Fluorescence enhancement reaction of flavoxate hydrochloride (FX) in strong alkali solution was studied, the mechanism of the reaction was investigated, and a novel fluorimetric method for analysis of FX in drug sample was established. FX has no intrinsic fluorescence, but it can slowly produce fluorescence in strong alkali solution. Heating can promote the fluorescence enhancement reaction. In 3D fluorescence spectra of the decomposition product of FX, two fluorescence peaks, located respectively at excitation wavelengths λex/ emission wavelength λem =223/410 nm, and 302/410 nm, were observed. Using quinine sulfate as a reference, fluorescence quantum yield of the decomposition product was measured to be 0.50. The structural characteriza- tion and spectral analysis of the decomposition product reveal that ester bond hydrolysis reaction of FX is firstly occurred during heating process, forming 3-methylflavone-8-carboxylic acid (MFA), then a cleavage reaction of the γ-pyrone ring of MFA occurred, producing α, β-unsaturated ketone. This product includes adjacent hydroxyl benzoic acid group in its molecule, which can form intramolecular hydrogen bond under alkaline condition, so that increase the conjugate degree and enhance the rigidity of the molecule, and thereby cause fluorescence enhancement. Based on this fluorescence enhancement reaction, a fluorimetric method was proposed for the determination of FX. A linear calibration curve covered the concentration range 0.020 3-0.487 µg · mL. The regression equation was I(F) = 23.9 + 5357.3 c, with correlation coefficient r = 0.999 7 (n = 8), detection limit D = 1.1 ng · mL(-1). The method was applied to the analysis of FX tablets, with a spiked recovery rate of 100.2%. The reliability of the method was verified by a UV-spectrophotometric method.
Alkalies ; Calibration ; Chemistry, Pharmaceutical ; Flavoxate ; analogs & derivatives ; chemistry ; Fluorescence ; Limit of Detection ; Reproducibility of Results ; Solutions ; Tablets

In order to confirm the role that the 49th amino acid residue plays in enzymatic inactivity of Glutamine 49 phospholipase A2(Gln49-PLA2),site-directed mutagenesis of its 49th amino acid gene codon was conducted using PCR.Aspartic acid 49 phospholipase A2(Asp49-PLA2-Q49D-PLA2),the mutant of Gln49-PLA2 was expressed in E.coli with pET32a+ vector.The fusion protein,expressed as inclusion body,after being denatured,was on-column refolded and purified by immobilized metal affinity chromatography(IMAC),and then cleaved by Factor Xa.The mature Q49D-PLA2 mutant was obtained by Hitrap SP cation exchange and Superdex 75 gel filtration chromatography,with the recovery rate of 1.3%,and the specific activity of the mature Q49D-PLA2 mutant was 72 U/mg.It has been demonstrated that the 49th glutamine amino acid residue is the main reason in enzymatic inactivity of Gln49-PLA2 and the results are helpful for denatured protein refolding,especially in rich disulfide bonds conditions.

OBJECTIVETo study the role of protein kinase C-delta (PKC-delta) in hyperthermia-induced apoptosis in human tongue squamous cell carcinoma Tca8113 cells.

METHODSTca8113 cells were treated at 43 degrees C in a heating water bath for 0, 40, 80, 120 min after pretreatment with Rottlerin, a specific inhibitor of PKC-delta, and equal volume dimethyl sulfoxide (DMSO) for 30 min, respectively. The cells were stained by propidium iodide (PI) and Rhodamine 123 to analysis apoptotic rate and the changes of mitochondrial transmembrane potential by flow cytometry (FCM). The total proteins were extracted for Western blotting analysis of activation and proteolysis of PKC-delta, and for colorimetric assay of relative activity of Caspase-3.

RESULTSHyperthermia could induce proteolysis and activation of PKC-delta, and this was attenuated by Rottlerin. Apoptotic rate, decreasing of mitochondrial transmembrane potential and activity of Caspase-3 which being induced by hyperthermia in Tca8113 cells were inhibited by PKC-delta specific inhibitor Rottlerin. There were significantly statistical differences in apoptosis rates, mitochondrial transmembrane potential and activity of Caspase-3 between Rottlerin- and non-Rottlerin-pretreated cells after hyperthermia for 40, 80, 120 min (P < 0.01).

