Table of Content

05 July 2011, Volume 31 Issue 7

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Hypertonicity Induces Mucus Hypersecretion in Human Airway Epithelial Cells

Jing TU Juliy M.Perelman Victor P.Kolosov Xiang-dong ZHOU

2011, 31(7):  723-727. 

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Objective To investigate the effect of hypertonic conditions induces mucin(MUC)5AC secretion in HBE16 cells, and in which the possible mechanism of PKC-HSP70 pathways. Methods HBE16 cells were cultured in hypertonic saline conditions to induce mucus hypersecretion in vitro. Inhibitors of PKC isoforms were used such as G?6976 (PKCμ inhibitor), Safingol (PKCα inhibitor), LY333531 (PKCβinhibitor) and rottlerin (PKCδ inhibitor) which were served to interference HBE16 cells. HSP70-2 were determined by Western blot analysis. RT-PCR was used to detecte the transcriptional level of HSP70-2. Mucin(MUC)5AC protein content in supernatant were analysed by ELISA. Results The level of HSP70-2, HSP70-2mRNA and MUC5AC proteins werer strongly increased after cells were exposed to hypertonic conditions, and the expression content were increased in a time-dependent manner. (all p < 0.05). The expressions in G?6976 group were significantly lower than that in hypertonic group (p < 0.01) and were still higher than that in control (p < 0.05). Treatment with Safingol, LY333531 and Rottlerin exerted no effects on hypertonicity-induced the content of HSP70-2, HSP70-2mRNA and MUC5AC proteins. Conclusion Hypertonic saline induces the MUC5AC mucin hypersecretion in human bronchial epithelial cells. HSP70-2 mediates hypertonic stress induces mucin hypersecretion through PKCμ dependent signaling pathways.

Quantities and phenotypes of tumor infiltrating gdT cells are related to clinical staging of head and neck tumor

Shan-shan YIN Xing-ming CHEN Ning KANG Xiao-wei CHEN Zhi-qiang GAO Wei HE

2011, 31(7):  728-735. 

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Objective This study was designed to explore the relationship between infiltrated Foxp3+gdT cells and the clinical stage of head and neck tumor. Methods Head and neck tumor biopsies were mechanically minced to get tumor infiltrating lymphocytes (TILs). Peripheral blood mononuclear cells (PBMCs) from the patients were isolated by density gradient centrifugation. TILs and PBMCs were stimulated by immobilized anti-pan-TCRgd in vitro for two to four weeks. Freshly isolated and expanded cells underwent immunofluorescence staining and flowcytometry analysis. Commercial head and neck tumor tissue chips were used for immunohistochemical staining. Results Immunofluorescence staining of TILs freshly isolated from 15 head and neck tumor samples showed the gdT/CD3+T ratio as 0.37%~12.35%, Foxp3+ gdT/gdT ratio as 0.13%~34.38%. Following in vitro expansion, gdT/CD3+T cell ratio increased to 20.38%~82.87%. Expanded cells primarily belonged to the Vd1 subset. Tumor tissue chips were divided into four groups according to the T stage of the clinical TNM staging system. Subsequent immunohistochemical results indicated a correlation between the quantity of Foxp3+gdT cells and tumor TNM stage. Conclusion Our data demonstrate the existence of Foxp3+ gdT cells in the head and neck tumor tissue, and suggest that the quantity of tumor infiltrating gdT regulatory cells might be associated with the prognosis of head and neck tumor.

Association between cytotoxic T lymphocyte-associated antigen 4 gene haplotype and Drug users with HIV Infection

Ying LIU Bo DIAO Yuan-ying SHEN Fang ZHOU Yong FENG Fan LUO Wei HOU

2011, 31(7):  736-739. 

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Objective:To investigate the association of gene polymorphism of cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) with drug users with HIV infection.Methods The A+49G transition polymorphism at position 49 (exon 1) and C-318T transition polymorphism at position 318 in promoter of the CTLA-4 gene were determined by polymerase chain reaction with sequence specific primers (PCR-SSP) method in 24 drug users,41 drug users with HIV Infection patients and 204 healthy controls.Results Compared with control group, in drug users group the frequency of C/C(12.50% VS 51.47%)( P<0.01) significantly reduced; in drug users with HIV infection group,no significant differences in the distribution of genotype and allele frequencies were observed in A+49G gene polymorphisms (p>0.05); however,there are significant differences in C-318T gene polymorphisms in drug users with HIV infection group(p<0.01),the frequency of C/T (68.3% VS 45.6%) and T/T（21.95% VS 2.94%）increased significantly,in contrast in C/C（9.76% VS 51.47%）significantly reduced. Compared with drug users group, no significant differences in the distribution of genotype and allele frequencies were observed in A+49G and C-318T gene polymorphisms (p>0.05) in drug users with HIV infection group.Conclusion There is close associativity between CTLA-4 Alleles and drug taking with HIV infection,in C-318T genotype, C/T and T/T were positively correlated with occurence of drug taking with HIV infection, but C/C was negatively correlated with it.

