ABSTRACT
Recently, there has been considerable interest in using 4-methylumbelliferone (4-MU) to inhibit hyaluronan (HA) synthesis in mouse models of cancer, autoimmunity and a variety of other inflammatory disorders where HA has been implicated in disease pathogenesis. In order to facilitate future studies in this area, we have examined the dosing, treatment route, treatment duration and metabolism of 4-MU in both C57BL/6 and BALB/c mice. Mice fed chow containing 5% 4-MU, a dose calculated to deliver 250 mg/mouse/day, initially lose substantial weight but typically resume normal weight gain after 1 week. It also takes up to a week to see a reduction in serum HA in these animals, indicating that at least a 1-week loading period on the drug is required for most protocols. At steady state, more than 90% of the drug is present in plasma as the glucuronidated metabolite 4-methylumbelliferyl glucuronide (4-MUG), with the sulphated metabolite, 4-methylumbelliferyl sulphate (4-MUS) comprising most of the remainder. Chow containing 5% but not 0·65% 4-MU was effective at preventing disease in the experimental autoimmune encephalomyelitis (EAE) mouse model of multiple sclerosis, as well as in the DORmO mouse model of autoimmune diabetes. While oral 4-MU was effective at preventing EAE, daily intraperitoneal injections of 4-MU were not. Factors potentially affecting 4-MU uptake and plasma concentrations in mice include its taste, short half-life and low bioavailability. These studies provide a practical resource for implementing oral 4-MU treatment protocols in mice.
Subject(s)
Encephalomyelitis, Autoimmune, Experimental/drug therapy , Hyaluronic Acid/antagonists & inhibitors , Hyaluronic Acid/biosynthesis , Hymecromone/administration & dosage , Hymecromone/pharmacokinetics , Administration, Oral , Animals , Biological Availability , Disease Models, Animal , Encephalomyelitis, Autoimmune, Experimental/chemically induced , Encephalomyelitis, Autoimmune, Experimental/immunology , Encephalomyelitis, Autoimmune, Experimental/prevention & control , Half-Life , Hyaluronic Acid/blood , Hymecromone/blood , Hymecromone/metabolism , Mice , Mice, Inbred BALB C , Mice, Inbred C57BLABSTRACT
Concanavalin A (Con A) was dissociated into dimeric and monomeric subunits by incubation at 37 degrees C in acetate buffer of pH 3.8 containing 0.5% sodium dodecyl sulfate. The dimer was isolated in pure form by a density gradient ultracentrifugation method. Several properties of the dimer were determined including the formation of a precipitin with anti-Con A antibodies, the molecular weight, the lack of a binding site for glycogen, the lack of mitogenic activity for spleen lymphocytes, and the lack of inhibition by alpha-methyl D-glucoside. The latter findings differ from results reported by other investigators.
