Does endotoxin-activated complement alter myocellular sodium homeostasis during sepsis?

Abstract

Inappropriate complement activation is closely related to tissue injury and organ dysfunction during systemic infection. It is not clear, however, if endotoxin-induced complement activation is responsible for changes in myocellular sodium homeostasis during sepsis. Rats underwent cecal ligation and puncture (CLP) or sham operation. Twenty-four hours after operation, fast-twitch extensor digitorum longus (EDL) muscles were isolated, incubated at 30 degrees C for 1 hour in Krebs-Henseleit buffer (KHB) (pH 7.4), and used to measure intracellular Na+ and K+ contents. Blood samples were collected to measure serum hemolytic complement activity and endotoxin levels. In addition, EDL muscles isolated from normal animals were incubated at 30 degrees C for 1 hour with zymosan-activated (10 mg/mL at 37 degrees C for 1 hour) rat sera, with lipopolysaccharide (LPS)-activated (LPS from Escherichia coli 055:B5, 10 or 200 microg/mL at 37 degrees C for 30 minutes) rat sera, with heat-inactivated (56 degrees C for 30 minutes) rat sera, with LPS (1 or 20 microg/mL), or in KHB. EDL muscles isolated from normal animals were also incubated with septic sera collected 6 or 24 hours after CLP with or without administration of soluble complement receptor type 1 (20 mg/kg, intraperitoneally). Myocellular Na+ and K+ contents ([Na+]i and [K+]i) were assayed using "washout" technique. Soluble C5b-9 complex levels in zymosan-activated or LPS-activated human sera were determined by enzyme-linked immunosorbent assay to evaluate the degree of complement activation induced by zymosan or LPS. Myocellular [Na+]i and [Na+]i/[K+]i ratios increased significantly 24 hours after CLP as compared with sham operation and were associated with decreased serum hemolytic complement activity and increased serum endotoxin levels. Zymosan-activated rat sera at sublytic concentrations markedly increased [Na+]i and [Na+]i/[K+]i ratios in isolated EDL muscles relative to heat-inactivated rat sera. LPS-activated rat sera, however, did not alter these two indices. In addition, myocellular [Na+]i and [Na+]i/[K+]i ratios were equivalent among normal EDL muscles incubated with septic sera, soluble complement receptor type 1-treated septic sera, or KHB. These results collectively suggest that polymicrobial sepsis, as produced by CLP, alters sodium homeostasis in fast-twitch skeletal muscles in association with changes in systemic complement activation and circulating endotoxin levels. Although endotoxin can activate the complement cascade, endotoxin-induced complement activation does not appear to be responsible for changes in myocellular sodium homeostasis observed during sepsis in rats.