Table of Content

21 April 2009, Volume 30 Issue 2

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肾病学专题

Inhibition of Endothelium-dependent Vasodilatation by Asymmetric Dimethyl Arginine in Human Resistance Vessels

ZHANG Dong-liang;LIAO Hua;ZHANG Li;LIU Wen-hu

2009, 30(2):  121-125. 

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Objective To study the inhibitory effects of asymmetric dimethyl arginine(ADMA) on endothelium-dependent vasodilation(EDVD) of human radial arteries. Methods Vascular rings were obtained from patients under maintenance hemodialysis(MHD) during internal arteriovenous fistula operation. According to the conditions of the endothelium, the arteries were divided into two groups: the endothelium intact group, and the endothelium deprived group, in which the endothelium was extracted mechanically. The arteries were then mounted on wire myographs. Contraction of the arteries were invoked with 10^-5 mol/L phenylephrine(PE), while EDVD was caused by a dose of 10^-5 mol/L acetylcholine(ACh) for the endothelium intact group or 10^-7 mol/L sodium nitroprusside(SNP) for the endothelium deprived group. Different concentrations of ADMA(10^-7~10^-3 mol/L) were used to test its inhibitory effects on EDVE. Cumulative concentration-response curves were then constructed for AMDA. Results For the endothelium intact group, the dilatation reached 67.10%±18.63%, when the arteries were subjected to 10^-5 mol/L ACh after PE(10^-5 mol/L)treatment. The inhibitory effects of ADMA on ACh-invoked EDVD were concentration-dependent. The relative contraction rates according to the concentrations of ACh were 7.32%±8.60%(ACh 10^-7 mol/L), 20.03%±13.49%(ACh 10^-6 mol/L), 29.93%±11.78%(ACh 10^-5 mol/L), 43.30%±11.29%(ACh 10^-4 mol/L), 80.21%±18.16%(ACh 10^-3 mol/L). For the endothelium deprived group, ADMA had a weak inhibitory effect on the endothelium independent vasodilatation(EIVD) caused by SNP. There were no concentration-dependent changes according to the concentration of ADMA. Relative constriction rates were 2.76%±1.98%(ADMA 10^-7 mol/L), 2.27%±1.82%(ADMA 10^-6 mol/L), 3.38%±2.99%(ADMA 10^-5 mol/L), 3.59%±3.66%(ADMA 10^-4 mol/L), 4.16%±3.67%(ADMA 10^-3 mol/L). Conclusion The results suggest that ADMA can inhibit EDVD caused by ACh. This effect is ADMA concentration-dependent, and endothelium-dependent.

The Increasing Carotid Intima-media Thickness and the Possible Risk Factors in Patients with End-stage Renal Disease

ZHANG Kun-ying;LIU Hui-lan;DUAN Xiao-feng;LI Guo-gang;XU Feng-bo

2009, 30(2):  126-129. 

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Objective The aim of our study is to find out whether atherosclerosis of dialysis patients is effectively accelerated due to the fact that many such patients at the start of dialysis treatment. Methods Using B-mode ultrasonography, we conducted a cross-sectional study to compare intima-media thickness in the common carotid arteries(CC), in the area of bifurcation(CB) and in the proximal part of internal carotid arteries(CI) for 45 MHD patients, of which 23 patients had chronic kidney diseases not subjected to hemodialysis(CKD-non-HD) and 20 healthy volunteers of matching age. Results There were no significant differences among the IMT values of CC, CB and CI as control in dialysis patients and patients starting dialysis treatment. We also failed to find any difference in atherosclerotic risk factors, such as lipids, smoking and blood pressures between both groups. Conclusion In our study we found no differences in carotid atherosclerosis lesions between MHD patients and CKD-non-HD patients. Patients with end-stage renal disease are at high risk for cardiovascular diseases so we should intervene earlier and more actively long before dialysis treatment in order to reduce the atherosclerotic risk factors.

Association of Serum OPN and ucMGP with the Presence of Cardiovascular Calcification in MHD Patients

LI Guo-gang;LIU Hui-lan;ZHANG Kun-ying;DUAN Xiao-feng

2009, 30(2):  130-132. 

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Objective To elucidate the association of osteopontin(OPN) and the circulating inactive form of matrix Gla protein(ucMGP) with the presence of cardiovascular calcification in maintenance hemodialysis(MHD)patients. Methods Serum OPN and ucMGP in 55 MHD patients and 22 health subjects at matching age were measured using ELISA. The intimal and medial thicknesses(IMT) in carotid artery were measured by doppler ultrasonography and the cardiac valve calcification by colour doppler ultrasonography. Results The levels of OPN and ucMGP in MHD patients were significantly higher than those in healthy ones. The concentrations of OPN were positively correlated with the IMT(r=0.55, P<0.05). The levels of OPN and ucMGP were higher in cardiac calcification group. Conclusion The levels of OPN and ucMGP may be associated with the presence of atherosclerosis and valvular calcification in MHD patients,and which may be used as markers for cardiovascular calcification.

Role of Endogenous H₂S in the Accumulation of Extracellular Matrix of the Diabetic Nephropathy

HUANG Wen;LI Xiao-lu;CHEN Shen;ZHANG Hong;YANG Min

2009, 30(2):  133-136. 

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Objective To explore changes of fibronectin(FN) in the kidney tissues of diabetic nephropathy)(DN) rats, and to study the role of endogenous H₂S in the pathological development of diabetic nephropathy. Methods Twenty-eight rats(male, Sprague-Dawley) were randomly divided into four groups: control group, diabetic nephropathy group, DN+NaHS(H₂S donor) group and DN+DL-propargylglycine(PAG, H₂S synthetase inhibitor) group. Relative location and expression of FN in kidney tissues were measured respectively by Western blotting and immunohistochemistry. Results FN was positive in both the intraglomerular mesangial region and the extracellular matrix(EMC) in kidney tissues of DN group, and its expression level increased significantly. Western blotting: DN group (48 896±779) vs control group(38 531±902), P<0.01. Compared with that of DN group and DN+PAG group, NaHS significantly inhibited the over-expression of FN, Western blotting: DN+NaHS group(42 737±1 196) vs DN group(48 896±779) and DN+PAG group(48 252±1 068), P<0.01. Conclusion Endogenous H₂S can alleviate the pathological progression of DN by inhibiting the accumulation of ECM.

