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

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

The hematopoietic cell phosphatase (HCP or SHP-1), the SH2 domain contain protein tyrosine phosphatase, is a crucial negative regulator in the process of hematopoietic cell development, proliferation and receptor-mediated mitogenic signaling pathways, and its mutation is responsible for the over-expansion and inappropriate activation of myelomonocytic population in motheaten mice. The aim of the study was to evaluate the role of the HCP gene in leukemogenesis. Bone marrow and/or peripheral blood from 32 acute myeloid leukemia (AML) patients, 9 acute lymphocytic leukemia (ALL) patients, 8 leukemia cell lines and 50 normal controls were analyzed by reverse transcription-polymerase chain reaction (RT-PCR) based on single strand conformation polymorphism (SSCP) and sequencing. RT-PCR showed that all samples expressed HCP gene, only one missense mutation at codon 225 (AAC to AGC, Asn to Ser) within N-terminal SH2 domain was found in an ALL patient. In addition, four polymorphic base substitutions were detected in codon 69, 85, 86 and 266, respectively. In conclusion, mutation of HCP gene is an infrequent genetic aberration which may only play a role in pathogenesis of a small part of leukemia, however, its significance needs to be further clarified.
Acute Disease ; Cell Line, Tumor ; Humans ; Intracellular Signaling Peptides and Proteins ; Leukemia ; enzymology ; genetics ; Mutation ; Polymorphism, Single-Stranded Conformational ; Protein Tyrosine Phosphatase, Non-Receptor Type 6 ; Protein Tyrosine Phosphatases ; genetics

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

To analyze the genetic characteristics of echovirus 6 (E6) isolated from meningitis and encephalitis cases in Shandong Province, China, we collected cerebrospinal fluid samples from meningitis and encephalitis cases in Shandong Province from 2007 to 2012 for virus isolation. Viral RNAs were extracted from positive isolates, and complete VP1 coding regions were amplified by RT-PCR and sequenced. Homology comparison and phylogenetic analysis were performed. Six isolates were identified as E6 by microneutralization assay and molecular typing. The homology analysis showed that the six isolates had 78. 6%-99. 8% nucleotide and 95. 5%-100. 0% amino acid identities with each other, as well as 76. 9%-78. 4% nucleotide and 92. 3%-95. 1% amino acid identities with the prototype strain (D' Amori). The phylogenetic analysis based on the integrated VP1 sequences indicated that all Shandong E6 isolates could be separated into four clusters, designated as A, B, C, and D. The six E6 isolates belonged to clusters A, B, and D. Our study reveals high genetic differences between Shandong E6 isolates and suggests different transmission lineages of E6 co-circulated in Shandong Province.
Amino Acid Sequence ; Child ; Child, Preschool ; China ; epidemiology ; Echovirus 6, Human ; classification ; genetics ; isolation & purification ; Encephalitis ; epidemiology ; virology ; Female ; Genetic Variation ; Humans ; Infant ; Male ; Meningitis ; epidemiology ; virology ; Molecular Sequence Data ; Phylogeny ; Sequence Alignment ; Viral Proteins ; chemistry ; genetics

The SH2 domain containing inositol 5'-phosphatase (SHIP) was initially described as a 145 kD protein phosphorylated on tyrosines upon growth factor and cytokine stimulation. SHIP is predominately expressed in hematopoietic cells, and is a crucial negative regulator in the development of hematopoietic cells. To evaluate the role of the SHIP gene in human leukemogenesis, expression and mutation of SHIP gene in bone marrow and/or peripheral blood from 32 patients with acute myeloid leukemia (AML), 9 patients with acute lymphoblastic leukemia (ALL), as well as human hematopoietic cell lines were analyzed by reverse transcription-polymerase chain reaction (RT-PCR), single strand conformational polymorphism (SSCP) and sequencing. The RT-PCR showed that all samples expressed SHIP gene. Mutations of SHIP gene were detected in 7 out of 32 AML patients (22%) and one out of 9 ALL patients (12%). Interestingly, two missense mutations that had been observed in one AML patient at diagnosis disappeared after complete remission (CR). In addition, Akt phosphorylation was prolonged and increased following IL-3 stimulation in this patient sample. In conclusion, data of this study demonstrate the mutation of the SHIP gene in acute leukemia for the first time and suggest a possible role of the mutation of this gene in the development of acute leukemia. SHIP serves as a tumor suppressor by negatively regulating the PI3K/Akt signaling pathway in hematopoietic cells.
Cell Line ; Humans ; Leukemia, Myeloid, Acute ; genetics ; Mutation ; PTEN Phosphohydrolase ; Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases ; Phosphoric Monoester Hydrolases ; genetics ; physiology ; Phosphorylation ; Polymorphism, Genetic ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; genetics ; Protein-Serine-Threonine Kinases ; metabolism ; Proto-Oncogene Proteins ; metabolism ; Proto-Oncogene Proteins c-akt ; Tumor Suppressor Proteins ; physiology

OBJECTIVETo evaluate the role of vascular resection and reconstruction in the treatment of hilar cholangiocarcinoma.

METHODS117 patients with potentially resectable hilar cholangiocarcinoma underwent exploration. Twenty-one patients had exploration or drainage only due to distant metastases, and the other 96 patients received surgical resection. Thirty-one of those had vascular resection and reconstruction, including portal vein resection alone in 21 patients, combined hepatic artery and portal vein resection in 2 and hepatic artery resection alone in 8. Therefore, the patients were divided into four groups: non-surgical resection (21), portal vain resection (21), hepatic artery resection (10) and non-vascular resection (65) and their clinical data were reviewed retrospectively.

RESULTSThe hepatic artery resection group had significantly higher perioperative morbidity and mortality rate (80.0% and 20.0%) than non-vascular resection group (16.9% and 1.5%), respectively, (P < 0.05), while no significant difference was found between the portal vein resection alone group and the non-vascular resection group (P > 0.05). Of all resected vessel specimens, vascular wall invasion beyond the adventitia was pathologically confirmed in 82.6% of the portal veins and 50.0% of the hepatic arteries. The 1-, 3- and 5-year survival rates were 59.0%, 34.0%, and 16.0% in the non-vascular resection group, versus 44.0%, 23.0% and 11.0% in the portal vein resection alone group (P < 0.05) and 18.0%, 0 and 0 in the hepatic artery resection group (P < 0.01), respectively, with a significant difference among the three groups. The 1-, 3- and 5-year survival rates in the non-surgical resection group were 13.0%, 0 and 0, respectively, which were similar to those in the hepatic artery resection group. Though a significant difference in survival rates existed between the portal vein resection alone group and non-resected group (P < 0.001), no significant difference was found between the hepatic artery resection group and non-resected group (P > 0.05).

CONCLUSIONBoth portal vein and hepatic artery resection can improve resection rate for hilar cholangiocarcinoma, and portal vein resection may improve the prognosis in selected patients. However, hepatic artery resection can not improve survival and may even lead to an increase of perioperative morbidity and mortality.

Adult ; Aged ; Bile Duct Neoplasms ; mortality ; surgery ; Bile Ducts, Intrahepatic ; Cholangiocarcinoma ; mortality ; surgery ; Female ; Follow-Up Studies ; Hepatic Artery ; pathology ; surgery ; Humans ; Male ; Middle Aged ; Neoplasm Invasiveness ; Portal Vein ; pathology ; surgery ; Reconstructive Surgical Procedures ; mortality ; Retrospective Studies ; Survival Rate ; Vascular Surgical Procedures ; mortality