CONCLUSIONActivated PKC-delta may facilitate hyperthermia-induced apoptosis in Tca8113 cells, and may be one of the mechanisms of apoptosis induced by hyperthermia.

Acetophenones ; Apoptosis ; Benzopyrans ; Carcinoma, Squamous Cell ; Humans ; Protein Kinase C ; Protein Kinase C-delta

OBJECTIVETo improve the accuracy of preoperative evaluation of cervical lymph node metastasis in thoracic esophageal squamous carcinoma.

METHODSForty-two patients with thoracic esophageal squamous carcinoma underwent neck ultrasonography. Enlarged lymph nodes with their long axis greater than 10 mm and a short-to-long axis ratio greater than 0.5 were considered as metastatic.

RESULTSPreoperative neck ultrasonography revealed the enlarged lymph nodes in 16 patients, but only in 5 (31%) cases the nodes were palpable. Among them 9 were classified as metastatic (cM(1-LN)), including 4 patients with palpable nodes. In 5 cM(1-LN) patients surgical intervention was canceled and the remaining 37 patients underwent trans-thoracic esophagectomy. Cervical node metastasis (pM(1-LN)) was confirmed pathologically in 6 surgical patients, 4 with tumors invading the adventitia (pT3) and the other 2 into the surrounding structure (pT(4)) (pT(1), pT(2) vs. pT(3), pT(4), P = 0.020). All 6 pM(1-LN) patients had concomitant mediastinal node metastasis and 4 of them had upper abdominal node metastasis. Statistically significant relationship was detected between cervical and abdominal nodal status (r = 0.536, P = 0.007). In comparison with the results of pathological examination and treatment response, the accuracy and sensitivity were 81% and 95% (P = 0.043), 36% and 82% (P = 0.081), respectively, for palpation and ultrasonography. Five out of 39 (13%) patients had their therapy changed due to ultrasonographic findings.

CONCLUSIONSNeck ultrasonography for cervical lymphadenopathy is of high sensitivity and accuracy, which plays an important role in the preoperative evaluation and therapeutic decision-making.

Adult ; Aged ; Carcinoma, Squamous Cell ; diagnostic imaging ; secondary ; surgery ; Esophageal Neoplasms ; pathology ; surgery ; Female ; Head and Neck Neoplasms ; diagnostic imaging ; secondary ; surgery ; Humans ; Lymph Node Excision ; methods ; Lymph Nodes ; diagnostic imaging ; Lymphatic Metastasis ; diagnosis ; Male ; Middle Aged ; Neck ; diagnostic imaging ; Sensitivity and Specificity ; Ultrasonography

Objective To analyze the viral genetic characteristics of hantaviruses carried by Microtus maximowixzii in Yakeshi of Inner Mongolia Autonomous Region and its relationship with Hantaan virus (HTNV) and Seoul virus (SEOV) viruses as well as to identify the natural host of Khabarovsk virus (KHAV).Methods HV specific RNAs were detected by RT-PCR.Complete S and M segment were amplified from the RNA-positive samples.Phylogenetic analysis were performed to estimate the genetic characterization and the relationship with other hantaviruses.Results Fifty two Microtus maximowixzii voles were captured in Yakeshi areas.Of those voles,hanta-viral RNA was tested positive in 5 samples (9.62％).Complete S and M segments sequences were obtained from 5 and 2 lung samples,respectively.The complete S segment was consisted of 1848 to 1861 bp,and the M segment consisted of 3662 bp.These viruses were closely related to each other with 92.5％-96.4％ for the S segment sequences and 88.9％-95.4％ for the M segment sequences.They shared a higher identity with KHAV found previously in Yakeshi and KHAV of Russia.However,they were obviously different from the other hantavirus species.The 5 strains had the consistent secondary structure of nucleocapsid protein (NP) and glycoprotein (GP).When further comparing their secondary structures with those of HTNV and SEOV,our results indicated that there were no obvious differences in NP between KHAV and both HNTV,SEOV but with obvious difference in GP.Based on the S and M segment sequences,phylogenetic analyses revealed that these 5 strains clustered together with KHAV and formed a distinct lineage.Furthermore,all known KHAV strains could be divided into two small branches with a nucleotide divergence more than 5.3％.Conclusion Our research data revealed that KHAV was highly endemic among Microtus maximowixzii in Yakeshi area which supported the notion that Microtus maximowixzii had been the natural host of KHAV in the area.

