Female ; Humans ; Neoplasm Metastasis ; Uterine Cervical Neoplasms ; pathology

OBJECTIVETo identify genes that differentially expressed in Lin(-)CD(34)(-) and Lin(-)CD(34)(+) cells.

METHODSWith Lin(-)CD(34)(-) cells as tester and Lin(-)CD(34)(+) cells as driver, cDNA subtractive library for Lin(-)CD(34)(-) cells was constructed using suppression subtractive hybridization technique. Part of clones in the library were sequenced and the homologue analysis was conducted against the DNA database in GenBank.

RESULTS593 clones containing an average of 300 - 500 bp insert were identified. Of them, 53 randomly selected ESTs were sequenced. Homologue analysis revealed that 37 ESTs represented 10 known genes, and the other 16 ESTs represented 4 novel sequences.

CONCLUSIONPart of specifically expressed genes in Lin(-)CD(34)(-) cells were identified, which maybe related to Lin(-)CD(34)(-) cells' specific characteristics.

Antigens, CD34 ; metabolism ; Cloning, Molecular ; Gene Expression Profiling ; Gene Library ; Hematopoietic Stem Cells ; cytology ; metabolism ; Humans ; In Vitro Techniques ; Nucleic Acid Hybridization

OBJECTIVETo find out the prevalence of sleep disturbances for children aged 2 to 12 years old in Chengdu.

METHODSTotally 1 600 children aged 2-12 years old were selected from 5 districts in Chengdu and investigated by using questionnaire.

RESULTSAll 1 526 survey papers were returned. The average time of every day sleep in each age group (infant group, pre-school age group and school age group) were 12.12 hours, 10.42 hours and 9.47 hours. The sleep time of the children in those three groups were much less than the standard one. The proportion of the prevalence of sleep disturbance was 37.88%. Among them, there were snoring in 5.57%, choke/gargling in 1.25%, sleep inquietude in 7.86%, mouth breathing in 4.59%, sweating in 21.36%, member spasm in 2.82%, molar teeth in 8.26%, night talking in 4.02%, somnambulate in 0.2%, bedwetting in 1.95%, and difficulty falling asleep in 10.75%. There were significant differences shown in different sexes and ages, and in incidence of symptoms of some sleep disturbances. The affecting factors were the co-sleeping, tonsillitis, bronchitis, pollen allergy and their parent's snore.

CONCLUSIONThe prevalence of sleep disturbances being higher and more severe than before might be due to the less sleeping time in Chengdu in children aged 2 to 12 years old. More attention should be paid by parents, the Ministry of Education and the children's doctors.

Child ; Child, Preschool ; China ; epidemiology ; Female ; Humans ; Male ; Prevalence ; Sleep Wake Disorders ; epidemiology ; Surveys and Questionnaires

OBJECTIVETo study the in vitro antitumor activity of bcl-2 fully phosporothioated antisense oligodeoxynucleotide (bcl-2 ASODN) to malignant lymphoblastic cells.

METHODSProliferation and apoptosis of Raji cells incubated with bcl-2 ASODN were evaluated by MTT assay, flow cytometry (FCM) and electron microscopy, and the level of bcl-2 protein and mRNA expression were assessed by FCM and RT-PCR, respectively.

RESULTSMTT assay demonstrated that bcl-2 ASODN could partially inhibit the growth of Raji cells. After incubated with ASODN for 48 hours, Raji cells exhibited characteristic morphologic changes of apoptosis, including cytoplasm membrane blebbing, chromatin condensation crescents formation and nuclear fragmentation. The apoptosis rate of Raji cells treated with 20 micromol/L bcl-2 ASON for 72 hrs was 43.86% which is significantly higher than that of control (10.05%). The bcl-2 ASODN induced apoptosis of Raji cells was accompanied by declined expression of bcl-2 mRNA, which decreased to 0.88% at 72 hrs and was significantly lower than that of control (79.54%).

CONCLUSIONbcl-2 ASODN induced Raji cells apoptosis by downregulating bcl-2 protein.

