RESUMEN
BACKGROUND: Adoptive immunotherapy with cytotoxic T cells has shown promising clinical results in patients with metastatic melanoma and post-transplant-associated viral infections. Cell transfer therapies often require the ex vivo expansion of large numbers of reactive lymphocytes. Therefore interleukin-2 (IL-2), a potent T-cell mitogenic cytokine that critically affects the features and effectiveness of T cells, is frequently added to cell culture media. METHODS: We examined the influence of various IL-2 concentrations on cell growth, cytotoxicity, cytokine release and surface marker expression of tumor-infiltrating lymphocytes (TIL) during a standard 14-day rapid expansion phase. The study was conducted under good manufacturing practice (GMP) conditions, using approved reagents in a class 10000 laboratory. RESULTS: T-cell cultures grown in very high IL-2 concentrations (600-6000 IU/mL) expanded massively and maximally secreted interferon (IFN)-gamma in response to antigenic stimulation, but exhibited only low direct cytotoxicity. On the other hand, TIL cultures grown in low concentrations of IL-2 throughout the rapid expansion phase expanded to a lower extent and barely secreted IFN-gamma but displayed high cytotoxic activity. A combined approach of starting with 10-120 IU/mL IL-2 during the first week, followed by increasing the IL-2 concentration to 6000 IU/mL during the second week, results in T cells that expand well, maximally produce IFN-gamma and are highly cytotoxic against tumor cells. DISCUSSION: Fine tuning of the IL-2 concentration during ex vivo expansion of T cells can yield high numbers of T cells with optimal features for clinical use.
Asunto(s)
Inmunoterapia Adoptiva , Interleucina-2/metabolismo , Linfocitos Infiltrantes de Tumor/patología , Melanoma/patología , Linfocitos T Citotóxicos/patología , Antígenos de Diferenciación/biosíntesis , Antígenos de Neoplasias/metabolismo , Proliferación Celular , Técnicas de Cocultivo , Medios de Cultivo , Citotoxicidad Inmunológica , Humanos , Interferón gamma/metabolismo , Interleucina-2/inmunología , Activación de Linfocitos , Linfocitos Infiltrantes de Tumor/inmunología , Linfocitos Infiltrantes de Tumor/metabolismo , Melanoma/inmunología , Melanoma/metabolismo , Metástasis de la Neoplasia , Linfocitos T Citotóxicos/inmunología , Linfocitos T Citotóxicos/metabolismoRESUMEN
The effect of an inhibitor of IL-1, purified from a human myelomonocytic cell line (M20) on the development of human erythroid cell development was studied. The inhibitor, is a protein of 52 kD molecular weight that is distinct immunologically and functionally from other reported IL-1 inhibitors. The experiments were performed in a two-phase culture system that allows separation of the erythroid cell development into an erythropoietin (EPO)-independent phase, where early erythroid-committed BFUe proliferate and differentiate into the more mature progenitors, CFUe, and EPO-dependent phase, where CFUe further proliferate and mature into hemoglobin-containing orthochromatic normoblasts. The results indicated that in both developmental stages the M20-derived inhibitor reversibly blocked cell proliferation without interfering with cell differentiation.
