Production of interleukin-1-alpha and interleukin-2 by mononuclear cells in healthy adults in relation to different experimental conditions and to the presence of growth hormone.

Several reports support the view that the growth hormone (GH) possesses a number of important immunomodulatory properties. This study was undertaken to determine in vitro the role of the GH on interleukin (IL) production. Cultures of blood peripheral lymphocytes obtained from human normal adults were performed in RPMI medium in the presence or absence of phytohemagglutinin (PHA), heated normal serum (NHS) 1% and GH 12.5-500 microgram/l. After incubation from 15 h to 4 days at 37 degrees C in a humidified atmosphere containing 5% CO2, cells were discarded and the supernatants were tested for their contents of IL-1 alpha and IL-2 measured using the Amersham radioimmunoassay system. The results of these in vitro experiments show that: (1) the bulk cultures from peripheral lymphocytes are suitable to study the IL-1 and IL-2 production; (2) in optimal conditions for IL production (incubation during 48 h in the presence of PHA and NHS) no effect of GH was observed on IL production; (3) in the absence of PHA GH acts at physiological doses (less than 100 ng/ml) by increasing the IL production. This effect of GH was optimized with a short time of incubation (16 h) and in the simplest conditions of cultures, that is to say in the absence of serum and of PHA: thus in the presence of GH 100 ng/ml the IL-1 production increases from 0.53 to 3.86 fmol (tubes) and IL-2 increases from 0.18 to 3.22 fmol (tubes). These differences are significant (p less than 0.001). We conclude that GH acts in vitro on mononuclear cells to induce IL production.

Production of interleukin-1 alpha and interleukin-2 by mononuclear cells from children with growth delay in relation to the degree of growth hormone deficiency: effects of substitutive treatment.

Interleukin-1 alpha (IL-1 alpha) and interleukin-2 (IL-2) levels were measured by radioimmunoassay in samples of conditioned medium from mononuclear cells taken from 20 normal subjects (14 adults ranging in age from 20 to 45 years and 6 children ranging in age from 3 to 11 years) and from 49 children with growth delay. Cultures were performed with 10(6) cells/ml in medium containing 1% normal human serum and 4.8 g/l phytohemagglutinin M. The incubation was performed for 48 h in an atmosphere containing 5% CO2. In normal subjects, the production of IL-1 alpha was 38.5 +/- 9.8 fmol/ml of conditioned medium (mean +/- SEM) in 14 adults and 41.6 +/- 3.0 fmol/ml in 6 children. The production of IL-2 was 46.9 +/- 6.5 and 57.3 +/- 10.5 fmol/ml, respectively. In the 16 patients with growth hormone (GH) deficiency studied before treatment, the production of ILs was significantly decreased in relation to the degree of deficiency. We observed a positive correlation between the production of IL-1 alpha and the values of insulin-like growth factor I but not with serum GH values. IL-1 alpha production was normalized after 15 days of substitutive GH therapy and IL-2 was normalized after 3 months of therapy. In 10 other patients with GH deficiency (4 with total and 6 with partial isolated GH deficiency) studied after long-term GH treatment (5 months or more), the mean of IL production was not significantly different from that of GH-deficient children treated for 3 months.(ABSTRACT TRUNCATED AT 250 WORDS)

In vitro effect of human growth hormone on lymphocyte transformation and lymphocyte growth factors secretion.

Cultures of human blood peripheral lymphocytes were performed in the presence or absence of human growth hormone, and also of phytohemagglutinin and normal human serum 10%. After incubation for 48 h, the supernatants were tested for their ability to promote the uptake of [3H]thymidine into lectin-activated lymphocytes. Supernatants from lymphocyte-free control samples, treated in the same manner, were assayed under the same experimental conditions. Variance analysis of the different dose-response relationships was performed. The results of these in vitro experiments confirm that physiological levels of GH inhibit the lectin-induced lymphoproliferation and that lymphocytes secrete an 'activity' able to stimulate the incorporation of [3H]thymidine into lectin activated lymphocytes. Furthermore we show that: 1) Secretion of this lymphocyte-stimulating activity is increased by physiological levels of GH; 2) This lymphocytic secretion is not radioimmunoassayable IGF-I; 3) Using fast protein liquid chromatography (FPLC), this activity appears in fractions with various molecular weights.

