<0.05）,and those in Ⅰ,Ⅱ and Ⅲ groups were increased in turn（P<0.05）. The relative expressions of ADAMTS-4 mRNA and ADAMTS-5 mRNA were higher in Ⅲ,Ⅳ and Ⅴ level groups than those in Ⅰ and Ⅱ level groups（P<0.05）,and those in Ⅱ level group was higher than those in Ⅰ level group（P<0.05）. There was no statistical significance for the other indicators（P>0.05）. Spearman correlation analysis showed that the relative expressions of MMP-3 mRNA,MMP-9 mRNA,ADAMTS-4 mRNA,ADAMTS-5 mRNA,IL-1α mRNA and IL-1β mRNA were positively correlated with Pfirrmann classification（P<0.05）. Pearson correlation analysis showed that the relative expressions of MMP-3 mRNA,MMP-9 mRNA,ADAMTS-4 mRNA and ADAMTS-5 mRNA were positively correlated with the relative expressions of IL-1α mRNA and IL-1β mRNA（P<0.05）. Conclusions MMP-3,MMP-9,ADAMTS-4,ADAMTS-5,IL-1α and IL-1β are related to the occurrence and development of intervertebral disc degeneration,which can reflect the severity of the disease.

High prolactin caused by risperidone related factors

CHEN Qiuying, ZHONG Yinghua, ZHAO Nan, YIN Haibo, XU Yang, SHU Ming

2022, 37(2):  122-125.  DOI: 10.3969/j.issn.1673-8640.2022.02.005

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Objective To study high prolactin（PRL） caused by risperidone-related factors,and to guide clinicians using drugs rationally. Methods Totally,90 patients with schizophrenia were enrolled after treatment of risperidone for 6 weeks. The general data,including age,sex,body mass index（BMI）,positive and negative syndrome scale（PANSS） score,plasma concentration of risperidone,fasting blood glucose,total cholesterol（TC）,triglyceride（TG） and the level of PRL,were collected. According to plasma concentrations of PRL,the research subjects were classified into high PRL group（PRL≥30 ng/mL） and normal PRL group（PRL<30 ng/mL）. Results The proportion of females in high PRL group was higher than that in normal PRL group（P<0.001）,and PANSS score and fasting blood glucose were lower than those in normal PRL group（P<0.001）. Age,BMI,TC,TG and plasma concentration of risperidone had no statistical significance between the 2 groups（P>0.05）. Logistic regression analysis showed that high PRL was associated with sex（female）,PANSS score and fasting blood glucose [odds ratios（OR） were 15.843,0.466 and 0.053,95% confidence intervals（CI） were 5.329-47.105,0.259-0.838 and 0.014-0.202,P<0.05]. Age,BMI,TC,TG and plasma concentration of risperidone were not risk factors for high PRL（OR=1.016,1.623,0.449,0.650 and 0.399,P>0.05）. PRL in female patients was higher than that in male patients（P<0.001）. Conclusions Female patients taking risperidone are easier to be high PRL,and patients taking risperidone with low PANSS score and low fasting blood glucose are tended to be high PRL. Clinicians should use individualized medication.

Correlation between metabolic syndrome and moderate-to-severe periodontitis complicated with psoriasis