Asunto(s)
Células Precursoras Eritroides/citología , Células Precursoras Eritroides/efectos de los fármacos , Interleucina-1/antagonistas & inhibidores , Leucemia Mielomonocítica Aguda/patología , Proteínas de Neoplasias/farmacología , Humanos , Monocitos/química , Monocitos/citología , Células Tumorales CultivadasRESUMEN
Previous studies have described an IL-1 Inhibitor produced by a myelomonocytic line developed in our laboratory (Eur J Immunol 1986; 16: 1449). This IL-1 Inhibitor was secreted by the M20 line constitutively in addition to IL-1, from which it could be separated. We have recently shown that the M20 IL-1 Inhibitor is distinct from the IL-1ra. In vitro this factor inhibited IL-1 induced proliferative responses as well as PGE2 secretion by IL-1 induced fibroblasts. We also showed for the first time (Lymphokine Research 1988; 7(3): 268) that an IL-1 inhibitor can reduce IL-1 induced inflammatory effects. This study describes the specific effect of the M20 IL-1 Inhibitor on IL-1 induced parameters of inflammation: fever, leukocytosis and local foot pad swelling or lymph node enlargement. Purified preparations of the IL-1 Inhibitor, when injected together with IL-1, or before the IL-1, reduced fever, leukocytosis, foot pad swelling and lymph node enlargement caused by IL-1. Similar responses were obtained by injection of IL-6 or TNF, but were unaffected by the IL-1 Inhibitor, when injected together. These results indicate that the M20 IL-1 Inhibitor acts specifically on IL-1 induced responses in vivo. The potential importance of this factor as an anti-inflammatory and immune regulatory factor, is supported by the findings of this study.
Asunto(s)
Antiinflamatorios no Esteroideos/farmacología , Inflamación/tratamiento farmacológico , Interleucina-1/antagonistas & inhibidores , Animales , Temperatura Corporal/efectos de los fármacos , Fiebre/inducido químicamente , Fiebre/tratamiento farmacológico , Humanos , Inflamación/inducido químicamente , Interleucina-1/farmacología , Interleucina-6/farmacología , Leucocitosis/inducido químicamente , Leucocitosis/tratamiento farmacológico , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C3H , Receptores de Interleucina-1/antagonistas & inhibidores , Factores de Tiempo , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
We have previously described an IL-1 Inhibitor derived from the M20 myelomoncytic cell line. This line also secretes several molecules of IL-1. We have shown that this factor is specific to IL-1 in vitro, as well as in vivo. In vitro IL-1 induced proliferative responses of mouse thymocytes, human T cells and fibroblasts and IL-1 stimulated PGE2 secretion from fibroblasts, were all inhibited by the M20 IL-1 Inhibitor. In vivo, the IL-1 Inhibitor reduced parameters of acute inflammation such as fever, leukocytosis and local inflammation. This study describes additional effects of the M20 IL-1 Inhibitor on inflammatory serum reactants. Levels of corticosterone and fibrinogen were increased by injection of IL-1, and decreased by the IL-1 Inhibitor. IL-1 reduced zinc and iron plasma levels and elevated copper plasma levels. The M20 IL-1 Inhibitor reversed these changes in a dose dependent manner. Similar effects produced by IL-6 and TNF were unaffected by the M20 IL-1 Inhibitor. Our results indicate that the M20 IL-1 Inhibitor acts specifically on IL-1 induced responses in vivo. Therefore we conclude that this IL-1 Inhibitor has a great potential as an anti-inflammatory agent.
Asunto(s)
Antiinflamatorios no Esteroideos/farmacología , Interleucina-1/antagonistas & inhibidores , Metales/sangre , Animales , Antiinflamatorios no Esteroideos/sangre , Cationes/sangre , Cobre/sangre , Corticosterona/sangre , Fibrinógeno/metabolismo , Interleucina-1/farmacología , Hierro/sangre , Ratones , Ratones Endogámicos BALB C , Zinc/sangreRESUMEN
An interleukin-1 (IL-1) inhibitor produced by the M20 myelomonocytic cell line has been shown to be active in various in vitro and in vivo IL-1 induced parameters. This inhibitor has been purified from the conditioned medium by gel filtration through a Sephacryl S-300 column or dye ligand chromatography on Affi-Gel blue column, followed by isoelectric focusing in free solution in the pH range 3-5 using the Rotofor cell. When gel filtration by FPLC with the Superose 12 column was used as the final step, the combined sequence of purification procedures resulted in a 1600-fold purification of the IL-1 inhibitor. The purified IL-1 inhibitor has a molecular weight of approximately 52 +/- 4 kDa and a pI of 4.15 +/- 0.1. By SDS-PAGE analysis the inhibitor preparation thus obtained showed the presence of two protein bands, while a few closely spaced protein bands were seen by analytical isoelectric focusing in polyacrylamide gels (pH 3-6). Some of these bands in PAGIF might correspond to different degrees of glycosylation of the inhibitory protein. Although the M20 IL-1 inhibitor has not yet been purified to homogeneity, it should be stressed that the procedures used, allowed us to remove the great majority of the proteins present in the medium in which the M20 cells were cultured, and to recover in satisfactory yield the inhibitor which we consider likely to be present in the conditioned medium in subnanomolar concentrations.
