Lymphocyte and Fibroblast Chalones: Some Chemical Properties

Houck, J.C.; Kanagalingam, K.; Hunt, Carl V.; Attallah, Abdelfattah M.; Chung, Anthony

doi:10.1126/science.140460

Cited by 33 publications

(7 citation statements)

References 7 publications

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“…The existence of a number of growth inhibitors has been postulated (1,2), and growth-inhibitory fractions have been obtained from various mammalian cells and tissues (3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14). Nevertheless, there are doubts as to the significance of endogenous growth inhibitors because most of the inhibitor preparations have low specific activity, the inhibitors have been difficult to purify, and with some preparations the causes of growth inhibition have, at least in part, turned out to be trivial (15)(16)(17).…”

mentioning

confidence: 99%

Purification of kidney epithelial cell growth inhibitors.

Holley¹,

Bőhlen²,

Fava³

et al. 1980

Proc. Natl. Acad. Sci. U.S.A.

154

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Two high molecular weight growth inhibitors have been isolated from the culture medium of BSC-1 cells, epithelial cells of African green monkey kidney. The purified kidney epithelial cell growth inhibitors, at ng/ml concentrations, reversibly arrest the growth of BSC-1 cells in the GI phase of the cell cycle. Their action is selective; they are most active on BSC-1 cells, are less active as inhibitors of the growth of rat lung and human breast epithelial cells, and do not inhibit the growth of 3T3 mouse embryo fibroblasts and human skin fibroblasts in culture. Their growth inhibitory action on BSC-1 cell cultures is counteracted by epidermal growth factor or calf serum.One of the areas of ignorance that remains in our broad understanding of the control of growth of mammalian cells is the subject of endogenous growth inhibitors. The existence of a number of growth inhibitors has been postulated (1, 2), and growth-inhibitory fractions have been obtained from various mammalian cells and tissues (3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14). Nevertheless, there are doubts as to the significance of endogenous growth inhibitors because most of the inhibitor preparations have low specific activity, the inhibitors have been difficult to purify, and with some preparations the causes of growth inhibition have, at least in part, turned out to be trivial (15-17). There is no doubt that highly active growth inhibitors exist for mammalian cells, because interferon and corticotropin are extremely active as growth inhibitors in certain cell cultures (18,19), but the inhibitory activities of these two substances could be incidental to their other biological activities. Because of the potential significance of growth inhibitors and because purified inhibitors could be very useful in the manipulation of growth of mammalian cells, we have undertaken further studies in this area.Density-dependent regulation of growth of BSC-1 cells (African green monkey kidney epithelial cells) has been found to result in part from the accumulation of growth inhibitors in the culture medium (20). The present paper describes the isolation of high molecular weight growth inhibitors from the culture medium of these cells. The growth inhibitors have very high specific activities, approximately equal to the specific activities of interferon and corticotropin in their respective cell culture assays. The new growth inhibitors appear to be selective in the types of cells on which they act. MATERIALS AND METHODSCell Cultures. BSC-1 cell stock cultures were maintained as previously described (21) in Dulbecco's modified Eagle's medium (DME medium) (22) Preparation of Conditioned Medium. Dense BSC-1 cell cultures, used for preparation of conditioned medium, were grown in 15-cm Falcon tissue culture plates in DME medium with 1% calf serum. The medium was changed once a week until the cells reached a density of approximately 1.5 X 105 cells per cm2. The cultures were then maintained in DME medium with 0.1% calf serum. To prepare conditioned medium, the cultu...

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mentioning

confidence: 99%

Purification of kidney epithelial cell growth inhibitors.

Holley¹,

Bőhlen²,

Fava³

et al. 1980

Proc. Natl. Acad. Sci. U.S.A.

154

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“…An (1978) described which binds avidly to RNA (11). This sse differs from ours in being readily inactivated by proteases, and it is also active on phytohemagglutinin-stimulated human lymphocytes which are known to be thiol independent.…”

Section: Discussionmentioning

confidence: 65%

Production of autoinhibitory factors by mouse lymphoma cells in vitro and its relationship to thiol dependence.

Tanapat¹,

Gaetjens²,

Broome³

1978

Proc. Natl. Acad. Sci. U.S.A.

