AIM: To detect the serum proteomic patterns in patients of breast cancer by the method of SELDI-TOF-MS and CM10 ProteinChip, and to screen the biomarker candidates, build and validate the diagnostic models, and evaluate its clinical value in surveillance and follow-up after operation. METHODS: The SELDI-TOF-MS technology and CM10 ProteinChip were used to detect the proteomic patterns of serum from 63 breast cancer patients and 40 healthy women. The biomarker candidates were screened and the diagnostic models were constructed by ZJU-PDAS software. Meanwhile, the model was blind-validated in another 23 patients and 20 healthy women. At the same time, 16 serum samples were detected to evaluate its value in surveillance and follow-up after operation. RESULTS: The best model was composed by two protein peaks (BC1/3.9 kD and BC2/5.6 kD) with its sensitivity and specificity of 87.30% (55/63) and 95.00% (38/40), respectively. The sensitivity and specificity in the blind-validation of new cases were 95.65% (22/23) and 85.00% (17/20), respectively. The diagnostic efficacies were the same to the patients of different stages (P>0.05). The expression of BC1 increased while BC2 decreased after operation. The expression of BC2 in the patients with recurrence or metastasis was higher than that in the tumor-free survivors (P<0.05). CONCLUSION: This method shows its potential in detection, surveillance and follow-up after operation. The method is also useful for screening the novel and better biomarkers in breast cancer.

Anthracosis ; complications ; Bacterial Proteins ; genetics ; Humans ; Methicillin Resistance ; genetics ; Methicillin-Resistant Staphylococcus aureus ; drug effects ; genetics ; Penicillin-Binding Proteins ; Respiratory Tract Infections ; complications ; microbiology ; Sputum ; microbiology

Objective To observe the changes in contrast sensitivity(CS) and color vision in patients with primary acute angle-closure glaucoma(PACG).Methods CS and color vision tests were performed in 35 patients(35 eyes) with preclinical stage of PACG,32 patients(32 eyes) with acute attack stage of PACG and 15 normal subjects(30 eyes).Then patients were followed up 1 month(31 eyes) and 3 months(28 eyes) after operation.Results The results of CS and color vision tests were abnormal in PACG patients when they were compared with those of normal people.The threshold of CS at 18.0 c/d was increased(P

This study aims to construct a eukaryotic expression system for envelope gene of Jaagsiekte sheep retrovirus, observes its localization in 293T cells, and investigates the potential in inducing malignant transformation of NIH3T3 cells. By RT-PCR, the full-length cDNA of envelope gene of Jaagsiekte sheep retrovirus (exJSRV-env) was amplified from the extract of naturally infected sheep lung. The clone of target gene was sub-cloned into eukaryotic expression system pEGFP-C1, and validated by PCR, restriction endonuclease, and sequencing. Bioinformatic analysis concerning biological function and cellular localiza tion of exJSRV-env was also performed. The recombinant clone of exJSRV-env was transfected into 293T cells and NIH3T3 cells by Lipofectamine LTX. The expression and celluar localization in 293T cells were validated by confocal microscopy. Soft agar colony formation assay was employed to test the anchorage-independent growth of NIH3T3. DNA sequencing and restriction enzyme digestion with Kpn I and Hind III indicated the correct construction of the recombinant plasmid, which was named pEGFP-C1/exJSRV-env. Amino acid sequence alignment of exJSRV-env with reference sequences found 85%-100% homogeneity. A YRNM motif was discovered at the cytoplasmic tail of envelope gene, which is exclusively found in exogenous viruses. Phylogenetic tree analysis showed that our clone of exJSRV-env clustered closely with pathogenic exogenous Jaagsiekte sheep retroviruses. Fluorescence microscopy indicated typical membrane localization of exJSRV-env protein. NIH3T3 cells transfected with exJSRV-env lost contact inhibition, and acquired colony forming ability in soft agar. This study indicated that envelope protein of Jaagsiekte sheep retrovirus can induce malignant transformation of mouse fibroblast cell NIH3T3. Discoveries of this study provide a basis for further structural and functional research on Jaagsiekte sheep retrovirus envelope protein.
Amino Acid Sequence ; Animals ; Betaretrovirus ; chemistry ; classification ; genetics ; physiology ; Cell Transformation, Viral ; Green Fluorescent Proteins ; genetics ; metabolism ; Mice ; Molecular Sequence Data ; NIH 3T3 Cells ; Phylogeny ; Retroviridae Infections ; veterinary ; virology ; Sequence Alignment ; Sheep ; Sheep Diseases ; virology ; Transformation, Genetic ; Tumor Virus Infections ; veterinary ; virology ; Viral Envelope Proteins ; chemistry ; genetics ; metabolism

Objective To study specific diagnosis of Spirometra erinaceieuropaei . Methods An enzyme linked immunosorbent assay (ELISA) was studied using highly pure gene engineering antigen expressed by the recombination of the cloned cysteine proteinase gene of Spirametra erinaceieuropaei with expression vector pMAL TM c2. Six sera from patient infected with Spirometra erinaceieuropaei were detected using this method. Results and Conclusion The results showed that the gene engineering antigen reacted strongly with the sera from Spirometra erinaceieuropaei infected patients, but did not with the sera from Cysticercus cellulosae infected patients.