Clinical Feature and Survival Analysis of 174 Cases with Wilms Tumor

JIANG Ye-ping;SHEN Ying;SUN Ning

2009, 30(2):  137-141. 

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Objective To analyze the clinical features and prognosis of 174 cases with Wilms tumor, and to investigate the distribution of postoperative time of death and evaluate the survival rate of the patients at different clinical stages, in order to estimate the prognosis scientifically and improve the survival rate of the affected children. Methods A total of 174 Wilms tumor patients who received treatment in Beijing Children's Hospital between May 1997 and July 2008 were analyzed retrospectively. Kaplan-Meier method was used in the evaluation of survival rates to reveal the connections between clinical staging and prognosis. Results The age of onset below 5 years was in 77.0% of the 174 patients, while the male-female ratio was 1.72∶ 1. Most patients complained of abdominal mass, pathologically, FH constituted 96.9% of all cases, while UH constituted 3.1%; 73.3% deaths happened within 2 years, postoperatively. The 4 years survival rate postoperation was predicted to be 97.0% in patients at stage Ⅰ, 91.3% at stage Ⅱ, 81.5% at stage Ⅲ, and 70.5% was at stage Ⅳ, only 62.5% of the patients at stage Ⅴ. Conclusion Wilms' tumor is seen mostly in children at 5 years old or younger. The morbidity of male and female is almost the same. The postoperative death is usually within 2 years. Clinical staging is important for prognosis. A trend of decline in survival rates with the increase in clinical staging has been found. Stage Ⅰ patients' survival rate is significantly higher than that of stages Ⅲ-Ⅴ.

Determinants of Arterial Distensibility in Non-diabetic Hemodialysis Patients

ZHOU Yi-lun;YU Ze-xing;GUO Rui-jun;SUN Fang;MA Li-jie;PENG Li-ren;JIA Hui-min;WANG Qing-tao;CUI Tai-gen

2009, 30(2):  142-145. 

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Objective To evaluate the arterial stiffness in non-diabetic hemodialysis patients and to investigate the risk factors for arterial stiffness. Methods A total of 80 non-diabetic hemodialysis patients were recruited. The distensibility coefficient(DC) of the common carotid artery was evaluated by an ultrasonic phase-locked echo-tracking system. Serum albumin, total cholesterol(TC), high density lipoprotein(HDL), low density lipoprotein(LDL), triglyceride(TG), glucose, creatinine, high sensitive C-reactive protein(hs-CRP), calcium, phosphorus, and intact parathyroid hormone(iPTH) were determined with standard methods. Results Carotid DC was(14.47±5.19)(10^-3/kPa) in the non-diabetic hemodialysis patients studied. In stepwise multiple regression analysis, systolic blood pressure(β=-0.340, P<0.01), age(β=-0.248, P<0.05), and serum calcium(β=-0.199, P<0.05) were independently correlated with carotid DC. Conclusion Systolic blood pressure, age, and serum calcium were independent determinants for carotid arterial stiffness in non-diabetic hemodialysis patients.

Lipid Abnormalities and Risk Factors in Maintenance Hemodialysis Patients

LI Han;WANG Shi-xiang

2009, 30(2):  146-149. 

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Objective To investigate the changes of lipid profile, and explored the risk factors of lipid abnormalities in maintenance hemodialysis(maintenance hemodialysis, MHD) patients. Methods A total of 205 cases on MHD were enrolled in this study. Among them, 51 patients were below 45 years of age. The following data were collected: age, gender, dialysis duration, serum albumin, lipid profile, etc. T test, χ² test and binary logistic regression analysis were performed for statistics. Results Among the 205 MHD patients, 145 MHD patients had lipid abnormalities. The prevalence of lipid abnormalities in MHD patients was 70.7%. The main characteristics were high density lipoprotein cholesterol(HDL-C) decreasing(HDL-C<1.03 mmol/L), low density lipoprotein cholesterol(LDL-C) increasing(HDL-C<1.03 mmol/L) and triglyceride(TG) increasing(TG≥1.69 mmol/L), and their prevalence were53.2％(109/205), 28.3％(58/205) and 26.8％(55/205), respectively. At the boundary of 45 years old, all the MHD patients were divided into two groups, young(n=51) and middle-and old-aged group(n=154). Among the 51 young-aged MHD patients, 36 cases had lipid abnormalities, the prevalence of lipid abnormalities in young-aged MHD patients was 70.6%, which was no significant different as compared with middle-aged and old-aged MHD patients(χ²=0.001, P=0.979). There were no significant differences in terms of age, serum albumin, lipoprotein a, and the primary diseases of chronic renal failure, such as chronic glomerulonephritis, chronic interstitial nephritis, diabetic nephropathy, hypertensive nephropathy, lupus nephritis, polycystic kidney, obstructive nephropathy and chronic pyelonephritis, between lipid abnormalities and lipid normalities patients. The level of dialysis duration, male female ratio and kidney transplant failure ratio were significantly higher in MHD patients with lipid abnormalities than in those without lipid abnormalities. Logistic regressive analysis revealed that male, dialysis duration and transplanted kidney failure were the important risk factors of lipid abnormalities in MHD patients. Conclusion The prevalence of lipid abnormalities in MHD patients was quite high. The main characteristic were HDL-C decreasing, LDL-C and TG increasing.The prevalence of lipid abnormalities in young MHD patients was similar to middle-aged and old-aged MHD patients. Male, dialysis duration and kidney transplant failure were the important risk factors of lipid abnormalities in MHD patients.

Factors Associated with Hyperkalemia in Patients with Maintenance Hemodialysis

2009, 30(2):  150-153. 

