BACKGROUND:Angiotensin II receptor antagonists have been found to exerct a stronger protective effect on bone than angiotensin converting enzyme inhibitors. OBJECTIVE:To investigate the effect of different concentrations of irbesartan (angiotensin II receptor antagonist) on the differentiation and mineralization of mouse preosteoblasts. METHODS:Mouse preosteoblast cel lines MC3T3-E1 in logarithmic phase were selected and cultured in the osteogenic induction medium containing 0 (control group), 0.001, 0.01, 0.1 mmol/L irbesartan, respectively. Ten days later, the cel differentiation was observed by alkaline phosphatase staining. The mineralization was observed by alizarin red staining after 21 days of culture. mRNA expressions of osteocalcin, alkaline phosphatase and Runt-associated transcription factor 2 in osteoblasts were detected by real-time PCR at 1, 4, 7, 14 and 21 days of culture. RESULTS AND CONCLUSION:The activity of alkaline phosphatase in al the irbesartan groups (0, 0.001, 0.01, 0.1) was higher than that in the control group (P<0.05), which was the most obvious in 0.01 mmol/L. The number and area of calcium nodules in each irbesartan group were significantly higher than those in the control group (P<0.05), especial y in 0.01 mmol/L. Compared with the control group, 0.01 mmol/L irbesartan significantly upregulated the mRNA expressions of osteocalcin, alkaline phosphatase and Runt-associated transcription factor 2 (P<0.05). These results suggest that 0.01 mmol/L irbesartan significantly promotes the differentiation and mineralization of osteoblasts.

Antibodies, Monoclonal ; administration & dosage ; Antibodies, Monoclonal, Humanized ; Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Breast Neoplasms ; diagnosis ; drug therapy ; metabolism ; pathology ; surgery ; Chemotherapy, Adjuvant ; Cyclophosphamide ; administration & dosage ; Doxorubicin ; administration & dosage ; Female ; Humans ; Neoplasm Recurrence, Local ; Paclitaxel ; administration & dosage ; Receptor, ErbB-2 ; metabolism ; Trastuzumab

0.5 cm or beneficial anatomical vari- ations displayed on MRCP,were obviously improved and there were no significantly different among the 4 types hilar eholangiocarcinoma.Conclusion MRCP could accurately make the preoperative diagnosis and type of hilar cholangiocarcinoma; the image of second branch of bile duct and the variation of the confluence of hepatic hilar displayed on MRCP has great clinical significance for operative regimes of hilar cholangiocar- cinoma,especially for typeⅣ.It does benefit not only to improve the resection and radical rate of some hilar cholangiocarcinomas, but also to select suitable method of biliary enteric anastomosis and avoid injuring the bile duct in operation.

In this study, a rapid and sensitive analytical method was developed for the determination of 10 major compounds (procyanidin B1, catechin, procyanidin B2, rutin, isoquercitrin, kaempferol-3-O-rutinoside, astragalin, quercitrin, quercetin, and kaempferol) in Tetrastigma hemsleyanum by using ultra-performance liquid chromatography coupled with triple-quadrupole tandem mass spectrometry (UPLC-MS/MS) in multiple-reaction monitoring (MRM) mode. UPLC-MS/MS assay with negative ion mode was performed on a Waters CORTECS C18 (2.1 mm x 100 mm, 1.6 μm) with the mobile phase consisting of acetonitrile (A) and 0.1% aqueous formic acid (B) in gradient elution at a flow rate of 0.25 mL · min(-1) and the column temperature was set at 45 °C. Under the optimized chromatographic conditions, good separation for 10 target compounds were obtained including chiral isomer procyanidins B1 and B2 were completely separated within 8.5 min. Satisfactory linearity was achieved with wide linear range and fine determination coefficient (r > 0.996 6), the overall recoveries were ranged from 95.44%-110.40% with the RSD ranging from 2.37%-8.69%. It is the first report about simultaneous analysis of 10 major flavonoids components in Tetrastigma hemsleyanum by using UPLC-MS/MS method, which affords highly sensitive, specific, speedy and efficient method for quality control of Tetrastigma hemsleyanum
Acetonitriles ; Chromatography, High Pressure Liquid ; Flavonoids ; chemistry ; Kaempferols ; Quercetin ; analogs & derivatives ; Rutin ; Tandem Mass Spectrometry ; Vitaceae ; chemistry

OBJECTIVEThe SH2 domain containing inositol 5'-phosphatase (SHIP) is predominately expressed in hematopoietic cells, and is a crucial negative regulator in the development of hematopoietic cells. This paper is to evaluate the role of the SHIP gene in human leukemogenesis.

