In this paper,300 bacteria strains and 31fungi strain were isolated from Qinghai plateau.The numbers of cellulose degradation organisms in soil is 2.6?10 5/g. A strain Trichoderma koningii No.0143 which produces cellulase was isolated from 11 fungi in the east area in QingHai,its FPA activity was 15u/g. It can be used in enzymatic feed.

OBJECTIVETo compare the changes of nucleus pulposus after in vitro culture of rabbit whole intervertebral disc and spinal motion segment.

METHODSTwenty-one New Zealand white rabbits which were randomly divided into organ group with 8 rabbits and segment group with 13 rabbits. Fifty intervertebral discs and 50 spinal motion segments were harvested respectively under aseptic conditions from two groups. These specimens were maintained in organ culture with hyperosmotic media (410 mOsm/kg), then 10 discs of the two groups were observed respectively by HE staining, immunohistochemistry of collagen type III, proteoglycan content and cells viability of nucleus pulposus before culture and at 3, 7, 14, 21 days after culture.

RESULTSHE staining showed the intervertebral disc tissue structure remained intact after culture of 21 days organ group and 14 days segment group,but there was severely degenerated of 21 days segment group. The intensity value of type II collagen immunohistochemical staining in the nucleus pulposus were not changed significantly between 21 days organ group and 14 days segment group (P > 0.05), but the staining of segment group at 21 days became shallower, there was significant difference compared with before each time points and organ group at 21 days (P < 0.05). PAS/AB staining of proteoglycan of nucleus pulposus showed that there were not decrease of tinting strength of two groups within 7 days, but the strength weakened slightly of two groups at 14 days, and the tinting strength became weaker at 21 days segment group, the change is more obvious than the organ group. The intensity value of fluorescence staining of nucleus pulposus cells was not changed significantly within 7 days of two groups (P > 0.05), the intensity value decreased slightly at 21 days organ group and 14 days segment group, but there were no significant difference compared with before time points (P > 0.05) however at 21 days segment group the intensity decreased as cells viability of nucleus pulposus decreased,and there was a significant difference compared with before each time points and organ group at 21 days (P < 0.05).

CONCLUSIONIt is not obviously degenerated of the discs of organ group cultured within 21 days and segment group cultured within 14 days, but there was significant degeneration of the intervertebral disc of segment group after cultured 21 days, so the rabbit spinal motion segment can be used on research about the biomechanics of intervertebral disc as a vitro experimental model within 14 days.

Animals ; Collagen Type II ; analysis ; Female ; Immunohistochemistry ; Intervertebral Disc ; chemistry ; pathology ; Male ; Organ Culture Techniques ; Rabbits

Objective To describe the hemodynamic changes during piggyback liver transplantation (PBLT), and to analyze the hemodynamic correlation with various degrees of cirrhosis according to Childpugh classification. Methods Between March 1999 and June 2004, 180 patients underwent PBLT procedure in our institution, and 95 cases were selected and divided according to Child classification. The intraoperative hemodynamics of different time points were retrospectively analyzed, including mean artery pressure (MAP), heart rate (HR), central vein pressure (CVP) and mean pulmonary artery pressure (MPAP). Results Hemodynamic changes were minimal before and during anhepatic phase in all the patients. At reperfusion, a hemodynamic disturbance occurred featured by decrease of MAP and increase of MPAP. Comparison between different Child class showed that in the Child C group, MAP were lower and HR were higher before new liver phase, while CVP and MPAP were higher during new liver phase. Conclusion Hemodynamic changes were minimal before and during anhepatic phase for PBLT, while they were more severe during reperfusion, and they also correlates with the different Child class before transplantation. The more severe of the cirrhosis before transplantation according to Child classification, the greater hemodynamic changes during the operation.

OBJECTIVEThis study was designed to detect the expression of bcl-2 and p53 proteins in colorectal carcinomas and to determine their association with the patient survival and stage of the diseases.

METHODSImmunohistochemistry method was used to detect the expression of bcl-2 and p53 proteins in 93 cases of colorectal carcinoma. The stain results were obtained by analyzing the clinic-pathological characteristics of patients.

