Table of Content

15 September 2013, Volume 28 Issue 9

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Change and clinical significance of peripheral blood cells and C reactive protein in patients with avian influenza A(H7N9)virus infection

LI Xin,SHEN Zhen,ZHANG Bei,LU Meijuan,HU Xiangnan,HU L鼀in,ZHANG Jun.

2013, 28(9):  745-748.  DOI: 10.3969/j.issn.1673-8640.2013.09.003

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Objective To investigate the changes of peripheral blood cells and C reactive protein (CRP) in patients with avian influenza A(H7N9)virus infection, and provide the relative reference for clinical diagnosis,treatment and monitor. Methods A total of 18 patients with avian influenza A(H7N9)virus infection (H7N9 group) were enrolled in the study, 20 patients with avian influenza A (H1N1)virus infection in 2009 (H1N1 group) were enrolled as control group, and 20 healthy subjects were also enrolled as healthy control group. H7N9 group was sub-classified into mild group (12 cases) and severe group (6 cases). The peripheral blood cells in all groups were quantified, and the serum CRP in H7N9 group was also determined. Results For H7N9 group,the white blood cell (WBC) count did not change much in the early period of infection onset,while the lymphocytes (Lym) decreased significantly when compared with healthy control group (P<0.05),together with the decreasing of platelets (PLT) and eosinophils (Eos)(P<0.05). For the severe group,the neutrophils (Neu) increased significantly when compared with those of healthy control group (P<0.05). However,no statistical significance was observed for all the parameters between the mild and severe groups (P>0.05). The clinical characteristics of peripheral blood cells in H7N9 group were similar with those in H1N1 group in 2009,while the Lym decreased more significantly(P<0.05). The PLT, Lym and Eos all increased significantly after treatment in the mild group when compared with those of pre-treatment(P<0.05). The serum CRP increased significantly in H7N9 group after administration,which was more obvious in the severe group than in the mild group (P<0.05). Similarly,the serum CRP decreased after treatment in the mild group when compared with that of pre-treatment (P<0.05). Conclusions The serum CRP level may be a parameter on distinguishing the mild group from severe group. However,the clinical characteristics of peripheral blood cells in patients with avian influenza A (H7N9)virus infection are similar with those in patients with avian influenza A(H1N1)virus infection in 2009. Clinical information regarding different aspects should be carefully collected for clinical diagnosis, prognosis assessment and prevention of influenza outbreaks.

Changes and clinical significance of biochemistry markers in patients with avian influenza A (H7N9) virus infection

CHEN Zhijin ,LI Haicong,FANG Huanying,ZHOU Yi,ZHANG Jun.

2013, 28(9):  749-754.  DOI: 10.3969/j.issn.1673-8640.2013.09.004

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Objective To investigate the changes and clinical significance of biochemistry markers in patients with avian influenza A (H7N9) virus infection (2013) before and after treatment. Methods A total of 12 biochemistry markers, including cardiac markers [cardiac troponin T(cTnT), creatine kinase isoenzyme (CK-MB),myoglobin(MYO) and B-type natriuretic peptide(NT-proBNP)],liver function markers [aspartate aminotransferase(AST),alanine aminotransferase(ALT),gamma-glutamyl transpeptidase(GGT),alkaline phosphatase(ALP),total protein(TP) and total bile acid(TBA)] and kidney function markers [creatinine(Cr) and urea nitrogen(BUN)] were determined in 12 cured patients (H7N9 cure group),5 dead patients (H7N9 death group) with avian influenza A (H7N9) virus infection. A total of 50 patients with avian influenza A (H1N1) virus infection (2009) (H1N1 group) and 12 healthy subjects (control group) were enrolled, and the results were analyzed and compared. Results The levels of CK-MB, MYO and NT-proBNP increased in H7N9 cure group and H1N1 group compared with those in control group before the treatment (P<0.05),however,the level of cTnT was normal (P>0.05).The levels of AST, ALT and GGT increased (P<0.05), while those of ALP and TP decreased (P<0.05). The levels of Cr and BUN were normal (P>0.05). Compared with H1N1 group, the increase and decrease degrees for the levels of ALT, ALP, GGT and TP were different, and there was no statistical significance for the other markers (P>0.05). The levels of CK-MB and NT-proBNP in H7N9 death group were higher than those in H7N9 cure group, and the levels of TBA and Cr increased (P<0.05). The levels of CK-MB, MYO, NT-proBNP, AST and TBA decreased after the treatment compared with those before the treatment, and the levels of ALP and TP increased (P<0.05). Compared with the control group, except AST, ALT and GGT, the other markers had no statistical significance in H7N9 cure group after the treatment (P>0.05), and except TP, the other markers had difference in H7N9 death group after the treatment (P<0.05). Conclusions The biochemistry markers may be helpful for the auxiliary diagnosis, progress monitoring and prognosis among the patients with avian influenza A (H7N9) virus infection. The clinical characteristics of biochemistry markers in patients with avian influenza A (H7N9) virus infection are similar with that in patients with avian influenza A (H1N1) virus infection (2009).

Application of real-time fluorescence RT-PCR to detect avian influenza A(H7N9) virus

HUA Zheyun, CHU Wei, SONG Lili, WANG Yu, GAO Shuna, LAO Baofa.