METHODSExpression of SHIP gene in bone marrow and/or peripheral blood from 32 patients with acute myeloid leukemia (AML), 9 with acute lymphoblastic leukemia (ALL), as well as human hematopoietic cell lines was analyzed by reverse transcription-polymerase chain reaction (RT-PCR), single strand conformational polymorphism (SSCP) and DNA sequencing.

RESULTSRT-PCR showed that all samples expressed SHIP gene. Mutations of SHIP gene were detected in 7 (22%) of 32 AML patients and one (12%) of 9 ALL patients. Interestingly, two missense mutations that had been observed in a AML patient at diagnosis disappeared after complete remission (CR). In addition, in vitro Akt phosphorylation was prolonged and increased following IL-3 stimulation of this patient's cells.

CONCLUSIONOur data demonstrate for the first time the mutation of SHIP gene in acute leukemia and suggest a possible role of the mutation of this gene in the development of acute leukemia. SHIP may serve as a tumor suppressor by negatively regulating the PI3K/Akt signaling pathway in hematopoietic cells.

Blotting, Western ; Cell Line, Tumor ; DNA Mutational Analysis ; HL-60 Cells ; Humans ; Inositol Polyphosphate 5-Phosphatases ; Interleukin-3 ; pharmacology ; K562 Cells ; Leukemia, Myeloid, Acute ; genetics ; metabolism ; Mutation ; Oncogene Protein v-akt ; metabolism ; Phosphoric Monoester Hydrolases ; genetics ; metabolism ; Phosphorylation ; drug effects ; Polymorphism, Single-Stranded Conformational ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; U937 Cells

Wound combined with total body irradiation (TBI) injury results in impairment of tissue repair and delayed processes of healing, so it has been considered as an important and representative model of impaired wound healing, but the mechanism is not fully clarified. Fibroblasts in wound are the most important cells participating in tissue repair, whereas its radiosensitivity is not high. To understand whether TBI injury has direct damaging effects on fibroblasts in wound, fibroblasts in wound combined with TBI injury and in wound of simple incision injury were isolated and cultured, and parameters associated with tissue repair were determined. The results showed that the abilities of proliferation, attachment and adhesion of fibroblasts isolated from wounds combined with TBI injury significantly decreased as compared with those of simple incision injury, nevertheless, apoptotic ratio of fibroblasts isolated from wounds combined with TBI injury increased significantly. These data suggest that TBI injury may cause direct damaging effects on fibroblasts in wounds, which might be one of the dominant reasons for impairment of wound healing when it is combined with TBI injury.
Animals ; Disease Models, Animal ; Fibroblasts ; metabolism ; physiology ; radiation effects ; Radiation Injuries, Experimental ; metabolism ; Rats ; Rats, Wistar ; Skin ; injuries ; Whole-Body Irradiation ; Wound Healing ; physiology

A qualitative analytical method of liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry (HPLC-Q-TOF-MS) was developed for identification of multi-constituents and an analytical method was developed for simultaneously determining 4 major compounds (rutin, isoquercitrin, kaempferol-3-0-rutinoside, and astragalin) in Tetrastigma hemsleyanum Diels et Gilg. The HPLC-Q-TOF-MS assay was performed on a Welch Ultimate XB-C18 column (4.6 mm x 150 mm, 5 microm) with the mobile phase consisting of acetonitrile (A) and water containing 0.1% Formic acid (B) in gradient mode at a flow rate of 0.8 mL x min(-1). The column temperature was at 30 degrees C, and negative ion mode was used for TOF-MS. The UPLC-QqQ-MS assay was performed on a Waters CORTECS C18 (2.1 mm x 100 mm, 1.6 microm) with the mobile phase consisting of acetonitrile (A) and water containing 0.1% formic acid (B) in gradient mode at a flow rate of 0.25 mL x min(-1). The column temperature was at 45 degrees C, and MRM mode was used for QqQ-MS. Based on the retention time and MS spectra, 24 compounds were identified or tentatively characterized by comparing with reference substances or literatures. For quantitative the linear range of 4 detected compounds were good (r > 0.9966), and the overall recoveries ranged from 98.27% to 101.58%, with the RSD ranging from 3.15% to 5.88%. The results indicated that new approach conbined HPLC-Q-TOF-MS and UPLC-QqQ-MS was applicable in qualitative and quantitative quality control of Tetrastigma hemsleyanum.
Acetonitriles ; chemistry ; Chromatography, High Pressure Liquid ; methods ; Formates ; chemistry ; Tandem Mass Spectrometry ; methods ; Vitaceae ; chemistry ; Water ; chemistry

OBJECTIVETo improve computer-assisted imaging analysis for quantitatively measuring brain slice volume of rats and mice in comparison with conventional measuring methods,and to evaluate its usefulness in assessment of focal cerebral ischemia.

