OBJECTIVETo explore the effect of Lignum Sappan (LS) containing serum on the proliferation cycle arrest of human lung cancer cell line PG and its molecular mechanism.

METHODSThe lung cancer PG cells were divided into four groups, i.e., the blank control group, the LS group, the LS plus cisplatin group, and the cisplatin group. They were cultured by RPMI-1640 with 20% blank serum, RPMI-1640 with 20% LS containing serum, RPMI-1640 with 20% LS containing serum plus 1 microg/mL cisplatin, and RPMI-1640 with 20% blank serum plus 1 microg/mL cisplatin, respectively. The morphology of PG cells was observed using light microscope and laser scanning confocal microscope in each group. The cell cycle arrest was observed using flow cytometry. The expression of P16 and Rb1 mRNA was tested by PCR method.

RESULTSUnder the light microscope and laser scanning confocal microscope, the apoptosis degree of PG cells in the LS group was significant, but less than that of the LS plus cisplatin group as well as the cisplatin group. Compared with the blank control group, the proportion of PG cells increased at G0/ G1 and S phases (P < 0.05) and decreased at G2/M phase (P < 0.01) in the LS group; The proportion of PG cells increased at G2/M and S phases (P < 0.05, P < 0.01) and decreased at G0/G1 phase (P < 0.01) in the LS plus cisplatin group as well as the cisplatin group. Compared with the LS group, the proportion of PG cells increased at G2/M and S phases (P < 0.05, P < 0.01) and decreased at G0/G1 phase (P < 0.01) in the LS plus cisplatin group as well as the cisplatin group. There was no statistical difference in PG cells at each phase between the cisplatin group and the LS plus cisplatin group (P > 0.05). The expression of P16 and Rb1 mRNA increased in the LS group, when compared with the blank control group. They also increased in the cisplatin group and the LS plus cisplatin group, higher than that of the LS group (P < 0.05). There was no statistical difference in the expression of P16 and Rb1 mRNA between the cisplatin group and the LS plus cisplatin group (P > 0.05).

CONCLUSIONLS containing serum induced PG cell apoptosis by up-regulating the mRNA transcription levels of P16 and Rb1, thus resulting in PG cell arrest at G0/G1 and S phases, which was different from the manner of cisplatin (achieved by arresting PG cells at G2/M and S phases through regulating cyclinB1 mRNA transcription).

Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Cisplatin ; pharmacology ; Drugs, Chinese Herbal ; pharmacology ; Humans ; Lung Neoplasms ; pathology

Objective To prepare the standard molecular weight fragment mixtures. Methods Primers were designed to prepare clones which contained different sizes of standard molecular weight fragments. The template used for amplification of insert fragments was the pMD18-T vector. Bacteria culture and plasmid extraction were used to obtain abundant target fragment. Unlabeled DNA fragments were prepared by double digestion of the recombinant plasmids, and the fluorescent adaptor was prepared by annealing with two partial reverse complimentary DNA fragments. The unlabeled fragments and fluorescent adaptor were connected by DNA ligation reaction assisted with T4 DNA ligase. In this way, different sizes of standard molecular weight fragments were prepared. Standard molecular weight fragment mixture was finally prepared by mixing all the fragments together before purification. Results Ten standard molecular weight fragments of different sizes were prepared. The sizes of each fragment are 80bp, 124bp, 194bp, 224bp, 254bp, 304bp, 349bp, 399bp, 424bp and 454bp. The internal standard could accurately determine the size of PCR products amplified with the DNATyper15 kit. Conclusion Using this method, the standard molecular weight fragment mixture which meet the requirements of research and laboratory use was prepared, perfectly providing a new method for preparation of the DNA molecular weight standards. The peaks and the size of the prepared DNA internal lane standard are correct, which can be used to calculate the DNA fragments size in capillary electrophoresis.