2013, 28(9):  755-757.  DOI: 10.3969/j.issn.1673-8640.2013.09.005

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Objective To assess the application effect of real-time fluorescence reverse transcription-polymerase chain reaction (RT-PCR) to detect avian influenza A(H7N9) virus, and to provide reliable data to laboratory detection for avian influenza A(H7N9) virus surveillance, prevention and control. Methods A total of 137 being monitored and suspected specimens from hospitals were collected, and the nucleic acids of avian influenza A(H7N9)virus in specimens from upper and lower respiratory tracts were detected by 4 kinds of real-time fluorescence RT-PCR kits. Results The positive rate of avian influenza A(H7N9)virus was 3.65%(5/137). Among the positive specimens,the positive rate of upper respiratory tract was 60%(3/5), and the positive rate of lower respiratory tract was 100%(3/3). The coincidence rate of 4 kinds of kits was 100%. Conclusions Real-time fluorescence RT-PCR can detect the nucleic acid of avian influenza A(H7N9)virus effectively. Specimens from lower respiratory tract contribute to improve the positive detection rate of avian influenza A (H7N9) virus.

Analysis and significance of T lymphocyte subsets in peripheral blood of patients with avian influenza A (H7N9) virus infection

MAO Huijun,CHEN Daihong,ZHANG Min,WANG Gang,QIU Jianping

2013, 28(9):  758-761.  DOI: 10.3969/j.issn.1673-8640.2013.09.006

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Objective To investigate the changes of peripheral blood T lymphocyte subsets in avian influenza A (H7N9) virus infection patients. Methods According to the prognosis,18 patients with avian influenza A (H7N9)virus infection (infection group) were classified into 13 cases of cure group and 5 cases of dead group. There were 20 healthy subjects as the control group. Specific primer polymerase chain reaction (PCR) was used to detect the avian influenza A (H7N9)virus,and the T lymphocyte subsets in peripheral blood were detected by flow cytometry. Results Compared with the control group,the absolute numbers of CD3⁺T cell, CD8⁺T cell and CD4⁺T cell and CD4⁺/CD8⁺ ratio of the infection group decreased significantly (P<0.05). In the cure group,the absolute numbers of CD3⁺T cell, CD8⁺T cell and CD4⁺T cell were lower than those of the control group (P<0.05). In the cure group,the absolute numbers of CD3⁺T cell, CD8⁺T cell and CD4⁺T cell before the treatment were significantly lower than those after the treatment (P<0.05), and the absolute numbers of CD3⁺T cell and CD4⁺T cell and CD4⁺/CD8⁺ ratio were lower than those in the control group (P<0.05).In the dead group, the absolute numbers of CD3⁺T cell, CD8⁺T cell and CD4⁺T cell before and after the treatment were lower than those in the cure and control groups (P<0.05) , and there was no statistical significance for the all parameters before and after the treatment (P>0.05). Compared with the dead group,the absolute numbers of CD3⁺T cell, CD8⁺T cell and CD4⁺T cell in the cure group were higher (P<0.05). Conclusions There is the imbalance of T lymphocyte subsets in avian influenza A (H7N9)virus infection patients. In the dead group,the imbalance of T lymphocyte subsets is serious. The imbalance may be correlated with the pathogenesis of avian influenza A (H7N9)virus infection.

Investigation on APTT and PT reference intervals in children and adolescents for STA-Compact hemostasis system

ZHENG Jianxin,ZHENG Zhaojing,YANG Hui, LI Huaiyuan, FU Qihua

2013, 28(9):  762-764.  DOI: 10.3969/j.issn.1673-8640.2013.09.007

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Objective To investigate activated partial thromboplastin time (APTT) and prothrombin time (PT) reference intervals in children and adolescents by retrospective analysis. Methods Patients in Shanghai Children′s Medical Center for surgical procedures and aged from 1 day old to 18 years old were enrolled for this study after excluding patients with hematological disorders, patients with cardiopulmonary diseases and patients with diseases of liver and kidney. A coagulation screening test was performed for all patients who were not treated with anti-coagulation or anti-platelet drugs for the past 2 weeks with a STA-Compact hemostasis system. There were 4 166 children eligible for this study, and all the children were classified into ≤30 d, 31 d-1 year, 2-6 years, 7-14 years and 15-18 years groups, respectively. The SPSS 17.0 software was used to analyze APTT and PT data. Results The sex distribution had no difference among the 5 groups. No statistical significance was observed on APTT results between the group of ≤30 d with the groups of 31 d-1 year, 2-6 years and 7-14 years, and on PT results among the group of 2-6 years with the group of 7-14 years (P>0.05). There were statistical significance among the other groups (P=0.000). Conclusions Through retrospective analysis, the reference intervals of APTT and PT for STA-Compact hemostasis system in children and adolescents are established.

