Animals ; Animals, Newborn ; Apoptosis ; drug effects ; Brain ; drug effects ; pathology ; Disease Models, Animal ; Estrogens ; pharmacology ; Female ; Hypoxia-Ischemia, Brain ; drug therapy ; In Situ Nick-End Labeling ; Male ; Rats ; Rats, Wistar

Objective To study the effect of exogenous prostaglandin E 1 (PGE 1) on the superoxide dismutase(SOD) and nitric oxide(NO) levels in brain tissue of neonatal rats with hypoxic-ischemic brain damage(HIBD).Methods Sixty 7-day old newborn Wistar rats to establish HIBD models,intraperitoneally and subcutaneous injected PGE 1 and TMP,then the rats were killed after hypo- xia and ischemia for 48 hours.Take cerebral cortex of arteria carotis ligation side and made them into homogenate to detect SOD and NO levels in brain tissue.Results SOD level in HIBD group was lower,and NO level was higher than those of normal group(P

AIMTo explore the role of humoral immunity in the pathophysiological process of freezing injury and the possible immune interference in the preventation and treatment of frostbite.

METHODSSevere experimental freezing injury model was made in Wistar rats( n = 20). The concentration of three types of immunoglobulin (IgG, IgA and IgM), two types of complement components (C3 and C4), and circulating immune complex (CIC) were measured respectively before and at 4h, 1d, 3d, and 5d after frostbite. At the same time, the tissue immune complex (TIC) in skeletal muscle and the contents of the red blood cell immune complex (RBC-IC) were also observed and then was the red blood cell immune adherence activity (RCIA).

RESULTSSerum IgG concentration decreased rapidly to the lowest level at 4 h after frostbite IgA concentration dropped to the nadir on 1 day after freezing. Decreases of both immunoglobulins were maintained during the 5 days after frostbite. The fate of both C3 and C4 were the same as those immunoglobulins. Freezing had rather less effect on IgM level. CIC concentration in serum, expressed as the percent of prefreezing increased rapidly and to the zenith on the 3 days post-freezing. By immunofluorescence microscopy, thin continuous linear pattern (IgG) was demonstrated along the SM on the first day post-freezing. Granular and nodular deposits (IgG) appeared along the SM as the time proceeded after frostbite. RBC-IC contents, expressed as the erythrocyte IC rosette rate, increased significantly and to the zenith on the 3 d post-freezing, while RCIA depressed to the nadir at the same time.

CONCLUSIONThe freezing frostbite is an immune complex related disease which have not been reported by others before.

Animals ; Antigen-Antibody Complex ; analysis ; immunology ; Frostbite ; blood ; immunology ; Immunoglobulin A ; immunology ; Immunoglobulin G ; immunology ; Immunoglobulin M ; immunology ; Immunoglobulins ; immunology ; Male ; Rats ; Rats, Wistar

Clinical evidence of hematopoietic restoration with umbilical cord blood (UCB) grafts indicates the UCB can be a useful source of hematopoietic stem cells for routine bone marrow reconstitution. Considering (10 +/- 5) x 10(8) nucleared cells per cord blood unit, there is a potential limitation for the use of cord blood in adults, which, however, can be overcome by ex vivo expansion of cells. A prerequisite for expansion is the significantly higher recovery of MNC, CD34+ cells and colony-forming cells (CFC) by thawing cryopreserved MNC. Cooling rate always acts as a critical factor that can affect the recovery of cells. Although the rate of - 1 degrees C/min is adopted in most of the cryopreservations, no data has been reported about the detailed effects of different cooling rates. The aim of the study was to reveal the different effects of cooling rates on cryopreservation of hematopoietic stem cells from cord blood. UCB samples were collected, and cryopreserved as mononuclear cells (MNC) with different cooling rates of - 0.5 degrees C/min, - 1 degrees C/min, - 5 degrees C/min, and the recovery and viability of MNC and CD34+ cells, the clonogenic capacity and the ex vivo expansion potential of UCB progenitor cells were evaluated after thawing. With - 1 degrees C/min cooling rate, the recovery of MNC reached 93.3% +/- 1.8% , viability 95.0% +/- 3.9% , recovery of CD34+ cells 80.0% +/- 17.9% , and clonogenic recovery were 87.1% +/- 5.5%, 88.5% +/- 8.9%, 86.2% +/- 7.4% for BFU-E CFU-GM CFU-MK, respectively. After 14 days of liquid culture, no significant difference was detected in CFC expansion between fresh and cryopreserved MNC cells with - 1 degrees C/min cooling rate, but this was not the case with - 0.5 degreesC/min and - 5 degrees C/min. In conclusion, it was demonstrated that controlling the rate at - 1 degrees C/min is more suitable for cryopreservation of hematopoietic stem cells than - 0.5 degrees C/min and - 5 degrees C/min.
Cell Survival ; physiology ; Cells, Cultured ; Cryopreservation ; methods ; Erythroid Precursor Cells ; cytology ; Fetal Blood ; cytology ; Flow Cytometry ; Granulocyte-Macrophage Progenitor Cells ; cytology ; Hematopoietic Stem Cells ; cytology ; Humans

