Hypoxia-induced 15-hydroxyeicosatetraenoic acid (15-HETE) is an essential mediator to constrict pulmonary arteries (PA). The signaling pathway involved in 15-HETE-induced PA vasoconstriction remains obscure. The aim of the present study was to test the hypothesis that hypoxic PA constriction induced by 15-HETE was possibly regulated by the extracellular signal-regulated kinase-1/2 (ERK1/2) pathway. PA ring tension measurement, Western blot and immunocytochemistry were used in the study to determine the possible role of ERK1/2 in 15-HETE-induced PA vasoconstriction. The organ bath for PA rings tension study was employed. Adult male Wistar rats were raised in hypoxic environment with fractional inspired oxygen (FIO2, 0.12) for 9 d. PA 1~1.5 mm in diameter were dissected and cut into 3 mm long rings for tension study. ERK1/2 up-stream kinase (MEK) inhibitor PD98059, which blocks the activation of ERK1/2, was used. The results showed that pretreatment of PD98059 significantly blunted 15-HETE-induced PA vasoconstrictions in the rings from hypoxic rat. Moreover, in endothelium-denuded rings, PD98059 also significantly attenuated 15-HETE-induced vasoconstriction. Phosphorylation of ERK1/2 in pulmonary arterial smooth muscle cells (PASMCs) of rat was enhanced evidently when stimulated by 15-HETE. Thus, the data suggest that ERK1/2 signaling pathway is involved in 15-HETE-induced hypoxic pulmonary vasoconstriction.
Animals ; Flavonoids ; pharmacology ; Hydroxyeicosatetraenoic Acids ; antagonists & inhibitors ; pharmacology ; Hypoxia ; physiopathology ; MAP Kinase Signaling System ; physiology ; Male ; Muscle, Smooth, Vascular ; cytology ; Myocytes, Smooth Muscle ; drug effects ; Pulmonary Artery ; cytology ; drug effects ; physiopathology ; Rats ; Rats, Wistar ; Vasoconstriction ; drug effects

OBJECTIVETo study the effect of glutamine on intestinal barrier function by examining the changes of plasma D-lactic levels and diamine oxidase (DAO) levels in plasma and intestinal tissue after glutamine intervention in young rats with endotoxemia.

METHODSEighty 18-day-old rats were randomly divided into endotoxemia and glutamine intervention groups (n=40 each). Endotoxemia was induced by lipopolysaccharide (LPS) injection. Plasma and small intestine homogenate were collected 1.5, 6, 24 and 72 hrs and 7 days after LPS injection. The glutamine intervention group was immediately administered with oral glutamine (2 g/kg) after LPS injection. Afterwards, glutamine was administered once daily. Plasma D-lactic and DAO levels and intestinal DAO levels were measured.

RESULTSPlasma DAO activity in the glutamine intervention group was significantly lower than that in the endotoxemia group 6 and 72 hrs after LPS injection (P<0.05). In contrast, the intestinal DAO activity in the glutamine intervention group was significantly higher than that in the endotoxemia group 6, 24 and 72 hrs and 7 days after LPS injection (P<0.05 or 0.01). Plasma D-lactic levels in the glutamine intervention group were significantly lower than those in the endotoxemia group 6, 24 and 72 hrs and 7 days after LPS injection (P<0.01).

CONCLUSIONSGlutamine may reduce the permeability of intestinal mucosa, and thus provides protective effects on intestinal barrier function in rats with endotoxemia.

Administration, Oral ; Amine Oxidase (Copper-Containing) ; metabolism ; Animals ; Behavior, Animal ; Endotoxemia ; metabolism ; Female ; Glutamine ; administration & dosage ; pharmacology ; Intestines ; drug effects ; metabolism ; Lactic Acid ; blood ; Male ; Rats ; Rats, Wistar

OBJECTIVETo obtain the monoclonal antibody against hexon protein of human adenovirus.

METHODSBALB/c mice were immunized with purified recombinant hexon protein, and the spleen cells of the mice were isolated and fused with myloma cells. Four hybridoma cell strains were screened by indirect ELISA and cultured, and the sensitivity, specificity and virus neutralizing activity were analyzed with ELISA, Western blotting and neutralizing test.

RESULTSThe mouse ascites produced by these hybridoma cells contained specific monoclonal antibodies against hexon protein of human adenovirus as identified by ELISA and Western blot, and the antibody generated by 4C6 strain showed human adenovirus type 3-neutralizing activity.

