Objective:To investigate the expression of HIF-1?and its relationship with angiogenesis in osteosarcoma.Methods: Osteosarcoma MG-63 cells were cultured in vitro under hypoxia and mimic hypoxia conditions.Thirty paraffin-embedded osteosarcoma tissues and 20 fresh frozen osteosarcoma specimens were collected.The mRNA and protein expression of HIF-1?and VEGF were detected by RT-PCR,Western blotting,ELISA,and immunohistochemistry methods.The mean vessel density(MVD)were also calculated.Results:The mRNA level of HIF-1?had no change under hypoxia and minic hypoxia conditions,whereas the protein expression was increased dramaticaly.The mRNA and protein expression of VEGF was significantly increased under hypoxia and minic hypoxia conditions.The positive rate of HIF-1?mRNA(90%)and VEGF(100%)in 20 fresh frozen tissues were higher than those of the para-tumor tissues(P

OBJECTIVETo further explore the effect of Jiangu Erxian Pill (JGEXP) on proliferation and cell cycle of human osteoblast on the basis of previous clinical and experimental studies.

METHODSHuman primary osteoblast were isolated and cultured. The cell proliferation was tested by 3H-thymine incorporation and methyl thiazolyl tetrazolium (MMT) method and the cell cycle was determined by flow cytometry technique.

RESULTSIn the medium and high dosage JGEXP groups, the cell proliferation rate and index, and percentage of diploid synthesis phase (S phase) cells were significantly higher than those in the blank control group (P < 0.01 or P < 0.05), and similar to those in the estrogen group; and the cell apoptosis rate and percentage of G0-G1 stage cells were lower than those in the blank control group (P < 0.01 and P < 0.05).

CONCLUSIONJGEXP could effectively promote the cell proliferation and differentiation, and prevent the cell apoptosis of osteoblast in vitro.

Animals ; Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Division ; drug effects ; Cells, Cultured ; Drugs, Chinese Herbal ; pharmacology ; Female ; Humans ; Osteoblasts ; cytology ; Random Allocation ; Rats ; Rats, Sprague-Dawley

Adenocarcinoma ; drug therapy ; Antibodies, Monoclonal ; adverse effects ; Drug Resistance, Neoplasm ; Female ; Fever ; chemically induced ; Humans ; Liver Neoplasms ; secondary ; Lung Neoplasms ; drug therapy ; Middle Aged ; Quinazolines ; therapeutic use

OBJECTIVE: To investigate the biological characteristics of endothelial progenitor cells (EPCs) from the umbilical cord blood (UCB), and to evaluate their oncogenicity after long-term culture in vitro. METHODS: The mononuclear cells (MNCs) were isolated from the UCB and cultured in MCDB131 medium supplemented with 20% FBS, VEGF and other growth factors. Morphology of the EPCs was observed, and the growth curve of the EPCs was investigated. Surface antigens of the EPCs were analyzed by the flow-cytometer. The capability of intaking the acetylated low-density lipoprotein (acLDL) of the EPCs was detected using fluoresencent chemical method. The vasoformative capability and genetic stability of EPCs were cultured in matrigel, and examined by karyotype analysis. The oncogenicity of EPCs was verified by the tumorigenesis test in athymic mouse and soft agar. RESULTS: EPCs were successfully derived from the UCB, and could be passaged to at least 42(nd) generation and had strong abilities of proliferation, acLDL intake and vasoformation, but there was not oncogenicity. They expressed endothelial cell-surface antigens and maintained normal karyotype. CONCLUSION The EPCs with proliferative potential can be isolated from the UCB. They can be passaged in long-term cultures without oncogenicity, and can maintain normal karyotype. The EPCs can be served as a new type of cells in cell and gene therapy.
Animals ; Antigens, Surface ; analysis ; Cell Line ; Cell Proliferation ; drug effects ; Cells, Cultured ; Endothelial Cells ; cytology ; metabolism ; Fetal Blood ; cytology ; Flow Cytometry ; HeLa Cells ; Humans ; Infant, Newborn ; Intercellular Signaling Peptides and Proteins ; pharmacology ; Karyotyping ; Mice ; Mice, Nude ; Neoplasms, Experimental ; pathology ; Stem Cells ; cytology ; metabolism ; Vascular Endothelial Growth Factor A ; pharmacology

OBJECTIVETo observe metabolomic changes in urine of chronic superficial gastritis (CSG) patients with Pi-qi deficiency syndrome (PQDS) or Pi-Wei dampness-heat syndrome (PWDHS), thereby providing scientific evidence for syndrome typing of them.

METHODSUrine samples were collected from CSG patients with PQDS/PWDHS and healthy volunteers, 10 in each group. Proton nuclear magnetic resonance spectroscopy (1H-NMR) based metabonomic analysis was performed on urine samples. Contents of related biomarkers were analyzed by principal component analysis (PCA), partial least square discriminant analysis (PLS-DA), and urivariate statistical analysis.

