Herpes simplex virus type 1-mediated induction of human immunodeficiency virus type 1 provirus correlates with binding of nuclear proteins to the NF-kappa B enhancer and leader sequence.

Herpes simplex virus type 1 (HSV-1) infection induces expression of the human immunodeficiency virus type 1 (HIV-1) provirus in the chronically infected T-cell line ACH-2. The HSV-1-mediated induction correlates with the appearance of two NF-kappa B-specific proteins of 55 and 85 kDa in the nucleus and with the binding of 50-kDa nuclear protein to the LBP-1 binding site of the untranslated leader sequence of the HIV-1 long terminal repeat. The HSV-1-induced LBP-1 binding protein, designated HLP-1, is present exclusively in HSV-1-infected, but not in phorbol-12-myristate-13-acetate- or tumor necrosis factor alpha-treated ACH-2 cells. Both the NF-kappa B and LBP-1 target sequences, when inserted either alone or together 5' of a heterologous minimal promoter (thymidine kinase), confer inducibility by HSV-1 infection in a transient transfection assay. Thus, it appears that the HSV-1-mediated activation of HIV-1 provirus is brought about by the binding of both NF-kappa B and HLP-1 specific proteins to two distinct regions of HIV-1 long terminal repeat.     

Macrophage control of herpes simplex virus type 1 replication in the peripheral nervous system

After corneal infection, herpes simplex virus type 1 (HSV-1) invades sensory neurons with cell bodies in the trigeminal ganglion (TG), replicates briefly, and then establishes a latent infection in these neurons. HSV-1 replication in the TG can be detected as early as 2 days after corneal infection, reaches peak titers by 3-5 days after infection, and is undetectable by 7-10 days. During the period of HSV-1 replication, macrophages and gammadelta TCR+ T lymphocytes infiltrate the TG, and TNF-alpha, IFN-gamma, the inducible nitric oxide synthase (iNOS) enzyme, and IL-12 are expressed. TNF-alpha, IFN-gamma, and the iNOS product nitric oxide (NO) all inhibit HSV-1 replication in vitro. Macrophage and gammadelta TCR+ T cell depletion studies demonstrated that macrophages are the main source of TNF-alpha and iNOS, whereas gammadelta TCR+ T cells produce IFN-gamma. Macrophage depletion, aminoguanidine inhibition of iNOS, and neutralization of TNF-alpha or IFN-gamma all individually and synergistically increased HSV-1 titers in the TG after HSV-1 corneal infection. Moreover, individually depleting macrophages or neutralizing TNF-alpha or IFN-gamma markedly reduced the accumulation of both macrophages and gammadelta TCR+ T cells in the TG. Our findings establish that after primary HSV-1 infection, the bulk of virus replication in the sensory ganglia is controlled by macrophages and gammadelta TCR+ T lymphocytes through their production of antiviral molecules TNF-alpha, NO, and IFN-gamma. Our findings also strongly suggest that cross-regulation between these two cell types is necessary for their accumulation and function in the infected TG.     

Role of Nitric Oxide in the Pathogenesis of Encephalomyocarditis Virus-Induced Diabetes in Mice