RESULTSFifty-seven percent (53/93) of the colorectal carcinomas were bcl-2 protein positive. The positive rate of bcl-2 protein in lymph node involvement cases was lower (15/37) than the cases without node involvement (38/58, P<0.01). The positive rate of p53 protein was 43% (40/93) in colon-rectum carcinomas. No significant correlation was observed between p53 protein expression and clinic-pathological manifestations (P>0.05) but the survival was significantly worse (P=0.0001) in the p53 protein positive cases. Neither bcl-2 nor p53 alone was correlated with stage of the disease. When combined bcl-2/p53 status was analyzed, a group with bcl-2(+) and p53(-) had the best prognosis. This group was significantly associated with earlier Dukes' stages (P=0.1763). In multivariate Cox regression analysis, lymph node involvement and p53 protein expression were two independent factors correlated with survival time.

CONCLUSIONThe expression of bcl-2 and p53 represent biological characteristics of colorectal carcinomas. Assessment of both bcl-2 and p53 status may be valuable in predicting the prognosis of patients.

Biomarkers, Tumor ; metabolism ; China ; epidemiology ; Colorectal Neoplasms ; diagnosis ; metabolism ; mortality ; Female ; Humans ; Male ; Middle Aged ; Prevalence ; Prognosis ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Risk Assessment ; methods ; Risk Factors ; Survival Analysis ; Survival Rate ; Tumor Suppressor Protein p53 ; metabolism

OBJECTIVETo prevent and manage biliary complications after orthotopic liver transplantation (OLT).

METHODSNinety-five patients of OLT performed at our institute from February, 1999 to December 2002 were retrospectively analysed. Recipient operation was performed using standard method combined with veno-venous bypass in 12 patients and piggyback method in 78 patients and living-related liver transplantation in 5 patients. Biliary reconstruction was performed by end-to-end choledochocholedochostomy (C-C) over a T-tube in 55 patients and without a T-tube in 36 patients while the remaining 4 patients underwent Roux-en-Y choledochojejunostomy (CRY). C-C and CRY were performed by the interrupted or continuous suture with 5 - 0 or 6 - 0 Vicryl or PDS. Routine examination of liver function, Doppler ultrasonography and cholangiography were performed during the follow-up period.

RESULTSBiliary complications occurred in 7 patients (7.3%). Two patients with bile leakage at the anastomotic site developed biliary peritonitis on the seventh and tenth postoperative day and needed reoperation. One patient developed anastomotic biliary stricture one month after the operation and was cured by endoscopic stenting. Two patients developed bile leakage after T-tube removal. One of the two patients was treated conservatively and the other underwent a exploratory laparotomy to ligate the T-tube tract and drain the peritoneal cavity. One patient died of biliary vast syndrome five months after OLT and one patient died of biliary tract necrosis secondary to hepatic artery thrombosis on the tenth postoperative day. One - 42-month (mean 11.4 months) follow-up revealed no biliary stricture in 74 patients. No biliary stone and biliary sludge were detected by Doppler ultrasound and/or cholangiography. Serological examinations proved that liver grafts functioned well in these patients.

CONCLUSIONSTo prevent biliary complications, it is crucial to protect biliary mucosa and arterial blood supply of the common bile duct while harvesting the graft and to obtain perfect mucosa-to-mucosa apposition of no-tension end-to-end anastomosis of the bile duct. Endoscopic dilation and stenting are effective for post-OLT extrahepatic biliary stricture.

Adult ; Aged ; Biliary Tract Diseases ; etiology ; prevention & control ; therapy ; Biliary Tract Surgical Procedures ; adverse effects ; methods ; Female ; Follow-Up Studies ; Humans ; Liver Transplantation ; adverse effects ; methods ; Male ; Middle Aged ; Postoperative Complications ; etiology ; prevention & control ; therapy ; Retrospective Studies ; Young Adult

OBJECTIVETo find out how GATA-1 and GATA-2 behave in the bone marrow of patients with Monge's disease.

