Mitogen requirements for the in vitro propagation of cutaneous T-cell lymphomas.

We attempted to generate continuous in vitro cultures from patients with mycosis fungoides and the Sezary syndrome (cutaneous T-cell lymphomas, CTCL). Using conventional culture techniques without mitogen stimulation. multiple attempts (32 specimens from 25 patients) failed to replicate T cells. but 6 B-lymphoblastoid cultures were established. Athymic nude mice injected by a variety of routes with CTCL cells from 1 3 patients failed to develop tumors; however. the B-lymphoblastoid cultures were tumorigenic. Lymphocytes from 6 healthy donors and CTCL cells from 7 patients were seeded in growth medium supplemented with one of the following mitogens: phytohemagglutinin (PHA). concanavalin A (Con-A). lymphocyte-conditioned medium (LyCM). pokeweed mitogen (PWM), and staphylococcal protein-A. Normal lymphocytes failed to replicate for more than a few days except in LyCM, where viability and replication remained absolutely dependent on the continued presence of the mitogen. In contrast, mitogen-induced proliferation of CTCL cells was variable, and 1 or more of 4 mitogens induced replication in 4 CTCL cultures. There was partial correlation between the ability of mitogens to stimulate CTCL cells in lymphocyte transformation assays and their ability to act as growth factors. CTCL cells were more resistant to the toxic effects of mitogens than lymphocytes from normal donors, permitting the use of mitogens as long-term growth factors. Two long-term CTCL cultures were established. one after stimulation with Con-A, and the other with LyCM. The cultures required mitogens only for initial growth and have proliferated without mitogens for over a year. The cultures retained many of the features of the CTCL cells present in the patients from whom they were derived, but differed from each other in morphology, E-rosette formation. DNA content, and tumorigenicity. Cytochemical studies provided further evidence of their T-ceIl origin. Both cultures released macrophage inhibitory factor.