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Objective To investigate the factors related to hyperkalemia in stable hemodialysis patients. Methods A total of 102 patients on maintenance hemodialysis were enrolled in the study. Serum potassium levels before and after hemodialysis, glucose, carbon dioxide binding-capacity were determined by standard methods; serum aldosterone levels were measured by radioimmunoassay in the morning at supine position and drug administrations(ACEI, ARB, β-receptor blocker), residual urine volume, history of diabetes mellitus were recorded. The data were analyzed by Pearson correlation test and Logistic regression analysis. Results The prevalence of hyperkalemia was 35%(36/102) in our dialysis center. Hyperkalemia was positively correlated with the dietary potassium(r=0.66, P=0.000) intake, but not with serum aldosterone level, usage of RAS blockers, and history of diabetes. In Logistic regression analysis, the dietary potassium intake was identified as a solely independent determinant for hyperkalemia. Conclusion The dietary potassium intake was an independent factor associated with hyperkalemia in maintenance hemodialysis patients.

肾病学专题

Clinical Aspects of Hemodiafiltration on Patients with Maintain Renal Replacement Therapy

YIN Dao-xin;ZHANG Dong-liang;LIU Wen-hu

2009, 30(2):  154-160. 

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Hemodiafiltration(HDF) is an extracorporeal renal-replacement technique using a highly permeable membrane, in which diffusion and convection are combined to enhance solute removal in a wide spectrum of molecular weights. HDF can improve clinical conditions and reduce morbidity compared with conventional hemodialysis. In this review, it summarizes recent views about clinical aspects of HDF, the main observational studies and ongoing randomized-controlled trials.

Endothelin-1 Promotes Human Pulmonary Artery Smooth Muscle Cells Proliferation by Up-regulation of Capacitative Ca²⁺ Entry

WANG Cong;;WANG Chen;WANG Yue-xiu;;LIU Jie;LI Ji-feng;;WANG Jun;

2009, 30(2):  161-166. 

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Objective This study is designed to investigate the possible signal transduction pathways that related the effect of endothelin-1(ET-1) on HPASMCs proliferation, namely, its effect on the Ca²⁺ signal pathway. Methods The cultured HPASMCs were divided into following two groups: the control group cultured with smooth muscle basal media (SMBM); the endothelin-1(ET-1) groups incubated with SMBM and ET-1(0.01~1 μmol/L, for 72 h). Cell number was determined with a hemocytometer. MTT assay was used to exam cell viability. ［Ca²⁺］_i was measured by calcium image. Real Time-PCR was performed to detect the expression of TRPC1 gene. Results The cell viability, cell number, CCE, and TRPC1 mRNA transcription were significantly higher after treatment of ET-1. Conclusion ET-1 induced HPASMCs proliferation via up-regulation of TRP channels expression, CCE and ［Ca²⁺］_i.

Role of Transient Potential Vanilloid Receptor 1 Channels in Hypoxia-induced Cytosolic Ca²⁺ Increase of Human Pulmonary Arterial Smooth Muscle Cells

WANG Yue-xiu;;WANG Chen;WANG Cong;;LIU Jie;WANG Jun;

2009, 30(2):  167-171. 

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Objective To investigate the role of transient potential vanilloid receptor 1(TRPV1) channels on cytosolic Ca²⁺ concentration(340 nm/380 nm ratio of the fluorescence image) of hypoxic human pulmonary artery smooth muscle cells(HPASMCs) and its possible signal pathway. Methods 4',6-diamidino-2-phenylindole(DAPI) staining and flow cytometry were used to detect the proliferation of HPASMCs under normoxia or hypoxia (3％O₂, 72 h) conditions. Cytosolic Ca²⁺ concentration(［Ca²⁺］cyt) was measured with a dynamic digital imaging system. Results The capability of HPASMCs proliferation significantly increased under hypoxic condition. Capsazepine(a TRPV1 channel inhibitor) could inhibit the HPASMCs proliferation. Hypoxia markedly increased resting cytosolic Ca²⁺ concentration and enhanced cyclopiazonic acid-induced capacitative Ca²⁺ entry(CCE) in HPASMCs. Capsazepine(10 μM) decreased CCE under hypoxia condition, but there was no effect on normoxia cultured group. Conclusion These results suggest that TRPV1 may be a critical pathway or mediator in hypoxia-induced increase of cytosolic Ca²⁺ concentration, CCE and the proliferation of HPASMCs.

Hypoxia Promotes Human Pulmonary Artery Smooth Muscle Cells Proliferation Through Ca²⁺-NFAT Pathway

LI Ji-feng;;WANG Cong;;LIU Jie;WANG Yue-xiu;;WANG Jun;WANG Chen;

2009, 30(2):  172-176. 

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Objective To investigate the possible signal transduction pathways involved in hypoxia-induced human pulmonary artery smooth muscle cell(HPASMCs) proliferation. Methods Cultured HPASMCs were divided into the following groups, the control group; the hypoxia group(cultured in 3% O₂ for 24 h, 48 h and 72 h) and the treatment groups(hypoxia for 48 h and treated with 2 mmol/L EDTA or 4 μmol/L VIVIT). Cell number was determined with a hemocytometer. ［Ca²⁺］_i was measured by calcium imaging. The translocation of NFATc3 in immunofluorescence-stained HPASMCs was detected by confocal microscopy. Results The cell number, ［Ca²⁺］_i and NFATc3 translocation were significantly higher in hypoxia-treated groups. EDTA(a chelator of Ca²⁺) or VIVIT(the specific inhibitor of NFAT) significantly inhibits the hypoxia-induced proliferation of HPASMCs. Conclusion Ca²⁺-NFAT pathway maybe involved in hypoxia-induced HPASMCs proliferation.

A Method of Isolating Endothelial Progenitor Cells from Mice

WAN Jun;;WANG Jun;XU Xiao-xue;LIU Jie;ZHAI Zhen-guo;WANG Chen;

2009, 30(2):  177-181. 