The study was aimed to investigate the promotive effect of LRP16 gene on K562 cell proliferation. Open reading frame of LRP16 gene was amplified using reverse transcription-polymerase chain reaction (RT-PCR) and ligated to pGEM-T plasmid to construct LRP16 ORF-pGEM-T recombinant vector. Then, LRP16 ORF identified by sequencing was inserted into pcDNA3.1+ plasmid to construct LRP16 ORF-pcDNA3.1+ recombinant expression plasmid which was transfected into K562 cell lines to make overexpression of LRP16 gene in K562 cells. Survival of cells was determined by MTT assay and growth curve of cells was drawn, the cell cycle was detected by flow cytometry. The results showed that LRP16 ORF was successfully amplified, then the LRP16 ORF-pcDNA3.1+ recombinant plasmid was constructed. The K562 cell line with overexpression of LRP16 gene was established. The promotive effect of LRP16 gene overexpression on proliferation of K562 cells was observed and the effect partially related to the enhancement of cells from G0 to S phase induced by LRP16 gene. It is concluded that LRP16 gene overexpression shows a promotive effect on proliferation of K562 cells.
Cell Proliferation ; Genetic Vectors ; Humans ; K562 Cells ; Neoplasm Proteins ; genetics ; Open Reading Frames ; Plasmids

The main purpose of the this study was to find the candidate cis-elements in negative regulation region throngh analysing the DNA sequences of lrp16 gene promoter so as to provide the experimental basis for screening drugs with inhibitory effect on lrp16 gene expression. The open reading frame (ORF) sequences in uncoding DNA and mRNA sequences of 5' flanking region in lrp16 gene were cloned by the data in GeneBank and Internet; the possibly existing cis-element in thsi region was searched in databank of human transcriptional factor by using TESS and Genomax online promoter analysis software; the drugs related to inhibition of lrp16 gene expression were screened by using SAGE and GEO databank. The results showed that there were many cis-elements in the negative regulation region, including T-Ag, PU.1, c-Ets, XPF-1, P2 alphaA, IL6-6RE and RAR. In cultured cell lines, hormone or its inhibitor such as corticosteroid, tamoxifen, forskolin, phenylephrine, inflammatory factors such as IFNgamma and TNFalpha, and chemotherapeutics 5-fluorouracil could down-regulate the lrp16 gene expression as compared with absent ones. It is concluded that cis-elements including T-Ag, PU.1, c-Ets, XPF-1, P2 alphaA, IL6-6RE and RAR may inhibit lrp16 expression and hormone or its inhibitor such as corticosteroid, tamoxifen, forskolin, phenylephrine, inflammatory factors such as IL6, IFNgamma and TNFalpha, and chemotherapeutics 5-fluorouracil may participate in the regulation of lrp16 gene expression in negative manner.
Cell Line ; Computational Biology ; Gene Expression Regulation ; Humans ; Neoplasm Proteins ; drug effects ; genetics ; Open Reading Frames ; Regulatory Elements, Transcriptional

