Objective Analyze the clinical feature of patients failed for diagnosis through endobronchial ultrasound transbronchial needle aspiration(EBUS-TBNA).Optimize the indication and increase diagnosis rate of EBUS-TBNA.Methods A total of 669 patients failed for diagnosis of EBUS-TBNA were included.Fifty-three of them(7.92％) were not exactly diagnosed.Perioperation clinical data and clinical feature were collected and evaluated based on specific disease,lesion location,size and operator' s experience.Results The undiagnosis rate was higher in lymphoma (77.78％),tuberculosis (23.08％) and sarcoidosis(9.09％) when analyzed from specific diseases.If the lesion location was taken into consideration,15.38％ upper paratracheal lymph nodes(R2) could not be diagnosed exactly by EBUS-TBNA,and the bilateral hilar lymph nodes(15.00％ for right,11.54 for left) were followed.Size of the lesion was not associated with the diagnosis rate.The operator's experience could also affect the results.The undiagnosis rate was highest in the first 10 cases among all operators.After at least 10 EBUS-TBNA processes,the undiagonsis rate stayed near 7.50％,which was close to the average.Conclusion It is necessary to select suitable indications for EBUS-TBNA based on the disease,lesion location and operatior experience,and cooperate with mediastinoscopy to rise diagnosis rate.

Objective To establish a mouse model by transplanting subcutaneously human pancreatic cancer tissue fragments.Methods Surgically resected pancreatic cancer tissue fragments from patients were transplanted into NOD/SCID mice subcutaneously,and then the growth of the tumor and transplanting it into the next generation were observed.The growth rate,HE staining and immunohistochemistry staining of Ki67 and VEGF were compared.Results We have obtained 13 cases ofpancreatic cancer tissues and 6 cases of biopsy specimens.In 5 cases transplantation was successful,in onemouse model passing to fourth generation,in 4 models to second generation.With the increase of generaions,tumor growth accelerated.HE staining showed later passage cells behavior in an identical manner as the primary cells.Immunohistochemistry staining showed that expressions of Ki67 and VEGF are increasing.Conclusions Through transplanting human pancreatic cancer tissue fragments directly,we have constructed mouse model of pancreatic cancer successfully.With the passage of subculture,the malignant degree and invasiveness may increase.

A simple and cost-effective indirect competitive enzyme-linked immune sorbent assay (ic-ELISA) was developed to rapidly screen the content of aflatoxin B1 (AFB1) in lotus seeds, and the results were confirmed by ultra-fast liquid chromatography-tandem mass spectrometry( UFLC-MS/MS). Matrix-matched calibration expressed a good linearity ranging from 0. 171 to 7. 25 µg · L(-1) for AFB, with R2 > 0.978. The medium inhibitory concentration( IC50 ) for AFB1 was 1.29 µg · L(-1), the recovery for AFB1 was 74.73% to 126.9% with RSD < 5%, and the limit of detection (IC10) was 0.128 µg · L(-1). The developed ic-ELSIA method was applied to rapid analysis of AFB, in 20 lotus seeds samples and the results indicated that the contents of AFB, in samples 1-15 were in the range of 1. 19- 115. 3 µg · kg(-1) and in 40% of the samples exceeded the legal limit(5 µg · kg(-1)), while the contamination rate of AFB, in samples 16-20 was 40%. Pearson correlation coefficient(r) reached 0.997 for AFB1 content in the samples detected by ic-ELSIA and UFLC-MS/MS methods. The results proved that the developed ic-ELISA method is simple, sensitive and reliable, and can be used for rapid and high-throughput screening of AFB1 in lotus seeds
Aflatoxin B1 ; analysis ; Drug Contamination ; Enzyme-Linked Immunosorbent Assay ; methods ; Loteae ; chemistry ; Seeds ; chemistry

