Objective:To investigate the effect of DKK1 on linearly patterned programmed cell necrosis (LPPCN) and vasculogenic mim-icry (VM) and the related molecular mechanism in non-small cell lung cancer (NSCLC). Methods:A total of 173 human NSCLC speci-mens were collected to detect LPPCN by H&E staining, detect VM with CD31/PAS double staining, and investigate DKK1 and related protein expression by immunohistochemistry. The clinical pathological significance of LPPCN, VM, and DKK1 and the correlation of them were analyzed. Human NSCLC H460-DKK1 cells were engrafed in nude mice to evaluate the influence of DKK1 up-regulation on VM and LPPCN in vivo. Results:Approximately, 14.45%(25/173) of NSCLC had VM and 49.71%(86/173) had LPPCN. 25.6%(22/86) of NSCLC cases in LPPCN-positive group formed VM. Both of VM and LPPCN were all correlated with poor differentiation, late TNM stage, easy recurrence and metastasis and poor prognosis in NSCLC. DKK1 expression in the VM-positive group and the LPPCN-positive group was higher than that in the VM-negative group and the LPPCN-negative group, respectively. DKK1, LPPCN, and VM were positive-ly correlated with VE-cadherin, MMP-2,β-catenin nuclear expression and Twist1. H460-DKK1 transplantation tumor model confirmed that DKK1 promotes the expression of VM and LPPCN and related proteins in NSCLC. Conclusion:The increase of theβ-catenin and Twist1 expression induced by DKK1 may promote the formation of LPPCN and VM in NSCLC.

Objective To study the antigenicity of human thyrotropin receptor(hTSHR)amino terminus (amino acid 29～280)and its association with Graves' disease.Methods Total thyroid RNA was prepared from human normal thyroid tissue.RNA was then reversely transcripted and cDNA was subjected to PCR amplification.PCR product was cloned into pcDNA3.1 and the recombinant plasmid was named pcDNA3.1/hTSHR188～940bp. Balb/c mice were immunized with peDNA3.1/hTSHR188～940bp. The levels of serum thyroxin,anti-TSHR antibody(TRAb)and thyroid stimulating antibody(TSAb)were measured,and the pathological changes of thyroid tissue were also observed.Results A 753 bp fragment encoding hTSHR ectodomain amino end was obtained after PCR amplification.Confirmed by Hind Ⅲ restriction enzyme digestion and DNA sequencing,pcDNA3.1/hTSHR188～940bphad been constructed successfully,with the correct sequence and direction of hTSHR188～940bp.In the Balb/c mice treated with pcDNA3.1/hTSHR188～940bp,elevated TRAb in week 6(0.148±0.018)were observed compared with those at week o(0.106±0.006,P＜0.01),and kept a higher level till week 10(0.134±0.011,P＜0.01).T4 and TSAb index values were significantly increased in week 10.Serum T4 concentration increased from(41.02±7.97)μg/L in week 0 to(62.20±12.77)μg/L in week 10(P＜0.01);TSAb index values rose from 0.864±0.076 at week 0 to 1.392±0.615(P＜0.01).Thyroid pathological examination showed that proliferated thyroid follicular epithelial cells and foll icular eapacity increased.Inflammatory cells were occasionally found.Conclusions There are antigen epitopes in hTSHR ectodomain amino acid 29～280,which can stimulate the production of TSAb.And the latter induces hyperthyroidism and Graves' disease like manifestations.It suggests that hTSHR ectodomain amino acid 29～280 is closely associated with Graves' disease,and maybe one of important etiological factors leading to the disease.

OBJECTIVE: To develop the single-tube one-step methylation variable position (MVP) analysis technology-single-tube post-digestion PCR-melting curve analysis (PDP-MCA). METHODS: Based on differentially methylated region (DMR) reported previously as the model, a set of primers with different melting temperatures of products in the two sides of MVP were designed. By using the FastDigest methylation-sensitive restriction enzyme (MSRE), DNA digestion, multiplex amplification, MCA detection and MCA profiles were performed in a single reaction tube. Same samples (peripheral venous blood, semen, and vaginal fluid, 5 samples each type) were tested by single-tube one step MVP and traditional MSRE-PCR MCA technology. To verify the feasibility of this method, the results were compared with that of the traditional technology. The MCA/HRM profiles of different samples were analyzed and compared. RESULTS: When the melting temperature of the fragments had a differential of 2 degrees C, the MCA melting peaks separated well, and MCA detection after multiplex amplification was successful. The single-tube PDP-MCA assay was developed, which integrated multiple reactions (digestion, amplification and detection) into one tube. By this method, the sample-specific profiles and data were analyzed in 2 h, which is similar to that of the traditional method. The rapid classifications of the samples were also realized. CONCLUSION Multiplex MVPs can be analyzed in a single closed-tube. The single-tube PDP-MCA technology is a simple, fast, and automatable method. It can be used for detection of DNA methylation variations.
DNA/isolation & purification* ; DNA Methylation/genetics* ; DNA Primers/genetics* ; Humans ; Multiplex Polymerase Chain Reaction/standards* ; Nucleic Acid Denaturation