Analysis on risk factors for aspirin resistance in patients with acute cerebral infarction

DENG Lin,QIU Lijun,GU Qing,WANG Genfa

2013, 28(9):  765-769.  DOI: 10.3969/j.issn.1673-8640.2013.09.008

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Objective To investigate the aspirin resistance (AR) and its risk factors in patients with acute cerebral infarction. Methods A total of 330 patients with acute cerebral infarction were enrolled. According to the platelet inhibition rate of arachidonic acid (AA) by thrombelastography, the patients were classified into AR group (platelet inhibition rate ≤20%), aspirin semi-resistance (ASR) group (platelet inhibition rate 20%-50%) and aspirin sensitivity (AS) group (platelet inhibition rate >50%). The incidence rate of AR was analyzed, the difference of clinical characteristics was analyzed, and Logistic regression analysis was performed for the risk factors of AR and ASR. Results The 33.9% patients had AR, and 19.7% patients had ASR. The high sensitive C reactive protein (hsCRP) level in AR+ASR group increased obviously compared with that in AS group(P<0.05).In the patients older than 65 years old,the incidence rate of AR+ASR in high risk patients increased significantly (P<0.05). Logistic regression analysis showed that platelet counts [odd ratio (OR)=0.996,95％ confidence interval (CI): 0.991-0.999,P=0.041] and hs-CRP levels(OR=0.972,95％CI: 0.959-0.996,P=0.014) were risk factors for AR and ASR. Conclusions The incidence rate of AR in patients with acute cerebral infarction may be related to platelet count and inflammation.

The relationship between gene polymorphism of platelet glycoprotein Ⅱb/Ⅲa and therapeutic effect of platelet transfusion

GU Ping,WANG Jing,ZHANG Fan,YAO Ruen,FU Qihua.

2013, 28(9):  770-774.  DOI: 10.3969/j.issn.1673-8640.2013.09.009

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Objective To investigate the relationship between gene polymorphism and therapeutic effect of platelet transfusion by detecting the gene polymorphism of platelet glycoprotein(GP) Ⅱb/Ⅲa and platelet antibody in 98 patients. Methods A total of 9 sites of 5 exons in GP Ⅱb/Ⅲa gene were amplified by polymerase chain reaction(PCR),and PCR products were directly sequenced to analyze the gene polymorphism. Platelet antibody was detected by solid phase agglutination assay. Results Only T18809G variation existing in Exon 26 of GP Ⅱb gene was found in 98 patients,which causing Ile843Ser polymorphism and producing HPA-3aa/ab/bb genotypes. All of the 3 genotypes can cause platelet antibodies,resulting platelet transfusion refractoriness. The positive rate of platelet antibody in 98 patients was 52.0%, the total effective rate of 1 468 platelet transfusion was 77.8%,and the effective rate of 65 matched platelet transfusion was 83.1%. There were no significant differences among the 3 genotypes. Conclusions The specificity and associated platelet antibody are the main immune factors for causing platelet transfusion refractoriness, and HPA-3 polymorphism in GP Ⅱb gene is associated with platelet transfusion refractoriness.

Distribution of 16S rRNA methylase genes in ESBLs-producing Enterobacteriaceae

GAO Hui,WANG Yang,HUANG Yunkun, ZHU Wenmei, WANG Jia, YAO Yao

2013, 28(9):  775-779.  DOI: 10.3969/j.issn.1673-8640.2013.09.010

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Objective To investigate the distribution of 16S rRNA methylase genes in extended-spectrum beta-lactamases (ESBLs)-producing Gram-negative Enterobacteriaceae. Methods A total of 70 isolates of Gram-negative Enterobacteriaceae were identified by VITEK-32 automated microbial system. The Kirby-Bauer method was used to detect ESBLs. Polymerase chain reaction(PCR) was used to detect 6 kinds of 16S rRNA methylase genes including armA, rmtA, rmtB, rmtC, rmtD and npmA. DNA sequencing was used to detect the positive productions of PCR with GenBank as control. Results From the 70 isolates of ESBLs-producing Gram-negative Enterobacteriaceae, 9 isolates carried 16S rRNA methylase genes, including 5 for armA gene, 4 for rmtB gene and 2 for both armA and rmtB genes, and no rmtA, rmtC, rmtD or npmA gene was detected. Conclusions The distribution of 16S rRNA methylase genes is various in the hospitals from different regions.

Research on the mechanisms of fluconazole resistance in clinical isolates of Candida glabrata

ZHANG Wei,YING Chunmei

2013, 28(9):  780-783.  DOI: 10.3969/j.issn.1673-8640.2013.09.011

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Analysis of multidrug-resistant strain hospital infections

LI Jixia,GONG Yanwen.

2013, 28(9):  784-788.  DOI: 10.3969/j.issn.1673-8640.2013.09.012

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Objective To understand the distribution trend of multidrug-resistant strains, in order to provide the reference of decreasing the incidence of multidrug-resistant strain infection. Methods All kinds of samples from Inpatient Department in Jinan Military General Hospital from January 2006 to December 2011 were collected. The strains were identified by VITEK-32 or VITEK-2 COMPACT 60(after November 2007), and the susceptibility testing was determined by Kirby-Bauer method. The data were analyzed statistically by WHO-NET 5.4 software. Results From January 2006 to December 2011, multidrug-resistant strain infection had increasing trend, and the infection rate of extended spectrum beta-lactamases (ESBLs)-producing strains was highest. The infection rates of pandrug-resistant Acinetobacter baumannii (PDR-AB) and extensively drug-resistant Pseudomonas aeruginosa (XDR-PA) increased obviously. Multidrug-resistant strains were mainly in Departments of Health, Neurosurgery, Intensive Care Unit, Surgery and Hematology among the elder and serious patients with long-term use of antibiotics and immunosuppressant. The main clinical manifestation was respiratory tract infection. Conclusions Multidrug-resistant strain hospital infections have increased year by year, and the monitoring of drug resistance should be strengthened. The prevention should be established in order to reduce the rate of hospital infections.

