OBJECTIVETo study the influence of Benzo (a) pyrene on cell cycle distribution of quiescent diploid human embryonic lung fibroblast (HELF) cells.

METHODSHELF cells were synchronized at G0 phase of cell cycle by 0.5% serum starvation for 48 hours and identified by flow cytometry (FCM). Cells were treated with 20 micromol/L benzo (a) pyrene for 4 h and detected for the changes of cell cycle distribution 0 h, 24 h, 48 h after treatment respectively. HELF cells were treated with 0, 5, 10, 20 micromol/L Benzo (a) pyrene for 24 h and detected for cell cycle regulators p53, p21 and p16 expression changes using Western Blotting method. On the other hand, the dynamic changes of these regulators were also been detected within 24 h after 20 micromol/L Benzo (a) pyrene treated for 4h.

RESULTS0.5% Serum starvation for 48 hours could effectively synchronize HELF at G0 stage and G0 reached 78%. Well-modulated control cells entered into cell cycle to synthesize DNA and cells at S phase reached 43.9% 24 h after serum re-stimulate, while 20 micromol/L B (a) P treated cells were arrested in G1 stage. Control cells entered into the G1 stage of next cell cycle another 24 h later, B (a) P treated cells recovered from G1 arrest, 26.5% of which reached S phase, having a delay of almost 24 h compared with controls. After a series of B (a) P concentrations acting for 24 h, we found that P53 and P21 expression increased dramatically. On the other hand, P53 and P21 increased 4 h after treatment, P53 recovered to normal level after 12 h while P21 kept increasing in 24 h. P16 initially decreased and became normal at 24 h.

CONCLUSIONB (a) P induced quiescent HELF cells undergoing a reversible G1 arrest related with p53-p21 pathway.

Benzo(a)pyrene ; toxicity ; Carcinogens ; toxicity ; Cell Cycle ; drug effects ; Cells, Cultured ; Diploidy ; Fibroblasts ; cytology ; drug effects ; Flow Cytometry ; G1 Phase ; drug effects ; Humans ; Lung ; cytology ; embryology

OBJECTIVETo explore the differences of the silica-induced inhibition on cellular proliferation and hprt gene mutagenesis between lung fibroblasts and alveolar type II cells.

METHODSThe proliferation inhibitive cytotoxicity was detected by MTT (3-[4,5-Dimethylthiazolzyl]-2,5-Diphenyl Tetrazolium Bromide) colorimetric method. Mutation in the hprt gene was screened by culture in the presence of the toxic purine analog, 6-thioguanine (6-TG).

RESULTSUnder the same circumstances of silica exposure, alveolar type II cells was more sensitive than lung fibroblasts for proliferation inhibition. The median proliferation inhibition concentration (IC50) of silica on epithelial was 140 micrograms/cm2, whereas IC50 of silica on fibroblasts was 282 micrograms/cm2. At the same doses of silica, the hprt gene mutation frequency in type II cells (84.2 x 10(-6))-156.6 x 10(-6) was statistically higher than that in fibroblasts (67.6 x 10(-6)-114.3 x 10(-6), P < 0.05).

CONCLUSIONThere were significant differences of both silica-induced cell proliferation inhibition and hprt gene mutation between rat lung fibroblasts and type II epithelial cells. In vitro, cultured rat alveolar type II cells were more sensitive in cytotoxicity and hprt gene mutagenesis to silica dust than lung fibroblasts were.

Animals ; Cell Proliferation ; drug effects ; Epithelial Cells ; drug effects ; Fibroblasts ; drug effects ; Hypoxanthine Phosphoribosyltransferase ; genetics ; Lung ; cytology ; drug effects ; metabolism ; Mutation ; Pulmonary Alveoli ; cytology ; drug effects ; Rats ; Silicon Dioxide ; toxicity

Animals ; Beryllium ; toxicity ; Cells, Cultured ; DNA Damage ; drug effects ; Lung ; cytology ; Male ; Rats ; Rats, Sprague-Dawley

