Table of Content

30 June 2015, Volume 30 Issue 6

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Talk about hemoglobin A_1c

JU Yi

2015, 30(6):  547-552.  DOI: 10.3969/j.issn.1673-8640.2015.06.001

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Hemoglobin A_1c(HbA_1c)is confirmed by the 2 epidemiological surveys, the UK Prospective Diabetes Study(UKPDS)and the Diabetes Control and Complications Trial Research(DCCT), as an important marker which is closely related to diabetes and its complications. HbA_1c has been used more and more widely in clinical laboratories. Due to the diversity of HbA_1c detection technologies, there is large difference among results, and thus it is unable to meet the demand of clinical diagnosis and treatment. Along with the International Federation of Clinical Chemistry and Laboratory Medicine(IFCC)and American Association for Clinical Chemistry(AACC)to carry out the HbA_1c standardization, testing quality in laboratories has been significantly improved. Since 2010, HbA_1c has been included in the diagnostic criteria for diabetes by American Diabetes Association(ADA), the International Diabetes Federation(IDF) and World Health Organization(WHO). In China, HbA_1c detection begins relatively late in clinical application,quality management and standardization, but the rate of progress of work and performance are very obvious. Although HbA_1c has temporarily not been included in the Chinese diabetes diagnosis guide, we believe that as long as we keep the unity and cooperation, we will create standardization in our own way and serve the clinicians and patients with diabetes mellitus better.

The variation and significance of IL-17 and IL-23 in adult patients with primary nephrotic syndrome

ZHOU Yunjiao, ZHANG Lihong, LI Peng, YE Xianwu, NIU Jianying

2015, 30(6):  554-558.  DOI: 10.3969/j.issn.1673-8640.2015.05.002

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Objective　To investigate the changes of interleukin 17 (IL-17) and interleukin 23 (IL-23) in adult patients with primary nephrotic syndrome (PNS) and their roles and clinical significance of IL-17 and IL-23 variations in the occurrence and development of PNS. Methods　A total of 35 steroid-effective hospitalized patients with PNS and 20 healthy subjects (healthy control group) were enrolled in the study. The patients with PNS were treated with prednisone according to the conventional dose. T cell subsets from peripheral blood and the levels of IL-17,IL-23 and IL-6 from serum and 24 h urine were detected in healthy control group and the patients with PNS before and after treatment. The correlations of 24 h urinary protein quantification with T cell subsets in peripheral blood and the levels of IL-17,IL-23 and IL-6 in both serum and 24 h urine were analyzed,respectively. Results　Compared with healthy control group, the percentage of CD3⁺ or CD3⁺CD4⁺ cells from patients with PNS significantly decreased (P<0.05), and the levels of IL-17 and IL-6 from serum as well as IL-17, IL-23 and IL-6 from 24 h urine significantly increased(P<0.05). After treatment, the percentage of CD3⁺ or CD3⁺CD4⁺ cells significantly increased (P<0.05), and the levels of IL-17, IL-23 and IL-6 in 24 h urine significantly decreased (P<0.05). There were positive correlations between 24 h urinary protein quantification and the levels of IL-17 and IL-23 in both serum and 24 h urine respectively (P<0.01). Conclusions　IL-17 and IL-23 may contribute to the proteinuria of PNS. Hormone therapy can improve the disorder of cell immune function and the imbalance of T cell subsets. Monitoring the levels of IL-17 and IL-23 in serum and 24 h urine may be helpful for disease judgement and clinical therapy.

Evaluation on the diagnostic significance of DKK1, GP73 and AFP in primary hepatic carcinoma by Logisitic regression analysis and ROC curve analysis

ZONG Yingying, XU Hao, XU Wei, GU Yuming, ZU Maoheng, WAN Haoguang, WU Kang

2015, 30(6):  559-563.  DOI: 10.3969/j.issn.1673-8640.2015.06.003

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To investigate the diagnostic significance of serum Dickkopf-1 (DKK1), Golgi protein 73 (GP73) and alpha fetoprotein (AFP) in primary hepatic carcinoma (PHC) by Logistic regression analysis and receiver operating characteristic (ROC) curve analysis.

Enzyme-linked immunosorbent assay (ELISA) and electrochemiluminescence immunoassay (ECLIA) were used to determine the serum levels of DKK1,GP73 and AFP in 81 patients with PHC, 50 patients with chronic hepatitis (CH), 50 patients with liver cirrhosis (LC) and 50 healthy controls(HC). A statistical model for diagnosing PHC was established, and the results were analyzed by Logisitic regression analysis and ROC curve analysis.