Construction of lentiviral expression vector expressing mouse Islet-1 gene and investigation of Islet-1 mediated induction of C3H10T1/2 cell differentiation into cardiomyocyte-like cells

Shen-shen ZHI Jie TIAN Guan-xin LIU Rong LU Jian-ping LIN Jian-ping LIU Jing ZHU

2011, 31(7):  740-745. 

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Object To construct lentiviral expression vectors targeting mouse gene Islet-1 and to study the effect of Islet-1 on stem cell differentiation. Methods Islet-1 gene was obtained by PCR and insected into pLenO-WPI Vector. The positive plasmid was selected and infected 293T cells with helper plasmid to produce recombinact lentivirus. Islet-1 expression and its related gene, gene and protein markers of heart, liver, bone, and nerves systems were measured by Real-time quantitative PCR and Western blot after C3H10T1/2 was infected by the lentiviruses. Results PCR and sequencing showed that the right DNA fragment and Islet-1 had been inserted and can be detected in both in gene and protein levels. Cardiac development related genes GATA-4, MEF2C; NKx2.5 increased in 1week and reached the highest level in 2 weeks, and cTnT reached a high level in 3 weeks and increasedover time. There were no expressions of gene and protein markers of liver, bone, brain. Conclusion Lentiviral expression vector carrying mouse Islet-1 gene was constructed. Islet-1 promoted C3H10T1/2 cells differentiate into cardiomyocyte-like cells specifically, that laid the foundation for further investigation of Islet-1.

Regulation of inflammatory cytokine gene transcription in macrophage induced by P.yoelii MIF

Ding-ding SHAO Shan ZENG Xiang ZHONG Heng WANG

2011, 31(7):  746-750. 

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Objective Plasmodium derived macrophage migration inhibitory factor (MIF) ortholog was considered to have potential to regulate host immune cell. Plasmodium yoelii derived MIF (PyMIF) has highly conserved crystal structure and similar bioactivity with host mouse MIF. However, the regulatory mechanism, including the function of PyMIF on host macrophage, is not clear so far. Here, we report the regulation of PyMIF on host inflammatory cytokines transcription in macrophage, and the compared analysis of regulation activity between PyMIF and host mouse MIF (MmMIF). Methods Both PyMIF and MmMIF were expressed and purified as recombinant proteins and were removed the endotoxin by C8 reversed-phase column. Then the endotoxin-free protein was added to cultured mouse monocyte / macrophage cell line RAW264.7. After incubation, the total RNA was extracted and applied to "RT2 Profiler? PCR Array Mouse Inflammatory Cytokines & Receptors" analysis. Results PyMIF significantly up-regulated 17 inflammatory cytokine genes transcription level and down-regulated 3 genes (CCR6、CCR8 and IL1F6) transcription. The regulated gene quantity by PyMIF was more than MmMIF. Most genes regulated by PyMIF were involved in a Th2 response bias or in expanding inflammatory response. Conclusions There is much difference between PyMIF and MmMIF in regulating host macrophage activity. These results provided more information about Plasmodium MIF function on host immune cell, and will be meaning for deep understanding the role of Plasmodium MIF during infection.

Preparation of MUC1-Targeted Nanoparticles and Evaluation of Its Cytotoxicity In Vitro

Chen-chen YU Yan HU Jin-hong DUAN Chen WANG Hai-yan XU Xian-da YANG

2011, 31(7):  751-755. 

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Objective To construct MUC1-targeted nanoparticles and evaluate its cytotoxicity in vitro. Methods MUC1 aptamers were conjugated to the surface of nanoparticles which were made of poly (lactic-co-glycolic-acid) (PLGA) nanoparticles and loaded with paclitaxel (PTX). The MUC1-targeted nanoparticles (Apt-NP-PTX) were characterized by UV spectrophotometry and dynamic light scattering. Using MUC1-overexpressing MCF-7 breast cancer cell as experimental group, and HepG2 as control group, the specificity of the Apt-NP-PTX was detected by flow cytometry. The cytotoxicity and IC50 of Apt-NP-PTX against MCF-7 were evaluated using a standard MTS assay. Results The Apt-NP-PTX were about (225.3± 9.2) nm in size with an encapsulation efficiency of 83.6% ± 1.7%. The Apt-NP-PTX enhanced in vitro drug delivery and cellular toxicity to MUC1+ cancer cells, as compared with non-targeted nanoparticles that lack the MUC1 aptamer (P < 0.01) , with IC50 of 1.52 mg/L and 4.10 mg/L, respectively. Conclusion the Apt-NP-PTX can effectively enhance the PTX delivery to MUC1- overexpressing MCF-7 cells in vitro.