The D variant of encephalomyocarditis virus (EMC-D virus) causes diabetes in mice by destroying pancreatic β cells. In mice infected with a low dose of EMC-D virus, macrophages play an important role in β-cell destruction by producing soluble mediators such as interleukin-1β (IL-1β), tumor necrosis factor alpha (TNF-α), and nitric oxide (NO). To investigate the role of NO and inducible NO synthase (iNOS) in the development of diabetes in EMC-D virus-infected mice, we infected iNOS-deficient DBA/2 mice with EMC-D virus (2 × 10² PFU/mouse). Mean blood glucose levels in EMC-D virus-infected iNOS-deficient mice and wild-type mice were 205.5 and 466.7 mg/dl, respectively. Insulitis and macrophage infiltration were reduced in islets of iNOS-deficient mice compared with wild-type mice at 3 days after EMC-D virus infection. Apoptosis of β cells was decreased in iNOS-deficient mice, as evidenced by reduced numbers of terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling-positive cells. There were no differences in mRNA expression of antiapoptotic molecules Bcl-2, Bcl-xL, Bcl-w, Mcl-1, cIAP-1, and cIAP-2 between wild-type and iNOS-deficient mice, whereas expression of proapoptotic Bax and Bak mRNAs was significantly decreased in iNOS-deficient mice. Expression of IL-1β and TNF-α mRNAs was significantly decreased in both islets and macrophages of iNOS-deficient mice compared with wild-type mice after EMC-D virus infection. Nuclear factor κB was less activated in macrophages of iNOS-deficient mice after virus infection. We conclude that NO plays an important role in the activation of macrophages and apoptosis of pancreatic β cells in EMC-D virus-infected mice and that deficient iNOS gene expression inhibits macrophage activation and β-cell apoptosis, contributing to prevention of EMC-D virus-induced diabetes.Type 1 diabetes results from absolute insulin deficiency caused by destruction of insulin-producing pancreatic β cells. The D variant of encephalomyocarditis virus (EMC-D virus) induces diabetes in genetically susceptible strains of mice by infecting and destroying β cells (13-18). In mice infected with a low dose (1 × 10² PFU/mouse) of EMC-D virus, macrophages play a central role in the destruction of pancreatic β cells (4, 5, 13-15), as evidenced by a significant increase in the incidence of diabetes if macrophages are activated prior to viral infection and complete prevention of EMC-D virus-induced diabetes if macrophages are inactivated prior to viral infection (4). Additional studies found that selective EMC-D viral infection of pancreatic β cells results in an initial recruitment of macrophages into the islets, followed by infiltration of other immunocytes, including T cells, natural killer cells, and B cells (5).EMC-D virus infects and activates macrophages without replication (13) and induces the production of soluble mediators such as interleukin-1β (IL-1β), tumor necrosis factor alpha (TNF-α), and inducible nitric oxide synthase (iNOS), which play important roles in the destruction of β cells (14). These infected macrophages express significantly more iNOS than either IL-1β or TNF-α (13). Treatment of EMC-D virus-infected mice with the tyrosine kinase inhibitor AG126, which inhibits nitric oxide (NO) production in EMC-D virus-infected macrophages, decreases the expression of IL-1β and TNF-α in the pancreatic islets and the incidence of diabetes and insulitis compared with those in vehicle-treated control mice (13). As well, treatment of EMC-D virus-infected mice with an iNOS inhibitor decreases the incidence of diabetes (14). These results suggest that iNOS and NO significantly contribute to the destruction of pancreatic β cells in mice infected with a low dose of EMC-D virus, although their roles are not fully understood.To directly test whether iNOS and NO play a critical role in the pathogenesis of EMC-D virus-induced diabetes in mice, we used iNOS knockout (KO) DBA/2 mice. We found that iNOS-deficient mice infected with EMC-D virus (2 × 10² PFU/mouse) showed a significantly lower incidence of diabetes. There was reduced expression of IL-1β and TNF-α in macrophages and decreased infiltration of immunocytes in the islets of iNOS-deficient mice, resulting in reduced apoptosis of β cells compared with that in EMC-D virus-infected wild-type mice. This study provides direct evidence of a role of NO in the activation of macrophages by EMC-D viral infection and in the pathogenesis of low-dose (2 × 10² PFU/mouse) EMC-D virus-induced diabetes.     

Differential effects of a Toll-like receptor antagonist on Mycobacterium tuberculosis-induced macrophage responses

We previously showed that viable Mycobacterium tuberculosis (Mtb) bacilli contain distinct ligands that activate cells via the mammalian Toll-like receptor (TLR) proteins TLR2 and TLR4. We now demonstrate that expression of a dominant negative TLR2 or TLR4 proteins in RAW 264.7 macrophages partially blocked Mtb-induced NF-kappa B activation. Coexpression of both dominant negative proteins blocked virtually all Mtb-induced NF-kappa B activation. The role of the TLR4 coreceptor MD-2 was also examined. Unlike LPS, Mtb-induced macrophage activation was not augmented by overexpression of ectopic MD-2. Moreover, cells expressing an LPS-unresponsive MD-2 mutant responded normally to Mtb. We also observed that the lipid A-like antagonist E5531 specifically inhibited TLR4-dependent Mtb-induced cellular responses. E5531 could substantially block LPS- and Mtb-induced TNF-alpha production in both RAW 264.7 cells and primary human alveolar macrophages (AM phi). E5531 inhibited Mtb-induced AM phi apoptosis in vitro, an effect that was a consequence of the inhibition of TNF-alpha production by E5531. In contrast, E5531 did not inhibit Mtb-induced NO production in RAW 264.7 cells and AM phi. Mtb-stimulated peritoneal macrophages from TLR2- and TLR4-deficient animals produced similar amounts of NO compared with control animals, demonstrating that these TLR proteins are not required for Mtb-induced NO production. Lastly, we demonstrated that a dominant negative MyD88 mutant could block Mtb-induced activation of the TNF-alpha promoter, but not the inducible NO synthase promoter, in murine macrophages. Together, these data suggest that Mtb-induced TNF-alpha production is largely dependent on TLR signaling. In contrast, Mtb-induced NO production may be either TLR independent or mediated by TLR proteins in a MyD88-independent manner.     

Role of Hck in the pathogenesis of encephalomyocarditis virus-induced diabetes in mice

Soluble mediators such as interleukin-1beta, tumor necrosis factor alpha (TNF-alpha), and inducible nitric oxide synthase (iNOS) produced from activated macrophages play an important role in the destruction of pancreatic beta cells in mice infected with a low dose of the D variant of encephalomyocarditis (EMC-D) virus. The tyrosine kinase signaling pathway was shown to be involved in EMC-D virus-induced activation of macrophages. This investigation was initiated to determine whether the Src family of kinases plays a role in the activation of macrophages, subsequently resulting in the destruction of beta cells, in mice infected with a low dose of EMC-D virus. We examined the activation of p59/p56(Hck), p55(Fgr), and p56/p53(Lyn) in macrophages from DBA/2 mice infected with the virus. We found that p59/p56(Hck) showed a marked increase in both autophosphorylation and kinase activity at 48 h after infection, whereas p55(Fgr) and p56/p53(Lyn) did not. The p59/p56(Hck) activity was closely correlated with the tyrosine phosphorylation level of Vav. Treatment of EMC-D virus-infected mice with the Src kinase inhibitor, PP2, resulted in the inhibition of p59/p56(Hck) activity and almost complete inhibition of the production of TNF-alpha and iNOS in macrophages and the subsequent prevention of diabetes in mice. On the basis of these observations, we conclude that the Src kinase, p59/p56(Hck), plays an important role in the activation of macrophages and the subsequent production of TNF-alpha and nitric oxide, leading to the destruction of pancreatic beta cells, which results in the development of diabetes in mice infected with a low dose of EMC-D virus.     

