This study was aimed to construct a pEGFP-N1 vector of von Willebrand factor cleaving protease (ADAMTS13, a disintegrin and metalloprotease with a thrombospondin type 1 motifs 13) so as to pave the way for further studying its synthesis and secretion. Human full-length cDNA sequence of ADAMTS13 was acquired by polymerase chain reaction (PCR) with Phusion(®) High-Fidelity (NEB), then the PCR product was double digested with EcoRI and XhoI. After digestion, the ADAMTS13 cDNA sequence was purified and recombined with the pEGFP-N1 vector. The DNA sequence analysis showed that ADAMTS13 was ligated to the pEGFP-N1 vector correctly. After transient expression in HeLa cells, the expression of EGFP could be detected by fluorescent microscopy, and the expression of ADAMTS13 protein could be detected by SDS-PAGE and Western blot. It is concluded that the ADAMTS13-pEGFP-N1 vector is successfully constructed, and it can be widely used in further research on the mechanism of the synthesis and secretion of ADAMTS13.
ADAM Proteins ; genetics ; ADAMTS13 Protein ; Gene Expression ; Genetic Vectors ; Green Fluorescent Proteins ; genetics ; HeLa Cells ; Humans ; Transfection

Objective To understand the patients of congenital microtia malformation families knowledge of skin expander and influencing factors. Methods Self-made questionnaire to sample survey of 500 cases of our department (Plastic Surgery Hospital, Chinese Academy of Medical Sciences, the second microtia concer) patients′ families. Results 47.8％(239/500) of 500 patients of expander knowledge level is high, 41.2％(206/500) pass the exam, 11.0％(55/500) fall the exam, only 13.4％(67/500) really have a comprehensive understanding on expander achieve excellent. Scores of male and female were (16.06 ± 1.99) points and (16.39 ± 2.16) points, t = 1.752, P > 0.05, there was no statistically significant difference comparing the 2 group. Patients′ families score of different cultural levels, respectively (14.06 ± 2.36), (14.98 ± 2.02), (16.54 ± 2.00), (16.73 ± 1.88) points, F = 21.736, P < 0.01, difference of four groups was statistically significant. Different age patients′families score ( 16.21 ± 1.96), (16.62 ± 2.14), (14.86 ± 2.11), (13.98 ± 2.02), (13.73 ± 1.88) points, F = 15.685, P > 0.05, there was no statistically significant difference comparing the 5 groups. Patients with different professional families score (13.25 ± 2.19), (13.79±2.27), (16.08±1.89), (14.10±2.08), (14.13±2.35), (14.45±2.09), (14.56±1.75), (16.84± 1.81) points, F = 2.737, P < 0.01, difference of eight groups was statistically significant. Conclusions Congenital microtia patients′families skin expander knowledge needs to be improved, it is necessary to take various forms, conduct for families of expander knowledge through propaganda and education.

Animals ; Aorta ; pathology ; Atherosclerosis ; blood ; etiology ; metabolism ; Chemokine CCL2 ; metabolism ; Cholesterol ; blood ; Cholesterol, Dietary ; administration & dosage ; Cholesterol, HDL ; blood ; Cholesterol, LDL ; blood ; Dehydroepiandrosterone ; pharmacology ; Diet, Atherogenic ; Immunohistochemistry ; Rabbits ; Random Allocation ; Triglycerides ; blood ; Vascular Cell Adhesion Molecule-1 ; metabolism

Full-length NS5A gene of the hepatitis C virus was amplified by PCR using plasmid pBAC25 containing HCV nonstructural gene as template. The amplified fragment (about 1.34 kb) was cloned into plasmid pQE32, and the recombinant plasmid pQENS5A was expressed in JM109 strain. The NS5A protein was purified by NiSO4 metal chelating resin, and characterized by Western-blot. Its antigenecity was determined by ELISA. The positive detection rate of anti-NS5A was 75% (69/92) in ninety-two clinic sera. The positive rate of anti-NS5A was 82.5% (33/40) in fourty positive standand sera, and the negative rate of anti-NS5A was 100% (40/40) in fourty negative standand sera. The results showed that the Full-length NS5A proteinn had the higher sensitivity and specificity in the detection of HCV antibody in sera, we suggested that NS5A protein was a useful antigen for blood screening.

This study aimed to explore Semaphrin4D (Sema4D) expression and clinical significance in non-small cell lung cancer (NSCLC), and to define the roles and mechanisms of Sema4D in regulating the malignant behaviors of A549 cells by small interfering RNA (siRNA). Firstly, immunohistochemistry revealed that Sema4D was more frequently expressed in NSCLC than in lung benign lesion (P<0.05) and its overexprssion was associated with low differentiation (P<0.05), poor pTNM staging (P<0.05) and occurrence of lymph node (LN) metastasis (P<0.05). Endogenous Sema4D expression was suppressed by Sema4D siRNA in A549 cells overexpressing Sema4D. Protein levels of Sema4D, total Akt and p-Akt were examined by Western blotting. Cell proliferation, migration and invasion abilities were measured by MTT assay and Transwell assay respectively. Results showed that Sema4D siRNA significantly suppressed phosphorylation of AKT in A549 cells, but it did not alter total AKT expression. In addition, efficient down-regulation of SemaD significantly inhibit cell proliferation (P<0.05), migration (P<0.05) and invasion (P<0.05) in A549 cells. These findings suggest that Sema4D might serve as a reliable tool for early prediction of NSCLC poor prognosis. Sema4D could play an important role in promoting tumor proliferation, migration and metastasis in the NSCLC, by influencing the Akt protein phosphorylation. Inhibition of Sema4D may be a useful approach for the treatment of NSCLC.