Investigation of venous blood cells from 3 866 children in Heilongjiang Province to get reference ranges for clinical usages

Juan DU Wei WU Ling QIU Tao XU Da-wei HUANG Li-ya XIN Qian CHEN Xin-qi CHENG Geng WANG Shao-mei HAN Guang-jin ZHU

2011, 31(7):  756-761. 

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Objective We aimed to establish all hepatocytes related reference ranges for normal children by investigating 8 parameters referred to children’s venous whole blood in Heilongjiang Province. Methods Fasting venous blood samples of 3 866 healthy children aged 8 to 16 (1 837 male and 2 029 famale)were taken into EDTA-K2 anticoagulation tube, and the related reference values were measured by Sysmex XT-1800i hematoanalyzer. Software SPSS 15.0 was used for statistical analysis to descript the deviations between ages、races and regions. Results The means(reference ranges) of venous blood of 3866 health children in the Heilongjiang Province were as follows: White blood cells count to 6.9 (3.9～9.9)×109/L,Red blood cells count to 5.0 (4.2～5.8)×1012/L for male and 4.6 (4.0～5.2)×1012/L for female,the hemoglobin concentration was 146(124～168)g/L for male and 135(117～153)g/L for female, the Hematocrit is 43.9%(37.5%～50.5%) for male and 41.2%(36.4%～46.0%) for female, the Mean corpuscular volume was 88(80～97)fl, the mean corpuscular hemoglobin was 29(25～33)pg，the mean corpuscular hemoglobin concentration was 329(311～347）g/L，the platelet count was 277(161～393)×109/ L .Among health children, when compared with Korean ethnic minority group, all indexes like RBC、HCT、MCV and PDW of Han nationality did not show any differences with statistical significance. While there were remarkably differences on WBC、HGB、MCH、MCHC and PLT( P<0.05).Comparing between different ages and groups, all parameters except WBC presented with significant differences (P<0.05), positive correlation was found between RBC、HGB、HCT、MCV、MCH and age, negative correlation was found between MCHC、PLT and age. Conclusions Basic data bank for reference ranges of venous blood of healthy children in Heilongjiang Province had been initially established .And the differences were found among different ages、nations and regions of the children aged 8 to 16 is of great value in clinical purposes.

Overexrpession of miR-16 promotes apoptosis in Neuro2a cells

Chang LIU Wei LIU Ming WANG Bin YIN Jian-gang YUAN

2011, 31(7):  762-766. 

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Objective To investigate the role of miR-16 in cell cycle and apoptosis in Neuro2a cells and explore the possible function of miR-16 in tumor. Methods The overexpression plasmid pcDNA3.1-miR-16 of miR-16 was cloned and introduced into Neuro2a cells. Real-time PCR was chosen to test the overexpression of miR-16. Cell cytometry was used to study the role of miR-16 in cell cycle and apoptosis, and apoptosis cells were quantitatively examined by TUNEL. Results After tranfection of pcDNA3.1-miR-16, the mature miR-16 was significantly overexpressed (P<0.05). Cell cytometry results showed that the cell cycle didn’t change after miR-16 overexpression while cell apoptosis were found and confirmed by TUNEL. Conclusion There was obvious apoptosis of Neuro2a cell after overexpression of miR-16, thus, miR-16 might involve in cure of tumor.

The effect of curcumin through suppression of IκBα phosphorylation on proliferation of Esophageal Squamous Cell Carcinoma cell lines

Fang TIAN Yu-rong CHAI Ya-nan JIANG Xiao-yan ZHANG

2011, 31(7):  767-772. 