Serum DKK1 levels in PHC group [5.29(4.27-7.45)ng/mL]were significantly higher than those in CH group[4.11(3.82-4.52)ng/mL], LC group [3.66(3.61-4.15)ng/mL]and HC group[3.42(3.28-3.68)ng/mL](P<0.01). Serum GP73 levels in PHC group[(215.44±68.66) ng/mL]were significantly higher than those in CH group [(121.02±49.10)ng/mL], LC group[(168.39±37.91)ng/mL]and HC group[(57.36±19.94)ng/mL](P<0.05). The sensitivities and specificities of DKK1, GP73 and AFP were 71.6%, 77.8%, 58.0% and 90.0%, 86.7%, 78.7%, respectively. The model had a sensitivity of 87.7% and a specificity of 90.0%. The areas under ROC curves (AUC) of AFP, DKK1 and GP73 were 0.76, 0.86 and 0.87, respectively, and the AUC of the model was 0.93, having statistical significance with AFP, DKK1 and GP73 (P<0.01).

DKK1 and GP73 can be as serum markers for the diagnosis of PHC, and the combined determination of DKK1, GP73 and AFP could improve PHC diagnostic performance. Moreover, Logisitic regression analysis combined with ROC curve analysis is a simple and effective statistical method, and it can be used in the analysis and evaluation of multi-index combined diagnosis of PHC.

Changes of serum concentrations of soluble Fas and soluble Fas ligand in patients with different degree Alzheimer disease

CHEN Tongwei

2015, 30(6):  564-566.  DOI: 10.3969/j.issn.1673-8640.2015.06.004

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To investigate the changes of serum concentrations of soluble Fas(sFas) and sFas ligand(sFasL) in patients with different degree Alzheimer disease (AD).

A total of 100 AD patients, 54 patients with old-age diseases (17 cases of diabetes mellitus, 18 cases of hypertension, 15 cases of hyperlipemia and 4 cases of other endocrine diseases) and 46 healthy subjects were enrolled as old-age disease control group and healthy control group.Serum concentrations of sFas and sFasL were detected by enzyme-linked immuosorbent assay(ELISA). The AD patients' cognitive functions were evaluated by Clinical Dementia Rating Scale (CDR), mini-mental state examination(MMSE) and Hachinski ischemia score(HIS).

The levels of serum sFas and sFasL were significantly higher in AD group than in 2 control groups (P<0.01). As the disease aggravated, the serum levels of sFas and sFasL increased gradually. Serum sFas level was positively correlated with age, course of disease and the scores of MMSE, while it was negatively with the scores of HIS (r=0.856, 0.834, 0.799 and -0.714, P<0.01). Serum sFasL level was positively correlated with age and course of disease, while it was negatively with the scores of MMSE and HIS (r=0.863, 0.857, -0.801 and -0.745, P<0.01).

The abnormal levels of sFas and sFasL among AD patients may indicate being involved in the apoptosis pathological change, and the serum levels of sFas and sFasL might be related with the intelligent level and severity of cerebral ischemia.

Comparison on in vitro antibacterial activities for five kinds of trditional Chinese medicine granules and decoctions against five kinds of clinical drug-resistant strains

LI Aiwen, TAN Junqing, WANG Kangchun, HUANG Shuangwang, ZHOU Bing, LIU Qibo

2015, 30(6):  567-570.  DOI: 10.3969/j.issn.1673-8640.2015.06.005

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To compare in vitro antibacterial activities for 5 kinds of traditional Chinese medicine granules and decoctions (Baicalin, Forsythia, Mint, Dandelion and Sophora) against 5 kinds of clinical drug-resistant strains.

According to the related researches on traditional Chinese medicine against resistant bacteria, 5 kinds of traditional Chinese medicine granules and decoctions were selected, and 5 kinds of clinical drug-resistant strains from blood plate were collected. They included meticillin-resistant Staphylococcus aureus (MRSA), Pseudomonas aeruginosa, Escherichia coli, Klebsiella pneumoniae and Acinetobacter baumannii. The disc agar diffusion method was used, and the diameter of bacteriostatic ring was measured. The bacteriostatic effects were analyzed statistically.

Five traditional Chinese medicines showed different antibacterial activities, with Baicalin being the most potent. The same kind of traditional Chinese medicine granule and decoction was compared. For MRSA, Baicalin, Forsythia, Mint, Dandelion and Sophora had antibacterial effects, and there was no statistical significance (P>0.05). For Pseudomonas aeruginosa and Acinetobacter baumannii, Baicalin, Forsythia and Mint had antibacterial effects, and there was no statistical significance (P>0.05). For Escherichia coli and Klebsiella pneumoniae, Baicalin had antibacterial effects, and there was no statistical significance (P>0.05).

The same concentrations of Baicalin, Forsythia, Mint, Dandelion and Sophora decoctions have not obvious difference for antibacterial effect.

Prognosis prediction of thrombelastography in patients undergoing percutaneous coronary intervention

GU Meixiu, SUN Lin, WANG Beili, REN Xiaoci, YAO Ke, GUO Wei, PAN Baishen

2015, 30(6):  571-574.  DOI: 10.3969/j.issn.1673-8640.2015.06.006

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To evaluate the prognosis prediction of thrombelastography (TEG) in patients undergoing percutaneous coronary intervention (PCI).