This paper aimed to study the protective effect of ginsenoside Rg_1 on endotoxin(LPS)-induced apoptosis of lung epithelial cells and its mechanism of action. Mouse lung epithelial cells(MLE-12) were first treated with LPS. The autophagy changes and apoptosis and the relationship with concentration and time of LPS were observed. Then,the level of autophagy in MLE-12 was regulated at a specific concentration and action time of LPS,and the changes of apoptosis were observed. Secondly,ginsenoside Rg_1 and autophagy inhibitor 3-MA were added respectively at the same concentration and action time of LPS. The lung epithelial cells were grouped to observe the effect of ginsenoside Rg_1 on LPS-induced apoptosis of lung epithelial cells and its mechanism. In the animal experiment,the mice were grouped and tested by apoptosis protein,lung injury score and HE staining section to verify whether ginsenoside Rg_1 has a protective effect on LPS-induced lung injury. The results showed that apoptosis and autophagy increased as the rise of concentration after treatment with LPS for 12 h. The apoptosis increased gradually,and the autophagy increased first and then decreased over time at the LPS concentration of 25 g·L-1. The apoptosis of LPS group was higher than that of control group,and LPS+3-MA group increased further,while apoptosis decreased significantly in LPS+RAM(rapamycin,autophagy promoter) group. The autophagy increased in LPS group,decreased in LPS+3-MA group and increased in LPS+RAM group. The apoptosis of LPS group was higher than that of control group,and the apoptosis of LPS+Rg_1 group decreased. The apoptosis of LPS+Rg_1+3-MA group increased again. The autophagy of LPS group further increased after administration of ginsenoside Rg_1,but decreased after administration of 3-MA. In the in vivo experiments in mice,the apoptosis of LPS group increased significantly compared with the control group,while LPS + ginsenoside Rg_1 group decreased. Lung injury score and HE staining also conformed to the above trend. LPS can induce the apoptosis of lung epithelial cells in a time-dependent and concentration-dependent manner. The autophagy of lung epithelial cells increases with the rise of LPS concentration. At the specific concentration of LPS,autophagy increases first and then decreases after 12-16 hours. Proper increase of autophagy in lung epithelial cells within a certain period of time can reduce the apoptosis induced by LPS,while inhibition of autophagy can increase apoptosis. Ginsenoside Rg_1 has a protective effect on lung cancer epithelial cell apoptosis induced by autophagy.
Animals ; Apoptosis ; Autophagy ; Cells, Cultured ; Epithelial Cells ; drug effects ; Ginsenosides ; pharmacology ; Lipopolysaccharides ; Lung ; cytology ; Mice

OBJECTIVETo investigate the cytotoxicity of the nickel-refining dusts for Chinese hamster lung (CHL) cells and the effects of nickel-refining dusts on the gap junctional intercellular communication (GJIC) of CHL cells.

METHODSThe cytotoxicity of the nickel-refining dusts for the CHL cells was determined in two nickel-refining dusts samples with the CHL cells as the target cells by MTT method while the effects of nickel-refining dusts on the CJIC of the CHL cells were investigated using the scrape-loading and dye transfer (SLDT) technique.

RESULTSThere were no significant difference in the CHL proliferation between all dosage groups in the two samples and the control group at 6 and 12 hours (P > 0.05). The survival rate of cells in all dosage groups were all decreased at 36 hours (P < 0.05), presenting the dosage-reaction relationship and the time-reaction relationship. IC(50) was 21.36 and 23.07 micro/ml for the two samples respectively at 36 hours. Compared with the control group, the transport of Lucifer Yellow (LY) from the injury line to the adjacent cells was decreased when the CHL cells were treated with nickel-refining dusts of 25.00, 50.00 and 100.00 microg/ml (P < 0.01).

CONCLUSIONThe nickel-refining dusts have cytotoxicity for the CHL cells cultivated in vitro, can inhibit the growth of the cells and at a certain concentration can inhibit the GJIC function of CHL cells.

Animals ; Cell Communication ; drug effects ; Cell Survival ; drug effects ; Cells, Cultured ; Cricetinae ; Cricetulus ; Dose-Response Relationship, Drug ; Dust ; Gap Junctions ; drug effects ; Lung ; cytology ; Nickel ; toxicity

OBJECTIVETo explore the DNA methylation levels of genome in cFb transdifferentiation induced by SiO2 in rats.

METHODSThe primary macrophages and fibrocytes of SD rats were co-cultured directly and indirectly, which were exposed to SiO2 at the doses of 25, 50 and 100 g/ml. The transdifferentiation of cFb was identified with immunohistochemical assay. The genomic DNA methylation levels of cFb were detected with HPLC.

