Interferons alpha and beta as cellular regulatory molecules.
Explore the source record for details and available documents.
Animals↗
Biomedical subjects
L M Pfeffer
Publications and source records attributed to L M Pfeffer.
Effect of anti-Ig on cytosolic Ca2+ in Daudi lymphoblastoid cells.
We examined the response in the free intracellular calcium concentration ([Ca2+]i) of Daudi (human lymphoblastoid) cells to antibodies against human immunoglobulins (anti-Ig), and the relationship of [Ca2+]i to anti-Ig-induced capping. At 80 microM intracellular quin-2 (a fluorescent probe for [Ca2+]i), anti-Ig (10 micrograms/ml) caused a rapid increase in [Ca2+]i from 100 to 600 nM; the signal returned to baseline with approximately 1 min. At 450 microM intracellular quin-2, [Ca2+]i rose to only approximately 250 microM, and the signal declined gradually, returning to base line after greater than 7 min. In subsequent experiments, the lower concentrations of quin-2 were employed. Plots of the amplitude of the [Ca2+]i transients and of the binding of 125I-anti-Ig to Daudi cells versus the concentrations of anti-Ig showed similar saturation kinetics, with half-saturation occurring at 2-3 micrograms/ml. Part of the calcium in the transient is derived from the extracellular medium, and part from the nonmitochondrial intracellular stores. Caffeine (4 mM) and 8-(diethylamino)octyl 3,4,5-trimethoxybenzoate HCl (0.5 mM) suppressed the release of calcium from internal stores and the entry of calcium from outside the cells, but permitted capping in more than half of the cells. Phorbol esters (1-2 nM) inhibited both capping and the anti-Ig-induced decrease in [Ca2+]i. None of these agents blocked the binding of anti-Ig to the cells. It appears that receptor capping is not dependent on the anti-Ig-induced transient increase in calcium concentration.
Aminoquinolines↗
Interferon-beta inhibition of concanavalin A-stimulated calcium uptake and exchange in HeLa cells.
Addition of the lectin concanavalin A (ConA) to HeLa-S3 tumor cells in culture stimulates calcium exchange and uptake. Pretreatment of the cells with interferon-beta (IFN-beta) at a concentration of 640 U/ml for 24 h markedly inhibits this ConA-stimulated calcium exchange and uptake, but not the basal level of calcium exchange. These findings suggest that a number of IFN effects on cell function may reflect an impaired ability of IFN-treated cells to respond to molecules such as growth factors and peptide hormones with increased calcium fluxes.
Calcium↗
Interferon stimulates cholesterol and phosphatidylcholine synthesis but inhibits cholesterol ester synthesis in HeLa-S3 cells.
Treatment of human HeLa-S3 cells (an epidermoid carcinoma line) with human beta-interferon (640 units/ml) selectively alters lipid metabolism by increasing cholesterol synthesis per mg of cell protein as measured by 1-hr pulse-labeling of cells with [3H]acetate. Cholesterol synthesis in interferon-treated cells is increased approximately equal to 60% at 24 hr after the beginning of treatment and approximately equal to 450% at 48 hr. Continuous labeling of interferon-treated cells with [14C]acetate shows increased accumulation of label in cholesterol when normalized per mg of cell protein, as well as an increase in the specific activity of cholesterol in the treated cells. In contrast, interferon treatment decreases the accumulation of [14C]acetate into cholesterol esters. The [14C]acetate labeling of sphingomyelin, phosphatidylethanolamine, and triglycerides shows no change compared to untreated controls. The labeling of phosphatidylcholine was moderately increased in treated cells. The interferon-induced changes in lipid metabolism are a part of a coordinated response of cells to interferon treatment, characterized by reduced cell proliferation and cell motility and an increase in cell size and mass. The increased cholesterol synthesis is consistent with a model in which beta-interferon treatment of HeLa cells inhibits the endocytosis of cholesterol-containing low density lipoprotein, which results in an increase in cholesterol synthesis.
Acetates↗
Effects of beta interferon on human fibroblasts at different population doubling levels. Proliferation, cell volume, thymidine uptake, and DNA synthesis.