CHEN Liang, LI Lin, LI Xia, ZHANG Yan, WU Jiongrui, GAO Yiming

2022, 37(2):  126-129.  DOI: 10.3969/j.issn.1673-8640.2022.02.006

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Objective To investigate the correlation between metabolic syndrome（MS） and moderate-to-severe periodontitis complicated with psoriasis. Methods A total of 50 patients with moderate-to-severe periodontitis,50 patients with psoriasis and 50 patients with moderate-to-severe periodontitis complicated with psoriasis were enrolled. Totally,50 healthy subjects were enrolled as control group. Blood pressure,glycated hemoglobin A_1c（HbA_1c）,glucose（Glu）,triglyceride（TG）,total cholesterol（TC）,high-density lipoprotein cholesterol（HDL-C） and low-density lipoprotein cholesterol（LDL-C） were determined. Multivariate Logistic regression analysis was used to assess the risk factors for disease development in each group. Results TC and LDL-C in moderate-to-severe periodontitis group and psoriasis group were higher than those in control group（P<0.05）. TG,LDL-C,TC and HbA_1c were higher in moderate-to-severe periodontitis complicated with psoriasis group than those in moderate-to-severe periodontitis group（P<0.05）,and TG,LDL-C and TC were higher than those in psoriasis group（P<0.05）. In moderate-to-severe periodontitis complicated with psoriasis group,TG,LDL-C,TC,Glu and HbA_1c were higher than those in control group（P<0.01）. Multivariate Logistic regression analysis showed that elevated LDL-C was a risk factor for the development of moderate-to-severe periodontitis [odds ratio（OR）=1.873,95% confidence interval（CI） 1.113-3.154],elevated TC was a risk factor for the development of psoriasis（OR=1.527,95%CI 1.012-2.304）,and elevated TC and HbA_1c were risk factors for the development of moderate-to-severe periodontitis complicated with psoriasis（OR=2.728 and 3.108,95%CI 1.572-4.732 and 1.283-7.533,respectively）. Conclusions Patients with moderate-to-severe periodontitis complicated with psoriasis show more significant disorders of MS-related components than patients with periodontitis or psoriasis alone,and elevated TC and HbA_1c are risk factors for the development of moderate-to-severe periodontitis complicated with psoriasis.

Prognostic role of intestinal fatty acid-binding protein in critically ill patients

LIU Jichun, ZHANG Yanju

2022, 37(2):  130-133.  DOI: 10.3969/j.issn.1673-8640.2022.02.007

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Objective To investigate the prognostic role of intestinal fatty acid binding protein（IFABP） in critically ill patients. Methods A total of 123 critically ill patients were enrolled,and the clinical data,including age,sex,basic diseases and quick sequential organ failure assessment（qSOFA） score,were recorded within 24 h of admission. According to the outcome of 28 d,they were classified into survival group（95 cases） and death group（28 cases）. Totally,35 healthy subjects were enrolled as control group. IFABP and C-reactive protein（CRP） levels were determined. The efficacy of each index was evaluated by receiver operating characteristic（ROC） curve,and the risk factors of death in critically ill patients were screened by multivariate Logistic regression analysis of binary variables. Results The levels of serum IFABP and CRP in death group,survival group and control group decreased gradually（P<0.05）. ROC curve analysis showed that the areas under curves（AUC）of IFABP,CRP and qSOFA score in the mortality risk of critically ill patients were 0.865,0.750 and 0.808,respectively. Logistic regression analysis showed that IFABP≥36.20 ng/mL,CRP≥36.82 mg/L and qSOFA score≥2.0 were risk factors for death in critically ill patients [odds ratios（OR） were 10.668,3.775 and 9.048,95% confidence intervals（CI） were 3.498-32.533,1.156-12.325 and 1.818-45.038,respectively]. Conclusions IFABP may be an effective predictor of mortality risk in critically ill patients,and its increase may indicate the risk of death in critically ill patients.

Relationship between procalcitonin level and all-cause death in maintenance hemodialysis patients

GU Feng, WU Yanfen, ZHAO Xinhui, HOU Zhaoyuan, WANG Zhihong, QI Hualin

2022, 37(2):  134-140.  DOI: 10.3969/j.issn.1673-8640.2022.02.008

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Objective To investigate the relationship between procalcitonin（PCT） level and all-cause death in patients undergoing maintenance hemodialysis（MHD）. Methods A total of 160 patients undergoing MHD were enrolled,and their clinical data,including sex,age,body mass index（BMI）,smoking history,primary disease,comorbidities,dialysis duration,New York Heart Association（NYHA） cardiac function classification and laboratory indicators,were collected. Receiver operating characteristic（ROC） curve was used to determine the optimal cut-off value of death and survival in MHD patients. The patients with PCT> optimal cut-off value were taken as high PCT group,and those with PCT≤optimal cut-off value were taken as low PCT group. The influencing factors of high PCT were analyzed by binary Logistic regression. Kaplan-Meier survival curve was used to analyze the survival rate of MHD patients. Cox proportional risk regression analysis was used to evaluate the relationship between PCT and all-cause death in MHD patients. Results The dialysis duration and CRP levels in high PCT group were higher than those in low PCT group（P<0.01）,but there was no statistical significance for the other indicators（P>0.05）. Binary Logistic regression analysis showed that increased CRP level was a risk factor for high PCT [odds ratio（OR）=1.182,95% confidence interval（CI）1.043-1.339,P=0.009]. Kaplan-Meier survival curve analysis showed that the survival rate of high PCT group was lower than that of low PCT group（Log-rank χ²=6.707,P=0.01;Breslow test value was 6.828,P=0.009）. Cox proportional risk regression analysis showed that high triglyceride（TG） level was a protective factor for all-cause death in MHD patients [hazard ratio（HR）=0.166,95%CI 0.071-0.387]. High PCT level was a risk factor for all-cause death in MHD patients（HR=4.409,95%CI 1.757-11.064）. Conclusions High PCT level may be an independent risk factor for all-cause death in MHD patients.

