Monique Parant

In vitro spleen cell responsiveness to various analogs of MDP (N-acetylmuramyl-l-alanyl-d-isoglutamine), a synthetic immunoadjuvant, in MDP high-responder mice

Abstract N -Acetylmuramyl- l -alanyl- d -isoglutamine (MDP), a synthetic immunoadjuvant, was incubated with spleen cells of DBA/2 or Balb/c mice and optimal responses were obtained after 4 or 5 days of culture in a serum-free medium supplemented with 2-mercaptoethanol. In contrast, lymphocytes of (C57B1/6 × AKR)F 1 hybrids responded weakly under the same conditions. The results reported here show that like in the case of DBA/2 and Balb/c strains, spleen cells of Swiss mice and of inbred AKR and CBA mice could be stimulated in vitro whereas C57B1/6 and LPS-refractory C3H/He mice did not respond. Fourteen synthetic MDP analogs (eight known to be adjuvant active and six devoid of activity) were tested in DBA/2 high-responder mice. A good correlation was observed between in vitro stimulation and the presence or absence of adjuvant activity in vivo of these compounds.

Priming effect of muramyl peptides for induction by lipopolysaccharide of tumor necrosis factor production in mice.

Lipopolysaccharide‐induced necrosis of grafted tumors was potentiated by several hydrophilic and lipophilic muramyl dipeptide (MDP) derivatives administered a few hours prior to small amounts of lipopolysaccharide (LPS) in spite of low titers of induced circulating tumor necrosis factor (TNF). However, pretreatment with MDP derivatives did increase the level of TNF in the blood of mice challenged by a greater dose of LPS. The TNF amount in 2 h postendotoxin mouse serum reached a peak when the glycopeptide had been given 6 h before the challenge, being approximately 100‐fold above that obtained in unprimed mice. The cytotoxic activity in mouse serum was inhibited by rabbit antibodies raised against recombinant mouse TNF. Although there exists a toxic synergism between BCG or MDP and endotoxin, the effect of certain MDP derivatives was not related to an increased susceptibility to the toxicity of LPS.

Nonspecific activation of murine spleen cells in vitro by a synthetic immunoadjuvant (N-acetyl-muramyl-L-alanyl-D-isoglutamine).

Abstract We reported previously that synthetic N -acetyl-muramyl- l -alanyl- d -isoglutamine (MDP) displayed marked adjuvant activity but was devoid of mitogenicity in vitro . The data reported here establish that, under different cultural conditions, thymidine uptake and blast cells can be increased by MDP in spleen cells of DBA/2 and Balb/c mouse strains. Optimal responses were obtained on culture in a serum-free medium supplemented with 2-mercaptoethanol for 4 or 5 days. This effect was also obtained with spleen cells of Balb/c nude mice. When the synthetic MDP was compared to a natural water-soluble adjuvant (neo-WSA), extracted from Mycobacterium smegmatis cells, both were found to stimulate [ 3 H] thymidine incorporation by mouse spleen cells. However, with the neo-WSA, the effect peaked on Day 2 and was weak or absent on Days 4 and 5. When the cells were cultured in a medium containing fetal calf serum, neo-WSA activation was completely abolished, while MDP-mediated stimulation was decreased.

Indirect and selective down-regulation of serum tumor necrosis factor-α release by interleukin-1β

A selective inhibition of LPS-induced tumor necrosis factor-alpha (TNF) response in mice was caused by an injection of recombinant human interleukin-1 (IL-1). The decrease in serum TNF level reached 70 to 80 percent of the controls receiving LPS alone when IL-1 was given simultaneously or prior to the challenge. At the same time serum IL-6 release was more elevated. Ex vivo assays have shown that macrophages from IL-1 treated animals did not respond to LPS when stimulated immediately after harvesting but recovered their normal responsiveness after being cultured for 2 hours and then washed. In vitro with or without addition of IL-1, mouse elicited macrophages responded equally to LPS in releasing TNF. In the absence of a direct and lasting effect on TNF-producing cells, the host reaction responsible for the inhibitory effect of IL-1 could be related to the overproduction of corticosterone that occurred after IL-1 injection, since it was not observed in adrenalectomized animals. Indeed the blockade of corticoid secretion by indomethacin prevented the inhibition of TNF production induced by IL-1 administration before LPS challenge. TNF administration did not result in elevation of corticosterone level and in contrast to IL-1 enhanced the TNF response to LPS injection. In vitro and ex vivo assays have shown this enhanced response to LPS was linked to a direct and prolonged effect of TNF on TNF-producing cells. Muramyl dipeptide (MDP) which was used as a known priming agent for enhanced cytokine release had a similar effect on TNF-producing cells.(ABSTRACT TRUNCATED AT 250 WORDS)

MDP Derivatives and Resistance to Bacterial Infections in Mice

Among the numerous biological response modifiers (BRMs) capable of increasing nonspecific immunity to infections, MDP (or muramyl dipeptide) derivatives represent remarkable tools for experimental studies and some of them appear to be suitable for human use because of immunostimulating properties can be separated from some unwanted side effects.

Changes in Rabbit Febrile Responses to Muramyl Dipeptide (MDP) After Coupling to a Synthetic Carrier

Muramyl dipeptide (MDP) is a small molecular weight synthetic glycopeptide (< 500), which has been shown to be an immunoadjuvant, and to induce a biphasic febrile response in the rabbit—probably via the release of endogenous pyrogen—accompanied by a marked leukopenia. Macromolecularization by coupling to a synthetic carrier (MW = 60,000) potentiates the immunostim‐ulant properties of MDP but also its pyrogenicity. The present study demonstrates that such a conjugate induced the release of endogenous pyrogen in vivo and in vitro at lower dosage levels than free MDP. Further experiments showed that there existed several differences between free and conjugated MDP. Thus, after intravenous administration of the conjugate, the fever pattern was monophasic with a prompt defervescence and not accompanied by leukopenia at dosage levels inducing similar increase in body temperature. In addition, when fever was recorded after intracerebroventricular administration, the increase in sensitivity was much greater in the case of free MDP than of MDP‐A‐L.

Dose-dependent efficacy of vaccination against K. pneumoniae in high and low antibody responder lines of mice

Innate and acquired resistance to Klebsiella pneumoniae infection were investigated in high (HI) and low (LI) antibody responder lines of mice. The two lines were very susceptible to infection since even small inoculum doses of a virulent strain provoked a 100% mortality within a few days. However the mean survival time was significantly longer in LI than in HI. (HI X LI) F1 hybrids were more resistant than both parental lines. Immunization with heat killed K. pneumoniae was able to confer full protection on the mice in the two lines. However there was a large difference in the number of killed bacteria required to induce the protective effect in HI and in LI mice. The dose-effect relationship for protection correlated with that of antibody production. The protective role of antibodies was confirmed by the survival of HI and LI mice, when antibodies were passively given prior to lethal challenge. The results are in agreement with the fact already demonstrated, that the defect of LI mice in antibody responsiveness is a quantitative one. Therefore a satisfactory immune protection against K. pneumoniae could be obtained in LI mice by adapting the vaccination procedure.

On the Nature of Some Nonspecific Host Responses in Endotoxin-Induced Resistance to Infection

“Tolerance” to endotoxins can be established by a previous injection of an unrelated O antigen which increases the host’s resistance and his capacity for clearing the second dose of endotoxin. It has been shown that these responses are nonspecific, i.e., not mediated by antibodies, but are produced by degradation and detoxification of endotoxic lipopolysaccharides [1, 2, 3].