Objective:Osteoprotegerin (OPG) is a key molecule negatively regulating osteoclast differentiation and activation; and the conserved mycobacterial heat shock 70 (HSP70) peptide p111-125 has also been found to inhibit inflammation reactions in chronic arthritis. BaculoDirectTM baculovirus expression system was selected to express recombinant OPG-HSP70 in insect cells.Methods:The human functional fragment (p22-194) of OPG and functional fragment (p111-125) of mycobacterial HSP70 gene were cloned into the transfer vector pENTRTM/SD/D-TOPO. The recombinant plasmid was performed an LR reaction with the BaculoDirectTM Linear DNA to generate recombinant baculovirus DNA. The cultured Sf9 insect cells were directly transferred with the recombinant baculovirus DNA,and the pure recombinant baculovirus was obtained. Then recombinant baculovirus was infected Sf9 insect cells again to express the OPG-HSP70 gene.Results:The target protein was detected at the time of 48h post infection,reached at highest yield at the time of 72h post-infection. A 28kDa protein immunostaining band was detected by Western blotting from lysate of those cells.And the purified protein was obtained by using Ni-NTA system. Functional stuies on the fusion protein showed it significantly reduce osteoclast cell number[(3.10?0.640) cells under each microscope field in treatment group by comparing to (10.70?0.817)cells in the control group] in the osteoclast inhibition test,and reduce the inflammation reaction in a delayed type hypersensitivity (DTH) mice model (P

OBJECTIVETo observe the effect of Baichanting Compound (BC) on dopamine (DA) in striatum of Parkinson's disease (PD) mice, and to screen the optimal component proportion.

METHODSThe PD model was established in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) induced C57BL/6 mice. By using uniform design, they were intervened by three extracts of BC in different proportions [Acanthopanax senticosus extract (X1): white peony root extract (X2): Uncaria rhynchophylla extract (X3) = 30.00: 34.92: 82.50, 48.00: 19.98: 72.19, 18.00: 44.88: 61.88, 36.00: 29.94: 51.56, 54.00: 15.00: 41.25, 24.00: 39.90: 30.94, 42.00: 24.96: 20.63). Equal volume of 5% carboxymethylcellulose sodium was administered to mice in the model group and the normal group by gastrogavage. All medication was lasted for 20 successive days. The dopamine (DA) content was determined by ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS). Except 10 in the normal group, 20 PD model mice were screened and divided into the model group and the BC group (with the optimal proportion) according to random digit table. BC extract in optimal proportion was administered to mice in the BC group by gastrogavage, while equal volume of 5% carboxymethylcellulose sodium was administered to mice in the model group and the normal group by gastrogavage. All medication was lasted for 20 successive days. Praxiology was observed in each group. DA content in striatum was also detected. Results Compared with the normal group, the DA content in striatum decreased significantly in the model group (P < 0.01), suggesting a successful PD modeling. Compared with the model group, the DA content in striatum increased significantly in 1 and 2 groups (P<0.05). According to results of quadratic polynomial stepwise regression statistics, the regression equation obtained was: Y = 0.265 + 0.026 X 2 - 0.056 X 3 + 0.334 x 10(-3) x X1 x X3 + 0.691 x 10(-3) X X3(2). X3 extract was the main factor influencing the effectiveness (P < 0.01). The optimal proportion of BC was predicted by the regression equation: X1 = 54.00 mg/(kg x d), X2 = 44.88 mg/(kg x d), the X3 = 82.50 mg/(kg x d). The pole climbing time was shortened, times of autonomic activities increased, DA content was elevated, all with statistical difference in BC groups (P < 0.01, P < 0.05).

CONCLUSIONBC could increase DA content in PD model mice with the optimal proportion as 54.00: 44.88: 82.50.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine ; Animals ; Disease Models, Animal ; Dopamine ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Mass Spectrometry ; Mice ; Mice, Inbred C57BL ; Motor Activity ; Parkinson Disease ; drug therapy ; metabolism

Objective To screen differentially expressed genes in placentas with hepatitis B virus (HBV)infection and to discuss the molecular mechanism of HBV intrauterine infection.Methods Thirty placenta tissue specimens from HBsAg and HBV DNA positive pregnant women were used as the study group and 30 placenta tissue specimens from normal pregnant women with HBsAg and HBV DNA negativity were served as the control group.The suppression subtractive hybridization(SSH)technique was used.Total RNAs of placenta tissue of the study group were mixed as the tester,and total RNAs of placenta tissue of the control group were mixed as the driver.A subtractive cDNA library was constructed by PCR-selective cDNA subtraction systems.Amplifications of the library were carried out with E.coil strain DH5? by reverse spot hybridization.RT-PCR confirmed that phosphatidylinositol 3-kinase(PI3K)was up-regulated in placenta tissue with HBV infection.Results Colony PCR showed that the clones contained 200-1000 bp inserts. Thirty five clones were confirmed by reverse spot hybridization and analyzed by sequencing and bioinformatics.Thirty three known genes and 2 genes with unknown function were obtained.RT-PCR preliminarily confirmed that PI3K gene was up-regulated in HBV infected placenta.Conclusions The differentially expressed genes in placentas with hepatitis B virus(HBV)infection using SSH technique has been screened out successfully.These differentially expressed genes encoding proteins participating in cell vital metabolism and malformation,and signal conduction-antiapoptosis pathway.This finding brings some new clues for studying the mechanisms of HBV intrauterine infection.

