Decreased corticosensitivity in quiescent Crohn's disease: an ex vivo study using whole blood cell cultures.

Corticosensitivity influences the degree and the duration of an inflammatory reaction by altering target cell responses to endogenous and/or exogenous glucocorticoids. Indeed, different clinical responses to glucocorticoids have been observed among patients with Crohn's disease, suggesting different degrees of corticosensitivity in these subjects. The purpose of this study was to compare the corticosensitivity of patients with quiescent Crohn's disease to that of healthy subjects (HS). Nineteen patients with quiescent Crohn's disease and 14 HS were studied; all patients were steroid-free for at least six months; 7 of the 19 were corticosteroid-dependent (CSD) and treated with nonglucocorticoid immunosuppressants at the time of the study. Corticosensitivity was measured by the inhibition of LPS-induced cytokine secretion in whole blood cell cultures treated with increasing concentrations (10(-9) to 10(-6) M) of dexamethasone. Tumor-necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), and interleukin-1 beta (IL-1 beta) were measured using specific immunoassays. Crohn's disease patients had a markedly decreased dexamethasone-mediated inhibition of TNF-alpha (P < 0.01), IL-6 (P < 0.001), and IL-1 beta (P < 0.01) compared to healthy subjects, with a shift of the dexamethasone dose-response curve to the right. No significant differences in the basal and LPS-stimulated secretion of the three cytokines were observed between CSD and non-CSD patients, and both subgroups of patients had similar degrees of dexamethasone-mediated cytokine inhibition. We conclude that patients with Crohn's disease have a significant decrease in the corticosensitivity of their leukocytes. This may be related to a specific genetic/constitutional background and/or could be acquired, due to inflammation-related endocrine and/or immune factors.

Tumour necrosis factor (TNF) gene polymorphism influences TNF-alpha production in lipopolysaccharide (LPS)-stimulated whole blood cell culture in healthy humans.

TNF-alpha is involved in infectious and immuno-inflammatory diseases. Different individuals may have different capacities for TNF-alpha production. This might determine a predisposition to develop some complications or phenotypes of these diseases. The aims of our study were to assess the inter-individual variability of TNF-alpha production and to correlate this variability to a single base pair polymorphism located at position -308 in TNF gene. We studied 62 healthy individuals. TNF-alpha production after LPS stimulation was evaluated using a whole blood cell culture model. The TNF gene polymorphism was studied by an allele-specific polymerase chain reaction. Other cytokines produced in the culture, soluble CD14 concentrations and expression of CD14 on blood cells were also measured. Among the 62 individuals, 57 were successfully genotyped. There were 41 TNF1 homozygotes and 16 TNF1/TNF2 heterozygotes. TNF-alpha production after LPS stimulation of whole blood cell culture was higher among TNF2 carriers than among TNFI homozygotes (929pg/ml (480-1473pg/ml) versus 521 pg/ ml (178-1307 pg/ml); P<0.05). This difference was even more significant after correction of TNF-alpha production for CD14 expression on blood cells. In conclusion, the single base pair polymorphism at position -308 in the TNF gene may influence TNF-alpha production in healthy individuals.

Effects of dexamethasone on the profile of cytokine secretion in human whole blood cell cultures.

EXPERIMENTAL OBJECTIVES: The interaction between the endocrine and immune systems is a very intriguing area. Endogenous glucocorticoids, as end-effectors of the hypothalamo-pituitary-adrenal axis, inhibit the immune and inflammatory responses and are used as immunosuppressive drugs in many inflammatory, autoimmune and allergic diseases. The aims of this study were to investigate the effects of dexamethasone on the profile of cytokine secretion in whole blood cell cultures from healthy subjects and to analyse the gender-related sensitivity to dexamethasone on each cytokine secretion. RESULTS: There was a significant inhibition by dexamethasone (from 1 to 100 nM) on the secretion of monokines (IL-1beta, IL-6, IL-8 and TNF alpha) and lymphokines (IL-2, IL-4, IL-10 and IFN gamma), either after LPS or PHA stimulation (P < 0.01). Interleukin 4 and IL-10 were less inhibited than IFN gamma (P < 0.05 at 1 nM, P < 0.01 at 10 nM and P < 0.001 from 100 nM to 10 microM). No gender difference was observed in the rate of inhibition of the secretion of each cytokine. CONCLUSION: This study shows that the inhibition of cytokine secretion by dexamethasone is more marked on Th1-type cytokines than on Th2-type cytokines. These data support the idea that glucocorticoids may induce a shift from the Th1 to Th2 profile of cytokine secretion.

