Objective To investigate the apoptosis response to mechanical stretch in human alveolar type Ⅱ epithelial cells A549.Methods First,A549 cells were given different stretch(0%,5%,15%,30%) at frequency of 0.5 Hz for 4 h.Then,A549 cells were stretched for different periods(0 h,2 h,4 h,8 h) at stretch 15%,0.5 Hz.Last,A549 cells were stretched for different frequency(0 Hz,0.2 Hz,0.5 Hz and 1 Hz) at stretch 15% for 4 h.The early apoptosis was detected by flow cytometry. Results A549 cells submitted to cyclic stretch caused the early apoptosis in a time-,stretch-and frequency-dependent manner.In details,at 0.5 Hz and 4 h stretch-dependence occurred,at 15% stretch and 0.5 Hz time-dependence occurred,and at 15% stretch and 4 h frequency-dependence occurred. Conclusion Mechanical stretch can induce the early apoptosis in human alveolar type Ⅱ epithelial cells A549,which may be one of the mechanisms of ventilator-induced lung injury.

Discharge against medical advice (DAMA) conflicts with the purpose of disease treatment in children. Some research has shown that there are high proportions of extremely preterm infants and infants with asphyxia or congenital malformation in neonates with DAMA. This suggests that the sustainable development of neonatology needs cooperation and co-development with obstetrics, neonatal surgery, and radiology to reduce the rate of DAMA. With reference to the current status of research in both China and other countries, this article reviews the causes for DAMA and the strategies for reducing the rate of DAMA, in order to provide a theoretical basis for effectively reducing the rate of DAMA from the neonatal intensive care unit, improving treatment outcomes of the neonates, and increasing hospitals' comprehensive benefits.
Ethics, Medical ; Health Services Needs and Demand ; Humans ; Infant, Newborn ; Insurance, Health ; Intensive Care Units, Neonatal ; Patient Discharge ; Prenatal Care ; Treatment Refusal

OBJECTIVETo investigate the contents of lead, cadmium, zinc and manganese in the follicular fluid and semen of infertile couples that are not professionally exposed to the four heavy metallic elements.

METHODSIn vitro fertilization pre-embryo transfer (IVF-ET) was carried out in wives, and follicular fluid collected after routine oocyte retrieval. Semen was obtained from husbands by masturbation. The contents of zinc in the follicular fluid and semen were determined by the flame atom absorption method and those of lead, cadmium and manganese by graphite furnace atomic absorption spectrometry.

RESULTSThe average concentrations of lead, cad- mium, zinc and manganese were 151.06 microg/L, 2.02 microg/L, 0.54 mg/L and 28.54 microg/L in the follicular fluid, and 250.23 microg/L, 7.42 microg/L, 189.11 mg/L and 82.15 microg/L in the semen. The follicular fluid samples in which lead, cadmium, zinc or manganese was detected constituted 43.8% (21/48), 22.9% (11/48), 75.0% (36/48) and 50.0% (24/48), and the semen samples accounted for 70.2% (33/47), 31.9% (15/47), 100.0% (47/47) and 46.8% (22/47), respectively.

CONCLUSIONThe results suggest that the average contents of lead, cadmium, zinc and manganese are higher in the semen than in the follicular fluid in the non-professionally exposed infertile couples, and so is the percentage of the samples containing each of the elements, with the exception of manganese.

Adult ; Cadmium ; analysis ; Environmental Exposure ; analysis ; Female ; Follicular Fluid ; chemistry ; Humans ; Infertility, Female ; metabolism ; pathology ; Infertility, Male ; metabolism ; pathology ; Lead ; analysis ; Male ; Manganese ; analysis ; Metals, Heavy ; analysis ; Semen ; chemistry ; Spectrophotometry, Atomic ; Zinc ; analysis

Acquired Immunodeficiency Syndrome ; Blood Donors ; China ; HIV Infections ; Humans ; Prejudice ; Rural Population

OBJECTIVETo construct the prokaryotic and eukaryotic expression vectors pET32/E5 and pcDNA3.1/E5 for transformation into E. coli BL21 and NIH(3)T(3) cells respectively to observe the expression of human papillomavirus type 16 E5 protein (HPV16 E5).

METHODSHPV16 E5 gene was amplified by PCR from clinical isolates of HPV 16 and inserted into the plasmid pET32a(+) followed by digestion with BamH I and Hind III. The recombinant plasmid pET32/E5 was transformed into E. coli JM109 and selected with ampicillin. The positive clones containing the recombinant plasmid pET32/E5 were verified by restriction endonucleases BamH I and Xho and sequence analysis. The expression of HPV16 E5-TRX fusion protein in E. coli BL21(ED3) was identified by SDS-PAGE and Western blotting. The digestion product of BamH I and Xho was purified and inserted into the eukaryotic expression vector pcDNA3.1(+) to construct pcDNA3.1/E5, which was identified by sequencing and transfected into NIH3T3 cells. The NIH(3)T(3) cells with stable expression of HPV16 E5 were selected by G418 and confirmed by RT-PCR.