A comparative study of three determination methods on the effect of apoE genotyping

XIONG Yu,QIAN Shiyun.

2013, 28(9):  789-792.  DOI: 10.3969/j.issn.1673-8640.2013.09.013

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Objective To evaluate the 3 determination methods [multiplex amplification refractory mutation system polymerase chain reaction (Multi-ARMS PCR), polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP)] on the effect of apolipoprotein E (apo E) genotyping. Methods apo E genotypes of 518 healthy subjects were analyzed by Multi-ARMS PCR, PCR-RFLP and PCR-SSCP respectively, and the results were confirmed by DNA sequencing. Results The results of the 3 determination methods were not entirely consistent with the results of DNA sequencing (Multi-ARMS PCR Kappa=0.964, PCR-RFLP Kappa=0.991, PCR-SSCP Kappa=0.913, and Kappa was closer to 1, the consistency was better). Conclusions Among the 3 determination methods, PCR-RFLP is the most reliable one for apoE genotyping.

Change analysis on serum GPDA in upper gastrointestinal bleeding

ZHANG Gui,TANG Zesheng,LAI Baoyi,SHI Qingxin.

2013, 28(9):  793-795.  DOI: 10.3969/j.issn.1673-8640.2013.09.014

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Objective To investigate the relationship of serum glycyl-proline dipeptidyl aminopeptidase(GPDA) and upper gastrointestinal bleeding(UGIB). Methods Serum GPDA and urea concentrations were measured in 33 patients with UGIB(UGIB group)and 33 healthy subjects (control group). The UGIB group was classified according sex and age (<60 years old and ≥60 years old). The results were analyzed statisically. Results Serum GPDA and urea concentrations between UGIB group and control group were significant statistically (P=0.000). The concentrations of serum GPDA and urea in different age groups had statistical significance (P<0.01). There was no statistical significance between male and female groups (P>0.05). There was a negative correlation of GPDA and urea concentrations in UGIB group [coefficient of correlation (r)=-0.585, P<0.01]. Receiver operating characteristic (ROC) curve showed that the areas under the ROC curve (AUC) were 0.893 and 0.769 for GPDA and urea. When the cut-off values of GPDA and urea were 69.20 U/L and 8.25 mmol/L, the sensitivities were 97.0% and 63.6%, and the specificities were 69.7% and 97.0%. Conclusions Serum GPDA reduces significantly when UGIB, and GPDA may play an important role in the course of UGIB.

Analysis on the laboratory determination results of hand-foot-mouth disease in Fuyang in 2011

GAO Yong,HAN Mingfeng,LI Xiuyong, CHEN Xiaofeng, TANG Zhenhua.

2013, 28(9):  796-800.  DOI: 10.3969/j.issn.1673-8640.2013.09.015

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Objective To analyze the laboratory determination results of hand-foot-mouth disease to reveal the enterovirus infection and the results of routine determination items in children with hand-foot-mouth disease at Fuyang, and to provide the reference for diagnosis, treatment, prevention and control. Methods Both throat swab and vesicle fluid were collected from 497 children with hand-foot-mouth disease (severe case: 102 cases and common case: 395 cases). Specific RNA of enterovirus primer (EV), enterovirus 71 (EV71) and coxsackievirus A16(CA16)were detected by real-time fluorescence polymerase chain reaction (RT-PCR). Simultaneously, white blood cell (WBC),C reactive protein(CRP),creatine kinase MB isoenzyme(CK-MB), aspartate aminotransferase (AST),lactic dehydrogenase (LDH) and glucose (Glu) were determined, and the results were analyzed. Results Among the 497 children, 339 cases were positive for EV, 286 cases were positive for EV71, and only 22 cases were positive for CA16.The positive rates were 68.21%, 57.55% and 4.43%, respectively. In the 102 hand-foot-mouth disease severe cases,EV71 positive rate was the highest(95 cases, 93.14%). There was no statistical significance in EV, EV71 and CA16 positive rates between boys and girls (P> 0.05).The age distribution was mainly in the group under 3-year-old (398 cases, 80.08%). In the group under 3-year-old, there were 96 severe cases (94.12%), and the EV71 positive rate was the highest (94.79%, 91/96). The increasing cases of WBC was 80 cases (78.43%),CRP 81 cases (79.41%),Glu 52 cases (50.98%) and AST 64 cases (62.75%) in the severe cases of hand-foot-mouth disease. In the common cases, the increasing cases of WBC was 70 cases (17.72%),CRP 87 cases (22.03%),Glu 26 cases (6.58%) and AST 78 cases (19.75%). The positive rates in the severe cases were higher than those in the common cases (P<0.01). The levels of CK-MB and LDH increased in the common and severe cases of hand-foot-mouth disease,and the positive rates of CK-MB and LDH were obviously higher than the other routine determination items. Conclusions The hand-foot-mouth disease appeared in Fuyang in 2011 is mainly related to the EV71 infection, and EV71 is the main pathogen causing the severe cases of hand-foot-mouth disease. The monitoring should be strengthened especially in the group under 3-year-old. The severe cases should be early recognized in order to reduce the occurrence of the severe cases and the death.

