Blocking of interferon synthesis in murine macrophages by pretreatment with interferon

The influence of pretreatment with interferon (IFN) on subsequent IFN synthesis was investigated in macrophage cultures of DBA/2 and C57BL/6 mice. The doses of IFN alpha/beta for pretreatment ranged from 10,000 U/ml to 100 U/ml and the incubation time was between 18 and 2 h. No blocking effect was observed for chemical induction with poly I:poly C or CMA. However, for viral infection with NDV, blocking was observed. This inhibition of IFN synthesis was dependent on the dose and time of IFN pretreatment and of the titer of the inducing virus. Similarly in mouse fibroblast cultures no blocking activity was observed for induction with poly I:poly C/DEAE-dextran. Again, with NDV as inducer, pretreatment with IFN resulted in inhibition of interferon synthesis. Thus, our data show that blocking occurs only with a viral inducer and suggest that it is caused by an antiviral effect.     

Modulation of natural killer cell activity in mice after interferon induction: depression of activity and depression of in vitro enhancement by interferon.

Splenic natural killer (NK) cell activity of BALB/c and C3H mice was assayed after administration of the interferon inducers Escherichia coli endotoxin or Newcastle disease virus (NDV). As expected, the NK cell activity rose early in response to the interferon inducers. At 1 to 3 days after an injection of endotoxin, NK activity was hyperesponsive to interferon stimulation. At 5 to 9 days after injection of either endotoxin or NDV, splenic NK activity was depressed, and the spleen cells showed a relative refractoriness to in vitro interferon stimulation. It is postulated that this phenomenon may be related to hyporeactivity, the inability to reinduce interferon after an initial period of interferon production.     

Regulation by endogenous interferon of 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 locus If-1, with C57BL/6 carrying the 'high-producer' allele If-1h whereas BALB/c have the 'low-producer' allele If-1l. The IFN produced consists of 90% IFN-beta and there are 10-fold differences between macrophages from If-1h and If-1l mice. Recently, we observed that interleukin-6 (IL-6) is coinduced by NDV in macrophages and seems to be under the same genetic control. Noninduced macrophages have been shown to secrete low amounts of antiviral activity endogenously when cultured in the presence of the macrophage-colony-stimulating factor (M-CSF). Here, we report that the amount of this endogenous IFN varies between macrophages from different mouse strains. Macrophages from BALB/c were found to secrete 5-10 times more endogenous IFN compared to C57BL/6. The antiviral activity could be identified as IFN-beta. Interestingly, we observed that endogenous IFN specifically down-regulates NDV-induced IFN and IL-6 production. Preculture of BALB/c macrophages in M-CSF plus anti-IFN-beta to neutralize the biological effects of the endogenous IFN provoked a 30- to 50-fold increase in NDV-induced cytokine production, resulting in a nearly complete abrogation of the genetically determined difference since the same treatment only caused a 6-fold increase in C57BL/6 macrophages following NDV infection. This increase in cytokine gene expression was specific for NDV and marked by a strong additional activation of IFN-alpha genes. Addition of mouse recombinant IFN-alpha 4 to anti-IFN-beta-treated macrophages for 18 h prior to NDV infection down-regulated again IFN gene expression and reestablished the genetic differences between macrophages from If-1h and If-1l mice.(ABSTRACT TRUNCATED AT 250 WORDS)     

Downregulation of Newcastle disease virus (NDV)-dependent IFN-alpha/beta production in macrophages by IFN-induced gene products of the locus If-1.

