Table of Content

25 March 2008, Volume 28 Issue 3

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研究论文

Decreased phosphorylation of cPKCγ-interacting ERK1/2 in the hippocampus of hypoxic preconditioned mice

Nan ZHANG; Song HAN; Jun-fa LI

2008, 28(3):  209-212. 

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Objective To determine the changes in phosphorylation of conventional protein kinase Cγ (cPKCγ)-interacting extracellular signal regulated kinase 1/2 (ERK1/2) in the hippocampus of hypoxic preconditioned (HPC) mice. Methods Healthy male BALB/c mice were used to develop "auto-hypoxia"-induced HPC mice and randomly divided into 3 groups as follows: normoxic control (H0), early (H3) and delayed hypoxic preconditioned groups (H6). Co-immunoprecipitation and Western blot were applied to quantitatively analyze the phosphorylation levels of cPKCγ-interacting ERK1/2 in cytosolic and particulate fractions of hippocampus of HPC mice. Results ERK1/2 could be co-immunoprecipitated by cPKCγ in the hippocampus of mice. The phosphorylation levels of cPKCγ interacting ERK1/2 at tyrosine 204 decreased significantly in the hippocampus of H3 and H6 groups when compared with that of the H0 group (P<0.05). Conclusion cPKCγ could interact with ERK1/2 in the hippocampus of mice, and their interaction might be involved in the development of cerebral hypoxic preconditioning of mice.

Injury role of sulfur dioxide on myocardium ischemia and reperfusion rats

Su-qing ZHANG; Jun-bao DU; Hong-fang JIN; Bin GENG; Shu-kui LI; Chun-yu ZHANG; Chao-shu TANG

2008, 28(3):  213-216. 

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Abstract: Oxidative stress and calcium over loading are thought to be important mechanism of ischemia and reperfusion (I/R) injury and main target of prevention and cure on studies for I/R injury. However, so far, mechanism of I/R injury yet is not completely clear. Sulfur dioxide (SO2) is a common outdoor air pollutant and is associated with day to day changes in hospitalization rates for lung disease. Our group found that SO2 could relax vascular smooth. We presume SO2, especially; endogenous SO2 may have important significance in physiology and pathology in I/R injury. In this a study, we investigated the action of SO2 on I/R hearts of rats. Methods: Hearts of rat was divided in three groups, I/R group; SO2 group; and HDX group. The Langendorff-prepared rat hearts model was used. The heart was under ischemia for 2 hours under hypothermic (40C) subsequent reperfusion/rewarming for 60min. Heart function was monitored by MacLab system for data collection. Results: In SO2 group recovery of heart function（LVP, ±dp/dtmax, HR, CF）decreased compared with that of group of I/R (p<0.05 or p<0.01); In HDX group recovery of heart function（LVP, ±dp/dtmax, HR, CF）increased compared with that of group of I/R (p<0.05 or p<0.01); In SO2 group activity of lactate dehydrogenase (LDH), Creatinekinase (CK), glutamic oxaloacetic transaminase (GOT), and content of myohemoglobin (Mb) in coronary flow (CF); content of Malonedialdehyd (MDA) and conjugated diene (CD), and GOT activity in tissue of cardiac muscle increased compared with I/R group (p<0.05 or p<0.01). In HDX group activity of LDH, CK, GOT, and content of Mb in CF; content of MDA and CD, and GOT activity in tissue of cardiac muscle decreased compared with those of I/R group (p<0.05 or p<0.01). In SO2 group content of glutathione (GSH) decreased compared with I/R group (p<0.05); In HDX group content of GSH increased compared with that of I/R group (p<0.05). In SO2 group content superoxide anion (O2-.), and hydroxyl radical (.OH) were produced by tissue of cardiac muscle, content of hydrogen peroxide (H2O2) and activity of superoxide dimutase (SOD) in tissue of cardiac muscle did not change compared with those of I/R group (p>0.05); In HDX group O2-., .OH were generated by cardiac muscle tissue, content of H2O2 and SOD activity in cardiac muscle tissue did not differ from those of I/R group (p>0.05). Conclusions: (1) endogenous SO2 was involved in isolated heart I/R injury of rat, and this injury was aggravated by exogenesis SO2. (2) Endogenous SO2 produced by tissue possibly increased in isolated heart I/R model of rat. (3) The fact that SO2 damage cardiac muscle tissue was through at least partly through increasing lipid peroxide of cardiac muscle tissue. (4) The fact that SO2 increased lipid peroxide of cardiac muscle tissue was through at least partly through decreasing GSH produced by cardiac muscle.