OBJECTIVETo observe the role of interleukin (IL) 21 alone, IL12 alone, and IL21 plus IL12 for inducing the antitumor activity of peripheral blood mononuclear cells (PBMCs) in patients with endometrial cancer.

METHODSPBMCs were isolated from peripheral blood in patients with endometrial cancer in vitro, and kept the culture with low-level IL2. IL2-stimulated PBMCs were cocultured under different conditions (with anti-IL21 antibody, IL21 alone, IL12 alone, or IL21 plus IL12) for 72 h. The cytotoxicity of PBMCs was then examined by lactate dehydrogenase(LDH) released assay. CD4(+) CD25(+) FOXP3(+) regulatory (Treg) cell and CD4(+) IL17A(+) T-helper (Th17) cell proportion were determined with flow cytometry. Cell proliferation and apoptosis were measured by cell counting kit-8(CCK-8)assay and flow cytometry, respectively.

RESULTSIn comparison to control group, both IL21 and IL12 significantly enhanced the cytotoxicity of PBMCs. The IL21 plus IL12 group had superior effect to IL21 alone and IL12 alone. IL21 and IL12 significantly decreased the percentages of Treg cells and the rate of PBMCs apoptosis. IL21 or IL12 had no significant effect on the differentiation of Th17 cells and the proliferation of PBMCs.

CONCLUSIONSIL21 and IL12 can enhance the cytotoxicity of PBMCs in patients with endometrial cancer, which can be further strengthened with treatment of IL21 plus IL12. Such effects may be achieved by inhibiting the differentiation of Treg cells and the apoptosis of PBMCs, but not by the differentiation of Th17 cell.

Adult ; Aged ; Apoptosis ; drug effects ; Cell Proliferation ; drug effects ; Cells, Cultured ; Endometrial Neoplasms ; immunology ; pathology ; Female ; Humans ; Interleukin-12 ; pharmacology ; Interleukins ; pharmacology ; Leukocytes, Mononuclear ; drug effects ; immunology ; pathology ; Middle Aged ; T-Lymphocytes, Regulatory ; immunology

OBJECTIVETo analyze the value of intraoperative ultrasonography (IOUS) in the diagnosis and therapy of insulinoma.

METHODSFrom January 2000 to December 2007, the application of intraoperative ultrasonography used in 44 cases with insulinoma who came from department of general surgery, First Affiliated Hospital of Zhengzhou University were retrospectively analyzed. There were 19 males and 25 females in the group. Every case accepted operation and examination of IOUS.

RESULTSTumor was accurately located and its adjacent structure was also clear by IOUS in 43 cases, the other one was islet cell hyperplasia, the detection rate of tumor was 100%. The complications: one case occurred pancreatic fistula, one occurred pancreatitis, and there was no biliary fistula and hemorrhage.

CONCLUSIONSPresently, IOUS is one simple and effective method to local insulinoma, and it could improve the success rate of operation and reduce complications.

Adolescent ; Adult ; Aged ; Female ; Humans ; Insulinoma ; diagnostic imaging ; surgery ; Intraoperative Care ; Male ; Middle Aged ; Pancreatic Neoplasms ; diagnostic imaging ; surgery ; Retrospective Studies ; Treatment Outcome ; Ultrasonography ; Young Adult

OBJECTIVETo explore the relationship between two exonic polymorphisms of DNA repair gene XPC and the susceptibility to lung cancer.