We have found that in cultured mouse bone-marrow-derived macrophages (BMDM), endogenous IFN-beta specifically regulates Newcastle disease virus (NDV) induced interferon (IFN)-alpha/beta synthesis, possibly by influencing the activity of genes within the regulatory locus If-1. Comparison of anti-IFN-beta-treated BMDM from C57BL/6 mice and the congenic line B.6C-H28c carrying the high (h) or low (l) producer allele of If-1, respectively, revealed a much stronger response of the If-1l allele to exogenous IFN-alpha treatment. Twenty IU rIFN-alpha 4 were sufficient to induce nearly complete suppression of NDV-induced IFN-alpha and IFN-beta production in BMDM from B6.C-H28c mice, but had no effect on the IFN-alpha/beta response induced by Sendai virus, another member of the paramyxovirus group. Simultaneous treatment of BMDM with cycloheximide inhibited the suppressive effect of rIFN-alpha 4, indicating that IFN induced the expression of one or several new proteins encoded by gene(s) within the If-1l locus which are responsible for the NDV-specific downregulation of IFN-alpha/beta production. A time course analysis indicated that the suppressive activity of IFN-induced If-1l gene products took 12 h to develop. It was preceded by an opposite priming effect, leading to enhancement of the early IFN-alpha/beta response to NDV measured 5 h after infection. This priming effect in BMDM was, however, only visible during an 8-h period of IFN-alpha treatment, whereas in the continued presence of IFN for 12 h or longer, priming was superimposed by the inhibitory action of the If-1l gene products.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of Corynebacterium acnes on interferon production in mouse peritoneal exudate cells.

Corynebacterium acnes, an organism closely related to C. parvum, has been recognized to have a striking effect on the reticuloendothelial system, as well as on both humoral and cellular immunity. In mice previously exposed to C. acnes, serum interferon levels induced by injection of Newcastle disease virus (NDV), Chikungunya virus (CV), and polyinosinic-polycytidylic acid are suppressed. When peritoneal macrophages and lymphocytes from animals exposed to C. acnes were cultivated in vitro, their capacity to produce interferon in response to NDV and CV was reduced. Furthermore, the interferon-producing capacity of these cells in tissue culture was inhibited after exposure to C. acnes to vitro. Exposure of separated populations of peritoneal macrophages and lymphocytes to C. acnes in vitro demonstrated that the interferon response to NDV by both cell types is inhibited. Peritoneal macrophages appear to be the major contributor to the interferon response in this system. Finally, this inhibitory effect was shown to occur after exposure to a purified cell wall preparation of C. acnes organisms, as well as a lipid extract of this preparation.     

Effect of thymus extract and of late thymectomy on interferon production

Mice aged 10 days do not produce interferon in response to injection of Newcastle disease virus (NDV). A single injection of a fraction of calf thymus extract, obtained by the method of Gyulling et al., into newborn mice conferred the ability to produce interferon on mice at the age of 10 days. In response to injection of NDV, adult rats produce interferon in high titers. The intensity of interferon synthesis was unchanged 2–3 months after thymectomy.Laboratory of Physiology and Pathophysiology and Laboratory of Virology, Kiev Research Institute of Otolaryngology. (Presented by Academician of the Academy of Medical Sciences of the USSR N. N. Gorev.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 84, No. 10, pp. 439–440, October, 1977.     

Increased Toxicity of Double-Stranded Ribonucleic Acid in Virus-Infected Animals

Virus-infected mice were significantly more susceptible to the toxic effects of double-stranded ribonucleic acid (RNA) than uninfected mice. A dramatic increase in mortality was observed after injection of either synthetic (polyriboinosinic·polyribocytidylic acid) or natural (mycophage) double-stranded RNA in mice infected with Newcastle disease virus (NDV) or vesicular stomatitis virus (VSV). With the exception of endotoxin, interferon inducers other than double-stranded RNA, such as tilorone-hydrochloride and chlorite-oxidized oxyamylose, did not show this increased toxicity in virus-infected animals. The increased susceptibility of virus-infected animals to the toxic effects of double-stranded RNA appears to be related to the levels of interferon induced by the virus infection, either systemically, in the blood stream (after inoculation of NDV), or locally, in the brain (after infection with VSV).     

Interferon responses of various populations of lymphocytes of inbred mice with various degrees of sensitivity to the western equine encephalitis virus

The sensitivity of mice of three lines: CBA, BALB/C, and C57BL/6 (males aged greater than or equal to 4-5 and one week) to peripheral inoculation with Western equine encephalomyelitis (WEE) virus was studied. The C57BL/6 line was found to be the most and CBA the least sensitive. The first generation hybrids between these lines are intermediate in their sensitivity to WEE virus. A correlation was established between the increased sensitivity to WEE virus of C57BL/6 mice and the defect of the interferon system in these mice manifested by the extremely low capacity of their thymocytes for production of beta and alpha-interferons, as well as by a significant decrease of interferon responses of splenocytes to Newcastle disease virus, L-1 (alpha-interferon), lafarin, beta-interferon, concanaxalin-A (alpha-interferon) after immunization with antigens of different origin.     