RESULTSThe pET32/E5 and pcDNA3.1/E5 vectors were constructed successfully. E.coli BL21(DE3) transformed by the recombinant plasmid pET32/E5 expressed HPV16 E5-TRX fusion protein efficiently. In the presence of 1 mmol/L IPTG at 28 degrees C, HPV16 E5-TRX recombinant protein accounted for about 10% of the total bacterial proteins. NIH3T3 cells stably expressing HPV16 E5 were harvested by selection with 250 g/ml of G418. HPV16 E5 gene from pcDNA3.1/E5-transfected NIH(3)T(3) cells was amplified by RT-PCR, and sequence analysis demonstrated the acquisition of the full-length gene fragment.

CONCLUSIONSThe prokaryotic and eukaryotic vectors for the HPV16 E5 gene have been successfully constructed. The acquisition of E .coli and NIH(3)T(3) cells with stable expression of HPV16 E5 protein may facilitate subsequent research of the biological properties and the transformation mechanism of HPV16 E5 protein on specific cells.

3T3 Cells ; Animals ; Escherichia coli ; genetics ; metabolism ; Eukaryotic Cells ; metabolism ; Genetic Vectors ; Humans ; Mice ; Oncogene Proteins, Viral ; biosynthesis ; genetics ; Papillomaviridae ; genetics ; isolation & purification ; Papillomavirus Infections ; virology ; Plasmids ; genetics ; Prokaryotic Cells ; metabolism ; Recombinant Fusion Proteins ; biosynthesis ; genetics

OBJECTIVETo evaluate clinical therapeutic effect of post-stroke shoulder pain treated by acupuncture combined with Tuina.

METHODSThree hundred cases of post-stroke shoulder pain were randomly divided into an acupuncture and Tuina group and a rehabilitation group by double-center randomized controlled clinical trial method. In acupuncture and Tuina group, normalized electroacupuncture and Tuina therapy were applied, that was electroacupuncture at main points, such as Chize (LU 5), Quze (PC 3), Shaohai (HT 3), Jianyu (LI 15), Jianliao (TE 14) and Jianjing (GB 21),etc., combined with traditional Tuina manipulations; in rehabilitation group, the rehabilitation methods such as the electrostimulation through nervus cutaneus and the squeezing and stabilizing manipulations of Proprioceptive Neuromuscular Facilitation (PNF), etc. were applied. The treatment courses of both groups were 6 weeks. The main therapeutic effect indices were the Assessment Face Scale (AFS) for pain when shoulder was in passive motion and the Fugl-Meyer Motor Assessment for upper limbs active function; the secondary indices were the moditied Rankin Scale (mRS) and the clinical incidences of shoulder-hand syndrome of hemiplegia and shoulder joint subluxation of hemiplegia.

RESULTSAfter 6 weeks treatment and 12 weeks follow-up, AFS score, Fugl-Meyer motor assessment of upper limbs active function and mRS evaluation in acupuncture and Tuina group were more obviously improved than those in rehabilitation group (P < 0.05, P < 0.01). Although the clinical incidences of shoulder-hand syndrome of hemiplegia and shoulder joint subluxation of hemiplegia in acupuncture and Tuina group was equal to those in rehabilitation group [3.55% (5/141) vs 8.45% (12/142), 1.42% (2/141) vs 5.63% (8/142), both P > 0.05], the data indicated that there was a superiority tendency in acupuncture and Tuina group.

CONCLUSIONThe combined therapy of electroacupuncture and Tuina is a normative manipulation, and the therapeutic effect is satisfying for post-stroke shoulder pain, superior to that of comprehensive rehabilitation treatment.

Aged ; Amobarbital ; Combined Modality Therapy ; Drug Combinations ; Electroacupuncture ; Female ; Humans ; Male ; Middle Aged ; Range of Motion, Articular ; Secobarbital ; Shoulder Pain ; etiology ; physiopathology ; therapy ; Stroke ; complications

OBJECTIVEDue to its minimal-invasive approach, endovascular procedure had replaced surgery in treating Budd-Chiari syndrome (BCS). The interventional therapy was a safe and effective treatment in adults with BCS and the cure rate was high. However Budd-Chiari syndrome in children and adolescents is rare. Published literature on interventional procedure for Budd-Chiari syndrome in children and adolescents is scarce. The aim of the study was to present results of percutaneous transluminal angioplasty (PTA) and stents placement in children and adolescents with BCS and to evaluate the efficacy and safety in these patients of this approach.