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Objective To investigate an appropriate method for isolating EPCs from mice bone marrow. Methods The mononuclear cells from mice bone marrow collected by density gradient separation were cultured in M199 medium after examination by flow cytometry. After 7 days' culture, cells were stained with DIL-acLDL and FITC-UEA-1, and tested the abilities of migration and incorporation into the vascular network. Results The spindle cells grew dominantly after 7 days' culture. About 0.029%±0.008% of total mononuclear cells detected by flow cytometry were CD34⁺CD133⁺VEGFR2⁺ cells. The percentage of cells labeled both with DIL-acLDL and FITC-UEA-1 was 86.085%±5.622%. The number of migratory cells reached 19.458±2.251/ field of view. The average cell number incorporating into the vascular network was 67.750±8.823/field of view detected by cell migration test. Conclusion The above method is a relatively ideal one for isolating and assessing EPCs from mice bone marrow, which could offer a better support to further research on EPCs.

The Biologic Significance of Wnt Signaling Pathway in Lung Disease

YU Xiao-min;;WANG Chen;WANG Jun

2009, 30(2):  182-184. 

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The Wnt signaling pathway is a highly conserved signaling pathway. It mainly consists of the canonical Wnt/β-catenin pathway and the non-canonical pathway〔the Wnt/planar cell polarity(PCP) pathway and the Wnt/Ca²⁺ pathway〕. Wnt pathway is involved in a wide range of biological functions, such as cell adhesion, migration, proliferation and apoptosis. Recently, accumulating evidence have suggested that the activation of Wnt signaling pathway is substantially involved in the pathogenesis of several pulmonary diseases, such as idiopathic pulmonary arterial hypertension, asthma, idiopathic pulmonary fibrosis and lung cancer. This review focuses on the biological significance of Wnt signaling pathway in lung diseases.

基础研究

Effect of Estrogen on PDZK1 Protein in Breast Cancer Cells

WANG Ying;YANG Xiao-mei;ZHENG Jun-fang;CHEN Peng;ZHAO Jing-jing;HE Jun-qi

2009, 30(2):  185-188. 

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Objective Exposure to estrogen is an important independent determinant factor of breast cancer. Many estrogen responsive proteins play an important role in breast cancer occurrence and development. Our study is to investigate the effects of 17β-estradiol(E2) and ICI182,780 (the estrogen receptor antagonist, Faslodex), at different concentrations and different times, on the expression level of the PDZK1(PDZ domain containing 1) protein in ER-MDA-MB-231, MCF-7 cell lines which were ER(+) and MDA-MB-231 cell lines which were ER(-). Methods MDA-MB-231, ER-MDA-MB-231 and MCF-7 cell lines were all incubated for 48 hours in RPMI 1640 medium which was free of phenol red supplemented with 10% charcoal-stripped fetal bovine serum, which aimed to remove the estrogen effects of fetal bovine serum and estrogen-like effects of phenol red in the medium respectively. Western Blot was used to analyse the expression levels of PDZK1 protein in MCF-7 and ER-MDA-MB-231, two ER(+) breast cancer cell lines, at different time points with different doses of 17β-estradiol and ICI182,780. The ER(-) cell line, MDA-MB-231, was used as control. Results Physiological 17β-estradiol could obviously up-regulate PDZK1 protein expression level in ER(+) breast cancer cell lines. The Western Blot analysis showed that after the treatment of 17β-estradiol at the concentration of 10^-8 mol/L for 72 hours, the PDZK1 protein expression level was significantly up-regulated in the two strains of MCF-7 and ER-MDA-MB-231 breast cancer cell lines, which was time and dose dependent. Compared with 17β-estradiol, the effects of ICI182,780 was the exactly opposite: after the treatment with ICI182,780 at the concentration of 10^-6 mol/L for 36 hours, the PDZK1 protein expression level was suppressed in ER-MDA-MB-231 significantly. The ICI182,780 effects were obvious when the concentration was 10^-5 mol/L. In MDA-MB-231 breast cancer cell line which was ER(-), the expression level of PDZK1 did not change as much as the ER(+) breast cancer cell lines, when treated with the same concentration of 17β-estradiol and ICI182,780 at the same time. Conclusion Our study demonstrates that the PDZK1 protein is estrogen responsive. The expression level of PDZK1 protein in ER(+) breast cancer cell lines was mediated through ER signaling transduction pathway. 17β-estradiol and ICI182,780 may affect the cell growth and the drug resistance by regulating the expression level of PDZK1. This finding may provide a new theoretical basis for ER(+) breast cancer endocrinotherapy.

Location of DNaseⅠHypersensitive Site in the Promoter of Egfr Gene

LI Shan-shan;LI Xiang-hui;LI Yan;ZHAO Bo;ZHANG Xiu-fang;WANG Ze-sheng;ZHANG Yu-xiang

2009, 30(2):  189-194. 

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Objective To predict the binding sites of transcription factors in DNaseⅠ hypersensitive site(DHS) of epidermal growth factor receptor(EGFR) gene promoter and to elucidate the regulation mechanism of egfr gene transcription, we used a new molecular technique to locate the precise DNaseⅠ cutting sites. Methods Cervical cancer cell line HeLa and breast cancer cell line MDA-MB-231, which are both positive for EGFR were tested; while leukemia cell line K562, which is EGFR negative, was used for the negative control. The HeLa, MDA-MB-231 and K562 cell nuclei were all treated with DNaseⅠ at the concentrations of 5 kU/L and 10 kU/L separately. Then ligation-mediated PCR(LM-PCR) technique was performed as follows. DNA extracted from DNase Ⅰ-treated or non-treated cell nuclei were filled-in to form blunt ends, and were ligated to the adaptors. The target DNA was then amplified by nest PCR, using the adaptors-specific primers and egfr promoter-specific primers. DHSs were predicted by the length of nest PCR products. Ten clones from each PCR products were randomly picked up and sequenced to confirm that the products were from egfr gene promoter. After analysis of the sequencing results, the precise DNaseⅠ cutting sites in the promoter of egfr gene were obtained. To further predict the binding sites of the transcription factors in DHS, we used bioinformatics software called transcription element search software(TESS) to analyze potential transcription binding sites on DHS. Results After the analysis of sequencing results, we have detected eight different precise DNaseⅠ cutting sites in the egfr gene promoter of HeLa and MDA-MB-231, but no specific DNaseⅠ cutting sites was found in the egfr gene promoter of K562. In the egfr gene promoter of HeLa and MDA-MB-231, the DHS distribution was similar, mainly located from 300 bp to 500 bp (-300 bp~-500 bp ) upstream of the transcription start site, which may be the key regulation region of the egfr transcription. Results showed that 16 transcription factors possibly bind to DHS in the promoter of egfr gene in the HeLa and MDA-MB- 231 cell lines. Among these transcription factors predicted by TESS, three of them, namely Sp1, TCF and YY1 were already reported by others. In our experiment, we found eight DNaseⅠ cutting sites in the egfr gene promoter in the HeLa and MDA-MB-231, but no specific site was found from the K562 cells. The results showed that in the DHS of EGFR⁺ cells, the chromatin conformation were looser and more accessible to the transcription factors, but which were not in the EGFR^- cells. By analyzing the sequence of DHS, we predicted more potential transcription factors possibly binding to DHS in the promoter of egfr gene. Conclusion We have established a simple method to detect the DHS in gene promoter region. Meanwhile, we can obtain more precise DHS location using this method. This method will be useful for the research of the regulation mechanism of specific gene transcription.

