Irène Florentin

Effects of interleukin-6 on cytochrome P450-dependent mixed-function oxidases in the rat

Intravenous treatment of male rats with recombinant human interleukin-6 (rhIL6) at 50, 100 and 200 micrograms/kg (corresponding to 4, 8 and 16 x 10(4) U/animal, respectively) reduced the activities of hepatic microsomal cytochrome P450-dependent monoxygenases to varying degrees. Ethylmorphine-N-demethylase activity fell to 53% of control values, an effect similar to that induced by 2.5 mg/kg Escherichia coli lipopolysaccharide (LPS). Ethoxycoumarin-O-deethylase activity was also sensitive to inhibition, whereas IL6 had little effect on the activities of other P450-dependent enzymes, including ethoxyresorufin-O-deethylase. Pentoxyresorufin dealkylase activity, which is representative of the cytochrome P450 IIB 1/2 subfamily, was unaffected by IL6 whereas LPS reduced it to 33.7% of control values. Another hepatocyte-related parameter, serum concentration of alpha 1-acid glycoprotein (AGP), was increased by up to 3.5-fold over baseline by IL6 and 10-fold by LPS. Recombinant human interleukin-1 beta (rhIL1 beta) (10 micrograms/kg, corresponding to 5 x 10(4) U/rat) and recombinant human tumor necrosis factor alpha (rhTNF) (150 micrograms/kg corresponding to 24 x 10(4) U/rat) were both as potent as LPS (2.5 mg/kg) in increasing serum AGP levels and reducing hepatic microsomal monoxygenase activities. IL6 did not potentiate the effects of rhIL1 beta. Hepatic microsomal glucuronyltransferase activities were little affected by LPS and unaffected by rhIL6. Finally, rhIL6 was more potent after i.p. injection than after i.v. or s.c. injection. These results suggest that the effects of LPS, TNF and IL1 on the mixed-function oxidase system in vivo may be due partly to an induction of IL6 in vivo. The different sensitivities of the enzymes to IL6 but not to IL1 or TNF may be due to the involvement of two distinct mechanisms.

Inhibition of human monocyte TNF production by adenosine receptor agonists

Adenosine receptor agonists and agents enhancing pericellular concentrations of adenosine possess antiinflammatory properties. In the present study, we found that R-phenylisopropyladenosine (R-PIA), 5-N-ethylcarboxamido adenosine (NECA), other agonists of adenosine receptors and dipyridamole, an adenosine uptake inhibitor, inhibited tumor necrosis factor (TNF) production by endotoxin-stimulated human monocytes in a concentration-dependent manner with no inhibition of interleukin-6. The rank order of agonist potency is characteristic of neither A1 nor A2 receptors and suggests the involvement of another receptor subtype. The effect of R-PIA on TNF was in part abolished by the antagonist 8-sulfophenyltheophylline. In endotoxin-treated rats, R-PIA pretreatment (2.5 mg/kg) reduced serum TNF levels by 98%, with no modification of serum IL6 levels. TNF inhibition could be an important mechanism by which adenosine analogs exert their antiinflammatory action.

Acute-phase response, interleukin-6, and alteration of cyclosporine pharmacokinetics.

Administration of interleukin‐6 partially reproduces the inhibitory effects of the acute‐phase response on cytochrome P450‐dependent drug metabolism. The aim of the study was to determine whether endogenous cytokine has such an effect in patients treated by cyclosporine, which is metabolized by the cytochrome P4503A subfamily.

When Should the Immune Clock Be Reset

Immune defenses are organized along both 24-h and yearly time scales. Two circadian systems have been isolated in man, which can be desynchronized: (1) the circulation of T, B, or NK lymphocyte subsets in peripheral blood and (2) the density of epitope molecules (CD3, CD4, ...) at their surface, which may relate to cell reactivity to antigen exposure. The in vitro response of murine splenocytes to interleukin 2, interferon (IFN), or cyclosporin A strongly depended upon circadian time of exposure. Temporally optimized delivery of biologic response modifiers (BRM) may be guided by immunologic marker rhythms. An alternative yet complementary strategy was sought with IFN: since high doses were shown as more effective than low doses against several malignancies, this drug was given at the presumed less toxic time, so that its dose could be increased. Continuous drug delivery was circadian modulated in 8 cancer patients. Dose intensities twice to fourfold higher than those usually recommended were safely infused to ambulatory patients. Chronotherapy with BRM may represent a necessary step for optimizing the immunologic control of malignancies.