Analysis of molecular typing,virulence genes and antibiotic susceptibility of uropathogenic Escherichia coli in patients with recurrent urinary tract infection

PAN Yunqi, LI Yungai, WANG Jianqiang, WU Qiong, TANG Jin

2022, 37(2):  141-145.  DOI: 10.3969/j.issn.1673-8640.2022.02.009

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Objective To investigate molecular typing,virulence genes and antibiotic susceptibility of uropathogenic Escherichia coli in patients with recurrent urinary tract infection（rUTI）. Methods Totally,60 patients with rUTI were enrolled,and the isolates of Escherichia coli were isolated. Pulsed-field gel electrophoresis（PFGE）,multi-locus sequence typing（MLST）and phylogenetic groups were performed. Polymerase chain reaction（PCR） and sequencing were used to analyze common Escherichia coli virulence genes（feoB,chuA,iutA,iroN,ireA,fimH,foc,sfa,aer and cnf）. The antibiotic susceptibilities of Escherichia coli to 13 common antimicrobial agents were analyzed. Results The research subjects were classified into relapse and reinfection groups by PFGE. A total of 30 isolates from relapse group and 60 isolates from reinfection group were collected for further analysis. MLST indicated that ST131 was the predominant type followed by ST405. The major phylogenetic groups were B2 and D. Among the 10 virulence genes,feoB（98.8%） was the most common virulence gene followed by fimH（93.3%）. Except for iutA（P=0.020）,no statistical significance of the other virulence genes was found between the 2 groups（P>0.05）. Conclusions The predominant types of uropathogenic Escherichia coli are ST405 and ST131 in Shanghai Sixth People's Hospital. Two virulence genes,feoB and fimH,are highly prevalent. The carry rate of iutA is high in relapse group,which can be as the treatment target for rUTI.

Analysis of drug resistance of rare CRE and clinical application of carbapenem screening test in Henan

SUN Ying, LI Yi, YAN Wenjuan, JING Nan, MA Bing

2022, 37(2):  146-149.  DOI: 10.3969/j.issn.1673-8640.2022.02.010

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Objective To investigate the determination and drug resistance characteristics of rare carbapenem-resistant Enterobacteriaceae（CRE）in Henan No.3 Provincial People's Hospital,to investigate the clinical application of CRE enzyme screening test,and to provide a reference for rational clinical use of antibiotics. Methods The non-repetitive rare CRE isolated in 2019 were collected,and the results of drug resistance sensitivity test in vitro were collected. There were 22 isolates randomly collected,and the carbapenem-producing enzyme patterns of 22 isolates were determined by ethylenediaminetetraacetic acid（EDTA）,3-aminophenylboric acid（APBA） carbapenemase inhibition test and colloidal gold immunochromatography. Results A total of 1 111 isolates of CRE were determined in 2019,which included 49 isolates of Enterobacter cloacae（15.4%）,14 isolates of Klebsiella acidogenes（19.2%） and 12 isolates of Citrobacter（13.8%）. The drug resistance rate of imipenem was 100%. Enterobacter,Klebsiella acidogenes and Citrobacter were sensitive to tigecycline and polymyxin B. The drug resistance rates of Proteus and Providencia which were naturally resistant to polymyxin B and tigecycline were 5.3% and 14.3%,respectively. Totally,13 isolates of NDM type metalloenzyme,3 isolates of KPC type serinase,1 isolate of OXA type serinase,1 isolate of KPC+IMP+NDM type and 1 isolate of KPC+NDM type were determined in the 22 isolates of CRE. Conclusions The situation of drug resistance of CRE in Henan is serious,and it should be monitored in time and used antibiotics reasonably according to the results of drug resistance sensitivity test. The carbapenem screening test should be optimized to provide a reference for the accurate use of drugs in CRE.

