RESUMEN
We performed aCGH, SKY/FISH, molecular mapping and expression analyses on a permanent CD8+ NK/T cell line, 'SRIK-NKL' established from a lymphoma (ALL) patient, in attempt to define the fundamental genetic profile of its unique NK phenotypes. aCGH revealed hemizygous deletion of 6p containing genes responsible for hematopoietic functions. The SKY demonstrated that a constitutive reciprocal translocation, rcpt(5;14)(p13.2;q11) is a stable marker. Using somatic hybrids containing der(5) derived from SRIK-NKL, we found that the breakpoint in one homologue of no. 5 is located upstream of IL7R and also that the breakpoint in no. 14 is located within TRA@. The FISH analysis using a BAC which contains TRA@ and its flanking region further revealed a approximately 231kb deletion within 14q11 in the der(5) but not in the normal homologue of no. 14. The RT-PCR analysis detected mRNA for TRA@ transcripts which were extending across, but not including, the deleted region. IL7R was detected at least at mRNA levels. These findings were consistent with the immunological findings that TRA@ and IL7R are both expressed at mRNA levels and TRA@ at cytoplasmic protein levels in SRIK-NKL cells. In addition to rcpt(5;14), aCGH identified novel copy number abnormalities suggesting that the unique phenotype of the SRIK-NKL cell line is not solely due to the TRA@ rearrangement. These findings provide supportive evidence for the notion that SRIK-NKL cells may be useful for studying not only the function of NK cells but also genetic deregulations associated with leukemiogenesis.
Asunto(s)
Antígenos CD8/metabolismo , Mapeo Cromosómico , Hibridación Fluorescente in Situ , Células Asesinas Naturales/metabolismo , Hibridación de Ácido Nucleico , Cariotipificación Espectral , Linfocitos T/metabolismo , Línea Celular , Rotura Cromosómica , Cromosomas Humanos Par 14 , Cromosomas Humanos Par 5 , Análisis Citogenético , Humanos , Reacción en Cadena de la Polimerasa , Translocación GenéticaRESUMEN
T-cells and NK cells arise from common pluripotent stem cells, with shared early developmental pathway and surface markers. Distinguishing between them is becoming difficult, but critical for study. A large family of NK cells, including classical NK, NK-T, and NK-CTL exists. Natural cytotoxicity receptors (NKp46, NKp44, NKp30) have been proposed as specific for classical NK cells, but were expressed at protein and mRNA level by CD8+NK/T cell line SRIK-NKL, suggesting more widespread expression. We investigated and found expression of these markers at the protein and mRNA level in multiple human cell lines.
Asunto(s)
Linfocitos T CD8-positivos/metabolismo , Regulación de la Expresión Génica/fisiología , Células Madre Hematopoyéticas/metabolismo , Células Asesinas Naturales/metabolismo , Glicoproteínas de Membrana/biosíntesis , Receptores Inmunológicos/biosíntesis , Biomarcadores/metabolismo , Linfocitos T CD8-positivos/citología , Línea Celular , Células Madre Hematopoyéticas/citología , Humanos , Células Asesinas Naturales/citología , Glicoproteínas de Membrana/genética , Receptor 1 Gatillante de la Citotoxidad Natural , Receptor 2 Gatillante de la Citotoxidad Natural , Receptor 3 Gatillante de la Citotoxidad Natural , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Receptores Inmunológicos/genéticaRESUMEN
The distinction between T cells and NK cells is difficult, and becoming more complex, as the diversity of the human lymphocyte repertoire is evident. We report the establishment of a permanent CD8+ NK/T cell line (SRIK-NKL) from a patient with leukemic phase of acute lymphoblastic lymphoma having characteristics of both NK and T cells, and extensively describe its phenotype, including cytotoxic activity, NK cell receptor expression, and other molecules critical for immune function. We further compare SRIK-NKL to other available NK/NK-T cell lines. SRIK-NKL may be useful for studying NK cell development, functions, and modulation, leading to novel strategies for treatment of autoimmune disease, infection, and cancer.