Objective To set up the isolated perfused rat liver model. Methods Rat livers were harvested after the cannulation of the portal vein and bile duct. The reperfusion solution was Krebs-Henseleit solution containing bovine albumin serum and sodium taurocholate. The original CZ-1 isolated perfused rat liver system contained two subsystems: recirculating perfusion system and heat-exchange system. Then we modified the original CZ-1 system and omitted the heat-exchange system.The modified CZ-1 system consisted of a thermostatically regulated water bath,a peristaltic pump,a 4-neck round-bottom flask, a flow meter, an in-line manometer, a glass organ chamber, an iron support and a set of recirculating pipe line. Then the livers were connected via the portal vein to the modified CZ-1 system for 120 min. After 120 min reperfusion, bile production was evaluated. Routine HE staining and electron microscopic examination of hepatic tissues were also performed. Results The was not significantly different from that reported by references. Hepatic tissues in reperfusion group were also morphologically normal Conclusion The CZ-1 isolated perfused rat liver system was cost-effective and reliable to use. It was easy to run and is the ideal model for investigation of organ preservation solution.

Objective To investigate the effects of extracts of Ginkgo biloba leaves ( EGb761 ) on learning,memory and the hippocampal long-term potentiation (LTP) in chronic alcoholic rats.Methods 72 SD rats were randomly divided into control group,chronic alcoholism model group,low-dose EGb761 (100 mg/kg) and high-dose EGb761 (200 mg/kg) groups on average.55％ alcohol was given ( 10 ml/kg) by gavages once every day for 6 weeks to create the model of chronic alcoholism.In low-dose and high-dose EGb761 groups,EGb761 was used for gastric infusion 30 minutes before model.The control group and the model group were infused with the same volume of distilled water.Morris water maze (MWM) was used to study the spatial learning and memory in rats.Electrophysiological methods were used to observe the changes of LTP through the comparison of field excitatory postsynaptic potential (fEPSP) slope in hippocampal CA3 region before and after high frequency stimulation (HFS).Results ( 1 ) The escape latency (EL) of model group ( (93.31 ± 14.21 ) s) was longer than that of the control group( (35.28 ±6.51 )s).The mean slope of fEPSP (relative to baseline) after HFS in model group was (96.91 ±7.65) ％,in control group was ( 177.22 ± 10.12) ％ ; Compared with control group,the ability of learning and memory as well as LTP obviously descended in model group.(2) Compared with model group,EGb761(100,200 mg/kg) could shorten the EL.The slope of fEPSP after HFS in either low-dose (( 126.16 ± 5.84)％ )or high-dose (141.31 ± 5.75)％ EGb761 groups was enhanced significantly compared with the model group (96.91 ± 7.65 ) ％,P =0.034,P =0.009 ).Conclusion EGb761 can significantly improve the alcohol-induced memory impairment by means of accelerating the recovery of the pathological synaptic plasticity.

Objective To discuss the application of transanal placement of ileus tube in treatment of colonic obstruction in left colorectal cancer.Methods Thirty-two patients with left malignant colorectal obstruction were divided into ordinary colonoscope group (16 cases) and superslim endoscope group (16 cases) by random digits table.The catheter success rate,catheter operating time and exposure to X-ray time was compared between two groups.Results Fifteen cases were successful and 1 case was failed in ordinary colonoscope group.Thirteen cases were successful and 3 cases were failed in superslim endoscope group.There was no significant difference in the catheter success rate between two groups (x2 =1.143,P =0.285).The catheter operating time was (42 + 15),(20 +6) min in ordinary colonoscope group and superslim endoscope group,and there was significant difference between two groups (t =3.895,P =0.005).The exposure to X-ray time was (20 + 12),(5 + 2) min,and there was significant difference between two groups (t =-3.596,P =0.007).Conclusion Transanal placement of ileus tube is more successful and convenient by superslim endoscope combined with anesthesia than by ordinary colonoscope.

Objective By using primary cultured mouse renal tubular epithelial cells (TECs) to develop an in vitro model of simulated ischemia-reperfusion (IR) injury.Methods The outer medulla of C57BL/6J mouse kidney was flushed and primary cultured after digestion in type Ⅰ collagenase,and then immunocytochemical staining was used to verify TECs.Primary cultured TECs were immersed in mineral oil to simulate the ischemic process,and 60 min later the whole culture medium was added to simulate reperfusion process.The cells were collected and RAN was extracted at indicated time points after medium replacement.The expression of TNF-α,IL-1β and IL-6 was detected by using real-time fluorescence quantitative RT-PCR. The culture supernatants were collected at 24 h after medium replacement for detection of the expression of cytokine protein by using ELISA.Results Primary cultured TECs were identified by cobblestone-shaped morphology and then verified by cytokeratin 18 (CK18) staining.In TECs of IR group after medium replacement the mRNA expression of TNF-α,IL-1β and IL-6 was higher than in control group.The expression of TNF-α after medium replacement was increased to a peak level at 0.5 h,about (24.45 ±6.51) times (P＜0.01 ) higher than the control group,and gradually declined thereafter.The mRNA expression of IL-1β after medium replacement kept an increasing tendency,about ( 15.27 ± 4.29) times (P＜0.05) higher than the control group at 6 h,and that of IL-6 after medium replacement was increased to a peak level at 3 h,about ( 11.19 ±4.55) times (P＜0.01) higher than the control group. In the IR group at 24 h after medium replacement,the protein expression of NF-α,IL-1β and IL-6 in the supernatants was significantly higher than in the control group.Conclusion High purity of primary cultured TECs was achieved from the outer medulla of mouse kidney by separation and digestion.The in vitro model of simulated IR in primary cultured mouse renal TECs was successfully created using paraffin oil.