Ex vivo cytokine production by whole blood cells from cancer patients.

It has been hypothesized that the development of cancer might partially result from a diminution of immunocompetence. Using ex vivo cytokine production by whole blood (WB) cells after polyclonal activation, we compared cytokine production levels of cancer patients to those of healthy controls. Seventeen patients without any prior treatment and attending the hospital for oncological surgery (for cancers of several origins) were enrolled in the study. WB was collected in heparinized tubes, diluted 1/10 in RPMI 1640 and incubated for 2, 4, 24, 48, and 72 h at 37 degrees C in the presence of 5 micrograms/ml PHA and 25 micrograms/ml LPS. Cytokine levels in the supernatant were measured by specific immunoassay kits. IL-10 levels after 24 h of culture, IFN-gamma and GM-CSF levels after 24 and 72 h of culture, and LIF levels after 72 h of culture were significantly lower in cancer patients than in healthy controls. No significant difference was observed for IL-1 beta, IL-2, IL-4, IL-8, and TNF-alpha production at any culture time. Our results suggest that the putative immunosuppression of cancer patients might be reflected by their reduced production of immunostimulated cytokines.

Characterisation of monoclonal antibodies against human interleukin-10 and their use in an ELISA for the measurement of this cytokine.

Biological and biochemical characteristics of monoclonal antibodies (MABs) raised against human interleukin-10 (IL-10) are described as well as their use in the design of a specific ELISA for the measurement of the cytokine. 21 murine anti-human interleukin-10 (IL-10) MABs were obtained by fusion of splenocytes from mice immunized against human recombinant IL-10 with SP2/0 myelomatous cells. These antibodies define three major antigenic areas on the IL-10 molecule, one of which comprises epitopes involved in receptor binding and induction of biological activity. They recognize recombinant human IL-10 with affinities ranging from 1.3 x 10(-7) to 3 x 10(-11), as well as natural IL-10. Most of them also recognize viral IL-10 (vIL-10) encoded by the Epstein-Barr virus (EBV). A specific human-IL-10 ELISA has been developed using two MABs (18 and 19) as capture antibody and one MAB (17) as detector. The sensitivity (3 pg/ml), precision (intra-assays < 4%), reproducibility (interassay < 3%), and accuracy (recoveries, ranging between 84 and 107%, in several fluids) of the assay, plus its excellent performance in dilution tests, and the lack of interference when in the presence of possible cross-reactive substances, permits accurate cytokine measurement in biological fluids such as serum, plasma, bronchoalveolar lavage, urine and culture supernatants. Using the assay, IL-10 was measurable in the plasma of patients with septic shock (range 11-2740 pg/ml) whereas IL-10 plasma levels were < 7.8 pg/ml in healthy volunteers.

An ELISA for the measurement of human leukemia inhibitory factor in biological fluids and culture supernatants.

A new monoclonal antibody-based ELISA for leukaemia inhibitory factor/human interleukin for DA cells (LIF/HILDA) measurements is described. The sensitivity (56 pg/ml after 4 h incubation, 14 pg/ml after 24 h incubation), precision (intra-assays < 5%), reproducibility (interassay < 10%), and accuracy (recoveries, ranging between 98 and 119%, in several fluids) of the assay, plus its excellent performance in dilution tests, and the lack of interference when in the presence of possible cross-reactive substances guarantee accurate cytokine measurement in biological fluids such as serum, plasma, synovial fluid, follicular fluid, urine and culture supernatants. Using the assay, LIF/HILDA was measurable in supernatants after in vitro whole blood stimulation with phytohemagglutinin (PHA), OKT3, and phorbol myristate acetate (PMA) but not with lipopolysaccharide (LPS) or Ca ionophore. LIF/HILDA production was not measurable until after 24 h of culture, when cytokine levels were seen to increase linearly in the supernatant to reach values of up to 40 ng/ml after 96 h of culture. Finally, a good correlation was found (r = 0.96; p < 0.0001; y = 23.1x + 233) between the LIF/HILDA values obtained using the ELISA and DA-1a bioassay.

Novel method for the measurement of cytokine production by a one-stage procedure.