The data from 115 patients undergoing PCI were collected, and the patients were classified into 2 groups according to maximum amplitude-adenosine diphosphate (MA-ADP), MA-MDP<50 mm group and MA-ADP≥50 mm group. The percentages of adverse cardiovascular events, reinfarction and restenosis were followed up for 1 year.

There was no statistical significance in the clinical characteristics of the 2 groups (P>0.05). Logistic regression analysis showed that MA-ADP was the risk factor of adverse cardiovascular event recurrence in patients undergoing PCI (OR=3.547, 95% CI: 1.103-11.412, P=0.034). Compared with MA-ADP normal patients (<50mm), the incidences of adverse cardiovascular events and reinfarction in patients with high MA-ADP (≥50mm) significantly increased (P<0.05).

MA-ADP can predict the prognosis after PCI. MA-ADP=50 can be the cut-off point in the prognosis prediction of patients undergoing PCI.

Analysis on the correlations of 25-hydroxyvitamin D concentrations with HCV RNA loads and cirrhosis severity in hepatitis C patients

SONG Binbin, ZHOU Yan, WU Wenhao, WU Jiong, ZHANG Chunyan, WANG Beili, GUO Wei, PAN Baishen

2015, 30(6):  575-580.  DOI: 10.3969/j.issn.1673-8640.2015.05.007

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Objective　To investigate the correlations of serum 25-hydroxyvitamin D[25（OH）D]concentrations with hepatitis C virus（HCV）RNA loads and cirrhosis severity in hepatitis C patients. Methods　A total of 271 hepatitis C patients and 218 healthy subjects were enrolled， and their serum samples were collected. Serum 25（OH）D concentrations and HCV RNA loads were determined. Moreover， serum alpha 2-macroglobulin（α2M）， haptoglobin（HPT）， apolipoprotein A1（apo A1）， total bilirubin（TBil）and gamma-glutamyl transpeptidase（GGT）concentrations were determined. According to Fibrotest classification， the severity of cirrhosis was evaluated. The 25（OH）D concentrations between hepatitis C and healthy control groups were compared. Furthermore， the difference of serum 25（OH）D between HCV RNA positive group and HCV RNA negative group was also evaluated. The correlation between serum 25（OH）D concentrations and HCV RNA loads in HCV RNA positive group was analyzed. Hepatitis C patients were classified according to the quartile concentrations of 25（OH）D， and the correlation between serum 25（OH）D concentrations and Fibrotest classification was investigated. Results　Serum 25（OH）D concentrations were significantly higher in healthy control group than in hepatitis C group[means were（60.42±1.34） nmol/L and（49.31±1.39） nmol/L， P＜0.01]. The proportions of patients with the deficiency of 25（OH）D（＜50 nmol/L）and the ones with severe deficiency（＜25 nmol/L） in hepatitis C group were 41.33%（112/271）and 14.40%（39/271）， which were significantly higher than those in healthy control group[27.98%（61/218） and 3.67%（8/218）]（P＜0.01）. No significant difference for serum 25（OH）D concentrations was observed between HCV RNA positive and HCV RNA negative groups[means were 48.98±21.59 and（49.89±25.27） nmol/L， P＞0.05]. Linearity regression revealed that there was no correlation between the logarithmic of HCV RNA loads and the concentration of 25（OH）D in HCV RNA positive group（r=0.056， P=0.412）. Moreover， there was no statistical significance for the proportion of patients with different Fibrotest classifications（F0-F1， F1-F3 and F3-F4）between the highest quartile concentration and the lowest quartile concentration of 25（OH）D [means were 49.12±5.80， 48.27±4.90 and（49.89±8.20） nmol/L， P＞0.05]. Conclusions　The hepatitis C patients are prone to encounter 25（OH）D deficiency compared with healthy subjects， which implys that these patients are suggested to uptake vitamin D supply. Furthermore， no clinical correlation is found between the logarithmic of HCV RNA loads， cirrhosis severity and the concentrations of 25（OH）D.

Study on the application significance of ischemia modified albumin in early diagnosis of acute coronary syndrome

ZHANG Ping, ZHOU Hongxing, BAI Yang, XU Zhe, SUN Yingxin, WANG Xin, WANG Sujian, ZHANG Hong

2015, 30(6):  581-584.  DOI: 10.3969/j.issn.1673-8640.2015.06.008

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To observe the changes of ischemia modified albumin(IMA) in patients with acute coronary syndrome(ACS), and investigate the application significance of IMA in early diagnosis of ACS.