CONCLUSIONThe monoclonal antibodies against hexon protein with high specificity have been successfully obtained, and these antibodies can be useful in developing assays for early diagnosis of HAdV3 infection and also in study of therapeutic drugs of the infection.

Adenoviruses, Human ; chemistry ; immunology ; Animals ; Antibodies, Monoclonal ; biosynthesis ; immunology ; Antibodies, Viral ; biosynthesis ; immunology ; Blotting, Western ; Capsid Proteins ; biosynthesis ; genetics ; immunology ; Enzyme-Linked Immunosorbent Assay ; Escherichia coli ; genetics ; Humans ; Hybridomas ; secretion ; Mice ; Mice, Inbred BALB C ; Recombinant Proteins ; biosynthesis ; immunology

Objective Data on the leukapheresis from 26 pediatric patients with hematologic or solid malignancies was retrospectively evaluated to screen predictive factors affecting the efficacy of peripheral blood stem cell(PBSC) collection from donors,as well as hematopoietic recovery in recipients.Methods We present our experience with 49 apheresis from 26 granulocyte-colory Stimulating factor mobilized donors and analyzed the correlations between the mobilization,the leukocyte count in the donor peripheral blood and the MNC and CD_(34)~+ cell yields in collecting products and the neutrophil and platelet recovery of recipients.Results The process of mobilization and apheresis were well tolerated by our pediatric donors.The median numbers for harvested MNCs and CD_(34)~+ cells were 4.5?10~8/kg and 1.9?10~6/kg of recipient body weight,respectively.Mobilizing dose positively affected the number of mononuclear ceus(MNC) but not CD_(34)~+ cells in the apheresis products.The CD_(34)~+ cell number in the apheresis product was influenced significantly by donor circulating MNC on the day of harvest and correlated with recipient′s engraftment after PBSC was reinfused.Conclusions The MNC yield was stable and met with the demand for autologous stem cell transplantation while the CD_(34)~+ cell number varies obviously from each donor.Since a rapid engraftment was associated with a high number of CD_(34)~+ cells collected,which was in turn predicted by the level of the pre-apheresis CD_(34)~+ cells in the peripheral blood of donors,it is necessary to monitor the donors′ CD_(34)~+ cell during mobilization to determine the optimal time for apheresis.J Appl Clin Pediatr,2006,21(3):148-150

15-hydroxyeicosatetraenoic acid (15-HETE) plays an important role in hypoxia-induced pulmonary vasoconstriction. Release of nitric oxide (NO) is apparently decreased and activity of endothelial nitric oxide synthase (eNOS) is impaired in chronic hypoxia. However, little is known whether 15-HETE contributes to eNOS/NO pathway in the constriction induced by 15-HETE. We examined the response of rat pulmonary artery (PA) rings to 15-HETE, the production of NO, total eNOS expression and the phosphorylation of eNOS in bovine pulmonary artery endothelial cells (BPAECs) stimulated by 15-HETE. Rat PA rings were divided into three groups: endothelium intact group, endothelium denuded group, and nitro-L-arginine methyl ester (L-NAME, 0.1 mmol/L, an inhibitor of eNOS) group. Constrictions to 15-HETE were significantly enhanced in endothelium denuded group and L-NAME group (both P< 0.05 vs endothelium intact group, n= 9); BPAECs were incubated in different conditions to test nitrite production by Greiss method. Nitrite production was significantly reduced by 1 mumol/L 15-HETE (P<0.05), and increased by the lipoxygenase inhibitors, 10 mumol/L cinnamyl 3,4- dihydroxy-[alpha] -cyanocinnamate (CDC, P< 0.05) and 0.1 mmol/L nordihydroguiairetic acid (NDGA, P< 0.01 ); Western blot analysis of extracts from BPAECs incubated with 15-HETE in different time was carried out to test total eNOS expression, and the expression was changed unobviously. Immunoprecipitation (IP) and Western blot analysis of cell extracts from BPAECs treated with 2 mumol/L 15-HETE in different length of time were accomplished, using phospo-eNOS-threonine 495 (Thr495, an inhibitory site) antibody for IP, and eNOS or 15-lipoxygenase (15-LO) antibodies for Western blot. 15-HETE depressed eNOS activity by increasing the levels of phospho-eNOS-Thr 495. The data suggest that eNOS/NO pathway is involved in PA constrictions induced by 15-HETE and that 15-HETE depresses eNOS activity by phosphorylation in Thr495 site. The protein interaction between phospho-eNOS (Thr495) and 15-LO is discovered for the first time.
Animals ; Cattle ; Down-Regulation ; drug effects ; Endothelium, Vascular ; cytology ; drug effects ; enzymology ; Hydroxyeicosatetraenoic Acids ; pharmacology ; In Vitro Techniques ; Male ; Nitric Oxide Synthase Type III ; metabolism ; Pulmonary Artery ; cytology ; enzymology ; physiology ; Rats ; Rats, Wistar