METHODSThe accurate volumes of rat and mouse brain slices were measured by weight and special gravity measuring. The areas of brain slices were measured by imaging analysis, then the slice volumes of right and left hemispheres were calculated by multiplying the adjusted thickness of the slices. In addition, the brain slice volumes of right and left hemispheres from focal cerebral ischemic mice were compared to assess ischemic injury using the imaging analysis.

RESULTArea measurement by computer-assisted imaging analysis was linear with different accurate areas (r=1.000). Slice volumes measured by imaging analysis correlated well with the accurate volumes measured by special gravity method, r=0.809 (n=45, P<0.001) in rats, and r=0.844 (n=74, P<0.001) in mice. The brain volumes in ischemic hemispheres were larger than in non-ischemic hemispheres in ischemic mice.

CONCLUSIONComputer-assisted imaging analysis can measure the brain slice volumes accurately and compare right and left hemisphere volumes quantitatively.

Animals ; Brain ; pathology ; Brain Ischemia ; pathology ; Image Processing, Computer-Assisted ; Male ; Mice ; Mice, Inbred ICR ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo explore PTEN gene expression and its clinical significance in acute leukemia.

METHODSThe expression levels of PTEN mRNA in 5 leukemia cell lines, 87 patients with acute leukemias (AL), including 59 acute myeloid leukemia (AML), 26 acute lymphoblastic leukemia (ALL), and 2 acute hybrid leukemia, 21 AL in complete remission (AL-CR), 31 chronic myelogenous leukemia (CML) and 14 normal controls were assayed by RT-PCR.

RESULTSPTEN mRNA was detected in K562 cell line, but not in Kasumi-1, HL-60, U937, Nalm-6 cell lines. The expression ratio of PTEN mRNA between CML (61.29%) and normal control (78.57%) had no statistical difference (P > 0.05). The expression ratios of PTEN mRNA in AL (18.39%) and AL-CR (42.86%) were significantly lower than that in normal control (P < 0.01 and P < 0.05, respectively), AL also has a lower expression ratio than that of AL-CR (P < 0.05). The decreased level of PTEN mRNA had a positive correlation with poor-prognostic factors (high white blood cell count of > or = 20 x 10(9)/L and chromosome abnormality).

CONCLUSIONThere is down-regulated expression of PTEN gene in AL. PTEN gene may play a role in leukemogenesis.

Cell Line, Tumor ; Humans ; Leukemia ; genetics ; metabolism ; PTEN Phosphohydrolase ; genetics ; metabolism ; RNA, Messenger ; genetics ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo generate a gene delivery plasmid carrying the dominant negative form of the protein phosphatase 2A catalytic subunit a (DN-PP2Aca) driven by a hepatocellular carcinoma (HCC) tissue-specific promoter and investigate its ability to inhibit growth of cultured hepatoma cells.

METHODSThe gene delivery plasmid was constructed by PCR-amplifying DN-PP2Aca from wild-type PP2Aca using site-directed mutagenesis and then ligating the sequence-verified amplicon downstream of an alpha-fetoprotein enhancer and phosphoglycerate kinase promoter (AFpg) in the luciferase reporter vector pGL3-Basic. Following transfection into two AFP+ hepatoma cell lines (HepG2 and HepG3) and two AFP- hepatoma cell lines (SK-HEP-1 and L02), the transcriptional activity of the AFpg-driven DN-PP2Aca plasmid was tested using luciferase reporter gene assay and western blotting. The effect on cell growth was tested using MTT assay. Between group differences were assessed by t-test.

RESULTSThe AFpg-driven DN-PP2Aca plasmid showed high transcriptional activity and protein expression in both HepG2 and Hep3B cells. At 72 h after transfection, the proliferation capacities were repressed by 42.65%+/-3.99% (P = 0.0002) and 39.87%+/-3.91% (P = 0.0002) in AFP+ HepG2 and Hep3B cells, respectively (vs. untransfected). In contrast, the plasmid was transcriptionally inactive in and had no effect on proliferation of AFP- cells.

CONCLUSIONThe AFpg-driven DN-PP2Aca plasmid exhibits selective cytotoxicity against AFP+ hepatoma cells, and may represent a useful gene therapy strategy to treat HCC.

Carcinoma, Hepatocellular ; genetics ; metabolism ; Enhancer Elements, Genetic ; Genetic Therapy ; Genetic Vectors ; Hep G2 Cells ; Humans ; Liver Neoplasms ; genetics ; metabolism ; Mutation ; Promoter Regions, Genetic ; Protein Phosphatase 2 ; genetics ; alpha-Fetoproteins ; genetics