Objective To design a new experimental facility to induce rat diffuse axonal injury model. Methods Rat diffuse axonal injury was induced by a new experimental facility,which was developed to let the rat head spin 90 degree at the moment to cause shearing injury.Vital sign and behavior of rat were measured simultaneously.The rats were sacrificed at 2 h,6 h,12 h,24 h,36 h,72 h and 10 d after injury,respectively,and the brain tissues were removed to prepare paraffin section.Then silver staining and HE staining were conducted to investigate changes of axonal fibers. Results There were unconsciousness,respiratory rhythm disturbance and hyporeflexia of pupil light reflex immediately after injury,and reactiveness decrease and activity retardation still existed even after resuming consciousness.At anatomical scene,subarachnoid hemorrhage or cerebroventricular haemorrhage were widespread.At an early stage,there were swelling,collapse,and axonal retraction ball formation at cortico-medulla junction,callosum,brainstem,and cerebellar white matter under microscope.But at the later stage,gitter cell proliferation and nest-like aggregation were major pathophysiological changes at focal brain tissue. Conclusion The new experimental facility is suit able to be used to induce rat diffuse axonal injury,since it is convenient,controlable,and precise.

Objective To investigate the effects of lymphangiogenesis on cancer growth and metastasis.Methods Human lymphatic endothelial cells(HLyECs)were isolated and purified from human lymph node by magnetic beads coated with antibody against human VEGFR3.Human breast cancer cell line(MDA-MB-435)or human osteosarcoma cell line(MG-63)was inoculated alone or co-inoculated with HLyECs subcutaneously into nude mice.The weight of tumor and lung surface metastasis were detected;peri-tumor lymphatic vessels were shown by Evans blue,intra-tumor lymphatic vessels were shown by immunohistochemistry for human and mouse PDPN.Conditioned medium of tumor on proliferation of HLyECs was evaluated by MTT assay.Results Compared with inoculating tumor cells alone,the growth and metastasis of MDA-MB-435 were promoted by co-inoculating HLyECs.The peritumoral and intra-tumoral lymphatic vessel density of MDA-MB-435 were increased by co-inoculating HLyECs.Both hPDPN-and mPDPN-positive lymphatic vessels were found.But co-inoculating HLyECs had no effect on lymphatic vessels density of MG-63.Neither intra-nor peri-tumor lymphatic vessels were found.The proliferation of HLyECs was increased by conditioned medium of MDA-MB-435 but not by MG-63.Conclusion Growth and lymphangiogenesis of breast cancer can be enhanced by co-inoculating lymphatic endothelial cells.

Objective To study the dynamic changes of transendothelial electrical resistance (TEER) on our in vitro model of brain blood barrier (BBB) made from primary culture of BALB/c mouse brain microvascular endothelial cells (BMVEC), and to explore the relation between TEER and BBB permeability, and search for the best culture condition. Methods BMVEC were isolated from BALB/c mouse and cultured on a transwell insert with special micro-pore. The cells were identified with immunohistochemical methods and electron microscope. TEER over BMVEC was measured after BBB model establishment for determining the 3H-Glucose permeability of BBB in vitro. Results BMVEC cultured in the transwell insert exhibited typical "flagstone" appearance and in a tight monolayer structure under electron microscope. Immunohistochemical detection of ZO-1 protein, a marker antigen of tight junction, showed smooth, continuous and tight junctions between confluent BMVEC. TEER over BMVEC monolayer increased to (346?10) ?/cm2 when the permeability for 3H-Glucose was decreased to the minimum. Conclusion BBB model in vitro made from primary culture of BMVEC in transwell has the basic characteristics of BBB in morphology, electrical resistance and permeability.

Objective To study the therapeutic effect of continuous renal replacement therapy (CRRT) in renal failure after liver transplantation.Methods Renal function in 82 patients who underwent CRRT in the perioperative period of liver transplantation were retrospectively analyzed.Results There were significant differences in ALT,TB,BUN and Cr before and after the treatment (P ＜ 0.05).The differences were significant in glutamic-pyruvic transaminase (ALT),creatinine phosphate kinase (CPK) and C reactive protein (CPR) before and after the treatment (P ＜0.05).There were significant improvements in K+,Na+,Cl-,HCO3-and CVP before and after the treatment (P ＜ 0.05),while the differences were not significant in other biochemical parameters (P ＞ 0.05).This research also looked at the effect of timing of CRRT on renal function recovery.Based on the RIFLE classification of AKI,the ratio of renal function recovery in RIFLE-Ⅰ was significantly higher than RIFLE-F (P ＜ 0.05).Conclusion CRRT treatment significantly improved the prognosis of patients with acute renal failure after liver transplantation.