RESULTSUnder the condition of indirect co-culture, as compared with control group, the genomic DNA methylation levels of cFb exposed to SiO2 at the doses of 25, 50 and 100 g/ml reduced by 19.9%, 26.9% and 30.3%, respectively (P < 0.05); as compared with cFb exposed to 100 g/ml SiO2, the genomic DNA methylation levels of cFb exposed to 5-aza-dC decreased by 22.0% (P < 0.05). Under the condition of ThinCert(TM) direct co-culture, as compared with control group, the genomic DNA methylation levels of cFb exposed to SiO2 at the doses of 25, 50 and 100 g/ml reduced by 22.2%, 30.2% and 36.7%, respectively (P < 0.05); as compared with cFb exposed to 100 g/ml SiO2, the genomic DNA methylation levels of cFb exposed to 5-aza-dC decreased by 20.6% (P < 0.05).

CONCLUSIONUnder the co-culture condition in vitro, SiO2 could reduce the genomic DNA methylation levels of cFb. The ThinCert(TM) direct co-culture can be used to study the silicosis fibrosis.

Animals ; Cell Transdifferentiation ; drug effects ; Cells, Cultured ; Coculture Techniques ; DNA Methylation ; Fibroblasts ; cytology ; drug effects ; Genome ; drug effects ; Lung ; cytology ; drug effects ; Male ; Rats ; Rats, Sprague-Dawley ; Silicon Dioxide ; adverse effects

OBJECTIVETo assess the severity and reversibility of the chronic toxicity of a novel recombinant human granulocyte colony-stimulating factor (rhG-CSFa) in rats and the dose-effect relationship.

METHODSA total of 100 Sprague-Dawley rats (equal numbers of male and female) were randomly divided into five groups (20 rats in each group): four groups were treated with rhG-CSFa at 500, 100, 10, 1 µg/kg, respectively, and one group was treated with vehicle only to serve as the control. The rats were received subcutaneous injections of rhG-CSFa or vehicle daily for 13 weeks. During the course of the chronic toxicity study, the physical status, body weight, and food consumption were monitored. Half of the rats in each group (n = 10) were sacrificed after the last rhG-CSFa administration, and the other half were sacrificed at five weeks after the last rhG-CSFa administration. Urinalyses, blood biochemistry, hematological analysis, histopathological examination, and immunological tests were performed for each of the rats.

RESULTSThe hematological analyses revealed that the mean white blood cells count, neutrophils count, and neutrophils percentage were increased in male rats at the dose of 10 µg/kg or higher, and these were related with the biological activity of rhG-CSFa. Some small abnormalities were observed in the spleen of a few rats when used highest dose (500 µg/kg, a dosage of 200 folds higher than the normal clinical dosage), but these abnormalities were recovered within 5-week recovery period. No other rhG-CSFa-related abnormalities were observed in this chronic toxicity study.

CONCLUSIONNo significant toxicity and immunogenicity are observed with rhG-CSFa administration to rats in the chronic toxicity studies.

Animals ; Bilirubin ; urine ; Blood Chemical Analysis ; Dose-Response Relationship, Drug ; Female ; Granulocyte Colony-Stimulating Factor ; genetics ; toxicity ; Humans ; Lung ; cytology ; drug effects ; Male ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Recombinant Proteins ; Spleen ; cytology ; drug effects ; Trachea ; cytology ; drug effects

OBJECTIVETo study DNA and nucleus damage in human embryo lung fibroblast (HLF) exposed to hydroquinone (HQ) and its genotoxicity.

METHODSHLF were treated with HQ (0, 10, 20, 40, 80 micro mol/L, respectively) for 3 h and DNA damage was detected by comet assay. HLF was also treated with the same concentrations of HQ for 1 h and micronucleus test was performed after they were cultured for 24 h.

RESULTSComet assay showed that percentage of cells with tails in each groups treated with varied doses of HQ was 12%, 19%, 42%, 79% and 95%, respectively, with mean tail length of 7.87, 9.35, 11.03, 19.28 and 23.32 micro m, respectively, in an obvious dose-dependent manner (P < 0.05). Very significant increase in percentage of cells with tails and length of their comet tail were observed in those groups treated with HQ of 20, 40 and 80 micro mol/L (P < 0.01). And, proportion of high and severe DNA damage increased with dose of HQ. HQ could also induce formation of micronucleus and abnormal nucleus in all groups treated by varied doses of HQ, with rates of micronucleus and abnormal nucleus of 2%, 3%, 10%, 9% and 15%, and 6%, 7%, 16%, 27% and 28%, respectively, in a significant dose-dependent manner. There was significant increase in rates of micronuclei and abnormal nuclei in cells treated with HQ at doses of 20, 40 and 80 micro mol/L (P < 0.05).