ObjectiveTo investigate the difference of pathogenicity between the two genotypes of human metapneumovirus(hMPV) for the further research.MethodsAt various time after hMPV infection in BALB/c mice,viral titers of lung tissue were measured by real-time RT-PCR,pathology was assessed by a histopathological scoring system,airway responsiveness was assayed by animal lung function monitoring equipment.Pathogenicity was then measured by detailed evaluation through the results above.Results There is no significant difference in weight of mice between control group and experimental group through dynamic monitoring ; though the difference was exists in airway responsiveness and pathological changes of mice between control group and experimental group,the differences were not statistically in airway reaction,pathological changes and virus drops among the three groups of experimental group.ConclusionThere is no difference in pathogenicity between the two subtypes of hMPV in infection of BALB/c mice,viral genotype do not appear to be associated with pathogenicity.

OBJECTIVETo observe the protective effect of electroacupuncture (EA) at "Zusanli" (ST 36) on inflammatory injury induced by intestinal ischemia/reperfusion (I/R) in rats.

METHODSForty-eight Wistar rats were randomly divided into a sham injury group, a model group, an EA group and a sham EA group, 12 rats in each group. Intestinal I/R rat models were established by method of clamping with occlusion of superior mesenteric artery (SMA) for 45 min followed by reperfusion. The EA group was treated with EA (2.5 mA, 2 Hz/100 Hz, 0.5 h) at "Zusanli" (ST 36) 30 min before reperfusion, and at the same time, the sham EA group was treated with fast insertion at two non-meridian acupoints on skin surface (2 cm horizontally away from linea alba abdominis and about 5 cm paralleled to cartilago ensiformis downward). No interventions were added on the sham injury group and the model group. The degree of pathological injury in intestines, water rate of intestines, diamine oxidase (DAO) activity and intestinal mucosal blood flow (IMBF) were examined at 1 h and 3 h after reperfusion.

RESULTSAt 1 h and 3 h after reperfusion, the intestinal pathological injury in EA group was significantly attenuated compared with that in model group, and the intestinal water rate of (74.00 +/- 2.11)% and (78.78 +/- 0.80)% in EA group were significantly lower than (80.69 +/- 1.66)% and (83.17 +/- 2.08)% in model group (both P < 0.01), but DAO of (68.83 +/- 4.31) U/L and (47.84 +/- 5.57) U/L as well as IMBF of (152 +/- 5.8) PU and (139.8 +/- 6.1) PU in EA group were significantly higher than DAO of (32.86 +/- 4.72) U/L, (17.01 +/- 2.96) U/L as well as IMBF of (124.7 +/- 8.3) PU and (89.4 +/- 13.2) PU in model group (all P < 0.01). Meanwhile, the above mentioned changes in sham EA group showed no significant differences compared with those in model group (all P > 0.05).

CONCLUSIONElectroacupuncture can not only reduce the inflammatory injury induced by intestinal IR but also increase intestinal blood supply so as to protect the intestine function.

Acupuncture Points ; Amine Oxidase (Copper-Containing) ; metabolism ; Animals ; Electroacupuncture ; Inflammation ; therapy ; Intestines ; blood supply ; metabolism ; pathology ; Male ; Rats ; Rats, Wistar ; Reperfusion Injury ; therapy

Objective Aselectivitystudywasconductedthroughtheexaminationofradionuclides137Cs(Cesium-137)for foodbasedondifferentdetectionconditions.Methods Atotalof48foodsampleswereselectedfromthreeareasincluding Qinshan nuclear power plant,Sanmen nuclear power plant and Hangzhou and Zhoushan respectively.1 37 Cs of these samples were determined by γspectrometry and Phosphoric acid ammonium molybdate method.The level of 48 foods were statistically analyzed,and then the time consuming,sample size requirements,influence factors were comprehensively discussed,thustheselectionreferenceproposaloftheexaminationmethodcouldbeprovided.Results Therewereno significant difference for the data of two examination method (P>0.05 ).The limit of detection of the γspectrometry was lower (P <0.05 ).Compared with Phosphoric acid ammonium molybdate method,γspectrometry had lower limit of detection,and could detect a variety of radionuclides at a time,but need more sample and time-consuming when multi-sample were detected.The limit of detection of the Phosphoric acid ammonium molybdate method was high,and the chemicalprocessingstepswerecumbersomeandwaseasytobeinterferedby134Cs.Conclusion Thelimitofdetectionof the γspectrometry is low,and the sensitivity of the Phosphoric acid ammonium molybdate method is high.Most food are recommended to be detected by γspectrometry in the practical work,and the food which were difficult collected,less ash or low content,are recommended to be detected by Phosphoric acid ammonium molybdate method.