Asunto(s)
Citocinas/fisiología , Interleucina-1/antagonistas & inhibidores , Monocitos/química , Línea Celular , Medios de Cultivo , Humanos , Punto Isoeléctrico , Peso MolecularRESUMEN
Interleukin-1 (IL-1) is an important mediator in inflammation and immunological processes. The findings of native IL-1 inhibitors suggest a negative feedback mechanism to down-regulate IL-1 mediated acute inflammation. IL-1 inhibitors were also found elevated in disease states associated with high IL-1 levels. We have previously described one such IL-1 inhibitor derived from the human M20 myelomonocytic cell line. In this paper we present several biological and biochemical characteristics of the M20 IL-1 inhibitor. Various in vitro activities of the inhibitor are described and its IL-1 specificity in these assays is demonstrated. Purification of the inhibitor was performed by DEAE-high performance liquid chromatography, isoelectric focusing, gel filtration and dye ligand chromatography column. This protein factor has a MW of 52 +/- 4 kDa and a pI of 4.15 +/- 0.1. The inhibitor has no cross-reactivity against a panel of known cytokines (IL-1 alpha, IL-1 beta, IL-2, sIL-2R, IL-6, tumor necrosis factor (TNF), interferon-gamma (IFN-gamma)) and is distinct from the IL-1 receptor antagonist (IL-1ra). The purified IL-1 inhibitor was destroyed by trypsin, 2-mercaptoethanol, sodium dodecyl sulfate and extremes in pH and in temperature. Only IL-1 induced (but not the IL-2, IL-6 or TNF induced) thymocyte proliferation and PGE2 production by fibroblasts were inhibited by the inhibitor, thus showing specificity to IL-1 in these assays.
Asunto(s)
Citocinas/inmunología , Interleucina-1/antagonistas & inhibidores , Monocitos/química , Bioensayo , Línea Celular , Citocinas/química , Citocinas/farmacología , Humanos , Concentración de Iones de Hidrógeno , Activación de Linfocitos/efectos de los fármacos , Mercaptoetanol/farmacología , Desnaturalización Proteica , Temperatura , Tripsina/farmacologíaRESUMEN
OBJECTIVES: To investigate the possibility of local interleukin-1 (IL-1) and IL-1 inhibitor production by human granulosa and cumulus cells and to assess their direct effects on the steroidogenesis of these cells in vitro. DESIGN: Prospective study. PARTICIPANTS: Normal ovulatory women undergoing ovulation induction for in vitro fertilization. INTERVENTION: Pretreatment of patients with gonadotropin-releasing hormone analogue, human menopausal gonadotropin, and human chorionic gonadotropin (hCG). MAIN OUTCOME MEASURES: Retrieval and isolation of granulosa luteal cells and follicular fluid (FF). Granulosa luteal cells and cumulus cells cultured and analyzed by fluorescent activated cell sorter. Follicular fluid separated and bioassayed for IL-1 and IL-1 inhibitory activity. Steroid measurement performed. Interleukin-1 inhibitor purified. Interleukin-1 and IL-1 inhibitor bioassay performed. Statistical analysis made and interpreted. RESULTS: Interleukin-1, but not IL-1 specific inhibitory activity, was found in granulosa and cumulus cell cultures and also in FF, only after its purification on a high-pressure liquid chromatography column. Under nonstimulated conditions, neither IL-1 nor IL-1 inhibitor had any effect on basal progesterone (P) or estradiol (E2) secretion. However, IL-1 inhibitor demonstrated significant (P < 0.01) inhibition of hCG-stimulated P secretion (from 200 to 110 ng/10,000 cells per 24 hours). In addition, IL-1 demonstrated a significant (P < 0.05) and dose-dependent inhibition of hCG-stimulated E2 production (from 6,832 +/- 460 to 4,237 +/- 141 pg/10,000 cells per 24 hours). CONCLUSIONS: Interleukin-1 may exert a significant local autocrine regulatory role in the human ovary.