METHODSGenotypes were determined by the primer introduced restriction analysis-PCR(PIRA-PCR) and the PCR-restriction fragment length polymorphism(PCR-RFLP) approaches, respectively, in 320 histologically-confirmed lung cancer cases and 322 age and sex frequency-matched cancer-free controls.

RESULTSMultivariate logistic regression analysis revealed that individuals carrying at least one 499Val variant allele (Ala/Val + Val/Val genotypes) had a significantly increased risk for lung cancer (adjusted OR=1.54; 95%CI: 1.11-2.14), compared with the wild-type genotype (499Ala/Ala). Furthermore, individuals with both putative risk genotypes had a significantly higher risk (adjusted OR=2.55; 95%CI: 1.45-4.52), compared with those with both wild-genotypes. In addition, a potential super multiplicative gene-environment interaction between Ala499Val genotypes and smoking on lung cancer risk was unveiled. The odds ratios of lung cancer for individuals with both putative risk genotypes were 2.63 (95%CI=1.23-5.62) in nonsmokers and 7.36 (95%CI=3.19-17.0) in smokers, respectively.

CONCLUSIONThese findings support the hypothesis that these two XPC variants may contribute to the risk of developing lung cancer.

Asian Continental Ancestry Group ; genetics ; China ; DNA-Binding Proteins ; genetics ; Exons ; genetics ; Female ; Genetic Predisposition to Disease ; genetics ; Genotype ; Humans ; Lung Neoplasms ; ethnology ; genetics ; Male ; Middle Aged ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length ; Polymorphism, Single Nucleotide ; Risk Factors

OBJECTIVETo measure the diameter and length of infrarenal inferior vena cava (IVC) in Shandong Peninsula adult through digital subtraction angiography (DSA) for better vena cava filter (VCF) choice and placement.

METHODSFrom April 2008 to June 2010, 83 discontinuous patients (49 males and 34 females, mean age 56.4 years) with deep venous thrombosis (DVT) of lower extremity were placed VCF through DSA according to ACCP-8. During operation, diameter and length of infrarenal IVC were measured. At the same time, the renal vein location and the type of the IVC were identified to help the VCF choice.

RESULTSAll the VCFs were placed successfully, no complications occurred. The diameter of infrarenal IVC was 10 to 26 mm with a mean of (19 ± 5) mm. The average length from beginning of IVC to the lower renal vein was (10.6 ± 2.8) cm. The renal vein was located between the first and second lumbar vertebra, the IVC beginning was located between the fourth and fifth lumbar vertebra.

CONCLUSIONSDiameter and length measurement of infrarenal IVC is helpful to the VCF selection and the domestic VCF research. Vena cava angiography is very important to the accurate placement of VCF.

Asian Continental Ancestry Group ; Female ; Humans ; Male ; Middle Aged ; Phlebography ; Vena Cava Filters ; Vena Cava, Inferior ; diagnostic imaging

OBJECTIVETo investigate the process and HPLC fingerprint of Charred Radix Scutellariae, and lay a foundation of Charred Radix Scutellariae quanlity control mode.

METHODSelect HPLC-UV fingerprint. Chromatogram condition: Hypersil C18 column (5.0 mm x 200 mm, 5 microm), mixtures of methanol, 0.4% phosphoric acid and acetonitrile as mobile phase in a gradient mode. Flow rate: 1.0 mL x min; Detection wavelength was set at 277 nm.

RESULTThere were no evident differences among Charred Radix Scutellariae that were normatively manufactured and processed.

CONCLUSIONThis test proves the process is feasible, this method recurs well and can be used to provide scieitific basis for the normative process and quanlity control mode of Charred Radix Scutellariae.

Charcoal ; chemistry ; Chromatography, High Pressure Liquid ; methods ; Flavanones ; analysis ; Flavonoids ; analysis ; Hot Temperature ; Plant Roots ; chemistry ; Plants, Medicinal ; chemistry ; Quality Control ; Scutellaria baicalensis ; chemistry ; Technology, Pharmaceutical ; methods