A cell model is desired for adipocyte differentiation investigation and for high-throughput screening of anti-obesity and anti-diabetes molecules from chemical resources due to the world wide epidemic of obesity and diabetes. In order to establish such a cell model, a plasmid of pPPARgamma2-promoter-EGFP was constructed by inserting a 660bp sequence of mouse PPARgamma2 promoter into the Ase I and Kpn I sites of pEGFP-N3 and transferred into 3T3-L1 preadipocyte cells. The cells were induced to differentiate and the expression of PPARgamma2 was detected by the microscopic observation of EGFP and by RT-PCR assays. The results showed that the EGFP gene expression patterns were similar to that of pPPARgamma2's, which indicated that the EGFP gene was transferred into the mouse 3T3-L1 preadipocyte cells, and its expression was under the control of pPPARgamma2 promoter. RT-PCR assays showed that the EGFP expression authentically represented the stable expression of PPARgamma2. In conclusion, a preadipocyte cell line expressing EGFP under the control of the promoter of adipocyte-specific expression gene PPARgamma2 was generated. The cell line provides a powerful approach for the research of adipocyte differentiation and for the high-throughput screening of anti-obesity and anti-diabetes chemicals.
3T3-L1 Cells ; Adipocytes ; cytology ; Animals ; Cell Differentiation ; Green Fluorescent Proteins ; genetics ; Mice ; PPAR gamma ; genetics ; Promoter Regions, Genetic ; Stem Cells ; cytology

OBJECTIVETo examine dietary zinc supplementation could alleviate the damage of alcoholic liver disease and the relationship with the expression of hepatocyte nuclear factor 4alpha (HNF-4alpha).

METHODS40 adult C57 BL/6 mice were randomly divided into four groups (n = 10): control, zinc, ethanol and zinc plus ethanol, which were sacrificed after fed four different diets for 6 months. Zinc sulfate was added in the drinking water of the Zinc and Zinc Plus Ethanol group and the content was 75 mg/L. Liver regeneration was assessed by immunohistochemical staining of proliferating cell nuclear antigen (PCNA), and the expression of HNF-4alpha was determined by RT-PCR and Western blot. And as to assess the status of oxidative stress of the mice, malondialdehyde (MDA) and superoxide dismutase (SOD) were detected.

RESULTSCompared with the control group, the expression level of HNF-4alpha decreased significantly in the ethanol group (P < 0.05), and the content of MDA increased significantly in this group, while the content of SOD declined significantly (P < 0.05). Compared with the ethanol group, the number of PCNA-positive hepatocytes increased significantly, and the expression level of HNF-4alpha also increased in the zinc plus ethanol group (P < 0.05), and the content of SOD increased in this group, while MDA decreased significantly (P < 0.05).

CONCLUSIONLong term ethanol exposure can lead to oxidoreduction imbalances which can be reversed by zinc supplementation. We suppose that zinc-enhanced liver regeneration is associated with an increase in HNF-4alpha, suggesting that dietary zinc supplementation may have beneficial effects in alcoholic liver disease.

Animals ; Dietary Supplements ; Hepatocyte Nuclear Factor 4 ; metabolism ; Liver ; metabolism ; Liver Diseases, Alcoholic ; metabolism ; therapy ; Male ; Malondialdehyde ; metabolism ; Mice ; Mice, Inbred C57BL ; Superoxide Dismutase ; metabolism ; Zinc Sulfate ; pharmacology ; therapeutic use

OBJECTIVETo investigate the target cells and molecules with sodium valproate induced differentiation of human glioma cells.

METHODSNude mice bearing human glioma xenogenic graft subcutaneously were treated with sodium valproate. The expressions of HDAC1 and Tob genes of xenografts were analyzed with semiquantitative RT-PCR. The CD133+ cells (BTSCs) were isolated from glioma specimens by immunomagnetic sorting, and cultured in the medium containing FCS or in the serum-free medium supplemented with growth factors, respectively, followed by treatment with sodium valproate in vitro for 21 days. The cell surface markers were detected with flow cytometry and confocal microscopy.

RESULTSSodium valproate inhibited the growth of subcutaneous xenografs bearing on nude mice (P<0.05), and up-regulated the HDAC1 expression (P<0.01), down-regulated the Tob expression (P<0.05). The cell surface markers of BTSCs were detected by flow cytometry after sodium valproate treatment for 21 days. In the FCS group, the GFAP or beta-tubulin III positive cells increased significantly (P<0.01), but in the growth factor group, no statistical differences were observed in the GFAP or beta-tubulin III expression (P>0.05). The results of confocal microscopy indicated that the GFAP+ or beta-tubulin III+ cells coexpressed with Nestin.

CONCLUSIONSHDAC1 and Tob genes were the potential target molecules in reversion of the differential inhibition of human glioma cells with sodium valproate. The BTSCs undergoing the processes of differentiation were the target cells for sodium valproate.