Asunto(s)
Células de la Granulosa/metabolismo , Interleucina-1/metabolismo , Células Cultivadas , Estradiol/biosíntesis , Femenino , Líquido Folicular/química , Humanos , Interleucina-1/análisis , Interleucina-1/antagonistas & inhibidores , Interleucina-1/fisiología , Fase Luteínica , Progesterona/biosíntesis , Estudios ProspectivosRESUMEN
The present study was performed to evaluate the correlation between follicular fluid levels of interleukin 2 (IL-2) and IL-2 soluble receptor (sIL-2R), oestradiol, progesterone and testosterone levels, oocyte fertilization, embryo quality and pregnancy rates. Twenty-eight patients with a pure tubal factor and undergoing in-vitro fertilization and embryo transfer were randomly chosen and treated with gonadotrophin releasing hormone agonist (GnRHa) in the midluteal phase (long protocol) coupled with follicular phase administration of human menopausal gonadotrophin. Transvaginal follicular aspiration was performed 36 h after human chorionic gonadotrophin administration, followed 48 h later by embryo transfer. One hundred and twenty-three follicular fluids were sampled. The mean follicular fluid levels (+/- SD) were 2.30 +/- 0.80 fmol for IL-2, 458.2 +/- 236.0 units/ml for sIL-2R, 28.5 +/- 58.1 ng/ml for oestradiol, 2360.5 +/- 2846 ng/ml for progesterone and 7.22 +/- 7.08 ng/ml for testosterone. There was a significant (P less than 0.01) correlation between IL-2 and testosterone levels. No correlation was found between the lymphokines and serum oestradiol, follicular fluid progesterone, oocyte fertilization, embryo quality and pregnancy. It may be concluded that significant concentrations of IL-2 and sIL-2R exist in follicular fluid. Wide variations in follicular IL-2 and sIL-2R concentrations of different follicles were found in the same patients.
Asunto(s)
Fertilización In Vitro , Líquido Folicular/metabolismo , Fase Folicular/fisiología , Interleucina-2/metabolismo , Progestinas/metabolismo , Receptores de Interleucina-2/metabolismo , Estradiol/metabolismo , Femenino , Fertilización/fisiología , Humanos , Oocitos/fisiología , Embarazo/fisiología , Progesterona/metabolismo , Solubilidad , Testosterona/metabolismoRESUMEN
Data has accumulated suggesting reciprocity between cytokines and the reproductive system. The present study was performed in order to evaluate the correlation between interleukin 1 (IL-1) and tumour necrosis factor (TNF) concentrations in follicular fluid and its oestradiol, progesterone and testosterone levels. A total of 39 follicular fluid samples, from eight patients undergoing in-vitro fertilization and embryo transfer were evaluated. All of the patients were treated by a midluteal (long) protocol involving a gonadotrophin releasing hormone agonist (GnRHa) coupled with follicular phase human menopausal gonadotrophin. Mean levels in follicular fluid of IL-1, TNF, oestradiol, progesterone and testosterone were 1.58 +/- 0.42 fmol/0.1 ml, 4.69 +/- 4.18 pg/ml, 28.5 +/- 58.1 ng/ml, 2360.5 +/- 2846.3 ng/ml and 7.22 +/- 7.08 ng/ml respectively. There was a significant (P less than 0.01) positive correlation between IL-1 and progesterone levels. There was no significant correlation between the different lymphokines and oestradiol secretion, oocyte fertilization, embryo quality and pregnancy rates. It is concluded that IL-1 and TNF exist in follicular fluid. It may be hypothesized that IL-1 has a local regulatory action, possibly promoting luteinization.