[GH-dependency of lymphocyte multiplication factor with a low molecular weight]. / GH-dépendance d'un facteur de multiplication lymphocytaire de petit poids moléculaire.

Liquid chromatography (FPLC) separation at pH 6.85 shows that the fractions in human serum which stimulate the multiplication of human lymphocyte are mainly in the 28-42 kd and the 1.3-1.9 kd fractions. The low molecular weight fraction is absent or very low in both GH-deficient and GH-resistant children and T lymphocyte-deficient children, and appears after appropriate treatment, i.e. GH or marrow transplantation. The data suggest the GH-dependency of 1.3-1.9 kd factor(s) produced by immuno-competent lymphocytes.

[Effect of serum fractions on the incorporation of tritiated thymidine in activated lymphocytes: I. Human serum from normal adult subjects]. / Effet des fractions sériques sur l'incorporation de thymidine tritiée dans le lymphocyte activé: I. Sérum humain provenant de sujets adultes normaux.

Fractionation of normal human adult serum using Centrisart or gel filtration at neutral pH shows that serum thymidine activity, measured as the ability to stimulate the 3H-thymidine uptake into lectin-activated lymphocytes, is represented by several molecular forms distinct from the endogenous Sm-C. The main thymidine activity is found in the 50-70 K range.

[Effect of serum fractions on the incorporation of tritiated thymidine in activated lymphocytes: II. Comparison between cord blood and the blood of newborn infants]. / Effet des fractions sériques sur l'incorporation de thymidine tritiée dans le lymphocyte activé: II. Comparaison entre le sang du cordon et le sang du nouveau-né.

After serum Sephacryl 300 gel filtration, the major circulating forms of TA in serum are macromolecular. However different patterns are found in cord blood and in the blood of newborns at 24 hr of life. The low values found in cord blood are not due to inhibiting factor(s). TA in serum fractions does not parallel immunoreactive Sm-C levels. The pattern of immunoreactive Sm-C are similar in both cord blood and newborn blood.

Effect of bromocriptine treatment on circulating somatomedins in tall adolescents.

Somatomedin and bromocriptine levels were measured before and after 3 months of bromocriptine treatment (5 mg/day) in the sera of 16 tall adolescents with excessive adult height prediction. Somatomedins were measured by RIA for somatomedin C and IgFII and by measuring thymidine incorporation into human lectin-activated lymphocytes. Mean +/- SEM levels of bromocriptine after three months of treatment were 0.61 +/- 0.08 ng/ml. No changes in radioimmunoassayable somatomedin C were observed after bromocriptine intake respectively 1.34 +/- 0.17 U/ml before and 1.4 +/- 0.01 U/ml during treatment. On the opposite a significant decrease of thymidine activity (p less than 0.002) from 1.45 +/- 0.17 U/ml to 1.12 +/- 0.19 U/ml was observed. No changes of IgFII levels were observed in the sera of the 8 patients where they were measured. In order to test a possible direct effect of bromocriptine on peripheral tissues bromocriptine mesylate was added in somatomedin bioassays. Inhibitory effect of bromocriptine in vitro is seen at higher levels (1 microM) compared to the circulating one (1 nM). This study demonstrates that the major effect of bromocriptine which is the acceleration of bone maturation is not related to changes in somatomedin C.

Somatomedin, transferrin and amino-acids in serum following injection of human growth hormone in children with growth disease.

17 children with growth retardation (12 with idiopathic hypopituitary dwarfism, 2 with craniopharyngioma and 3 constitutionally short) were studied for three days following a single intramuscular injection of human growth hormone. Somatomedin activity was bioassayed using both sulphate incorporation into chick embryo cartilage and thymidine uptake by human lectin-activated lymphocytes. In hypopituitary patients it showed a significant response, maximal 24 hours after the injection, and significantly correlated for the two bioassays. The aminoacid content of the incubation medium used for thymidine bioassay appeared as an important factor: both glutamine and nonessential aminoacids are required to obtain significant stimulation by low serum concentrations, thus increasing the sensitivity of the assay but reducing the differences between normal and hypopituitary sera. Transferrin levels in serum were significantly lower in hypopituitary dwarfs. They did not rise in the three days following hGH. Aminoacid levels were lower in idiopathic GH deficient patients than in other groups, and did not show short term increase in the fasting samples collected after hGH administration.