Circannual rhythm in natural killer activity and mitogen responsiveness of murine splenocytes

Seasonal variations were observed in murine splenic natural killer (NK) cell activity and also in murine lymphocyte responsiveness to mitogens, namely, concanavalin A, phytohemagglutinin, and lipopolysaccharide. The maximum and minimum splenic NK cell activities were observed in January-February and July-August, respectively. Conversely, maxima and minima of lymphoproliferative responses to all the three mitogens occurred in April-June and January-February, respectively. Such variations, when inferential statistics are used, appeared to be accounted for by circannual and other low-frequency (infradian) bioperiodicities. More specifically, the circannual rhythm in murine NK cell activity was demonstrated in data from a total of 356 mice collected over a period of 5 years. The various components of the immune system are characterized by a multifrequency time structure. The understanding of the organization of the immune system along the yearly scale may have bearings on that of the seasonal incidence of numerous infectious diseases and on the success/failure of immunotherapy.

Nitric oxide mediates the depression of lymphoproliferative responses following burn injury in rats

Among the multiple biological activities of nitric oxide (NO) an immunoregulatory role consisting of the mediation of macrophage suppressive activity, has recently been evidenced. In the present work, we investigated whether NO was implicated in immunosuppression following burn injury. Thermal injury affecting 20-25% of the total body surface area in Wistar rats, provoked a biphasic depression of spleen cell proliferative responses to phytohemagglutinin (PHA) and concanavalin A (Con A). We show that these responses are fully restored on day 4 after burn and only by 55% on day 10 when spleen cells were stimulated in the presence of NG-monomethyl-L-arginine (NMMA), a potent inhibitor of the macrophage inducible NO synthase. Nitrite content in culture supernatant, as an indicator of NO release (in the absence of NMMA), was significantly augmented in Con A-stimulated spleen cells from burned rats as compared to normal spleen cells. These results show for the first time that NO is implicated, at least in part, in an immunosuppression state which is not linked to an infectious disease.

Nitric oxide inhibits spleen cell proliferative response after burn injury by inducing cytostasis, apoptosis, and necrosis of activated T lymphocytes: role of the guanylate cyclase.

We previously showed that an overproduction of nitric oxide (NO) by macrophages was responsible for the collapse of lymphoproliferative responses after burn injury in rats. First, we demonstrate here that 10 days post-burn, the inhibition of splenocyte response to concanavalin-A results from cytostatic, apoptotic, and necrotic effects of NO on activated T cells. This was evidenced by various criteria at the levels of DNA, mitochondria, and plasma membrane. Inhibition of NO synthase by S-methylisothiourea (10 microM) normalized all the parameters. Second, we show that two soluble guanylate cyclase (sGC) inhibitors, LY83583 and ODQ, restored the proliferative response in a concentration-dependent manner. LY83583 (0.5 microM) rescued T cells from apoptosis. Similar results were obtained with KT5823 (5 microM) a specific inhibitor of protein kinase G (PKG). In contrast, neither LY83583 nor KT5823 inhibited NO-induced necrosis. These results suggest that NO blocked T cells in the G1 phase and induced apoptosis through a sGC-PKG-dependent pathway and necrosis through an independent one.

The lymphocyte inhibiting factor extracted from the thymus (LIFT): inhibition of the DNA synthesis in vitro.

