Levels of 2',5'-oligoadenylates in the tissues of pathogen-free mice

2',5'-Oligoadenylate (2-5A) has been reported to exist in a variety of cells and tissues. Since it has been suggested that these compounds play a role in normal cell metabolism as well as in the interferon response, we sought to determine the levels of 2-5A in healthy mouse tissue. Methodology developed to recover 2-5A from tissue included TCA extraction, fractionation on silica and and C18 cartridges and HPLC. The 2-5A content in HPLC fractions was determined by an enzyme-linked immunoassay. This protocol gave high recovery of 2-5A and allowed detection of less than 10 fmole per fraction. Pathogen-free NIH Swiss mice were injected intravenously with saline, poly(I).poly(C) or encephalomyocarditis virus (EMCV) and examined for 2-5A content, 2-5A synthetase levels and circulating interferon. Levels of 2-5A were determined to be 400-800 fmole/g in spleen, liver, kidney and brain of the saline-treated mice. Mice that were treated with poly(I).poly(C) and EMCV showed substantially higher amounts of 2-5A (3-16 pmole/g). Cores represented up to 98% of the total 2-5A recovered from poly(I).poly(C)-injected mice. The levels of synthetase also markedly increased in mice infected with EMCV or injected with poly(I).poly(C). However, the levels of 2-5A did not correlate directly with the increase in enzyme levels. Tissues having similar amounts of synthetase had substantially different levels of 2-5A, implying that factors other than enzyme content influence the intracellular concentration of 2-5A.