Differential effect of poly rI.rC and Newcastle disease virus on the expression of interferon and cellular genes in mouse cells

The expression of type I murine interferon (MuIFN) genes and several other cellular genes was examined in poly rI.rC induced and Newcastle disease virus (NDV) infected mouse cells. Northern analysis of RNA from induced L cells revealed that the MuIFN-alpha s are expressed efficiently in NDV infected cells but only at low levels in poly rI.rC induced cells. MuIFN-beta 1, however, is expressed equally well in cells treated with poly rI.rC or infected with NDV. As shown by the use of a probe specific for poly rI.rC, interferon induction correlates with the cellular uptake of poly rI.rC into the cells. The relative levels of alpha and beta 1 mRNAs in the cells reached a maximum at 10 hr after the induction which indicates coordinate expression of alpha and beta 1 interferon genes. The effect of viral infection on the expression of two murine genes coinduced with interferon (pMIF20/11 and pMIF3/10) and several cellular genes was also examined. While pMIF20/11 is an inducible gene, the pMIF3/10 gene is expressed constitutively in mouse L cells. Viral infection, but not poly rI.rC treatment, enhanced the expression of the pMIF3/10 gene, as well as two other cellular genes; H-2 and c-myc, however, the expression of beta-actin gene was unaltered. These data indicate that enhancement of gene expression in virus infected cells in not limited to the interferon system.     

Mx genes show weaker primary response to virus than other interferon-regulated genes.

Some interferon (IFN)-regulated genes are induced as a primary response to virus as well as secondarily through virus-induced IFN. Here we investigated whether this dual control mechanism would also regulate the activity of the human and mouse Mx genes that encode proteins with intrinsic antiviral potentials. To distinguish between a primary response to virus and a secondary response to virus-induced IFN, we studied virus-induced Mx gene expression in cell lines that lack a functional IFN system and in cells with blocked protein synthesis. In contrast to the two IFN-regulated human genes ISG56 and ISG15, the human MxA gene showed almost no primary response to Newcastle disease virus (NDV) or influenza virus. Similarly, direct activation of the mouse Mx1 gene by NDV or influenza virus was not significant in mouse embryo cells or explanted peritoneal macrophages. A moderate primary Mx1 response to NDV was observed in the permanent cell line L1210. Lack of a strong IFN-independent Mx response to virus indicates that this mode of gene regulation does not play a significant role in Mx-mediated resistance to viral disease.     

Interferon production in Propionibacterium acnes treated mice

Interferonogenic properties of Propionibacterium acnes (PA) was studied in vivo and in vitro using CBA, BALB/c and 129AoBoy strains of mice. IFN was induced only in CBA strain after i.v. PA injection. BALB/c and 129AoBoy mice did not produce IFN. In the sera of CBA mice, obtained after i.p. injection of PA, interferon was not found. However, spleen cells of these mice produced IFN beginning from the 3rd day after injection. This interferon response lasted until the 10th week. Furthermore, in vivo studies showed enhancement effect of PA i.p. injection on IFN synthesis when NDV was introduced i.v. as inducer. The increased interferon level was also observed in the peritoneal cells isolated from PA--pretreated mice, induced in vitro with NDV or PA.     

Comparative study of the interferon-inducing capacity of Newcastle disease and Sendai viruses in human and animal bone marrow cells

Sendai and Newcastle disease virus (NDV) induced high and approximately similar interferon production in bone marrow cells of man, rats, and mice. In contrast to NDV, Sendai virus induced no interferon production in chick bone marrow cells. Interferon production by the bone marrow cells did not differ in its time course and requirements from production of leukocyte interferon.     