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Objective This study was to detect whether curcumin inhibits NF-κB signaling pathway through suppression IκBα phosphorylation and evaluate the effects on proliferation and cell cycle in EC9706 and Eca109 cell lines. Methods ESCC cells were treated with different concentrations of curcumin and then the number of viable cells were determined by MTT. Cytoplasmic extracts or whole cell extracts were examined for the expression of pIκBα and cyclinD1 using Western Blotting after treated with curcumin at different time. ESCC cells were treated with curcumin, 5-FU or combinations thereof for 72h. The cells were stained with PI and analyzed cell cycle by flow cytometry. Results The results showed that curcumin inhibited two ESCC cells growth in a concentration-dependent manner. The Western blotting results indicated that curcumin inhibited IκBα phosphorylation and downregulated the expression of cyclinD1 in the two ESCC cell lines in a time-dependent manner. CyclinD1 was declined in the forepart of curcumin-treated cells. Curcumin-treated cells showed an increase in the percentage of cells in the G0/G1 phase and a decrease in the percentage of cells in the S phase. While combination with 5-FU, the results were more prominence. Conclusion Therefore, curcumin inhibits the phosphorylation of IκBα and downregulates the activation of NF-κB signaling pathway, leading to suppression of proliferation in ESCC cell. Therefore, curcumin may prove useful in the treatment of ESCC because it is a pharmacologically safe compound without side effects.

Association of methylenterahydrofolate reductase gene C677T polymorphism and plasma homocysteine and folate concentration with coronary heart disease

Xiao-ping HU Chun-lian LIU Ling WU Xiao-jing DAI Liang PENG Hai-yan JIAO Yin-tao CHEN

2011, 31(7):  773-776. 

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Objective To investigate the correlation between methylenterahydrofolate reductase (MTHFR) gene C677T polymorphism, homocysteine (Hcy), folate and coronary heart disease (CHD) in the Han population of Ningxia, China. Methods In this case-control study, we selected 202 patients with CHD and 199 normal controls in Han population from Ningxia. The gene polymorphism of MTHFR C677T was detected by PCR-RFLP assay. Plasma Hcy concentrations were measured by fluorescence polarization immunoassay (FPIA) and serum folate and vitamin B12 concentrations by chemiluminescent immuno assay (CLIA). Results (1) In CHD and control groups, the frequencies of MTHFR C677T genotype were CC (homozygous wild) 23.3% vs 20.7% , CT (heterozygous) 52.3% vs 54.5% and TT (homozygous mutant) 24.4% vs 24.8% respectively. There was no difference in the genotype and allele frequency distribution of MTHFR C677T polymorphism between two groups. (2) In CHD group, the plasma Hcy concentration of individuals with CC genotype was lower than that of T allele carriers (10.84 μmol/L vs 12.24 μmol/L, P< 0.01). The plasma folate concentration of individuals with CC genotype was higher than that of T allele carriers (5.38 μg/L vs 3.72 μg/L, P=0.01). Conclusion There was no significant difference between CHD patients and normal people in genetic polymorphism of MTHFR C677T in Han population of Ningxia. The polymorphism of MTHFR C677T was related to the level of plasma Hcy and folate, which possibly were the risk factor of CHD.

Decreased bone remodeling in type 2 diabetic osteoporosis rats

2011, 31(7):  777-782. 

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Objective To explore the characteristic of bone remodeling by establishing type 2 diabetic osteoporosis rat model. Methods Female Wistar rats were randomly divided into sham operation group (NS); bilateral ovariectomy group (NOVX); diabetic sham operation group (DS ) and diabetic ovariectomized group (DOVX ). Ovariectomy was performed in both control and diabetic rats after type 2 diabetic models had been made . Venous blood samples were collected at 0, 4, 8 and 12wks after ovariectomy，fasting plasma glucose(FPG)、fasting insulin(FINS)、estrogen(E2) (B-ALP),tartrate-resistant acid phosphatase (TRACP5b) were all measured. BMD were measured , osteoblast and osteoclast cell were isolated and cultured from rats of all groups. Results At 0wks，FPG had significantly increased when DOVX' 13.1±4.9 mmol/L compared with NS’4.4±0.6mmol/L（all P<0.05）. at 4wks, BMD had significantly decreased when DOVX'0.073±0.012g/cm2 compared with DS'0.098±0.016g/cm2（P<0.05）.at 4、8 and 12wks, there were no differences in plasma B-ALP levels when DOVX rats compared with NS rats.at 4 and 8wks,compared with NS rats, DOVX rats had significantly higher plasma TRACP5b concentration (all p<0.05). For the activity of Osteoblast , there were no differences when DOVX compared with NS , NOVX and DS group . osteoclast numbers were higher in DOVX rats than those from NS rats(P <0.05).Conclusion There were actived bone resorption and relatively slow bone formation in type 2 diabetic osteoporosis rats.

Therapeutic effects of autologous RetroNectin activated CIK killer cell immunotherapy for patients with advanced colon carcinoma

Ben-ling XU Quan-li GAO Long YUAN Rui-hua FAN Cheng-juan ZHANG Yong-ping SONG

2011, 31(7):  783-787. 