RESULTSPLS-DA analysis showed that metabolites among CSG patients with PQDS/PWDHS and healthy volunteers could be mutually distinguished. Seven differentially identified metabolites were screened from urines of CSG patients with PQDS and healthy volunteers included glutamate, methionine, α-oxoglutarate, dimethylglycine, creatinine, taurine, and glucose. Four differentially identified metabolites were screened from urines of CSG patients with PWDHS and healthy volunteers included 2-hydroxybutyric acid, trimethylamine oxide, taurine, and hippuric acid. Eleven differentially identified metabolites were screened from urines of CSG patients with PQDS and PWDHS included fucose, β-hydroxybutyric acid, alanine, glutamate, methionine, succinic acid, citric acid, creatinine, glucose, hippuric acid, and lactic acid.

CONCLUSIONThe metabolic differences of CSG patients PQDS and PWDHS mainly manifested in glycometabolism, lipid metabolism, and amino acids catabolism, and 1H-NMR based metabonomics may be used in classified study of Chinese medical syndrome typing.

Biomarkers ; urine ; Discriminant Analysis ; Gastritis ; urine ; Hot Temperature ; Humans ; Hydroxybutyrates ; Ketoglutaric Acids ; Least-Squares Analysis ; Medicine, Chinese Traditional ; Metabolome ; physiology ; Metabolomics ; Principal Component Analysis ; Proton Magnetic Resonance Spectroscopy ; Qi ; Syndrome

AIM:To investigate the status of RAS/BRAF mutations and microsatellite instability ( MSI ) and their associations with clinicopathological features and prognosis of the patients with stage Ⅲ colorectal cancer ( CRC ) . METHODS:The surgical patients with stage ⅢCRC (n=281) were followed up.The mutations of RAS/BRAF were ex-amined by PCR amplification-Sanger sequencing , and MSI status was detected using immunohistochemistry ( IHC) in their archival paraffin-embedded tissue specimens .The relationships of the status with the clinicopathological features and prog-nosis of the patients were statistically analyzed .RESULTS: Among 281 patients, the mutations of RAS/BRAF were ob-served in 136 cases (48.4%), including 116 cases (41.3%) of KRAS mutations.RAS/BRAF mutations were highly cor-related with the level of carcino-embryonic antigen (P<0.05).Moreover, 18 cases (6.4%) of MSI-high (MSI-H) pa-tients were determined by IHC, and MSI-H status was more common in N2b patients (P<0.05).Correlation study found that the mutation rate of BRAF was higher in MSI-H tumors than that in MSI-low ( MSI-L)/microsatellite stability ( MSS) counterparts (P<0.01), although no association between KRAS/NRAS mutations and the MSI status was observed .The prognosis in the patients with wild-type RAS/BRAF or MSI-H was better than the patients with any mutation or MSI-L/MSS (P<0.01).CONCLUSION:Mutant RAS/BRAF and MSI may serve as fairly good indicators for prognosis of the patients with stage Ⅲ CRC.

This study was aimed to clone the gene coding mouse fibroblast growth factor receptor-1 (fgfr1), to construct the recombinant lentiviral vector of truncated form fgfr-1 (Δfgfr1) carrying enhanced green fluorescence protein (EGFP) and to investigate its expression in eukaryotic cells (293FT cells). The full length fgfr1 gene was cloned by RT-PCR using brain tissue of BALB/c fetal mouse as template and inserted into PCR-Blunt vector, a truncated fgfr1 fragment was produced by site-directed mutagenesis for deleting intracellular phosphorylated domain, then was subcloned into a lentiviral vector and cotransfected into 293FT packaging cells together with envelope plasmid and packaging plasmid by lipofectamine 2000. Viruses were gathered and concentrated using ultracentrifuge, and then transfected into 293FT cells. Expression of EGFP was detected by fluorescent microscopy and flow cytometry (FCM), and the truncated FGFR1 protein was detected by Western blot. The results demonstrated that mouse fgfr1 gene was cloned and the lentiviral expression vector LV-IRES-EGFP-Δfgfr1 and control vector LV-IRES-EGFP were successfully constructed. The lentiviral particles were correctly packaged, and the virus titers were above 10(8) TU/ml in the supernatant after concentration. Expression of EGFP was detected by fluorescent microscopy in 293FT cells post transfection, and the transfection efficacy was > 95% determined by FCM. Expression of FGFR1 protein detected by Western blot was significantly higher than that in control group. It is concluded that the truncated gene fgfr1 along with the gene coding EGFP is successfully inserted into a lentiviral vector to construct a recombinant lentiviral vector, which can be expressed in eukaryotic cells.
Animals ; Cell Line ; Genetic Vectors ; Humans ; Lentivirus ; genetics ; Mice ; Mice, Inbred BALB C ; Plasmids ; Receptor, Fibroblast Growth Factor, Type 1 ; genetics ; Recombinant Fusion Proteins ; genetics ; Transfection

OBJECTIVETo investigate the hearing change when chronic tinnitus acute aggravated and the affect of prognosis due to hearing change.