Objective　Coronary artery bypass grafting (CABG)i n 13 patients was analyzed. Methods　9 patients were performed C ABG with cardiopulmonary bypass, 4 patients undergone off-pump coronary artery bypass(OPCAB). Among the 4 patients, 3 undergone transmyocardial laser revascula rization concomitantly. 2 patients with single-vessel disease, 3 with double-v essel disease, 7 with triple-vessel disease and 1 with left main coronary arter y disease. The average bypass per patient was 2.3. Results　All patients survived, 11 patients were angina free, 2 were angina relief. C onclusion　CABG is a safe operation, OPCAB may reduce blood transfusion and complication, patients recover more quickly after OPCAB compared with those with CABG.

Objective To explore the application efficacy of artificial nose in CCU patients with tracheal intubation.Methods Totals of 62 patients with tracheal intubation were randomly divided into the group of artificial nose and group of electrothermal and constant temperature moist,then the sputum viscosity,irritating cough,airway administration hours,average intubation time,consumables costs,and the incidence of ventilator-associated pneumonia in two groups were observed and compared.Results In artificial nose group,the incidence of irritating cough was28.1％,the incidence of ventilator-associated pneumonia was 15.6％,airway administration hours was( 2.3 ± 0.5 ) h,average intubation time was ( 112 ± 6.5 ) h,consumables costs was (44.2±6.7)yuan all better than that of electrothermal and constant temperature moist group,which was 71.9％,37.5％,( 3.5 ± 0.6 ) h,( 133 ± 7.8 ) h,and ( 56.3 ± 1.5 ) yuan,respectively,differences were statistically significant ( x2 =12.25,3.925 0; t =8.660 5,11.699 8,8.212 9,respectively; P ＜ 0.01 or P ＜0.05).Sputum viscosity status of artificial nose group was that Ⅰ grade 12.5％,Ⅱ grade 62.5％,Ⅲ grade 25.0％,whilethat of electrothermal and constant temperature moist group was 46.9％,25.0％,28.1％,respectively,and the differences was statistically significant ( x2 =11.559 0,P ＜ 0.01 ).Conclusions Artificial nose using in patients with endotracheal intubation can achieve the desired efficacy of airway humidification,improve the efficacy of airway administration,save energy and time and significantly reduce the incidence of ventilator-associated pneumonia,which likely to be generally used in clinical.

BACKGROUNDNowadays bone marrow represents the main source of mesenchymal stem cells (MSCs). We identified a new population of MSCs derived from human placenta and compared its biological characterization with bone marrow derived MSCs.

METHODSMononucleated cells (MNC) were isolated from the human placenta tissue perfusate by density gradient fractionation. Individual colonies were selected and cultured in tissue dishes. At the same time, human bone marrow derived MSCs were identified. Culture-expanded cells were characterized by immune phenotyping and cultured under conditions promoting differentiation to osteoblasts or adipocytes. The hematopoietic cytokines were assayed using reverse transcriptase polymerase chain reaction (RT-PCR).

RESULTSHuman placental MSCs exhibited fibroblastoid morphology. Flow cytometric analyses showed that the placental MSC were CD29, CD44, CD73, CD105, CD166, HLA-ABC positive; but were negative for CD34, CD45, and HLA-DR. Functionally, they could be induced into adipocytes or osteocytes. Moreover, several hematopoietic cytokine mRNA was found in placenta-derived MSCs by RT-PCR analysis, including IL-6, M-CSF, Flt3-ligand and SCF. These results were consistent with the properties of bone marrow derived MSCs.