Objective To analyze the results of the Objective Structured Clinical Examination(OSCE) of medical interns in Department of Gynecology and Obstetrics,and to assess the role of OSCE in clinical teaching and practice. MethodsAtolal of 104 medical interns of five-year education program in 2002 and seven-year education program in 2003 in School of Medicine had taken part in OSCE of gynecology and obstetrics.The OSCE consisted of six stations: gynecologic examination,obstetric examination,oral test,interrogation of standardized-patients(SP),drawing partogram,and non-stress test(NST) explanation.After gathering the data of each station,the software of SPSS 11.0 was used to make statistical analysis. Results The median scores were as follows: 92.0 for gynecologic examination,91.0 for obstetric examination,83.5 for oral test,80.0 for interrogation of SP,80.0 for drawing partogram,and 70.0 for NST explanation.There was no significant impact of familiarity to OSCE on the results of the test.The results showed that medical interns did good job in basic performance,but lack of clinical practice skills in some complicated practice of gynecology and obstetrics.Conclusion The grading system and stations setup of OSCE in Department of Gynecology and Obstetrics in Shanghai Sixth People's Hospital show objectivity and equity.The OSCE can be widely applied in the after-department examination and even graduation examination.

The mutations in the dual oxidase 2 (DUOX2) and dual oxidase maturation factor 2 (DUOXA2) genes can cause congenital hypothyroidism (CH). This study reports the pedigree with goitrous congenital hypothyroidism (GCH) due to the coexistence of heterozygous mutations in the DUOX2 and DUOXA2 genes. The two sisters with GCH were diagnosed with CH at neonatal screening and were enrolled in this study. The DUOX2, DUOXA2, and thyroid peroxidase (TPO) genes were considered for genetic defects screening. Family members of the patients and normal controls were also enrolled and evaluated. The two girls harbored compound heterozygous mutations, including a new mutation of c.2654G>T (p.R885L) in the maternal DUOX2 allele and c.738C>G (p.Y246X) in the paternal DUOXA2 allele, that has been previously reported. The germline mutations from the families were consistent with an autosomal recessive inheritance pattern. No mutations in the TPO gene and the controls were observed.
Alleles ; Congenital Hypothyroidism* ; Female ; Germ-Line Mutation ; Humans ; Infant, Newborn ; Inheritance Patterns ; Iodide Peroxidase ; Mass Screening ; Neonatal Screening ; Oxidoreductases ; Pedigree ; Siblings

OBJECTIVETo analyze and evaluate the fatigue of shoulder skeletal muscle caused by different lifting loads with surface electromyography (sEMG).

METHODSAccording to the loading standard of1 Repetition Maximum (1RM), ten male volunteers performed 3 tasks of upper limb flexion, i.e. 10%, 50% and 90%-1RM. During action process, the signals of Upper Trapezius (UT), Lower Trapezius (LT), Serratus Anterior (SA) and Anterior Deltoid (AD) were recorded by sEMG. The Mean Amplitude (MA) served as an index to evaluate the changes in skeletal muscle fatigue.

RESULTSThe scores of Borg were 15.6, 15.9 and 15.2 for 3 loads of 10%-1RM, 50%-1RM and 90%-1RM, respectively (P > 0.05). The mean amplitudes (MAs) of Upper Trapezius, Lower Trapezius, Anterior Deltoid and Serratus Anterior in shoulders increased obviously. Under the load intensity of 10%-1RM, the MAs of Upper Trapezius and Anterior Deltoid increased significantly (P < 0.05), which were 0.898 and 0.736, respectively. After the exhaustion, the contribution of mean amplitude in shoulder muscle did not change significantly (P > 0.05).

CONCLUSIONThe low-load action for long time can induce easily the fatigue of upper trapezius and anterior deltoid.

Adult ; Electromyography ; Humans ; Lifting ; Male ; Muscle Fatigue ; physiology ; Muscle, Skeletal ; physiology ; Shoulder ; physiology ; Workload

