RESUMEN
The receptor activator of nuclear factor-kappaB ligand (RANKL) and interleukin-1beta are osteoclast activating factors which are abnormally expressed in bone marrow stromal cells and plasma cells of multiple myeloma patients. In this work we analyzed RANKL expression in human bone marrow mesenchymal stromal cells and the effect of the bisphosphonate ibandronate on RANKL expression after IL-1beta activation of ERK pathway. Mesenchymal stromal cells were obtained from bone marrow iliac aspirates from multiple myeloma patients at stages II/III and non-osteoporotics control donors; these cells were maintained under long-term culture conditions. Cells were cultured in the presence or the absence of 5 ng/ml IL-1beta and/or 5 microM ibandronate, during selected periods. mRNA for RANKL and protein levels were assayed by RT-PCR and Western blot, respectively. Human bone marrow stromal cell line HS-5 was used for assessing IL 1beta- and ibandronate-ERK phosphorylation responses. Multiple myeloma mesenchymal stromal cells differentiate from control cells by increased basal RANKL expression. IL-1beta up regulated RANKL expression showed dependent on activated MEK/ERK pathway. Finally, the bisphosphonate ibandronate, that hindered activation of the MEK/ERK pathway significantly inhibited both basal and IL-1beta dependent RANKL expression by cells. Results indicate that RANKL expression involves the MEK/ERK pathway in multiple myeloma mesenchymal stromal cells, and that early obstruction of this path, such as that achieved with ibandronate, significantly deters RANKL protein expression.
Asunto(s)
Células de la Médula Ósea/efectos de los fármacos , Células de la Médula Ósea/metabolismo , Difosfonatos/farmacología , Mieloma Múltiple/metabolismo , Ligando RANK/metabolismo , Células del Estroma/efectos de los fármacos , Células del Estroma/metabolismo , Anciano , Conservadores de la Densidad Ósea/farmacología , Células de la Médula Ósea/citología , Células Cultivadas , Inhibidores Enzimáticos , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Femenino , Humanos , Ácido Ibandrónico , Interleucina-1beta/metabolismo , Masculino , Persona de Mediana Edad , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Ligando RANK/genética , Transducción de Señal/fisiología , Células del Estroma/citologíaRESUMEN
A stable cyclized 9-mer peptide (cP) containing the active site of alpha-alpha fetoprotein (alphaFP) has been shown to be effective for prevention of estrogen-stimulated tumor cell proliferation in culture or of xenographt growth in immunodeficient mice. cP does not block 17beta-estradiol (E2) binding to its receptors, but rather appears to interfere with intracellular processing of the signal that supports growth. To obtain insight on that mechanism we studied the effect of cP on the proliferation of MCF-7 cells in culture. Proliferation in the presence of 2 microM E2 is decreased up to 40% upon addition of 2 microg ml(-1) cP to the medium; the presence of cP did not increase cell death, cP reduced also the proliferation of estrogen-dependent ZR75-1 cells but had no effect on autonomous MDA-MB-231 cells, cP did not modify the number of binding sites for labeled E2 or affected cell death. We detected increased nuclear p21Cip1 immunoreactivity after cP treatment. Our results suggest that cP acts via p21Cip1 to slow the process of MCF-7 cells through the cycle.