This study was purposed to investigate lrp16 gene expression in leukemia cell lines and bone marrow cells of leukemia patients and explore the relationship between lrp16 gene expression and development of leukemia. Reverse transcriptase-polymerase chain reaction (RT-PCR) was employed to test the lrp16 mRNA expression in 4 leukemia cell lines, including K562 (CML), HL-60 (APL), MOLT4 (ALL) and U937 cell lines, as well as in bone marrow-derived cells from 115 patients with leukemia. The effect of lrp16 gene expression on genesis and progression of leukemia was analyzed according to clinicopathological features. The results indicated that positive expression of lrp16 mRNA was found in all 4 leukemia cell lines. For leukemia patients, the positive expression rate of lrp16 mRNA in all AML patients was 38% (16/42), in which the positive rates in AML patients with complete remission (CR) and AML patients without remission were 13% (4/30) and 100% (12/12) respectively. The positive expression rate of lrp16 mRNA in ALL patients was 38% (10/26), in which the positive rate in ALL patients with CR and ALL patients without remission were 16% (3/18) and 87% (7/8) respectively. The positive expression rate of lrp16 mRNA in CML patients was 36% (9/25), in which the positive rates in CML patients with CR and CML patients without remission were 20% (4/20) and 100% (5/5) respectively. The positive rate of lrp16 mRNA in CLL patients was 31% (7/22), in which the positive rate in CLL patients with CR and CLL patients without remission were 11% (2/17) and 100% (5/5) respectively. There was no difference of lrp16 gene expression between leukemia subtypes, but there was statistical significant difference in lrp16 gene expression between CR patients and non CR patients (p < 0.001). It is concluded that the lrp16 gene is a leukemic oncogene and closely relates to genesis and progression of leukemia, which may be an indicator for evaluating clinical efficacy of leukemia therapy.
Adolescent ; Adult ; Aged ; Bone Marrow ; metabolism ; pathology ; Female ; HL-60 Cells ; Humans ; K562 Cells ; Leukemia ; metabolism ; pathology ; Male ; Middle Aged ; Neoplasm Proteins ; genetics ; metabolism ; RNA, Messenger ; genetics ; Young Adult

OBJECTIVETo evaluate the efficiency of PGE(1) in relieving the circulatory disorder of ischemic skin flap.

METHODSNew Zealand rabbits were employed in the study with skip flaps each with the size of 2.5 x 6.0 cm(2) being raised from the back. PGE(1) cream in different concentrations, i.e. 0.2%, 0.4%, 0.8% was respectively topically applied to the skin flaps forming 3 groups (n = 10 in each group), while pure cream without PGE(1) was applied to those in control group (n = 30). The PGE(1) was applied 1 hour after the flap was opened, raised and sutured back. Blood perfusion in the flap was measured with Laser Doppler flowmetry before and 5, 10, 15, 20, 30, 45 and 60 mins after PGE(1) application. The tissue samples from the skin flap were harvested at 2 hours after PGE(1) application for immunohistological staining, and the cross sectional area of capillary lumens was measured under microscope. The survival area of the flap was assessed on the 3(rd) day after operation for the calculation of relative survival length of the flap. Clinically, PGE(1) ointment was applied onto the skin flap vulnerable to necrosis, and the outcome of the flap was observed thereafter.

RESULTSThe blood perfusion in animal skin flaps was increased evidently after PGE(1) application, especially at 30 mins after PGE(1) usage when compared with that in control group (P < 0.05). The capillaries in the skin flap in PGE(1) application groups were dilated obviously after drug usage as observed under microscope (P < 0.05). The survival area and relative survival length in groups 1 and 2 on the 3(rd) post-operational day were much more increased when compared with those in other groups (P < 0.01). Clinically, the skin flaps treated with PGE(1) survived well even in the distal end of the flaps.

CONCLUSIONThe blood perfusion and the survival rate of the skin flaps could be improved by local application of PGE(1) in concentrations of 0.2% or 0.4%.

Adult ; Alprostadil ; administration & dosage ; Animals ; Female ; Humans ; Ischemia ; drug therapy ; Male ; Rabbits ; Surgical Flaps ; blood supply

BACKGROUNDStudies of interleukin (IL)-4 and IL-6 in the exhaled breath condensate (EBC) of asthmatic patients are limited. This study was to determine the effect of inhaled corticosteroid (ICS) treatment on IL-4 and IL-6 in the EBC of asthmatic patients.

METHODSIn a prospective, open-label study, budesonide 200 μg twice daily by dry powder inhaler was administered to 23 adult patients with uncontrolled asthma (mean age 42.7 years) for 12 weeks. Changes in asthma scores, lung function parameters (forced expiratory volume in 1 s [FEV1], peak expiratory flow [PEF], forced expiratory flow at 50% of forced vital capacity [FEF50], forced expiratory flow at 75% of forced vital capacity, maximum mid-expiratory flow rate) and the concentrations of IL-4 and IL-6 in EBC were measured.