Asunto(s)
Fertilización In Vitro , Líquido Folicular/química , Hormonas Esteroides Gonadales/análisis , Interleucina-1/análisis , Factor de Necrosis Tumoral alfa/análisis , Adulto , Fase de Segmentación del Huevo/fisiología , Estradiol/análisis , Femenino , Fertilización/fisiología , Humanos , Progesterona/análisis , Radioinmunoensayo , Testosterona/análisisRESUMEN
This study aimed to assess the effect of the M20 interleukin-1 (IL-1) inhibitor on normal and leukemic hematopoietic cells. The M20-derived IL-1 inhibitor was found to inhibit the growth of various hematopoietic cells. The in vitro proliferation of myeloid cell lines in serum-containing medium or proliferation of these cells induced by IL-1 in serum-free medium (measured by 3H-TdR) were inhibited by the M20 IL-1 inhibitor. In addition, growth of normal progenitors and fresh leukemic cells stimulated by granulocyte-macrophage colony-stimulating factor (GM-CSF) (as measured by colony and liquid systems) was also inhibited by this factor. After the removal of the IL-1 inhibitor at the peak of growth inhibition, leukemic and normal progenitor cells retain their ability to grow and develop into GM-CSF colonies. These results show that the growth inhibition phenomena were reversible and did not result from a cytotoxic effect. Our data suggest that the M20-derived IL-1 inhibitor might function as a true negative growth regulator of normal and leukemic hematopoietic cells.
Asunto(s)
Granulocitos/efectos de los fármacos , Células Madre Hematopoyéticas/efectos de los fármacos , Interleucina-1/antagonistas & inhibidores , Leucemia/patología , División Celular/efectos de los fármacos , Factor Estimulante de Colonias de Granulocitos y Macrófagos/farmacología , Granulocitos/patología , Células Madre Hematopoyéticas/patología , Humanos , Interleucina-1/farmacología , Interleucina-6/farmacología , Células Tumorales CultivadasRESUMEN
Several inhibitors of IL-1 have been described. Four appear to be the same: one purified from urine of patients with monocytic leukemia, another from IgG-stimulated monocytes, a third from PMA-induced U937 cels, and a fourth from keratinocytes. Because these IL-1 inhibitors compete with bona fide IL-1 for occupancy of IL-1 receptors, they are now called the IL-1 receptor antagonist (IL-1ra) or IL-1 receptor antagonist protein (IRAP). We have described another IL-1-specific inhibitor produced by the human myelomonocytic leukemia cell line, M20. This inhibitor specifically blocks IL-1-induced effects both in vitro and in vivo. In the present study, we compared the M20 IL-1 inhibitor with IL-1ra using neutralization in an IL-1 bioassay and immunoblotting with an anti-IL-1ra antibody that recognizes natural IL-1ra. Neutralization experiments, immunoblotting, and western blotting obtained after transfer from SDS-PAGE revealed that anti-IL-1ra does not recognize the M20 IL-1 inhibitor. In addition, the isoelectric point and molecular weight of the M20 IL-1 inhibitor were different from those of the IL-1ra. From these data, we conclude that the M20 IL-1 inhibitor is antigenically unrelated to the IL-1ra but is a distinct and specific IL-1 inhibitor.