The serum concentrations of IMA, MB isoenzyme of creatine kinase MB (CK-MB) and cardiac troponin T(cTnT) were continuously monitored at 2, 6, 12, 24 and 48h after onset in ACS patients with acute chest pain, including 43 cases of acute myocardial infarction (AMI) and 54 cases of unstable angina pectoris (UAP), and were compared with 42 suspected cases of ACS (control group). Their applications for the early diagnosis of ACS were evaluated.

IMA was higher in patients with UAP and AMI at 2 h after chest pain onset than healthy control group(P<0.05). The serum IMA reached a peak level at 12 h after the onset of chest pain and dropped to the same level as that of control group at 24 h. CK-MB and cTnT had no statistical significance in any determination time among the UAP patients. Receiver operating characteristic (ROC) curves showed that the areas under the curve (AUC) of IMA, CK-MB and cTnT were 0.812, 0.781 and 0.648, respectively at 2 h after chest pain onset.

IMA may be a sensitive biomarker for the myocardial ischemia of ACS patients, which can be used into the early diagnosis of AMI.

Clinical significance of detecting serum adiponectin in patients with acute myocardial infarction

JIANG Xiaohong

2015, 30(6):  585-587.  DOI: 10.3969/j.issn.1673-8640.2015.06.009

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To investigate the clinical significance of detecting serum adiponectin (APN) in patients with acute myocardial infarction (AMI).

A total of 40 patients with AMI, 40 patients with unstable angina pectoris (UAP), 40 patients with stable angina pectoris (SAP) and 40 healthy subjects were enrolled in this study. Serum high-sensitivity C reactive protein (hs-CRP), cardiac troponin I (cTnI) and APN levels were determined and compared in all patients and healthy subjects. The AMI patients were classified into high APN group and low APN group according to the median of APN levels, and the occurrence of major cardiovascular events were compared between the 2 groups.

Serum levels of APN decreased in turn among healthy subjects and patients with SAP, UAP and AMI. The levels of hs-CRP and cTnI sequentially increased in these subjects. Correlation analysis showed that APN in AMI patients was negatively correlated with hs-CRP and cTnI (r=-0.599 and -0.527, P<0.05). The major cardiovascular events in low APN group were significantly more than those in high APN group in the follow-up 12 months (P<0.05).

Serum APN levels in patients with AMI may be associated with the occurrence and development of AMI. Thus, the monitoring of serum APN levels in AMI patients has clinical significance for the diagnosis, treatment and prognosis of AMI.

The significance of urinary neutrophil gelatinase-associated lipocalin in the early diagnosis of diabetic nephropathy

CAI Leiming, LI Qian, TAN Longyi

2015, 30(6):  588-590.  DOI: 10.3969/j.issn.1673-8640.2015.06.010

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To investigate the significance of urinary neutrophil gelatinase-associated lipocalin (NGAL) in the diagnosis of diabetic nephropathy(DN).

A total of 120 diabetes mellitus patients were enrolled in the study. According to 24 h urine albumin excretion rate (UAER), the patients were classified into 3 groups, normal albuminuria (NA, 46 cases), microalbuminuria (MA, 39 cases) and clinical albuminuria (CA, 35 cases) groups. Another 50 healthy subjects were enrolled as normal control (NC) group. Serum NGAL concentrations were determined by enzyme-linked immunosorbent assay (ELISA). Serum creatinine (SCr) levels were determined by Jaffé reaction-rate method. Serum cystatin C (Cys C) concentrations were determined by latex-enhanced immune turbidity method. The results were analyzed comparatively.

Urine NGAL concentration in NA group was significantly higher than that in NC group(P<0.05). No statistical significance was observed in concentrations of serum Cys C and SCr between NA and NC groups (P>0.05). Urinary NGAL and serum Cys C levels in MA group were significantly higher than those in NC grpup (P<0.05). No statistical significance of SCr was found between MA and NC groups (P>0.05). Urinary NGAL, serum Cys C and SCr levels in CA group were all significantly higher than those in NC group(P<0.05).

Urinary NGAL may be a biomarker for early renal damage in diabetes mellitus.

Clinical significance of human papillomavirus genotyping by gene chip technology in Shanghai

MIAO Yingxin, GAN Jiemin, CHEN Jie, SHI Hong, JIANG Wenrong, ZHAO Hu

2015, 30(6):  595-598.  DOI: 10.3969/j.issn.1673-8640.2015.06.013

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To study the situation and genotype distribution of human papillomavirus(HPV) in Shanghai, and to investigate the relationships of HPV genotype, single-infection and multi-infection with the severity of cervical intraepithelial pathological changes.

Cervical epithelial cell samples were collected from 846 outpatients and healthy examination subjects and determined for HPV genotyping by gene chip technology and thin-prep cytology test(TCT). Biopsies under colposcopy were taken on subjects with abnormal cytology results.

