Synthesis and secretion of a high molecular weight form of nerve growth factor by skeletal muscle cells in culture.

Abstract

Rat skeletal muscle cells and a cloned myogenic cell line synthesize and secrete in culture a molecule that is immunologically and biologically indistinguishable from the active form of nerve growth factor (NGF) from mouse submandibular gland. This protein can be detected in medium conditioned by muscle cells both before and after fusion and in the soluble fraction of muscle cell homogenates. Chromatographic data also reveal that the molecular properties of muscle cel NGF differ from those of the growth factor purified from mouse submandibular glands. Musce cell NGF has a molecular weight between 140,000 and 160,000, whereas purified mouse gland NGF has a molecular weight of G-200 and G-75, from Pharmacia; D-arabinofuranosylcytosine (Ara C) and cycloheximide, from Calbiochem; bovine serum albumin (3X crystallized), trypsin, and horse heart ferricytochrome c, from Sigma; 3H20 and NaI25I from New England Nuclear; and human IgG from Pentex. Rat tail collagen was prepared by the procedures of Ehrmann and Gey (7). Calf skin collagen without preservative was purchased from Worthington. NGF was isolated and purified from male mouse submandibular glands by modifications (1) of the method of Bocchini and Angeletti (8). All preparations were electrophoretically homogeneous as previously described (1).Antisera to NGF were prepared in rabbits (1). IgG was purified by diluting serum with an equal volume of 0.15 M NaCl in 10 mM sodium phosphate, pH 7.4. This solution was brought to 50% saturation with ammonium sulfate, and the precipitate was redissolved in and dialyzed against 10 mM sodium phosphate/15 mM NaCl at pH 7.5. The solution was applied to a column of Abbreviations: NGF, nerve growth factor; Ara C, D-arabinofuranosylcytosine.

Tissue engineering the mechanosensory circuit of the stretch reflex arc: Sensory neuron innervation of intrafusal muscle fibers

“…The basic serum-free medium composition developed in our laboratory for the co-culture of embryonic motoneurons and myocytes was used as a starting point for the development of a medium that enhanced synaptic connectivity between neurons and myotubes in vitro [54-56]. The addition of 10 new growth factors known to play a role in neuromuscular junction formation and synaptic maintenance led to increased recovery and development of neurons and myocytes as well as increased development of synaptic structures in the system [57] (Table 2).…”

Tissue engineering intrafusal fibers: Dose- and time-dependent differentiation of nuclear bag fibers in a defined in vitro system using neuregulin 1-β-1

“…The addition of nerve growth factor (NGF) to the formulation came as a result of experiments done by Murphy et al showing an active form of NGF was secreted by rat skeletal muscle cells and a myogenic cell line [42]. They postulated the molecule acted in a support role for associated nerves.…”

“…Using media supplemented with NGF, myotube formation progressed at the usual rate, but the longevity of the culture increased. NGF was added to the medium to assist in myocyte recovery and to more accurately represent the plausible environment that exists inside a muscle spindle in vivo due to secretion of the factor by schwann cells and myocytes and to also serve as a starting point for future sensory neuron / muscle co-culture experiments [42,43] (Table 1).…”

Myoblasts and myoblast‐conditioned medium attract the earliest spinal neurites from frog embryos.

“…NGF is released from new-born rat skeletal muscle (Murphy,Singer,Saide,50 Pantazis, Blanchard, Byron, Arnason & Young, 1977 (Letourneau, 1978), did not attract the earliest spinal neurites from Xenopus. NGF has no effect on embryonic motoneurones in the rat (Smith & Appel, 1983).…”