Migration inhibitory factor and interferon in the circulation of mice with delayed hypersensitivity

When mice infected with Mycobacterium tuberculosis strain BCG were inoculated intravenously with old tuberculin (OT) or living BCG cells, both migration inhibitory factor (MIF) and interferon appeared in the circulation within a few hours. In such animals, which showed delayed hypersensitivity by footpad tests, as little as 1.5 mg of OT or as few as 1.7 × 10⁶ bacteria per mouse were capable of eliciting circulating MIF and interferon. Uninfected animals inoculated with large doses of OT or living BCG cells did not produce MIF or interferon. When nonspecific stimuli such as bacterial lipopolysaccharide (LPS; from Salmonella typhimurium strain LT-2), heat-killed Brucella abortus, Newcastle disease virus (NDV), and polyinosinic acid:polycytidilic acid (poly I:C) were inoculated intravenously into BCG-infected mice, MIF was produced in the circulation of animals challenged with LPS or Brucella but not in those challenged with NDV or poly I:C, although all the stimuli were capable of eliciting an interferon response. The interferon elicited in BCG-infected mice by specific antigen differed in at least one important property from the viral inhibitor produced by the nonspecific stimuli. The interferon which appeared after injection of OT or living BCG cells was destroyed by treatment at pH 2 for 24 hr at 4C, whereas the interferons produced after injection of the nonspecific stimuli were stable under the same conditions. The MIF activity in plasma from sensitized mice inoculated with specific antigen was also destroyed by treatment at pH 2. When mouse plasma containing both MIF and interferon activity was filtered through Sephadex G-100, both mediators were excluded in the same peak fractions. Sensitization of mice with complete Freund adjuvant instead of infection with BCG cells produces a different pattern of response. Although hypersensitive to specific antigen by footpad swelling tests, mice sensitized with complete Freund adjuvant failed to produce MIF or interferon when they were inoculated intravenously with OT or living BCG cells.     

Interferon-hyporesponsiveness in mice in connection with dose interval and site of administration of Newcastle disease virus. Reflexion in serum complement levels.

Intraperitoneal administration of Newcastle disease virus (NDV) resulted in enhanced serum levels of complement not accompanied by an increase of interferon levels, when measured at 24 hours' intervals. On the other hand, intravenous injection of NDV caused a drop of complement levels of short duration with an accompanying increase of interferon levels. Hyporeactivity to induction of serum interferon could not be achieved by intraperitoneal administration of NDV, but an incomplete hyporeactivity could be achieved by intravenous administration of NDV. It might be assumed that production of interferon in mice occurs in different separated compartments depending on the route of inoculation of the inducer.     

Effect of verapamil on interferon production, mastocyte degranulation and histamine release--studies in mice sensitized with ovalbumin

The effect of verapamil, calcium antagonist, on the IFN production, the histamine release and degranulation of mastocytes were studied. The mastocytes were harvested from mice sensitized with ovalbumin, treated with verapamil and induced with Newcastle virus (NDV) to the interferon production (IFN). It has been shown that the percentage of degranulation was lower in mastocytes of the mice treated with verapamil, both induced with NDV and non-induced ones. The titer of interferon was lower when the mice induced with virus were injected with verapamil. The inhibitory effect of this drug on histamine release has only been found on the 8th day after the sensitization.     

Effect of ethylene diamine tetraacetate (EDTA) on the interferon-inducing activity of Newcastle disease virus (NDV)in vivo

Summary Circulating interferon levels increased and persisted for prolonged periods of time when groups of mice were stimulated with a mixture of Newcastle disease virus (NDV) and EDTA, or pretreated with EDTA and then injected with NDV, instead of injection with NDV alone.     

Protective effectiveness of the blood sera and immune system cells from mice immunized with live or inactivated influenza virus

The results of the study of comparative protective role of humoral and cell-mediated immunity factors obtained from inbred donor mice immunized with live or inactivated influenza virus are presented. The superiority of the live virus over the inactivated preparation as the inducer of not only humoral but especially cell-mediated immune response was demonstrated by the effectiveness of passive intranasal protection of the infected mice, by the degree of inhibition of virus reproduction in the lungs of the protected mice, and by the capacity for interferon production by the cells of the immune system of donor mice.     

X-linked resistance of mice to high doses of herpes simplex virus type 2 correlates with early interferon production.

