RESUMEN
Red cell production is primarily determined by the action of erythropoietin. Additional erythropoiesis-regulatory factors include molecules and cellular interactions occurring within the bone marrow (BM) microenvironment. Sotatercept (ACE-011) is an activin receptor ligand trap that binds several members of the TGF-ß superfamily. Treatment with ACE-011 reverses bone loss and reduces the degree of osteoporosis, but it is accompanied by elevated hemoglobin and hematocrit levels. The mechanisms underlying the beneficial effects of ACE-011 on red cell production remain unknown. This study explores the means by which ACE-011 promotes erythropoiesis. We showed that ACE-011 does not directly affect erythroid differentiation of human CD34(+) cells in vitro. We next tested whether ACE-011 acts indirectly by affecting BM accessory cells. Conditioned media produced by BM stromal cells (SCs) inhibited erythroid differentiation of CD34(+) cells while maintained their ability to proliferate. However, conditioned media from SCs treated with ACE-011 partially restored erythropoiesis, coinciding with changes in the molecular and secretory profile of SCs, including the expression and secretion of erythropoiesis-modulatory factors. We conclude that inhibitory factors produced by BM SCs in vitro might control erythropoiesis in vivo and that agents that reverse these microenvironmental signals could provide an approach to attenuate anemia in clinical conditions.
Asunto(s)
Receptores de Activinas Tipo II/antagonistas & inhibidores , Células Precursoras Eritroides/efectos de los fármacos , Eritropoyesis/efectos de los fármacos , Células del Estroma/efectos de los fármacos , Adulto , Células Sanguíneas/citología , Células Sanguíneas/efectos de los fármacos , Células Sanguíneas/metabolismo , Células de la Médula Ósea/citología , Células de la Médula Ósea/efectos de los fármacos , Células de la Médula Ósea/metabolismo , Células Cultivadas/citología , Células Cultivadas/efectos de los fármacos , Células Cultivadas/metabolismo , Ensayo de Unidades Formadoras de Colonias , Medios de Cultivo Condicionados/química , Citocinas/biosíntesis , Citocinas/genética , Citocinas/metabolismo , Evaluación Preclínica de Medicamentos , Células Precursoras Eritroides/citología , Células Precursoras Eritroides/metabolismo , Eritropoyesis/fisiología , Sangre Fetal/citología , Perfilación de la Expresión Génica , Humanos , Técnicas In Vitro , Células K562/citología , Células K562/efectos de los fármacos , Células K562/metabolismo , Ligandos , Especificidad de Órganos , ARN Mensajero/biosíntesis , Proteínas Recombinantes de Fusión , Células del Estroma/fisiologíaRESUMEN
We previously demonstrated that RB18A, a member of TRAP220/DRIP205/PBP family, in vivo acted as a cofactor of transcription by differently regulating p53wt transactivating activity on physiological promoters. Using p53-negative cells transfected with different constructs, we herein demonstrated that RB18A down-regulated p53wt-dependent apoptosis. This biological regulation was due to a specific diminution of p53wt protein level, as level of p53mut and GAPDH proteins was not modified. This p53wt diminution was dependent on proteasome activity, as inhibited by MG-132 inhibitor. This specific p53wt degradation was correlated with an increase in expression of MDM2, which promoted p53wt degradation into proteasome. RB18A up-regulated MDM2 expression by activating MDM2 promoter, even in absence of p53wt. Altogether, these data emphasized that RB18A could regulate p53wt function not only by direct interaction between both proteins, but also by up-regulating promoter activity of MDM2, a p53-regulating partner.
