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Introducción: Los antígenos plaquetarios humanos (HPA) se expresan en 6 glucoproteínas plaquetarias diferentes. Se ha descrito que estos antígenos pueden estimular la producción de aloanticuerpos una vez expuestos a plaquetas humanas con diferentes HPA, lo que provoca complicaciones clínicas como la trombocitopenia neonatal aloinmune y la púrpura postransfusional. Métodos: Se realizó el estudio a 11 muestras de pacientes en espera de trasplante renal de genotipo de los antígenos HPA-1,2,3 a/b mediante PCR multiplex, mientras que para el estudio de genotipo de los antígenos HPA-5a/b se utilizó la técnica de PCR con secuencia específica de primer. Los productos de ADN amplificados fueron visualizados mediante electroforesis en gel de agarosa y electroforesis capilar. Resultados: El análisis de los fragmentos de ADN amplificados revelaron resultados similares por ambos métodos. Para los antígenos HPA-1,-2, el 63 por ciento de las muestras fueron homocigóticas para el fenotipo (a) mientras que se observó heterocigocidad en todos los casos para el genotipo HPA-3. En el sistema HPA-5, el 54 por ciento fueron homocigóticas para el fenotipo (a) y el 46 por ciento, heterocigóticas. Para el genotipo del HPA-15, el 4 por ciento fueron homocigóticas para el fenotipo (b) mientras que el 96 por ciento resultaron ser heterocigóticas. Conclusiones: Estos resultados muestran similitudes para los genotipos HPA 1, 2,3 a/b, HPA 5a/b y HPA15 a/brespecto a lo planteado en la literatura(AU)
Introduction: Human platelet antigens (HPA) are expressed in 6 different platelet glycoproteins. It has been described that these antigens can stimulate the production of alloantibodies once exposed to human platelets with different HPA, which causes clinical complications such as neonatal alloimmune thrombocytopenia and postransfusional purpura. Methods: The study was performed on 11 samples of patients awaiting kidney transplantation of genotype of the HPA-1,2,3 a/b antigens by multiplex PCR, while for the genotype study of the HPA-5a/b antigens was used the PCR technique with primer-specificsequence. The amplified DNA products were visualized by agarose gel electrophoresis and by capillary electrophoresis. Results: The analysis of DNA fragments amplified by agarose electrophoresis and capillary electrophoresis revealed similar results in both methods. For the HPA-1, -2 antigens, 63 percent of the samples were homozygous for phenotype (a) while heterozygosity was observed in all cases for the HPA-3 genotype. In the HPA-5 system, 54 percent were homozygous for the phenotype (a) and 46 percent were heterozygous. For the genotype of HPA-15, 4 percent were homozygous for phenotype (b) while 96 percent proved heterozygous. Conclusions: These results show similarities for the genotypes HPA 1, 2.3 a/b, HPA 5a/b and HPA15 a/bwith respect to report in literature(AU)
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Humanos , Masculino , Femenino , Antígenos de Plaqueta Humana/genética , Técnicas de Genotipaje/métodosRESUMEN
The clinical application of doxorubicin (DOX) in cancer chemotherapy is limited by its life-threatening cardiotoxic effects. Chrysophanol (CHR), an anthraquinone compound isolated from the rhizome of L., is considered to play a broad role in a variety of biological processes. However, the effects of CHR׳s cardioprotection in DOX-induced cardiomyopathy is poorly understood. In this study, we found that the cardiac apoptosis, mitochondrial injury and cellular PARylation levels were significantly increased in H9C2 cells treated by Dox, while these effects were suppressed by CHR. Similar results were observed when PARP1 activity was suppressed by its inhibitors 3-aminobenzamide (3AB) and ABT888. Ectopic expression of PARP1 effectively blocked this CHR׳s cardioprotection against DOX-induced cardiomyocyte injury in H9C2 cells. Furthermore, pre-administration with both CHR and 3AB relieved DOX-induced cardiac apoptosis, mitochondrial impairment and heart dysfunction in Sprague-Dawley rat model. These results revealed that CHR protects against DOX-induced cardiotoxicity by suppressing cellular PARylation and provided critical evidence that PARylation may be a novel target for DOX-induced cardiomyopathy.