Construction of Eukaryotic Expression Vector pcDNA3.1(+)-CYP2J3 and Its Expression in Myocardial Cell of Rats

MA Li-quan;ZENG Xiang-jun;WANG Hong-xia;YOU Hong-jie;LU Ling-qiao;ZHANG Li-ke;ZHENG Shao-peng

2009, 30(2):  195-198. 

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Objective To clone rat CYP2J3 gene and recombine eukaryotic expression vector, pcDNA3.1(+)-CYP 2J3, transfect the vector by FuGENE HD transfection agent, and measure its expression level in myocardial cells of rats. Methods The total RNA was extracted from rat liver. The cDNA of CYP2J3 was amplified by RT-PCR and OE-PCR, and inserted into pBS-T vector. The recombinants were checked by PCR and digestion of restriction endonuclease. The CYP2J3 gene was confirmed by DNA sequencing and cloned into eukaryotic expression vector of the pcDNA 3. 1(+). Resulting in pcDNA3.1(+)-CYP 2J3. Cultured myocardial cells of rats were transfected with pcDNA 3. 1(+)-CYP2J3 by FuGENE HD transfection agent. RT-PCR was used to detect the expression of pcDNA 3. 1(+)-CYP2J3. Results CYP2J3 gene was obtained by RT-PCR and OE-PCR. The recombinant eukaryotic expression vector for CYP2J3 gene had been successfully constructed. The transfected myocardial cell of rats could express CYP2J3. Conclusion Successful cloning of CYP2J3 gene construction of pcDNA3. 1(+)-CYP2J3, and lasting expression in cultured myocardial cells of rats at 24 h, 48 h and 72h after transfecting pcDNA3. 1(+)-CYP2J3 may provided the foundation for the further study of CYP2J3 gene, especially its function study in cells.

Texture Characteristic Comparison between Patients with Multiple Sclerosis and Those with Alzheimer's Disease Based on MR Imaging

ZHAO Yuan-yuan;TONG Long-zheng;JIANG Gui-lian;YU Chun-shui

2009, 30(2):  199-202. 

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Objective Multiple sclerosis(MS) is a chronic inflammatory disease of the central nervous system, which leads to severe progressive neurological dysfunction. Alzheimer’s disease(AD) is a severe neurodegenerative disease of the central nervous system. The previous researches shown that there were significant differences between MRI texture characteristics of MS and normal control(NC). To compare texture characteristics of MS and other cerebral parenchyma diseases, the differences of cerebral parenchyma texture characteristics on MR images between patients with Multiple Sclerosis(MS) and Alzheimer’s disease(AD) were analyzed. Methods In this research, the total sample set was 39 with MR images of group AD, group MS and group NC 13 for each. Texture analysis of gray level co-occurrence matrix was performed on MR T1-weighted images, and the region of interest(ROI) was cerebral parenchyma. Texture characteristics, such as energy, contrast, entropy and inverse different moment were calculated in order to test if there was a significant difference between these three groups. Results Monotonic trend of texture values existed in AD, MS and NC group the mean values were 0.002 2 vs 0.003 8 vs 0.008 3 for energy, 2.863 3 vs 2.678 9 vs 1.317 8 for entropy, 47.524 1 vs 44.396 1 vs 36.809 0 for contrast, 0.221 7 vs 0.249 8 vs 0.338 3 for inverse different moment. There were significant differences for energy, entropy and inverse different moment between every two groups of AD, MS and NC. Concerning contrast, there was a significant difference between groups AD and NC, groups MS and NC, but not difference between AD and MS. Conclusion The texture differences between MR image of MS and NC could reveal the pathological changes of brain substance for patients with MS, which may be helpful to aid diagnose of MS. At the same time, the texture difference may reveal pathological differences between AD and MS, which may be helpful to differentially diagnose AD, MS and other pathological changes. The data of our subjects is limited, so a larger sample set maybe require to confirm the above conclusion in further studies.

The Methylation Status and Expression of MGMT and hMLH1 in Glioma and Their Clinical Significance

CHEN Hui-yuan;JIANG Tao;Yuan Fang;CUI Yun;LI Gui-lin;HUANG Lei

2009, 30(2):  203-207. 

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Objective To explore the methylation status and expression of hMLH1 and MGMT in glioma, and the role of them in the glioma oncogenesis. Methods Methylation status of the MGMT and hMLH1 promoter regions was assayed in 78 glioma tissues by methylation-specific PCR(MSP). The expression of MGMT and hMLH1 in the glioma was investigated by immunohistochemical technique using monoclonal antibody. Results Methylation in the MGMT and hMLH1 promoter regions was found in 43(55.1%) and 32(41.0%) cases respectively. The median of expression rates of MGMT and hMLH1 in tumor cells were 15.9% and 84.6% respectively. Methylation status in the promoter region showed no obvious correlation with the percentage of expression in tumor cells, and the sex, age, and pathological type. Conclusion As proteins of DNA repair system, the expression of MGMT is much lower than that of hMLH1, suggesting that a number of different pathways may exist in gliomas which is associated with disturbances of DNA repair system.