The HPV positive rate was 29.55%, and the most were high-risk HPV single-infection. A total of 24 types of HPV were detected, and the most prevalence types were HPV52, 58, 16, 53, 51 and 55 in turn. The prevalences of HPV18, HPV68 and HPV59 were different between in single-infection and in multi-infection. There were 13 types of high-risk HPV, and only one low-risk HPV was detected in high-grade cervical intraepithelial neoplasia (CIN) [including cervical squamous cell carcinoma or adenocarcinomas, CIN 3 and CIN2, as ≥CIN2]. The prevalence of HPV16 ≥CIN2 was 46.88%, which was a far higher proportion than in healthy subjects and in low-grade cervical intraepithelial pathological changes (including atypical squamous cell of undetermined significance, normal and CIN1, as<CIN2) (9.63%, P<0.05).

HPV52, 58 and 16 are the most popular types in Shanghai. HPV16 is popular in ≥CIN2 group. HPV16 is the most important predictor of high-grade cervical intraepithelial pathological changes.

Evaluation on multiplex PCR for detecting the common serotypes of one kind of Streptococcus pneumoniae

PAN Fen, ZHAO Shengyuan, QIN Huihong, ZHU Junying, XIAO Shuzhen, ZHANG Hong

2015, 30(6):  599-602.  DOI: 10.3969/j.issn.1673-8640.2015.06.014

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To evaluate the performance of multiplex polymerase chain reaction (PCR) for detecting the common serotypes of one kind of Streptococcus pneumoniae.

A total of 50 isolates of Streptococcus pneumoniae were analyzed by multiplex PCR with traditional quellung reaction as reference method for detecting the common serotypes of Streptococcus pneumoniae.

The 50 isolates were typed by traditional quellung reaction with 9 isolates being 19F, 19A, 23F, 6A/B and 14, respectively, and 5 isolates being 15B/C. However, according to the results of multiplex PCR, 8 of 50 isolates were serotype 19F, and there also were 19A (10 isolates), 23F (9 isolates), 6A/B (9 isolates), 14 (10 isolates) and 15B/C (4 isolates), which showed coincidence rate 96%. The value of Kappa was 0.952.

Multiplex PCR has good consistency with traditional method, and it has the advantages of easy, rapid performance and low costing, which could provide the reference for monitoring the serotypes of Streptococcus pneumoniae in clinic.

Performance evaluation of Cobio Palio 300 urine biochemical analyzer for the determinations of 5 urinary micro-proteins

HUANG Yan, XU Xiaorong, ZHANG Jue

2015, 30(6):  603-606.  DOI: 10.3969/j.issn.1673-8640.2015.06.015

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To evaluate the performance of Cobio Palio 300 urine automatic biochemical analyzer (Palio 300) for the determinations of 5 kinds of urinary micro-proteins.

By Palio 300 and Siemens BN-prospec, 120 urine samples were determined for urinary micro-albumin (mAlb), α₁ microglobulin (α₁-MG), β₂ microglobulin (β₂-MG), transferrin (TRF) and immunoglobulin G (IgG). The results were compared and analyzed statistically. In addition, the medium-value and high-value mixed urine was used for these items. Each sample was determined repeatedly for 20 times, and the coefficient of variation (CV) was calculated.

Four items showed CV<6.00%, except β₂-MG CV=8.66%. The results were consistent with those of Siemens BN-prospec. The correlation coefficients (r) of 5 items were >0.9 (mAlb 0.953, α₁-MG 0.915, β₂-MG 0.927, TRF 0.918 and IgG 0.935, P<0.001).

Palio 300 has good repeatability, and its result is consistent with Siemens BN-prospec in the determinations of 5 urinary micro-proteins.

Research on devR regulated cytotoxicity of Mycobacterium tuberculosis

DANZHENG Jiancuo, MA Yueyun, ZHOU Lei, FU Xiaorui, SU Mingquan, HAO Xiaoke

2015, 30(6):  607-612.  DOI: 10.3969/j.issn.1673-8640.2015.06.016

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To identify the influence of devR expression on the growth and cytotoxicity of tuberculosis.

Culture wild Mycobacterium bovis Bacillus Calmette-Guérin (BCG), devR knockout BCG (ΔdevR) and devR compensated BCG (devR.com) were identified for devR sequence, and the growth curves at different times were drawn under absorbance (A) 600 nm. A549 cells and Raw264.7 cells were infected with the strains at multiplicity of infection (MOI)=10. The apoptosis ratio was detected by flow cytometry, and lactate dehydrogenase (LDH) released by host cells was measured by enzyme-linked immunosorbent assay(ELISA) at 0, 24, 48 and 72 h. The expressions of apoptosis related genes, bcl-2 and bad, were determined by real-time quantitation polymerase chain reaction (PCR).