ObjectiveTo investigate the effect and underling mechanism of water-soluble CO-releasing molecules (CORM-3)on the alleviation of allograft rejectionafter mouse kidney transplantation.Methods A mice kidney transplantation model was established using C.FVB-Tg (Itgax-DTR/GFP)57Lan/J or C57BL/6J (H-2Kb) mice as donors,and Balb/c (H-2Kd) mice as recipients.After donor nephrectomy,kidney was preserved in UW solution which contained CORM-3 or iCORM (inactive CO-releasing molecules) for 24 h in 4℃.Recipient survival after removal of both na? ve kidneys,serum creatinine as well as graft histology was observed.In the C.FVB-Tg(ItgaxDTR/GFP) 57Lan/J donors,rDCs were acquired in vitro and selected by magnetic cell sorting (MACS) after graft nephrectomy.The expression of activation markers,CD80 and CD86,on rDC was assessed by using flow cytometry.ResultsThe graft medium survival time was 40.5 days in the iCORM group and 70 days in the CORM-3 group respectively (P＜0.05).CORM-3 preserved the graft function as shown by significantly lower serum creatinine (P＜0.05; or P＜0.01) and alleviated graft pathology injury.Diffuse infiltration of mononuclear cells in the interstitial tissues,moderate tubulitis and partial glomerular sclerosis were found in the iCORM graft kidney,while the CORM-3 graft kidney displayed almost normal histology.Meanwhile,CORM-3 suppressed the expression of CD80 and CD86 in donor-derived rDC.ConclusionCORM-3 can alleviate allograft rejection,prolong the graft survival,and improve kidney function in mouse kidney transplantation,probably via inhibiting rDC activation.

ObjectiveTo investigate the role of aldehyde dehydrogenase 2 (ALDH2) in the protection against tubular epithelial cells (TEC) ischemia/reperfusion (IR)injury induced by pretreatment with ethanol.Methods Mouse primary cultured TECs were pretreated with 50 mM ethanol 3 h before simulation of in vitro IR.Lactate dehydrogenase (LDH) release was assessed to evaluate the protection of ethanol pretreatment on IR injury.Thereafter,TECs were transfected with a negative control siRNA (NC) or an ALDH2-siRNA. The ALDH2 protein levels and ALDH enzymatic activities were assessed 48 h after transfection.Ethanol pretreatment and in vitro IR were performed on those transfected TECs.LDH release was assessed to evaluate the role of ALDH2 in the ethanol pretreatment-induced protection against IR injury.ResultsEthanol pretreatment significantly reduced the LDH release in TECs upon IR insult.As compared with NC group and INTERFERin group,the ALDH2 protein levels were decreased by 82.1％,ALDH enzymatic activities were decreased hy 67.3％,and the protective effect induced by ethanol pretreatment was almost completely abrogated in ALDH2-siRNA group.ConclusionEthanol pretreatment protects TECs against IR injury through ALDH2 dependent pathways.

Background and purpose:Epithelial ovarian cancer has the highest mortality rate of gynecologic cancers and overall survival rates have improved little in the last 20 to 30 years. CXXC ifnger protein 5 (CXXC5) plays an important role in AML (acute myeloid leukemia) and MDS (myelodysplasia). However, little is known about its clinical signiifcance and biological function in epithelial ovarian cancer. This study aimed to investigate the expression of the CXXC5 in ovarian cancer and the effect of the CXXC5 on ES-2 cell proliferation. Methods:①The alteration of CXXC5 in cancer genomics data of TCGA (Cancer Genome Atlas) was analyzed.②The CXXC5 protein in the tissue chips was detected containing 37 benign ovarian cyst and 173 malignant tumor samples. The relationship between the expression of the CXXC5 with the clinicopathological features of patients with ovarian cancer was analyzed by SPSS software;③The cells with the highest CXXC5 expression quantity from 5 ovarian cancer cells were selected by re-al-time quantitative PCR (qRT-PCR) and Western blot.④ES-2 cells with shRNA stable transfection were construted us-ing the strategy of lentivirus infection and analyzed cell proliferation by cell counting kit-8(CCK8). Results:①Through the TCGA database, CXXC5 ampliifcation was found in 7 of 563 cases.②The CXXC5 expression in ovarian malignant carcinoma (39.3%) was higher than that in benign ovarian cyst (13.5%, P=0.003), the histologic type was highly asso-ciated with CXXC5 (43%in serous, 22.9%in mucinous, 23.5%in endometrioid, 67%in clear cell, P=0.014) and there was a signiifcant correlation between CXXC5 and lymph node metastasis (positive vs negative, P=0.022).③The ES-2 cells with shRNA stable transfection had a growth disadvantage (P<0.05). Conclusion:The CXXC5 gene might have an advantage in proliferation of epithelial ovarian carcinoma and be expected to become the biomarker of poor prognosis.