Using ampicillin to predict imipenem susceptibilities of Enterococcus faecalis and Enterococcus faecium

GUO Lanfang, GUO Yan, ZHENG Yonggui, YANG Yang, YIN Dandan, SHI Qingyu, HU Fupin

2022, 37(2):  150-154.  DOI: 10.3969/j.issn.1673-8640.2022.02.011

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Objective To evaluate the feasibility of ampicillin to predict the imipenem susceptibilities of Enterococcus faecalis and Enterococcus faecium. Methods Totally,127 non-duplicated isolates of Enterococcus faecalis and 124 isolates of Enterococcus faecium were collected from 23 hospitals. The susceptibilities of Enterococcus faecalis and Enterococcus faecium to penicillin,ampicillin and imipenem were determined by broth microdilution and disk diffusion methods. Results For Enterococcus faecalis,which were both sensitive or resistant to penicillin and ampicillin,the categorical agreement（CA） of ampicillin-imipenem by broth microdilution was 100.0%,and there was no very major error（VME） and major error（ME）. For Enterococcus faecalis,which were resistant to both penicillin and ampicillin,the CA of ampicillin-imipenem by disk diffusion was 77.8%,and the ME accounted for 22.2%. For penicillin-resistant but ampicillin-sensitive Enterococcus faecalis,the CA of ampicillin-imipenem were 57.1% and 81.8% by broth microdilution and disk diffusion. Conclusions Ampicillin is better than penicillin to predict the susceptibilities of Enterococcus faecalis and Enterococcus faecium to imipenem. The susceptibilities to ampicillin can not be used to predict the susceptibilities of penicillin-resistant but ampicillin-sensitive Enterococcus faecalis and Enterococcus faecium to imipenem. Enterococcus faecalis and Enterococcus faecium isolates being sensitive to penicillin are predictably susceptible to ampicillin.

Inhibitory elimination effects of different enhancers on urea,heme and cholate in fluorescent PCR

LIU Wenxia, WANG Wei, LIN Songyang, LI Zhenhong

2022, 37(2):  155-158.  DOI: 10.3969/j.issn.1673-8640.2022.02.012

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Objective To investigate the inhibitory elimination effects of different polymerase chain reaction（PCR） enhancers on cholate,urea and heme. Methods Different concentrations of PCR enhancers [spermidine,formamide,PEG6000,NP40,Tween-20,betaine,trehalose,bovine serum albumin（BSA）,gelatin,ammonium sulfate,tetramethylammonium chloride（TMAC）,glycerol and Triton X-100] were added to the amplification system containing cholate,urea and heme. The anti inhibitory effects of different PCR enhancers were determined by fluorescent PCR and analyzed according to the cycle threshold（Ct） value. Results The 6.5 mmol/L spermidine and 20% glycerol could eliminate the inhibitory effect of 6.98 mmol/L cholate on PCR. The 6.5 mmol/L spermidine,100 mmol/L ammonium sulfate and 0.2 mol/L betaine could eliminate the inhibitory effect of 0.99 mol/L urea on PCR. The 300 mmol/L BSA,6.5 mmol/L spermidine,20% PEG6000 and 50% glycerol could eliminate the inhibitory effect of 6.98 mmol/L heme on PCR. The 40% formamide,10% NP40,40% Tween-20, 1.5 mol/L trehalose,2% gelatin,200 mmol/L TMAC and 20% Triton X-100 could not reduce the inhibitory effects of 6.98 mmol/L cholate,0.99 mol/L urea and 6.98 mmol/L heme on PCR. Conclusions Adding different types of PCR enhancers can effectively improve the determination sensitivity and accuracy of fluorescent PCR.