AC133 Antigen ; Actins ; analysis ; Animals ; Antigens, CD ; analysis ; Brain Neoplasms ; metabolism ; pathology ; prevention & control ; Cell Differentiation ; drug effects ; Flow Cytometry ; Gene Expression ; drug effects ; Glial Fibrillary Acidic Protein ; analysis ; Glioma ; metabolism ; pathology ; prevention & control ; Glycoproteins ; analysis ; Histone Deacetylases ; genetics ; Humans ; Intermediate Filament Proteins ; analysis ; Mice ; Mice, Nude ; Microscopy, Confocal ; Nerve Tissue Proteins ; analysis ; Nestin ; Peptides ; analysis ; Reverse Transcriptase Polymerase Chain Reaction ; Tumor Cells, Cultured ; Tumor Suppressor Proteins ; metabolism ; Valproic Acid ; pharmacology ; Xenograft Model Antitumor Assays ; methods

AIMTo study the antitumor peptide components in the stems and leaves of mistletoe (Viscum coloratum (Kom.) Nakai), the primary structure of the novel peptide was elucidated.

METHODSCation exchange, gel filtration and HPLC were employed for isolation and purification. Matrix Assisted Laser Desorption Ionization-Time of Flight-Mass Spectrometry was used to determine the mass. The complete amino acid sequence of the novel peptide was obtained by Edman degradation combined with enzyme digestion. The antitumor activity of the peptide in vitro was studied with MTT method.

RESULTSThe primary stucture of the peptide named as viscotoxin B2 is KSCCKNTTGRNIYNTCRFAGGSRERCAKLSGCKIISASTCPSDYPK. The IC50 value of viscotoxin B2 on the Rat Osteoblast-like Sarcoma 17/2.8 cells in vitro is 1.6 mg x L(-1).

CONCLUSIONViscotoxin B2 in V. coloratum, which has high similarity with viscotoxins from V. album, showed antitumor activity.

Amino Acid Sequence ; Animals ; Antineoplastic Agents, Phytogenic ; chemistry ; isolation & purification ; pharmacology ; Bone Neoplasms ; pathology ; Inhibitory Concentration 50 ; Molecular Sequence Data ; Molecular Weight ; Osteosarcoma ; pathology ; Peptides ; chemistry ; isolation & purification ; pharmacology ; Plant Leaves ; chemistry ; Plant Proteins ; chemistry ; isolation & purification ; pharmacology ; Plant Stems ; chemistry ; Plants, Medicinal ; chemistry ; Rats ; Tumor Cells, Cultured ; drug effects ; Viscum ; chemistry

OBJECTIVETo investigate the long-term outcomes of off-pump coronary artery bypass grafting (OPCAB) in patients aged over 75 years and analyze the risk factors affecting the outcomes of the procedure.

METHODSClinical data were reviewed for 97 consecutive patients aged 75 years or above receiving OPCAB at our center between November, 2000 and November, 2013. The perioperative data including length of ICU stay, duration of mechanical ventilation, incidence of postoperative complications and mortality rate of the patients were analyzed. The follow-up data of the patients were also analyzed including all-cause mortality rate and major adverse cardiac and cerebral events (MACCE, including myocardial infarction, cerebrovascular event, and repeated revascularization).

RESULTSThe perioperative mortality rate was 3.09% (3/97) in these patients. Of the 97 patients analyzed, 91 (93%) were available for follow-up for 29-192 months (with a median of 95.61∓34.07 months). The 10-year survival rate of the patients was 62% with a 10-year MACCE-free survival rate of 47.4%. During the follow-up, 6 (6.8%) patients underwent repeated revascularization procedures, 12 (12.37%) had cerebrovascular accidents and 5 (5.15%) had myocardial infarction. Logistic regression analysis showed that hypertension (OR=1.388, P=0.043) and diabetes (OR=1.692, P=0.017) were independent predictors of MACCE, and incomplete revascularization did not increase the risk of postoperative MACCE.

CONCLUSIONOPCAB is safe and effective in elderly patients with good long-term outcomes. Hypertension and diabetes are independent risk factors of MACCE, and adequate control of blood pressure and blood glucose can reduce the incidence of postoperative MACCE. Incomplete revascularization is not detrimental to the long-term outcomes of OPCAB in elderly patients.

OBJECTIVETo study the curative effects of pirfenidone (PF) on pulmonary fibrosis induced by paraquat (PQ) in mice and to provide the theoretical basis for clinical treatment.

