Objective:To develop a fixed clamp apparatus for Macrophage system and PLEKHQ1 gene knockout mice, in order to provide a safe, simple, convenient and time-saving experimental tool for mice tail blood collection and injection.Methods:Clamp for macrophage system and PLEKHQ1 gene knockout mice was made by 50 ml syringe with slot according to the drawing. Clamp base was made by 50 ml plastic tube and appropriate size of the plastic tap.Results:Compared with current mice clamp in the market, the mice fixed clamp was a simple, low-cost, safe, easy and timesaving tool.Conclusion: The mice clamp for mice tail blood collection and injection provided a safe and convenient tool, and greatly improved the efficiency of the experiment.

Objective:To construct of eukaryotic expression vector of RNA interference specific for CXCR4. Methods: Genome sequences of CXCR4 gene were retrieved from Genbank and cDNA was designed coding expression of shRNA (small hairpin RNAs) for CXCR4 gene. The cDNA was synthesized and inserted into plasmid pWH1, and the recombinant pWH1-CXCR4 expression vector was identified by enzyme cutting method. Then, pWH1-CXCR4 expression vector were transfected into salivary mucoepidermoid carcinoma Mc3 cells. At last, the expression of CXCR4 in Mc3 transfected with pWH1-CXCR4 expression vector was evaluated by RT-PCR and flow cytometry analysis. Result:Compared with Mc3 cells and Mc3 cells transfected with plasmid pWH1, the Mc3 cells transfected with pWH1-CXCR4 expression vector showed a lower mRNA and protein expression of CXCR4. Conclusion:The constructed CXCR4 shRNA expression vector can block the expression of CXCR4 in salivary mucoepidermoid carcinoma cells, and it may be helpful for further study on the significance of CXCR4 on the proliferation, metastasis and experimental treatment of salivary mucoepidermoid carcinoma.

Objective:To observe the influence of Naoxintong on expression of glucose-regulated protein 78(GRP78) and GRP94 in the striatum of rats with focal cerebral ischemic injury,so as to investigate their possible protective mechanism on cerebral ischemic injury.Methods:Ninety rats were equally randomized into 3 groups(n=30): sham operated group,ischemic group(Focal transient cerebral ischemia model was established with intraluminal occlusion of left middle cerebral artery) and Naoxintong pre-treated group(Treated with Naoxintong 5 d before ischemic injury).The expression of GRP78,GRP94 in rats striatum was detected by histological method,immunohistochemistry staining and semiquantitative RT-PCR at the different time points(6,12 and 24 h after ischemic treatment).Results:The focal transient cerebral ischemia model was successfully established in rats.Histological results showed that the degree of focal cerebral ischemic injury in Naoxintong pre-treated group was significantly lower than that in ischemic group.Immunohistochemistry staining and RT-PCR results illustrated that the expression of GRP78 and GRP94 in ischemic group was lower than that in sham operated group at each time point after ischemic treatment((P

Objective To study the value of color Doppler ultrasound in diagnosis of deep vein thrombosis (DVT) of lower extremity and pulmonary embolism (PE) in spinal cord injury (SCI). Methods 60 PE patients (PE group) and 35 SCI patients without PE (control group) received color Doppler ultrasound examination for DVT of lower extremity. Results PE group included 36 SCI patients (PE-SCI group) and 24 no-SCI patients (PE-no-SCI group). There were 15 cases with lower extremity thrombosis in PE-SCI group, and 9 cases in PE-no-SCI group (P>0.05), while there were 5 cases in the control group. There was significantly different in lower extremity thrombosis between PE group and the control group (P<0.01). In PE group, the detection rate was not significantly different between acute PE (detected 11 cases out of 37 cases) and chronic PE (detected 10 cases out of 23 cases) (P>0.05). 31 cases were rechecked as lower extremity venous valve regurgitation and calf muscle vein dilation (51.7%) in PE group while 8 cases in the control group (22.9%) (P<0.01). Conclusion There is not significantly different in the detection rate of DVT of lower extremity in PE patients with and without spinal cord injury, which are higher than in the patients without PE. Color Doppler ultrasound is necessary to check DVT in acute and chronic PE patients.

