Objective 　To screen for gene mutation of exon 18 in Chinese patients with Wilson disease. Methods　PCR-SSCP was used to screen exon 18 in 45 Wilson disease patients among 39 Chinese families and 10 normal controls. Those with abnormality were further analyzed by necleotide sequence analysis. Results　There were 16 mobility shift with two different styles in exon 18. All abnormal mobility shifts were sequence analysed. No gene mutation was found. Conclusions　Our result suggest that, contrary to findings in Caucasians, exon 18 is not a frequent mutation point in Chinese patients with Wilson disease.

【Objective】 To investigate the molecular basis of hepatolenticular degeneration (Wilson disease, WD) and to attempt to construct the feasibility of gene diagnosis in the disease. 【Methods】 We have performed the molecular biological study on this disease for 10 years by molecular geneti c techniques. 【Results】 ①Location of WD gene in Ch inese: Using pairwise linkage analysis and multipoint linkage analysis method, w e constructed a genetic map of DNA markers within D13q14.2-3 which refined the location of WD gene by restriction fragment length polymorphism(RFL P) and microsatellite polymorphism analysis; ②Screen for mutations of WD gene in Chinese people: we detected the structure of 21 exons of WD ge ne in 45 patients from 39 pedigrees by PCR-SSCP(Single strand conformation poly morphism) and PCR-DNA sequencing technology, found a new mutation in exon 5 and nuclcotide sequence analysis showed it is a T insertion. We also conformed the Arg778Leu in exon 8, the highest frequence mutation point in Chinese people, wit h mutation rate 22.8% in total;③Carrier detection and presymptomatic diagnosi s of WD: Based on DNA recombination technology, we peformed successfully the gen e diagnosis in all individuals of 79 families with WD and built up a helpful spe cific enzyme cut method (PCR-Msp1) to detect the carrier and presympomatic patients in Chinese pe ople with WD. 【Conclusion】 These results showed that the location of WD gene within D13q14.2-3 is the same in Chinese as in Caucasians, but the g ene high m utation point,the gene diagnosis method and its pathogenesis are markly different.

Venous thromboembolism includes deep venous thrombosis and pulmonary embolism. It is a more common and preventable complication in neurology. The prevention of venous thromboembolism is an important component in the treatment of the patients with cerebral hemorrhage. The measures include mechanical prevention and drug prevention. The mechanical prevention measures include intermittent pneumatic compression devices and pressure gradient elastic stockings. Studies have suggested that anticoagulants also plays an important role in the prevention of venous thromboembolism. The comprehensive and systematic understanding of the prevention of venous thromboembolism wil help to guide the clinical therapy and improve the outcomes of patients after primary intracerebral hemorrhage.

Objective: To study if the immune-enhancing effect of Arg was mediated via liver.Methods: The direct effect of Arg on T cell proliferation was measured by 3H-TdR incorporation. Rat hepatocytes were primarily cultured in serum-free DMEM/F12 medium, and then cultured in medium containing Arg(?mol/L:0,7.5,75,750,7 500), and the supernatant was collected at 0, 24, 48, 72 hours, and added to splenocytes culture, and T cell proliferation, NK cell activity and IL-2 activity,〔Ca 2+〕i were measured respectively.Results: Arg had no direct effect on splenocyte proliferation. The hepatocyte culture supernatant significantly increased the lymphocyte〔Ca 2+〕i ,IL-2 activity and T lymphocyte proliferation; 7 500 ?mol/L was most effective. Conclusion: Arg may enhance immune function via secretion of bioactive molecules by hepatocytes.