Content and Dissolution Characteristics of Trace Elements in Ornithogalum Caudatum Ait and Three Anticancer Chinese Herbs

BAI Jin-fa;LIU Zhi-qiang;ZHANG Rui-ping;SONG Feng-rui;LIU Shu-ying

2009, 30(2):  208-211. 

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Objective To provide basis for the anticancer effect of Ornithogalum Caudatum Ait, the content and dissolution rates of 16 elements in Ornithogalum Caudatum Ait and three anticancer Chinese herbs(Herba Hedyotis Diffusa, Herba Scutellariae Barbatae and Radix Sophorae Tonkinensis) were investigated and compared. Methods Samples were analyzed by inductively coupled plasma atomic emission spectrometry(ICP-AES), atomic absorption spectrometry(AAS) and atomic fluorescence spectrometry(AFS). Results 1 There were many essential elements in Ornithogalum caudatum Ait and the content of Ca, Mg, Fe, and Zn were higher than those of others; 2 The ratios of Cu/Zn in water extract of Ornithogalum Caudatum Ait and the three anticancer Chinese herbs were much lowe than the normal value in human serum; 3 The differences in the contents of Se in water extracts of four Chinese herbs were very small and the contents were low. The contents of Ge in four crude herbs and their water extracts were very low. Conclusion The anticancer and anti-inflammatory effects of Ornithogalum Caudatum Ait is probably achieved by adjusting the balance among Cu, Zn, and Ca elements. The results provide basis for further analysis of the relevance between the elements in Ornithogalum Caudatum Ait and its therapeutic effect.

Resveratrol on the Growth of Human Skin Cancer Cell Line A431 and Its Underlying Mechanism

JIAO Hui-qin;WANG Rui;SUN Hai-mei;LIU Yong;GUO Min;SU Hong-xing;ZHOU De-shan

2009, 30(2):  212-217. 

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Objective To investigate the effects of Res on the growth of skin cancer cell line A431 and benign epidermal keratinocyte cell line HaCaT in human and its mechanism. In addition, to evaluate the potential value of Res as a natural antitumor drug. Methods A431 and HaCaT cells were incubated in HDMEM medium containing 10% serum and 1% SP, then they were each treated with different concentrations of Res and DDP. MTT assay was used to measure the inhibitory rate. Phase contrast microscope was used to observe the morphological changes of A431 cells treated with Res. The effects of Res on A431 cell apoptosis was evaluated by flow cytometry, the expression of Caspase-3 and Bcl-2 proteins were analysed by SP immunohistochemistry. Results 1) The results of MTT assay showed that both Res and DDP could inhibit A431 cell proliferation in time-dependent and dose-dependent manners. At their respective IC50, the inhibitory rate of DDP was higher than that of Res on HaCaT cells(P<0.05). 2) Inhibitory effects of Res on the growth of A431 cells could be seen under Phase contrast microscope, with the increase in Res concentration, the adhesion ability of A431 cell decreased, many cells shed from the vessel wall and floated on the culture medium. Flow cytometry detected that typical apoptosis took place in A431 cells, when treated with 15 mg/L and 20 mg/L Res for 48 h, the apoptosis rate of A431 cells showed a dose-dependent manner, When treated with 15 mg/L for 24 h, 48 h and 72 h, the apoptosis rate of A431 cells showed a time-dependent manner; Compared with the control group, after treatment with Res, the expression of caspase-3 increased and Bcl-2 decreased in A431 cell. Conclusion Res, which is less toxic than DDP, can inhibit proliferation and induce apoptosis of A431 cell. The possible molecular mechanisms of apoptosis might be related to the increasing expression of caspase-3 and the decreasing expression of Bcl-2.

Experimental Research of the Relationship between Insulin Resistance and Non-alcoholic Fatty Liver Disease

HU Zhi-tian;CHE Nian-cong;XIA Rong;YUAN Meng

2009, 30(2):  218-221. 

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Objective To observe the insulin resistance(IR) changes in non-alcoholic fatty liver disease(NAFLD), and to investigate the relationship between IR and NAFLD. Methods Experimental animal models of NAFLD were established, which were treated with insulin sensitizer-Metformin and Rosiglitazone. Fasting blood glucose and fasting insulin were tested by the end of the second week, fourth, sixth and eighth week. Insulin resistance index and insulin sensitivity index were calculated. Results There wasn't a significant difference between the fasting blood glucose of each group by the end of week four, NAFLD was formed in model group and treatment group. In the first two weeks, IR was not found in the model group. But by the end of the fourth week, sustained peripheral IR was found in this group. IR was not found in the treatment group in the first four weeks, but the results in the end of sixth week and eighth week showed that sustained IR was found in this group. Conclusion IR is not the (only) factor in forming NAFLD, maybe it is the result of NAFLD. The function of Insulin sensitizer is not reliable to pathological changes in liver, the drug can not prevent the formation of NAFLD. These results require further in-depth clinical and experimental research.

Expression of Inducible Nitric Oxide Synthase and Heme Oxygenase-1 in Pulmonary Intravascular Macrophages of Rats with Hepatopulmonary Syndrome

GAO Jun;WANG Yu;ZHANG Zhong-tao;LI Jian-she;MA Xue-mei;ZHAO Li-zhen

2009, 30(2):  222-226. 