Cellular aging had no effect on the ability of beta interferon to increase cell volume and population doubling time in 76-109 cells, a line of human skin fibroblasts. However, DNA synthesis in cells at high population doubling levels (PDL 55-70) was inhibited after 72 h of beta interferon treatment (1,000 U/ml) while no inhibition of DNA synthesis was observed in cells at middle population doubling levels (PDL 30-40).
Cell Division↗
Interferon inhibition of thymidine incorporation into DNA through effects on thymidine transport and uptake.
Replenishment of medium after 72 hr of growth of HeLa-S3 cells in dense suspension cultures increased [3H]-thymidine uptake into cells and incorporation into DNA, with the levels reaching a peak approximately 12 hr following medium change; beta interferon inhibits the enhanced uptake of [3H]-thymidine and labeling of DNA in a dose-dependent manner. Some reduction in these processes is observed at a concentration as low as 1 u/ml, and approximately 75% inhibition at 640 u/ml. Kinetic analysis has revealed that the rate of labeling of the acid-soluble pool with [3H]-thymidine, measured either at 22 degrees C or 37 degrees C, is reduced in interferon-treated (640 u/ml, 24 hr) HeLa-S3 cells. At 22 degrees C, the initial rate of thymidine transport at a high (500 microM) thymidine concentration, determined within the first 30 sec of [3H]-thymidine addition was depressed by 44% in interferon-treated HeLa cells. At 37 degrees C, labeled precursors accumulate in acid-soluble material for approximately 8 min after the addition of [3H]-thymidine, after which an apparent equilibrium level is attained. At this temperature, the rate of thymidine uptake and the apparent equilibrium level attained were depressed by 70% in interferon-treated HeLa cells. The reduced incorporation of [3H]-thymidine into DNA in interferon-treated HeLa-S3 cells can be largely explained by interferon inhibition of thymidine transport and phosphorylation.
Biological Transport↗
Interferon suppresses pinocytosis but stimulates phagocytosis in mouse peritoneal macrophages: related changes in cytoskeletal organization.
Treatment of thioglycolate-elicited macrophages with mouse beta-interferon markedly reduces pinocytosis of horseradish peroxidase and fluorescein isothiocyanate (FITC)-dextran but stimulates phagocytosis of IgG-coated sheep erythrocytes. Experiments with FITC-dextran have revealed that the overall decrease in pinocytosis is due to a nearly complete inhibition of pinocytosis in a large fraction of interferon-treated macrophages. In the remaining cells pinocytosis continues at a rate similar to that in untreated control cells. A considerable reduction in the number of cells pinocytosing FITC-dextran was observed within 12 h from the beginning of interferon treatment. Measurement of the overall level of pinocytic activity with horseradish peroxidase showed a progressive decline through 72 h of treatment. In the interferon-sensitive subpopulation, there were marked changes in cytoskeletal organization. Microtubules and 10-nm filaments were aggregated in the perinuclear region while most of the peripheral cytoplasm became devoid of these cytoskeletal structures as observed by fluorescence and electron microscopy. In addition, interferon treatment of macrophages appeared to disrupt the close topological association between bundles of 10-nm filaments and organelles such as mitochondria, lysosomes, and elements of the Golgi apparatus and endoplasmic reticulum. Such alterations in the distribution of microtubules and 10-nm filaments were not seen in the interferon-insensitive subpopulation. We have investigated the mechanism of the interferon-induced enhancement of phagocytic activity by binding IgG-coated sheep erythrocytes to mouse peritoneal macrophages at 4 degrees C and then initiating a synchronous round of ingestion by warming the cells to 37 degrees C. Thioglycolate-elicited macrophages that had been treated with mouse beta-interferon ingested IgG-coated erythrocytes faster and to a higher level than control cells in a single round of phagocytosis. In interferon-treated cultures, phagocytic cups became evident within 30 s of the shift of cultures from 4 degrees to 37 degrees C, whereas in control cultures, they appeared in 2 min. Cytochalasin D, an inhibitor of actin assembly and polymerization, abolished phagocytic activity in both control and beta-interferon-treated macrophages. However, to inhibit phagocytosis completely in thioglycolate-elicited interferon-treated macrophages, twice as much cytochalasin D was required in the treated as in control cultures.(ABSTRACT TRUNCATED AT 400 WORDS)
Actins↗
Growth rate of control and beta-interferon-treated human fibroblast populations over the course of their in vitro life span.