The sensor of naringin(NG), a non-electroactive substance, was prepared based on the molecular imprinting technique.Using cyclic voltammetry technique (scan rate is 100 mV/s), the naringin imprinting sensitive film, poly-o-aminophenol was coated on the surface of a graphite electrode in the presence of naringin which was considered as the template, and characterized by SEM and X-ray reflective spectrophotometry (XRR).Using K_3Fe(CN)_6 as an electroactive marker, the electrochemical properties of the NG sensor were investigated by CV, electrochemical impedance spectroscopy (EIS), differential pulse voltammmetric and chronoamperometric.The results showed that the imprinted electrode was significantly different from the non imprinted electrode in morphologies and electrochemical properties, and a linear relationship between the peak current and the naringin concentration was found in the range of 6.0 × 10 ~(-5)-1.4 × 10 ~(-4) mol/L with a detec tion limit of 1.6 × 10 ~(-5) mol/L.Moreover, the imprinted electrode exhibited a good selectivity and rapid response to the naringin template molecules, as well as an excellent reproducibility(RSD = 1.8 %, n=5).

Objective To investigate the change of emotion and striatum dopamine transporter(DAT) expression in adult male rats experiencing maternal deprivation, and to explore whether DNA methylation is involved in the regulatory mechanism of DAT expression. Methods Newborn rats were randomly divided into 2 groups: a maternal separation group (n=16) and a control group (n=14). The maternal deprivation group were separated from their mother for 6 hours (09∶00-15∶00) per day from postnatal day 1 to 14, while the controls (n=14) without the deprivation. When the rats in the 2 groups were 12 week, their spontaneous anxiety levels and exploratory ability in novel environments were assessed by an elevated plus maze and an open field test. DAT mRNA expression in the striatum was detected by reverse transcription-PCR, and its DNA methylation level was measured by bisulfated DNA sequencing. Results Maternally-deprived rats showed lower ability of exploring in a new environment and lower levels of anxiety than the controls. The expression of DAT mRNA in the striatum of the maternal separation group (0.236±0.043) was significantly lower than that in the control group (0.480±0.107) (P＜0.05). However the DNA methylation level in the promoter region of DAT was not significantly different between the 2 groups. Conclusion Maternal deprivation influenced the emotion and expression of dopamine transporter in adult rats and DNA methylation may not be involved.

Objectives:In this study,the effects of glutamine(Gln) and branched chain amino acids(BCAA) enriched formulas on free radical metabolism and immunity in traumatized rats were investigated. Methods:After injury,twenty one male Wistar rats were randomly divided into three groups,and fed with rations containing casein,a commercial amino acids(17AA),and a new amino acids formula(20 AA) respectively.The rations were isonitrogenous and isocaloric.Before operation and on days 3,7,14 postoperation,body weight,dietary intake,the concentrations of MDA,the activities of SOD in plasma were measured.At last,the animals were killed,the hydroyproline and spleen lymphocyte blastogenesis were determined. Results:①After injury,body weight of rats were reduced significantly,the concentrations of MDA in plasma were increased,while the SOD activities were decreased. ②Compared with 17AA group, the levels of hydroyproline in sponge were increased in 20AA group.③There were better effects of reducing plasma MDA levels and enhancing plasma antioxidase activities in 20 AA group than 17 AA group.④The weight of thymus and spleen and spleen lymphocyte blastogenesis were more obviously increased in 20AA group than in 17AA group. Conclusions:The new amino acids preparations can increase the antioxidase activities,enhance immunity and promote wound healing.

Objective To analyze the condition and characteristics of hand-foot-mouth disease ( HFMD) from 2010 to 2012 in Renqiu city.Methods Surveillance and detecetion of HFMD was collected according to Renqiu city system for diseases control and prevention .The pathogen of HFMD severe case was deteceted .Results 12 293 cases including 735 severe cases were recorded in Renqiu city from 2010 to 2012,The highest of the resident population was in 2012 and the lowest one was in 2010(r=0.47,P<0.05).The total morbidity presented the obvious seasonal char-acteristic,which reached the summit in June ,July,August.The population morbidity was the clustered children .The average incidence rate of severe cases was 5.98%.The incidence rate in 2012 and 2011 was higher than that in 2010 (r=0.43,0.39,all P<0.05).There was significant difference of the pathogens types in severe cases among three years with the pathogen of CoxA 16 in 2010,2011 and humantero virus 71 viruses in 2012.Conclusion The inci-dence of HFMD presents the increasing and seasonal characteristics with the prevalence in the scattered children and the pathogens of CoxA16 in 2010,2011,humantero virus 71 in 2012.

