Effects of Clostridium difficile toxin B on human monocytes and macrophages: possible relationship with cytoskeletal rearrangement.

Toxin B from Clostridium difficile is cytopathic in vitro for various types of cells, including polymorphonuclear cells, lymphocytes, and monocytes. Since intestine lamina propria is rich in macrophages, we studied the effect of toxin B on human monocytes and on human macrophages generated in vitro by long-term culture of purified circulating blood monocytes. Upon addition of toxin B, human monocytes exhibited few modifications whereas macrophages adopted a stellate morphology, with rounding up of the perikaryon. Toxin B made microfilaments of actin disappear and induced an important reorganization of vimentin and a redistribution of tubulin. Membrane area increased by approximately 16%. Toxin B did not affect the viability of human mononuclear phagocytes and did not exert any significant lytic effect. It profoundly altered the phagocytic function of macrophages. When activated by gamma interferon in the presence of toxin B, monocytes were more cytotoxic for U-937 target cells than control monocytes activated in absence of toxin. Finally, the combined treatment of monocytes with gamma interferon and toxin B increased significantly the secretion of tumor necrosis factor alpha, whereas toxin B alone was unable to induce tumor necrosis factor production. These results suggest that morphological and functional alterations induced in human mononuclear phagocytes by toxin B from C. difficile are due to the disorganization of the cytoskeleton and the resulting impairment of the membrane traffic equilibrium.

Activation of human monocytes by Streptococcus mutans serotype f polysaccharide: immunoglobulin G Fc receptor expression and tumor necrosis factor and interleukin-1 production.

Streptococcus mutans serotype f polysaccharide (poly f) was prepared from S. mutans whole cells by autoclaving. The poly f was purified by chromatography on DEAE Trisacryl M and Bio-Gel P100, treated with insoluble pronase, and resubjected to chromatography on DEAE Trisacryl M. Normal human blood monocytes, stimulated in vitro with purified poly f, produced extracellular tumor necrosis factor alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta) in a dose-dependent fashion as determined by a heterologous two-site sandwich enzyme-linked immunosorbent assay. Poly f also increased the expression of monocyte cell surface receptors for the Fc part of human immunoglobulin G, activity which is correlated with an increase of the phagocytic activity of the stimulated monocytes. Polymyxin B had no effect on TNF-alpha and IL-1 beta release. Neutralization assays with anti-recombinant human TNF-alpha and anti-recombinant human IL-1 beta immunoglobulin G confirmed the fact that the cytotoxic and mitogenic mediators released by the poly f-stimulated monocytes were mainly TNF-alpha and IL-1 beta.