CONCLUSIONThese observations implicate the postpartum human placenta as an important and novel source of multipotent stem cells that could potentially be used for investigating mesenchymal differentiation and regulation of hematopoiesis.

Bone Marrow Cells ; physiology ; Cell Differentiation ; Cells, Cultured ; Female ; Humans ; Mesenchymal Stromal Cells ; physiology ; Placenta ; cytology ; Pregnancy

Osteosarcoma is the most common primary malignant bone cancer in children and adolescents. Emerging evidence has suggested that the capability of a tumor to grow is driven by a small subset of cells within a tumor, termed cancer stem cells (CSCs). Although several methods have been explored to identify or enrich CSCs in osteosarcoma, these methods sometimes seem impractical, and chemotherapy enrichment for CSCs in osteosarcoma is rarely investigated. In the present study, we found that short exposure to chemotherapy could change the morphology of osteosarcoma cells and increase sarcosphere formation in vitro, as well as increase tumor formation in vivo. Furthermore, methotrexate (MTX)-resistant U2OS/MTX300 osteosarcoma cells were larger in size and grew much more tightly than parental U2OS cells. More importantly, U2OS/MTX300 cells possessed a higher potential to generate sarcospheres in serum-free conditions compared to parental U2OS cells. Also, U2OS/MTX300 cells exhibited the side population (SP) phenotype and expressed CSC surface markers CD117 and Stro-1. Notably, U2OS/MTX300 cells showed a substantially higher tumorigenicity in nude mice relative to U2OS cells. Therefore, we conclude that chemotherapy enrichment is a feasible and practical way to enrich osteosarcoma stem cells.
Animals ; Antigens, Surface ; metabolism ; Antimetabolites, Antineoplastic ; pharmacology ; Bone Neoplasms ; metabolism ; pathology ; Cell Line, Tumor ; Cell Proliferation ; Drug Resistance, Neoplasm ; Humans ; Methotrexate ; pharmacology ; Mice ; Mice, Nude ; Neoplasm Transplantation ; Neoplastic Stem Cells ; drug effects ; pathology ; Osteosarcoma ; metabolism ; pathology ; Phenotype ; Proto-Oncogene Proteins c-kit ; metabolism

OBJECTIVEWith the extremity soft tissue sarcoma close to neurovascular bundle, combined en bloc resection and brachytherapy or simple en bloc resection were performed to evaluate the treatment outcome of the combined en bloc resection and brachytherapy.

METHODSRetrospectively investigation was performed for the extremity soft tissue sarcoma close to neurovascular bundle between 2000 and 2009. Inclusion criteria were primary extremity soft tissue sarcoma, MRI showed that the reaction zone involved the main neurovascular bundle, and the reaction zone closed less than 1 cm to the main neurovascular bundle. 86 cases were included in the study. There were 41 men and 45 women. The average age was 38.5 years old (Range from 15 to 73). There were malignant fibrous histiocytoma, synovial sarcoma, fibrosarcoma, liposarcoma, clear cell sarcoma, epithelioid sarcoma, leiomyosarcoma, rhabdomyosarcoma and vascular sarcoma etc. The stage were IA (8), IIA (12), IIB (10), IIC (7), III (43) and IV (6).

RESULTSDuring an average follow-up of 53 months (range 24 - 102 months), the distant metastasis rate 32.56% (28/86) and the lymph node metastasis rate was 6.98% (6/86). The local recurrence rates was 13.95% (12/86). In the group of combined en bloc resection and brachytherapy with 38 cases, the local recurrence rates was 5.26% (2/38). Four cases had wound infection and six cases had wound delay healing. The MSTS functional score was 21.11 ± 1.79. In the group of simple en bloc resection with 48 cases, the local recurrence rates was 20.83% (10/48). One case had wound infection and four cases had wound delay healing. The MSTS functional score was 84.23% (26.11 ± 1.79). The local recurrence rates was significant different between.

CONCLUSIONWith the extremity soft tissue sarcoma close to neurovascular bundle, combined en bloc resection and brachytherapy could decrease the local recurrence rate.

Adolescent ; Adult ; Aged ; Brachytherapy ; Female ; Follow-Up Studies ; Humans ; Male ; Middle Aged ; Neoplasm Recurrence, Local ; Retrospective Studies ; Sarcoma ; radiotherapy ; surgery ; Soft Tissue Neoplasms ; radiotherapy ; surgery ; Young Adult