Apoptosis ; drug effects ; Cell Division ; drug effects ; Dose-Response Relationship, Drug ; Flow Cytometry ; Gene Expression Regulation, Neoplastic ; drug effects ; Humans ; Oligonucleotides, Antisense ; pharmacology ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; metabolism ; RNA, Messenger ; drug effects ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Tumor Cells, Cultured ; drug effects ; metabolism

OBJECTIVEImpulse oscillometry (IOS) is a new method for determination of breathing mechanics, which features convenient operation, good repeatability and wider range analysis. As there is no standardized normal value in China at present, this study will provide a normal value of lung function determination by impulse oscillometry for children in Chengdu area.

METHODTotally 549 children were chosen at random from Chengdu area, with 292 boys and 257 girls who were 4 to 14 years old. The subjects were assigned into 10 age groups according to their chronological age with one year difference between every two adjacent groups. The respiratory total impedance (Zrs), viscosity resistance (Rrs) and elastic resistance (Xrs) at various oscillation frequency were measured by the Master Screen IOS which was manufactured by German Jaeger Company. The measured data were treated with the linear stepwise multiple regression, and established the prediction equation. At the same time, paired comparison was carried out with the measured data and equation obtained from this study, Lechtenboerger equation and prediction equation obtained from Guangzhou area.

RESULTThe total impedance and airway resistance were negatively correlated with the children's height and age. Zrs (male) = -0.756 + 189.586/height, r = -0.782, P < 0.001; Zrs (female) = -0.497 + 152.468/height, r = -0.726, P < 0.001. Rrs became the same in trend; while Xrs were proportional to the height, e.g. the values increased as the height increased. The difference of the airway resistance (R(5)-R(20)) was negatively correlated with the children's height: R(5)-R(20) (male) = 0.601 - 0.0034 x height, r = -0.677, P < 0.001; R(5)-R(20) (female) = 0.549 - 0.0031 x height, r = -0.658, P < 0.001. Among the relationships with many impulse oscillometry parameters, height ranked at first place; age at second. The multiple regression equation of IOS primary index was established. Both the measured data and the correlation coefficient of the study obtained equation were greater than the coefficient correlation of the Lechtenboerger equation, but had no significant difference compared with that of prediction equation in Guangzhou area.

CONCLUSIONThe normal value in impulse oscillometry in children in Chengdu area is different from the predicted parameters in other countries. The equation obtained from this study seems to be more suitable for the children in its local area. It is recommended to apply the predicted value from the corresponding population in the determination of the lung function by impulse oscillometry.

Airway Resistance ; physiology ; Child ; China ; Electric Impedance ; Female ; Humans ; Male ; Oscillometry ; methods ; Respiratory Function Tests ; methods ; Respiratory Physiological Phenomena

Bone marrow mesenchymal stem cells (MSCs) are multipotent tissue stem cells that can be induced in vitro to differentiate into a variety of cells such as osteoblasts, chondrocytes and adipocytes. MSCs are useful vehicles for both cell and gene therapy for a variety of diseases. Here, we injected human MSCs with enhanced green fluorescent protein (EGFP) into the striatum of Parkinson disease (PD) rat and examined their survival, migration, differentiation, and the behavior changes in PD rats, which will provide a theoretical foundation and technical method for clinic PD therapy by stem cells. The results showed that human bone marrow MSCs can survive in rat brain for a long time (exceeding 70 d). MSCs were found in multiple areas of the rat brain including the striatum, the corpus callosum, contralateral cortex and even the brain vascular wall. Immunocytochemical staining suggested that implanted cells expressed human neurofilament (NF), neuron-specific enolase (NSE) and glial fibrillary acid protein (GFAP). At the same time, remission in abnormal behavior of the PD rats appeared. Rotation scores decreased gradually from 8.86+/-2.09 r/min pre-transplantation to 4.87+/-2.06 r/min 90 d post-transplantation (statistic result showed P<0.05).
Animals ; Bone Marrow Cells ; cytology ; Cell Differentiation ; Cell Movement ; Corpus Striatum ; Green Fluorescent Proteins ; administration & dosage ; Humans ; Male ; Mesenchymal Stromal Cells ; cytology ; Parkinson Disease ; therapy ; Rats ; Rats, Wistar ; Stem Cell Transplantation ; methods ; Transplantation, Heterologous

