ABSTRACT
Background: The conventional cytogenetic approach to demonstrate Philadelphia (Ph) chromosome at times does not yield enough number of metaphases or are of suboptimal quality. Further, the rapid molecular tests have completely pushed this simple technique into disrepute. Aims: This study aimed to evaluate usefulness of phytohemagglutinin (PHA)-stimulated peripheral blood culture for detection of Ph chromosome in chronic myeloid leukemia (CML) patients. Materials and Methods: Fifty-six patients, including 11 newly diagnosed cases of CML and 45 patients of CML on imatinib therapy showing the presence of Ph chromosome in unstimulated samples, were included in the study. Cytogenetic analysis was done on unstimulated samples, i.e. bone marrow aspirate, 24- and 48-h peripheral blood culture, and compared with PHA-stimulated 72-h peripheral blood culture. Results: The preparations from PHA-stimulated peripheral blood culture samples in all 56 patients yielded high number of good-quality metaphases. All the 11 (100%) newly diagnosed patients and 39/45 (87%) of the patients on imatinib therapy showed the presence of Ph chromosome in PHA-stimulated samples. Addition of PHA-stimulated 72-h peripheral blood culture preparation can be of use for increasing the diagnostic yield in cases of CML with suboptimal results on conventional cytogenetics from bone marrow aspirate sample.
Subject(s)
Adult , Humans , Karyotyping/methods , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/diagnosis , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/pathology , Leukocytes/drug effects , Male , Middle Aged , Philadelphia Chromosome , Phytohemagglutinins/metabolismABSTRACT
Phytohemagglutinin (PHA) is known to increase the synthesis of bovine leukemia virus (BLV) particles and viral antigens in short-term culture of BLV-infected neoplastic and non-neoplastic lymphocytes. This stimulation of BLV expression has been shown to be due to enhanced transcription of the viral genome by a PHA-induced protein. We have investigated the binding of 125I-labelled PHA to BLV-infected bovine B lymphocytes and subsequent events that may lead to the stimulation of BLV-p25 synthesis. We found that PHA binding to the infected cells were rapid, but only a small fraction of the bound PHA is translocated to nucleus. However, bound PHA dissociated rapidly from the cell membrane, but not from the nucleus when PHA is removed from the culture medium. Furthermore, continuous presence of PHA was not essential for optimal stimulatory activity, instead a minimum incubation with PHA for 6 hr. followed by culturing the infected cells in its absence, was sufficient to exhibit maximal stimulatory activity. Our results raise the possibility that interaction of PHA with the plasma membrane probably triggers synthesis of a protein which in turn enhances the transcription of BLV genome in vitro.
Subject(s)
Animals , B-Lymphocytes/metabolism , Cattle , Gene Expression Regulation, Viral , Leukemia Virus, Bovine/genetics , Phytohemagglutinins/metabolism , Retroviridae/genetics , Retroviridae Proteins, Oncogenic/biosynthesisABSTRACT
El método de rosetas con eritrocitos de pollo recubiertos de IgG se ha utilizado junto con separación por flotación en ficoll-hypaque para obtener subpoblaciones de células enriquecidas o disminuidas de linfocitos T con receptor para Fc de IgG (Tg). Estas poblaciones celulars se han utilizado para determinar producción de LIF a diferentes concentraciones de los mitógenos, utilizando un método en microgota de agarosa. De esta manera, hemos podido observar una respuesta semejante en las diferentes poblaciones celulares hacia concanavalina A y una respuesta diferencial hacia fitohemaglitinina donde por lo general se observa una respuesta negativa de la población enriquecida con células Tg mientras que las poblaciones de linfocitos totales o aquellas en las que se han eliminado las Tg dan una respuesta positiva al segundo mitógeno