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Objective To explore the role of pulmonary intravascular macrophages(PIM ) in the pathogenesis of hepatopulmonary syndrome(HPS). Methods Sprague-Dawley(SD) rats were divided into two test groups, control group and CCl₄ group. Arterial blood was collected for measurement of blood gases, venous blood for hepatic function and endotoxin levels. Pathology of liver and lung was done. The mesenteric lymph nodes were dissected for bacteriology studies. Immunolocalization of macrophages was performed using monoclonal macrophage antibody ED1 on paraffin sections. Proteins of iNOS and HO-1 of lung tissue were examined by immunohistochemistry. By real-time polymerase chain reaction(PCR) using SYBR Green Ⅰ, the expression levels of iNOS and HO-1 mRNA in lung tissues were measured. Results There was no inflammation, edema, fibrosis, alveolar collapse and hyaline membrane formation in the lung control group. All the lungs from CCl₄ group showed widened alveolar wall architectures, widespread dilatation of alveolar capillaries and decreased alveolar volume. PaO₂(81.39±2.36)mmHg and PaCO₂(30.55±2.87)mmHg decreased significantly in CCl₄ group compared with(98.99±3.41)mmHg and(36.26±2.88)mmHg in the control group(P<0.05). Alveolar-arterial oxygen difference(A-aDO₂) increased significantly in CCl₄ group(30.79±5.73) as compared with the control(3.79±0.86) (P<0.05). Endotoxin level increased significantly in CCl₄ group(0.30±0.13)ZU/L compared with control(0.05±0.02)ZU/L(P<0.05). Culture-positive mesenteric lymph nodes were found in 62.5% of CCl₄ group. There were not Culture-positive mesenteric lymph nodes in the control. All lungs from CCl₄ group showed accumulation of large mononuclear macrophage-like cells within the lumen of numerous small muscular and nonmuscular pulmonary vessels. Using immunohistochemical analysis, iNOS and HO-1 expressions were localized to PIM in CCl₄ group. The mRNA expression of iNOS(0.03±0.01) and HO-1 (0.16±0.04) increased significantly in CCl₄ group than(0.01±0.01) and(0.07±0.02) in the control(P<0.05). Conclusion Cirrhosis results in bacterial translocation from the gastrointestinal tract and endotoxemia. High concentrations of endotoxin in the pulmonary bloodstream activate PIM. PIM aggregates and expresses iNOS and HO-1. Thus the increasing amount of NO and CO released in the lungs results in HPS.

临床研究

Vascular Calcification in Rat Models of Type 2 Diabetic Nephropathy

WANG Ying;LIU Hui-lan;AN Na

2009, 30(2):  227-230. 

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Objective To investigate the vascular calcification and its risk factors in type 2 diabetic rat models. Methods Models of type 2 diabetes were established in rats by high-energy diet and an intraperitoneal injection of streptozocin(STZ, 30 mg/kg). The rats in control group were fed with regular diet. Calcification of rat aortas were indentified by Von Kossa staining and atomic absorptiometry. Fasting blood glucose, insulin, 24 h urine albumin excretion and creatinine clearance rate were monitored. Results The rats fed with high-energy diet markedly shaved obesity, hyperglycaemia, hyperinsulinemia, insulin resistance and albuminuria. Although HE and Von Kossa staining didn't demonstrate calcium deposition, the calcium content detected by atomic absorptiometry in model group was significantly higher than that of control group(P＜0.01). We didn't find any connections between calcium content and weight, blood sugar, plasma insulin, albuminuria or Ccr. Conclusion The aorta calcium level of type 2 diabetic rat model increased significantly. The cause and mechanism of calcification remain to be elucidated.

Application of Endoscopic Great Saphenous Vein Harvesting during Coronary Artery Bypass Grafting

LI Hong-li;XU Dong;SHANG Xue-bin;LIU Yan-hui;LIU Fei;ZHANG Ke-feng;YAO Qing

2009, 30(2):  231-234. 

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Objective To observe the effects of endoscopic saphenous vein harvesting technique on the prevalence of leg-wound complications after coronary artery bypass grafting, and assess the feasibility of this technique and the potential trauma of the conduits. Methods Between January 2007 and February 2008, 118 patients underwent primary coronary artery bypass grafting, and 40 patients''saphenous vein taken using endoscopic harvest system(EVH), and the others by open vein harvesting(OVH), based on patient's willingness. The operation risk factors were compared between the two groups, and the proximal vein segment that underwent endoscopic harvesting was examined with electron-microscope. Results There was no significant difference in risk factors of incision complications between the two groups, but the incidence of various incision complications was significantly lower in the EVH group(12.5%) than in the OVH group(39.7%)(P<0.01). The time of harvesting was similar between the two groups. The length of conduits was(30.3±7.45)cm and (37.7±8.01)cm with significant difference between the two groups(P<0.05). Actually, there was increase in the use of sequential grafting technique in the EVH group for revascularization. Histological studies suggested that there were no extra vein trauma associated with EVH. At the same time, we have accumulated rich experience and skills in this procedure, and realized anatomical features of vein grafts. However, because of limited experience, the patency rate of conduits and the incidence rate of myocardial infarction in the perioperative or postoperative period were not compared between the two groups, more precise observations might be required. Conclusion The decrease in incision complications of EVH was unquestionably superior to those of OVH, especially for those patients having risk factors of incision complications. With increasing experience, the EVH technique might replace conventional saphenous vein harvesting methods and become the standard option.

Botulinum Toxin A Combining Home Rehabilitation for the Treatment of Spastic Cerebral Palsy

GAO Bao-qin;WANG Gui-fen;WANG Ya-jie;DENG Ya-xian;YANG Li-ying;YANG Wei-li;DENG Xin;TIAN Zhao-xia;HUANG Hai-tao;XIE Tian;ZHOU Da-wei;XU Jing

2009, 30(2):  235-238. 

