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Document Type

Thesis - University Access Only

Award Date

1989

Degree Name

Master of Science (MS)

Department / School

Animal Science

First Advisor

Douglas C. McFarland

Abstract

Satellite cells, isolated from the superficial pectoralis muscle of growing Nicholas tom turkeys, were cloned to obtain a pure population of myogenic cells. These cultures proliferated rapidly and differentiated (fused) into myotubes typically containing 92-98% fused nuclei. Competitive binding assays were performed on near-confluent (approximately 700 nuclei/mm2) satellite cell or myotube cultures by adding 200,000 dis/min [1251] IGF-1 (sp act 250-300 Ci/g) to each of 4 replicate 35 mm diameter wells, along with increasing concentrations of unlabeled IGF-1, IGF-11, or insulin. Following incubation, the cultures were washed to remove the unbound hormones, solubilized with 0.5 N NaOH, and the radioactivity specifically bound was determined. Total and fused nuclei number as well as total protein and DNA were determined in parallel cultures. Our results indicate that turkey satellite cell and myotube cultures possess specific binding sites for IGF-1. Displacement of [1251] IGF-1 was in the order of IGF-1 > IGF-11 > insulin. Although the [1251] IGF-1 association constants were similar for turkey satellite cells and myotubes, a 2.8-fold decrease in the number of receptors on a per nuclei basis was observed as satellite cells differentiated (fused) to form multinucleated myotubes. The 50% inhibition constants for IGF-1, IGF-11, and insulin were 3.7 x 10 -9 M, 7.5 x 10.a M, and 8.7 x 10.s M for satellite cells and 3.1 x 10 -9 M, 7.5 x 10 -8 M, and 9.6 x 10. -8 M for myotubes, respectively. Receptor crosslinking analysis was performed on near-confluent satellite cell cultures- incubated with 4.2 x 10 -11 M [1251] IGF-1 in the presence or absence of 1 x 10 -7 M IGF-1, IGF-11, or insulin and cross-linked with disuccinimidyl suberate. Receptor subunit species of Mr 130 KDa (alpha) and Mr 98 KDa (beta) were observed with electrophoresis under reducing conditions (100 mM dithiothreitol) and at 300 KDa (native receptor complex) under non-reduced conditions. Autoradiographic bands were displaced with IGF-1 but not with I equimolar IGF-11 or insulin, and banding patterns typical of the type II receptor (Mr 220 KDa native and Mr 260 KDa reduced) were not observed. Therefore, it appears that turkey satellite cells possess a type I IGF receptor.

Library of Congress Subject Headings

Satellite cells

Muscle cells

Turkeys -- Growth

Growth factors

Format

application/pdf

Number of Pages

98

Publisher

South Dakota State University

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