Analysis on the results of five trace elements in whole blood of children in Panyu district，Guangzhou city

XIAO Xing,HE Jinhua,LI Yuguang, GAO Hui, HU Shufen.

2013, 28(9):  801-804.  DOI: 10.3969/j.issn.1673-8640.2013.09.016

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Objective　To provide the reference for the reasonable supplement and its study of trace element levels by investigating the distribution and variation regularity of Cu, Ca, Mg, Zn and Fe of children in Panyu district, Guangzhou city. Methods　Five trace elements(Cu, Ca, Mg, Zn and Fe) in whole blood of 3 476 children were detected by atomic absorption spectrometer. The children were classified into 5 age groups：infancy group (112 months old, 743 cases), toddler age group (>12 years old, 916 cases), preschool age group (36 years old, 988 cases)，school age group(711 years old, 579 cases) and adolescence group(1218 years old, 250 cases). Based on sex, each group was subclassified into female and male groups. The levels of the 5 trace elements were analyzed among different sex and age groups. Results　The abnormal rates of Cu and Mg were low. The total deficiency rates were 1.5% and 0.26%，and the total exceeding standard rates were 0.08% and 0.06%.It was common for the deficiency of Zn and Fe, and their total deficiency rates was 21.0% and 19.7%，and with the increasing of age, their deficiency rates were downward. The total exceeding standard rates of Zn and Fe were 1.8% and 0%.The total deficiency rate of Ca was 6.7%，and with the increasing of age, the deficiency rate was upward. The total exceeding standard rate of Ca was 1.9%. In preschool age and adolescence groups, the levels of Mg and Fe had statistical significance between males and females （t＝2.563, 2.172, 2.822 and 4.537, P<0.01）. The levels of the other trace elements had no statistical significance between males and females in each age group（P>0.05）. The levels of Zn and Fe were upward with the increasing of age, and the levels of Cu and Ca were downward. The level of Mg was stable. Conclusions　The abnormal rates of Zn, Fe and Ca in whole blood of children in Panyu district are high. The distributions of Cu and Mg become reasonable. The clinical guidance and research of Zn，Fe and Ca should be strengthened.

Study on the correlationship of ferritin concentration with insulin resistance in patients with type 2 diabetes mellitus

XU Jing,HU Xiaobo.

2013, 28(9):  805-808.  DOI: 10.3969/j.issn.1673-8640.2013.09.017

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Objective To investigate the relationship of serum ferritin (SF) with type 2 diabetes mellitus (T2DM) in Chinese Han population with different ages. Methods A total of 100 patients with T2DM (T2DM group),100 patients with high ferritin and non-T2DM (high SF group) and 100 healthy subject controls (control group) were enrolled for the determinations of fasting blood glucose (FBG),fasting insulin (FINS),SF and glycosylated hemoglobin (HbA_1c) and blood parameters [hemoglobin (Hb),hematocrit (Hct),mean corpuscular volume (MCV) and mean cell hemoglobin (MCH)]. The correlationship was analyzed. Results The SF,FBG,HbA_1c,FINS and homeostasis model assessment of insulin resistance (HOMR-IR) in T2DM group were significantly higher than those in control group (P<0.05). SF had a significant correlation with FBG ,HbA_1c ,FINS and HOMR-IR (r＝0.140, 0.171, 0.189 and 0.316,P=0.003,0.023,0.004 and 0.001). The Hb, FBG, HbA_1C, FINS and HOMA-IR in T2DM group were significantly higher than those in high SF group (P<0.05). The SF and MCV in high SF group were significantly higher than those in control group (P<0.05). Conclusions Iron overload leads to the increasing of SF, which is correlated with insulin resistance in patients with T2DM.

Clinical significance of the determinations of serum MMP-13 and TGF-β1 in chronic liver disease

FAN Yongxi,WAN Furong

2013, 28(9):  809-810.  DOI: 10.3969/j.issn.1673-8640.2013.09.018

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Objective To determine the matrix metalloproteinase-13(MMP-13) and transform growth factor beta 1 (TGF-β1) in patients with chronic liver disease, and to analyze the correlation and clinical significance. Methods A total of 50 chronic hepatitis patients, 50 cirrhosis patients, 60 liver carcinoma patients and 50 healthy subjects (healthy control group) were enrolled and determined for MMP-13 and TGF-β1 by enzyme-linked immunosorbent assay (ELISA). Results The MMP-13 in the chronic hepatitis patients, cirrhosis patients, liver carcinoma patients and healthy controls were 287.05±29.80, 328.04±62.03, 329.02±70.04 and (279.03±32.80) ng/L. The TGF-β1 in the chronic hepatitis patients, cirrhosis patients, liver carcinoma patients and healthy controls were 0.99±0.11, 0.96±0.12, 2.22±1.08 and (0.65±0.09) μg/L. The expressions of MMP-13 and TGF-β1 in patients with chronic hepatitis, cirrhosis and liver carcinoma were higher than those in healthy controls. The serum MMP-13 in cirrhosis and liver carcinoma patients were much higher than those in chronic hepatitis patients (P<0.01). The TGF-β1 in liver carcinoma patients was higher than those in cirrhosis and chronic hepatitis patients (P<0.01). In comparison to healthy controls, the TGF-β1 in the cirrhosis and chronic hepatitis patients had no statistical significance (P>0.05). Conclusions In the development of chronic liver diseases, the MMP-13 and TGF-β1 express excessively. The dynamic analysis of their levels may be helpful to the diagnosis and monitoring of chronic disease.