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Objective To observe the curative effects and economical values of botulinum toxic A(BTX-A) combining home rehabilitation for treatment of spastic cerebral palsy(CP). As well as the application of gait analysis apparatus for the effective treatment of CP. Methods One hundred and forty eight children with CP were randomly divided into treatment group (83 cases) and control group(65 cases), and choose another 30 children with normal exercise and non-cerebral palsy as normal group. The treatment group received intramuscular injection of BTX-A and home rehabilitation. The control group received rehabilitation by professional physicians. Physician rating scale(PRS) and medical cost of both groups were assessed. Using gait analysis test platform, 30 cases of the treatment group received effectiveness evaluation before and 1 month after therapy. Results The PRS of treatment group on 3, 6 months after therapy improved more significantly than the control group(P<0.000). The cost in the treatment group was less than that of the control group(P<0.000). The ankle, knee joint movements and gait speed in the treatment group improved more significantly(P<0.000). Conclusion Rehabilitation combining intramuscular injection of BTX-A was better than single rehabilitation and its cost is lower than professional rehabilitation. It is objective, reliable and accurate in evaluating the curative effect with gait analysis apparatus.

临床研究

Allelic Imbalance in the Clonal Evolution of Prostate Carcinoma

LI Guang;ZHANG Hai-feng;XU Miao-sheng

2009, 30(2):  239-242. 

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Objective To observe the genetic basis of the clonal evolution of prostate carcinoma. Methods The pattern of allelic loss in 25 matching primary and metastatic prostatic adenocarcinoma were analyzed. DNA sample for the analysis of allelic loss pattern were prepared by tissue microdissection. The oligonucleotide primer pairs for the microsatellite DNA markers were D8S133, D8S136, D8S137, ANK1 on chromosome 8p12-21, LPLTET on chromosome 8p22, and D17S855 on chromosome 17q21. Results The overall frequency of allelic imbalance was 79% in primary tumors and 88% in paired metastases. Of 24 informative cases, 14 patients(58%) showed the same pattern of allelic loss or retention in matching primary and metastatic tumor at all marker locus; discordant allelic loss was observed in the remaining 10 patients(42%). Four patients showed loss of the same allele at one or more marker loci and both primary and metastatic tumors, but discordant allelic loss was observed at other marker loci. Five patients showed allelic loss in at least one genetic marker in the metastatic tumor but not in its matching primary tumor. Five patients displayed loss in one or more marker loci in a primary tumor but not in the matching metastases. Conclusion These data suggest that different patterns of allelic deletion may be acquired during cancer progression to metastases. The differences in genetic composition between primary prostate carcinoma and its metastases may be related to intrinsic cancer heterogeneity, overall genetic instability, and clonal divergence.

Fibrosarcoma Caused by Postoperative Radiotherapy for a Pituitary Adenoma Report of a Case and Review of the Literature

WANG Cui-zhi;LUO Jing;SUN Wei;DAI Jin-dong

2009, 30(2):  243-248. 

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Objective To study the clinical and histological characteristics of fibrosarcoma caused by postoperative radiotherapy for pituitary adenoma. Methods We analyzed a case of post-irradiation fibrosarcoma 8 years after surgery followed by irradiation for pituitary adenoma. Results The patient underwent transsphenoidal resection of the recurrent tumor. This led to significant relief in the patient's symptoms, but the tumor recrudescent a year later original. Humor was histologically verified to be a well differentiated fibrosarcoma(NCI grading system Ⅰ). The recrudescenttumor had high grade characteristics as a poorly differentiated fibrosarcoma(NCI grading system Ⅲ). The tumor cells responded positively in response to immunohistochemical staining for vimentin, but negatively for pan-CK, S-100, SMA, GFAP(-), CD117, GH, PRL, LH, FSH, TSH, ACTH. Conclusion Radiotherapy for pituitary adenomas can rarely induce fibrosarcomas. They should be suspected in patients following sellar irradiation who show abrupt new symptoms and has appropriate radiological findings. The secondary tumors are easy to regrow and the prognosis is poor.

Effects of Swimming on Jaundice and Emotion of Neonate

YUAN Xiu-zhen;CUI Zhi-qing;HAO Lan-xiang

2009, 30(2):  249-251. 

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Objective To explore the effects of swimming on jaundice and emotion of the neonate. Methods One thousand full-term neonates were randomized to control group(shower only) and observation group(swimming) 500 for each, and evaluation and observation of conditions of jaundice and the emotion of neonates were performed. Results Cases of jaundice in observation group were significantly fewer than the control group, and good emotion were more common than in the observation group. There was significant difference between those two groups(P<0.05). Conclusion Neonatal swimming can help to recover from the physiological jaundice and to promote the fadeaway of neonatal jaundice, which can keep the fine emotion and sleep of the neonate.

Evaluation on Efficacy of Sulfentanil Combined with Ropivacaine for Brachial and Jugular Plexus Nerve Block

CAI Yuan-xing;LI Lei;SUN Lei;CHENG Qing-hao;SHI Jian;DU Ping;LI Bao-qin

2009, 30(2):  252-255. 

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Objective To evaluate the efficacy and safety of sulfentanil combined with ropivacaine in brachial and jugular plexus nerve block. Methods Forty ASA Ⅰ, Ⅱ patients without contraindications of brachial and jugular plexus nerve block, aged 18~75, and weighing 47~98 kg, underwent elective cervical and upper limbs operations were randomly divided into two groups(n=20). 1% ropivacaine 10 mL+0.9% NS 20 mL(group A) and sufentanil 10.0 μg(1 mL) combined with 1% ropivacaine 10 mL+0.9% NS 19 mL(group B) were given separately. The efficacy of analgesia, sedation and locomotor block were evaluated by: 1Visual analogue scales(VAS)(0=no pain, 10=severe pain); 2 Ramsay sedation scales; 3 Bromage motor scores; 4 side effects such as nausea, vomiting, tickle and respiratory depression were also recorded at the points of before injection, 5 min, 10 min,15min, 20 min, 30 min, 1 h, 2 h and 4 h after injection. Results The demographic data(age, height, bodyweight, MAP and HR before operation) and durations of operations were comparable among two groups(P>0.05). VAS of 5 min, 1 h, 2 h and 4 h after injection in group B were lower than group A(P＜0.05); and Ramsay sedation scales 10 min till 30 min after injection were higher than group A(P＜0.05). Conclusion It has been demonstrated in this study that sulfentanil combined with ropivacaine for brachial and jugular plexus nerve block is safe and more efficient for early analgesia and sedation, and longer for analgesia than to use ropivacaine alone.