Phosphorothioated bFGF oligonucleotides inhibits proliferation of HepG2 and Hep2 cells in vitro

Hong-liang HUANG; Jun-jian XIANG; Yong TANG; Hong WANG; Ning DENG; Ying-jie JIN

2008, 28(3):  217-221. 

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Objective to explore effects of bFGF on proliferation of human hepatoma HepG2 and larynx carcinoma Hep2 cell lines by bFGF phosphorothioate oligonucleotides. Methods Oligonuleotides were transfected into HepG2 and Hep2 cell lines with jetPEI (polyethyleneimine). The relative cell proliferation was from MTT analysis. Location of FITC-labeled phosphorothioate oligonucleotides was observed with fluorescence microscope, and positive rates of tumor cells transfected with antisense/sense phosphorothioate oligonucleotides were analyzed by Flow CytoMeter. Results Positive cells transfected by FITC conjugated bFGF phosphorothioate oligonuleotides were above 90%, and above mean 3000 FITC were detected in each cell by FACS and mainly located in nuclei. Antisense phosphorothioate oligonuleotide inhibited the growth of HepG2 and Hep2 cells by 46.6% and 25.5% respectively, equivalent to that corresponding antisense phosphodiester oligonuleotide did by 50.6% and 34.6%. Mismatch nucleotide in antisense phosphorothioate oligonuleotide reduced it's inhibition to 17.9％ and 10.4％ respectively to HepG2 and Hep2 cells. Sense phosphorothioate oligonuleotides with inhibited the growth of HepG2 and Hep2 cells by 70.6% and 67.8% respectively in a dose-dependent manner, more efficiently than antisense phosphorothioate oligonuleotides. However, sense phosphodiester oligonuleotides just affected the growth of HepG2 and Hep2 cells by 19.7% and 9.3% respectively. And inhibition of sense phosphorothioate oligonuleotide didn't be reduced by mismatch nucleotide. Conclusions These findings suggested that phosphorothioate modification didn't significantly influence antisense oligonuleotide binding to nucleic acid and inhibiting tumor cell proliferation in a antisense manner, but sense oligonuleotide with phosphorothioate modification biding to bFGF or other proteins reduces tumor cell proliferation.

Establishment of PC12 cell lines stably suppressing the DAPK expression

Xia GUO; Ya-jun WANG; Hai-tao ZHANG

2008, 28(3):  222-226. 

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Aim To establish the stable inhibition of DAPK gene expression PC12 cell lines and observate the character Methods 5 pairs of shRNA were designed by using web soft ware, then they were synthesized and inserted into the pDsRed1-N1-U6 vector. The recombinant plasmids were transfected into PC12 cell and screening the monoclone by G418 and establishing the stable PC12 cell lines. DAPK expression was detected by Western blot. MTT, FCM assay were used to assess the cell lines growth. Results The recombinant plasmids were successfully constructed and transfected into PC12 cell, Western blot showed that the RNA interference effect of the pDsRed1-N1-U6-F2 was the best among them. Conclusion It is a feasible way to establish the stable inhibition of DAPK gene expression PC12 cell lines by shRNA expression vectors.

Collagen remodeling and expression of MMP-2,MMP-9,TIMP-2 in chronic hibernating myocardium of rabbits

Yan CHEN; Fu-qing SUN; Shu-mei LI; Fei-long ZHANG; Liang-long CHEN

2008, 28(3):  227-231. 

