OBJECTIVETo construct a eukaryotic vector which could express bone morphogenetic protein-7 (bmp-7) in MC3T3-E1.

METHODSBone morphogenetic protein-7 gene was obtained by RT-PCR from human embryo kidney. And after sequencing and electrophoresis the obtained aim DNA fragment was inserted into eukaryotic expression plasmid pcDNA3.1 (+) by using restricted endonuclease and ligase. The DNA sequence of the newly-constructed plamids was proved right by the gene technic company. And then the new plasmids containing right sequence aim gene were transfected into MC3T3-E1 cells by Lipofectamine 2000. 72 h after transfecting, RT-PCR was performed to show the transfected cells containing the aim gene, and the whole protein of the transfected cells were gathered and used as samples in the next Western blot to test the expression of bmp-7 gene.

RESULTSDNA sequencing indicated the sequence of the obtained bmp-7 was identical to the reported ones in GeneBank. The electrophoretic map of the products of RT-PCR and restriction enzyme digestion played another evidence that the newly-constructed plasmids were bmp-7/pcDNA3.1(+). The results of Western blot showed that the transfected cells could express BMP-7.

CONCLUSIONThe construction of a eukaryotic vector which could express BMP-7 in MC3T3-E1 was successful.

Bone and Bones ; Genetic Vectors ; Humans ; Plasmids ; Transfection

BACKGROUNDTumor necrosis factor a receptor 1 (TNFalphaR1) plays an important role in the signal pathway of apoptosis. The objective of this study was to investigate the effects of TNFalphaR1 knockout on the up-regulation of erythropoietin receptor (Epo-R) and the coordinated anti-apoptosis functions during myocardial ischemia-reperfusion injury in mice.

METHODSThe ischemia-reperfusion injury model for cardiomyocytes was performed by ligating the left circumflex branch artery of TNFalphaR1 knockout (P55(-/-)) C17 B6 mice, as well as wild-type (P55(+/+)) C17 B6 mice. Triphenyltetrazolium chloride (TTC) staining was performed to observe the damaged area of the heart. TUNEL staining and DNA fragmentation were used to identify apoptosis. Mitochondrial Bcl-2 and Bax as well as expression of Epo-R and its downstream genes (Jak-2, stat-5, Akt, IkB-alpha, HIF-1alpha) were measured by Western blotting. The gene knockout mice were assigned into those undergoing the apoptosis surgical model group (KO group), and those subjected to sham operation (KOs group). Similarly, wild-type mice were either exposed to the surgical model (WT group) or subject to a sham operation (WTs group).

RESULTSThe myocardial damage ratio of the wild-type group after the operation was significantly higher than that of the knockout group, (50.5 +/- 6.4)% vs (36.9 +/- 6.9)%, P < 0.01. Similarly, TUNEL positive ratio of the wild-type group was significantly higher than that of the knockout group, (63.1 +/- 5.6)% vs (42.1 +/- 4.7)%, P < 0.01. The gray value ratios of Epo-R, Jak-2, stat-5, Akt, IkB-alpha, HIF-1 and mitochondrial Bcl-2 in the KO group were significantly higher than those of the WT group, P < 0.05; however, mitochondrial Bax was significantly lower than that of the WT group significantly (P < 0.05).

CONCLUSIONSUsing the ischemia-reperfusion injury model in mice, cardiomyocytes of TNFalphaR1 knockouts exhibited anti-apoptotic characteristics. This information could be used to coordinate the prevention of myocardial apoptosis by up-regulating and activating the Epo-R pathway.

Animals ; Apoptosis ; Blotting, Western ; Disease Models, Animal ; I-kappa B Proteins ; metabolism ; In Situ Nick-End Labeling ; In Vitro Techniques ; Janus Kinase 2 ; metabolism ; Male ; Mice ; Mice, Knockout ; Myocardial Reperfusion Injury ; genetics ; metabolism ; pathology ; Myocytes, Cardiac ; metabolism ; pathology ; NF-KappaB Inhibitor alpha ; Oncogene Protein v-akt ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Receptors, Erythropoietin ; metabolism ; Receptors, Tumor Necrosis Factor ; genetics ; metabolism ; STAT5 Transcription Factor ; metabolism ; Up-Regulation ; bcl-2-Associated X Protein ; metabolism

The regulation mechanism of arecoline on rat hepatic CYP2E1 was studied in vivo. After oral administration of arecoline hydrobromide (AH; 4, 20 and 100 mg x kg(-1) x d(-1)) to rats for one week, the hepatic CYP2E1 mRNA level remained unchanged, but the hepatic CYP2E1 protein content was dose-dependently increased. Additionally, although the hepatic CYP2E1 activity was induced by AH treatment, the induction was attenuated with the increase in dosage. The results indicate that the effect of arecoline on rat hepaticdoes not involve transcriptional activation of the gene, but largely involves the stabilization of CYP2E1 protein against degradation or increased efficiency of CYP2E1 mRNA translation, and additionally involve the post- ranslational modification of CYP2E1 protein. Furthermore, the CYP2E1 response is fairly equal among the different species, the induction of rat hepatic CYP2E1 by arecoline suggests that there is a risk of metabolic interaction among the substrate drugs of CYP2E1 in betel-quid use human.
Animals ; Arecoline ; pharmacology ; Cytochrome P-450 CYP2E1 ; metabolism ; Cytochrome P-450 CYP2E1 Inducers ; pharmacology ; Humans ; Liver ; drug effects ; metabolism ; RNA, Messenger ; Rats