Comparing to BCG and devR.com, the growth of ΔdevR was significantly accelerated at early phase (2-10 d, P<0.01). Apoptosis and necrosis ratio of A549 cells and Raw264.7 cells infected by ΔdevR were significantly higher than those infected by BCG and devR.com(P<0.05). All 3 strains could up-regulate the expressions of bcl-2 of A549 cells, and ΔdevR stimulated higher level of bcl-2 expression than BCG and devR.com. They also could increase the bad expression of Raw264.7 cells. ΔdevR induced more bad gene expression than BCG and devR.com.

The knockout of devR could enhance the cytotoxicity of BCG.

Studies of 16S rRNA methylase gene and aminoglycoside modifying enzyme gene in Pseudomonas aeruginosa

PENG Jinghong, HOU Yanqiang, XIE Duoshuang, LIU Jun, ZHENG Rong

2015, 30(6):  613-616.  DOI: 10.3969/j.issn.1673-8640.2015.06.017

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To investigate 16S rRNA methylase gene and aminoglycoside modifying enzyme gene in aminoglycoside-resistant clinical isolates of Pseudomonas aeruginosa, and to investigate the mechanism of drug resistance to aminoglycosides.

A total of 35 isolates of aminoglycoside-resistant clinical isolates of Pseudomonas aeruginosa were collected, and the identification and susceptibility testing in vitro were performed by VITEK 2 Compact automatic microbial analysis system. Five kinds of 16S rRNA methylase genes(arm-A, rmt-A, rmt-B, rmt-C and rmt-D) and 6 kinds of aminoglycoside modifying enzyme genes [aac(3)-Ⅰ, aac(3)-Ⅱ, aac (6')-Ⅰb, aac(6')-Ⅱ, ant (3″)-Ⅰand ant (2″)-Ⅰ] were detected by polymerase chain reaction (PCR).

Aminoglycoside-resistant Pseudomonas aeruginosa showed being multi-drug resistant to a variety of antibiotics. Only the resistance rate to imipenem was low (5.7%). There were 21 isolates of Pseudomonas aeruginosa carried rmt-B 16S rRNA methylase gene, accounting for 60.0%, and 30 isolates of Pseudomonas aeruginosa carried aminoglycoside modifying enzyme genes, accounting for 85.7%, among them, 28 isolates carried aac(3)-Ⅱ aminoglycoside modifying enzyme genes, accounting for 80.0%, and 18 isolates carried ant (3″)-Ⅰ aminoglycoside modifying enzyme genes, accounting for 51.4%. The other 8 isolates were not detected any genes.

Aminoglycoside-resistant isolates of Pseudomonas aeruginosa carry rmt-B 16S rRNA methylase gene and aac(3)-Ⅱ and ant (3″)-Ⅰ aminoglycoside modifying enzyme genes. Furthermore, the multi-drug resistances of Pseudomonas aeruginosa are serious.

Relationship between STEC biofilm formation ability and the expression of virulence genes

XIE Fandi, NIU Shumeng, SHI Jinjie, ZHANG Wenyan

2015, 30(6):  617-621.  DOI: 10.3969/j.issn.1673-8640.2015.06.018

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To investigate the relationship between the biofilm formation ability of the patients with diarrhea of Shiga toxin-producing Escherichia coli (STEC) and virulence gene expression.

STEC were detected and isolated from 520 patients with diarrhea. The antibiotic drug sensitivity in vitro and ability to form biofilm were quantitatively analyzed. Shiga toxin related gene stx1 and stx2 were amplified and detected by multiplex polymerase chain reaction (PCR). The expressions of eae and ehxA genes were detected by real-time quantitative reverse transcriptase (RT)-PCR. A total of 98 isolates of Escherichia coli from fecal specimens in healthy subjects were included as control isolates.

A total of 328 isolates of Escherichia coli were isolated from patients with diarrhea. The stx1 and stx2 gene positive bacteria (STEC) had 78 isolates, accounting for 23.8%. There were 57 isolates of STEC formed biofilm (73.1%). The relative expression of eae gene was 6.57(2.82-27.8), and the expression of ehxA gene was 1.81(1.07-5.28). There were 21 isolates of STEC unformed biofilm (26.9%).The relative expression of eae gene was 3.41(0.96-12.1), and the expression of ehxA gene was 1.76(0.89-6.06). A total of 18 isolates of Vibrio parahaemolyticus were isolated, and 10 isolates were formed biofilm (55.6%). There were 3 isolates of Proteus mirabilis.Among 98 control isolates, 39 isolates were formed biofilm (39.7%). The relative expression of eae gene was 1.02(0.86-3.47), and the expression of ehxA gene was 1.03(0.88-3.03). Biofilm-formed STEC resistance to ampicillin / sulbactam, trimethoprim and ciprofloxacin was significantly higher than that of biofilm-unformed STEC and control isolates.

The STEC biofilm formation rate is significantly higher than that in healthy subjects' colonization Escherichia coli. The expression of eae gene in biofilm-formed STEC is significantly improved. The expression of ehxA gene has no significant difference with biofilm-unformed STEC.