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Objective To observe the interstitial collagen and the expression of MMP-2,MMP-9 and TIMP-2 after chronic hibernating myocardium(CHM) in rabbits. Methods Twenty-seven rabbits were assigned to the following two groups: chronic hibernating myocardium group(CHM，n=15)， sham-operatioin group(SHAM，n=12). The interstitial collagen volume fraction(ICVF) and the changes in the type and proportion of collagen fibers were observed. The expression of MMP-2,MMP-9 and TIMP-2 was assessed by immunohistochemistry and western bolt. Results Comparing with the SHAM group, ICVF was significantly increased（P < 0.01）; the ratio of collagen I to Ⅲ and collagen ratio score(CRS) were greatly increased (P < 0.01); the expression of MMP-2 and MMP-9 were markedly increased （P < 0.01）in CHM group, while TIMP-2 was greatly decreased（P < 0.01）. Conclusions During the progress of CHM, the balance of MMPs/TIMPs tended to disorder , and this may be one of the potencial mechanisms of interstitial remodeling .

Role of transforming growth factor-?1 tnducing myofibroblast formation in hypoxic pulmonary vascular remodeling

Bo-wan LI; Yong-liang JIANG; Ai-guo DAI

2008, 28(3):  232-237. 

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Objective To investigate transforming growth factor-┈1(TGF-┈1) induces myofibroblast formation in hypoxic pulmonary vascular remodeling of rats. Methods Forty male Wistar rats were exposed to hypoxia for 0, 3, 7, 14 or 21days. Mean pulmonary arterial pressure (mPAP), vessel morphometry, right ventricle hypertrophy index (RVHI) were measured. Immunocytochemistry were used to measure the expression of メ-smooth-muscle actin(メ-SMA) and TGF-┈1 in pulmonary artery walls, however, in situ hybridization were used to measure the expression of TGF-┈1 mRNA in pulmonary artery walls. Ultrastructural characteristics of cell phenotype in alveolar wall vessels were observed by electron microscopy. Cell culture observed human embryonic lung fibroblasts(KMB17) phenotype after induction of hypoxia and TGF-┈1. Results (1) mPAP increased significantly after 7-day of hypoxia[(18.41【0.37)mmHg,P<0.05]. Pulmonary artery remodeling index and right ventricle hypertrophy became evident after 14-day of hypoxia. (2) The distribution of nonmuscular, partially muscular, and muscular vessels(39%、46%、15% ) is significantly different （PX2＜0.005＝ from hypoxia 7d than that in the control group (60%、35%、5% ).（3）The intra-acinar pulmonary arteries with cells expressing メ-SMA increased most with hypoxic time by immunocytochemistry. TGF-┈1 mRNA expression in pulmonary arterial walls was increased significantly after 14 days of hypoxia(0.385【0.028, P<0.01),but showed no obvious changes after 3 or 7 days of hypoxia. In pulmonary tunica adventitia and tunica media, TGF-┈1 staining was poorly positive in control rats, but was markedly enhanced after 3 days of hypoxia(0.198【0.031,P<0.01), reaching its peak after 7 days of hypoxia(0.267【0.035,P<0.01). (4) The myofibroblast phenotype was confirmed by electron microscopy, which revealed microfilaments and a well-developed rough endoplasmic reticulum.(5) Cell culture showed that hypoxia induced KMB17 phenotype transforming and TGF-モ1 accelerated its transforming.Conclusion It is reasonable to believe that transforming growth factor-┈1(TGF-┈1) induces myofibroblast formation are the important causes in hypoxic pulmonary vascular remodeling of rats.

Expression of phosphorylation of p27kip1 positively correlated with malignancy of Non-Hodgkin's Lymphomas

2008, 28(3):  238-242. 

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Objective To investigate the expression of the phosphorylated P27kip1 and correlation with Non-Hodgkin's lymphomas. Methods The expressions of the phosphorylated P27kip1 was detected in lymphoid nodes by immunohistochemical technique. Results The average labeling indexs (LIs) of phosphorylated P27kip1 in NHLs were significantly higher than those in benign lymphoid nodes (outside of the germinal center). The expression of phosphorylated P27kip1 in aggressive lymphomas was higher than that in indolent group; and the expression of phosphorylated P27kip1 was significantly correlated with expression of Ki-67. The expression of Thr187 phosphorylated P27Kip1 was significantly correlated with expression of P27kip1 while the Ser10 phosphorylated P27kip1 negtively correlated with the expression of P27kip1. Conclusions The high expression of phosphorylated P27kip1 plays key roles in the occurrence and development of NHL; phosphorylated P27kip1 LI and P27kip1 LI could be good parameters to determine the malignance of NHL.