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@#[Abstract] Objective: To establish a method for in vitro isolation and culture of T lymphocytes from peripheral blood of cynomolgus monkeys that induced by human CD3 antibody based on the foundation of protein homology of CD3 from human, cynomolgus monkey and porcine. Methods: The amino acid sequences of human, cynomolgus monkeys and porcine CD3 proteins were obtained from NCBI, and the sequence, homology and phylogenetic tree were analyzed by DNAMAN software. Western blotting was used to detect the expression of CD3 protein on T cell membranes from the three species. PBMCs of healthy cynomolgus were isolated and divided into three groups: group A was stimulated with anti-human CD3Ab alone, group B was stimulated with IL-2 alone, and group C was costimulated with human CD3Ab and IL-2. Cell morphology and growth status were observed under inverted microscope and the cell growth curve was plotted. Cell viability was detected by trypan blue staining and the expressions of CD3, CD4 and CD8 on T cell surface were detected by flow cytometry. Results: The homology of the amino acid sequence of human CD3 protein to cynomolgus monkey and porcine were 86.9% and 65.6% respectively. The expression levels of CD3 protein on cynomolgus and porcine T cell membrane were 79% and 17% contrast to human, respectively. Cells of group A did not proliferate. Proliferation, viability and CD3 expression [(93.8±3.6)% vs (70.3±4.7)%, P<0.01] in T cells of group C were significantly higher than those in group B. Growth curve of T cells in group C showed an S-shape, which is consistent with Logistic growth curve. T cells in group C exhibited high purity and expressed high level CD3; moreover, the CD8+T cell took a high proportion. Conclusion: The membrane of T lymphocytes from peripheral blood of cynomolgus can express CD3 protein that highly homological to human. Co-stimulation of human CD3Ab, IL-2 and 1% PHAcan induce the proliferation and differentiation of T lymphocytes of cynomolgus, and obtain T lymphocytes with good growth status, high proliferation ability and high purity.
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Objective For Genistein has been reported to inhibit many tumors ,we investigate the role of autophagy in the proliferation inhibition to Hela cells by Genistein and the machanism of autophagy plays in this process.Methods Human cervical cancer Hela cells were divided into control group,Genistein group and 3-MA+Genistein group,the control group were cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum(FBS),Genistein group were cultured in various concentrations Genistein(25,50,100μmol/L),3-MA+Genistein group were treated with 5mmol/L 3-MA for 1h before cultured in 100μmol/L Genistein.The proliferation inhibitory rate of Hela cells was detected by MTT method.The ultrastructure changes of Hela cells was observed under transmission electronic microscope(TEM).The levels of autophagy-associated protein P62 and Beclin-1 were detected by Western blotting analysis.The expressions of autophagy-associated proteins LC3A/B in Hela cells were determined by fluorescent staining to analyse the autophagy induced by Genistein in Hela cells.Results Compared with control group ,the proliferation inhibitory rate of Hela cells was 20.9%±1.3%,33.5%±1.6% and 46.5%±3.2% when cultured in 25,50,100μmol/L Genistein(P<0.01).After treated with various concentrations Genistein for 48h, we observed a dose-dependent increase in the expression of Beclin-1 and decrease of P62.Confocal laser scanning microscopy confirmed the fluorescent density of LC3A/B expression in Hela cells treated with 100μmol/L Genistein increased significantly as compared with control group.TEM showed there are many vacuoles and double-membrane autophagosomes which involved cytoplasmic components in Hela cells treated with 100μmol/L Genistein.The proliferation inhibitory rate of Hela cells of Genistein group is decreased as compared with those in 3-MA+Genistein group[(46.5±3.2)% vs (58.2±2.2)%,P<0.01].Conclusion Genistein could inhibit Hela cells proliferation and induce autophagy.
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To identify linear epitopes on the non-structural protein 3AB of foot-and-mouth disease virus (FMDV),BABL/c mice were immunized with the 3AB protein and splenocytes of BALB/c mice were fused with myeloma Sp2/0 cells.Two hybridoma monoclonal antibodies (mAbs) cell lines against the 3AB protein of foot-and-mouth disease virus (FMDV) were obtained,named C6 and E7 respectively.The microneutralization titer was 1∶1024 for mAb C6,and 1∶512 for E7.Both mAbs contain kappa light chains,and were of subclass IgG2b.In order to define the mAbs binding epitopes,the reactivity of these mAbs against FMDV were examined by indirect ELISA.The results showed that both mAbs can react with FMDV,but had no cross-reactivity with Swine Vesicular Disease (SVD) antigens.The titers in abdomen liquor were 1∶5×106 for C6 and 1∶2×106 for E7.In conclusion,the mAbs obtained from this study are specific for the detection of FMDV,can be used for etiological and immunological researches on FMDV,and have potential use in diagnosis and future vaccine designs.
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Objective To evaluate the ability of 3-AB to sensitize the human esophageal carcinoma cell strain (CaEs-17) to radiation in v/tro and its mechanisms. Methods CaEs-17 cells were treated with 3-AB at 0, 2.5, 7.5 mmol/L and given irradiation O, 3, 6, 9, 12 Gy. 3-AB concentration in each group was made dose-survival curve using multi-target single-hit maiths model by clonogenie assay. MTT assay was performed to observe the survival of irradiated cells.comet assay and metaphase chromosome analysis were used to measure the DNA damage degree and chromosome aberration of CaEs-17 cell after 3-AB treatment and irradiation. Results Cell survival experiments showed SER of 1.21, 1.52 for 2.5 mmol/L, 7.5 mmol/L 3-AB respectively using multi-target single-hit maths model. The survival fraction of irradiated CaEs-17 cell was decreased after 3-AB treatment. DNA damage and the chromatid breakage number of irradiated CaEs-17 cells were increased after 3-AB treatment. Conclusions 3-AB, a PARP inhibitor, can enhance the radiosensitivity of human esophageal carcinoma cell strain (CaEs-17). DNA damage repair inhibition by 3-AB might be one of the mechanisms.