A new one-step culture-immunoassay procedure is described for testing cytokine production by immunocompetent cells in whole blood (WB) without the need for an isolation step. Briefly, WB samples or distilled water were added to RPMI medium containing specific anti-cytokine peroxidase-labelled monoclonal antibodies and incubated in micro-well plates coated with specific capture monoclonal antibodies, directed against distinct epitopes of the cytokine, and containing dried polyclonal activators (5.625 micrograms LPS + 1.125 micrograms PHA) or dried standards respectively. The optimalisation of the assay is described for an extended measurement range. The best compromise between sensitivity and linearity was obtained with the addition of 50 ng/well for TNF-alpha and IL-6 or 100 ng/well for IFN-gamma of unconjugate antibodies to the corresponding conjugate. The kinetics of individual production of each cytokine in WB of normal healthy donors showed values entering the standard range following incubation times of between 2 and 8 h for TNF-alpha, 2 and 4 h for IL-6, and 4 and 24 h for IFN-gamma. The sensitivity, the precision (intra-assay CVs) and the reproducibility (interassay CVs) of the assays were as follows: 70 pg/ml, < or = 14% and < or = 11% for TNF-alpha; 25 pg/ml, < or = 11% and < or = 16% for IL-6; 25 pg/ml, < or = 19% and < or = 20% for IFN-gamma. The accuracy (% of recovery) of the assays was in the order of 100% and between 40 and 60% in the absence or presence of polyclonal activators, reflecting the occurrence of an active production/consumption mechanism during the activation.

Immunomodulatory properties of cyclic hexapeptide oxytocin antagonists.

The thymic repertoire of neuroendocrine 'self' antigens has been previously described on the basis of the intrathymic expression of neurohypophysial (NHP)- and tachykinin-related peptide signals and receptors. According to that model, the cryptocrine signalling between thymic epithelial/nurse cells and thymocytes through NHP-related signals and receptors constitutes one accessory pathway in the process of T-cell differentiation and/or activation. A pharmacological manipulation of that novel type of cell-to-cell signalling was tested by the investigation of the immunomodulatory properties of novel cyclic hexapeptide oxytocin (OT) antagonists (MSD Research Laboratories). These compounds were found to significantly inhibit the productions of cytokines (mainly IL-1 beta and IL-6) elicited by anti-CD3 treatment of human whole blood cell cultures. Cytokine productions were more significantly reduced by OT antagonists in whole blood cell cultures derived from female volunteers than in those obtained from male donors, suggesting an influence of the gonadal steroid environment on the expression of NHP peptide receptors by immune cells. These observations support the concept of novel immunomodulating approaches through immune-specific neuropeptide antagonists, as well as the pharmacological value of such strategies in selective immunotherapy.

Direct stimulation of cytokines (IL-1 beta, TNF-alpha, IL-6, IL-2, IFN-gamma and GM-CSF) in whole blood. I. Comparison with isolated PBMC stimulation.

Production of interleukin 1 beta (IL-1 beta), interleukin 6 (IL-6), tumor necrosis factor alpha (TNF-alpha), interleukin 2 (IL-2), interferon gamma (IFN-gamma) and granulocyte-macrophage colony-stimulating factor (GM-CSF) after stimulation by lipopolysaccharide (LPS) and phytohemagglutinin (PHA) was studied in 1/10 diluted whole blood (WB) culture and in peripheral blood mononuclear cell (PBMC) culture. Cytokines IL-1 beta, TNF-alpha and IL-6 are preferentially stimulated by LPS whereas IL-2, IFN-gamma and GM-CSF are stimulated by PHA. Combination of 5 micrograms/ml PHA and 25 micrograms/ml LPS gave the most reliable production of the six cytokines studied. IL-1 beta, TNF-alpha and IL-6 represent a homogeneous group of early-produced cytokines positively correlated among themselves and with the number of monocytes in the culture (LeuM3). Furthermore, IL-1 beta was negatively correlated with the number of T8 lymphocytes. IL-2, IFN-gamma and GM-CSF represent a group of late-produced cytokines. Kinetics and production levels of IL-6 and GM-CSF are similar in WB and PBMC cultures. In contrast, production levels of TNF-alpha and IFN-gamma are higher in WB than in PBMC whereas production levels of IL-6 and IL-2 are lower in WB than in PBMC. Individual variation in responses to PHA + LPS was always higher in PBMC cultures than in WB cultures. The capacity of cytokine production in relation to the number of mononuclear cells is higher in WB, or in PBMC having the same mononuclear cell concentration as WB, than in conventional cultures of concentrated PBMC (10(6)/ml). Because it mimics the natural environment, diluted WB culture may be the most appropriate milieu in which to study cytokine production in vitro.