METHODSThe levels of mRNA in mononuclear cells (MNC) and proteins of GATA-1 and GATA-2 in the bone marrow of patients with Monge's disease and controls were determined by RT-PCR and immune cytolysis chemical method.

RESULTS(1) All patients and controls expressed GATA-1 mRNA (Monge's disease 1.033 +/- 0.146, Control 0.458 +/- 0.076) and GATA-2 mRNA (Monge's disease 0.451 +/- 0.073, Control 0.185 +/- 0.074). All patients expressed both GATA-1 (positive cell counts 77.3 +/- 33.3, positive score 135.4 +/- 75.4) and GATA-2 ( positive cell counts 29.4 +/- 11.4, positive score 48.4 +/- 19.7). All the controls expressed GATA-1 (positive cell counts 18.1 +/- 11.3, positive score 24.2 +/- 13.4) while 12 of 20 controls expressed GATA-2 ( positive cell counts 5.4 +/- 3.0, positive score 7.3 +/- 4.2). The expression of mRNA and proteins of GATA-1 and GATA-2 in Monge's disease were higher than in controls (P < 0.01). (2) There was a positive correlation between GATA-1 and Hb (P < 0.01), as did between mRNA and proteins of GATA-1 and GATA-2. (3) Both the proteins of GATA-1 and GATA-2 located only in the cytoplasm but not the nucleus.

CONCLUSIONSTwo of inherent genes, GATA-1 and GATA-2 which were expressed at higher levels in patients with Monge's disease than in controls might play significant roles in the pathogenesis of Monge's disease.

Adult ; Altitude Sickness ; metabolism ; GATA1 Transcription Factor ; metabolism ; GATA2 Transcription Factor ; metabolism ; Humans ; Male ; Polycythemia ; metabolism ; RNA, Messenger ; metabolism

OBJECTIVETo prepare microencapsulated cells releasing human tissue inhibitor of metalloproteinase-2 (TIMP-2), and investigate their biological characteristics in vitro.

METHODSChinese hamster ovary (CHO) cells were stably transfected with a human TIMP-2 expression vector, encapsulated in barium alginate microcapsules and cultured in vitro. Morphological appearance of the microcapsules was observed under a light microscope. Cell viability was assessed using MTT (3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyltetrazolium bromide) assay. Enzyme linked immunosorbent assay (ELISA) and reverse zymography were used to confirm the release of biologically active TIMP-2 from the microcapsules. Cryopreservation study of the microencapsulated cells was carried out using dimethyl sulfoxide (DMSO) as preservative agent.

RESULTSThe microcapsules appeared like a sphere with diameter of 300 - approximately 600 microm. The surface of the capsule wall was clearly smooth. The microencapsulated cells survived well and kept proliferating over the 6 weeks observed. No significant difference in TIMP-2 secretion was found between encapsulated and unencapsulated cells. Reverse zymography confirmed the bioactivity of MMP (matrix metalloproteinase) inhibition of TIMP-2. The cryopreservation process did not damage the microcapsule morphology nor the viability of the cells inside.

CONCLUSIONMicroencapsulated engineered CHO cells survive at least 6 weeks after preparation in vitro, and secrete bioactive TIMP-2 freely from the microcapsules.

Animals ; CHO Cells ; Cells, Immobilized ; Cricetinae ; Cryopreservation ; Humans ; Microspheres ; Recombinant Proteins ; biosynthesis ; genetics ; Tissue Engineering ; Tissue Inhibitor of Metalloproteinase-2 ; biosynthesis ; genetics ; Transfection