Asunto(s)
Neoplasias de la Mama/patología , Proliferación Celular/efectos de los fármacos , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Estradiol/farmacología , Péptidos Cíclicos/farmacología , alfa-Fetoproteínas/farmacología , Animales , Neoplasias de la Mama/metabolismo , Femenino , Humanos , Ratones , Ratones SCID , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
This study was aimed to obtain additional information on the activity of a cyclized 9-amino acid peptide (cP) containing the active site of alpha fetoprotein, which inhibits the estrogen-stimulated proliferation of tumor cells in culture and of xenografts in immunodeficient mice. Breast cancer cells cultured in the presence of 2 nM estradiol were exposed to cP for different periods and their proliferation, estradiol binding parameters, clustering tendency and expression of E-cadherin and p21Cip1 were analyzed by biochemical and cell biology methods. The proliferation of MCF7 cells was significantly decreased by the addition of 2 microg/ml cP to the medium. cP did not increase cell death rate nor alter the number of binding sites for estradiol nor the endogenous aromatase activity of MCF7 cells. cP also decreased the proliferation of estrogen-dependent ZR75-1 cells but had no effect on estrogen-independent MDA-MB-231 cells. An increased nuclear p21Cip1 expression detected after cP treatment suggests that cP slows MCF7 cell proliferation via this regulator. We propose that cP could represent a novel breast cancer therapeutic agent whose mechanism of action is different from that of tamoxifen or of inhibitors of aromatase.
Asunto(s)
Antineoplásicos/farmacología , Proliferación Celular/efectos de los fármacos , Estradiol/farmacología , Glándulas Mamarias Humanas/efectos de los fármacos , alfa-Fetoproteínas/farmacología , Animales , Antineoplásicos/química , Western Blotting , Neoplasias de la Mama/tratamiento farmacológico , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/biosíntesis , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/efectos de los fármacos , Femenino , Técnica del Anticuerpo Fluorescente , Humanos , Ratones , Ratones Desnudos , Péptidos/farmacología , Ensayos Antitumor por Modelo de Xenoinjerto , alfa-Fetoproteínas/químicaRESUMEN
Se realiz¢ una investigaci¢n descriptiva y transversal en el rea de salud del Policl¡nico Docente Lu¡s A. Turcios Lima de Pinar del R¡o con el objetivo de conocer algunos aspectos epidemiol¢gicos de la lepra durante el decenio de 1991 al 2000. El universo estuvo representado por los 25 pacientes diagnosticados con esta enfermedad durante los a¤os se¤alados. La informaci¢n se obtuvo del departamento de estad¡sticas y de la revisi¢n de las historias de salud individual. Se presentaron los datos en medidas de frecuencias absolutas y medidas de res£menes cualitativos tales como proporciones y tasas. Se encontr¢ que la lepra ha tenido un comportamiento irregular en esta poblaci¢n durante el decenio, predominando en el grupo de 45 a 59 a¤os y en el sexo femenino. El mayor porciento de los enfermos provienen de zonas urbanas, adem s predomin¢ la lepra indeterminada, siendo el principal modo de detecci¢n de la enfermedad el estudio de los contactos de enfermos(au)
Asunto(s)
Humanos , Masculino , Femenino , Adulto , Persona de Mediana Edad , Lepra/epidemiología , Epidemiología DescriptivaRESUMEN
Cultures of skin fibroblasts show variation of androgen binding with culture conditions; binding variations are usually avoided by using confluent cultures. In this work, we analysed the effect of cell density and mitogenic agents on the level of androgen receptor (AR) of cultured human skin fibroblasts. Results demonstrated that in cultures of human skin fibroblasts, cellular binding of dihydrotestosterone was higher in cells grown at low than at high cell density. The reduction in binding resulted from a decrease in the number of high affinity receptors and not from a change in receptor affinity. Immunocytochemistry for AR showed greater staining intensity in cells grown at low than at high cell density. Additionally, immunoblot analysis demonstrated more AR protein in low cell density cultures. On the other hand, it was observed that cells grown at low cell density showed diminished androgen binding capacity after 24 h of treatment with insulin-like growth factor (IGF-l), basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), or granulocyte-colony stimulating factor (G-CSF); this effect of growth factors was not observed in cells grown at high cell density. In conclusion, we found that cell density of cultures and mitogenic agents can regulate AR binding activity in human fibroblasts. While we do not yet know how changes in cell density affect the amount of AR, we conclude that the mechanism could be mediated by activation of the tyrosine kinase pathway, as the effect was reproduced by mitogens.