Asunto(s)
Interleucina-1/antagonistas & inhibidores , Receptores Inmunológicos/antagonistas & inhibidores , Bioensayo , Línea Celular , Electroforesis en Gel de Poliacrilamida , Humanos , Inmunoglobulina G , Radioisótopos de Yodo , Focalización Isoeléctrica/métodos , Receptores de Interleucina-1RESUMEN
The effect of human recombinant interleukin-1 (IL-1) on the regulation of lipoprotein lipase (LPL) was studied in rat heart mesenchymal cell cultures. A time-dependent reduction in enzyme activity occurred with a 30% fall after 1 h. The suppression of enzyme activity was accompanied by a commensurate reduction in enzyme mass. The reduction in LPL activity was most prominent in the heparin releasable pool; IL-1 treatment resulted in a 7.2-8.3-fold decrease in the functional compartment and a 2.5-2.8-fold decrease in residual cellular activity. The effect of IL-1 could be prevented by the addition of the IL-1 inhibitor. However, in contradistinction to the effect of tumor necrosis factor (TNF), there was no change in LPL mRNA in cultures treated with IL-1. The present results show that the regulation of LPL in mesenchymal heart cell cultures by IL-1 occurs posttranscriptionally, as has been shown in 3T3 cells. The more pronounced effect on LPL activity in the functional pool suggests that IL-1 treatment might have influenced also the processing and/or transport of the enzyme to the cell surface.
Asunto(s)
Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Interleucina-1/farmacología , Lipoproteína Lipasa/antagonistas & inhibidores , Miocardio/enzimología , Animales , Animales Recién Nacidos , Células Cultivadas , Electroforesis , Immunoblotting , Lipoproteína Lipasa/genética , Mesodermo/efectos de los fármacos , Mesodermo/enzimología , ARN Mensajero/efectos de los fármacos , Ratas , Proteínas Recombinantes/farmacología , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
PBMC concentrated from 12 patients with CLL were incubated for up to 6 days with various doses of TPA or RA in an attempt to induce differentiation. The results show a great heterogeneity in the cellular response to both inducers. TPA produced two major changes in B-CLL cells, namely the development of some hairy-cell leukemia features with an increase in Ig secretion and the expression of Leu M5 and TAC receptors. In contrast to TPA, RA induced only modest or no changes in surface, but increased numbers of large macrophages were seen as assessed by latex-bead ingestion, expression of Fc receptors and cytochemistry. These changes were more obvious after exposure to RA than to TPA. Thus, while TPA induces mostly differentiation and hairy-cell features in B-lymphocytes, RA induces activation of the monocyte pool with no obvious changes in the lymphocytic compartment.
Asunto(s)
Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Linfocitos/efectos de los fármacos , Monocitos/efectos de los fármacos , Acetato de Tetradecanoilforbol/farmacología , Tretinoina/farmacología , Fosfatasa Ácida/análisis , Adulto , Anciano , Antígenos de Superficie/análisis , Femenino , Humanos , Inmunoglobulinas/metabolismo , Leucemia Linfocítica Crónica de Células B/inmunología , Linfocitos/enzimología , Linfocitos/inmunología , Linfocitos/ultraestructura , Masculino , Persona de Mediana Edad , Monocitos/enzimología , Monocitos/inmunología , Monocitos/ultraestructuraRESUMEN
Elevated proportions of monocytes have previously been found in the blood of healthy women during the ovulation period as well as in other conditions associated with increased blood estradiol (E2). This phenomenon was explained, in part, by an augmenting effect which physiological concentrations of E2 may have on the development of granulocyte-macrophage (GM) colonies derived from normal peripheral blood mononuclear cells. To analyze this effect, we tested possible alternatives for the interaction between E2, colony-stimulating factor (CSF) and GM colony progenitor cells. E2 was found not to interact synergistically with CSF, but pre-treatment of the progenitor cells with E2 resulted in higher numbers of colonies in response to CSF. Moreover, E2 did not induce higher secretion of CSF but treatment with anti-CSF antibodies abolished the enhancing effect of E2. Based on these results, we suggest that the augmenting effect of E2 on GM colony formation is mediated by inducing the colony precursor cells to be more responsive to CSF. These findings may help to elucidate some of the complex relationships between estrogens, immune responses and hemopoiesis.