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Objective To study the therapeutic safety and effects of autologous RetroNectin activated CIK cell immunotherapy for patients with advanced colon carcinoma. Methods CIK cells were generated from peripheral blood mononuclear cells (PBMC) and incubated in the presence of IFN-γ followed by Retronectin, mouse antihuman CD3 monoclonal antibody and IL-2. Then, CIK cells were transfused intravenously every week for at least 4 times. Immunophenotypic characteristics of CIK cells were analyzed by flow cytometry. IFN-γ, tumour necrosis factor α (TNF-α), IL-2, IL-10 and IL-4 concentrations in serum were determined, using the BDTM human Cytometric Bead Array kits (CBA). Results The median number of transferred cells per patient was 23.9×109/L(20.6×109~27.2×109), and the toxicity pro?le was favourable. After CIK cells infusion, the absolute median count of lymphocytes, CD3+, CD8+ and CD3+CD56+ cells signi?cantly increased in patient’s peripheral blood. One patients had partial response (PR) and four patients had stabilization of disease(SD). The downtrend of the CEA level was observed in the PR and two SD patients. Responding patients, after treatment, showed in the serum an increased production of IFN-γ (6.9 vs. 53.2 pg/ml) and TNF-α (3.2 vs. 24.6 pg/ml) that is statistically signi?cant with a p-value of 0.001 and 0.004, respectively. No signi?cant differences were seen in non-responders and in the expression of other cytokines. Conclusion Adoptive immunotherapy with Retronectin induced CIK cells is a safe therapy with some suggestion of ef?cacy that signi?cantly enhances immune functions increasing absolute numbers of effector cells without side effects. This study might provide valuable information on the conventional treatment strategy of malignancies.

Transplantation of neural stem cells from human adipose tissue-derived stromal cells for treatment of cerebral ischemic- reperfusion injury in rats

Bin LIU Jing DONG Ning LIU Rui-min WANG Jin-xia ZHANG Shi-ying LI Gui-liang CHEN

2011, 31(7):  788-792. 

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Objective To study the effects of neural stem cells from human adipose tissue-derived stromal cells transplantation for treatment of cerebral ischemic- reperfusion injury in rats. Methods:The middle cerebral artery occlusion (MCAO) 2h and reperfusion models were made by intraluminal vascular occlusion in rats.Adipose tissue-derived stromal cells were cultivated,proliferated and induced into neural stem cells in vitr.Transplantation treated group was transplanted neural stem cells(2×106 cells /ml ) through tail vein at 24h hours after MACO. Neurological function changes were by using Modified neurologic severity scores(MNSS), the volume of cerebral infarction and the change of cerebral pathology were observed by TTC and HE staining. Results The neurologic deficit scores in transplantation treated group were significantly lower than those in ischemia control group at 14d、21d、28d of MCAO (P<0.05,orP<0.01). The infarction volume of transplantation treated group (40.20±9.5 mm3) was less than that of ischemia control group (66.60±14.24mm3) at 14d (P<0.01). The number of degeneration and necrosis nerve cell in ischemia transplanted group is significantly reduced compered to ischemia-reperfusion group.Conclusion Transplantation of neural stem cells differentiated from human adipose tissue-derived stromal cells could improve neurological functionon of cerebral ischemic- reperfusion injury in rats and protective effect on neurons.

Comparison of different cartilage protein extraction strategies used by two-dimensional electrophorosis technique

Jun XIAO Ruo-feng YIN Jin-qian LIANG Zhan-jun SHI Zhi-hong WU Gui-xing QIU

2011, 31(7):  793-798. 

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Objective To explore an optimal cartilage protein extraction approach that can guarantee both the image quality and the result fidelity of the two-dimensional gel electrophoresis (2-DE) technique. Methods Knee cartilage samples were obtained from femoral condyles (n=17). Approaches used for protein samples of 2-DE were grouped: (1) Extracting protein directly from cartilage samples (Cartialge approach); (2) Total protein was treat with cetylpyridinium chloride (CPC) after being extracted from cartilage samples (Cartilage plus CPC approach); (3) Extracting protein from chondrocytes directly isolated from cartilage samples (directly extracted chondrocytes approch). (4) Extracting protein from cultured chondrocytes (cultured chondrocyters approach). Image qualities generated by 2-DE with different protein extracting approaches were compared and the capabilities of these approaches in generating believable proteomic results were also evaluated. Results Isoelectrofocusing was failed in the cartilage approach group. Cartilage plus CPC precipitation approach group can reach isoelectrofocusing, but the protein spots it generated were much less than those of both the chondrocyte approaches. The directly isolated chondrocytes approach group generated comparable high-quality gel images to that of the cultured chondrcoytes approach group. It also isolated some protein spots that were missed in the high-molecular or alkaline regions that of the cultured chondrocytes approach group. Results of mass spectrography revealed that the proteins located in these regions were collagen VI, TGF-β2 and annexin, etc. All were pathologic proteins correlated with the pathogenesis of osteoarthritis. Conclusion The approach of isolating chondrocytes from cartilage and extracting total protein directly for 2-DE analysis can avoid the problem of isoelectrofocusing disturbance and generate believable results through avoiding altered protein expression spectra under cell-culture environment, which was an optimal protein extraction strategy for 2-DE technique in investigations on cartilage-related diseases.

