Objective To construct the eukaryotic expression vector of linker for activation of T cells(LAT) gene in mouse. MethodsLAT cDNA of mouse mononuclear cells was amplified with polymerase chain reaction.The fragment was inserted into the eukaryotic expression vector of pCMV-HA plasmid.The recombinant plasmid was verified by DNA sequencing and used to transfect lymphocytes of the asthmatic model. Results The length of amplified fragment was 729 bp.The sequence of LAT gene was confirmed by Genbank.The LAT protein could be detected in the lymphocytes of asthmatic model.Conclusion The recombinant eukaryotic expression vector pCMV-HA-LAT can be successfully constructed.

Animals ; Apoptosis ; Extremities ; blood supply ; Kidney ; pathology ; Male ; Nitric Oxide ; physiology ; Random Allocation ; Rats ; Rats, Wistar ; Reperfusion Injury ; pathology ; physiopathology

Objective To investigate the effect of curcumin on the expression of cannabinoid receptor 1 (CBR1) and NR2B subunit-containing NMDA receptor in spinal cord dorsal horn and dorsal root ganglion (DRG) neurons in a rat model of neuropathic pain.Methods Seventy-two male Sprague-Dawley rats,weighing 200-230 g,were randomly divided into 4 groups ( n =18 each):sham operation group (S group),chronic constrictive injury (CCI) group,curcumin group (Cur group) and solvent control group (SC group).Neuropathic pain was induced by CCI.The sciatic nerve was exposed and 4 loose ligatures were placed on the sciatic nerve at 1 mm intervals in groups CCI,Cur and SC.Curcumin was injected intraperitoneally at 100 mg· kg- 1· d-1 for 14 consecutive days after operation in Cur group,while the equal volume of dimethyl sulfoxide was given instead of curcumin in SC group.Mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) were measured on 2 days before operation and on 1,3,7,10 and 14 days after operation.Six rats in each group were sacrificed on 3,7 and 14 days after operation and the lumbar segments of the spinal cord ( L4,5 ) and DRGs were removed to determine the expression of CBR1 and NR2B by immuno-histochemistry.Results Compared with S group,MWT was significantly decreased,TWL was significantly shortened,and the expression of CBR1 and NR2B in spinal cord dorsal horn and DRG neurons was up-regulated after operation in the other three groups (P ＜ 0.05 ).Compared with CCI group,MWT was significantly increased,TWL was significantly prolonged,the expression of CBR1 inspinal cord dorsal horn and DRG neurons was up-regulated,and the expression of NR2B in spinal cord dorsal horn and DRG neurons was down-regulated after operation in group Cur (P ＜ 0.05 ),and no significant change in the parameters mentioned above was found in group SC ( P ＞ 0.05 ).Conclusion Curcumin can alleviate neuropathic pain in rats,and up-regulation of CBR1 expression and down-regulation of NR2B expression in spinal cord dorsal horn and DRG neurons are involved in the mechanism.

Objective To investigate the effect of curcumin on apoptosis in hippocampal neurons and the expression of c-Jun N-terminal kinase 3 (JNK3) and postsynaptic density protein 95 (PSD95) in hippocampus during cerebral ischemia-reperfusion (I/R) in rats with spontaneous hypertension (SH) .Methods One hundred and thirty-five male rats (homologous with WKY) with SH and 90 male normotensive WKY rats, weighing 275-325 g,were used in this study. The WKY rats were randomized into 2 groups ( n = 45 each) : sham operation group (WS group) and cerebral I/R group (W-I/R group) . The rats with SH were randomly divided into 3 groups ( n = 45each) : sham operation group (S-S group), cerebral I/R group (S-I/R group) and curcumin group (S-C group) .Global cerebral ischemia was produced by 4 vessel-occlusion method. The bilateral common carotid arteries were only exposed but not ligated in W-S and S-S groups. Intraperitoneal corn oil 10 ml/kg was injected at 30 min of reperfusion in W-I/R and S-I/R groups. Intraperitoneal curcumin 100 mg/kg was injected at 30 min of reperfusion in S-C group. Three animals in each group were sacrificed at 2 h, 6 h, 1 d, 3 d and 7 d of reperfusion and their brains were harvested for determination of apoptosis in hippocampal neurons and the expression of JNK3 and PSD95in hippocampus. Results The number of apoptotic neurons was significantly increased in S-S group compared with W-S group ( P < 0.05) . The number of apoptotic neurons was significantly increased and the expression of JNK3was up-regulated in S-I/R group compared with S-S group ( P < 0.05) . The number of apoptotic neurons was significantly decreased and the expression of JNK3 was down-regulated in S-C group compared with S-I/R group (P <0.05) . There was no significant difference in the expression of PSD95 among all the groups ( P > 0.05) . Conclusion Curcumin can inhibit apoptosis in hippocampal neurons and the mechanism is related to down-regulation of the expression of JNK3 in hippocampus. The mechanism by which curcumin down-regulates the expression of JNK3in hippocampus may not be related to PSD95 pathway.