In order to investigate the anti-tumor activity of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors and the mechanism underlying the cell proliferation and apoptosis modulated in myeloma cells, the effects of mevastatin, an HMG-CoA reductase inhibitor, on cell growth, cell cycle progression and apoptosis in U266 human multiple myeloma (MM) cell line in vitro were explored by MTT colorimetric assay, morphologic observation, flow cytometry, DNA gel electrophoresis, and RT-PCR. The results demonstrated that mevastatin inhibited the growth of U266 cells in time- and dose-dependent manners. Cell cycle analysis showed that U266 cells underwent G(0)/G(1) arrest under exposure to mevastatin, but it did not affect p27 expression at both mRNA and protein level. Morphologic observations revealed cytoplasm shrinkage, nuclear condensation and fragmentation in mevastatin-treated cells, and fraction of annexin V(+)PI(-) cells increased significantly in the presence of the agent as determined by flow cytometric assay. In addition, mevastatin caused the collapse of mitochondrial transmembrane potential (Deltapsim), induced DNA fragmentation, and down-regulated the mRNA expression of bcl-2. The growth-inhibitory, cell cycle arresting, and proapoptotic effects of mevastatin in U266 cells could be effectively reversed by the addition of mevalonate (MVA), the immediate endproduct of the reaction catalyzed by HMG-CoA reductase. It is concluded that mevastatin suppresses proliferation by inducing G(0)/G(1) phase arrest and triggering apoptosis via down-regulation of bcl-2 and reduction of Deltapsim, which may be attributed to the inhibition of MVA pathway by mevastatin. Statins including mevastatin may find their future application in the treatment of MM.
Apoptosis ; drug effects ; Cell Division ; drug effects ; Cell Line, Tumor ; G1 Phase ; drug effects ; Genes, bcl-2 ; Humans ; Hydroxymethylglutaryl-CoA Reductase Inhibitors ; pharmacology ; Lovastatin ; analogs & derivatives ; pharmacology ; Multiple Myeloma ; drug therapy ; pathology

Ex vivo maintainance of human stem cells is crucial for many clinical applications. Current culture conditions provide some level support but cytokines induce most quiescent stem cells to proliferate and differentiate. Better control of primitive cells is needed to extend the time and range of manipulation of such cells. A recently identified plant lectin Flt3 receptor-interacting lectin (FRIL) present may a special ability to preserve primitive CB progenitors for extended periods in culture without exogenous cytokines. But the mechanisms of FRIL preserving quiescent primitive cells are still unknown. Recently a novel protein HTm4 and its alternatively spliced variant HTm4S, which serve as hematopoietic cell cycle regulators, have been identified. In this report we studied the effect of FRIL on the in vitro maintenance of quiescent human cord blood stem cells and the expression of the novel hematopoietic cell cycle regulator HTm4 and HTm4S in progenitor cells cultured in FRIL. We analyzed the proliferation and the HPP-CFC proportion of CD34(+) cells treated with FRIL. The human HTm4 and HTm4S mRNA expression was detected by semi-quantitative RT-PCR, and the cell cycle status of CB CD34(+) cells was analyzed by FACS. The results showed that incubation of CD34(+) cells in FRIL resulted in a low proliferation of progenitor cells and fewer cycling cells, but FRIL selectively maintained a higher number of primitive cells with proliferative potential in suspension culture. CB CD34(+) cells cultured in FRIL showed significant diversity in the expression of HTm4 and HTm4S during 0~14 d. On d 0, HTm4 was detected at high level, downregulated on d 1, but upregulated during d 3 to d 14, and reaching the highest level on d 7. But the expression levels of HTm4S changed little in the cells cultured in FRIL except the obviously increased expression on d 7. Exogenous expression showed that HTm4 was localized around the karyon while HTm4S scatted in the cytoplasm, respectively, which may be responsible for their difference in function. Thus, FRIL can preserve quiescent primitive CD34(+), and FRIL's ability to preserve quiescent primitive cells in a reversible manner may significantly expand the time and range of ex vivo manipulations of human stem cells for clinical applications. In other words, HTm4 and HTm4S may play a crucial role in the cell cycle modulation of CD34(+) progenitor cells maintained with FRIL in vitro.
Antigens, CD20 ; biosynthesis ; genetics ; Antigens, CD34 ; metabolism ; Cell Cycle ; Cell Cycle Proteins ; biosynthesis ; genetics ; Cell Separation ; Cells, Cultured ; Fetal Blood ; cytology ; Hematopoietic Stem Cells ; cytology ; Humans ; Mannose-Binding Lectins ; pharmacology ; Membrane Proteins ; biosynthesis ; genetics ; Plant Lectins ; pharmacology

