Mouse genotype affects inducible expression of cytokine genes.

The levels of circulating IFN in mice infected with Newcastle disease virus (NDV) are regulated by the If-1 locus. In this study we show that in NDV-infected C57BL/6 mice, which carry the If-1h allele and produce high levels of IFN, high levels of both IFN-alpha and -beta mRNA can be detected in the spleen. In contrast, only very low levels of IFN mRNA could be detected in spleens of infected BALB/c mice containing the If-1l allele and producing low levels of IFN or in B6.C-H28c mice that are congenic for the If-1l allele. The relative levels of all individual IFN-alpha 1, alpha 4, and alpha 6 mRNA in spleens of infected BALB/c were lower than in spleens of infected C57BL/6 mice, indicating that the If-1 locus affects the expression of all IFN-alpha subtypes and is not associated with the deletion or inactivation of a specific IFN gene. The relative levels of IFN regulatory factor-1 mRNA in infected mice carrying the If-1l and If-1h loci were comparable, suggesting that the If-1 regulation is not associated with the altered expression of the IFN regulatory factor-1 gene. Quantitative difference in the expression of IFN-alpha and -beta genes was also observed in in vitro-infected peritoneal macrophages isolated from either C57BL/6 or BALB/c mice. A surprise finding was that the If-1 locus also affected the NDV-induced expression of two other cytokine genes, TNF-alpha and IL-6. Priming of the macrophage cultures with murine IFN enhanced the expression of all cytokine genes, and the relative levels of IFN, TNF-alpha, and IL-6 mRNA induced by NDV in macrophages derived from C57BL/6 and BALB/c mice were comparable. We propose that the If-1 locus affects the early stages of a signal transduction pathway which are common to the virus-mediated induction of IFN, TNF-alpha, and IL-6 genes.     

Endogenous interferon specifically regulates Newcastle disease virus-induced cytokine gene expression in mouse macrophages.

In macrophages from inbred mice, the magnitude of the interferon (IFN) response to Newcastle disease virus (NDV) infection is under genetic control of the If-1 locus, which carries the allele for either high (h) or low (l) IFN production. Here, we report that the activity of genes within the If-1 locus is influenced by macrophage-derived endogenous IFN. In addition to various other biological effects, we observed that endogenous IFN specifically downregulated NDV-induced IFN and interleukin 6 production. Preculture of bone marrow-derived macrophages (BMM) from BALB/c (If-1l) mice in macrophage colony-stimulating factor plus anti-IFN-beta provoked a 30- to 50-fold increase in NDV-induced cytokine production compared with induced control cultures in macrophage colony-stimulating factor alone, whereas only a 4- to 6-fold increase was observed in anti-IFN-beta-treated BMM from C57BL/6 (If-1h) mice. This resulted in nearly complete abrogation of the genetically determined difference in the response to NDV. The increase was specific for NDV and was marked by strong additional activation of IFN-alpha genes. Studies using BMM from B6.C-H28c If-1l congenic mice gave results identical to those obtained with BALB/c BMM. Addition of 20 IU of recombinant IFN-alpha 4 to anti IFN-beta-treated macrophages from B6.C-H28c mice 20 h prior to NDV infection strongly downregulated the IFN-alpha, IFN-beta, and interleukin 6 responses. The genetic difference between macrophages from If-1h and If-1l mice was thus reestablished, since the same treatment caused only weak reduction of NDV-induced cytokine gene expression in BMM from C57BL/6 mice. These data suggest that the If-1h and If-1l alleles harbor IFN-inducible genes that, following activation, specifically suppress subsequent cytokine gene expression in response to NDV.     

Effect of mouse genotype on interferon production. I. Lines congenic at theIf-1 locus

The level of circulating Interferon induced in mice by Newcastle disease virus is controlled by a single codominant locus,If-1, with two alleles,If-1 ^l for low andIf-1 ^h for high production. This locus is linked to the histocompatibility locusH-28. Of three C57BL/6By lines congenic for the BALB/cBy allele atH-28, two are carrying the BALB/cBy allele and one, the C57BL/6By allele atIf-1. Thus, mouse strains that are genetically very similar but different in their production of NDV-induced circulating Interferon now are available.A preliminary report of these studies was presented at the ASM meeting at Miami Beach in May 1973.     

The mouse Rn locus: S allele of the renin regulator gene results from a single structural gene duplication.

Inbred strains of mice have been divided into two distinct phenotypic groups having different levels of renin activity regulated by androgen in the submaxillary gland (SMG). Strains carrying the Rnrs allele of the renin gene regulator, located on chromosome 1, have a high level of renin activity; strains carrying the Rnrb allele have a low level of renin activity. The level of SMG renin activity correlates with the level of renin mRNA. We have analyzed, by Southern blot hybridization, the organization of renin genes in both strains. Strains carrying the Rnrb allele, such as BALB/c or C57 Bl/6, or CH3 mice, have one renin structural gene per haploid genome, while those having the Rnrs allele, such as AKR or Swiss mice, have two renin genes. We have also identified renin genes in mice belonging to different biochemical groups: Mus spretus has one renin gene while M. vrania and M. musculus brevirostris have two renin genes.     

Genetic aspects of the immunomodulating action of interferon

The data of the study of alpha/beta interferon (IFN) effect in mice of different genotype were presented. CBA mice of H-2k genotype, C57B1/6 mice of H-2b genotype and their hybrid (CBA X C57B1/6) F1 have been used in the experiments. IFN has been injected intraperitoneally in a dose of 100-5000 U/mouse in combination with antigenic stimulation. It was shown that IFN enhanced stem cells migration from bone marrow in CBA, but not in (CBA X C57B1/6)F1 mice. At the same time the splenocytes from CBA mice were more sensitive to inhibition by IFN than splenocytes from C57B1/6 mice. This was found in antibody and immune rosette-formation tests. The effect of IFN on the immune system cells is probably predetermined by the individual genetic characteristics of a mouse strain.     