Inhibitory effect of Valproic Acid on cell cycle of Kasumi-1 cell line in nude mice

Xia TIAN Peng LIU Li-hong WANG Zhi-hua ZHANG Lei ZHAO Chang-lai HAO

2011, 31(7):  799-803. 

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Objective To investigate the influence of Valproic acid(VPA) on cell cycle of Kasumi-1 cells. Methods Kasumi-1 cell line model transplanted subcutaneously in nude mice were established and divided into control group and VPA group, to observe and record the growth of nude mice and their transplanted tumors and calculate volume and inhibition rate.Kasumi-1 cell of each group cell cycle changes were analyzed by flow cytometry.P21WAF1/CIPand pRB were detected by immunohistochemistry and Western blot. Results ①VPA obviously inhibited the growth of tumor, animals treaed with VPA showed a statistically significant decrease in tumor volume and weight compared with controls with 57.25% growth inhibition. ?VPA blocked cell in G0/G1 phase were detected by flow cytometry. ?The P21WAF1/CIP protein significantly increased with VPA group. pRB protein significantly decrease with VPA group. Conclusion We conclude that administration of VPA inhibit the growth of Kasumi-1 cell line tumor VPA can arrest Kasumi-l cell in Go/G1 phase through the regulation of P21WAF1/CIP and pRB.

Propofol Alleviates Cerebral Ischemia and Reperfusion Damage in Rats

Shou-lin ZHONG Hui-ping CHEN Zhi-liang ZHU Jing ZHOU Tong-xiang WANG

2011, 31(7):  804-808. 

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Objective To investigate the protective effect of propofol on cerebral ischemia and reperfusion damage in rats. Methods Eighteen adult male SD rats were randomly divided into 3 groups, ischemia group (n=6), sham injury group (n=6) and propofol group (n=6). Cerebral ischemia and reperfusion models were made by Longa method, monitoring the rectal temperature and blood sugar. The rats were excuted after 24 hours, whose brains were dying by TTC to measure the area of the cerebral infraction. Apoptosis and necrosis rate were detected by cytometry .Western blot was used to detect the expressions of the HIF-1α、caspase-3 and survivn in brain mantle and hippocampi. Results Apoptosis and necrosis appeared in the nerve cells in brain mantle and hippocampi after the cerebral ischemia and reperfusion damage, in the same time, the expressions of HIF-1α and caspase-3 increased and the survivn decreased .However, the Apoptosis and necrosis reduced in the propofol groups, and the expression of HIF-1α and caspase-3 were inhibited and the survivn increased. (HIF-1α/α-tublulin ratio 0.68±0.02 vs 0.84±0.03, P<0.05, caspase-3/α-tublulin ratio 0.29±0.03 vs 0.57±0.04, P<0.05, survivn/α-tublulin ratio 0.52±0.02 vs 0.19±0.02, P<0.05). Conclusion Propofol may inhibit hypoxia in the brain and the apoptosis of nerve cells in result of protecting the cerebral ischemia and reperfusion damage in rats.

The effect of Elafin in 16HBE cytoplasm on the mucin5AC production

Hua-sheng WEI Juliy M.Perelman Victor P.Kolosov Xiang-dong ZHOU

2011, 31(7):  809-813. 

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Objective To investigate the effect of Elafin in cytoplasm on epidermal growth factor(EGF)-induced mucin(MUC)5AC production and the mechamism of it. Methods After cultivating 16HBE cells in vitro, the constructed pEGFP-N1-Elafin eukaryotic expression vector was transfected into the 16HBE cells by LipofectamineTM2000,and 16HBE cells transfected the vector without Elafin gene as control group. The cells were then incubated with exogenous EGF.The mucin(MUC)5AC mRNA level was analyzed by RT-PCR,the MUC5AC protein production was assayed by ELISA,the NF-κB activity in nuclear was detected by immunofluorescence and laser scanning confocal microscope,the SUMO-1-p-IκBα production was assayed by co-immunoprecipitation(CO-IP) and Western blot. Results The MUC5AC mRNA level,the MUC5AC protein production(0.36±0.09) and the NF-κB activity of the experimental group were all significantly lower than those of the empty vector group(MUC5AC production of the later is 0.55±0.07),but the SUMO-1-p-IκBα production was significantly higher than that of the latter(all P﹤0.01). Conclusions Elafin prevents NF–κB activation via an effect on p-IκBα sumoylation,thus reducing epidermal growth factor-induced mucin5AC production.