OBJECTIVETo investigate the effects of Naoxinduotai capsule on the markers of prothrombotic state as plasminogen activator inhibitor-1 (PAI-1), von Willebrand factor (vWF) and alpha-granular membrane protein (CD62p) in essential hypertension patients of yang hyperactivity and blood stagnation.

METHODThe 62 essential hypertension patients of yang hyperactivity and blood stagnation were divided into Naoxinduotai capsule + perindopril group (treatment group, 30 subjects) and Perindopril group (control group, 32 subjects). Clinical symptoms, blood pressure and the blood plasma PAI-1, vWF, CD62p of the patients were observed. The blood plasma PAI-1, vWF and CD62p were measured by enzyme-linked immunosorbant assay (ELISA).

RESULTAfter 8 weeks treatment, in treatment group the clinical symptoms became better, and there was a significant difference with control group (P < 0.05). Blood pressure was significantly degraded, but there was not a significant difference with control group. The blood plasma PAI-1, vWF, CD62p level was degraded ,and there were significant differences with control group in all the three makers (P < 0.05).

CONCLUSIONNaoxinduotai capsule can treat the essential hypertension patients of Yang hyperactivity and blood stagnation, and it has conspicuous advantages in improving the clinical symptoms and the makers of prothrombotic state.

Aged ; Biomarkers ; blood ; Blood Pressure ; drug effects ; Capsules ; Female ; Humans ; Hypertension ; blood ; drug therapy ; physiopathology ; Male ; P-Selectin ; blood ; Plasminogen Activator Inhibitor 1 ; blood ; Platelet Membrane Glycoproteins ; metabolism ; Thrombosis ; blood ; Treatment Outcome

OBJECTIVETo research the relationship between the virulence factors of Saccharomyces albicans (S. albicans) and the random amplified polymorphic DNA (RAPD) bands of them, and establish the regression model by multiple regression analysis.

METHODSExtracellular phospholipase, secreted proteinase, ability to generate germ tubes and adhere to oral mucosal cells of 92 strains of S. albicans were measured in vitro; RAPD-polymerase chain reaction (RAPD-PCR) was used to get their bands. Multiple regression for virulence factors of S. albicans and RAPD-PCR bands was established.

RESULTSThe extracellular phospholipase activity was associated with 4 RAPD bands: 350, 450, 650 and 1 300 bp (P < 0.05); secreted proteinase activity of S. albicans was associated with 2 bands: 350 and 1 200 bp (P < 0.05); the ability of germ tube produce was associated with 2 bands: 400 and 550 bp (P < 0.05).

CONCLUSIONSome RAPD bands will reflect the virulence factors of S. albicans indirectly. These bands would contain some important messages for regulation of S. albicans virulence factors.

DNA ; Polymerase Chain Reaction ; Random Amplified Polymorphic DNA Technique ; Saccharomyces ; Virulence Factors

To better understand the effect of a new split variant of human asialoglycoprotein receptor (ASGPR H1b) on ASGPR ligands' binding ability, we established a functional cell line which expresses ASGPR. The full lengths of ASGPRH1a and H2c fragments from human liver were amplified by reverse transcript PCR (RT-PCR) and inserted into eukaryotic expression vector pIRES2EGFP, pCDNA3.1 (Zeo+) respectively. The recombinants were co-transfected into HeLa cells. After selection by using Neocin and Zeocin, a stably transfected cell line was established, which was designated 4-1-6. The transcription and expression of ASGPRH1a and H2c in 4-1-6 were confirmed by RT-PCR, Western blotting and immunofluorescence. The endocytosis function of the artificial "ASGPR" on the surface of 4-1-6 was tested by FACS. It was found that the cell line 4-1-6 could bind ASGPR natural ligand molecular asialo-orosomucoid (ASOR). After the eukaryotic plasmid H1b/pCDNA3.1 (neo) was transfected into cell line 4-1-6, H1b did not down-regulate the ligand binding ability of ASGPR. The eukaryotic expression plasmid H1b/pcDNA3.1 (neo) and H2c/pcDNA3.1 (neo) were co-transfected transiently into Hela cell. Neither single H1b nor H1b and H2c could bind ASOR. In conclusion, a functional cell line of human asialoglycoprotein receptor (ASGPR) which expresses both H1a and H2c stably was established. The new split variant H1b has no effect on ASGPR binding to ASOR. ASGPRH1b alone can't bind to ASOR, it yet can't form functional complex with ASGPRH2c.