Effects of a hydrosoluble bacterial extract from Escherichia coli (OM-89) on cytokine production by peripheral blood mononuclear cells from healthy subjects and patients with rheumatoid arthritis.

OM-89 is a bacterial extract from escherichia coli, proposed as an immunomodulating drug for the treatment of rheumatoid arthritis (RA). Since immunological mechanisms may play a role in its action, the immunological effects of OM-89 were evaluated in vitro on peripheral blood mononuclear cells (PBMC) derived from healthy subjects and RA patients. Results indicated that in the absence of OM-89, production of the monokines interleukin-1 beta (IL-1 beta) and tumor necrosis factor-alpha (TNF-alpha) is increased, while that of the lymphokines interleukin-2 (IL-2) and interferon-gamma (IFN-gamma is decreased by phytohemagglutinin (PHA)-stimulated PBMC from RA patients as compared with PBMC from healthy subjects. In the presence of PHA, OM-89 enhanced the production of IL-1 beta, TNF-alpha, IL-2, and IFN-gamma. IL-1 beta and IL-2 curves obtained using increasing amounts of OM-89 did not differ depending on the source of PBMC. By contrast, in the presence of increasing amounts of OM-89, TNF-alpha secretion significantly higher and IFN-gamma secretion significantly lower with PBMC from RA patients compared to PBMC from healthy subjects. These data indicate that OM-89 acts on monocytes and T cells directly and/or indirectly and suggest a possible clinical activity by OM-89 in RA relative to its immunological properties.

Expression of preprotachykinin-A and neuropeptide-Y messenger RNA in the thymus.

The preprotachykinin-A gene, the common gene of mRNAs encoding both substance-P (SP) and neurokinin-A (NKA), was shown to be expressed in Sprague-Dawley rat thymus by detection of specific mRNA in gel-blot analyses. In situ hybridization revealed dispersed PPT-A-labeled cells in sections from rat thymus, with a concentration of grains over a subpopulation of cells in the thymic medulla. Also, neuropeptide-Y mRNA-expressing cells were found in the rat thymus, primarily in the thymic medulla. Rat thymic extracts contained SP-like immunoreactivity (SP-LI), and the major part of the immunoreactivity coeluted with authentic SP and SP sulfoxide standards. SP-LI was also detected in human thymus, which contained between 0.09-0.88 ng SP-LI/g wet wt. Evidence for translation of preprotachykinin-A mRNA in the rat thymus was obtained from the demonstration of NKA-LI in thymic cells with an epithelial-like cell morphology. Combined with previous observations on the immunoregulatory roles of tachykinin peptides and the existence of specific receptors on immunocompetent cells, the demonstration of intrathymic synthesis of NKA suggests a role for NKA-LI peptides in T-cell differentiation in the thymus.

In vivo cell activation following OKT3 administration. Systemic cytokine release and modulation by corticosteroids.

A massive and self-limited release of tumor necrosis factor and interferon gamma was detected in the systemic circulation in 35 consecutive renal allograft recipients by specific radioimmunoassays very soon following the first injection of the monoclonal antibody OKT3 (anti-CD3). Peak serum TNF and IFN gamma levels were reached, respectively, at 1 and 4 hr following the first OKT3 injection. Abnormally high serum interleukin 2 levels were also observed 4 hr following the first OKT3 injection in a minority of patients (5 cases). OKT3 had no effect on interleukin 1 beta, interferon alpha, and granulocyte/macrophage colony stimulating factor serum levels, which in all patients remained within the normal range throughout the study. This selective OKT3-induced cytokine release, which only followed the first injection, was transient (i.e., lasting a few hours). It tightly paralleled the spontaneously reversible clinical syndrome characterized by high fever, headaches, and gastrointestinal symptoms that is invariably associated with the first OKT3 administration. Importantly, when administered in adequate dosages and with adequate timing, corticosteroids influenced both the cytokine release and the systemic reaction. Thus, the highest TNF, IFN gamma, and IL-2 serum levels were detected in patients who did not receive corticosteroids. Patients who received high-dose corticosteroids (1 g solumedrol bolus) concomitantly with the first OKT3 injection still had high TNF and IFN gamma levels. Conversely, when the same corticosteroid dose was injected 15-60 min prior to the first OKT3 injection, in all cases the increase of serum TNF and IFN gamma was significantly lower as compared with the above-described groups; IL-2 levels did not rise. These data offer a direct explanation for one major side effect of OKT3 and thus provide the basis for devising means to prevent its occurrence.