Autophagy Enhances Chemotheragy Resistance of CD133+ Colon Cancer Stem Cells to 5-FU

SI Xiao-li, LIU Wei, YANG Ai-jun, WANG Chen-yu, LI Min

2011, 31(7):  814-819. 

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Objective To observe the autophagy of CD133+ CSCs induced by 5-FU,and explore the effect of autophagy on the chemotherapy Methods The cell proliferative inhibition rates and IC50 of 5-FU on human colon cancer cell lines SW480 were measured by MTT assay. CD133+ CSCs were purified with MACS(magnetic activated cell-sorting system) CD133 cell isolation kit. Then the morphplpgical features of cells were observed using transmission election microscopy(TEM),and the autophagic vacuoles(AV) was observed using fluorescent microscope by monodansylcadaverin(MDC) staining. The effect of autophagy on the chemotherapey was evaluated by cell viability assay and colony-forming test. Results The vibility of human colon cancer cell line SW480 was obviously inhibited with different contrations of 5-FU,half of inhibition concentration(IC50) value was (1.09±0.18)μg/mL .Autophagsomes and autophagy vesica were observed in CD133+ CSCs. The combination treatment of 5-FU and 3-MA significantly decreased the viability of CD133+CSCs from (81.8±4.6)% to (56.3±5.5)%(P<0.05), and the colony-forming efficiency from (81.1±3.4)% to (64.4±4.8)% (P<0.05), compared with 5-FU alone. Conclusion 5-FU may induce autophagy in CD133+ CSCs. Inhibition of autophagy may be employed to increase the cytotoxic effect of CD133+ CSCs to 5-FU.

Establishment of kainic acid-induced mice models of medial temporal lobe epilepsy

Ting-ting HE Dan ZHANG Xiao-liang XING Long-ze SHA Li-ri JIN Yan SHEN Li-wen WU Qi XU

2011, 31(7):  820-826. 

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Objective To establish the kainic acid (KA)-induced mesial temporal lobe epilepsy (TLE) mice model and detect the characteristic changes. Methods Healthy male C57BL/6 mice were randomly divided into control group (non-injected), saline group (35 μL/g) and KA group (12 mg/kg) for intrahippocampal injections. 5 days and 5 weeks after injection,the mice brains were sectioned and stained by hemotoxylin-eosin (H&E) to detect the pathologic changes in the hippocampal area; the expression level of P-S6 protein, the marker of hippocampus mTOR signal pathway, was also determinated to verify the animal model. Results （1）Only KA-treated mice showed wet dog shakes, facial clonus and generalized tonic-clonic convulsions.（2）After injection of KA, neuronal cell loss was prominent in in CA1, CA3 and hilar area of the ipsilateral hippocampus; granule cell in dentate gyrus area dispersed, which are fertures of hippocampus sclerosis.（3）P-S6 expression level increased in both acute (P < 0.05) and chronic phases (P < 0.01) of MTLE models. Conclusion Intrahippocampal injection of KA in C57BL/6 mice could induce behavioural, histopathological changes which are similar to those clinic features of MTLE.

Clinical Significance of the Triage Role of HR-HPV for Atypical Squamous Cells, cannot Exclude High-grade Squamous Intraepithelial Lesion

Xiao-yan ZHANG Mei-lu BIAN Qing-yun CHEN

2011, 31(7):  827-830. 

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Objective To evaluate the possible role of HR-HPV testing for ASC-H in clinical practice.Methods Among 17 125 cervical smesrs in China-Japan friendship hospital from June 2004 to April 2007 ,those interpreted as ASC and those had HR-HPV testing were enrolled in the study.The possible role of triage of HR-HPV testing for ASC-H was evaluated according to the results of biopsy.Results CIN 2/3 was identified in 25 of 52(48.1%) patients with HR-HPV positive ASC-H compared with 48 of 190 (25.2%)patients with HR-HPV positive ASC-US（P <0.01）. CIN 2/3 was identified on a follow-up in a similarly small proportion of both patients with HR-HPV negative ASC-H and patients with HR-HPV negative ASC-US (5.9%、5.7%，respectively).The sensitivity of HR-HPV testing in detecting CIN 2/3 for ASC-US and ASC-H was 82.8%,96.2% ,respectively;and the negative prective rate was 94.3%,92.3% ,respectively. Conclusions Patients with ASC-H and negative HR- HPV testing may be more efficiently managed by follow-up with regular Pap and HR-HPV testing rather than universal colposcopy.HR-HPV testing for triage of ASC-H in routine practice may have the same important triage role as for ASC-US.

