Native heparin from rat peritoneal mast cells

Abstract

[35S]Heparin was produced in vitro by incubation of rat peritoneal mast cells with [35S]sulfate and in vivo by injection of [35S]sulfate into rats. The [35S]heparin together with nonlabeled heparin in the mast cells was isolated in native form by mild methods that avoided the use of proteolytic enzymes or high alkali concentrations. The heparin had low anticoagulant activity. Incubations of mast cells with [35S]sulfate for less than several hours in vitro resulted in [35S]heparin of approximately Mr=200,000 to 400,000 based on gel filtration, while longer incubations yielded [35S]heparin of approximately Mr=750,000 that was similar to the nonlabeled heparin in the mast cells. When [3H]serine was included in the in vitro incubations, 3H-labeled material was found to co-chromatograph with the [35S]heparin. None of the heparin could be degraded by any of several proteolytic enzymes, but incubation for 14 h at 25 degrees with 0.5m NaOH degraded all samples to a size of approximately Mr=40,000. One-third of the [3H]serine label continued to co-chromatograph with the [35S]heparin after alkali treatment, while the remaining two-thirds appeared as smaller molecules completely separated from the [35S]heparin. Thus, native heparin of the mast cell may be an unusual proteoglycan that is resistant to proteolytic enzymes.