Endoplasmic reticulum (ER) stress occurs in macrophage-rich areas of advanced atherosclerotic lesions and contributes to macrophage apoptosis and subsequent plaque necrosis. The purpose of the present study was to investigate the effects of caveolin-1 (Cav-1) on ER stress-induced apoptosis in cultured macrophages and the underlying mechanisms. RAW264.7 cells were incubated with thapsigargin (TG) to establish ER stress model. And Cav-1 expression was detected by Western blot. After being pretreated with filipin(III), a caveolae inhibitor, RAW264.7 cells were assayed with flow cytometry and confocal laser scanning microscopy to detect cell apoptosis. Moreover, p38 mitogen-activated protein kinase (MAPK) phosphorylation and C/EBP homologous protein (CHOP) expression were detected with Western blot. The results showed that Cav-1 expression was markedly increased at early stage of TG treatment (P < 0.05) and then decreased with prolonged or high dose TG treatments. The increasing of Cav-1 expression induced by TG in RAW264.7 cells was abolished under inhibition of caveolae by filipin(III) (P < 0.05). The effect of TG on apoptosis of RAW264.7 cells was further augmented after pretreatment with filipin(III) (P < 0.05). Western blotting showed that MAPK phosphorylation induced by TG was inhibited by filipin(III) in RAW264.7 cells (P < 0.05), whereas CHOP remained unchanged (P > 0.05). These results suggest that Cav-1 may play a critical role in suppressing ER stress-induced macrophages apoptosis in vitro, and one of the mechanisms may be correlated with the activation of p38 MAPK prosurvival pathway.
Animals ; Apoptosis ; drug effects ; Caveolin 1 ; genetics ; metabolism ; Cell Line ; Endoplasmic Reticulum Stress ; physiology ; Filipin ; pharmacology ; MAP Kinase Signaling System ; Macrophages ; cytology ; drug effects ; Mice ; Thapsigargin ; pharmacology ; Transcription Factor CHOP ; metabolism ; p38 Mitogen-Activated Protein Kinases ; metabolism

Aqueous dispersion and stability of Fe3O4 nanoparticles remain an issue unresolved since aggregation of naked iron nanoparticles in water. In this study, we successfully synthesized different Fe3O4 super-paramagnetic nanoparticles which were modified by three kinds of materials [DSPE-MPEG2000, TiO2 and poly acrylic acid (PAA)] and further detected their characteristics. Transmission electron microscopy (TEM) clearly showed sizes and morphology of the four kinds of nanoparticles. X-ray diffraction (XRD) proved successfully coating of the three kinds of nanoparticles and their structures were maintained. Vibrating sample magnetometer (VSM) verified that their magnetic properties fitted for the super-paramagnetic function. More importantly, the particle size analysis indicated that Fe3O4@PAA had a better size distribution, biocompatibility, stability and dispersion than the other two kinds of nanoparticles. In addition, using CNE2 cells as a model, we found that all nanoparticles were nontoxic. Taken together, our data suggest that Fe3O4@PAA nanoaparticles are superior in the application of biomedical field among the four kinds of Fe3O4 nanoparticles in the future.

OBJECTIVETo investigate the effect of microRNA143 on cell proliferation and K-ras expression in colorectal carcinoma.

METHODSNorthern blot was used to examine the expression of miR-143 in colorectal carcinoma and adjacent normal tissues. A miR-143 expression vector was constructed and transfected into a human colon adenocarcinoma cell line SW480. Cell proliferation was evaluated by MTT assay. RT-PCR and Western blot were used to examine the expression of K-ras oncogene in transfected cells.

RESULTSThe level of mature miR-143 was lower in tumors compared with adjacent normal tissues in 81% of colorectal carcinoma specimens. In transfected cells, the increased accumulation of miR-143 inhibited the cell proliferation, and resulted in approximately 40.3% decrease of K-ras protein levels, but had no effect on level of K-ras mRNA.

CONCLUSIONThe increased accumulation of miR-143 inhibits the proliferation of transfected cells, and results in down-regulation of K-ras protein in colorectal carcinoma.

Adenocarcinoma ; genetics ; metabolism ; pathology ; Cell Line, Tumor ; Cell Proliferation ; Colonic Neoplasms ; genetics ; metabolism ; pathology ; Down-Regulation ; Genes, ras ; Genetic Vectors ; Humans ; MicroRNAs ; genetics ; metabolism ; Plasmids ; RNA, Messenger ; metabolism ; Transfection ; ras Proteins ; metabolism