Objective To investigate the features of white matter impairment and its relationship with cognition in patients with amnestic mild cognitive impairment (aMCI).Methods Eighty-three cases of aMCI and 85 normal aging volunteers were scanned with diffusion tensor imaging (DTI) using MR system.All subjects completed the neuropsychological battery.We analyzed the differences between two groups using tract-based spatial statistics and the association between regions in difference and cognition using correlation analysis.Results There were significant differences between aMCI and normal control in the neuropsychological battery including the Mini-Mental State Examination(26.2 ± 2.6 vs 28.3 ± 1.3,F =43.224,P =0.000),Mattis Dementia Rating Scale-2 (131.4 ± 6.9 vs 138.0 ± 3.5,F =62.308,P =0.000),Auditory Verbal Learning Test-delayed recall(2.4 ± 1.6 vs 7.5 ± 2.0,F =324.018,P =0.000),Boston Naming Test(8.7 ± 1.4 vs 9.2 ± 1.0,F =6.821,P =0.010),Rey-Osterrich Complex Figure Test (12.1 ± 7.3 vs 18.5 ± 6.1,F =40.674,P =0.000),Symbol Digit Modulation Test (30.0 ± 10.1 vs 38.6 ± 9.8,F =30.786,P =0.000),Trail-Making Test Part B ((256.8 ± 124.5) s vs (178.1 ± 59.0) s,F =27.601,P =0.000).Significantly higher diffusivity indexes and radial diffusivity were also found in aMCI subjects compared to healthy elders in the parahippocampal,superior longitudinal fasciculus,inferior longitudinal fasciculus,superior fronto-occipital fasciculus,inferior fronto-occipital fasciculus,unciform fasciculus,corticospinal tract,corpus callosum,cingulum,corona radiate.We also found that axial diffusivity was significantly increased in the parahippocampal,superior longitudinal fasciculus,inferior longitudinal fasciculus,superior fronto-occipital fasciculus,inferior fronto-occipital fasciculus,unciform fasciculus,corticospinal tract and corpus callosum,whereas fractional anisotropy changes were not observed in aMCI.Diffusivity indexes values in bilateral frontal lobe (left r =0.67 ; right r =0.70),left cingulum (r =0.63),parietal white matter (r =0.69) and radial diffusivity values in left parietal (r =0.68) were significantly related to Trail Making Test A among aMCI (all P ＜ 0.05).Conclusions In aMCI patients,there was a wide range of white matter damage,with no brain region-specific.Executive function deficit was related to the white matter impairment in bilateral frontal lobe,left cingulate and parietal lobe.The specificity and sensitivity of four DTI parameters fordetecting white matter lesions are variant.Trial registration Clinical Research Center of Jiangsu Province (BL2013025)

ObjectiveTo examine whether the single nucleotide polymorphisms in inflammation-related genes are associated with the risk of amnestic mild cognitive impairment (aMCI).MethodsThe study recruited 116 aMCI patients and 93 matched healthy controls.All subjects underwent extensive assessment of cognitive function,genotyping was carried out on the platform of matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry.Results ( 1 ) There was prominent discrepancy between aMCI and controls in the memory,attention and executive functions,20 minutes delayed recall of auditory verbal memory test (AVMT) (3.0(0.0 ～ 10.0 ),8.0 (0.0 ～ 12.0),t =- 8.533,P ＜ 0.05 ),recall of Rey-Osterrieth complex figure test ( R-O CFT) (11.2 ±8.3,16.1 ±8.0,t=4.216,P＜0.05),digit span test (DST) (12.0(7.0 ～ 19.0),13.0(7.0 ～20.0),Z=-2.516,P＜0.05),trail making test A (TMTA) (80.0s(35.0 ～200.0)s,72.0s(29.0 ～512.0)s,Z=-3.113,P＜0.05),trail making test B (TMTB) ((180.1 ±72.7)s,(141,7 ±52.1)s,t=-4.385,P＜0.05 ).(2) No significant differences were found in frequencies of alleles,genotypes and hapolotypes of inflammation mediator genes ( interleukin 10,interleukin 1 A,interleukin 1 B,tumor necrosis factor,interleukin 6,α1- an-tichymotrypsin gene,transforming growth factor B1 ) between aMCI and controls (P ＞ 0.05 ).ConclusionThe results indicate that polymorphisms in the inflammation-related candidate genes do not appear to be involved in the risk of developing aMCI.