Fluorescence resonance energy transfer(FRET)is increasingly used to study inter-and intramolecular interactions in living cells.Since being proportional to the concentration of the donor-acceptor complex.FRET value must be normalized to exclude the influence of the ratio and the concentration of donor and acceptor for comparison.Different from the intra.molecular FRET which is simplified by the fact that the COncentration of the donor is equal to that of the acceptor,the inter-molecular FRET is usually too complicated for most existing measurements to quantify exactly.We deduced the exact proportion of the donor-acceptor complex based on a unique characteristic of homodimer,a special kind of the intermolecular interaction,developed an exact quantification measurement of the FRET.We proved the novel method Can generate more reliable estimation of FRET value by comparison with other methods using a homodimer,estrogen receptor alpha(ERa),as a FRET pair.

Objective To investigate the prevention and management of portal vein thrombosis following orthotopic liver transplantation.Methods Between May 1995 to September 2005,clinical data of 137 cases subject to orthotopic liver transplantation were analyzed.Among them,there were 10 cases of portal vein thrombosis(5 cases in gradeⅠ,4 cases in gradeⅡand 1 case in gradeⅢ). All patients received an eversion thromboendovenectomy(ETEV)with occlusion of the portal flow u- sing a Forgarty balloon.Ligation of the collateral circulation,especially spontaneous or surgical sple- norenal shunt,was made as approaches to improve portal flow.Heparin or low-molecule-weight hepa- rin as a prophylactic anticoagulation therapy was maintained during and after operation if prothrombin time was less than 18 s.Follow-up Doppler ultrasonography was used daily in the early postoperative period.Results After a follow-up of 2～66 months,overall incidence of portal vein thrombosis was 2.92%(4/137).Surgical thrombectomy and revascularization was carried out in 1 case.Thromboly- sis,balloon angioplasty and stent placement via hepatic artery were performed on 2 cases.No treat- ment was given in 1 patient without hepatic dysfunction and portal hypertention.Mortality related to portal vein thrombosis was 0.Conclusions Portal vein thrombosis might be avoided by performing a complete thrombectomy as often as possible,by ligation of portosystemic shunt during surgery,and by postoperative anticoagulation.Close follow-up by Doppler ultrasonography may make a prompt di- agnosis and reduce portal vein thrombosis-derived loss of grafts.

Background Retinal vascular recanalization is key to the treatment of retinal vascular occlusive disease.Studies confirmed that urokinase by intravitreal injection inhibits the expression of occludin protein at tight junction complexes among retinal capillary endothelial cells.Objective This study was to observe the effects of urokinase via eye local injection on the outward permeability of blood-retinal barrier by detecting the concentration of intravitreal Evans blue (EB).Methods Sixty healthy Sprague-Dawley (SD) rats were randomly assigned to four groups,and the right eyes of the rats were used as experimental eyes.Urokinase of 4 μl (350 U) and the equal volume of PBS (0.01 mol/L) was intravitreally injected separately in the intravitreal urokinase group and the intravitreal PBS group,and 10 μl urokinase (1000U) and the equal volume of PBS was injected via retrobulbar tissue respectively as the retrobulbar urokinase group and the retrobulbar PBS group.Twenty-four hours after injection of drugs,0.5％ EB 4 μl was intravitreally injected.Four hours later,the rats were sacrificed and the right eyeballs were excised for the extraction and drying.EB was extracted from dried vitreous by formamide.Then,the concentration of EB in formamide was determined by a formamide extraction-ultraviolet spectrophotometry method to calculate the concentration of EB in vitreous.The use and care of experimental animals followed the Regulations for the Administration of Affair Concerning Experimental Animals by State Science and Technology Commission (2011 version).Results The rat vitreous body showed the light blue color in intravitreal urokinase group and the retinal vessels were visible under the microscope,and that in the retrobulbar urokinase group presented blue color.However,in the intravitreal and retrobulbar PBS group,rat vitreous exhibited the deeper blue color and retinas were invisible.Absorbance of EB in formamide was 0.181 ±0.008,0.450±0.017,0.330±0.009 and 0.436±0.012 in the intravitreal urokinase group,intravitreal PBS group,retrobulbar urokinase group and retrobulbar PBS group,respectively.The intravitreal EB concentrations in the intravitreal urokinase group were (0.266±0.014)g/L,which was lower than (0.667±0.026) g/L,(0.496±0.015) g/L and (0.657±0.017) g/L of the intravitreal PBS group,retrobulbar urokinase group and retrobulbar PBS group,showing significant different among the four groups (F =100.406,P＜0.01),and the intravitreal urokinase group showed the lowest value in comparison with other three groups (all at P＜0.01).Conclusions Local application of urokinase around eye can augment the outward permeability of blood-retinal barrier in rats.Intravitreal assay of EB after intravitreal injection is a feasible approach to the determination of outward permeability of blood-retinal barrier.