Analysis on the change of CD4^＋CD25^＋Foxp3^＋ regulatory T cells in peripheral blood of patients with chronic hepatitis B

QING Keqin,YANG Chaoguo,LIU Rong

2013, 28(9):  811-814.  DOI: 10.3969/j.issn.1673-8640.2013.09.019

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Objective To investigate the frequency of CD4⁺CD25⁺Foxp3⁺ regulatory T cells (Treg)in peripheral blood of patients with chronic hepatitis B (CHB)and its significance in the hepatitis B virus (HBV)infection. Methods The frequencies of CD4⁺CD25⁺Foxp3⁺Treg in peripheral blood from 86 CHB patients,30 acute hepatitis B (AHB)patients and 30 healthy subjects were determined by flow cytometry.The correlations of HBV DNA loads with CD4⁺CD25⁺Foxp3⁺ Treg frequency and the frequency of CD4⁺CD25⁺Foxp3⁺ Treg after therapy with different therapeutic effects were analyzed respectively.CD4⁺CD25⁺Foxp3⁺ Treg frequencies among mild,moderate and severe active CHB patients were compared respectively. Results The percentages of CD4⁺CD25⁺Foxp3⁺ Treg in CHB patients,AHB patients and healthy subjects were 2.56%±1.12%,1.39%±0.68% and 1.45%±0.76%, respectively.The percentage in CHB patients was higher than those in AHB patients and healthy subjects (P<0.05),and there was no statistical significance between the percentages of AHB patients and healthy subjects (P>0.05).The percentages of CD4⁺CD25⁺Foxp3⁺ Treg among mild,moderate and severe active CHB patients had no statistical significance (P＞0.05).The levels of alanine aminotransferase (ALT)in CHB patients,AHB patients and healthy subjects were 285.6±168.3,(780.9±432.6) and (23.2±6.7) U/L, respectively.The levels of ALT in CHB patients and AHB patients were higher than that in healthy subjects(P<0.05).The level of ALT in CHB patients was lower than that in AHB patients(P<0.05).The frequency of CD4⁺CD25⁺Foxp3⁺ Treg was not correlated with the level of ALT (r=0.11,P>0.05).The HBV DNA loads in CHB patients,AHB patients and healthy subjects were (5.2±1.8)log₁₀copies/mL,(7.2±1.8)log₁₀copies/mL and negative respectively.The HBV DNA load in AHB patients was higher than that in CHB patients(P<0.05).There was a positive correlation between CD4⁺CD25⁺Foxp3⁺ Treg frequency and HBV DNA logarithm(r＝0.30,P＜0.05).CD4⁺CD25⁺Foxp3⁺ Treg frequency decreased obviously after anti-viral therapy in the patients with obvious good therapeutic effect,and it was lower than that in the patients with poor therapeutic effect (P<0.05). Conclusions The increase of CD4⁺CD25⁺Foxp3⁺ Treg in CHB patients plays an active role in chronic HBV.The regulation and control of function and amount of CD4⁺CD25⁺Foxp3⁺ Treg may provide a new therapy for CHB.

Analysis and significance of T lymphocyte subsets in peripheral blood of chronic hepatitis B patients with perC region 1896 mutation

MAO Huijun, QIU Jianping, LIN Feng, WANG Gang, ZHANG Min, CHEN Daihong

2013, 28(9):  815-819.  DOI: 10.3969/j.issn.1673-8640.2013.09.020

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Objective To investigate the relationship of hepatitis B virus (HBV) preC region 1896 mutation with peripheral blood T lymphocyte subsets. Methods A total of 196 patients with hepatitis B were classified into 146 cases of chronic hepatitis B (CHB) group and 50 cases of hepatitis B liver cirrhosis (LC) group. CHB group was classified into 80 cases of wild-type strain CHB group (CHB Ⅰ group) and 66 cases of preC region 1896 mutation CHB group (CHB Ⅱ group). LC group was classified into 19 cases of wild-type strain LC group (LC Ⅰ group) and 31 cases of preC region 1896 mutation LC group (LC Ⅱ group). In 196 patients with hepatitis B, the specific primers of polymerase chain reaction (PCR) was used to detect the HBV preC region 1896 mutation, and the T lymphocyte subsets in peripheral blood were detected by flow cytometry. A total of 20 healthy subjects were enrolled as healthy control group. Results In 196 hepatitis B patients, the rate of HBV preC 1896 mutation was 49.5%. The mutation rate in LC group (62.0%) was higher than that in CHB group (45.2%). Compared with the healthy control group, the CD4⁺T cell absolute number of CHB group decreased significantly (P<0.01). In LC group, the absolute number and percentage of CD3⁺T cell, the absolute number of CD8⁺T cell and the absolute number and percentage of CD4⁺T cell were significantly lower than those in the healthy control group (P<0.01). Compared with CHB Ⅰ group, the CD3⁺T cell absolute number and CD4⁺T cell absolute number of CHB Ⅱ group decreased significantly (P<0.05). Compared with CHB Ⅱ group,the absolute number of CD3⁺T cell and CD4⁺T cell in LCⅡgroup had significant decrease (P<0.01). Conclusions In CHB patients with HBV preC 1896 mutation, the imbalance of T lymphocyte subsets is more serious than that of CHB patients with wild-type strain. The imbalance may participate in the pathogenesis of chronic hepatitis caused by the HBV preC region 1896 mutation.

