RESUMEN
Previous findings from our laboratory demonstrated that when used at low concentration (0.1 microg ml(-1)), CsA as well as its analog PSC 833 were able to revert the MDR phenotype, while at high concentration (1 microg ml(-1)) were able to induce apoptosis. CsA induced apoptosis in leukemia cell lines sensitive (LBR-) and resistant to vincristine (LBR-V160), and doxorubicin (LBR-D160), while PSC 833 only induced apoptosis in vincristine-resistant cell line (LBR-V160). In this work, we investigated mitochondrial-associated mechanisms during CsA- and PSC 833-induced apoptosis. Mitochondrial function was evaluated by recording changes in its transmembrane potential, cytochrome c release, and caspase activation cascade. Results showed that CsA- and PSC 833-induced apoptosis was associated with mitochondrial depolarization, through potentiometric measurements with JC-1 and DiOC(6) probes. Collapse of mitochondrial potential in these cell lines after CsA treatment was followed by cytochrome c release to the cytosol, reaching an increase of 2.61-fold in LBR-, 1.98-fold in LBR-V160, and 3.01-fold in the case of LBR-D160. However, in the case of PSC 833 treatment, induction of apoptosis in LBR-V160 was associated with mitochondrial depolarization followed by a lower cytochrome c release of 1.15-fold as compared with untreated cells. Caspase 3 activation was clearly observed in LBR-, LBR-V160, and LBR-D160 after CsA treatment, while in LBR-V160, PSC 833 was less effective inducing activation of this caspase. Neither caspase 6 nor 8 activity was observed in these three cell lines. Our results suggest that during CsA- and PSC 833-induced apoptosis, mitochondrial dysfunction occurs. This is mediated through mitochondrial events, associated with an evident decrease in DeltaPsi(m), cytochrome c release and caspase 3 activation.
Asunto(s)
Antineoplásicos/uso terapéutico , Apoptosis/efectos de los fármacos , Caspasas/efectos de los fármacos , Ciclosporina/farmacología , Ciclosporinas/farmacología , Inmunosupresores/farmacología , Leucemia Linfoide/tratamiento farmacológico , Potenciales de la Membrana/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Animales , Caspasas/metabolismo , Resistencia a Múltiples Medicamentos/efectos de los fármacos , Resistencia a Múltiples Medicamentos/genética , Activación Enzimática/efectos de los fármacos , Citometría de Flujo , Leucemia Linfoide/enzimología , Leucemia Linfoide/metabolismo , Ratones , Ratones Endogámicos BALB C , Células Tumorales CultivadasRESUMEN
We have established and characterized a cell line (LBL) from a spontaneous murine T lymphoma LB. Histopathological analysis has demonstrated LB primary tumor infiltration in spleen, lymph nodes, liver, thymus, bone marrow and lung. However LBL cells infiltrated all these organs except lung. Two sublines with different growth behavior were derived from LBL cell line. One of them grew in suspension as clusters (LBLc) while the other one grew as adherent monolayers (LBLa). Growth rate, response to mitogenic stimuli and apoptosis induction were different among the parental cell line and the derived sublines. CD44 was expressed constitutively in LBL and LBLa cells. In contrast LBLc cells only expressed similar levels of this molecule when stimulated with PMA. LBLa cells showed hyaluronic acid (HA) binding properties, while LBL and LBLc cells were not able to bind HA even when activated with PMA. We postulate that differences in HA binding could be related with different infiltration behaviors.(Au)
Asunto(s)
Animales , Ratones , Adyuvantes Inmunológicos/metabolismo , Receptores de Hialuranos/metabolismo , Ácido Hialurónico/metabolismo , Leucemia de Células T/patología , Células Tumorales Cultivadas/patología , División Celular , Intervalos de Confianza , Citometría de Flujo , Leucemia de Células T/metabolismo , Ratones Endogámicos BALB C , Invasividad Neoplásica , Unión Proteica , Células Tumorales Cultivadas/metabolismoRESUMEN
We have established and characterized a cell line (LBL) from a spontaneous murine T lymphoma LB. Histopathological analysis has demonstrated LB primary tumor infiltration in spleen, lymph nodes, liver, thymus, bone marrow and lung. However LBL cells infiltrated all these organs except lung. Two sublines with different growth behavior were derived from LBL cell line. One of them grew in suspension as clusters (LBLc) while the other one grew as adherent monolayers (LBLa). Growth rate, response to mitogenic stimuli and apoptosis induction were different among the parental cell line and the derived sublines. CD44 was expressed constitutively in LBL and LBLa cells. In contrast LBLc cells only expressed similar levels of this molecule when stimulated with PMA. LBLa cells showed hyaluronic acid (HA) binding properties, while LBL and LBLc cells were not able to bind HA even when activated with PMA. We postulate that differences in HA binding could be related with different infiltration behaviors.
Asunto(s)
Animales , Adyuvantes Inmunológicos , Receptores de Hialuranos , Ácido Hialurónico , Leucemia de Células T , División Celular , Intervalos de Confianza , Citometría de Flujo , Leucemia de Células T , Ratones , Ratones Endogámicos BALB C , Invasividad Neoplásica , Unión ProteicaRESUMEN
We have established and characterized a cell line (LBL) from a spontaneous murine T lymphoma LB. Histopathological analysis has demonstrated LB primary tumor infiltration in spleen, lymph nodes, liver, thymus, bone marrow and lung. However LBL cells infiltrated all these organs except lung. Two sublines with different growth behavior were derived from LBL cell line. One of them grew in suspension as clusters (LBLc) while the other one grew as adherent monolayers (LBLa). Growth rate, response to mitogenic stimuli and apoptosis induction were different among the parental cell line and the derived sublines. CD44 was expressed constitutively in LBL and LBLa cells. In contrast LBLc cells only expressed similar levels of this molecule when stimulated with PMA. LBLa cells showed hyaluronic acid (HA) binding properties, while LBL and LBLc cells were not able to bind HA even when activated with PMA. We postulate that differences in HA binding could be related with different infiltration behaviors. (AU)