METHODTwenty-five patients [16 boys and 9 girls; average age of (14.5 ± 3.4) years old; age ranged from 5 to 17 years] with Budd-Chiari syndrome who were hospitalized from December 1990 to August 2012 were presented. All of them were diagnosed by color Doppler ultrasound scan while 12 of them had magnetic resonance venography (MRV) scan. All of the patients had undergone angiographic examination. Four cases with membranous obstruction of the inferior vena cava (IVC) were treated with PTA. One case with segmental block of IVC was treated with PTA and stent placement. Five cases with membranous obstruction of IVC and hepatic vein (/and accessory hepatic vein) were treated with PTA. Among 8 cases with membranous obstruction of hepatic veins, 6 cases were treated with PTA and the others with PTA and stent placement. Among 4 cases with blocks of 3 hepatic veins (HVs), one was treated with PTA, one with PTA plus catheter thrombolysis plus PTA, one with PTA and stent placement and the other one was unsuccessful. Three cases with obstruction of HV and accessory HV (AHV) were treated with PTA. Totally, 24 patients were treated with interventional approach and followed up.

RESULTThe procedure was successful in 24 patients. The involved veins (hepatic veins or IVC) were patented after interventional procedure. The pressure of hepatic vein was (42.1 ± 4.2) cm H2O (37-50 cm H2O) (1 cm H2O = 0.098 kPa) before the interventional therapy, while it was (17.3 ± 3.3) cm H2O (14-26 cm H2O) after it. The pressure of IVC was (30.6 ± 2.9) cm H2O (26-36 cm H2O) before the interventional therapy, while it was (18.8 ± 4.2) cm H2O (15-26 cm H2O) after it. The symptoms and signs vanished instantly after interventional procedure. There were no procedure-related complications. The rate of overall initial cure was 96%. The patients were followed up for a mean of 25.8 months (range 6 months to 8 years). Seven cases developed restenosis after first procedure. Five of them were treated with PTA, one with PTA plus catheter thrombolysis plus PTA, one with PTA and stent placement. All of the involved veins were patented again. Clinical symptoms were relieved. There were no procedure-related complications as well.

CONCLUSIONThe interventional procedure in children and adolescents with BCS is the same as in adults. Radiological therapeutic intervention is efficacious and safe in children and adolescents with BCS.

Adolescent ; Angioplasty ; Budd-Chiari Syndrome ; diagnostic imaging ; surgery ; therapy ; Catheterization, Peripheral ; Child ; Child, Preschool ; Female ; Follow-Up Studies ; Hepatic Veins ; diagnostic imaging ; surgery ; Humans ; Liver ; blood supply ; diagnostic imaging ; Male ; Phlebography ; methods ; Radiography, Interventional ; Retrospective Studies ; Stents ; Thrombolytic Therapy ; Treatment Outcome ; Vena Cava, Inferior ; diagnostic imaging ; surgery ; Venous Thrombosis ; therapy

OBJECTIVETo clone the urea membrane channel gene (ureI) from Helicobacter pylori (Hp) for its expression in E. coli, and evaluate the expression conditions and immunological features of the fusion protein.

METHODSureI gene cloned by PCR from Hp was inserted into the plasmid pET32a (+) to construct the recombinant plasmid pET32a/ureI, followed by identification by BglII and HindIII digestion and sequencing. E. coli BL-21+(DE3) was transformed with pET32a/ureI to obtain the engineered bacterium BL21+/UreI, which was cultured at different temperatures and induced with 1.0 mmol/L IPTG for expression of the recombinant protein. The expressed proteins were identified by SDS-PAGE and analyzed by Pro-gel analyzer 4.0. Western blotting was performed to evaluate the immunogenicity of the expressed protein.

RESULTSThe cloned gene fragment was about 650 bp in length, and BglII and HindIII digestion of pET32a/ureI yielded a 650-bp band. Sequence analysis revealed that the cloned ureI gene contained 646 bp without reading frame alterations. Comparison against GenBank indicated a homology of 100% of the cloned gene with ureI gene of the corresponding Hp strains, and also one no less than 98.5% with ureI gene from other strains. The engineered E. coli BL21+/UreI could express recombinant UreI (rUreI) with His tag, and the target protein accounted for 20.2% of the total bacterial proteins after 1.0 mmol/L IPTG induction of the bacterium at 37 degrees C for 14 h. SDS-PAGE and Western blotting showed that the recombinant UreI protein was produced mainly in the inclusion bodies and fused with his-tag (rUreI/his), which could react with human anti-Hp and mAb to his tag but not with mAb to Hp UreB.