ObjectiveTo summarize and investigate the incidence,reason,diagnosis and treatment of postoperative early cardiac arrhythmia after orthotopic liver transplantation(OLT).Method A retrospective study was made for the incidence,dignosis and treatment of cardiac arrhythmia (two weeks after OLT) following OLT from June 2004 to January 2012 in the Beijing You-An Hospital.ResultsIn 500 patients who received OLT,Cardiac arrhythmia developed in 82 patients (16.4％).Among these cardiac arrhythmia,35(7.0 ％) were sick sinus syndrome (including severe sinus bradycardia ),18 (3.6 ％ ) were paroxysmal supraventricular tachycardia,21(4.2 ％) were atrial fibrillation,8(1.6 ％,including 2 patients with torsades de pointes) were ventricular tachycardia and 4 (0.8 ％ )were cardiac arrest.Mortality rate after OLT relate to cardiac arrhythmia was 0.4％ (2 patients).Cardiac arrhythmia was mainly correlated with four factors:(1)whether patient had heart disease before OLT or not(x2 =15.82,P＜0.01),(2)Prolonged QT interval in patients with end-stage liver disease before OLT(x2 =11.00,P＜0.01).ConclusionsCardiac arrhythmia was common complication after OLT,and it can lead to death of recipients.Careful evaluation to recipients before OLT,controlling fluid load after OLT,keeping the balance of the electrolyte,acidity and alkalescence,giving intensive monitor to patients with heart disease before OLT and prolonged QT interval are the key factor to reduce incidence and mortality of cardiac arrhythmia.Application of medication and cardiac pacemaker can prevent cardiovascular accident after OLT.

Objective To observe the impact of mild hypothermia (MH)on the reactive oxygen species (ROS)and expression of cacpase-3mRNA and light chain 3 (LC3,a subunit of immunoglobulin)in hippocampus nerve cells of rats after cardiopulmonary resuscitation (CPR).Methods A total of 65 healthy male Sprague Dawley (SD)adult rats were randomly (random number)divided into 2 groups:blank control group (n =5)and CPR group (n =60).Cardiac arrest (CA)was induced in rats of CPR group by asphyxia.The survival rats after CPR were randomly (random number)divided into 2 groups:normothermia CPR group (NT)and hypothermia CPR group (HT).Homeothermia of 37 ℃ was maintained in NT group after restoration of spontaneous circulation (ROSC),and hypothermal intervention to 32 ℃ was carried out in HT group for 4 hours immediately after ROSC.Both NT group and HT group were then randomly divided to 2 subgroups 12 hours and 24 hours after ROSC (NT-12,NT-24,HT-12,HT-24 subgroups).During observation,the neurological deficit (NDS)of rats was scored,then the bilateral hippocampi were obtained from rats'head,and monoplast suspension of fresh hippocampus tissue was made immediately to determine the level of intracellular ROS by flow cytometry.Transmission electron microscope was used to observe the ultrastructure changes of cellular nucleus and mitochondria.Reverse transcription-polymerase chain reaction (RT-PCR)was used to determine the expression of caspase-3mRNA and Western-blotting (WB)was used to determine the level of LC3 in frozen hippocampus tissue.Measured data were analyzed with paired sample T test and One-Way ANOVA.Results Of 60 rats with CA,44 were successfully resuscitated (73%)and 33 survived until the end of the experiment (55%).The NDSs of rats in NT and HT groups were significantly reduced in comparison with BC group (F=8.107,P<0.05),while the NDSs of rats in HT-12 subgroup and HT-24 subgroup were significantly increased in comparison with NDSs of rats in NT-12 subgroup and NT-24 subgroup,respectively (t=9.692,P<0.01;t=14.374,P<0.01 ).The ROS in hippocampus nerve cells of rats in NT group and HT group were significantly increased compared to BC group (F=16.824,P<0.05 ),whereas the ROS in HT-12 and HT-24 subgroups were significantly reduced compared to ROS in NT-12 and NT-24 subgroups,respectively (t =9.836,P<0.01;t =7.499,P<0.01).The expressions of caspase-3 mRNA in hippocampus nerve cells of rats in NT and HT groups were significantly increased compared to BC group (F=24.527,P<0.05),while the expressions of caspase-3 mRNA in rats of HT-12 and HT-24 subgroups were significantly reduced compared to NT-12 and NT-24 subgroups,respectively (t =6.935,P <0.01;t =4.317,P <0.01 ).The level of LC3B-II/I in hippocampus nerve cells of rats in NT and HT groups were significantly increased compared to BC group (F=6.584,P<0.05),while the levels of LC3B-II/I in rats of HT-12 and HT-24 subgroups were significantly reduced compared to NT-12 and NT-24 subgroups,respectively (t=10.836,P<0.001;t=2.653,P=0.02).Ultrastructure damage of nucleus and mitochondria in NT group was more evident compared to BC group,and eumorphism of nucleus and mitochondria were maintained in rats of HT group compared to NT group.Conclusions The mild hypothermia reduced the injury of nerve cells and improved the neurological function of rats survived from cardiac arrest likely by reducing ROS production of nerve cells and inhibition the expression of caspase-3mRNA and lowering the level of LC3 leading to reducing cellular apoptosis and massive autophagy in rats survived from cardiac arrest after CPR.