Simultaneous determination of 5 steroid hormones in human serum by ultra-performance liquid chromatography-tandem mass spectrometry and establishment of reference intervals in healthy adults

LU Youli, YANG Shuangshuang, ZHANG Meiwei, OU Meixian, DONG Chunxia, SHEN Weiwei, JIANG Fengli, LI Shuijun

2022, 37(2):  165-173.  DOI: 10.3969/j.issn.1673-8640.2022.02.015

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Objective To establish and validate a method for the simultaneous determination of testosterone（T）,dihydrotestosterone（DHT）,androstenedione（AD）,dehydroepiandrosterone（DHEA） and 17alpha-hydroxyprogesterone（17OHP） in human serum by ultra-performance liquid chromatography-tandem mass spectrometry（UPLC-MS/MS）,and to establish the reference intervals of 5 steroid hormones in healthy adults. Methods The serum samples were purified by SOLAμ HRP solid-phase extraction,and they were separated by Poroshell 120 EC-C18 analytical column. Mobile phase was constituted of 30% acetonitrile solution and 90% acetonitrile solution containing 0.05% formic acid with gradient elution. The electrospray positive ion multi-reaction monitoring mode was used for determination,and the isotope internal standard method was used to quantify the concentration of steroid hormones. The performance of the method was verified comprehensively,and the reference intervals suitable for the local population in Shanghai were established. Results The minimum limits of quantification of the 5 steroid hormones in human serum were 0.01-0.10 ng/mL. The standard curve range covered 1 000 times （r²≥0.999 5） and showed good linearity,and the sample after dilution can be reported to cover the range of 5 000 times. The average accuracy of 5 steroid hormones ranged from 95.0% to 105.6%,with the precision of <8%. None of the 8 structural analogs interfered with the target analytes,high triglyceride blood did not affect the determination results,and hemolysis resulted in low DHEA concentration. The extraction recoveries of the analytes were 60.5%-93.3%,and there was no obvious matrix effect after calibration of internal standard. Analytes demonstrated good stability at room temperature for 4 h,at -25--15 ℃ for 30 d or at -90--70 ℃ for 100 d. The method was used and participated in a survey of endocrine accuracy verification supplied by the National Center for Clinical Laboratories in 2019. The determination deviations of T and 17OHP were -5.00%-0.38%. The results of 367 physical examination subjects were analyzed and showed that there was statistical significance in the concentration of 5 steroid hormones between females and males（P<0.05）. There was statistical significance between the 2 age subgroups（aged 16-50 years and aged>50 years） of female groups（P<0.05）. Conclusions A UPLC-MS/MS has been established for the determination of 5 steroid hormones in human serum,and the reference intervals suitable for the Central Laboratory of Shanghai Xuhui Central Hospital have been verified.

RSV infection of air-liquid interface cultured human bronchial epithelial cell and its effect on HMGB1 and pMLKL expressions

WANG Juan, LIAO Huanjin, LI Yanning, GE Yiqin, LI Jia

2022, 37(2):  177-182.  DOI: 10.3969/j.issn.1673-8640.2022.02.017

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Objective To construct a respiratory syncytial virus（RSV） infected the model of air-liquid interface（ALI）cultured human bronchial epithelial cells（hBEC）,and to provide a cell model closer to the in vivo environment for further study of respiratory virus pathogenesis. By analyzing the effect of RSV infection on the expressions of high mobility group box protein 1（HMGB1） and phosphorylated mixed-lineage kinase domain-like protein（pMLKL）,to investigate the injury pathogenesis caused by RSV. Methods The hBEC was inoculated on Transwell membrane and cultured unsubmerged mode. After the confluence of 100%,cells were transferred to ALI culture. After the cells were differentiated and matured,they were infected respectively with RSV according to the following groups:6 h control group,6 h MOI 1.0 group,6 h MOI 3.0 group,24 h control group,24 h MOI 1.0 group,24 h MOI 3.0 group,each group with 3 repeated holes. The effect of RSV infection on the expressions of HMGB1 and pMLKL was confirmed by immunofluorescence staining. Results After 4-10 d of submerged culture,the cell confluence reached 100%. Then,cells were transferred to ALI interface culture. After about 4 weeks,the distribution of cell bands became clearer,and visible mucus was secreted to form a mucus layer. HE staining showed typical pseudostratified epithelium,and immunofluorescence staining showed that RSV successfully infected cells at MOI 3.0 24 h. Anti-HMGB1 and anti-pMLKL immunofluorescence staining showed that under the condition of infection,there was pink fluorescence in the nucleus [blue 4',6-diamidino-2-phenylindole（DAPI）and red HMGB1 fluorescence merged results],which confirmed the expression of HMGB1 after RSV infection. However,there was no pMLKL expression neither before nor after RSV infection. Conclusions The well-differentiated hBEC can be obtained by submerged and ALI culture,which maintain cell morphology and function for a long time,and provide a cell model for respiratory virus infection and other common respiratory disease studies. At MOI 3.0 24 h,RSV successfully infects the cell model and causes the expression of HMGB1.

