Partial purification from mammalian peripheral nerve of a trophic factor that ameliorates atrophy of denervated...

Atrophy in a denervated muscle results from the disuse caused by paralysis of the muscle, and from the loss of special nerve-derived trophic substances. Crude preparations of protein from rat or sheep sciatic nerves have been shown to prevent the nondisuse atrophy of the rat's extensor digitorum longus muscle when injected into the denervated muscle daily for 1 week. Aqueous extracts of sheep sciatic nerves were fractionated by gel-liquid chromatography. After each step of purification, the trophic activities of the various fractions were assayed in the rat. Cross-sectional areas of type IIB muscle fibers in the denervated extensor digitorum longus were measured to determine which injected fraction contained the active principle. Affinity chromatography on concanavalin A-agarose revealed that the trophic substance was a glycoprotein. Further fractionation by gel filtration indicated that the active substance had a molecular weight in the range of 90,000 to 130,000. Ion-exchange chromatography on DEAE-cellulose yielded an active fraction containing substances with isoelectric points between 7.0 and 7.2, determined by polyacrylamide gel isoelectric focusing. This active fraction was resolved into 15 bands on sodium dodecyl sulfate-gel electrophoresis. Two bands had apparent molecular weights of 91,300 and 127,400. The active factor was shown thus to be a glycoprotein, molecular weight approximately 100,000, isoelectric point approximately 7.0. It may be one of two protein bands that are similar to it in molecular weight.