A model is proposed for the structure of nuclease-resistant chromatin particles. The model is novel in that it proposes that the DNA in such a particle is wound about a protein core, made up of the hydrophobic regions of histone molecules.

No abstract

No abstract

Recent studies have established the ability ofhuman B lymphocytes to undergo GI-phase cell cycle progression and subsequent DNA synthesis upon exposure to factor(s) present in media conditioned by lectin-stimulated mononuclear cells. Procedures for the isolation of such a cytokine have been the focus of the present investigation. Conditioned medium from cells stimulated by lectin for 72 hr was fractionated by ammonium sulfate precipitation, ion-exchange chromatography, and gel filtration chromatography. During the isolation procedure the proliferation-stimulating activity of the column fractions was assayed concurrently on purified human T cells, purified human B cells, and murine thymocytes. T cell and B cell stimulatory factors present in the initial conditioned medium were found to copurify during ammonium sulfate precipitation, DEAE-Sephadex chromatography, and Bio-Gel P-30 gel filtration. However, partial separation of these two activities was achieved after Bio-Gel P-100 gel filtration. Analytic polyacrylamide gel electrophoresis of radiolabeled Bio-Gel P-100 column fractions demonstrated a distinct protein band of 14,000-15,000 daltons in those column fractions predominantly supporting T cell growth and a distinct protein band of 12,000-13,000 daltons for those fractions predominantly supporting B cell growth. The fractions associated with B cell mitogenic activity induced B cell S-phase entry in a proportion of B lymphocytes in the absence of any detectable IgM secretion.Soluble growth factors for normal epithelial, fibroblastic, hematopoietic, and lymphoid cells have been described and characterized (1). The best delineated lymphoid cell growth factors have been termed interleukin 1 (IL-1) and interleukin 2 (IL-2) (2). These factors have been demonstrated to function in a bimodal amplification network resulting in the proliferative expansion of activated T cells (3,4). The role of soluble factors in the induction of B cell proliferation has also recently been investigated (5-10). Studies have confirmed the capability ofculturing normal, Epstein-Barr virus-negative, B lymphoblastoid cell lines by using exogenously supplied growth-promoting agents (7,8). Investigation into the sources for these B cell growth-promoting agents has revealed that conditioned media derived from several culture systems contains the molecule(s) capable of stimulating B cell proliferation. Both lectin-stimulated normal mononuclear cells (derived from human peripheral blood or murine spleen cell preparations) and antigen-restricted normal helper T cells (grown in the presence of irradiated accessory cells) have been' shown to produce factors capable of supporting B cell growth (5-7). Similar B cell growth-supporting factors have also been observed in conditioned media from phorbol ester-stimulated EL-4 thymoma cells and lectin-stimulated T hybridoma cells (FS6 14.13) (7, 9). Yet it has been well documented that these conditioned media preparations contain multiple functional biological activities. The question that become...

Normal human B lymphocytes, prepared from peripheral venous blood, have been stimulated with intact antiIgM (,u chain specific) bound to an insoluble matrix. The activation event, in a subfraction of human B cells, was associated with subsequent receptivity to the mitogenic effects of exogenously added B-cell growth factor. The ability of the cell population to specifically absorb the B-cell growth factor was dependent upon the time of stimulation with the anti-IgM. Continuous replenishment of the growth factor resulted in the ability to maintain long-term growth-factor-dependent human B-cell populations. These cultured B lymphocytes were shown to specifically absorb the B-cell growth factor, suggesting the presence of membrane receptors for it. The cultured B lymphocytes were routinely maintained in logarithmic-phase growth, in the presence of growth factor, with a population doubling time of 36 hr. These cultured B cells have been utilized in a microassay for the assessment of B-cell growth factor activity that is accurate, sensitive, and precise.Experimentation has demonstrated that an activated human B lymphocyte may be stimulated to enter the S phase of the cell cycle by factor(s) present in media conditioned by lectin-stimulated peripheral blood lymphocytes (1-4). One factor capable of stimulating human B cell proliferation, presently known as B-cell growth factor (BCGF), is a trypsin-sensitive protein with a molecular weight of 12,000-13,000 (5, 6). The protein has a mildly acidic isoelectric point (pH 6.3-6.6), is relatively unstable at 56°C (6, 7), is unaffected by low concentrations of reducing agents, and is stable between pH 3 and pH 8 (7). The human BCGF possesses functional specificity in that its action is strictly proliferative. The factor also has target cell specificity in that its scope is limited to B lymphocytes (3, 5).Further experimentation on many aspects of the biological and biochemical characterization of the growth factor would be enhanced by the establishment of long-term cultures of normal human B cells dependent upon BCGF. In this report we describe a procedure for the polyclonal activation of human B lymphocytes by using anti-IgM (,. chain specific) bound to an insoluble matrix. Anti-immunoglobulin antibodies have previously been utilized in several experimental systems because of their efficacy in human lymphocyte activation; such studies have suggested an important role for the immunoglobulin receptor in B-lymphocyte regulation (8-11). This activation procedure is shown in the present paper to render a subset of cells subsequently responsive to the effects of BCGF and to provide conditions supporting the long-term growth of these B lymphocytes. The long-term BCGF-dependent populations provide an effective system for the sensitive, accurate assessment of BCGF activity and provide evidence for the presence of growth factor receptors on these cells. MATERIALS AND METHODSPreparation of Growth Factors. BCGF-containing growth factor preparations were prepared essentially as d...