Influence of different-brand calibrators on the comparability of serum alkaline phosphatase determination results

LIN Feiran, OU Yuanzhu, LIU Wenbin, YU Xiaoxuan, GE Danhong, ZHAO Ran

2022, 37(2):  183-187.  DOI: 10.3969/j.issn.1673-8640.2022.02.018

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Objective To discuss the influence of different-brand calibrators and matrix on the comparability of alkaline phosphatase（ALP） determination,and to provide a reference for further standardization of serum ALP determination and clinical result mutual accreditation. Methods ALP was calibrated on AU5800 analyzer and cobas c501 analyzer by 2 commonly used calibrators（AU calibration and C.f.a.s. calibration） and ALP national secondary reference material. Totally,25 fresh clinical samples and 5 concentration level samples of routine chemistry of Shanghai external quality assessment were determined for 2 times. Taking the test value after calibration of the reference material as the reference system,the bias and correlation between determination values after calibrating the matching calibration and reference material were calculated. Taking the determination value of the c501 as the reference system,the bias and correlation between determination values obtained by the 2 instruments after the same calibrator were calculated and calibrated. Results Taking the determination value after calibration of the reference material as the reference system,the absolute value of the bias of the 2 systems was <10%. The c501 had a negative bias,while the AU5800 had a positive bias. There was no statistical significance in the bias between the external quality assessment samples and the clinical samples. Taking c501 determination value as the reference system,after using reference material to calibrate clinical serum samples,the differences between the systems were reduced. After the calibration of the non-matching system,the differences between the systems were increased. The external quality assessment samples presented the opposite results from the serum samples. Conclusions The use of traceability and commutability reference materials is beneficial to the consistency and comparability of the determination results between different determination systems and laboratories. The use of non-matching system calibrators will increase the differences between systems and will not be interchangeable. In addition,the external quality assessment samples with matrix effect can not fully show the real determination results of each system.

Analysis of external quality assessment of parasite morphology from 2013 to 2020 in West China Hospital of Sichuan University

DENG Shanying, PEI Yuqing, ZHANG Chunying, XIE Heng, MENG Qiang, MA Ying

2022, 37(2):  188-193.  DOI: 10.3969/j.issn.1673-8640.2022.02.019

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Objective To improve the ability of clinical laboratory examiners to detect and identify parasites through reviewing and analyzing the results of the external quality assessment of parasite morphology. Methods A retrospective analysis of 200 samples from the external quality assessment of parasite morphology from 2013 to 2020 was conducted,of which 120 samples were from the National Center for Clinical Laboratories and the other 80 samples from the College of American Pathologists（CAP）. The accuracy of external quality assessment was calculated,the reasons for misidentification of parasites（eggs） were analyzed,and the morphological characteristics and identification points of rare and similar parasites were summarized. Results Of the 200 samples,195 samples were identified correctly（an accuracy rate of 97.5%）. Of the 80 CAP samples,77 samples were identified correctly,3 samples were identified incorrectly（an accuracy rate of 96.3%）,and 12 samples were not graded. Of the 120 samples from the National Center for Clinical Laboratories,118 samples were identified correctly,with only 2 identification errors（an accuracy rate of 98.3%）. Totally,30 parasites in 47 different life-history forms were involved in the external quality assessment of parasite morphology. The morphologically similar parasite eggs in identification included Diphyllobothrium latum eggs,Paragonimus westermani eggs,Hepatella hepatica/Fasciolopsis buski eggs,Ascaris lumbricoides eggs and hookworm eggs,hookworm eggs and Trichostrongylus orienatalis eggs,microfilaria-Brugia malayi and microfilaria-Wuchereria bancrofti. The rare parasite eggs identified were Hymenolepis nana eggs and Hymenolepis diminuta eggs. Food residue and parasite eggs in stool,as well as different kinds of cysts,were also identified. Conclusions Periodically retrospective analysis of the laboratory external quality assessment data of parasites,regular review of mixed egg samples under a microscope,review of parasite archives,and morphology review of rare and similar parasites（eggs） can improve the quality of parasite laboratory and improve the capabilities of clinical laboratory examiners.