OBJECTIVETo highlight the clinical features and diagnosis of chronic disseminated tuberculosis, with emphasizing the usefulness of several recently available diagnostic technologies in this setting.

METHODWe presented a case of chronic disseminated tuberculosis diagnosed with the combined application of interferon-gamma release assay T-SPOT. TB, 18F-fluorodeoxyglucose (FDG)-positron emission tomography (PET), and gene chip assay.

RESULTSA 53-year-old gentleman who had chronic cough for 7 years and fever for 2 weeks was referred to our hospital for further evaluation. 18F-FDG-PET/CT scan showed increased FDG uptake in multiple lesions involving bilateral lungs, supraclavicular, mediastinal and intro-abdominal lymph nodes and bones, mimicking metastatic malignancy. T-SPOT. TB assay revealed significant responses [ early secreting antigen target 6 (ESAT-6): 3 908 spot forming cells (SFCs)/10(6) peripheral blood mononuclear cells (PBMCs), culture filtrate protein (CFP-10): 3 400 SFCs/10(6) PBMCs]. Subsequent biopsy of supraclavicular lymph node, lung, and ilium revealed granulomas, while culture of the obtained tissue yeilded mycobacteria. Gene chip testing identified M. tuberculosis sensitive to isoniazid and rifampin. After 10 weeks of treatment for tuberculosis, the patient's condition was improved and a second T-SPOT. TB assay showed significantly reduced responses (ESAT-6: 1528 SFCs/10(6) PBMCs; CFP-10: 1460 SFCs/10(6) PBMCs).

CONCLUSIONSTimely diagnosis of chronic disseminated tuberculosis requires high index of suspicion. T-SPOT. TB assay, PET/CT, and gene chip assay may provide valuable information that facilitates further diagnostic procedures and treatment decision.

Enzyme-Linked Immunospot Assay ; Humans ; Leukocytes, Mononuclear ; Mycobacterium tuberculosis ; Oligonucleotide Array Sequence Analysis ; Positron-Emission Tomography ; Tuberculosis ; diagnosis

Objective To investigate the effect on radioactivity in drinking water around Qinshan nuclear power station (QNPS)in normal operational condition.Methods The field monitoring and laboratory analysis methods were adopted to detect the total radioactivity level in drinking water in 2015,according to different distances from the nuclear island and different types of water.Results The total alpha and total beta radioactivity level in drinking water around QNPS were 0.027(0.098)Bq /L and 0.263(0.071)Bq /L respectively,which were obviously lower than the national health standard limits(total alpha and total beta are 0.5,1.0 Bq /L respectively).Total radioactivity level had no relation with the distance from the nuclear island (P >0.05).The total alpha radioactivity in deep well water was the highest among the investigated three types of drinking water,and the highest value was 0.224 Bq /L.The beta radioactivity level in river water was the highest,and the highest value was 0.408 Bq /L.The total alpha radioactivity level was 0.017 (0.013)Bq /L in 2015, higher than the average level during 2010—2014.The beta radioactivity average level was 0.319 (0.102)and 0.289 (0.055)Bq /L,also higher than the average level during 2010—2014.Conclusion The total radioactivity in drinking water among nuclear power plant is in normal background level,so at present there is no effect of the radioactive contamination on drinking water around QNPS in nuclear power plant's normal operational condition.

Objective To establish a high performance liquid chromatography (HPLC) through the optimization of the chromatographic conditions, which can detect the contents of clenbuterol hydrochloride (CL) residues in animal edible product in a large quantity. Methods The animal edible product were extracted by perchloric acid, and then impurities were removed by liquid-liquid extraction (LLE) which used ethyl acetate- isopropanol. After the organic phase was concentrated, C18 column (150 mm×4.6 mm, 5 μm) was used to separate CL. Mobile phase were methanol-sodium dihydrogen phosphate, and then determined by HPLC. Results A good linear response was obtained over the range of 0.2-10.0 μg/mL with the correlation coefficient (r) 0.99984. The method determination limit was 0.15 μg/kg which was lower than the National standard method 0.5μg/kg. The retention time of the CL was 6.51 min, the chromatographic peak was good. The recovery rates spiked with standards 1.6-12 μg were 92.86％-100.93％, which was higher than National standard method (89.79％-92.36％) . The precision of intra-day and inter-day were both under 5％, which lower than National standard. Conclusion The optimized chromatographic conditions are suitable for the large quantity determination of clenbuterol hydrochloride in animal edible product.

Animals ; Apoptosis ; drug effects ; Cell Division ; drug effects ; Drugs, Chinese Herbal ; pharmacology ; Flow Cytometry ; In Situ Nick-End Labeling ; Liver ; cytology ; drug effects ; ultrastructure