CONCLUSIONSExposure to HQ could cause DNA and nucleus damage inducing genotoxic effects on HLF.

Cell Nucleus ; drug effects ; Comet Assay ; DNA Damage ; drug effects ; Embryo, Mammalian ; Fibroblasts ; cytology ; Humans ; Hydroquinones ; toxicity ; Lung ; cytology ; Micronucleus Tests

OBJECTIVETo investigate the effect of salvianolic acid B (SAB) on the proliferation of human embryonic lung fibroblast MRC-5, and the secretion of procollagen I and endogenous transforming growth factor-beta1, (TGF-beta1).

METHODSThe MRC-5 cells were randomly divided into four groups as follows: the control group: cells cultured with DMEM but with no TGF-beta1, or SAB; the TGF-beta1, group: cell cultured with 10 ng/mL TGF-beta1; the SAB1 group: cell cultured with medium with 10 ng/mL TGF-beta1 and 1 pmol/L SAB; the SAB2 group: cell cultured with medium with 10 ng/mL TGF-beta1, and 10 pmol/L SAB. The proliferation of cells was assayed by MTT incorporation. The concentration of amino-terminal propeptide of type I procollagen (PINP), a marker of collagen synthesis, was measured by radioimmunoassay. The endogenous TGF-beta1, levels were measured using ELISA.

RESULTSThe optical density, procollagen I contents, and endogenous TGF-beta1, levels significantly increased when compared with those of the control group (P<0.05). Compared with the TGF-beta1, group, the optical density was obviously lowered, the procollagen I contents and endogenous TGF-beta1, levels significantly decreased in the SAB1 group and the SAB2 group, and better in the SAB2 group, showing statistical difference (P<0.05).

CONCLUSIONSSAB could inhibit the proliferation of MRC-5 cells induced by TGF-beta1 and attenuate the roles of secreting collagen and endogenous TGF-beta1. It had the potential of postponing or delaying the progressive developing of pulmonary fibrosis.

Benzofurans ; pharmacology ; Cell Proliferation ; drug effects ; Cells, Cultured ; Collagen Type I ; metabolism ; Fibroblasts ; cytology ; drug effects ; Humans ; Lung ; cytology ; embryology ; Transforming Growth Factor beta1 ; pharmacology

OBJECTIVETo investigate the effects of dexamethasone on human lung fibroblast cell proliferation, cell cycles, and cell mitogen-activated protein kinases (MAPKs) passway.

METHODSDexamethasone was used at various concentration in culture medium. Cell number was counted using a hemacytometer. Whole cell propidium iodide staining and flow cytometric analysis were performed to determine cellular DNA content. MAPK proteins and activation were tested by Western blot analysis with antibodies to c-Jun N-terminal kinase (JNK), phospho-JNK, extracellular signal-regulated kinase (ERK), phospho-ERK, p38 and phospho-p38.

RESULTS1x10(-7) mol/L and 1x10(-6) mol/L dexamethasone suppressed the proliferation of lung fibroblast cells by 34% and 72%, respectively, than that of control. This suppression was dose-dependant. Dexamethasone suppressed cell cycle with accumulation of cells in G1/G0 stage. It increased from 81.9% to 90.1% compared with that of control. We did not find any apoptosis induced by dexamethasone for lung fibroblast cells. Using Western blot analysis, we found that dexamethasone resulted in decreased activity of ERK, but had no effects on JNK and p38.

CONCLUSIONSDexamethasone may suppresses the proliferation of lung fibroblast cells, which is partly resulted from the facts that it can inhibit ERK activation in MAPK-signaling pathway but has little effect on JNK and p38 pathway. Dexamethasone may not induce lung fibroblast cell apoptosis directly.

Cell Cycle ; drug effects ; Cell Division ; drug effects ; Cells, Cultured ; Depression, Chemical ; Dexamethasone ; pharmacology ; Fibroblasts ; cytology ; Humans ; Lung ; cytology ; MAP Kinase Signaling System ; drug effects ; Mitogen-Activated Protein Kinase Kinases ; drug effects