Objective To investigate the effect of curcumin on the apoptosis in spinal cord and dorsal root ganglion neurons in a rat model of diabetic neuropathic pain (DNP) . Methods One hundred and eight male SD rats weighing 200-230 g were randomly divided into 4 groups ( n = 27 each): control group (group C), DNP group, solvent control group (group SC) and curcumin group (group Cur) . Diabetes was induced with intraperitoneal streptozocin 70 mg/kg. Successful induction of diabetes was defined as blood glucose ＞ 16.7 mmol/L. Curcumin and com oil 100 mg/kg (23 mg/ml) were given intraperitoneally once a day for 14 consecutive days starting from 14 days after administration of streptozocin in Cur and SC groups respectively. Mechanical paw withdrawal threshold (MWT) and thermal paw withdrawal latency (TWL) were measured 2 d before and 14 d after streptozocin injection and 3, 7 and 14 d after curcumin injection. The pain threshold measured at 14 d after administration of streptozocin decreased by more than 15% of the baseline in all the rats. The expression of caspase-3 and Bcl-2 in spinal cord and dorsal root ganglion was determined at 3, 7 and 14 d after curcumin injection by immuno-histochemistry and Western blot, and the neuronal apoptosis rate was determined by TUNEL. Results Compared with group C, MWT and Bcl-2 expression were significantly decreased, TWL was significantly shortened, the neurona lapoptosis rate and caspase-3 expression were significantly increased in DNP, SC and Cur groups ( P ＜ 0.05).Compared with group DNP, MWT and Bcl-2 expression were significantly increased, TWL was significantly prolonged, the neuronal apoptosis rate and caspase-3 expression were significantly decreased in Cur group ( P ＜0.05) . There was no significant difference in the parameters mentioned above between DNP and SC groups ( P ＞0.05). Conclusion Curcumin can attenuate DNP by inhibiting the apoptosis in spinal dorsal hom and dorsal root ganglion neurons in rats, and the inhibition of caspase-3 expression and increase in Bcl-2 expression are involved in the mechanism.

Objective To investigate the changes and significance of concentration of soluble intercellular adhesion molecule-1(sICAM-1) in cerebrospinal fluid (CSF) of bacterial meningitis(BM) in rabbits. Methods A total of 36 rabbits were randomly divided into m eningitis group, meningitis cefotaxime-treated group and control group. BM indu ced by escherichia coli(Ec) via cerebellomedullary cistern inoculated. Normal sa line was injected in control group. CSF was sampled in different time. The conce ntration of CSF sICAM-1 was detected by ELISA.Results 1.There was a low concentration of sICAM-1 in CSF in 85 percent of normal rabbi ts.2.In meningitis group, there was a sharp rise in the concentration of CSF sIC AM-1 at 6 hours after Ec was inoculated, reached a peak level at 12 hours, and t here was higher concentration of CSF sICAM-1 between 6 and 24 hour than that at 0 hour.3.In meningitis cefotaxime-treated group, the concentration of CSF sICAM -1 at 6 hour and 12 hours was similar to meningitis group, the time that get pe ak level was at 24 hours that at 12 hours after making use of antimicrobial agen t.The peak level was higher than meningitis group. The concentration of CSF sICA M-1 decreased markedly at 48 hours that made use of antimicrobial agent 36 hour s later,but the concentration was still higher than the peak level of meningitis group.Afterwards, with the time of making use of antimicrobial agent lengthened ,the concentration of CSF sICAM-1 decreased gradually.4.Experimental results in dicated by correlating analysis to these data that there was positive correlatio n in the concentration of CSF sICAM-1 with the brain water content.Conclusions sICAM-1 participates in the pathological process of BM, and contributes to the damage of blood brain barrier and the formation of brain edema.There is importan t significance that drugs which can resist the discharge of sICAM-1 will be impl ied. J Appl Clin Pediatr,2005,20(2):163-165

Objective To investigate the effect of human cytomegalovirus (HCMV) on proliferation of colony forming unit T-lymphocyte (CFU -TL)and its interference methods. Methods Normal CFU - TL culture was used as blank control. Normal CFU- TL culture system plus inactivated HCMV fluid as inactivated HCMV control. The dilution of 1:10,1:100,1:1000 were added into CFU -TL colonies culture system directly as infected group. Astragalus (AMI) and ganciclovir(GCV) were added into culture system with HCMV dilution of 1:10 as experimental group. By methylcellulose semi-solid culture, different concentrations of HCMV - AD1699 affect CFU-TL and interfered by astragalus AMI, GCV. CFU - TL were surveyed. The effect of HCMV on CFU-TL proliferation was measured by MTT; HCMV-AD169 DNA in CFU-TL was found by PCR. Results 1. Compared with control group, the numbers of CFU -TL in the HCMV infection groups decreased significantly(P