The estimated growth rates for five lines of human fibroblasts (Human Genetic Mutant Cell Repository designations 0011, 2936B , 0038A , 2912A , and 3529), during the first few population doublings after establishment of the lines, were within the expected range, i.e., equivalent to doubling times from 16 to 28 hr, with a mean of 20 hr. The lines were derived from donors aged 8 fetal weeks, 20 days, 9 years, 26 years, and 66 years, respectively. The growth rates of the five lines declined as an exponential function of the population doubling level in all cases. The rate of decline of the growth rate varied for different lines and appeared to be related to the life span of the lines, which in turn was related to donor age. After 30 population doublings, the population doubling times had increased 1.3, 1.3, 1.7, 3.4, and 4.7 times for the five cell lines of the corresponding replicative life spans of 65, 57, 56, 31, and 28 population doublings. Sensitivity of the fibroblast lines to the cell growth-inhibitory effect of beta-interferon was independent of the population doubling level of the lines, i.e., interferon depressed the population growth rate of low population doubling level cells as much as that of middle or late population doubling level cells for any given line. However, the fibroblast lines showed differences in their sensitivity to the cell growth-inhibitory effect of interferon, probably as an expression of genotypic differences among the lines.
Adult↗
Comparison of the effects of alpha and beta interferons on the proliferation and volume of human tumor cells (HeLa-S3, Daudi, P3HR-1).
Treatment of three lines of tumor cells (epidermoid carcinoma: HeLa-S3, and Burkitt's lymphoma: Daudi and P3HR-1) with human alpha or beta interferon (HuIFN-alpha or HuIFN-beta) results in inhibition of cell proliferation as well as in increased modal cell volume and heterogeneity of the populations with respect to cell volume. In all three cell lines the IFN-alpha- or IFN-beta-induced changes in cell proliferation and in cell volume are inversely related to each other. However, these lines differ significantly in their sensitivity to the effects of IFN-alpha and IFN-beta.
Burkitt Lymphoma↗
Effects of beta interferon on concanavalin A binding and size of HeLa cells.
Treatment of HeLa-S3 cells with 640 u/ml of human beta interferon (HuIFN-beta) in suspension culture increases the cell volume and the heterogeneity of the population with respect to cell size, as measured by Coulter electronic volume analysis. Based on such analysis, the cell surface area is increased 11% by 24 h, and 42% by 48 h after beginning of interferon treatment. Binding of [125I] Concanavalin A (measured at 22 degrees C) is increased 75% by 24 h, and 120% by 48 h on a per-cell basis. It follows that interferon treatment for 24 h causes a 58% increase in the abundance of functional Con A receptor sites per unit area of cell surface. The further increase in Con A binding after treatment of cells with interferon for 48 h can be attributed to increased surface area of cells.
Cell Membrane↗
Assay of the inhibitory activities of human interferons on the proliferation and locomotion of fibroblasts.
Explore the source record for details and available documents.
Animals↗
Assays to measure plasma membrane and cytoskeletal changes in interferon-treated cells.
Explore the source record for details and available documents.
Cell Line↗
Interferon increases the abundance of submembranous microfilaments in HeLa-S3 cells in suspension culture.
Human beta (fibroblast) interferon inhibits the proliferation of human HeLa-S3 carcinoma cells in suspension culture. Accompanying this effect, the lateral mobility of cell surface receptors for concanavalin A is decreased and the rigidity of the plasma membrane lipid bilayer is increased. The present findings show a marked increase in the number of polymerized actin-containing microfilaments 3 days after treatment of HeLa-S3 cells with beta-interferon (640 units/ml). The cortical region of the treated enlarged cells contains a thick and dense meshwork of 40-70 A microfilaments. The actin nature of the filaments was verified by their ability to bind heavy meromyosin. These results support the concept that beta-interferon induces a coordinated response in the plasma membrane and the underlying microfilaments in both tumor and normal cells.