The application of the determination of urinary albumin by HPLC in patients with diabetes mellitus

SHI Mei,SHI Weifeng,LI Yanmei.

2013, 28(9):  820-823.  DOI: 10.3969/j.issn.1673-8640.2013.09.021

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Objective To investigate the application of urinary albumin determination by high performance liquid chromatography (HPLC) in patients with diabetes mellitus (DM). Methods Urine samples of 85 DM patients and 40 healthy subjects (healthy control group) were collected, and the urinary albumin concentrations were determined by HPLC and immunoturbidimetry respectively. The consistency of the results between the 2 methods was analyzed by the regression analysis and Bland-Altman analysis. Results HPLC linear regression equation was Y=2.948 2X+38.601,r²=0.994 4.The retention time (RT) of standard materials and samples were 2.42 min. Within-run coefficient of variation (CV) was 4.1%, and between-run CV was 5.6% at concentration of 40.1mg/L. Within-run CV was 4.8%, and between-run CV was 6.5% at concentration of 152.6 mg/L. The lowest detection limit was 2 mg/L. The urinary albumin concentration was significantly higher by HPLC [21.0(3.4-678.9)mg/L] than by immunoturbidimetry [8.2(2.0-442.2)mg/L] by 1.3-6.1 folds (P<0.01). There was no statistical significance between the concentrations of control group by HPLC and immunoturbidimetry(P>0.05). The regression analysis and Bland-Altman analysis demonstrated that the 2 methods were not consistent. Of all the 85 DM patients, according to microalbuminuria diagnosis standard [urinary albumin excretion rate (AER) >30 mg/24h], there was 48(56.5%) positive cases by HPLC and 27(31.8%) positive cases by immunoturbidimetry. Conclusions Immunity and no-immunity total urinary albumin can be determined by HPLC. Microalbuminuria in DM patients can be determined earlier by HPLC.

Study on the correlation between polyethylene glycol precipitation treatment and gel filtration chromatography in the detection of macroprolactin

WANG Xia, LIU Jinling,GAO Shuo

2013, 28(9):  824-827.  DOI: 10.3969/j.issn.1673-8640.2013.09.022

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Objective To investigate the feasibility of polyethylene glycol(PEG)precipitation treatment in the detection of macroprolactine(MPRL). Methods A total of 26 hyperprolactinemia (HPRL) patients were classified into 2 groups: genuine HPRL group (11 cases) and macroprolactinemia group (15 cases) according to the clinical and imaging data. By gel filtration chromatography (GFC), the components of prolactin (PRL) were separated, and their levels were determined. After PEG precipitation treatment, the concentration of monomer PRL was determined, the results were compared with those by GFC, and the correlation was analyzed. Results The chromatogram of macroprolactinemia group was different from that of genuine HPRL group. The genuine HPRL group was consisted of monomer PRL, but the macroprolactinemia group was consisted of big PRL. The 2 groups′ PRL before the treatment had no statistical significance (P>0.05).After the treatment,the PRL concentrations of genuine HPRL group were high than those of macroprolactinemia group (P<0.05). The 2 groups′ monomer PRL concentrations before PEG precipitation treatment and GFC had no statistical significance (P>0.05), and the correlation was good [correlation coefficient (r)=0.844, P<0.05]. Conclusions PEG precipitation treatment for the detection of MRPL has good correlation with GFC, and it is economical and convenient. It is recommended as the routine screening method.

A clinical consensus study of HBsAg quantitative results by two methods

SHENG Huan,HU Xiaobo,XU Jie

2013, 28(9):  828-834.  DOI: 10.3969/j.issn.1673-8640.2013.09.023

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Objective To evaluate the clinical application feasibility of hepatitis B surface antigen (HBsAg) quantitative determination by Roche and Abbott. Methods A total of 423 serum and plasma samples were collected and determined by Abbott ARCHITECT HBsAg and Roche HBsAg quantitative Elecsys, and the results were analyzed comparatively. The correlation was analyzed statistically. Performance evaluation on Roche and Abbott was also conducted. Results The intra-analysis and inter-analysis coefficients of variation (CV) were low. The HBsAg results of serum and plasma samples from the same patient correlated well (r=0.988, P<0.05). The correlation between Roche and Abbott was good (r=0.965,P<0.05). The 2 assays were not influenced by potential interferences such as lipemia, icterus and hemolysis. The limit of the 2 assays was 0.05 IU/mL. Through dilution, the 2 assays could report the common clinical high-value HBsAg results. The recoveries of the 2 assays were consistent. Conclusions Results from Roche and Abbott HBsAg quantitative results are consistent and have a good correlation, and they are suitable for the clinical application. HBsAg results are consensus for each other.