We used SDS-polyacrylamide gel electrophoresis (SDS-PAGE), immunoblotting of monoclonal antibody (McAb) J28 and peanutagglutinin (PNA) for detecting fetoacinar pancreatic protein (FAP) in extracts of 20 cases of fetal pancreas, 5 of pancreatic cancer and 4 of normal pancreas. The result showed that fetal pancreas and pancreatic cancer had a same level band about 110000 in Commassie blue stain, but absent in normal pancreas. The band could react with both McAb J28 and PNA PNA also could react with smaller glycoprotein in pencreatic cancer. It is suggested that McAb J28 and PNA combine with different sites of FAP.

Objective To conduct an empirical study for quantifying the anastomosis between two vessels after skin expanded technique by the method of angiography and to provide a precise basis for vascular study in skin flap.Methods Bilateral skin flaps based on deep iliac circumflex vessels were elevated from the abdominal wall including deep superior epigastric vessels.One was expanded at the boundary between two vessels and the other unexpanded.An X-ray image was obtained by carotid arterial injection of gelatin-lead oxide mixture.Three parallel lines with equal intervals perpendicular to long axis of the two vessels were designed at the communication area.Vessel anastomosis quantity was determined by counting the number of marks derived from the intersections of the lines and the vessels and statistical analysis was carried out.Results The marks of intersection in expanded group were more than unexpanded group with statistical significance.Conclusions The method for quantifying vessel anastomosis in skin flap is reliable.The principles of this procedure may also be applied to other experimental and elinical studies.

BACKGROUND: Alginic sodium diester (ASD) possesses neuroprotective function because of its selective calcium antagonist effects.OBJECTIVE: To compare the influences of ASD on intraneuronal Ca2+content and nerve cell apoptosis before and after reperfusion in focal cerebral ischemic rats.DESIGN: Randomized controlled observation.SETTING: Neurological Department of Xiangya Hospital Affiliated to South China University; Laser Orthopedic Surgery of the First Hospital Affiliated to Southern China University.MATERIALS: This experiment was carried out between November 2003and April 2004 at the Neurological Department of Xiangya Hospital Affiliated to South China University. A total of 65 male SD rats were recruited and randomized into 6 groups; 17 got lost during the experiment, and the other 48 rats completed the experiment with 8 rats in each group.METHODS: In sham operation group, an incision was made on rats' cervical skin and sutured. Right cerebral middle artery was occluded in rats of ischemic group, ASD 5 mg/kg preischemic group, ASD 5 mg/kg postischemic group, ASD 10 mg/kg preischemic group, and ASD 10 mg/kg postischemic group. After that, rats in all but ischemic group were subjected to intraperitoneal injection of various dosage of ASD or excipient 30minutes before reperfusion and 5 hours after reperfusion. FCM was used to determine intraneuronal Ca2+ content and rate of nerve cell apoptosis;meanwhile, neurological dysfunction was scored.MAIN OUTCOME MEASURES: [1] Influence of ASD on the score for neurological dysfunction, intraneuronal Ca2+ fluorescence intensity, and neuronal apoptosis in rats with right cerebral middle artery ischemia. [2]Correlation of behavioral obstacle score with intraneuronal Ca2+ fluorescence intensity and neuronal apoptosis in rats with right cerebral middle artery ischemia.RESULTS: Totally 65 rats were enrolled in this study, 17 of which got lost and the other 48 rats entered the result analysis. [1] Influence of ASD on the score for neurological dysfunction, intraneuronal Ca2+ fluorescence intensity, and neuronal apoptosis in rats with right cerebral middle artery ischemia: The score was obviously reduced in ASD 5 mg/kg preischemic group, ASD 5 mg/kg postischemic group, ASD 10 mg/kg preischemic group and ASD 10 mg/kg postischemic group as compared with ischemic group (1.80±0.21, 2.20±0.23, 1.20±0.11, 2.00±0.22, 3.40±0.65); moreover,functional improvement was more obvious due to pre-reperfusional administration than post-reperfusional administration. Intraneuronal Ca2+ concentration was reduced after ASD administration at different degrees and lower than that of ischemic group. Decrement of intraneuronal Ca2+ concentration was found most obvious due to 10 mg/kg ASD administration 30 minutes before reperfusion, approximately reduced by 70%; moreover, neuronal apoptosis rate on the ischemic side was obviously suppressed by ASD administration, displaying time-dependent manner, with apoptotic suppression effect more obvious in pre-reperfusional group than in post-reperfusional group (5.68%, 10.03%; 4.00%, 9.91%). [2] Correlation of behavioral obstacle score of right cerebral middle artery ischemic rats with intraneuronal Ca2+ fluorescence intensity and membrane associated protein/propidium iodide apoptosis: Obvious positive correlation was found between behavioral obstacle score and intraneuronal Ca2+ fluorescence intensity and detection rate of membrane associated protein/propidium iodide apoptosis (r=0.51,0.62, P ＜ 0.05); intraneuronal Ca2+ fluorescence intensity was also positively correlated with the detection rate of membrane associated protein/propidium iodide apoptosis (r=0.84, P ＜ 0.05).CONCLUSION: [1] ASD can exert anti-apoptosis effect by suppressing the increment of intraneuronal Ca2+ concentration, thus having neuroprotective function and ultimately improving neurological dysfunction. [2] Its effect displays time-dependent manner, and neurological functional improvement is more obvious by pre-reperfusional administration than by post-operational administration.