OBJECTIVE:To develop a method for simultaneous determination of irinotecan(CPT-11)and its active metabo-lite 7-ethyl-10-hydroxycamptothecin(SN-38)in human plasma.METHODS:After precipitated by acetonitrile and acidified with hy-drochloric acid,using camptothecin as internal standard,the plasma sample was determined by HPLC-FLD. The determination was performed on Waters Luna C18column with mobile phase consisted of 0.05 mol/L sodium dihydrogen phosphate-acetonitrile(70:30, V/V,adjusted pH to 4.0 by phosphoric acid)at flow rate of 1 mL/min. The excitation wavelength was set at 380 nm;the emis-sion wavelengths of CPT-11 and SN-38 were set at 480 nm and 535 nm,respectively. The column temperature was 25 ℃ and the sample size was 20 μL. RESULTS:The linear ranges were 200-1 000 ng/mL for CPT-11(r=0.999 4,n=5)and 5-45 ng/mL for SN-38(r=0.999 2,n=5). RSDs of inter-day and intra-day were 1.68%-5.57%. The relative recoveries of CPT-11 and SN-38 were 90.12%-106.93%(RSD<8%,n=5)and 92.07%-102.56%(RSD<6%,n=5);the extraction recoveries of CPT-11 and SN-38 were 72.23%-86.56%(RSD<6%,n=5)and 71.98%-83.44%(RSD<7%,n=5),respectively. The plasma concentra-tions of CPT-11 and SN-38 in 5 patients with colon cancer were 431.13-617.19,13.97-31.89 ng/mL(1 h after intravenous drip-ping)and 398.14-584.43,11.61-29.94 ng/mL(2 h after intravenous dripping). CONCLUSIONS:The method is simple,rapid, sensitive,reproducible and suitable for the determination of plasma concentration and pharmacokinetic study of CPT-11 and its metabolite SN-38.

Background As a common form of gastric cancer migration,lymph node metastasis largely affects the surgical treatment and prognosis of gastric cancer.Surgery is the fundamental curative option for gastric cancer that varies depending on different stages.The study aimed to compare the clinicopathological characteristics and lymph node metastatic patterns in patients of proximal gastric cancer with different T stages and investigate a reasonable radical gastrectomy approach in terms of the range of lymphadenectomy for proximal gastric cancer.Methods In our retrospective study,the data of 328 patients of proximal gastric cancer with different T stages were analyzed.By comparing the differences of lymph node metastatic rate and ratio,we investigated the clinicopathological characteristics and metastatic patterns of lymph nodes.Also,we were especially interested in the differences in survival rates between patients with and without No.5 and 6 group metastasis with the same TNM stage.Results The overall lymph node metastatic rate and ratio of advanced proximal gastric cancer were 73.4％ and 23.3％,respectively.The tumors of different T stages were statistically significant in size and differentiation degree (P ＜0.05),multivariate analysis showed that the depth of tumor invasion was an independent risk factor for lymph node metastasis in proximal gastric cancer (RR,12.025; 95％ CI,2.326 to 62.157; P=0.003).The overall survival rate of patients with No.5,6 group lymph node metastasis and those without was significantly different,but the differences in survival rates between patients with and without No.5 and 6 group metastasis with the same TNM stage were not statistically significant.Conclusions Different T stages in proximal gastric cancer showed different patterns and characteristics of lymph node metastasis.D2 lymphadenectomy in patients with early gastric cancer had little survival benefit because metastasis to level 2 nodes was rare.Therefore the range of the lymph node dissection in radical gastrectomy for earty gastric cancer was considered reasonable.Moreover,to meet the requirements of the lymph node dissection,total gastrectomy plus D2 lymphadenectomy or more are supposed to be applied for the advanced proximal gastric cancer patients.Precise T staging larqely determines the range of gastrectomy and lymphadenectomy.