Homocysteine downregulates the estrogen receptor alpha expression in aorta of ovariectomized rats by methylation modification

Jun-xu LIU Jing-ge ZHANG

2008, 28(3):  243-246. 

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Objective To observe the effects of homocysteine on the expression of estrogen receptor alpha (ERα) mRNA and the CpG island methylation of ERα gene promotor region in rats. Methods The serum levels of homocysteine in rats were measured by reversed- phase high performance liquid chromatography. RT-PCR and methylated specific PCR (MSP) assayed the mRNA expression and the methylation status in promoter CpG island of ERα gene in aorta and cultured rat aortic smooth muscle cells (RASMCs) respectively. Results The homocysteine levels were increased in ove and ove＋H groups (P<0.05 or P<0.01). The mRNA expression of ERα decreased in both ove and Hcy groups，while increased in cells treated with 5'-Aza. The methylation of CpG island in promoter region of ERα gene in aorta increased approaching 30% in ove group and 60% in ove+H group compared with 20% in sham group. The CpG island in ERα gene promotor region was completely demethylated in control RASMCs, and Hcy induced hypermethylation, while 5'-Aza produced demethylation. Conclusion Homocysteine downreglates the estrogen receptor alpha expression in aorta of ovariectomized rats by methylation modification manner.

Recombinant human glia maturation factor gamma induced neural stem cells in vitro into astrocytes

Xiao-ping LIAO; Zhang-liu CHEN; Shen-tao LI

2008, 28(3):  247-251. 

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Objective To investigate the function of human glia maturation factor gamma to induce the proliferation and differentiation of neural stem cells into astrocytes. Methods The neural stem cells were isolated from the cerebral cortex of 11-to 12-day embryoes of SD rats. After three passages of culture of the cells, the cells could react with nestin antigen, which demonstrated that the cells were neural stem cells. The neural stem cells were divided into 4 groups, control group with only medium, FBS group with culture medium and FBS, GMFG 5μg/L group with medium and 5μg/L of GMFG, GMFG 10μg/L group with medium and 10μg/L of GMFG. Results In the control group, the cells had no distinct differentiation within two days and slight branches were observed till the fourth day. In the FBS group, the cells differentiated into astrocytes within three days, which grew on the surface of the culture flasks. In the GMFG group, the cells differentiated into astrocytes within one day, and the differentiation of cells from the 10μg/L group was more obvious than that of cells from the 5μg/L group. The cells from the 10μg/L group reacted weakly with GFAP antigen. Conclusion GMFG could induce the proliferation and differentiation of neural stem cells into astrocytes effectively in vitro.

Association of Lys656Asn variant in leptin receptor gene with serum triglyceride level in obese population

Jin LU; Da-jin ZOU

2008, 28(3):  252-255. 

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Objective To investigate the association of Lys656Asn polymorphism in leptin receptor gene with serum triglyceride level. Methods The genotypes of Lys656Asn polymorphism in LepR gene were determined by polymerase chain reaction- single strand conformation polymorphism (PCR-SSCP) assay in 232 unrelated subjects of Shanghai Chinese population (including 130 subjects with simple obesity and 102 normal controls). The clinical data were also analyzed. Results No significant difference of genotype frequency of Lys656Asn polymorphism in LepR gene was observed between obese and non-obese group. In obese group, "G" Allele was associated with increased triglyceride level (1.74±0.91 vs 0.95±0.32, P<0.05). Logistic regression analysis showed that this gene polymorphism was independently associated with serum triglyceride level (P=0.026). Conclusion Lys656Asn variant in LepR gene is associated with serum triglyceride level in Shanghai Chinese obese population.

Comparative proteomics study of the kidney in lupus nephritis mouse

Meng-ru SHI; Quan LIN; Jian-xin LV

2008, 28(3):  256-259. 