The viability and life span of pollen were evaluated by TTC (2,3,5-triphenyl tetrazlium chloride) and the peroxidase solution, the stigma receptivity were estimated by benzidine-H2O2 method and the fruiting characteristics were investigated. The results showed that (1) Anoectochilus roxburghii and A. formosanus appeared the same up-and-down trend of the pollen viability, increased and then decreased. The storage temperature and storage time had significant impact on the pollen viability. With the extension of storage time, the pollen activity decreased. 4 degrees C refrigerator storage may be extended the pollen vitality. (2) The stigma had receptivity in 1st day and reached the highest level in the 4th day after blooming. A. roxburghii lost receptivity in the 8th day while A. formosanus lost receptivity in the 10th day after blooming. (3) The different pollination had significant impact on seed setting rate. The seed setting rate of artificial cross-pollination was higher than that of the artificial self-pollination. Collecting pollen in the 3rd day and carrying out artificial cross-pollination in the 4th day after blooming can significantly improve seed setting rate. The results provided technical assurance for A. roxburghii and A. formosanus breeding of new varieties and seed breeding.
Cell Survival ; China ; Flowers ; physiology ; Fruit ; growth & development ; Orchidaceae ; growth & development ; physiology ; Pollen ; growth & development ; Reproduction ; Temperature

In this study, the processes of pollination ecology of Fritillaria delavayi were investigated to document its reproductive characteristics. Some individuals of F. delavayi could produce seeds under bagging without emasculation (11%), but the rate was significantly lower than that of the natural control (87%). It is suggesting that pollination of F. delavayi largely depends on pollen vectors. Bombus sushikini was the only effective pollinator of F. delavayi and the visitation frequency was 0.003 time xXflower(-1) x min(-1). Flowering of F. delavayi in whole population lasted for 35 d and single flower for 11 d. Pollen viability and stigma receptivity lasted for 9 d and were relatively long compared with other Fritillaria genus plants. Consequently, bumblebee pollination and long floral longevity seem to be important for reproductive assurance of F. delavayi in harsh alpine environments.
Animals ; Bees ; physiology ; Fritillaria ; physiology ; Pollen ; physiology ; Pollination

We report one case of pediatric acute myeloid leukemia type 2 (AML-M2) who presented with karyotypic aberration of trisomy 21 with the t(5;ll) chromosomal translocation.The patient achieved complete remission after two cycles of chemotherapy of daunorubicin,cytarabine and etoposide.Then,follow-up cytogenetic analysis from bone marrow cell cultures demonstrated a normal karyotype of 46,XY.After 9 years,the patient relapsed and the karyotypic abnormalities of trisomy 21 with t(5;ll) reappeared.It was concluded that trisomy 21 with t(5;11) is a new unfavorable cytogenetic aberration in AML-M2.

Objective To investigate the clinical significance in MRP1/CD_9 expression in cervical squamous cancer tissues and normal cervical tissues.Methods The expression of MRP1/CD_9 were assayed by SABC immunohistochemical methods in 53 cases of cervical cancer tissues and 13 cases of normal cervical tissues.Results Positive expression of MRP1/CD_9 was detected in 13 normal cervical tissue.MRP1/D_9 ex- pression is down-regulated in cervical carcinoma(P

OBJECTIVETo investigate the relationship between peak concentration (Cmax) of gemcitabine at fixed-dose-rate and its hematological toxicity profile in patients with advanced non-small-cell lung cancer (NSCLC).

METHODSTwenty-one patients received gemcitabine at a fixed dose rate (1200 mg/m2 over 120 min) with carboplatin. Plasma concentrations of gemcitabine were measured by ion-pair reversed-phase high-performance liquid chromatography.

RESULTSThe mean value of Cmax in 21 eligible patients was(4.95+/-2.42) microg *ml(-1). The main hematological toxicity was grade III-IV thrombocytopenia and neutropenia. The mean percentages of reduction of WBC, NEC, PLTC and Hb of 21 patients were (38.3+/-38.1)%, (31.3+/-73.6)%, (31.8+/-53.5)% and (12.0+/-12.2)%, respectively. The C(max)of gemcitabine and the percentage of reduction in WBC showed a significant correlation (r2=0.4575, P<0.05). A significant correlation (r2=0.5671, P<0.05) was also observed between the percentage of reduction of PLTC and Cmaxof gemcitabine.

CONCLUSIONThe results of relationship between Cmax and toxicity profile suggest that gemcitabine administration should be individualized in order to decrease the occurrence of ADR.

Adult ; Aged ; Antimetabolites, Antineoplastic ; administration & dosage ; adverse effects ; pharmacokinetics ; Antineoplastic Combined Chemotherapy Protocols ; adverse effects ; pharmacokinetics ; therapeutic use ; Carboplatin ; adverse effects ; blood ; pharmacokinetics ; Carcinoma, Non-Small-Cell Lung ; drug therapy ; metabolism ; Chromatography, High Pressure Liquid ; Deoxycytidine ; adverse effects ; analogs & derivatives ; blood ; pharmacokinetics ; Female ; Humans ; Infusions, Intravenous ; Lung Neoplasms ; drug therapy ; metabolism ; Male ; Metabolic Clearance Rate ; Middle Aged ; Neutropenia ; chemically induced ; Thrombocytopenia ; chemically induced

To determine the pharmacokinetics of gemcitabine (2',2'-difluorodeoxycytidine) in Chinese non-small-cell lung cancer (NSCLC) patients. Six study subjects were administered gemcitabine at a fixed dose rate of 10 mg/m2 per min (1200 mg/m2,two hours infusion) and carboplatin, and plasma gemcitabine concentrations were measured by ion-pair reversed-phase high-performance liquid chromatography (HPLC). 3P97 Pharmaceutical Kinetics Software was used for the calculation of pharmacokinetic parameters. The obtained mean parameters, elimnation half life (t1/2) (10.67±3.38 min), area under the curve hematologic toxicology result showed that the regimen was effective on and tolerated by the patients.