Objective:To investigate the prognostic significance of plasma fibrinogen (FIB) and D-dimer in elderly patients with non-small cell lung cancer (NSCLC) before any anti-cancer treatments.Methods:97 cases of patients with lung cancer (lung cancer group) and 36 healthy subjects (control group) were enrolled;the level of plasma D-dimer and FIB were compared;the relationship between their levels and clinical pathological factors of NSCLC as well as prognosis was analyzed.Results:The levels of plasma FIB and D-dimer in lung cancer group were higher than those in healthy control group (P＜0.05).FIB in lung cancer group was related to TNM stage,and D-dimer was related to lymph node metastasis and TNM stage.Univariate analysis showed that FIB,D-dimer,tumor size,lymph node metastasis and TNM staging were associated with overall survival time (OS) and progression free survival (PFS).Multivariate analysis revealed that only D-dimer and FIB were independent prognostic factors of patients with NSCLC.Conclusion:Detection of fibrinogen and D-dimer in elderly patients with NSCLC before anti-cancer treatment would be useful for prognosis and might provide a certain value for individualized therapy.

Objective To explore the relationship between the amount of bleeding and the histopathologic grade of cerebral glioma with susceptibility weighted imaging (SWI), so that to assess the diagnostic value of SWI in grading cerebral glioma. Methods Totally 30 patients with cerebral glioma underwent SWI. The area of hemorrhage of every slice displayed in SWI was measured with software. The rate and amount of bleeding were compared between high and low grade tumors. Results There was no statistical significance in the incidence of detected bleeding on SWI (P=0.064), though it was higher in highly malignant group (80.00%) than that in low grade group (46.67%), while there was statistical significance in the amount of bleeding detected on SWI (Z=-2.275, P=0.026) between highly malignant group and low grade group. Conclusion The amount of tumor hemorrhage displayed in SWI is valuable for the preoperative grading of cerebral glioma.

BACKGROUND:Although drug treatment.physics-behavior therapy,and postoperative therapy have been commonly used to treat stress urinary incontinence(SUI),there is still no satisfactory treatment at present.OBJECTlVE:To build a stable animal model simulating stress urinary incontinence(SUI)by bilateral transaction of pudendal nerve and nerves innervating pelvic floor muscles,including iliococcygeous muscle and pubococcygeous muscle.METHODS:A total of 18 6-week-old female SD rats weighing(1 99.44±8.41)g were randomly divided into 3 groups:normal,model,and sham-surgery groups,with 6 rats in each group.Rats in the model group underwent bilateral transaction of pudendal nerves and nerves innervating iliococcygeous/pubococcygeous muscles,while rats in the sham-surgery group had same procedures except nerve transaction.The normal group did not undergo any operation.Each rat was subjected to measure leak point pressure(LPP)at 2 weeks after the operation.After the measurement of LPP,cross sections of connection area of bladder and urethra were sent to histology.RESULTS AND CONCLUSION:One rat in the sham-surgery group died at 1 week after the operation.The LPP of model group decreased significantly by approximately 33%compared with the normal group(P＜0 05):however,there was no significant difference in LPP between sham-surgery and normal groups(P＞0.05).The results of histology showed loosely arrangement and atrophy of urethral sttriated muscle fibers in rats of the model group.Bilateral transaction of pudendal nerves and nerves innervating to iliococcygeous/pubococcygeous muscles resulted in SUI in rats stably.

