Inducers of interferon and host resistance, V. In vitro studies.

Abstract

Previous reports'-6 from this laboratory recorded that double-stranded ribonucleic acid molecules from numerous sources are efficient inducers of interferon and of resistance to viral infection in vivo and in vitro. The present report describes the induction of resistance against several viruses in a variety of cell cultures by the complex of polyriboinosinic and polyribocytidylic acids (rI:: rC) and presents evidence to associate such resistance with interferon induction. The inactivity of the individual homopolymers, rI and rC, is reaffirmed, and interferoln induction by rI: rC in vitro is shown to be inhibited by exposure of cells to actinomycin D. Materials and Alethods.-(l) Polyriboinosinic acid (rI) and polyribocytidylic acid (rO) were purchased from Miles Laboratories, Elkhart, Indiana. The rI: rC complex was prepared by mixing rI and rC in equimolar coD.eentration in phosphate-buffered salin.e (0.006 M sodium phosphate 0.15 11l NaCl pH 7.0). (2) Cell cultures: The various primary, cell strain, and line cell cultures listed in Table 1 were prepared and maintained by ordinary procedures. T'he RK13 culture is a stable line of rabbit kidnley cells, and the WI-38 and HFL cultures are diploid cell strains of hu:man embryonic lung. The RK13 an-d WI-38 cultures are well documented and the HFL cell strain was developed by Dr. C. Baugh in these laboratories. Rabbit spleen cell suspensions were prepared according to Field et al.2 (3) The viruses used were prepared from the seed stocks of this laboratory. The rhinovirus serotypes were designated according to the number system of Hamparian et al.' (4) Induction of resistance to viral illfection was measured after overnight treatment of cell cultures with rI :rC. Following removal of inducer, interference with virus replication was measured by the plaque-reduction assay.' (5) Interferon induction in primary rabbit kidney cells or in rabbit spleen cell suspensions by rI: rC was assayed by t:ransfer of serial dilutions of the cell supernatant fluids to RK13 cultures in Falcon. flasks. 'rhis was followed by overnight incubation prior to removal and challenge with. vesicular stomatitis virus (VSV) with observation for reduction in plaque formation. RKI.3 cells were used for assay since they were relatively insensitive to iniduction of resistance to VSV by rI: rC, requiring more than 1 ,ug/ml. Hence, these cells were unaffected by the small residual amount of rI:rC in the interferon samples. The interferon titer was the highest dilution of sample which caused at least 50% reduction in plaque formationL. (6) Other pertinent methods are described in the text.