RESULTSBoth asthma scores and lung function parameters were significantly improved by ICS treatment. The mean IL-4 concentration in the EBC was decreased gradually, from 1.92 ± 0.56 pmol/L before treatment to 1.60 ± 0.36 pmol/L after 8 weeks of treatment (P < 0.05) and 1.54 ± 0.81 pmol/L after 12 weeks of treatment (P < 0.01). However, the IL-6 concentration was not significantly decreased. The change in the IL-4 concentration was correlated with improvements in mean FEV1, PEF and FEF50 values (correlation coefficients -0.468, -0.478, and -0.426, respectively).

CONCLUSIONSThe concentration of IL-4 in the EBC of asthmatic patients decreased gradually with ICS treatment. Measurement of IL-4 in EBC could be useful to monitor airway inflammation in asthmatics.

Administration, Inhalation ; Adult ; Asthma ; drug therapy ; physiopathology ; Breath Tests ; Budesonide ; administration & dosage ; Female ; Forced Expiratory Volume ; Humans ; Interleukin-4 ; analysis ; Interleukin-6 ; analysis ; Male ; Middle Aged ; Peak Expiratory Flow Rate ; Prospective Studies

BACKGROUNDThe number of Clara cells and the Clara cell 16-kDa protein (CC16) levels of the lung decrease in patients with chronic obstructive pulmonary disease (COPD). N-acetylcysteine (NAC) is a powerful antioxidant and can reduce the frequency of acute exacerbations of COPD. But the exact mechanism is unclear. The present study was designed to investigate the effects of NAC on Clara cells in rats with cigarette smoke exposure.

METHODSEighteen adult male Wistar rats were randomly divided into 3 groups, 12 exposed to cigarette smoke (CS) thrice a day, 10 cigarettes for 30 minutes each time for 1 week, without (CS group) or with (CS + NAC group) oral intake of NAC 80 mg x kg(-1) x d(-1), and another 6 rats exposed to fresh air (control group). Clara cells were observed by an electron microscope. The mRNA expression of CC16 and CC16 protein in lungs were determined by reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemistry respectively. The glutathion (GSH) level in plasma and lung tissue were tested by fluorimetry assay.

RESULTSCompared with the controls, the pathologic score of small airways significantly increased in the CS exposed rats (20.3 +/- 14.7 vs. 53.7 +/- 11.5, P < 0.05). The Clara cell particles in cytoplasm decreased in the CS group (P < 0.05). The percentage of CC16-positive cells in bronchioles in the CS group (27.8 +/- 4.3 and 29.5 +/- 2.4 in terminal bronchioles and respiratory bronchioles, respectively) significantly decreased as compared with the control group (37.1 +/- 3.8 and 43.8 +/- 5.8 in terminal bronchioles and respiratory bronchioles, respectively) (P < 0.05). No significant difference was observed in GSH level ((181 +/- 26) nmol/L in the control group vs. (170 +/- 18) nmol/L in the CS group) between the two groups. After treatment with NAC, the pathologic score of small airways (24.1 +/- 17.5) decreased (P < 0.05). Clara cell particles in cytoplasm of Clara cells increased and GSH level in plasma ((213 +/- 40) nmol/L vs. (170 +/- 18) nmol/L in the CS group) increased too (P < 0.05), while the increase in the proportions of CC16 positive cells in bronchioles (30.1 +/- 6.4 and 34.3 +/- 6.3 in terminal bronchioles and respiratory bronchioles, respectively) did not reach the statistical significance (P > 0.05). No significant difference was found in the expression of CC16 mRNA among the three groups. Correlation analysis indicated that the percentage of CC16-positive cells in bronchioles negatively correlated with the pathologic score of small airways (r = -0.592, P < 0.05), but not with GSH level.

CONCLUSIONSOne-week CS exposure decreased the number of Clara cells and the expression of CC16 in bronchioles in rats. NAC might provide protection of the Clara cells from oxidative damage and possibly through the elevation of the synthesis and secretion of CC16. These data indicate that NAC decreases airway inflammation induced by CS via induction of CC16.

Acetylcysteine ; metabolism ; Animals ; Bronchioles ; cytology ; drug effects ; metabolism ; Fluorometry ; Glutathione ; metabolism ; Immunohistochemistry ; Male ; Microscopy, Electron, Transmission ; Random Allocation ; Rats ; Reverse Transcriptase Polymerase Chain Reaction ; Smoking ; adverse effects ; Uteroglobin ; genetics ; metabolism