Objective To establish a simple and sensitive method for the determination of serum copper by spectrophotometry.Methods Nitro-PAPS was used as a coloring agent for serum copper in the presence of surfactants Tween-80 and Triton X-100 and the formed complex was measured by spectrophotometry.Results The maximum absorption wavelength of the complex was 570 nm and the molar absorption coefficient was 7.95?10~4 L/(mol?cm).The lineafity of the method was up to 63.0 ?mol/L and the recoveries ranged from 98.6% to 103.1%.The within-run and between-run CVs were 2.1%-3.3% and 2.7%-3.8%.The method(Y)was compared with an AAS method(X)and a correlation of Y=1.01X -0.27(r=0.998 2)was obtained.A reference interval(x~-?2s)determined with this method on 68 individuals was 9.7-24.1 ?moL/L.Conclusions A simple and sensitive method for serum copper has been established.It may used for the analysis of serum copper in clinical laboratories.

Objective To investigate the etiological factors, clinical features and prognosis of non- traumatic rhabdomyolysis(RML). Methods The medical records of 13 non-traumatic RML patients hospital- ized between 1995-2006 were reviewed. The etiological, clinical, laboratory and therapeutic data were anal- ysed. Results Among 13 patients with non-traumatic RML, multiple factors were responsible for rhabdomyol- ysis in eight patients and single etiologic factor in 5 patients. Different etiological factors were identified, in- cluding 6 with excessive exercise, 3 with hyperpyrexia, 3 with drugs(including illicit drugs, fenofibrate, cy- closporine), 3 complicated with inflammatory myopathy and 2 with limbs compression. Nine patients had myal- gia and muscle weakness, 6 patients had abnormality in nervous system, 4 patients had hyperpyrexia, 3 pa- tients had digestive symptoms. Nine patients were complicated by coagulation disorders and 6 with acute renal failure(ARF). The serum levels of creatine kinase(CK)were decreased to normal within one month in 6 patients, the patient whose rhabdomyolysis was induced by fenofibrate with diabetes and chronic renal failure showed to inadequate decrease within 60 days. Three patients whose problem was induced by inflammatory myopathy, CK levels decreased within 4 months and 6 months in 2 patients, respectively, but CK level was not returned to normal level in one patient during the 80 follow-up days. Three patients died from multiple causes, such as ARF, coagulation disorders,electrolyte and metabolic disturbances. Conclusion Excessive exercise is the most common cause of non-traumatic RML, followed by drugs and inflammatory myopathy. The prognosis is poor in patients with multiple etiological factors and ARF.

Objective To evaluate the effect of B-vitamins(B1,B6,B12)on diabetic neuropathic pain (DNP)in rats.Methods 104 male SD rals weighing 200-230 g were randomly divided into 13 groups(n=8 each):group Ⅰ control(group C);group Ⅱ DNP;group Ⅲ DNP+ normal saline(solvent of vitamins,group NS);group Ⅳ,Ⅴ,Ⅵ DNP+vitamin B1 10,33 or 100mg/kg,kg(group B1 10,group B133,group B1 100);group Ⅶ,Ⅷ,Ⅺ DNP+vitamin B6 10,33 or 100 mg/kg(group B6 10,group B633,group B6100);group Ⅹ,Ⅲ,ⅫDNP+vitamin B12 0.5,1.5 or 4.5 mg/kg (group B12 0.5,group B121.5,group B124.5)and group ⅩⅢ DNP+vitamin B1 10/B6 33/B12 1.5 mg/kg(group VBC).Diabetes was induced with intraperitoneal(IP) streptozocin mg/kg in group Ⅱ-ⅩⅢ.B-vitamins were give.IP once a day for 14 consecutive days starting from 14 d after IP streptozocin in group Ⅳ-ⅩⅢ.Venous blood samples were taken before(baseline)and 3 d after IP streptozocin for determination of blood glucose level. Successful induction of diabetes was defined as blood glucose > 14.6 mmol/L. Mechanical paw withdrawal threshold to yon Frey stimuli (MWT) and paw withdrawal latency to thermal nociceptive stimulus (TWL) were measured 2 days before and 14 days after IP streptozocin and on the 1, 3, 7, 14 days of B-vitamin administration. Animals with pain threshold measured at 14 days after IP streptozocin decreasing by less than 15% of the baseline were excluded from the study. The animals were sacrificed after the last pain threshold measurement and L4,5 lumbar segment of the spinal cord and dorsal root ganglions (DRG) were removed for determination of p-CREB expression using immuno-histuchemistry. Results MWT was significantly lower and TWL was significantly shorter and the expression of p-CREB was significantly higher in the other groups than in group C. B-vitamin administration significantly reduced thermal and mechanical hyperalgesia induced by diabetes and down-regulated the expression of p-CREB in a dose-dependent manner as compared with group DNP. The inhibitory effect of vitamin B complex against thermal and mechanical hyperalgesia was significantly stronger and the expression of p-CREB was significantly lower in group VBC as compared with group B110, group B633 and group B121 .5 respectively. Conclusion B-vitamains can attenuate DNP through inhibition of phospberylation of CREB in the spinal dorsal horn and DRG.