Cell Membrane↗
Beta-interferon-induced time-dependent changes in the plasma membrane lipid bilayer of cultured cells.
The time-dependent interferon-induced structural changes in the plasma membrane lipid bilayer have been investigated in cultured cells using spin label electron spin resonance techniques. Treatment of human HeLa-S3 cells in suspension culture with human beta 1 interferon (640 u/ml) for as short a time as 30 min causes an increase in the rigidity of the plasma membrane lipid bilayer (Landsberger, Pfeffer and Tamm, in preparation). The plasma membrane rigidity of interferon-treated cells returns to the level of control cells within 3 to 5 hr, but by 24 hr after beginning of treatment, the rigidity of the plasma membrane is increased again and remains so for at least 2 days. Mouse beta interferon causes both an early (Landsberger, Pfeffer and Tamm, in preparation) and a late increase in membrane rigidity in homologous mouse L-929 cells, but not in heterologous HeLa-S3 cells. Thus, the interferon-induced perturbation of membrane structure is species specific. The late increase in membrane rigidity may have a different underlying mechanism from that of the early increase. While the early and transient change probably is related to signal generation and transmission, the later and persistent change may be an aspect of the phenotype and interferon-treated cells and may reflect changes in the plasma membrane-cytoskeletal complex.
Cell Membrane↗
Interferon inhibits the redistribution of cell surface components.
Interferon treatment impairs the ability of cells to redistribute cell surface receptors for concanavalin A (Con A). The effect of interferon becomes evident within 3-6 h and is maximal within 36-48 h. Highly purified human fibroblast interferon (> 2 x 10(8) U/mg of protein sp act; concentration; 640 U/ml) caused approximately 85% inhibition of capping of fluorescein-conjugated Con A in interferon-sensitive HeLa-S3 cells at 36 h from the beginning of treatment.
Cell Membrane↗
Interferon effects on microfilament organization, cellular fibronectin distribution, and cell motility in human fibroblasts.
We have shown previously (Pfeffer et al., 1979, Exp. Cell Res. 121:111-120) that treatment of human fibroblasts, planted at a density of 2x10(3) cells/cm(2), with purified human fibroblasts interferon (640 U/ml) for 3 d at 37 degrees C decreases the overall rate of cell proliferation to 35-40 percent of the control value. In the present experiments we have characterized the phenotype of interferon-inhibited fibroblasts. The mean volume of trypsinized, interferon-treated cells was increased 31 percent abover that of control cells. The interferon-treated population was much more heterogeneous than the control population with respect to volume, and there was a considerable overlap in the volume distributions of the two populations. The cell surface area was, on the average, increased 65 percent after interferon treatment. More than 80 percent of the treated cells had enlarged nuclei, many of which were lobed, and the fraction of binucleated cells was increased fivefold. After interferon treatment, over 40 percent of the cells showed large actin-containing fibers in the form of multiple parallel arrays. Fewer than 5 percent of the control cells contained such large actin fibers. The number of actin fibers of all sizes was tripled in the treated fibroblasts on a per cell basis and, calculated per unit surface area of the cells, the number was increased 82 percent. In contrast, 10-nm filaments and microtubules did not appear to be increased in number per unit surface area of the cells. The increases per cell in the abundance of these structures were directly related to increased cell size. After interferon treatment, fibronection was distributed in arrays of long filaments covering most portions of the cell surface. Interferon treatment markedly decreased the rate of cell locomotion as well as membrane ruffling and saltatory movements of intracellular granules.
Cell Line↗
Growth characteristics of human skin fibroblasts in vitro: a simple experimental approach for the identification of hereditary adenomatosis of the colon and rectum.
Kinetic parameters of cell growth have been studied in cultured skin fibroblasts derived from individuals with hereditary adenomatosis of the colon and rectum. The growth parameters consisted of saturation density, plating density, cloning efficiency and colony-forming ability. The results suggest deployment of these selective assays to distinguish skin fibroblasts of presumptive ACR positive individuals from those of normal subjects in vitro.
Adenoma↗
Interferon effects on the growth and division of human fibroblasts.
Explore the source record for details and available documents.
Cell Division↗