[Objective]To investigate the effects of continuous long-term sleep deprivation on rat's behavior,memory and neurotransmitters in hippocampus,as well as determine the effects of prevention of electro-acupuncture.[Method]We used multi-platform water environment to make the rat model of insomnia.The experiment was divided into two sections.Sec-tion one:thirty rats were divided into 5 groups(six rats each group):tank control group(TC),sleep-deprived for continu-ous 7 days group(SD group),three days of recovery after SD group(3d after SD group),six days of recovery after SD group (6d after SD group),eleven days of recovery after SD group(11d after SD group).The term of recovery refers to normal feed-ing and sleeping when rats return to cages after sleep-deprivation.The open-field test was performed following the end time of each group experiment and then the rats were perfused and fixed.Section two:Eighteen rats were divided into three groups (six rats each group):tank control group(TC),sleep-deprived for continuous 7 days group(SD group),sleep-deprived rats treated with electro-acupuncture group(SD+EA group).Rats of SD group and SD+EA group were deprived sleep for con-tinuous 7 days. During the period of sleep deprivation,the rats of SD+EA group were treated with electro-acupuncture 20 minutes every morning.At the end time of experiment,rats were tested by the Morris water maze and then perfused and get specimens.[Results]Section one:the statistical analysis of outcome of open-field test showed:as compared to the TC group,the scores of rat's horizontal movement and vertical movement were significantly fewer in the SD group(P<0.05).Af-ter 3,6,or 11 days recovery of sleep,the scores increased significantly as compared with SD group(P<0.05). Although, the outcome of open-field test in the 3d after SD and 6d after SD groups had the significant difference with TC group(P<0.05),there was no different between the 11d after SD group and TC group(P>0.05).The quantitative analysis of gamma aminobutyric acid(GABA)expression in the hippocampus showed:the SD group shows lower level of GABA when com-pared to the TC group(P<0.05).After 3 days or 6 days of recovery,the expression of GABA is higher than the SD group(P<0.05),and similar to the TC group(P>0.05).In the 11days after SD group,the expression of GABA declined as compared to the TC group and the 6d after SD group(P<0.05).Section two:In the Morris water maze experiment,when compared to the TC group and the SD+EA group,the escape latency period was significantly prolonged and the times of across the plat-form were significantly reduced in the SD group(P<0.05).On the contrary,there were no statistical differences in the escape latency period or the across platform times between the SD+EA group and the TC group(P>0.05).The results of serotonin(5-HT)and GABA in the hippocampus showed:the 5-HT level in the SD group was no significant difference as compared to the TC group and the SD group on 5-HT level,while the 5-HT level in the SD+EA group was reduced as compared to the TC group(P<0.05). In the opposite,the GABA level in the SD group was decreased as compared to the TC group(P<0.05),while the SD+EA group and the TC group appear similarly level(P>0.05).[Conclusions]The effect of 7 days'contin-uous long-term sleep-deprivation may lead to the reduction in the ability of learning and memory behavior of rats.Our results is different with 4 days'continuously sleep-deprivation which was reported on the rats'expression of GABA and 5-HT. Electro-acupuncture stimulating the Sishengcong Acupoints seems preventing the harming of long-term sleep-deprivation on the ability of learning and memory,which probably is through adjusting the disorder of central neurotransmitter expression caused by sleep deprivation.

OBJECTIVETo explore the operative methods and effects of treatment of old acetabular fractures.

METHODSFrom October 2001 to October 2007, 26 patients with old acetabular fractures were treated with operation including 21 males and 5 females with an average age of 34 years ranging from 18 to 65 years. On the basis of the three-dimensional computed tomography, all cases were diagnosed and classified according to Letourne-Judet classification, 9 cases were posterior wall fracture, 3 cases were lateropulsition fracture, 7 cases were lateropulsition and posterior wall fracture, 2 cases were posterior column and posterior wall fracture, 2 were T-shape fracture, 3 were dual column fracture. These patients were treated through the anterior,posterior, combined anterior-posterior approaches. The time from injured to operation was 33 to 141 days (averaged 36.4 days). All the fractures were fixed with screws and AO reconstruction plates.