Plasmid-mediated fosfomycin resistance genes in Gram-negative bacilli

CAI Hongmei, TANG Yu, JIANG Xiaofei

2015, 30(6):  622-625.  DOI: 10.3969/j.issn.1673-8640.2015.06.019

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To investigate the prevalence of plasmid-mediated fosfomycin resistance genes in Gram-negative bacilli and analyze the correlation with drug resistance.

A total of 133 Gram-negative bacilli were collected, including Escherichia coli (52 isolates, 39.1%), Pseudomonas aeruginosa (36 isolates, 27.1%), Klebsiella pneumoniae (31 isolates, 23.3%) and Acinetobacter baumannii (14 isolates, 10.5%). The antimicrobial susceptibility test was performed by Kirby-Bauer (K-B) method, and the plasmid-mediated fosfomycin resistance genes(fosA, fosA3 and fosC2) were detected by polymerase chain reaction (PCR).

The fosfomycin-resistant rate of Escherichia coli was 9.6%(5/52), and the ones of Pseudomonas aeruginosa, Klebsiella pneumoniae and Acinetobacter baumannii were 36.1%(13/36), 74.2%(23/31) and 64.3%(9/14), respectively. Five of 133 isolates were positive for fosA, and 5 isolates were positive for fosA3, all belonging to Escherichia coli or Klebsiella pneumoniae, while there was no fosC2 gene. Seven of 10 isolates with plasmid-mediated fosfomycin resistance genes were resistant to fosfomycin, and the other 3 isolates were sensitive.

The fosfomycin-resistant rates of Pseudomonas aeruginosa, Klebsiella pneumoniae and Acinetobacter baumannii are higher than that of Escherichia coli. The fosA and fosA3 genes are detected in Escherichia coli and Klebsiella pneumoniae, whereas no fos gene is detected in Pseudomonas aeruginosa and Acinetobacter baumannii, highlighting the necessity of a comprehensive research in regard to fosfomycin resistance genes.

Study on carbapenemase and integrase genotypes of carbapenem-resistant Acinetobacter baumannii

BU Jingsong, WANG Yong

2015, 30(6):  626-630.  DOI: 10.3969/j.issn.1673-8640.2015.06.020

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To investigate the genotypes of carbapenemase and integrase by carbapenem-resistant Acinetobacter baumannii (CRAB).

A total of 93 isolates of CRAB and 16 isolates of carbapenem-sensitive Acinetobacter baumannii (CSAB) were collected. Carbapenemases were detected by modified Hodge testing. Metallo-beta-lactamase was detected by EDTA collaborative testing. Carbapenemase and integrase genotypes were amplified by polymerase chain reaction(PCR)and sequencing.

The carbapenemase phenotype test results showed that 62.4% isolates were positive, and metal enzymes were all negative. The PCR results for CRAB showed the detection rates of OXA-23 and Int1 were 100%, for OXA-51 was 96.8%, for OXA-58 was 1.1%, and OXA-24, VIM, IMP and Int2 were not found. The PCR results for CSAB showed the detection rate of Int1 was 37.5%.

CRAB mainly carrys OXA-23 and OXA-51 carbapenemase genes, and all isolates carry ClassⅠ integrase gene.

Anticancer activity and mechanism of osthole combined with cisplatin in human lung cancer cells

CHENG Lixia, ZHANG Benhong

2015, 30(6):  631-634.  DOI: 10.3969/j.issn.1673-8640.2015.06.021

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To investigate anticancer activity and mechanism of osthole combined with cisplatin in human lung cancer cell NCI-H460.

NCI-H460 cells were treated with osthole(50, 100 and 200 μmol/L), cisplatin (1.5, 3.0 and6.0 μmol/L)and osthole combined with cisplatin (50 μmol/L osthole+1.5 μmol/L cisplatin), and the cell proliferation was determined by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl tetrazolium bromide(MTT) assay. NCI-H460 cells were treated with osthole (50 μmol/L), cisplatin (1.5 μmol/L)and osthole combined with cisplatin (50 μmol/L osthole+1.5 μmol/L cisplatin), and then the induction of apoptosis was determined by Annexin Ⅴ/ propidium iodide(PI) staining, and the generation of reactive oxygen species (ROS) was determined by dihydroethidium (DHE) staining. Furthermore, N-acetylcysteine (NAC) was used to determine the apoptosis induced by above treatments. Untreated NCI-H460 cells were taken as control group.

Osthole and cisplatin showed the dose dependent inhibition of the NCI-H460 cell proliferation. The 50 μmol/L osthole combined with 1.5 μmol/L cisplatin can significantly inhibit NCI-H460 cell proliferation and induce its apoptosis. Compared to single treatments, combined treatment can significantly increase the ROS positive cells(P<0.05). Compared to the control group, NCI-H460 cells treated with NAC produced obviously the decreasing of ROS, apoptosis and cell proliferation(P<0.05).