Effect of purified somatomedins on thymidine incorporation into lectin-activated human lymphocytes.

These data describe the effects of purified preparations of several growth factors on thymidine incorporation into phyto-haemagglutinin-activated (PHA) human lymphocytes. The somatomedins selected for this study included human somatomedins A and C, insulin-like growth factors (IFG1 and IGF2) and multiplication stimulating activity (MSA). Assays were carried out with and without serum. Complementary assays were performed with a low-molecular serum ultrafiltrate added to somatomedin C and to MSA. We found that all the peptides tested, except MSA, stimulated thymidine incorporation into PHA-activated lymphocytes in a dose-dependent manner, even though different doses were required to obtain a response. The data reported point out the multiplicity and the interrelationships of the serum factors involved in the stimulation of human cells growth.

[Early effects of intramuscular growth hormone on plasma somatomedin activity (author's transl)]. / Effets à court terme de l'injection d'hormone de croissance sur l'activité somatomédine du plasma.

The early effects of intramuscular injection of human growth hormone (hGH) on plasma sulfation activity (Sm) have been followed for 72 hours: 1) after one injection of 6 mg in control children and in pituitary dwarfs (isolated idiopathic GH deficiency, multiple idiopathic pituitary deficiencies, and post-surgical or post-radiotherapic hypopituitarism); 2) after 6 injections of 1 mg every 12 hours in idiopathic pituitary dwarfs. Following injection of 6 mg, Sm decreases during 2-4 hours in all groups studied, then rises and peaks at 24 hours. The early decrease of Sm could relate to a rise of Sm-binding protein, as suggested by data obtained in the dog after intravenous injection of bovine GH. Following 6 injections of 1 mg, the rise of Sm is slower but higher and more protracted than with one injection of 6 mg. This fact suggests that the clinical effects of fractionation of treatment with HGH require further study. The lack of correlation between the biological data obtained and the clinical effects of hGH treatment upon the growth of pituitary dwarfs shows that the short-term hGH test used does not allow to foresee the effects of treatment.

Serum somatomedin activity measured as sulphation factor in peripheral, hepatic and renal veins of patients with alcoholic cirrhosis.

Serum somatomedin (SM) activity, measured as sulphation factor on chick embryo cartilage, and growth hormone (GH) levels were measured in peripheral, hepatic and renal veins of 23 patients with a alcoholic cirrhosis. SM activity (mean +/- SEM) was 0.65 +/- 0.05 U/ml in peripheral vein, 0.59 +/- 0.04 U/ml in hepatic vein, and 0.74 +/- 0.07 U/ml in renal vein. Mean GH levels were respectively 2.8, 2.5 and 3.1 ng/ml. Compared to peripheral vein, SM increase in renal vein was 19% (P less than 0.05). Serum SM activity was significantly lower in 13 patients with alcoholic hepatitis associated with cirrhosis than in other 10 patients (P less than 0.02 in hepatic blood and P less than 0.05 in peripheral blood). The decrease of SM activity seems related to cytolysis and hepato-cellular insufficiency. At last, in patients with alcoholic hepatitis, SM activity was lower in the hepatic vein than in the peripheral vein (P less than 0.05). The cause of this difference remains under discussion, no SM inhibitors being found in the serum samples used in this study.

Correlative study of radioreceptor assay and radioimmunoassay of serum growth hormone in children. II: Primary, genetic and intra-uterine growth retardation.

Growth hormone (GH) was measured by radioreceptorassay (RRA) and radioimmunoassay (RIA) in the sera of 24 children with idiopathic primary growth retardation (PGR), 15 with genetic short stature (GSS) and 11 with intra-uterine growth retardation (IUGR), and compared to results obtained in normal children. The average RRA/RIA ratio was close to normal in PGR (1.02 +/- 0.05) and in IUGR (1.06 +/- 0.07), and slightly though not significantly lower in GSS (0.86 +/- 0.06). Some variability in RRA/RIA ratio was found in individual patients of each group, and some sera gave a non-parallel displacement of the tracer when compared to the standard curve. But no genetic difference of RRA-assayable GH was found between the three groups studied and normal children.