Tilianin was separated and authenticated from the seeds of Dracocephalum moldavia, a Uygur medicine, by chromatographic technique and spectroscopic method. The purity of tilianin is more than 98% determined by HPLC area normalization method. Thin layer chromatography (TLC) method was used to separate tilianin from D. moldavia by mixture of chloroform-methanol (5: 1) as a developing solvent on high performance silicagel precoated plate (SGF254) and using aluminium trichloride as a chromogenic agent for qualitative identification of D. moldavia. To establish a HPLC method for quantitative analysis of D. moldavia, tilianin was used as a Quantitative marker and separated on a C18 (4.6 mm x 250 mm, 5 μm) column with acetonitrile-01% formic acid (25: 75) as the mobile phase and detected at 330 nm. The calibration curve of tilianin displayed ideal linearity over the range of 0.617 2-123.44 μg x mL(-1) with a regression equation of Y = 33.773X - 0.824 8 (r = 1). The average recovery of tilianin was 101.0% with RSD of 3.7%. The RSD values of intra-day and inter-day precision were less than 2%. The content of tilianin in 4 batches of the authenticated semen of D. Moldavia was between 0.016 and 0.187 mg x g(-1). The qualitative and quantitative method established is suitable for the quality evaluation and assessment of semen of D. Moldavia.
Chromatography, High Pressure Liquid ; Chromatography, Thin Layer ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; standards ; Flavonoids ; chemistry ; isolation & purification ; standards ; Glycosides ; chemistry ; isolation & purification ; standards ; Lamiaceae ; chemistry ; Magnetic Resonance Spectroscopy ; Quality Control

Objective:To investigate the influence of metastasis suppressor gene KAI1 on the proliferation,invasion and metastasis of endometrial carcinoma cell line AN3CA and HEC-1-B.Methods:The KAI1 cDNA was transfected into human endometrial carcinoma cells AN3CA and HEC-1-B via Lipofectamine 2000.The expression of KAI1 protein was ex- amined by Western blotting and flow cytometry before and after transfection.The proliferation ability of AN3CA and HEC- 1-B cells was observed by MTT assay and anchorage-independent growth assay.The changes of cell invasive ability were studied by transwell assays.Results:Stable expression of KAI1 protein was observed in AN3CA and HEC-1-B cells and on their surface after transfection with pcDNA3-KAI1 plasmid.Cells transfected with blank plasmid formed more colonies and had a larger size,with the colony forming rates being(54.2?3.1)% for AN3CA cells and(52.7?4.3)% for HEC- 1- B cells;the doubling time of AN3CA and HEC-1-B cells were 21.3 h and 20.1 h,respectively.Cells transfected with pcDNA3-KAI1 formed less colonies and had a smaller size,with the colony forming rates being(37.4?5.1)% for AN3CA cells and(32.1?3.7)% for HEC-1-B cells;the doubling time of AN3CA and HEC-1-B cells were 43.7h and 45.2 h,respectively.The cell proliferation abilities and colony-forming ability were significantly different between the two groups(P

BACKGROUNDThe optimal endpoint for catheter ablation of persistent atrial fibrillation (AF) remains ambiguous. This study investigated the impact of AF termination as a procedural endpoint and the termination mode on long-term clinical outcome.

METHODSTwo hundred and ninety-three patients who underwent stepwise ablation for persistent AF were categorized into the AF termination by ablation group and into the electrical cardioversion (CV) group. Subgroups were also analyzed based on different termination modes. Follow-up assessment included early recurrence and sinus rhythm (SR) maintenance.

RESULTSDuring initial ablation, 33 patients (11.3%) were directly converted to SR, 166 patients (56.7%) were converted to atrial tachycardia (AT) that subsequently restored SR with further ablation in 98 patients (33.4%), and a total of 162 patients (55.3%) underwent cardioversion due to persistent atrial arrhythmias. Comparison between termination by ablation and termination by cardioversion in patients exhibiting AF or AT revealed that no significant difference was observed in early recurrence (38.2% vs. 43.8%, P = 0.328) and SR maintenance (67.2% vs. 59.8%, P = 0.198) during the (23 ± 7) months follow-up. Even after repeat ablation, the SR maintenance continued to exhibit no statistical difference in above two groups (72.5% vs. 70.4%, P = 0.686). Further analysis of subgroups, however, demonstrated that patients with AF terminated directly to SR experienced better clinical outcomes than other subgroups (P < 0.05). Furthermore, atrial arrhythmias present during ablation have been implicated in prediction of recurrence mode: AF or AT (P < 0.05).

