Metabolic and pharmacological effects on nt-methylhistidine
turnover in skeletal muscle cells measured by gc-ms.
Thompson, Michael G., Robert M. Palmer, Amanda Thom, Karen Garden,
Gerald E. Lobley, and Graham Calder.
Rowett Research Institute, Bucksburn, Aberdeen, AB2 9SB U.K.,
Telephone Aberdeen (01224) 712751, Fax 01224 715349
APStracts 3:0098C, 1996.
A method employing gas chromatography-mass spectrometry has been
developed to measure Nt-methylhistidine (3-MH) synthesis and release
from skeletal muscle myotubes in vitro. It shows excellent linearity
(0.9999) over the range studied (0-4nmol), high recovery (92.6%) and
low coefficient of variation (1.6%). 3-MH release from myotubes was
essentially linear over a 96 h incubation whilst the loss of 3-MH
from cell protein accelerated with increasing time; an effect due, at
least in part, to decreasing rates of total protein synthesis. When
incubated in either glutamine-free or methionine-free medium for 48
h, 3-MH in cell protein and appearing in the medium were greatly
reduced when compared to the 48 h controls suggesting that
hypertrophy was greatly reduced. Similar but lesser trends were
observed with cAMP. In contrast, 12-0-tetradecanoylphorbol 13-acetate
(TPA), appeared to both stimulate 3-MH synthesis and inhibit its
release during a 48 h incubation. The development of this method
facilitates detailed investigation into the mechanisms through which
agents such as TPA regulate myofibrillar protein degradation.
Received 11 October 1995; accepted in final form 4 March 1996.
APS Manuscript Number C617-5.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 1 April 96