Diagnosis and treatment of 32 cases with heterotopic pancreas

Ying-chao GU Jian-chun YU

2011, 31(7):  831-833. 

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Objective To investigate the clinical features and treatment methods of ectopic pancreas. Methods Retrospective analysis of 32 pathologically confirmed patients with ectopic pancreas in Peking Union Medical College Hospital from 1992 to 2010. Results Thirty-two cases with heterotopic pancreas located in the stomach（15 cases）, duodenum（4 cases）, jejunum（9 cases） and ileum（4 cases）, and 26 cases were detected in recent 4 years. Eight cases were diagnosed before surgery, 6 cases were misdiagnosed as gastrointestinal stromal tumors, 5 cases were misdiagnosed as leiomyoma, 1 case was misdiagnosed as malignant gastric ulcer; the remaining 12 cases were found accidentally in other abdominal surgery. One case was confirmed by routine endoscopic biopsy，the rest 31 cases accepted surgical resection and confirmed by postoperative pathologic findings; all patients had no complications. Conclusions Gastrointestinal submucosal protuberant lesions should be considered the possibility of ectopic pancreas, endoscopic procedure or surgical resection are available if confirmed diagnosis is made.

Clinicopathologic features of Churg-Strauss syndrome with renal involvement

Bi-xia GAO Ming-xi LI Yu-bing WEN Xue-mei LI Xue-wang LI

2011, 31(7):  834-838. 

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Objective To study the clinicopathologic features of Churg-Strauss syndrome (CSS) with renal involvement. Methods The clinicopathologic data of patients with CSS in Peking Union Medical College Hospital in last 10 years were retrospectively collected and analyzed. The outcome of CSS cases with renal involvement after one month intervention was evaluated. Results Forty cases were enrolled with a mean age of （43.6±18.1）years. Antineutrophil cytoplasmic antibody (ANCA) was positive in nine of forty patients (22.5%), with specificity for myeloperoxidase(MPO) in six patients（15%）. Renal involvements were found in thirteen patients (32.5%), six cases (46.2%) manifested as isolated urinary abnormalities, seven cases（53.8%）manifested as renal insufficiency with estimated glomerular filtration rate(eGFR) <60 ml/min/1.73m2. The positive rates of ANCA were significant higher in patients with nephropathy than those without nephropathy (46.2% versus 14.8%, P < 0.05). The values of eGFR in patients with renal involvement were significantly increased after treatment with full dose prednisone and CTX for one month (73.5±36.5 ml/min/1.73m2 versus 102.0±35.3 ml/min/1.73m2, P<0.05). Proteinuria and hematuria in CSS patients were decreased considerably after treatment. Conclusion Renal involvements in CSS patients are not rare, the renal abnormalities are relatively mild and response well to the corticosteroid and CTX therapy. The renal impairment in CSS patients may be related to the ANCA, particularly antibodies against MPO.

Advances in the Application of Proteomics in Cervical Carcinoma Research

Li LUAN Yu-feng CHENG

2011, 31(7):  839-842. 

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Proteomic technology has been widely used in the research of medical sciences, including the study of the pathogenesis of carcinoma, discovery and identification of novel biomarkers and evaluating the response to treatment, as well as in the application of early diagnosis, clinical pathologic stages and screening therapeutic targets in cervical carcinoma. In the aspect of cervical carcinoma, many reports have demonstrated the differently expressed proteins from the proteome level, and these results provide reliable theoretic base for well understanding of the diagnosis and treatment in cervical carcinoma.

The research on Fluorescence in situ hybridization in non-urothelial tumors of urinary system

De-xin DONG Han-zhong LI

2011, 31(7):  843-846. 

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As a new technique, fluorescence in situ hybridization in non-urothelial tumors of urinary system has been widely used, and has got some results: The identification of benign and malignant adrenal tumors; pathological type of renal tumor, metastasis and prognosis of renal tumors; the pathogenesis of Wilms tumor; the gene diagnosis of VHL syndrome; the pathogenesis and prognosis of prostate cancer; the gene diagnosis of testicular cancer and other tumor.