CONCLUSIONSTermination as a procedural endpoint is not associated with favorable long-term SR maintenance in persistent AF. AF methods that convert arrhythmia directly to SR have, however, been linked with improved clinical outcomes, although conversions to AT may not be correlated. Atrial arrhythmias observed during the ablation may be used to predict the recurrence mode.

Adult ; Aged ; Atrial Fibrillation ; physiopathology ; therapy ; Catheter Ablation ; methods ; Electrophysiologic Techniques, Cardiac ; Female ; Humans ; Male ; Middle Aged

This study was purposed to summarize the clinical characteristics and laboratorial data of blastic plasmacytoid dendritic cell neoplasm (BPDCN) in pediatric patients in order to enhance understanding this disease in diagnosis and therapy. A rare case of BPDCN in children was enrolled in this study. The blood routine test, examination of bone marrow cell morphology, histopathology and immunophenotype of the skin lesions were performed and analysed, the single cell suspensions of the biopsied skin mass were detected by flow cytometry. The results showed that tumor cells expressed CD4, CD56, CD43 and CD123, while not expressed CD19, CD20, CD3, CD8, CD13, CD11b and myeloperoxidase (MPO). According to the clinical and laboratorial features and the results from histopathological and immunophenotype examinations, BPDCN was confirmed. It is concluded that BPDCN in children is an extremely rare hematopoietic malignancy with presenting a rapidly and fatally aggressive clinical course. The diagnosis of this disease is mainly based on the clinical presentations, pathologic and immunohistochemical features. BPDCN is a highly aggressive disease, its prognosis is very poor, its pathogenesis remans still unclear. A standard treatment protocol for BPDCN has not yet been established.
Adolescent ; Dendritic Cells ; Hematologic Neoplasms ; Humans ; Male ; Skin Neoplasms ; Waldenstrom Macroglobulinemia

Objective To explore the siRNA as a new antiviral therapy,evaluate the inhibition effect of siRNA based on vector on the HBV of HepG2.2.15 cell,and observe the side effect and toxicity of siRNA vector on cells and the off-target effect of siRNA.Methods Three pairs of siRNA duplexes targeting HBV C gene were designed as double strands,and the duplex were annealed and ligated into the p-Silencer-Cmv 4.1-hygro vector.The ligation products were used to transform JM109 cells.The clones with shRNA were obtained,and the vectors were purified.After the initial identification of the vector with agarose gel and the size of the inserted sequence got examined by native polyacrylamide gel electrophoresis,furthermore the sequencing was further carried out.The recombinant plasmids were purified with ultrapure Midipreps DNA Purification System.Then HepG2.2.15 cells were transfected with the plasmid mixed with siPort XP-1.The expression of HBsAg and HBeAg were detected by immunofluorescence and Western blot,and the HBV RNA was investigated by RT-PCR.Furthermore the real-time quantitive PCR was carried out to detect the changes of HBV DNA.In order to evaluate the toxicity of the shRNA,MTT was used to examine the growth rate and curve of cells.The ELISA was performed to detect the changes of interferon-? (IFN-?).Results The Western blot showed that the HBsAg and HBeAg protein were suppressed with (81.15?0.69)%,(88.12?0.92)% respectively by vector p-C2 on the third day of post-transfection.It had the similar result indicated by immunofluorescence.And the RT-PCR showed that the specific siRNA targeting HBV C gene could markedly suppress the expression of HBV mRNA and the HBV C gene mRNA was inhibited with 96.9%.The real-time quantitive PCR showed that the specific functional siRNA could markedly suppress HBV DNA copy with two orders of magnitude,while the siRNA vector had no effect on the growth of cell showed by MTT detection.Compared with the non-transfected group and p-NC group,the IFN-? level was almost the same with siRNA p-C1,p-C2,p-C3 groups.Conclusions The siRNA based on the expression vector can suppress the expression and replication of HBV in HepG2.2.15 cell.The inhibition effect was specific and had a certain dependency on siRNA concentration.No toxicity effect was found in the study.And the drug resistance wouldn′t happen because the silence was based on the split of gene.