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Objeetive To construct the cell DNA damage models for the human CML K562 cell line before or after imarlnib mesylate treatment and observe the repairing process dynamically for investigating the iniluence of imatinib mesylate on the repair function of K562 cells after cell DNA danlage.Methods The MTT assay was used to estimate the optimal pretreatment concentration of imatinib mesylate in K562 cells and Western blot was employed to evaluate the phosphorylation status in K562 cells after imatinib mesylate treatment to estimate BCR/ABL tyrosine kinase inhibition by imatinib mesylate.The comet assay was used to detect the DNA damage induced by hydrogen peroxide at various concentrations in K562 cells with or without the pretreatment of imatinib mesylate.A dynamic observation on the repairing process after cell DNA damage was made by the comet assay.Results The pretreatment by imatinib mesylate for K562 cells was optimized to be at a final concentration of 1 μmol/L for 24 h as revealed by the MTT assay additionly imatinib mesylate treatment at this concentration could effectively inhibit the phosphorylation of the BCR-ABL fusion protein at Tyr177(Deusityrate 0.100±0.018).When compared with the control group(Deusityrate 0.425±0.039),the BCR/ABL phosphorylation at Tyr177 was significantly decreased by (77. 11±5.59) % (t=4. 57,P＜0. 05). The cell DNA damage models for both imatinib mesylate-nontreated and imatinib mesylate-pretreated K562 cell groups were constructed with hydrogen peroxide treatment at a final concentration of 10 μmol/L for 10 min at 4℃ as confirmed by the comet assay. When compared with the control imatinib mesylatenontreatod K562 cell group,the time duration required for the DNA repair in imatinib mesylate-pretreated K562 cell group was significantly prolonged (F= 97.79,P＜0. 05 ). Conclusions The cell DNA damage models for the leukemic K562 cell groups before or after imatinib mesylate treatment were successfully constructed and the tyrosine kinase inhibitor imatinib mesylate for BCR/ABL fusion protein was revealed to attenuate the DNA repair capacity of the K562 cells after DNA damage.
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To investigate whether human amniotic membrane (HAM) preparations have the possibility to type I hypersensitivity and its allergenicity. In systemic active allergic test model, 30 guinea pigs were equally divided into three groups. Each 10 guinea pigs were immunized with fresh HAM homogenate, albumen solution (positive control) and PBS (negative control). After the animals were stimulated with corresponding allergen, observe their reaction till dying or 3 h, then obtain blood samples, to determine blood histamine concentrations using chemical fluorometry and four hemorheologic markers by hemorheology analysis system. The guinea pigs responded to fresh HAM homogenate in almost the same manner as to PBS, and no obvious allergic reaction was observed in the animals except those in positive control group. The blood histamine concentration and four hemorheologic markers showed no significant differences between HAM and PBS (P > 0.05), both were much lower than positive control group (P < 0.01). Fresh HAM won't lead to type I hypersensitivity for lack of allergen performance.
Subject(s)
Animals , Humans , Allergens , Allergy and Immunology , Amnion , Allergy and Immunology , Guinea Pigs , Histamine , Blood , Hypersensitivity, Immediate , Allergy and Immunology , Materials TestingABSTRACT
Objective:Our purpose is to clarify the mechanisms for total saponins of Panax ginseng(TSPG) inducing K562 cells to apoptosis.Methods:By using morphological observation and TUNEL,the effect of TSPG on apoptosis of K562 cells were studied.The expression of Fas was studied by immunocytochemistry,and sfas was assayed by ELISA.Results:The results indicated that 50,100?g/ml TSPG could induce K562 cells to apoptosis.Our experiment also showed after induced by TSPG,the ratio of positive K562 cells of expression Fas is increased ( P
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Objective To construct the transformed mouse BaF3-P210 cell line stably expressing BCR/ABL and to initially investigate the influence of BCR/ABL on the cell biological characteristics of BaF3 cell line. Methods The retroviral vector with bcr/abl gene was transfected into the packaging cell line. The BaF3 cells were infected with the collected viral supernatant. The transgenic BaF3-P210 cell line stably expressing BCR/ABL were screened and subcloned. The integration of the bcr/abl gene in the genome of the target cell was determined by PCR and DNA sequencing,trypan blue staining assay,flow cytometry and MTT assay. Results The bcr/abl gene was integrated into the BaF3 cell genome; RT-PCR and Western blot verified the stable expression of the bcr/abl gene and protein in the screened subclone cell line BaF3-P210. Compared with the control group,the cell proliferation rate was promoted (P
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Objective To clarify the mechanisms of total saponins of panax ginseng (TSPG) in induction of apoptosis of K562 cells and to provide the theoretical basis and the experimental evidence for its clinical application. Methods The effects of TSPG on apoptosis of K562 cells were studied by morphometry, flow cytometry, morphological observation, and immunocytochemistry. Results The results indicated that TSPG could markedly inhibit the proliferation of K562 cells and induce the apoptosis of K562 cells. Our experiment also showed that after induction by TSPG, the ratio of positive K562 cells with Fas expression increased. Conclusion The mechanisms of TSPG in the induction of K562 cell apoptosis may be related to the increased expression of Fas in K562 cells.