CONCLUSIONSWe have successfully cloned ureI gene and constructed the prokaryotic expression plasmid for efficient rUreI expression, and the fusion protein rUreI/his expressed in the inclusion bodies can react specifically with both Hp antibody and his-tag antibody.

Bacterial Proteins ; genetics ; metabolism ; Blotting, Western ; Cloning, Molecular ; Electrophoresis, Polyacrylamide Gel ; Escherichia coli ; genetics ; Helicobacter pylori ; genetics ; Humans ; Membrane Transport Proteins ; genetics ; metabolism ; Plasmids ; genetics ; Recombinant Fusion Proteins ; genetics ; isolation & purification ; metabolism

OBJECTIVETo investigate the role of Snitrosylation of protein disulphide isomerasec in methamphetamine (METH)-induced expression of alpha synuclein (αSN) in mouse hippocampus and striatum neurons.

METHODSForty C57BL/6 mice were randomized equally into saline control group, METH group, L-NNA (a NOS inhibitor) group and L-NNA plus METH group. All the agents were injected intraperitoneally at an interval of 12 h, and a total of 8 injections were administered; in L-NNA plus METH group, METH was injected 30 min after LNNA in each treatment. Western Blotting was used to detect the expression of nitric oxide synthase (NOS), αSN, PDI and Snitrosylation of protein disulphide isomerase (PDI-SNO) in the hippocampus and striatum of the mice, and nitric oxide (NO) levels were determined using a NO assay kit.

RESULTSIn METH group, the levels of NOS, PDISNO, αSN and NO all increased significantly compared with those in the control group (P<0.05). Combined treatment with L-NNA and METH, compared with METH alone, resulted in significantly lowered expression of NOS, NO, PDI-SNO and αSN in the hippocampus and striatum of the mice (all P<0.05). No significant differences were found in NOS, NO, PDI-SNO or αSN expressions among METH+L-NNA, L-NNA and control groups (P>0.05).

CONCLUSIONMETH induces the activation of NOS and increases NO level to cause the occurrence of PDI-SNO, leading subsequently to increased expression of αSN in mouse striatum and hippocampus. L-NNA, the inhibitor of NOS, can partly relieve nervous system toxicity induced by METH.

OBJECTIVETo evaluate the prevalence and distribution of C-kit, NPM1 and FLT3 gene mutations in patients with acute myeloid leukemia (AML), and to analyze the relationship between the gene mutations and their prognosis.

METHODSMutations in exon 8 and 17 of C-kit gene, exon 12 of NPM1 gene, exon 20 of FLT3-TKD gene, and exon 14/15 of FLT3-ITD gene were detected by direct sequencing. Clinical data was collected and followed up if the patient had accepted treatment in our hospital.

RESULTSAmong the 656 AML patients, mutations in C-kit exon 8 were found in 6 patients (0.9%), C-kit exon 17 in 33 (5.0%), NPM1 in 169 (25.8%), FLT3-TKD in 46 (7.1%), and FLT3-ITD in 178 (27.1%). Six subtypes of mutations were detected in C-kit exon 8, 8 in C-kit exon 17, 11 in FLT3-TKD, 15 in NPM1, of which 5 were not reported before. C-kit exon 17 mutations were more frequently detected in patients with t(8;21) and exon 8 in patients with inv(16) cytogenetic abnormality. No other gene mutations except FLT3 were detected in M(3) patients. NPM1 and ITD mutations were often detected in individuals with normal cytogenetics or M(5) and M(1) of FAB classification, and accompanied with high white blood cell counts in peripheral blood, high blast counts in bone marrow and low CD34 expression. The older the patients were when diagnosed, the more gene mutations and the higher white blood cell count were detected. More mutations were found in individuals with normal karyotype than that with other karyotypes. It appeared that FLT3-ITD was significantly associated with shorter overall survival (OS) (P = 0.004), NPM1 was not significantly associated with OS, but NPM1(+)/ITD(-) patients had the longest OS.

CONCLUSIONSOur results showed that the mutation types and amounts had particular distribution in MICM subtypes, and were associated with white blood cell counts in peripheral blood, blast counts in bone marrow and prognosis. Especially for patients with normal karyotype, the genetic mutations could be new molecule marker.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Asian Continental Ancestry Group ; genetics ; DNA Mutational Analysis ; Female ; Humans ; Karyotyping ; Leukemia, Myeloid, Acute ; diagnosis ; genetics ; Male ; Middle Aged ; Mutation ; Nuclear Proteins ; genetics ; Prognosis ; Proto-Oncogene Proteins c-kit ; genetics ; Young Adult ; fms-Like Tyrosine Kinase 3 ; genetics