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Objective To study the differential proteome of kidney between lupus nephritis mouse and normal mouse. Methods The proteins of kidney were separated by two-dimensional gel electrophoresis(2-DE). The gels stained by silver were scanned by ImageScanner and analyzed by PDQuest software. Results The experiments were repeated for three times. About 573±52 and 658±43 protein spots were found in the three maps of control group and LN group respectively; the match ratio is 83% and 87% respectively. Analyzed by PDQuest software,114 spots were found increased for two folds through comparing LN group to control group,and 140 spots were found decrease for 0.5 folds. Conclusions A significant differences in protein expression of LN mouse kidney,which may be related to the etiopathogenesis of LN,had been found.

Transcription factors Foxo3a expression on adipose tissue of KKay diabetic mice and the effects of treatment with rosiglitazone and metformin

Jing-bo ZENG;

2008, 28(3):  260-263. 

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Objective To detect the expression of Foxo3a in adipose tissue from KKay diabetic mice and the effects of treatment of rosiglitazone and metformin on FoxO3a expression in adipose tissue. Aim to understand what the role they play in the mechanism of insulin resistance. Method Study group consists of 21 KKay diabetic mice fed normal diet until 12 weeks, thereafter fed high fat diet for 4 weeks. mice were randomly divided into 3 groups one group without any treatment, one gaven rosiglitazone 12.5 mg/kg qd, one metformin 3g/kg/day bid for 4 weeks. Control group consists of 7 C57BL mice 16 week old. Dispatched the mice and took adipose tissue then put on tissue lytic solution to measure the protein concentration by Bradford method and to detect Foxo3a protein expression on adipose tissue by Western Blot with multiple colony antibody 1:1250. Quantitative analysis used by gel image analyzing instrument. Results Foxo3a was higher expressed in adipose from KKAy diabetic mice than that in control group (FoxO3a/b-actin ratio 1.76 ± 0.19 vs1.15 ± 0.10，P<0.001). Rosilitazone treatment decreased the expression of FoxO3a in adipose tissue significantly compare to KKAy diabetic mice (FoxO3a/b-actin ratio 1.43 ± 0.24, P<0.05). Metformin treatment can not significantly reduce FoxO3a expression in adipose tissue1.57 ± 0.23, compared to KKAy diabetes group. Conclusion Expression of FoxO3a in adipose tissue increased in KKAy diabetic mice. Rosiglitazone decreased the blood glucose and insulin resistance may through the pathway of FoxO3a. Metformin reduces blood glucose and insulin resistance may not related adipose FoxO3a expression.

技术与方法

Construction a Three Dimensional Culture Model for Inducing Embryonic Stem Cells Differentiation in vitro

Guo-feng WEI; Xiang-dong LI; Wei-jiang ZHANG; Peng QU

2008, 28(3):  264-268. 

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Objective To mimic the three-dimensional (3-D) growth pattern of stem cells in vivo, a 3-D culture system in vitro constructed by rat tail collagen scaffold was established. Methods Circular strands were prepared by mixing suspended mouse embryonic stem cells (mESCs) with rat tail collagen. Growth profile of mESCs within the collagen strand was observed under phase contrast microscope. Their metabolic activity was evaluated by glucose/lactic acid contents. To evaluate the effect of 3D culture system on ESCs differentiation, ES-derived cardiomyocytes were detected by immunohistochemistry, RT-PCR and transmission electron microscope (TEM), respectively. Results ESCs grew well within the 3-D culture system constructed by rat tail collagen. As time passage, cell connections can be found in those cell clusters formed within collagen stands, which indicated that tissue-like cultures should be produced during the process of 3-D culture in vitro. Further, ESCs cultured by 3-D collagen strand differentiated into cardiomyocytes spontaneously. Conclusions Collagen could provide an ideal growth matrix for ESCs proliferation in vitro and promote ESCs differentiation towards tissue-like structures. Thus, three dimensional culture system constructed by rat tail collagen could be applied to study ESCs differentiation mimicking in vivo.

临床园地

Clinical application of ultrasound-guided technique in double-injection interscalene brachial plexus block

Xu-lei CUI; Zhong-huang XU; Shao-hui CHEN; Hong-zhi REN; Ai-lun LUO; Yu-guang HUANG

2008, 28(3):  269-271. 