Asunto(s)
Estradiol/farmacología , Leucemia Mieloide/patología , Monocitos/citología , Adulto , Agregación Celular/efectos de los fármacos , División Celular/efectos de los fármacos , Línea Celular , Medios de Cultivo , Femenino , Humanos , Cinética , Monocitos/efectos de los fármacosRESUMEN
Culture supernatants from a myelomonocytic cell line (M20) were found to inhibit interleukin 1 (IL 1) activity in vitro. The factor, isolated from these supernatants, inhibited augmentation of phytohemagglutinin response of mouse thymus cells induced by IL 1 derived from several established cell lines. Various IL 1-dependent activities such as lymphocyte and fibroblast proliferation in vitro were also inhibited by the factor. The factor did not inhibit IL 2-induced or other proliferative responses not related to IL 1. Preliminary biochemical characterization of the factor indicated that the activity resides in a protein with a molecular mass of 52 kDa.
Asunto(s)
Interleucina-1/antagonistas & inhibidores , Linfocinas/metabolismo , Monocitos/metabolismo , División Celular/efectos de los fármacos , Línea Celular , Depresión Química , Fibroblastos/efectos de los fármacos , Humanos , Interleucina-1/análisis , Interleucina-2/análisis , Leucemia Mieloide/metabolismo , Leucemia Mieloide/patología , Activación de Linfocitos/efectos de los fármacos , Linfocinas/farmacología , Fitohemaglutininas/farmacologíaRESUMEN
Culture supernatants from normal human monocytes, monocyte hybrid cell lines, and myelomonoblastic cell lines were tested for human interleukin-1 (IL-1) activity. In the present study, we report the detection of IL-1 secreted by several cell lines of monocyte origin and compare their biological and biochemical characteristics. IL-1 activity was tested by the regular assay of phytohemagglutinin (PHA) response of mouse thymus cells. IL-1 was found to be constitutively secreted by U937 and the M20 cell lines, as well as by three of the monocyte hybrid cell lines. The activity was always augmented following dialysis and did not require the presence of serum for its secretion. We compared the IL-1 activity of the myelomonoblastic M20 and hybrid 1C4 cell lines to that of normal monocytes. We found differences in the kinetics of IL-1 secretion, the pattern of activity following dilution of concentrated supernatants, and augmentation of activity by various inducers. The differences described may be explained by concomitant secretion of IL-1 inhibitory factors, as well as the secretion of activities other than IL-1. Preliminary biochemical analysis showed that all three cell sources tested shared some species of molecules characterized by gel filtration and ion-exchange chromatography. However, some species of molecules expressing IL-1 activity were unique to the cell lines and were not found in normal monocytes.
Asunto(s)
Interleucina-1/biosíntesis , Monocitos/inmunología , Animales , Bioensayo , Línea Celular , Humanos , Células Híbridas/inmunología , Interleucina-1/análisis , Leucemia Mieloide Aguda/inmunología , Activación de Linfocitos , RatonesRESUMEN
The P component of amyloid is a normal serum protein designated SAP. SAP has substantial homology with C-reactive protein (CRP). However, unlike CRP, SAP is not an acute-phase reactant in man. Recent studies have established SAP as a major acute-phase protein in mice. Moreover, mice which have received tumour implants have also been found to have raised serum concentrations of SAP. The aim of the present study was to determine possible association between the serum level of SAP and human cancer. We found that patients with carcinoma of the breast have significantly increased serum concentrations of SAP. Moreover, in these patients SAP levels correlated with the severity of the disease. Patients with carcinoma of the colon, however, did not differ from healthy individuals in the serum level of SAP. Possible explanations for this discrepancy are discussed.
