Page 185 - 73_04

VOL. 73 (4), 987-1008, 2007 CONTRIBUTION OF TNF-a TO OBESITY...

main candidates. In this regard, TNF-a blocks skeletal muscle
differentiation causes sarcopenia and produces insulin resistance in
skeletal muscle of healthy humans and in primary cultures of mouse
skeletal muscle (23). Although ceramide and FFA have been reported
to produce insulin resistance in skeletal muscle, a direct effect of
TNF-a in this tissue has been the matter of controversia. Several
reports did not detect TNF-a inhibitory action on insulin-induced
glucose uptake, although TNF-a per se highly increased basal glucose
uptake. However, others observed an inhibitory effect on insulin
action without modifying basal glucose uptake in muscle cells.
Moreover, in most of these studies insulin stimulation of glucose
uptake was very poor because virtually all cultured skeletal muscle
cell lines, including L6 and C2C12 myotubes, have been found to be
deficient in GLUT4 expression.

Consequently, in our laboratory we developed primary cultures
of neonatal rat skeletal muscle that represented a suitable system for
investigating the molecular basis of TNF-a-induced insulin
resistance. When these cells were differentiated in low serum until
the formation of myotubes and maintained in low glucose medium
to mimick the physiological environment, responded to acute insulin
stimulation by increasing glucose uptake and GLUT4 translocation
to plasma membrane (24). Chronic exposure to TNF-a impaired both
insulin-stimulated glucose uptake and GLUT4 translocation, without
affecting the content of GLUT4 protein or the state of differentiation
of the myotubes (24), in agreement with the effect produced in
muscle in vivo. The molecular mechanism underlying TNF-a-
mediated insulin resistance could involve activation of stress kinases
and proinflamatory pathways, as was observed in neonatal myotubes
(24). Acute insulin stimulation also produced a transient phos-
phorylation of p38MAPK (3), but insulin activated the isoform a
meanwhile TNF-a activates the b. When chemical inhibitors were
used to evaluate the contribution of sustained activation of stress
kinases by TNF-a to insulin resistance, only the inhibition of
p38MAPK completely restores insulin-stimulated glucose uptake and
insulin signaling (24). In this regard, adenovirus-mediated
transfections of constitutively active MKK6/3 mutants in L6
myotubes have been reported to diminish glucose transport induced
by insulin via down-regulation of GLUT4 gene expression (25).

995