Radio-immunological study of the regulation of interleukin-2 production in peripheral blood mononuclear cell cultures from normal subjects and rheumatoid arthritis patients.

A radio-immunoassay for human T cell growth factor, also called Interleukin-2 (IL-2), has been carried out using a recombinant IL-2 preparation as tracer and a polyclonal rabbit antiserum. The assay is highly specific for IL-2: there is no cross-reaction with either type I and II interferons, epidermal growth factor or tumor necrosis factor alpha. Using the sequential saturation procedure the limit of sensitivity was 0.5 U/ml. Intra- and between-assay coefficients of variation were 8 and 11%, respectively. With this assay, IL-2 recovery in serum and peripheral blood mononuclear cell culture (P.B.M.C.) medium was 79 and 95%, respectively. In serum of 109 normal subjects and 102 rheumatoid arthritis patients mean IL-2 concentrations (+/- SD) were 1.5 +/- 0.5 U/ml and 1.4 +/- 0.4 U/ml respectively. The IL-2 production by P.B.M.C. in vitro was also studied. In unstimulated cultures, IL-2 release remained undetectable, i.e. below 0.5 U/ml. After stimulation of mononuclear cells from 36 normal subjects with increasing amounts of phytohemagglutinin (PHA), the 3H-thymidine incorporation followed a bell-shaped curve, the maximum response being observed at a 2.5 micrograms/ml PHA concentration. After a 72-hr mononuclear cell stimulation, IL-2 release increases with PHA concentrations ranging from 0 to 10 micrograms/ml. In patients with rheumatoid arthritis (R.A.), P.B.M.C. incorporated 3H-thymidine as in normal subjects. In contrast, mean +/- SEM IL-2 production by P.B.M.C. from patients with inactive RA (5 +/- 0.9) and active disease (1 +/- 0.5) was significantly lower than that from normal subjects (12 +/- 0.7 ng/ml).(ABSTRACT TRUNCATED AT 250 WORDS)

Production of interferon gamma by peripheral blood mononuclear cells from normal subjects and from patients with rheumatoid arthritis.

A radioimmunoassay for human interferon gamma (IFN-gamma) has been carried out using a recombinant glycosylated interferon (Hu IFN-gamma) as tracer, the N.I.H. reference preparation (Gg 23-901-530) and a polyclonal rabbit antiserum. The assay is highly specific for IFN-gamma: there is no cross-reaction either with interferons alpha and beta, Interleukins 1 and 2, tumor necrosis factor alpha and beta or with various brain peptides. The sequential saturation procedure allowed a sensitivity of 0.4 U/ml with intra and between assay coefficients of variation less than 8 and 12%, respectively. The in-vitro production of IFN-gamma by peripheral blood mononuclear cells (P.B.M.C.) was also measured. In unstimulated cultures, IFN-gamma production remained undetectable, i.e. below the 0.4 U/ml sensitivity level. After stimulation of P.B.M.C. from normal subjects with increasing amounts of PHA, both the 3H-thymidine incorporation and IFN-gamma release followed bell-shaped curves. There was no significant difference of 3H-thymidine incorporation between PHA stimulated cultures (0.2 and 2.5 ug/ml) from normal subjects (36 cases) and those with active (16 cases) or non-active (14 cases) rheumatoid arthritis. At two PHA concentrations of 0.2 and 2.5 ug/ml, mononuclear cells from patients with active disease produced significantly less IFN-gamma than those from either controls or cases with non-active disease.

[T lymphocyte activation induced in vivo by the first injection of OKT3 monoclonal antibodies]. / Activation lymphocytaire T induite in vivo par la première injection de l'anticorps monoclonal OKT3.

One of the major side effects induced by the in vivo administration of the murine monoclonal antibody OKT3 is a spontaneously reversible clinical syndrome associating in variable proportions depending on the patient: fever, chills, headaches, diarrhea and seldomly meningismus. Sera from 3 renal allograft recipients treated with OKT3 were studied and showed that a massive although transient release of some cytokines namely, Tumor Necrosis Factor alpha, Interleukin 2 and Interferon gamma is observed following the first OKT3 injection.

Production of tumour necrosis factor-alpha, interferon-gamma and interleukin-2 by peripheral blood mononuclear cells of subjects suffering from rheumatoid arthritis.

