Objective To investigate the expression of survivin and its correlation with tumor thymidine kinase and axillary lymph node metastasis in breast cancer. Methods Expression of survivin and cytosolic thymidine kinase(TK) in 60 cases of breast cancer(28 cases with axillary lymphonode metastasis) were measured by immunohistochemistry. Results There was expression of survivin in breast cancer; labeling index(LI) of TK in breast cancer cells with positive expression of survivin was significantly higher than that with negative expression of survivin;the positive percent of survivin expression in breast cancer with axillary lymph node metastasis was higher than that without axillary lymph node metastasis. Conclusions The expression of survivin in breast cancer is related to TK and axillary lymph node metastasis.

Objective: To construct subtracted cDNA libraries of stomach tumors and normal stomach tissue using suppression subtractive hybridization(SSH).Methods: cDNA Library subtraction was performed using the protocol described in the Clontech PCR-Select cDNA Subtraction Kit. cDNA was synthesized from 2 μg of poly A+RNA from the tumor and normal tissues using AMV reverse transcriptase. The tester and driver cDNAs were digested with RsaⅠ, a four-base-cutting restriction enzyme that yields blunt ends. The tester cDNA was then subdivided into two portions, and each was ligated with different cDNA adaptor. Two hybridizations were performed. In the first, an excess of driver was added to each sample of tester. Hybridization kinetics led to equalization and enrichment of differentially expressed sequences. During the second hybridization, the two primary hybridization samples were mixed together without denaturing and thus the templates were generated from differentially expressed sequences for PCR amplification. Using suppression PCR, only differentially expressed sequences were amplified exponentially and after second PCR amplification the background was reduced and differentially expressed sequences were further enriched. The cDNAs were then directly inserted into a T/A cloning vector to generate a stomach tumor-specific subtracted cDNA library. Results: The amplified library contained 800 positive clones. Plasmid inserts were PCR amplified and showed 250－700 bp inserts. Conclusions: The successfully constructed subtracted cDNA library of gastrocarcinoma and normal tissue enables us to compare two populations of mRNA and obtain clones of genes that expressed in one population but not in the other.The characterization of these genes will allow them to be exploited for their diagnostic and therapeutic potentials.

Objective To compare the clinical effects of proximal femoral nail antirotation (PFNA) and dynamic hip screw (DHS) in the treatment of intertrochanteric fractures in elderly patients.And provide a reasonable basis for clinical treatment of elderly patients with intertrochanteric fractures.Methods The clinical and follow-up records of 58 elderly patients with intertrochanteric fractures treated by PFNA and DHS were retrospectively reviewed.For times of operation,blood loss in operation,length of incision,incidents of complication and Harris's hip functional standard score were compared and analyzed.Results PFNA group operation time[(59.61 ± 8.27)min],amount of bleeding[(234.51 ± 38.80)ml] were both better than the DHS group of[(83.54 ± 11.12)min and (446.57 ±54.01) ml] respectively.There were significant differences (t =9.80,18.10,all P ＜ 0.05).The satisfactory rate was 92.8％ in PFNA group,higher than the DHS group's 83.3 ％ (x2 =6.18,P ＜ 0.05).There was significant difference between two groups in the incident of complication,DHS group was higher.Conclusion PFNA internal fixation with a minimally invasive,fixed solid and recovery fast,is the better internal fixation in the treatment of elderly intertrochanteric fracture patients.

12E7 Antigen ; Adolescent ; Adult ; Antigens, CD ; metabolism ; Biomarkers, Tumor ; metabolism ; Brain Neoplasms ; secondary ; Cell Adhesion Molecules ; metabolism ; Child ; Child, Preschool ; Diagnosis, Differential ; Extremities ; Female ; Follow-Up Studies ; Humans ; Immunohistochemistry ; In Situ Hybridization, Fluorescence ; Infant ; Ki-67 Antigen ; metabolism ; Male ; Neuroectodermal Tumors, Primitive ; metabolism ; pathology ; Oncogene Proteins, Fusion ; metabolism ; Repressor Proteins ; metabolism ; Sarcoma, Ewing ; metabolism ; pathology ; Sarcoma, Synovial ; diagnosis ; metabolism ; pathology ; surgery ; Soft Tissue Neoplasms ; diagnosis ; metabolism ; pathology ; surgery ; Vimentin ; metabolism ; Young Adult

