RESUMEN
<p><b>BACKGROUND AND OBJECTIVE</b>Recently, the theory of cancer stem cells (CSCs) has presented new targets and orientations for tumor therapy. The major difficulties in researching CSCs include their isolation and purification. The aim of this study is to identify and characterize the side population (SP) cells in small cell lung cancer (SCLC) cell line H446, which lays the foundation for the isolation and purification of CSCs.</p><p><b>METHODS</b>Fluorescence-activated cell sorting (FACS) was used to sort SP and non-SP (NSP) cells from H446. Both subgroups were cultivated to survey the capacity to form into suspended tumor cell spheres. Reverse transcription-polymerase chain reaction (RT-PCR) and real-time PCR were used to evaluate the expression levels of the mRNA of CD133, ABCG2, and nucleostemin in both subgroups. The capacity of proliferation and the differences in drug resistance of both subgroups and unsorted cells were tested by the MTT method. The differentiation ability of both subgroups was determined by FACS. Proliferation was determined by subcutaneous tumor formation in nude mice.</p><p><b>RESULTS</b>The percent of Hoechst 33342 negative cells was about (5.1 +/- 0.2)% in H446 by fluorescence microscopy. The percent of SP cells was (6.3 +/- 0.1)% by flow cytometry. SP cells had a stronger capability of forming into tumor spheres than NSP cells. The mRNA expression levels of ABCG2, CD133, and nucleostemin in SP cells were 21.60 +/- 0.26, 7.10 +/- 0.14, and 1.02 +/- 0.08 folds higher than that in NSP cells (P < 0.01, P < 0.01, and P > 0.05, respectively). In vivo, SP cells showed better proliferative ability and tougher viability when treated with drugs. SP cells can differentiate into NSP cells, but NSP cells cannot differentiate into SP cells. SP cells had a greater ability to form tumors.</p><p><b>CONCLUSION</b>The H446 cell line contained some SP cells with stem cell properties. CD133 and ABCG2 may be cancer stem cell markers of SCLC.</p>
Asunto(s)
Animales , Humanos , Masculino , Ratones , Antígeno AC133 , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2 , Transportadoras de Casetes de Unión a ATP , Genética , Metabolismo , Antígenos CD , Genética , Metabolismo , Biomarcadores de Tumor , Metabolismo , Diferenciación Celular , Línea Celular Tumoral , Proliferación Celular , Transformación Celular Neoplásica , Proteínas de Unión al GTP , Genética , Metabolismo , Glicoproteínas , Genética , Metabolismo , Neoplasias Pulmonares , Metabolismo , Patología , Ratones Desnudos , Proteínas de Neoplasias , Genética , Metabolismo , Células Madre Neoplásicas , Metabolismo , Patología , Trasplante , Proteínas Nucleares , Genética , Metabolismo , Péptidos , Genética , Metabolismo , ARN Mensajero , Metabolismo , Células de Población Lateral , Metabolismo , Patología , Trasplante , Carcinoma Pulmonar de Células Pequeñas , Metabolismo , PatologíaRESUMEN
This study was aimed to investigate the effects of rituximab (RTX), a chimeric human anti-CD20 monoclonal antibody, on lymphoma cell injury induced by X ray irradiation. The human Burkitt EBV-infected and moderate radioresistance lymphoma cells (Namalwa) were used in the this study. Cytotoxicity of rituximab combined with X ray irradiation on Namalwa cells was measured by sulforhodamine B (SRB)-staining; the apoptosis of Namalwa cells was detected by flow cytometry with FITC-Annexin V/PI double staining; the morphologic changes of cells were observed under transmission electron microscope (TEM) and the change of intracellular free calcium level ([Ca(2+)]i) in response to irradiation and rituximab was determined by means of the fluorescent dye fluo-3 and confocal microscopy. The results showed that the growth inhibition in Namalwa cells exposed to irradiation was enhanced by treatment with rituximab. Compared with irradiation alone, rituximab combined with irradiation significantly induced the cell apoptosis and a sustained rise of intracellular free calcium ([Ca(2+)]i) level in Namalwa cells; the serial apoptotic appearances of cells could be observed under TEM. It is concluded that rituximab can enhance the sensitivity of lymphoma cells on X ray irradiation as to induce cell more apoptosis, in this process the intracellular free calcium ([Ca(2+)]i), as an intracellular signaling molecule probably plays an important role.