BACKGROUND: Wnt signaling pathway plays an important regulative role in the embryonic development processes. Accordingly, it is of great significance to establish the cell model of Wnt signaling pathway so as to conduct study on it. OBJECTIVE: To establish Wnt signaling pathway cell model by transfecting L-M (TK-) cells with Wnt3a eukaryotic expression plasmid, and to investigate the effect of canonical Wnt signal pathway on the β-catenin subcellular distribution. METHODS: The eukaryotic expression plasmid pgk-Wnt3a-pcDNA3.0 after amplification was digested by restriction endonuclease first. Then it was transfected together with the control plasmid pgk-neo-pcDNA3.0 into L-M (TK-) cells via lipofection, after which the cell colony was screened by G418 for amplification. RT-PCR was used for detecting the expression products and the indirect immunofluorescence assay for observing the effect of Wnt3a on the β-catenin subcellular localization of L-M (TK-) cells. RESULTS AND CONCLUSION: The Wnt3a plasmid was verified by endonuclease digestion to have produced the expected plasmids after amplification. According to the RT-PCR detection to the 10 stably-transfected cell colonies achieved by 3 weeks of G418 screening, it was seen, on the L-Wnt3a cDNA, a strip of bright band of 320 bp in length, which showed that the products of amplification were exactly the expected fragments and that the Wnt3a plasmid was expressed on mRNA transcriptional level after being transfected with L-M (TK-) cells. In contrast, no expected band was found on the cDNA of L-M (TK-) calls transfecting the control plasmid. In addition, the immunofiuorescence assay detection showed that the protein expression of Wnt3a was found in the cytoplasm of the L-M(TK-) cells tranfecting Wnt3a plasmid, while for those transfecting the control plasmid, it was opposite. β-catenin, as showing by bright red fluorescence, was found to concentrate and enter into the nucleus of the L-M (TK-) cells transfecting Wnt3a plasmid, while for those transfecting the control plasmid, it was opposite. Cell model with continually activated Wnt signaling pathway is established. The stable expression of Wnt3a in L-M (TK-) cells transfected with pgk-Wnt3a-pcDNA3.0 is obtained. The expression of Wnt3a is able to promote the transfer of β-catenin from cytoplasms into nucleus in L-M (TK-) cells.

Objective To know the efficacy and safety of the blood sugar control by double-pump a-mong severe patients with duodenectomy. Methods Divided 60 severe patients with doudenectomy into the experimental group and the,control group randomly, there were 30 cases in each group. In the control group, pa-tients received intravenous infusion with the liquid containing glucose and neutral insulin (4g: 1 U), intravenous injection of 50U neutral insulin added into 50ml normal saline by micro-pump, the pump speed was In the ex-perimental group, sugar-containing liquid without insulin was infused within 24 hours by infusion pump with u-niform speed, a neutral insulin 50U mixed with 5Oral normal saline were injected by micro-pump according to the ratio fo glucose 4g: 1U neutral insulin through the same vein. Which also adjusted the pump speed according to maintaining blood sugar in the range of 4.4～8.3mmol/L. Compared the level of blood sugar control, treatment safety, efficacy and the workload of nurses between the two groups. Results The highperglycemic index and the time to achieve target values of blood sugar in the experimental group was shorter than in the control group respectively, while the time of keep blood sugar in idea level in the experimental group was longer than in the control group. There were no significant difference about incidence rale of hypoglycemia and workload of nurses between the two groups. Conclusions Double-pump can effective control blood sugar for severe patients with duodenectomy without burden nurses' workload, which is more reasonable than the traditional method.

Objective To evaluate the clinical application of CT-guided localization with combination of methylene blue and a Hookwire system for small pulmonary nodules (SPNs) before video-assisted thoracoscopic resection.Methods CTguided localization the SPNs before resection in 56 patients and 60 nodules,then underwent video-assisted thoracic surgery (VATS) resection.Among 56 patients,19 males and 37 females,aged from 35 to 81 years,mean age was (61.1 ±8.9)years.Results SPNs diameter (6.80 ±4.12) mm,distance from the parietal pleura (15.38 ±4.63) mm.CT-guided localization success rate was 100％,positioning time (10.76 ± 8.17) min,8.9％ (5/56) had micro pneumothorax aftet positioning,7.1％ (4/56) occurrence of needle tract bleeding,no conservative treatment.VATS resection rate was 100％.The pathology of 60 lesions were shown:Bronchiolo-alveolar carcinoma(BAC) were 33 lesions(55.0％),BAC and adenocarcinoma were 11 lesions(18.3％),Atypicaladenomatous hyperplasia (AAH) were 7 lesions (11.8％),Inflammation were 4 lesions (6.7％),Harmatoma were 3 lesions(5.0％),Tuberculoses were 2 lesions(3.3％).Conclusion CT-guided localization with combination of methylene blue and a Hookwire system before video-assisted thoracoscopic resection is a promising technique for small solitary pulmonary nodules.It could play an important role in accurate localization of small pulmonary nodules,and it is a safe technique with clinical application.