Objective In this study,we attempted to investigate whether Electroacupuncture(EA)could promote the autophagy function in macrophages of inflammatory skin tissues by activating CB2 receptor,thus relieving inflammatory pain induced by CFA in mice,and whether activation of CB2 receptor in NR8383 macrophages cell line can simulate the effect of EA on the autophagy function and mitochondrial damage.Methods Inflammatory pain model was induced by CFA injection into the planta the hind paw of wildtype and CB2 knockout mice.EA or sham EA was applied on the left Huantiao(GB30)and Yanglingquan(GB34)sites.Thermal hyperalgesia was determined with the Hargreaves test.Mechanical sensitivity was assessed with von Frey filaments.NR8383 microphage cell line was used to study the effect of CB2 activation on macrophage function induced by CFA.The expression level of autophagy protein LC3 and p62 in wildtype and CB2 knockout mice skin tissue and NR8383 cell line were determined by Western blot.And flow cytometry analysis was applied to detect damaged mitochondria and mitochondrial superoxide.Results CFA significantly reduced the thermal and mechanical pain threshold in both wildtype and CB2 knockout mice,comparing with the vehicle control groups(P<0.01).EA significantly inhibited thermal and mechanical hyperpathia induced by CFA in wildtype mice(P<0.05),but had no effect on CB2 knockout mice with CFA(P>0.05).CFA significantly increased the expression of p62 protein and decreased LC3-II/I ratio,which was inversed by EA in wildtype mice but wasn't affected by EA in CB2 knockout mice.CFA increased the expression of p62 protein and decreased LC3-II/I ratio in NR8383 cell line,which were inversed by CB2 agonist AM1241.CFA increased mitochondria damage,which were then attenuated by CB2 agonist AM1241.Conclusion The analgesic effect of EA on inflammatory pain induced by CFA was mediated by activation of CB2 receptor,which promoted the autophagy function and the clearance of damaged mitochondria in macrophage.

BACKGROUNDTo construct the full-length complementary DNA of HCV genome from an HCV infected patient.

METHODSFour HCV gene fragments (1.6, 3.5, 2.4 kb and 2.6 kb) were amplified by RT-PCR from serum of a Chinese patient and fused and connected together to produce a 9.2 kb subgenomic fragment, which was further cloned into a cassette vector with fixed 5-prime and 3-prime termini of HCV to make the full-length cDNA. The cDNA heterogeneity was analyzed by comparing the sequences of 4 isolated HVR1 regions. The prokaryotic expressed Core, NS3 protease, NS3 helicase were detected for their specific reactivities with patient serum by Western blot analysis. And the protease activity of NS3 was evaluated in a cell-based NS3/4A-SEAP expression system.

RESULTSThe cDNA covered the near full-length of HCV genome from the patient's serum. The difference among HVR1 regions indicated no selection of HCV variants during RT-PCR and the quasi-species characteristic of the amplified cDNA. The prokaryotic expressed viral proteins could be identified by patient serum. In the NS3/4A-SEAP system, NS3 could cleave the 4A-4B site between NS4A and SEAP proteins and resulted in the secretion of SEAP in culture media.

CONCLUSIONThese results suggest that the cloned HCV cDNA encodes a complete and functional open reading frame and will be useful for further construction of infectious cDNA clone.

Base Sequence ; Cloning, Molecular ; DNA, Complementary ; genetics ; Genome, Viral ; Hepacivirus ; genetics ; Hepatitis C ; virology ; Humans ; Molecular Sequence Data ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo study the chemical constituents from the stem bark of Trewia nudiflora.

METHODThe chemical constituents were isolated by silica gel and sephadex LH - 20 column chromatography, and the structures were elucidated by means of spectral analysis.

RESULTTen compounds were obtained from EtOAc fraction of EtOH extract and identified as stigmast-4-en-6beta-ol-3-one (1), stigmast-4-en-6alpha-ol-3-one (2), 7beta-hydroxysitosterol (3), 7alpha-hydroxysitosterol (4), schleicheol 2 (5), taraxerone (6), abbeokutone (7), beta-hydroxypropiovanillone (8), o-vanillyl alcohol (9), glycerol monopalmitate (10).

CONCLUSIONCompounds 1-5 and 7-9 were isolated from this plant for the first time.

Chromatography, Gel ; Drugs, Chinese Herbal ; chemistry ; Mallotus Plant ; chemistry ; Plant Bark ; chemistry ; Plant Stems ; chemistry

OBJECTIVETo analyze expression characteristics of human skin epidermal stem cell at different developmental stages, and to explore its biological significance.