METHODSThe pure tone threshold and acoustic immittance were used for every case in the acute aggravation of chronic tinnitus (AACT) group, and for some of AACT group members the auditory brainstem response (ABR), distort-product otoacoustic emission (DPOAE), electrocochleogram (EcochG), glycerin test and MRI were used at the same time for further diagnosis. For the chronic tinnitus, the patients accepted the intervention of tinnitus retraining therapy (TRT) both for the AACT group and the control group. As for the recent hearing loss appeared in the AACT group members, the patients were treated the cases as the sudden deafness, and then the hearing characteristic of the AACT group was analyzed and the effect of the chronic tinnitus both for the AACT group and control group after the TRT for 3, 6, 9 and 12 months was evaluated.

RESULTSThree kinds of hearing changes appeared when chronic tinnitus acute aggravated. Based on the dated high frequencies hearing loss ranged 4-8 kHz, recent hearing loss appeared in 1-2 middle frequencies near the impaired high frequencies; Based on the dated single middle frequency hearing loss, another middle frequency hearing impaired near the dated hearing loss frequency, the sawtooth-like audiogram changed to groove-like audiogram; Hearing fluctuation in low frequency area, hearing threshold increased 10-30 dB in 125-1000 Hz but kept unchanged in the high frequency area. The recent local frequency hearing loss healed after the vasodilator and neurotrophic drugs therapy, but the dated hearing loss can't rehabilitate. Along with the rehabilitation of recent local hearing loss, 25 of 32 cases chronic tinnitus victims get to habituation; this finding showed that the experience of tinnitus aggravation and relief accelerated the habituation. Until the 12 months after the TRT, the curative effect difference between AACT group and control group showed statistical significance.

CONCLUSIONSWhen the chronic tinnitus acute aggravated, 3 kinds of local area hearing impaired. Timely diagnosis and effective treatment had brought not only hearing rehabilitation but also promotion to habituation of chronic tinnitus.

Adult ; Audiometry, Pure-Tone ; Auditory Threshold ; Female ; Humans ; Male ; Middle Aged ; Prognosis ; Tinnitus ; diagnosis ; physiopathology ; therapy ; Treatment Outcome

OBJECTIVETo determine whether the single nucleotide polymorphism (SNP) on chromosome 12q24.31(rs2259816) is associated with coronary artery disease (CAD) in Han population of southwest China.

METHODSA case-control association study with 592 unrelated patients with coronary artery disease and 463 normal controls from Chinese Han population was performed. Genotype for the SNP on chromosome 12q24.31 (rs2259816) was determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP).

RESULTSThe genotypes of AA, AC, CC were both detected in the coronary artery disease group and the control group. The frequencies of A allele were 49.5% in case group and 43.8% in control group, showing statistically significant difference(OR=1.129, 95%CI:1.029-1.239, P=0.010).

CONCLUSIONThe replication study showed that the genetic polymorphism in rs2259816 is associated with coronary artery disease in Han population of southwest China.

Base Sequence ; Case-Control Studies ; Chromosomes, Human, Pair 12 ; genetics ; Coronary Artery Disease ; blood ; genetics ; Female ; Gene Frequency ; Genetic Predisposition to Disease ; genetics ; Humans ; Lipids ; blood ; Male ; Polymorphism, Single Nucleotide ; genetics

OBJECTIVETo prepare lidocaine nanoemulsion and investigate its transdermal delivery ability in vitro.

METHODSThe optimal Km (surfactant/cosurfactant) value and the component proportion were determined by pseudoternary phase diagrams combined with Origin software analysis. The diameter and distribution range were detected by Zeta particle size analysis instrument, and the morphology of the nanoemulsion was observed by electron microscope. The permeation flux of lidocaine was determined in vitro using the modified Franz diffusion cell combined with HPLC, and the cumulative transdermal absorption amount and the apparent skin transdermal velocity were compared among nanoemulsion, gel and tincture containing 5% lidocaine. The permeation mode of lidocaine nanoemulsion was analyzed.

RESULTSThe average drop size of lidocaine nanoemulsion was 29.8-/+14.4 nm, and 98% of the drop sizes ranged from 15.1 to 45.5 nm and 2% from 77.9 to 261.3 nm. The nanoemulsion drop showed a spherical morphology in a polydisperse system. The Kp value of the nanoemulsion (3.07-/+0.74 cm/h) was significantly higher than that of gel (1.27-/+0.35 cm/h) and tincture (0.97-/+0.18 cm/h), and the permeation rate of the nanoemulsion was 69.82-/+7.48 microg x cm(-2) x h(-1), which fitted the the Zero-order release dynamic procedure.

CONCLUSIONSThe component proportion of lidocaine nanoemulsion can be conveniently obtained through pseudoternary phase diagrams and Origin software analysis, and the drop size, distribution, morphology and system type can be determined by Malvern Zetasizer combined with electron microscopy. The results also indicate that the nanoemulsion system with high permeation rate may provide a new promising means for local anesthesia.

Administration, Cutaneous ; Anesthetics, Local ; administration & dosage ; metabolism ; Animals ; Emulsions ; Lidocaine ; administration & dosage ; metabolism ; Male ; Nanoparticles ; Particle Size ; Permeability ; Rats ; Rats, Wistar ; Skin Absorption ; drug effects