In order to investigate the role and the mechanism of protein tyrosine phosphatase (PTPase) signaling pathway in the regulation of proliferation, cell cycle and apoptosis in lymphoma cells, the effects of sodium orthovanadate, Na(3)VO(4), a specific PTPase inhibitor, were explored on Raji lymphoblast-like cell line by MTT assay and CFU-Raji culture, morphologic observation, DNA gel electrophoresis, FCM and RT-PCR. Results showed that MTT assay and CFU-Raji culture demonstrated that sodium or thovanadate inhibited the growth of Raji cells in a concentration-dependent fashion; morphologic observations showed that Raji cells exhibited cytoplasm shrinkage, cytoplasm membrane blebbing, nuclear fragmentation and chromatin condensation forming crescents along nuclear membrane characteristic of apoptosis in the presence of Na(3)VO(4); DNA gel electrophoresis revealed typical DNA ladder reminiscent of DNA cleavage at internucleosomal sites in Na(3)VO(4) treated cells; FCM and RT-PCR indicated that Na(3)VO(4) intervention increased the fraction of annexin V(+) PI(-) cells, reduced the value of mitochondrial transmembrane potential, induced G(2)/M arrest and down-regulated the expression of Bcl-2 and cyclin B1 at both mRNA and protein level in a concentration-dependent manner. It was concluded that PTPase pathway might be implicated in the regulation of cell proliferation, cell cycle and apoptosis, and PTPase specific inhibitor Na(3)VO(4) could induce Raji cell growth inhibition, G(2)/M arrest and apoptosis via down-regulation of Bcl-2 and cyclin B1, and reduction of mitochondrial transmembrane potential.
Apoptosis ; drug effects ; Cell Division ; drug effects ; Cyclin B ; analysis ; Cyclin B1 ; Enzyme Inhibitors ; pharmacology ; Humans ; Leukocyte Common Antigens ; analysis ; Membrane Potentials ; drug effects ; Mitochondria ; drug effects ; physiology ; Protein Tyrosine Phosphatases ; antagonists & inhibitors ; Vanadates ; pharmacology

To investigate the expression of telomerase in cord blood hematopoietic stem/progenitor cells during their committed differentiation in vitro and provide an index of monitoring the proliferating potential of the hematopoietic stem/progenitor cells and security for clinical application. Human CD34 positive cells were isolated from umbilical cord blood by using magnetic cell sorting system (MACS), and were induced to differentiation with hematopoietic growth factors (SCF + IL3 + IL6 + GCSF and SCF + IL3 + IL6 + EPO) in a liquid culture system. The telomerase activity and the cytalytic subunit of telomerase (hTERT) of the cells were analysed during different periods of culture by using TRAP-PCR, TRAP-ELISA, Western blot and RT-PCR techniques, respectively. The results showed that a peak of cell growth was achieved on day 14 - 21 during induction of differentiation in vitro. Total cell number could increase 1006.4 +/- 103.2 times and could not increase there after. Telomerase activity and hTERT expression were low in freshly isolated cord blood CD34(+) cells and increased after about 7 days of culture in addition of cytokine combinations of SCF + IL3 + IL6 + GCSF and SCF + IL3 + IL6 + EPO, respectively. The telomerase activity and hTERT decreased after 14 days of culture and were not detected after 28 days of culture. It was concluded that the hematopoietic stem/progenitor cells can be expanded in large number in vitro and do not have the character of immortality and the telomerase activity could be a useful index in hematopoiesis regulation.
Antigens, CD34 ; blood ; Blotting, Western ; Cell Differentiation ; DNA-Binding Proteins ; Enzyme-Linked Immunosorbent Assay ; Fetal Blood ; cytology ; Hematopoietic Stem Cells ; cytology ; enzymology ; Humans ; Polymerase Chain Reaction ; RNA, Messenger ; analysis ; Telomerase ; genetics ; metabolism