Establishment and performance evaluations of dot immuno-gold filtration assay for the quantitative determination of serum amyloid A

CHEN Changqiang,YANG Jing,GU Zhidong,WAN Yinglei,MAO Kezi,FENG Xiaojing,XU Jianxin,FAN Qishi.

2013, 28(9):  835-840.  DOI: 10.3969/j.issn.1673-8640.2013.09.024

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Objective To establish a dot immuno-gold filtration assay (DIGFA) for the quantitative determination of serum amyloid A (SAA),and evaluate its analysis performance. Methods Serum DIGFA-based SAA detection reagent (SAA-DOT) was established using 2 anti-SAA monoclonal antibodies being coated with nitrocellulose filter and labeled by colloidal gold, respectively. According to EP files, the SAA-DOT analysis performances (such as detection limit,precision,accuracy,linear range,et al) were evaluated. Results The detection limit of SAA-DOT was 5mg/L. The within-run coefficients of variation (CV) were 9.07% and 10.21% respectively, whereas the inter-day CV were 11.33% and 11.53% respectively, when serum SAA concentrations were 10 and 100 mg/L. The detection linear range was from 5 mg/L to 230 mg/L. SAA-DOT had a good correlationship with latex-enhanced rate nephelometry (LERN) reagent (r=0.995,P<0.05). The median of serum SAA was 5.7 mg/L, and the 95% confidence interval was 0-9.6 mg/L in healthy subjects. Conclusions The DIGFA-based SAA-DOT is convenient, and the reaction can be finished within 5 min. Its performance meets the State Food and Drug Administrration (SFDA) requirements for in vitro diagnostic reagents,which is suitable for the clinical applifcation of serum detection.

Effects of vasoactive intestinal peptide to the activation of GFAP after focal cerebral ischemia-reperfusion in rats

LIU Xu.

2013, 28(9):  841-844.  DOI: 10.3969/j.issn.1673-8640.2013.09.025

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Objective To investigate the effects of vasoactive intestinal peptide(VIP) on the dynamic changes of glial fibrillary acidic protein(GFAP) after focal cerebral ischemia-reperfusion in vivo and the proliferation of astrocyte, and discuss the neuroprotective effects of VIP. Methods In rats, VIP was given via injection, and middle cerebral artery occlusion (MCAO) model was set up by suture method. The evaluation and screening of rats with successful occlusion of MCAO were performed by neurological testing. The expression of GFAP was detected by immunohistochemistry. Results Compared with the control groups after VIP injection, GFAP expression significantly increased(P<0.05). The existing astrocyte response in the same ischemic area was upregulated. Conclusions VIP injection enhances the expression of GFAP and reduces the degree of cerebral ischemia-reperfusion injury. This study provides the evidence for the neuroprotective effects of VIP in vivo.

Investigation on the apoptosis mechanisms of liver cancer HepG₂ cells induced by Apollon antisense oligodeoxynucleotide

LI Yuguang,HE Jinhua, WANG Li, XIE Xingyi,CHEN Shunyi, CHEN Wujia,HUANG Guoxian.

2013, 28(9):  845-850.  DOI: 10.3969/j.issn.1673-8640.2013.09.027

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Objective To study the influence of inhibitor apoptosis protein (IAP) Apollon antisense oligodeoxynucleotide (ASO) on the proliferation and apoptosis on human liver cancer HepG₂ cells, and investigate the mechanisms. Methods Apollon ASO was incubated by liposome with human liver cancer HepG₂ cell for 48 h, and the proliferation inhibition was detected by WST-8. The expression of Apollon mRNA was detected by real-time fluorescence quantitative reverse transcription polymerase chain reaction (PCR). The early apoptosis rates were determined by flow cytometry. The morphology of HepG₂ cell apoptosis was determined by Hochest 33258 staining. The cell apoptosis was determined by mitochondrial membrane potential. The activities of cysteine proteinases (Caspase-3 and Caspase-9) were determined by colorimetric method. The expression of cytochrome C was analyzed by Western blot. Results After being treated by Apollon ASO on HepG₂ cells for 48 h, it was found that Apollon ASO could all suppress the proliferation of HepG₂ cells, which was depended on the concentration. Apollon ASO could induce HepG₂ cell apoptosis. The different concentrations of Apollon ASO reduced significantly the expression level of Apollon mRNA. The activities of Caspase-3 and Caspase-9 and the protein expression levels of cytochrome C increased. Through observing by fluorescence microscopy assay, nucelus high fluorescence cells were found, and the apoptosis cytoryctes were also observed. Conclusions Apollon ASO can reduce the expression level of Apollon mRNA and inhibit the proliferation of liver cancer HepG₂ cells, and it can induce the apoptosis by the way of mitochondria mediation.