Objective To evaluate the safety and efficacy based on cytochrome P450(CYP)3A5*1 gene polymorphisms in guiding the individualized medication of tacrolimus (FK506) after liver transplantation. Methods Clinical data of 100 consecutively enrolled recipients who underwent liver transplantation for the first time were analyzed and randomly divided into experimental group and control group, with 50 cases in each group. The donors and recipients in the experimental group received preoperative CYP3A5 gene detection, and determined the FK506 medication regimen according to the CYP3A5*1 genotype. The compliance rate of FK506 target blood concentration, the recovery rate of liver function in the two groups of recipients at 7, 14, 28 d and 3, 6, 9, 12 months postoperatively, as well as the number of FK506 dosage adjustment during the follow-up were observed. The 1-year graft survival rate and the incidence of complications were recorded in both groups of recipients, such as acute rejection, infection, acute kidney injury, gastrointestinal symptoms, de novo hypertension, de novo diabetes, colds and rash, etc. Results The differences of the compliance rate of FK506 target blood concentration between the two groups of recipients at 7, 14 d after operation were statistically significant (both P < 0.05). There was no statistically significant difference between the two groups in the compliance rate of FK506 target blood concentration at 28 d and 3, 6, 9, 12 months and the recovery rate of liver function at the 7 observation time points after operation (all P > 0.05). The difference between the two groups of recipients in number of FK506 dose adjustment during follow-up was statistically significant (P=0.021). There were no statistically significant differences in 1-year graft survival rate and incidence of complications between the two groups of recipients after operation and during follow-up (all P > 0.05). Conclusions It is safe to guide individualized medication of FK506 after liver transplantation according to CYP3A5*1 gene polymorphism. It can increase the compliance rate of FK506 target blood concentration of recipients in the early postoperative stage, and can effectively reduce the number of dose adjustment duringfollow-up.

Objective To explore the safety application of organs from infectious donors. Methods Clinical data of 67 donors and recipients undergoing orthotopic liver transplantation were retrospectively analyzed. According to the occurrence of infections and infection sites in donors, all recipients were divided into the bloodstream infection group (n=16, donors with non-drug resistant bacterial infections), non-bloodstream infection group (n=20, donors with other site infections) and non-infection group (n=31). Perioperative clinical parameters including preoperative model for end-stage liver disease (MELD) score, operative time, anhepatic phase, intraoperative blood loss and intraoperative blood transfusion were statistically compared among three groups. The recovery of liver function and coagulation function in the recipients was observed at postoperative 1, 3, 7, 14 and 21 d. The incidence rate of complications and mortality rate in the recipients were recorded within 1 month after liver transplantation. The recovery of postoperative infection-related parameters including white blood cell (WBC), neutrophil pet (NE%) and procalcitonin (PCT) level in the recipients was observed. The application rate and application time of restricted antibiotics were recorded. Results Perioperative clinical parameters in the recipients did not significantly differ among three groups (all P > 0.05). At each time point after liver transplantation, the liver function, coagulation function, incidence rate of complications and mortality rate in the recipients did not significantly differ among three groups (all P > 0.05). The NE% of recipients at postoperative 3 and 7 d in the bloodstream infection group was significantly higher than those in non-bloodstream infection and non-infection groups (all P < 0.05). The PCT levels of recipients at postoperative 3, 7 and 14 d in the bloodstream infection group were significantly higher than those in the non-bloodstream infection and non-infection groups (all P < 0.05). The application rate and application time of restricted antibiotics in the recipients with bloodstream infections were significantly higher or longer than their counterparts in the non-bloodstream infection and non-infection groups (all P < 0.05). Conclusions It is safe to apply liver grafts from donors with bloodstream infection of non-drug resistant bacteria or other site infections when antibiotics are applied as early as possible.