Asunto(s)
Apoptosis/fisiología , Proteínas Portadoras/fisiología , Regulación de la Expresión Génica , Proteínas Nucleares , Factores de Transcripción , Proteína p53 Supresora de Tumor/fisiología , Apoptosis/genética , Carcinoma Embrionario/patología , ADN Complementario/genética , Genes p53 , Humanos , Células K562/citología , Células K562/metabolismo , Neoplasias Pulmonares/patología , Subunidad 1 del Complejo Mediador , Proteínas de Neoplasias/biosíntesis , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/fisiología , Regiones Promotoras Genéticas , Proteínas Proto-Oncogénicas/biosíntesis , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas/fisiología , Proteínas Proto-Oncogénicas c-mdm2 , Proteínas Recombinantes de Fusión/fisiología , Transfección , Células Tumorales Cultivadas/citología , Células Tumorales Cultivadas/metabolismo , Proteína p53 Supresora de Tumor/biosíntesisRESUMEN
AIM: To study whether Cladonia furcata polysaccharide (CFP-1) might induce apoptosis in human leukemia K562 cells. METHODS: Inhibition of proliferation was measured by MTT assay. Morphological assessment of apoptosis was performed with fluorescence microscope and electron microscope. DNA fragmentation was visualized by agarose gel electrophoresis. The amount of apoptosis cells was measured by flow cytometry. RESULTS: CFP-1 (50 - 800 mg/L) inhibited K562 cell proliferation in a concentration-dependent manner. After incubation of K562 cells with CFP-1 300 mg/L for 5 d, morphological changes of typical apoptosis were observed and agarose gel electrophoresis of DNA revealed "ladder" pattern. CONCLUSION: CFP-1 induced apoptosis in human leukemia K562 cells.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Apoptosis/efectos de los fármacos , Células K562/citología , Plantas Medicinales/química , Polisacáridos/farmacología , División Celular/efectos de los fármacos , Humanos , Extractos Vegetales/farmacologíaRESUMEN
OBJECTIVE: To investigate the action of Ginsenosides (GS) in inducing transcription factor c-fos and GATA-1 to explore the mechanism of GS in hematopoietic cells. METHODS: The proliferation effects of GS on granulocytic (HL-60), monocytic (U937), erythrocytic (K562) and megaryocytic (Meg-01) cell lines were observed by using proliferation test of MTT and colony formation of progenitor cells. The combining reaction of transcription factors c-fos and GATA-1 with nuclear protein antigen were analyzed by Western Blot after being treated by GS. RESULTS: (1) GS (10 micrograms/ml) could stimulate and promote proliferation of 3 cell lines with significant difference between GS and non-GS control (P < 0.05 in all) in both MTT test and colony assay. (2) After treatment with GS, c-fos protein in HL-60, K562 and Meg-01 cell lines was increased by 1.5, 2.0 and 2.5 fold respectively, while U937 cell did not express c-fos. (3) Except that U937 cell hadn't expressed GATA-1, the other cell lines after the treatment by GS, the GATA-1 protein level was elevated to 1.5, 2.1 and 1.3 fold of that before treatment. CONCLUSION: The proliferation of three lines initiated by GS was involved in transcription factor c-fos or GATA-1, which could pay the role in the GS induced up-regulation correlated with proliferation and differentiation of hematopoiesis.
Asunto(s)
Caenorhabditis elegans , Medicamentos Herbarios Chinos/farmacología , Ginsenósidos/farmacología , Proteínas de Caenorhabditis elegans , División Celular/efectos de los fármacos , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Medicamentos Herbarios Chinos/aislamiento & purificación , Factores de Unión al ADN Específico de las Células Eritroides , Factores de Transcripción GATA , Factor de Transcripción GATA1 , Ginsenósidos/aislamiento & purificación , Células HL-60 , Células Madre Hematopoyéticas/citología , Humanos , Células K562/citología , Panax/química , Proteínas Proto-Oncogénicas c-fos/genética , Transactivadores/genética , Factores de Transcripción/metabolismo , Células U937/citologíaRESUMEN
The alpha1beta1 and alpha2beta1 integrins, extracellular matrix receptors for collagens and/or laminins, have similarities in structure and ligand binding. Recent studies suggest that the two receptors mediate distinct post-ligand binding events and are not simply redundant receptors. To discern the mechanisms by which the two receptors differ, we focused on the roles of the cytoplasmic domains of the alpha subunits. We expressed either full-length alpha1 integrin subunit cDNA (X1C1), full-length alpha2 integrin subunit cDNA (X2C2), chimeric cDNA composed of the extracellular and transmembrane domains of alpha2 subunit and the cytoplasmic domain of alpha1 (X2C1), chimeric cDNA composed of the extracellular and transmembrane domains of alpha1 subunit and the cytoplasmic domain of alpha2 (X1C2), alpha1 cDNA truncated after the GFFKR sequence (X1C0) or alpha2 cDNA truncated after the GFFKR sequence (X2C0) in K562 cells. Although the cytoplasmic domains of the alpha1 and alpha2 subunits were not required for adhesion, the extent of adhesion at low substrate density was enhanced by the presence of either the alpha1 or alpha2 cytoplasmic tail. Spreading was also influenced by the presence of an alpha subunit cytoplasmic tail. Activation of the protein kinase C pathway with phorbol dibutyrate-stimulated motility that was dependent upon the presence of the alpha2 cytoplasmic tail. Both the phosphatidylinosotide-3-OH kinase and the mitogen-activated protein kinase pathways were required for phorbol-activated, alpha2-cytoplasmic tail-dependent migration.