Using radio-immunoassay methods, the production of tumour necrosis factor-alpha (TNF-alpha), interleukin-2 (IL-2), and interferon-gamma (IFN-gamma) released by peripheral blood mononuclear cells (PBMC), maintained in culture and stimulated by phytohemagglutinin (PHA), was measured in normal subjects and patients with active or inactive rheumatoid arthritis (RA). Results indicated a dissociation between mitogenic response and secretion of mediators by PBMC under the influence of PHA in both normal controls and in patients with rheumatoid arthritis (RA). While [3H]thymidine incorporation was characterized by a rather bell-shaped curve with increasing concentrations of PHA, IL-2 and TNF-alpha displayed a linear dose-dependent increase. [3H]thymidine uptake by PBMC was in the same range in normal subjects as in patients with active and inactive RA, although cytokine secretion differed. The PBMC of patients with active RA produced less TNF-alpha, IL-2, and IFN-gamma than did those of the controls. In cases of inactive RA, the secretory response varied from subject to subject; mean values did not differ from those of normal subjects, except for those of IL-2 (p less than 0.01). The significance and the clinical relevance of these findings are discussed.

Effect of diclofenac on plasma levels of immunoreactive prolactin, follicle stimulating hormone, luteinizing hormone, thyrotropin, and beta-endorphin in humans.

Prostaglandins have been shown to modulate the secretion of several pituitary hormones, suggesting that therapeutic doses of nonsteroidal anti-inflammatory drugs may change basal hormone levels. In this study, plasma levels of prolactin, follicle stimulating hormone, luteinizing hormone, thyrotropin and beta-endorphin were determined in 6 healthy men after administration of diclofenac, a prostaglandin synthesis inhibitor. The subjects were given 75 mg intramuscularly and 50 mg orally at 08.00 h the first day, 50 mg orally at 08.00, 12.00 and 20.00 h the second day and an additional 50 mg orally at 08.00 h the third day. Blood samples were collected throughout these 3 days. Diclofenac resulted in a significant and sustained decrease in plasma level of prolactin (p less than 0.005). The other hormones did not demonstrate significant change following diclofenac administration. These data suggest that administration of a prostaglandin synthesis inhibitor, such as diclofenac, selectively alters basal pituitary secretion of prolactin in humans without a detectable effect on plasma levels of other pituitary hormones. This study supports the hypothesis that prostaglandins are necessary for maintaining basal level of prolactin secretion in man.

Thymopoietin and thymopentin enhance the levels of ACTH, beta-endorphin and beta-lipotropin from rat pituitary cells in vitro.

Thymopoietin and thymopentin are well characterized polypeptides influencing immunoregulation by several mechanisms. Proposed as a therapy in diseases with major immune abnormalities such as rheumatoid arthritis, thymopentin improved within 2 weeks some clinical parameters as pain and joint swelling. The hypothesis that this spectacular effect could be mediated through interactions with anti-inflammatory (ACTH) and pain relieving (beta-endorphin) hormones producing cells was tested on the rat isolated pituitary cell model. Thymopentin and thymopoietin can enhance in vitro the levels of ACTH, beta-endorphin and beta-lipotropin in a time- and dose-dependent fashion for physiological concentrations ranging from 10(-12) to 10(-8) mol/l. The action on pituitary cells was restricted to those molecules as no changes occurred in LH, FSH, GH, TSH and PRL levels, after otherwise identical experimental conditions.

In vitro effects of thymopentin on the gamma-interferon production by peripheral blood mononuclear cells from normal subjects and from patients with rheumatoid arthritis.

The in vitro production of gamma-interferon (gamma-IFN) by peripheral blood mononuclear cells (MNC) was measured using a specific radioimmunoassay in 16 patients presenting with active rheumatoid arthritis (RA), in 14 patients with inactive disease, and in 36 control subjects (CS). Unstimulated cultures produced undetectable levels of gamma-IFN and did not incorporate tritiated thymidine. In response to phytohemagglutinin (PHA) 0.2 microgram/ml, MNC from active RA produced 9 times less, and under PHA 2.5 micrograms/ml, 4 times less gamma-IFN than did MNC from inactive RA or from CS. The uptake of tritiated thymidine was, however, similar in the 3 groups. In unstimulated cultures of the 3 groups, thymopentin (TP-5), at all concentrations tested, did not influence either the levels of gamma-IFN or the uptake of tritiated thymidine. In the presence of PHA 0.2 microgram/ml and TP-5, lambda-IFN levels were increased in CS, unchanged in inactive RA and reduced in active RA, whereas no changes were observed in the uptake of tritiated thymidine. Our results show that under our experimental conditions, TP-5 was able to increase the levels of gamma-IFN produced by normal MNC in vitro, but could not reverse the profound defect observed in active RA.