Abstract The action of a lymphocyte inhibiting factor extracted from the thymus (LIFT) has been investigated by studying its capacity to inhibit in vitro , independently of any cytotoxic effect, DNA synthesis in different types of mouse lymphoid cells. [ 3 H]TdR incorporation was strongly depressed under the influence of the thymic extract in short term cultures of thymus, bone marrow cells as well as of spleen cells coming from: 1) normal CBA mice; 2) mice treated with PHA. Similarly in vitro PHA stimulation of spleen cells coming from normal mice was depressed when thymic extract was added to the culture. By contrast, the thymic extract does not significantly decrease [ 3 H]TdR uptake in spleen cells coming from: 1) normal CBA mice injected with LPS; 2) T lymphocyte deprived mice, whether or not their B lymphocytes were activated by in vivo LPS administration. These results suggest that the LIFT acts on T lymphocytes at different stages of their maturation and differentiation. The inhibiting effect of the thymic extract upon [ 3 H]TdR incorporation in bone marrow cells could be the result of an action on erythrocytic and leukocytic precursors.

Restoration of postburn impaired lymphocyte responsiveness by nonsteroidal anti‐inflammatory drugs is independent of prostaglandin E2 inhibition

Prostaglandin E2 (PGE2) has been implicated in postburn immunosuppression, which is responsible for septic complications. In the present work, seven nonsteroidal anti‐inflammatory drugs (NSAIDs), differing by their capacity to inhibit the cyclooxygenase pathway, were compared for their ability to restore T lymphocyte proliferative responses evaluated 4 days after thermal injury in rats. Salicylic acid, 5‐aminosalicylic acid, and niflumic acid, given daily, fully restored spleen cell responses to concanavalin A (Con A) and phytohemagglutinin. These drugs were active only at doses that were below the anti‐inflammatory doses and did not modify normal spleen cell responses. In these conditions, indomethacin slightly restored lymphocyte reactivity, whereas acetylsalicylic acid, ketoprofene, and piroxicam were ineffective. PGE2 production by Con A‐stimulated spleen cells from untreated burned rats and after treatment with niflumic acid or 5‐aminosalicylic acid did not correlate with the intensity of the proliferative response. Indomethacin, niflumic acid, and 5‐aminosalicylic acid were added in vitro to spleen cells from normal and burned rats, at concentrations from 10‐7 to 10‐4 M. PGE2 production was strongly depressed by indomethacin and niflumic acid and not modified by 5‐aminosalicylic acid. The proliferative response of normal spleen cells was depressed in a concentration‐dependent manner by niflumic acid and slightly inhibited at the highest concentrations of indomethacin. In contrast, indomethacin concentration dependently restored the burn‐impaired proliferative response, whereas niflumic acid further depressed it and 5‐aminosalicylic acid had no effect. These results demonstrate that only some NSAIDs are able to restore T lymphocyte reactivity impaired after thermal injury and that this property is not related to inhibition of PGE2 production. J. Leukoc. Biol. 55: 64–72; 1994.

Modulation of immune responses in mice by oral administration of niflumic acid

The in vivo effects of non-steroidal anti-inflammatory drugs (NSAIDs) on the host immune system are still poorly understood. However, through inhibition of prostaglandin synthesis, NSAIDs may exhibit immunomodulating properties. The present work was aimed at evaluating the influence of niflumic acid on immune responses when administered orally for 7 consecutive days to 8-week-old inbred mice. Immunological tests were performed 24 h after the arrest of the treatment. At a dosage of 50 mg/kg/day, niflumic acid exerted noticeable immunostimulating effects, as shown by an increase in plaque-forming cell numbers after in vivo immunization with sheep red blood cells, an augmentation of spleen cell proliferation responses to stimulation with T- or B-cell mitogens and of T-cell cytotoxic response to allogenic cells. Phagocytosis-induced chemiluminescence of peritoneal macrophages was also enhanced whereas interleukin-1 production by these cells was depressed, but without concomitant modification in interleukin-2 production by T-cells. Increasing the niflumic acid dosage to 75 mg/kg resulted in the disappearance of the immunostimulatory effects on lymphocytes responses. Macrophage activities were affected similarly in mice receiving 50 mg/kg. These results demonstrate that niflumic acid is able to stimulate in vivo several immunological functions and, consequently, to maintain host immune defenses. Interestingly, it depressed interleukin-1 production, known to play a major role in the inflammatory process.