Osthole combined with cisplatin can cause the apoptosis of NCI-H460 cells by inducing production of ROS.

Study of miR-29c inhibiting invasion and migration of ESCC cell by targeting Tiam1

CHU Qinghua, ZHANG Jie, SUN Fenyong

2015, 30(6):  635-640.  DOI: 10.3969/j.issn.1673-8640.2015.06.022

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To research microRNA(miR)-29c inhibiting invasion and migration of esophageal squamous cell carcinoma(ESCC) cell by targeting T-lymphom invasion and metastasis 1(Tiam1).

Invasion and migration capacity changes of ESCC cell with overexpression of miR-29c were observed through cell scratch and transwell experiments. Western blot analysis and sensor reporter assay were used to research the regulation and target site of miR-29c for Tiam1. In shRNA rescue experiments, Western blot analysis was used to detect the expression change. Immunofluorescence-laser scanning confocal microscope method was used to investigate the mechanisms of miR-29c regulating the invasion and migration capacities of ESCC cell.

Cell scratch and transwell experiment results showed that miR-29c inhibited the invasion and migration of ESCC cell. Tiam1 was predicted to be the target gene of miR-29c through bioinformatics methods. Western blot analysis and sensor reporter assay showed that miR-29c degradated Tiam1 through binding Tiam1 3'-untranslated region (UTR). The analysis of shRNA rescue experiments, including cell scratch experiment, Western blot and immunofluorescence analysis, showed that activated Rac1- guanosine triphosphate(GTP) reduced due to miR-29c inhibiting Tiam1 expression, which resulting migration capacity of ESCC cell diminished, while the inter ference of siTiam1 (artificially synthesized siRNA-specific molecule for Tiam1) can "rescue" the migration capacity of ESCC cell.

miR-29c lowers the level of Rac1 activation through degradating Tiam1 mRNA, resulting the change of cell motility status, thereby inhibiting ESCC cell invasion and migration.

Primary experimental study on the reference method for the counting of red blood cells and white blood cells

XU Lei, WANG Qing, MIAO Yingbo, SONG Ying, WANG Guofei, ZHU Peichao, GUO Xiaojun

2015, 30(6):  641-645.  DOI: 10.3969/j.issn.1673-8640.2015.06.023

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To investigate the reference method for red blood cell (RBC) and white blood cell (WBC) counts, and to establish standardized determination platform which will provide technical support for blood cell counting trueness verification test.

The reference method was established according to relevant documents of the International Council for Standardization in Hematology (ICSH) and the standard of the People's Republic of China (WS/T 245-2005). The S-CAL cell calibrator, 5C cell control, aperture instrument concentration control, processed whole blood and fresh whole blood samples were determined on Z2 particle counter. Parameters such as within-run and between-run coefficients of variance (CV), coincidence counting correction, stability and comparability were evaluated.

The method showed that within-run CV was<1%, and between-run CV met the requirements of the Clinical and Laboratory Standards Institute (CLSI) H26-A2. The results demonstrated the biases of coincidence counting correction in RBC and WBC counts were <1%. RBC and WBC of processed fresh whole blood samples were stable after storage for above 5 d. The biases of fresh whole blood RBC and WBC between Z2 particle counter and BECKMAN-COULTER ACT diff Ⅱ, BECKMAN-COULTER LH-750, SYSMEX K-4500, SYSMEX XE-2100 hematology analyzers were ≤±2% and ≤±4%.

The reference method for RBC and WBC counting by Z2 particle counter has been established basically. Precision, coincidence counting correction, accuracy, stability and comparability are conformed to the requirements. It provides the technical support for blood cell counting trueness verification test.

Research progress on immune factors related with HIV infectious disease development

WEI Shanqiu

2015, 30(6):  646-649.  DOI: 10.3969/j.issn.1673-8640.2015.06.024

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Host immune response is one of important factors that determines the progress of human immunodeficiency virus (HIV) infectious diseases. Different immunocytes or immune factors might play different roles, or even work oppositely, when the disease develops. So far, key factors affecting HIV infectious disease development clinical outcome are still unclear. This review gives an overview to the hot research in this field.

Application of nanomaterial-based field effect transistor biosensor in medical detections

CAI Bingjie, ZHANG Guojun

2015, 30(6):  650-654.  DOI: 10.3969/j.issn.1673-8640.2015.06.025

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The combination of nanomaterials and biosensing elements to selectively recognize biological molecules promotes the development of nanobiosensors. Nanomaterial-based field effect transistor biosensors are one of newly developed biosensing technologies. Compared with the conventional detection technologies, they offer several important advantages including high sensitivity and selectivity, rapid analysis, label-free detection, easy operation and so on. This review summarizes the working principle and fabrication of field effect transistor biosensors based on graphene, silicon nanowires and carbon nanotubes and their applications in medicine detection.