OBJECTIVEBoth of gp91(phox) (an isoform of nicotinamide adenine dinucleotide phosphate-reduced oxidases) and Src (a non-receptor protein tyrosine kinase) play a prominent role in mediating hypoxia/reoxygenation injury of neurons. The present study was designed to investigate the neuroprotective effect of equol, a predominant active metabolite of daidzein, against hypoxia/reoxygenation injury in rat pheochromocytoma cell line (PC12) and explore the underlying mechanisms.

METHODSPC12 cells exposed to hypoxia/reoxygenation injury were examined for reactive oxygen species (ROS) using dihydroethidium and 2', 7'-dichlorofluorescein diacetate and analyzed for changes in lactate dehydrogenase (LDH) activity and malondialdehyde (MDA) content. The expression levels of gp91(phox) and phosphorylated Src-Tyr416 (p-Src) were measured using Western blotting.

RESULTSEquol dose-dependently restored the cell viability and decreased LDH activity and MDA content in culture medium of PC12 cells exposed to hypoxia/reoxygenation. Pretreatment of the cells with 10(-5) and 10(-6) mol/L equol inhibited hypoxia/reoxygenation-induced increase of ROS. PC12 cells treated with equol prior to hypoxia/reoxygenation injury showed significant enhancement of the protein levels of gp91(phox) and p-Src.

CONCLUSIONEquol confers neuroprotection against hypoxia/reoxygenation injury in PC12 cells by inhibiting the generation of ROS very likely as a result of down-regulation of gp91(phox) and inhibition of Src phosphorylation.

Animals ; Cell Hypoxia ; Cell Survival ; Down-Regulation ; Equol ; pharmacology ; L-Lactate Dehydrogenase ; metabolism ; Malondialdehyde ; metabolism ; Membrane Glycoproteins ; metabolism ; NADPH Oxidase 2 ; NADPH Oxidases ; metabolism ; Neurons ; drug effects ; metabolism ; Neuroprotective Agents ; pharmacology ; PC12 Cells ; Phosphorylation ; Rats ; Reactive Oxygen Species ; metabolism ; src-Family Kinases ; metabolism

OBJECTIVETo investigate the role of oxidative stress in impaired intermediate-conductance Ca(2+)-activated potassium channels (IKCa)- and small-conductance Ca(2+)-activated potassium channels (SKCa)-mediated relaxation in diabetic resistance arteries.

METHODSRat diabetic model was induced by a high fat and high glucose diet and streptozotocin (STZ) injection. Endothelial function of mesenteric arteries was assessed with the use of wire myography. The expression levels of IKCa and SKCa in cultured human umbilical vein endothelial cells (HUVECs) treated with H2O2 and/or antioxidant alpha lipoic acid (ALA) were measured using Western blotting.

RESULTSIKCa- and SKCa-mediated vasodilatation in response to acetylcholine was impaired in the third-order mesenteric arterioles of diabetic rats. In cultured HUVECs, H2O2 significantly decreased the protein expression of IKCa and SKCa. ALA alleviated the impairment of both vasodilatation function of the mesenteric arterioles ex vivo and enhanced the expression of IKCa and SKCa challenged with H2O2 in cultured HUVECs.

CONCLUSIONOur data demonstrated for the first time that impaired IKCa- and SKCa-mediated vasodilatation in diabetes was induced, at least in part, by oxidative stress via down-regulation of IKCa and SKCa channels.

Animals ; Cells, Cultured ; Diabetes Mellitus, Experimental ; metabolism ; physiopathology ; Human Umbilical Vein Endothelial Cells ; drug effects ; pathology ; Humans ; Hydrogen Peroxide ; pharmacology ; Intermediate-Conductance Calcium-Activated Potassium Channels ; metabolism ; Male ; Mesenteric Arteries ; physiopathology ; Oxidative Stress ; Rats ; Rats, Sprague-Dawley ; Small-Conductance Calcium-Activated Potassium Channels ; metabolism ; Thioctic Acid ; pharmacology ; Vasodilation