Objective To study the pharmacokinetics of aristolochic acid A in Radix Aristolochiae and the compound preparation of Guanxinsuhe Capsule in mice in vivo after single-dose oral administration and observe the difference of aristolochic acid A absorption and distribution. Methods Aristolochic acid A assay was performed by RP-HPLC on a Waters apparatus with a DiamonsilTM C18 column (250 mm × 4.6mm, 5 μm), a mobil phase: a mixture of methanol-water-acetic acid (72: 27 : 1), flow rate: 1.0 mL/min, detection wavelength: 315 nm, and column temperature: 20 ℃. Results Mice were given Radix Aristolochiae and Guanxinsuhe Capsule by ig at the same level of 2. 5 mg/kg of aristolochic acid A, respectively, which were suspended in 0. 3% CMC-Na solution. Plasma concentrations were determined by RPHPLC. After single-dose ig administration of Radix Aristolochiae or Guanxinsuhe Capsule to mice, the mean plasma concentration-time courses of aristolochic acid A obtained fitted the one-compartment model.The main pharmacokinetic parameters of aristolochic acid A in Radix Aristolochiae, t1/2ka, t1/2 ke, tmax,AUC, Cmax are 5. 103 min, 43. 63 min, 17.89 min, 80. 45 (μg · min)/mL, and 0. 916 8 μg/mL; the rela tive pharmacokinetic parameters in Guanxinsuhe Capsule are 5. 294 min, 43.50 min, 18. 32 min, 33.08(μg · min)/mL, and 0. 381 8 μg/mL. Conclusion The Cmax of aristolochic acid A in Guanxinsuhe Capsule is significantly less than that in Radix Aristolochiae, which indicates that the compound compability could decrease the absorption of aristolochiae acid A.

Objective To investigate the roles of hemopoietic stem cells (HSCs) in the pathogenesis of psoriasis. Methods HSCs were isolated from the bone marrow of a patient with psoriasis vulgaris and a normal human control. Forward- and reverse-subtracted cDNA libraries were constructed by using suppression subtractive hybridization (SSH) technique between HSCs from the patient and control. Bacterial PCR and dot hybridization were performed to screen for positive clones followed by gene sequencing, identification and functional analysis. Real time quantitative PCR was carried out to measure the mRNA expression of interferon-γ and thymosin 10 (TMSB10). Results Nine genes were screened from the forward-subtracted cDNA library which encoded interferon-γ, tyrosine phosphatase, SUMO1 activase, etc, and 8 genes including the TMSB10-encoding gene were screened from the reverse-subtracted cDNA library. The relative expression level of interferon-γ in HSCs from the patient was 47.5 times that in HSCs from the control, while the level of TMSB10 from the control was 22.6 times that from the patient. Conclusion The abnormal expression of 17 genes which encode interferon-γ, thyrosin, and so on, may be involved in the development of psoriasis.

AIM: To study the effect of cocaine on caspase-3 in myocardiac cells of male rats in different age. METHODS: Three-week-old( n= 16), six-week-old( n= 16) and twelve-week-old( n= 16)male Sprague Dawley rats were all divided into control groups and experiment groups randomly, each group had eight animals, experiment groups were given cocaine hydrochloride (15 mg?kg -1 body weight) subcutaneously daily for four weeks.The ratio of heart weight to body weight (HW/BW,mg/g) were measured. DNA fragmentation of myocardia cells was determined by gel electrophoresis, and caspase-3 activity in myocardia cells was tested by flow cytometry (FCM). RESULTS: In three experiment groups, the DNA isolated from myocardial cells displayed clear ladder pattern. The HW/BW and the caspase-3 activity were increased significantly than those of control groups ( P