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Five different kinds of mouse lymphoma cells conditioned culture medium so that it became inhibitory to thiol-dependent cells but not to thiol-independent cells. Conditioning was rapid and appeared to be complete in 1-3 hr. The inhibitory effects could be reversed by concentrations of 2-mercaptoethanol higher than those usually required for growth promotion. Inhibitory factors were concentrated by ultrafiltration and chromatography on Sephadex G-10 which showed their apparent molecular weights to be 300-600. The factors were stable at 1000 and to proteases but were inactivated (after reduction) by iodoacetamide, indicating that they may be thiols or disulfides.Specific thiols and their corresponding disulfides promote the survival and replication in vitro of mouse lymphoid cells, a property which is retained in numerous lymphoma lines (1-4). Such substances, however, are either nonphysiological (particularly 2-mercaptoethanol which has been widely used in experimental systems) or are required at concentrations much greater than occur in vivo. On the other hand, thiol is not required by lymphoma cells when they are cocultivated with macrophages or other cells but, after separation, the supernatant medium does not continue to support growth (5, 6). The mechanism of these effects and their relationship to conditions in vio thus far have not been explained.We here present evidence that thiol-dependent lymphoma cells in vitro are subject to autoinhibition mediated through substances that they liberate into the medium. These have been partially characterized and appear to be thiols or disulfides. Growth promoting thiols (e.g., 2-mercaptoethanol) or cocultivated macrophages interrupt the autoinhibitory system, and these actions may therefore be significant for restoring cell proliferation. The experiments to be described were prompted by observations on the growth promoting effect of macrophages in cocultivation with lymphoma cells (6). Numerous attempts failed to show evidence that macrophages produced a stable conditioning of the medium, and an alternative explanation, that these cells might reverse an autoinhibition by L1210(V) cells, was therefore considered.In most of our experiments we used cells of the thiol-dependent lymphoma L1210(V), whose properties we compared with those of its thiol-independent variant L1210(A) (7). Three other cell lines were examined-EL4, RADA1, and ERLDand in these the same relationships between autoinhibition and thiol dependence were found. MATERIALS AND METHODSTumors. B6D2F1 mice were used to carry the following tumors in ascitic form: L1210(V), L1210(A), and EL4. ERLD (i) Supernatants were concentrated up to 20 times by using a UM05 or other ultrafiltration membrane (Amicon Co., Lexington, MA). Aliquots from these concentrated solutions were tested for their ability to support growth of the various cell lines in tissue culture.(ii) Supernatants were lyophilized to dryness and the residues were dissolved in water and chromatographed on Sephadex G-10.Measurement of Lymphoma Ce...

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“…endog enous inhibitor of lymphocyte response in vitro (lectin and alloantigcn stimulation) and in vivo (skin allograft rejection). Further studies with the ultrafiltrate from calf thymus [6,7] showed that the inhibitory activity could be converted to low molecular weight by treatment of the ultrafiltrates with ribonu cléase or low pH. These results suggest that the inhibitor was bound to high molecular weight carrier molecules.…”

Section: Introductionmentioning

confidence: 76%

Low Molecular Weight Inhibitors of Lymphocyte Transformation

Patt¹,

Gleisner²,

Barrantes³

et al. 1981

Pharmacology

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A variety of factors isolated from bovine thymus have been found to inhibit the transformation of human and mouse lymphocytes. The majority of this activity fractionates as low molecular weight material by ultrafiltration or column chromatography. Three distinct fractions of low molecular weight have been isolated. One fraction contains the polyamines spermine and spermidine. A second fraction contains thymidine or thymidine-like nucleotides. The third fraction appears to be polypeptide in nature, and gives an estimated molecular weight of 500–600, is heat and pH stable, and is easily extracted by solutions containing organic solvents. Preliminary steps in the isolation of this inhibitor are presented, and its relationship to other immunosuppressive and anti-mitotic agents is discussed.

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Lymphocyte and Fibroblast Chalones: Some Chemical Properties

Cited by 33 publications

References 7 publications

Purification of kidney epithelial cell growth inhibitors.

Purification of kidney epithelial cell growth inhibitors.

Production of autoinhibitory factors by mouse lymphoma cells in vitro and its relationship to thiol dependence.

Low Molecular Weight Inhibitors of Lymphocyte Transformation

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