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Objective To compare the efficacy of interscalene brachial plexus block using an ultrasound-guided method with neutostimulator-guided method. Methods：Ninety ASAⅠ~Ⅱ patients scheduled for surgery of the upper extremity were randomly allocated into three groups(n=30), Control group: nerve stimulator-guided and single-injection group; US group: ultrasound-guided with nerve stimulator confirmation and single-injection group; UD group : ultrasound-guided with nerve stimulator confirmation and double-injection group. 30ml 0.5% ropivocaine was injected in each group.The patient in group UD group received half the volume of ropivocaine injected around the two target nerves. Results The average onset time of sensory blockade in the UD group was significantly shorter than in the US group and Control group .The rate of satisfactory sensory blockade in the UD group was significantly higher than in the US group and Control group. Conclusions Ultrasound-guided interscalene brachial plexus block may shorten the onset time with fewer adverse events. The ultrasound-guided double-injection method may significantly improve the quality of blockade.

研究短文

Cardioprotective role by Erythropoietin in hypoxia and reoxygenation in ventricular cardiomyocytes of rats

Yuan-jun YIN; Ting-mei YE; Qin GAO; Qiang XIA

2008, 28(3):  272-274. 

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GABA receptors mRNA expressed by human mesenchymal stem cell derived neuron-like cells

Yan XU; Ping DUAN; Su-xia WU; Xue-fei HAN; Wen-hai YAN; Ying XING

2008, 28(3):  275-277. 

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JNK kinase inhibitor suppress renal tubular epithelial cell apoptosis induced by renal ischemia/reperfusion in rats

Yan WANG; An-zhou XIA; Huai-xue JI; Shu-hua XING

2008, 28(3):  278-279. 

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Clofibrate inhibit HMG-CoA reductase expression and cholesterol efflux in rat hippocampal neuron

Shu-kun ZHANG; Ling-ling JIANG Su-ling WANG; Rong-hui YANG; Yan-ning HOU

2008, 28(3):  280-281. 

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医学教育

Approaches to construct the faculty's professionalism in medical schools

Ping LIN; Yuan-zhi GUAN

2008, 28(3):  282-285. 

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Professional development of faculty is an essential component of strategy for medical schools to construct teaching team. Development of education is dependent upon the professional quality of teaching staff. Starting from connotation of faculty's professionalism, this paper compared present status of composition of teaching staff in China with that in foreign countries, explored the potential approaches for medical schools to strengthen the construction and sustainable development of teaching staff.

短篇综述

T-bet/GATA3 and diseases

Chun-xia ZHANG; Shao-min REN

2008, 28(3):  286-288. 

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Transcription factors T-bet/GATA3 are one of the most fascinating areas of immunology today. Rencently, information has been accumulated to clarity T-bet/GATA3 roles in etiology and progression of certain diseases. This review focuses on the relationship between T-bet/GATA3 and diseases.

Recent application of G-CSF in the transplantation of stem cell

Shi-zhu JIN; Ming-zi HAN; Feng-hua PEI

2008, 28(3):  289-293. 

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Granulocyte colony-stimulating factor(G-CSF) is specific hematopoietic regulating growth factor of granulocyte lineage.It can be used to treat different kinds of granulocytopenia . Recently a variety of basic and clinical researches demonstrated that G-CSF can mobilize bone marrow stem cell and haemopoietic stem cell in the peripheral blood, which give us a new idea of releasing、enriching、mobolizing、promoting migration and inducing cell differentiation in the stem cell transplantation. The recent application of G-CSF in stem cell transplantation is summarized in this view.

Related genes about embryonic stem cell's proliferation and differentiation

Kai-jun JIANG; Ju WANG; Zong-yao ZHOU

2008, 28(3):  294-296. 

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Embryonic stem cells，which are detached from inner cell mass(ICM), can maintain self-renewal undifferentiation status and differentiate into almost every type of cells. Lots of genes take part in the regulation of embryonic stem cell's proliferation and differen- tiation. In this paper, we review the related genes about embryonic stem cell's proliferation and differentiation, meantime draw off questions which need to be further illuminated.

协和大查房

Fever, hepatosplenomegaly and eosinophilia

Huan WU; Ying GE; Xiao-qing LIU; Guo-hua DENG

2008, 28(3):  297-304. 

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