Linomide suppresses experimental autoimmune neuritis in Lewis rats by inhibiting myelin antigen-reactive T and B cell responses

Abstract

Linomide (quinoline-3-carboxamide) is a synthetic immunomodulator that suppresses several experimental autoimmune diseases. Here we report the effects of Linomide on experimental autoimmune neuritis (EAN), a CD4+ T cell-mediated animal model of acute Guillain-Barré syndrome (GBS) in humans. EAN induced in Lewis rats by inoculation with bovine peripheral nervous system (PNS) myelin and Freund's complete adjuvant was strongly suppressed by Linomide administered daily subcutaneously from the day of inoculation. Linomide dose-dependently delayed the interval between immunization and onset of clinical EAN, as well as the severity of EAN symptoms. These clinical effects were associated with dose-dependent down-modulation of PNS antigen-induced T and B cell responses and with suppression of the proinflammatory cytokines IL-12, interferon-gamma (IFN-gamma) and tumour necrosis factor-alpha (TNF-alpha) mRNA. In PNS sections, Linomide suppressed IL-12 and TNF-alpha, and up-regulated IL-10 mRNA expression. These findings suggest that Linomide could be useful in certain T cell-dependent autoimmune diseases.

Fig. 1

Clinical scores of experimental autoimmune neuritis (EAN) in Lewis rats in two different experiments (a and b) treated with Linomide at 1.6, 16 and 160 mg/kg per day and in control rats receiving PBS. Symbols indicate mean clinical scores for each group on the indicated day.

Fig. 2

Body weight of experimental autoimmune neuritis (EAN) Lewis rats treated with Linomide at 1.6, 16 and 160 mg/kg per day and of control rats receiving PBS in two different experiments (a and b). Symbols indicate mean body weight of each group on the indicated day.

Fig. 3

Proliferation of lymph node mononuclear cells (MNC) at day 27 post-immunization from experimental autoimmune neuritis (EAN) rats treated with Linomide (1.6, 16 and 160 mg/kg per day) and control rats treated with PBS, after ex vivo culture without antigen or mitogen, and in the presence of phytohaemagglutinin (PHA), bovine peripheral nerve myelin (BPM) and P2 peptide, respectively. Means and s.d. are indicated. P values refer to comparisons between EAN rats treated with Linomide and PBS-treated EAN control rats. *P < 0.05; **P < 0.01; ***P < 0.001.

Fig. 4

Numbers of IFN-γ-secreting cells per 10⁵ lymph node mononuclear cells (MNC) at day 27 post-immunization from experimental autoimmune neuritis (EAN) rats treated with Linomide (1.6, 16 and 160 mg/kg per day) and controls, after ex vivo culture without antigen or mitogen, and in the presence of phytohaemagglutinin (PHA), bovine peripheral nerve myelin (BPM) and P2 peptide, respectively. Means and s.d. are indicated. P values refer to comparisons between EAN rats treated with Linomide and PBS-treated control EAN rats. *P < 0.05; **P < 0.01; ***P < 0.001.

Fig. 5

The effect of Linomide on anti-peripheral nervous system (PNS) myelin and anti-purified protein derivative (PPD) IgG antibodies. Serum specimens obtained on day 27 post-immunization from experimental autoimmune neuritis (EAN) rats treated with Linomide (1.6, 16 and 160 mg/kg per day) and from PBS-treated EAN controls. Means and s.d. are indicated. P values refer to comparisons between EAN rats treated with Linomide and PBS-treated control EAN rats. *P < 0.05; **P < 0.01.

Fig. 6

Numbers of cells expressing mRNA for IFN-γ (a), tumour necrosis factor-alpha (TNF-α) (b), IL-12 (c), IL-10 (d) and transforming growth factor-beta (TGF-β) (e) per 10⁵ lymph node mononuclear cells (MNC) from experimental autoimmune neuritis (EAN) rats treated with Linomide (1.6, 16 and 160 mg/kg per day) and PBS-treated control rats, after ex vivo culture without antigen or mitogen, and in the presence of phytohaemagglutinin (PHA), bovine peripheral nerve myelin (BPM) or P2 peptide (left ordinate), and per 100-mm² tissue section of sciatic nerve (right ordinate), obtained on day 27 post-immunization. mRNA expression was detected by in situ hybridization with ³⁵S-labelled synthetic oligonucleotide probes. Bars indicate 1 s.d. P values refer to comparisons between EAN rats treated with Linomide and PBS-treated EAN control rats. *P < 0.05; **P < 0.01.