RESULTSAll patients were followed up for 6 to 96 months, with an average time of 32.4 months. Evaluated according to Matta criteria, the results of scores was (5.04 +/- 1.04) on pain, (5.23 +/- 0.76) on range of motion, (4.92 +/- 1.16) on walking,and tatal (5.06 +/- 0.99) on average; The functional results of hip joints were excellent in 6 cases, good in 10 cases, fair in 6 cases, and poor in 4 cases. Sciatic nerve injury was found in 2 patients,lateral femoral cutaneus nerve injury in 3 patients, necrosis of femoral head in 1 patient,infection in 1 patient, and ectopic bone formation in 6 patients.

CONCLUSIONGood clinical results can be obtained by careful selection of operative indications of old acetabular fractures in combination with proper operative approach and correct reduction and fixation.

Acetabulum ; injuries ; surgery ; Adolescent ; Adult ; Aged ; Female ; Follow-Up Studies ; Fractures, Bone ; surgery ; Humans ; Male ; Middle Aged ; Postoperative Complications ; Time Factors ; Young Adult

To study the chemical constituents of Abacopteris penangiana (Hook.) Ching, various chromatographic techniques were used to isolate and purify the constituents. The structures of the obtained compounds were elucidated by spectroscopic data and physical-chemical properties. Two compounds were isolated from the n-BuOH soluble fraction of an acetone-H2O (4:1) extract of A. penangiana and were identified as 4'-hydroxy pneumatopterin B (I) and 6"-O-acetyl triphyllin A (II). Compounds I and II are new compounds.
Ferns ; chemistry ; Flavonoids ; chemistry ; isolation & purification ; Glycosides ; chemistry ; isolation & purification ; Molecular Structure ; Plant Components, Aerial ; chemistry ; Plants, Medicinal ; chemistry

OBJECTIVETo isolate and identify the high-yield poly-malic acid (PMLA) bacterial strains from the nature.

METHODSSamples were collected and cultured. The high-yield PMLA bacterial strains were screened through morphological observation, qualitative PMLA tests by HPLC and ITS sequence analysis on the isolated bacterial strains.

RESULTSA high-yield PMLA strain II 04 was isolated, the yield of PMLA of the strain reached to 26.23g/L in the rotary shaker at 25 degree for 7d. From morphological observation and ITS sequences analysis, the strain belonged to Aureobasidium pullulans, and named as Aureobasidium pullulans ZUCC-41.

CONCLUSIONA high-yield bacterial strain has been isolated from the nature and identified to be Aureobasidium pullulans.

Fermentation ; Malates ; metabolism ; Mitosporic Fungi ; isolation & purification ; metabolism ; Polymers ; metabolism

OBJECTIVETo set up animal models of the lung cancer induced by Yunnan tin mineral dusts (no radon) in F344 rats and to explore the process of carcinogenesis and pathologic alterations in various stages of malignant transformation in the animal models.

METHODSOne hundred and ninety F344 rats were randomly divided into Yunnan tin mineral dust group (100 rats), furfural physiological saline group (30 rats), physiological saline group (30 rats) and normal control group (30 rats). The intratracheal instillation with mass fraction of 6% suspension liquid mixture Yunnan tin mineral dusts, volume fraction of 2% furfural physiological saline and physiological saline 0.2 ml was performed in the rates once per week respectively except normal control group. Then the rats were sacrificed in batch periodically after one week. The last rat was exposed to the tin mine dusts for 100 weeks. The morphological process and tumor formation were dynamically observed under LM and TEM. Immunohistochemistry detection of cytokeratin of High MW and low MW was used for tumor classification. Pollak stein was used to evaluate the development of fibrosis of lung in the rats.

RESULTSBronchoalveolar inflammation occurred in the early stage after the intratracheal instillation of Yunnan tin mineral dust was performed in F344 rates. Along with reduction of inflammation, collagen fibrils increased at alveolar interstices. Simple hyperplasia, papillary hyperplasia and metaplasia of the epithelial cells in alveolar and bronchi were observed, followed by atypical adenomatous hyperplasia and squamous dysplasia. Lung cancer was induced in the end. Among the 14 cases of lung cancer, 9 cases were adenocarcinoma, 2 squamous cell carcinoma and 3 mixed carcinoma. No lung cancer occurred in other three control groups. There was a significant difference in the malignancy rate between the experimental group and the three control groups (P < 0.01). The squamous metaplasia and squamous carcinoma were found in alveoli that expressed cytokeratin of High MW. Lung fibrosis was found in 31 cases of in the tin mineral dust group. The greater the mineral dust deposit was, the more serious the alveolar fibrosis was.

CONCLUSIONYunnan tin mineral dusts without radon induce lung cancer in rates. The adenocarcinoma and squamous carcinomas induced in F344 rat lung can occur in the alveoli. The further study on whether type II alveolar epithelial cells are the origin cells of adenocarcinoma and some peripheral squamous lung carcinomas is worthwhile.

Animals ; Disease Models, Animal ; Dust ; Female ; Lung ; drug effects ; pathology ; Lung Neoplasms ; chemically induced ; pathology ; Male ; Rats ; Rats, Inbred F344 ; Tin ; adverse effects