Mice inoculated intraperitoneally with herpes simplex virus type 2 develop focal necrotizing hepatitis and eventually die from ascending myelitis and encephalitis. The genetics of resistance to the infection were analyzed in crosses between resistant C57BL/10 mice and susceptible BALB/c mice. It was shown that the resistance of C57BL/10 mice to hepatitis induction was influenced by an X-linked dominant gene as previously shown for the GR mouse strain. The course of infection in the liver pointed to early, natural defense mechanisms as being responsible for the difference between the mouse strains, whereas the clearance of virus from the liver, probably mediated by specific immunity, was exerted at the same time and with equal efficiency for all groups of mice. In mortality experiments, resistance was shown to be an autointerference phenomenon in that a considerable number of C57BL/10 mice survived an intraperitoneal injection of 10(6) PFU, whereas all mice were killed by 10(5) PFU. This resistance of C57BL/10 mice to high doses of HSV-2 was retrieved in all groups of F1 mice in crosses between C57BL/10 and BALB/c mice except the (BALB/c female X C57 male) male group, in which the mice receive the X chromosome from the susceptible BALB/c female. Thus, the autointerference phenomenon also seems to be influenced by loci on the X chromosome. A similar pattern of inheritance was observed when early interferon induction (4 to 5 h after infection) in response to HSV-2 was measured. The possible relevance of this early interferon response in conjunction with other potential natural defense mechanisms is discussed.     

Effect of maternal antibodies on influenza virus-specific immune response elicited by inactivated virus and naked DNA

While vaccines are effective in adults, they are less successful in newborns and infants. Neonatal unresponsiveness to vaccines could be owing to immaturity of lymphocytes and/or to inhibition by maternal antibodies. Unresponsiveness of newborn to vaccines can be overcame by genetic immunization. In the present study we investigated the effect of maternal antibodies on the anti-influenza virus protective response in progeny born to dams immunized with plasmid containing the hemagglutinin gene or UV-inactivated virus. The effect of maternal antibodies was studied in plasmid immunized F1 mice born to BALB/c dams, previously immunized with virus or plasmid and crossed with C57BL/6 males, as well as in offspring born to BALB/c dams immunized with plasmid and then immunized with UV-inactivated WSN virus. We have found that the inhibition period of the anti-HA antibody response in offspring born to dams immunized with DNA is shorter than that of offspring born to dams immunized with virus. Furthermore, there is a persistent inhibitory effect on B cells from offspring born to dams immunized with virus or injected with antiviral monoclonal antibodies (MoAb), after the decline of maternal antibody titers. The analysis of the haemagglutinin-specific clonotype reactivity pattern of offspring born to dams immunized with inactivated influenza virus or with a plasmid showed that clonotypes producing antibodies specific for the immunizing virus strain were predominant in offspring born to dams immunized with DNA compared to those born to dams immunized with virus. Maternal antibodies do not affect cell-mediated immunity. These findings might be used to design efficient vaccination schedules for newborns and infants.     

Distinguishing characteristics of interferon induction with poly(I)-poly(C) and Newcastle disease virus in human cells.

Exposure of a human diploid foreskin cell strain (FS-4) to polyinosinate-polycytidylate [poly(I)·poly(C)] resulted in an early rise of interferon production that peaked at about 4 hr after induction and decreased rapidly thereafter. Irradiation of cells with low to moderate doses of ultraviolet (uv) light immediately before induction with poly(I)·poly(C) increased the amount of interferon produced up to about tenfold. This enhancement was apparently due to interference with the shut-off process which in unirradiated cells leads to early termination of interferon production; in irradiated cells interferon production continued for much longer.Inoculation of FS-4 cells with Newcastle disease virus (NDV) resulted in interferon production that showed a slower rise and peaked only at about 10–15 hr after inoculation. Irradiation of cells at the time of induction with NDV resulted in a dose-dependent decrease of interferon production. However, a small fraction of the total amount of interferon produced in response to NDV, which appeared by about 5 hr after virus inoculation, was resistant to uv. This uv-resistant early peak of NDV-induced interferon was greatly enhanced in cells which 6 hr before virus inoculation had either been induced with poly(I)·poly(C) or incubated with interferon, while the appearance of the major, uv-sensitive peak of NDV-induced interferon was inhibited or delayed after the same treatments. In its characteristics the early peak of NDV-induced interferon resembled the poly(I)·poly(C)-induced interferon response. Poly(I)·poly(C)-induced, as well as the early and late NDV-induced interferons were all neutralized by an antiserum raised against poly(I)·poly(C)-induced interferon, suggesting that they represent products of the same structural gene(s). It is concluded that there may be more than one mechanism of interferon induction by a single virus.