METHODSHealth skin samples from 28-32 w fetuses (F group), 4-12 y children (C group), and 35-55 y adult (A group) were harvested, with 10 cases in each group. Epidermis were separated using trypsin digestion and EDTA, and human epidermal stem cells were isolated and purified with type IV collagen attachment method. The monoclonal antibody of integrin beta1 and keratin 19 were used for detection and identification of epidermal stem cells by immunohistochemical staining. Total RNA was extracted from above cells by Trizol one-step method, and were detected by formaldehyde denaturing agarose gel electrophoresis. Probes were prepared and hybridized into cDNA microarray for scanning fluorescent signals and analysis of images, with two-fold differential expression value for screening. Significantly up/down-regulated genes were selected for verification by real time RT-PCR.

RESULTSBy comparing expression profile between A and C groups, a total of 1808 genes with differential expression were detected, including 1089 up-regulated genes and 719 down-regulated genes, and they were classified into 128 categories. Among them, 1462 genes were known (found in GeneBank), 346 genes were unknown. A total of 4534 genes with differential expression were detected between C and F groups, in which 1783 genes were up-regulated and 2751 genes were down-regulated, and they were classified into 216 categories. Among them, 3577 genes were known (found in GeneBank), and 957 genes were unknown. There were 1104 genes with differential expression consistently detected in F, C and A groups, which were classified into 32 categories according to gene function. Among them, 94 genes were consistently up-regulated and 75 genes consistently down-regulated. Test results of real time RT-PCR were in accordance with above-mentioned results.

CONCLUSIONSGene expression profiles of epidermal stem cells cultured in vitro, harvested from fetuses, children, and adult, exhibit obvious difference. This may be closely related to different stages of proliferation and differentiation of human epidermal stem cell and self-repair ability of wound at different developmental stages.

Adult ; Cell Differentiation ; Child ; Child, Preschool ; Epidermis ; cytology ; growth & development ; Epithelial Cells ; cytology ; Fetus ; cytology ; Gene Expression Profiling ; Gene Expression Regulation, Developmental ; Humans ; Middle Aged ; Oligonucleotide Array Sequence Analysis ; methods ; Stem Cells ; cytology ; Transcriptome

OBJECTIVETo study the platelet morphology and function of an inherited macrothrombocytopenia disorder with abnormal large granules.

METHODSPlatelet size and structure were investigated by both light microscopy and electron microscopy. The platelet membrane expression of GP I b, GP II b, GPIII a, P-selectin and CD63 were analyzed by using respective monoclonal antibodies. Platelet 5-hydroxy-tryptamine was measured with spectrophotofluorometer.

RESULTSBoth the patient and her father had large granules in their platelets, with exocytosis being easily observed. The expressions of GP I b, GP II b and GP II a on the platelets were in normal range, while P-selectin and CD63 were somewhat increased. The abnormal large granules were not the alpha granules, lysosomes or dense bodies.

CONCLUSIONBoth morphological and functional abnormalities of the platelets from the patient are clearly distinguishable from other hereditary giant platelet disorders. It would probably represent a novel platelet disorder.

Adult ; Blood Platelets ; metabolism ; ultrastructure ; Female ; Humans ; Integrin beta3 ; biosynthesis ; Microscopy, Immunoelectron ; Platelet Glycoprotein GPIb-IX Complex ; biosynthesis ; Platelet Membrane Glycoprotein IIb ; biosynthesis ; Thrombocytopenia ; genetics ; pathology

Objective To investigate the association between the level of polymorphism of APOE gene and cognitive impairment in patients with CNS demyelinating diseases. Methods 56 patients with central nervous system demyelinating disease were applied APOE genotyping,MoCA and expanded disability status (EDSS) scale score. Patients with MOCA scores <26 were divided into cognitive impairment group, and those with MOCA scores ≥26 were divided into normal cognitive preserved group. Results The probability of cognitive dysfunction in patients with central nervous system demyelinating diseases was 53.57%. There was no significant difference in age, gender, and disease duration between the CI group and the CP group(P>0.05), the difference in age and education among groups is statistically significant (P<0.05). There was no statistical significance in the difference in age, sex, education years and EDSS score between APOEε4 gene positive group and APOEε4 gene negative group (P<0.05). The difference of visual space and attention between different cognitive domains is statistically significant(P<0.05). Years of schooling is a risk factor for cognitive dysfunction in patients with central nervous system demyelinating disease(P<0.01). Conclusion The central nervous system demyelinating disease is impaired cognitive function. Patients with APOEε4 gene positive are more severely impaired in visual space and attention than patients with negative APOEε4 gene.Years of education are the risk factors of cognitive dysfunction in patients with central nervous system demyelinating disease. The course of disease and disabled function may not be significant related to cognitive impairment.