Asunto(s)
Amiloide/sangre , Neoplasias de la Mama/sangre , Neoplasias del Colon/sangre , Neoplasias de la Mama/patología , Antígeno Carcinoembrionario/análisis , Neoplasias del Colon/patología , Femenino , Humanos , Masculino , Componente Amiloide P SéricoRESUMEN
In vitro maturation of human monocytes to macrophages was characterized by morphological criteria, cell size and lysosomal enzymes activity. Purified populations of monocytes were maintained in culture at either adherent or nonadherent conditions and their maturation to macrophages was observed in both cases. The addition of external factors such as hydrocortisone and vitamin D3 inhibited monocyte maturation. In the absence of external factors, nonadherent monocytes were inhibited in their maturation for up to 10 days when plated at crowded cell concentrations. In addition, the presence of human serum in the culture media had a higher inhibitory activity than similar concentrations of fetal calf serum. Supernates from crowded macrophages were also inhibitory for monocyte maturation. We suggest the possibility that cell crowding, as well as soluble factors found in the serum and probably secreted by macrophages, participate in the regulation of monocyte development by inhibiting their maturation. Once released from this inhibitory signal or environment, the monocytes mature to macrophages.
Asunto(s)
Macrófagos/fisiología , Monocitos/fisiología , Adolescente , Adulto , Adhesión Celular , Recuento de Células , Diferenciación Celular/efectos de los fármacos , Supervivencia Celular , Células Cultivadas , Colecalciferol/farmacología , Medios de Cultivo , Femenino , Hexosaminidasas/análisis , Histocitoquímica , Humanos , Hidrocortisona/farmacología , Macrófagos/citología , Macrófagos/efectos de los fármacos , Macrófagos/enzimología , Macrófagos/ultraestructura , Masculino , Microscopía Electrónica de Rastreo , Monocitos/citología , Monocitos/efectos de los fármacos , Monocitos/enzimología , Monocitos/ultraestructura , beta-N-Acetil-GalactosaminidasaRESUMEN
Primary cultures of cells derived from 13 patients with acute myelomonocytic leukemia (AMML) were studied with particular emphasis on in vitro proliferation, cell differentiation and the mode for establishment of cell lines. Using irradiated human macrophage monolayers to assist cell growth, we obtained four new cell lines of myelomonocytic origin. All the cell lines were characterized for cytochemical markers and response to phorbol esters (TPA), a differentiation inducing agent. In the absence of any inducing agent, spontaneous differentiation of blast cells into mature macrophages-like cells occurred in 8 out of the 13 primary cultures. Thus, maturation induction by agents such as TPA is not always required in order to obtain leukemic cell differentiation in vitro. The regulation of cell proliferation and differentiation by cellular interactions and by extrinsic soluble products is discussed in detail, in the light of these findings.
Asunto(s)
Línea Celular , Leucemia Mieloide/patología , Diferenciación Celular/efectos de los fármacos , Medios de Cultivo , Humanos , Macrófagos/efectos de la radiación , Acetato de Tetradecanoilforbol/farmacologíaRESUMEN
Monoclonal anti GP-70 antibodies (BI) were generated in mice and used for screening of various malignant and non-malignant cell lines. The reactivity of these monoclonal antibodies was compared with that obtained with the polyclonal anti GP-70 antibody described in earlier studies [1-3]. The results indicated complete similarity in reactivity of both of the antibodies used. Furthermore, the reactivity of BI antibodies with cell samples obtained from a variety of leukemia and lymphoma patients and with peripheral blood samples from healthy blood donors was also very similar to the pattern of specificity described in earlier reports for the polyclonal preparation. From these studies we conclude that the monoclonal antibodies can substitute the polyclonal anti GP-70 antibodies in the diagnosis and subtyping of B-type leukemias and lymphomas.