METHODSNinety adult healthy male ICR mice were randomly divided into six groups: control group, PQ group, 2 mg/kg Dexamethasone group, 25 mg/kg PF group, 50 mg/kg PF group and 100 mg/kg PF group, there were 15 mice in each group. The corresponding volume of normal saline was given to the each mouse in control group according to the weight, after 2 h 0.1% CMC was given to the each mouse of control group one time by intragastric administration, then the CMC was administrated at regular time until sacrifice. All mice for other 5 groups were exposed to 100 mg/kg PQ by intragastric administration. At 2 h after exposure to PQ, 0.02 ml/10 g dexamethasone and 25, 50, 100 mg/kg PF were given to mice for dexamethasone group and for 3 PF groups by intragastric administration each day for 49 days, respectively. The lung coefficient was calculated and pathological changes of lung tissue were observed by HE staining for each mouse. The hydroxyproline (HYP) level in lung tissue was measured for each mouse. The mRNA level of and the protein level of TGF-β(1) in lung tissue for each mouse were determined, and the protein level of TGF-β(1) in the bronchus-alveolus lavage fluid (BALF) of each mouse was detected.

RESULTSThe survival rates on the 3rd day in PQ group, 3 PF groups and dexamethasone group were 53.33%, 46.67%, 73.33%, 86.67% and 80%, respectively. The survival rates on the 3rd day in dexamethasone group, 50 mg/kg and 100 mg/kg PF groups were significantly higher than those of PQ group and 25 mg/kg PF group (P < 0.05). The lung coefficients of 3 PF groups were significantly lower than that of the PQ group (P < 0.05). The lung tissue HYP levels of dexamethasone group and 3 PF groups were 50.95 ± 11.65, 44.52 ± 9.48, 43.27 ± 6.01 and 40.82 ± 5.90 mg/g respectively, which were significantly lower than that (74.27 ± 3.68) of PQ group (P < 0.01). The TGF-β(1) protein levels of BALF in dexamethasone group, 50 and 100 mg/kg PF groups were 22.03 ± 7.27, 27.75 ± 5.84 and 21.31 ± 6.82 ng/ml respectively, which were significantly lower than that (52.52 ± 15.51) ng/ml of PQ group (P < 0.01) The expression level of TGF-β(1) mRNA in 100 mg/kg PF group decreased significantly, as compared with PQ group (P < 0.01).

CONCLUSIONPF could reduce the collagen deposition and pulmonary fibrosis induced by PQ in mice lungs.

Animals ; Disease Models, Animal ; Lung ; metabolism ; pathology ; Male ; Mice ; Mice, Inbred ICR ; Paraquat ; poisoning ; Pulmonary Fibrosis ; chemically induced ; drug therapy ; pathology ; Pyridones ; therapeutic use ; Transforming Growth Factor beta ; metabolism

The purpose of this study was to establish a method for the monitoring of minimal residual disease (MRD) in bone marrow samples of the children with T acute lymphoid leukemia (T-ALL), and to evaluate its value in clinical application. The immuno-phenotype of the leukemic cells were detected by flow cytometry with two sets of 4-color combinations of antibodies against TdT/CD5/cCD3/HLA-DR(+)CD19(+)CD33 and CD34/CD5/cCD3/HLA-DR(+)CD19(+)CD33 in 32 cases of de novo T-ALL and were compared with the results in 10 normal controls. The antibody combination in regions of the two-parameter plots where the leukemic cells appeared were different from the normal cells was screened as the effective combination which was used to monitor MRD in the bone marrows of the T-ALL children after the inductive treatment. The results indicated that the respective effective frequencies of antibodies against TdT/CD5/cCD3/HLA-DR(+)CD19(+)CD33 and CD34/CD5/cCD3/HLA-DR(+)CD19(+)CD33 were 90.6% and 62.5%. 32 cases of childhood T-ALL were successively screened for antibodies combinations of interest and were identified in 100% (32/32) of these cases. After inductive treatment, the positive rate in 129 times of MRD monitoring was 19.4% (25/129) by flow cytometry and 5.43% (7/129) by FAB morphology. It is concluded that monitoring MRD in patients with T-ALL by flow cytometry with two 4 color combinations of fluorescent antibodies is an quick and effective method. The sensitivity of this method is high and it may be of important significance for the treatment and prognostic evaluation in childhood T-ALL.
Adolescent ; Child ; Child, Preschool ; Female ; Flow Cytometry ; methods ; Humans ; Immunophenotyping ; Male ; Neoplasm, Residual ; diagnosis ; immunology ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; diagnosis ; immunology