Induction of natural killer cell activity of thoracic duct lymphocytes by polyinosinic-polycytidylic acid (poly(I:C)) or interferon

Natural killer (NK) cell activity of thoracic duct lymphocytes (TDL) was examined in normal mice and in mice treated with polyinosinic-polycytidylic acid (poly(I:C) and interferon (IFN). TDL from mice treated with phosphate-buffered saline (PBS) expressed little or no NK cell activity against YAC-1 target cells at effector-to-target ratios of up to 200:1, even after in vitro treatment with murine L-cell IFN. In contrast, TDL from poly(I:C)- or IFN-treated mice expressed significant NK activity, which correlated with the significantly higher NK activity of splenocytes from these mice compared to the NK activity of splenocytes from PBS-treated mice. These data indicate that although TDL from normal mice express no detectable NK cell activity, NK cell activity can be induced in TDL by in vivo treatment with poly(I:C) or IFN.     

Genetic control of the natural killer cell activity in SJL and other strains of mice

Murine natural killer (NK) cell activity against lymphoma targets can be classified into three major functional phenotypes, i.e., low, inducible, and high, according to the levels of endogeneous activity and the extent of augmentation by interferon (IFN) or IFN inducers, as previously described. The prototype strains identifying these three phenotypes are SJL, A.SW, and B10.S, respectively, all bearing the H-2s haplotype. In the present study, the genetic basis of the low phenotype of SJL mice was examined further. F1 hybrid offspring of crosses between SJL and a strain with the high NK phenotype (B10.S, B10.D2, B10, C3H/HeN, or D1.LP) invariably expressed the high NK phenotype, indicating recessiveness of the low phenotype. Crosses between SJL and another low NK strain, such as A/J, A/HeN, or I/St, resulted in offspring of either the inducible or the high NK phenotype. Such genetic complementation between the low NK pairs indicates that the low phenotype of SJL and that of the other strains have different genetic bases. F1 hybrid mice between SJL and an inducible strain, A/WySn, were inducible, but those between SJL and the second inducible strain, A.SW, had the high NK phenotype. Thus, the congenic A/WySn and A.SW have distinct genotypes resulting in the same inducible phenotype. According to analyses of the segregating offspring from backcrosses of (SJL X B10.S)F1 mice to SJL, a single gene difference is responsible for the low endogenous level of NK activity in SJL as opposed to the high endogenous level in B10.S, and that the difference in three genes accounts for the poor responsiveness of NK cells to IFN in SJL mice. Studies of the two congenic lines of SJL, i.e., SJL-Igha and SJL-nu, indicated that the Igh locus is irrelevant for the low NK phenotype of SJL, but the nu locus clearly is relevant; SJL mice homozygous for the nu allele were phenotypically inducible in contrast to the nu/+ or +/+ mice which are low. The nu gene homozygosity rendered SJL mice responsive to IFN, not only in NK activity against lymphomas but also in ADCC activity against antibody-coated lymphoma cells.     

Mechanisms of depression of splenic natural killer cell function in C57BL/6 mice infected with Leishmania donovani

C57BL/6 mice chronically infected with the protozoan parasite Leishmania donovani exhibit profoundly depressed splenic natural killer (NK) cell activity as measured by in vitro cytolysis of lymphoma target cells. Injection of infected mice with an interferon (IFN) inducer or in vitro treatment of infected splenocytes with IFN, a phorbol ester, or indomethacin failed to restore their NK activity to the degree shown by age-matched, uninfected mice. Fractionation of infected splenocytes by nylon wool, Sephadex G-10, or carbonyl iron and magnetism treatments was also unable to effect an increase in NK activity. Addition of infected splenocytes to uninfected ones in in vitro NK assays suppressed the NK activity of the latter, and the suppression could be partially or wholly abrogated by prior fractionation of infected splenocytes by the methods noted above. In vitro treatment of infected splenocytes with concanavalin A revealed the presence of NK activity in these cell populations. The results indicate that splenocytes in L. donovani-infected mice become insensitive to IFN stimulation; and the impairment of another, possibly IFN-independent pathway of NK-cell activation may also contribute to the observed L. donovani-induced depression in splenic NK activity in C57BL/6 mice.     

Expression of interferon-induced genes in different tissues of mice.

In vivo responses to interferon (IFN) in mice were determined by measuring the steady-state levels of induced mRNAs following injection of IFN and poly(I)-poly(C). With cDNA probes for mouse 2'-5' oligoadenylate synthetase (2-5A synthetase) and 1-8, constitutive expression of the corresponding mRNA was detectable in different organs of normal C3H/He mice. These mRNA levels were increased by as much as 15-fold over control levels in various tissues, including the brain, after IFN and poly(I)-poly(C) treatment, coincident with increases in 2-5A synthetase enzyme activity. The basal activity level of this enzyme could be reduced in normal mice by treatment with anti-mouse IFN (alpha + beta) antibody. This treatment also reduced the levels of 2-5A synthetase and 1-8 mRNAs. Thus, physiological levels of circulating IFN maintain elevated levels of IFN-induced mRNAs in mice. Furthermore, changes in 2-5A synthetase enzyme activity reflect the changes in gene expression in vivo.