Reduced physeal area and chondrocyte proliferation in Pasteurella multocida toxin-treated rats.

Pasteurella multocida toxin depresses weight gain in rats and pigs. It also affects tissues with rapidly dividing cells. In the present study, we investigated the role of this protein toxin on chondrocyte growth in vivo. Rats were divided into a single- or multiple-dose group and were given, respectively, either a single injection (0.15 or 0.6 micrograms/kg toxin subcutaneously) or multiple injections (0.01-0.2 micrograms/kg subcutaneously) of toxin. Bone (humerus) and other selected tissues were stained for bromodeoxyuridine immunoreactivity (BrDU-IR) in order to gauge cell proliferation. Physeal area was measured in rats from the multiple-dose group. Serum from single- and multiple-dose groups were tested for tumor necrosis factor alpha (TNF-alpha) and interleukin 6 (IL-6) activity using a bioassay system. Decreased weight gain, feed intake, and feed efficiency were observed in single- and multiple-dose groups of rats. Decreased BrDU-IR indices were present in the resting and proliferative zone chondrocytes of the humeral physis in rats from the multiple-dose group, as was decreased physeal area. Increased serum IL-6 bioactivity was present in rats after 24 hours, and no changes in TNF-alpha bioactivity were seen in any group. No alterations in BrDU-IR were seen in rats fed restricted (80% of control) diets. These studies show that sublethal doses of toxin decrease weight gain and affect growth of long bones through suppression of chondrocyte proliferation. These effects may be mediated by direct binding of the toxin to target cells or IL-6 but are not associated with altered feed intake or TNF-induced cachexia.

The effect of L-amino acid oxidase on activity of melphalan against an intracranial xenograft.

We have previously shown that diet restriction-induced depletion of large neutral amino acids (LNAAs) in murine plasma to 46% of control significantly enhances intracranial delivery of melphalan without enhancing delivery to other organs. Studies have now been conducted to determine whether more substantial LNAA depletion could further enhance intracranial delivery of melphalan. Treatment with L-amino acid oxidase (LOX) significantly depleted murine plasma LNAAs: phenylalanine, leucine, and tyrosine (> 95%); methionine (83%); isoleucine (70%); and valine (46%). Experiments evaluating the intracellular uptake of melphalan and high-pressure liquid chromatography quantitation of melphalan metabolites revealed, however, that melphalan is rapidly degraded in the presence of LOX, and that the timing of the administration of melphalan following the use of LOX to deplete LNAAs is crucial. Conditions were found under which LOX-mediated degradation of melphalan was minimized and LNAA depletion was maximized, resulting in a potentiation of the antitumor effect of melphalan on human glioma xenografts in nude mice. Such potentiation could not be obtained using diet restriction alone.