Measuring whole-body actin/myosin protein breakdown in mice using a primed constant stable isotope-infusion protocol.
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abstract
To measure actin/myosin protein breakdown, the 24 h excretion of N (tau)-methylhistidine (3MH) is used. However, in mice, this method is invalid. Therefore we have developed a liquid chromatography-MS technique to measure the tracer/tracee ratio and concentration of 3MH in plasma, enabling an in vivo primed constant infusion protocol with a deuterated stable isotope of 3MH. We tested this model by giving a primed constant infusion of L-[3-methyl-(2)H(3)]histidine, L-[phenyl-(2)H(5)]phenylalanine and L-[phenyl-(2)H(2)]tyrosine to three anaesthetized experimental groups: mice receiving saline intraperitoneally (i.p.) (CON), mice receiving saline i.p. and starved for 9 h (STA), and mice receiving lipopolysaccharide i.p. and starved for 9 h (STA+LPS). The contribution of myofibrillar to total protein breakdown was significantly lower in the STA group than the CON group (30+/-4% and 54+/-14% respectively; P <0.05), and was significantly higher in the STA+LPS group than the STA group (52+/-7% and 30+/-4% respectively; P <0.05). Whole-body myofibrillar protein breakdown, total protein breakdown, protein synthesis and net protein breakdown were not different between the groups. We conclude that this in vivo primed constant stable isotope-infusion protocol can give valuable information about the role of actin/myosin protein breakdown in mice.