Vaccinia virus infection attenuates innate immune responses and antigen presentation by epidermal dendritic cells

Langerhans cells (LCs) are antigen-presenting cells in the skin that play sentinel roles in host immune defense by secreting proinflammatory molecules and activating T cells. Here we studied the interaction of vaccinia virus with XS52 cells, a murine epidermis-derived dendritic cell line that serves as a surrogate model for LCs. We found that vaccinia virus productively infects XS52 cells, yet this infection displays an atypical response to anti-poxvirus agents. Whereas adenosine N1-oxide blocked virus production and viral protein synthesis during a synchronous infection, cytosine arabinoside had no effect at concentrations sufficient to prevent virus replication in BSC40 monkey kidney cells. Vaccinia virus infection of XS52 cells not only failed to elicit the production of various cytokines, including tumor necrosis factor alpha (TNF-alpha), interleukin-1beta (IL-1beta), IL-6, IL-10, IL-12 p40, alpha interferon (IFN-alpha), and IFN-gamma, it actively inhibited the production of proinflammatory cytokines TNF-alpha and IL-6 by XS52 cells in response to exogenous lipopolysaccharide (LPS) or poly(I:C). Infection with a vaccinia virus mutant lacking the E3L gene resulted in TNF-alpha secretion in the absence of applied stimuli. Infection of XS52 cells or BSC40 cells with the DeltaE3L virus, but not wild-type vaccinia virus, triggered proteolytic decay of IkappaBalpha. These results suggest a novel role for the E3L protein as an antagonist of the NF-kappaB signaling pathway. DeltaE3L-infected XS52 cells secreted higher levels of TNF-alpha and IL-6 in response to LPS and poly(I:C) than did cells infected with the wild-type virus. XS52 cells were productively infected by a vaccinia virus mutant lacking the K1L gene. DeltaK1L-infected cells secreted higher levels of TNF-alpha and IL-6 in response to LPS than wild-type virus-infected cells. Vaccinia virus infection of primary LCs harvested from mouse epidermis was nonpermissive, although a viral reporter protein was expressed in the infected LCs. Vaccinia virus infection of primary LCs strongly inhibited their capacity for antigen-specific activation of T cells. Our results highlight suppression of the skin immune response as a feature of orthopoxvirus infection.     

Augmentation of pathogenesis of coxsackievirus B3 infections in mice by exogenous administration of interleukin-1 and interleukin-2.

Two variants of coxsackievirus B3 (CVB3) which differ dramatically in the ability to induce myocarditis in BALB/c mice were studied. H3 virus infection of murine monocytes in vitro resulted in release of concentrations of interleukin 1 (IL-1) and alpha/beta interferon that were high compared with those of cells infected with the H310A1 virus variant. In vivo, H3 virus infection caused substantial inflammatory cell infiltration of the myocardium, and lymphocytes from these animals gave predominantly Th1-cell responses to either whole H3 virus or overlapping peptides of the CVB3 vp1 capsid protein, as determined by IL-2 production. In contrast, H310A1 virus infection produced minimal myocarditis and Th1-cell responses, but Th2-cell activation was more pronounced than in H3 virus-infected mice (as determined by IL-4 concentrations). Exogenous treatment of H310A1 virus-infected mice with either IL-1 or IL-2 restored both myocarditis susceptibility and Th1-cell responses to whole virus and vp1 peptides. Furthermore, H310A1 virus-infected mice given exogenous IL-1 showed substantial in situ IL-2 deposition in the myocardium. These results indicate that CVB3-induced myocarditis may depend upon release of specific cytokines during infection and that activation of Th1 cells may be an important factor in pathogenesis.     

Contact-dependent interference with invariant NKT cell activation by herpes simplex virus-infected cells

Invariant CD1d-restricted NKT (iNKT) cells play important roles in generating protective immune responses against infections. In this study, we have investigated the role of human iNKT cells in HSV-1 infection and their interaction with epidermal keratinocytes. These cells express CD1d and are the primary target of the virus. Keratinocytes loaded with α-galactosyl ceramide (α-GalCer) could stimulate IFN-γ production and CD25 upregulation by iNKT cells. However, both α-GalCer-dependent and cytokine-dependent activation of iNKT cells was impaired after coculture with HSV-1-infected cells. Notably, CD1d downregulation was not observed on infected keratinocytes, which were also found to inhibit TCR-independent iNKT cell activation. Further examination of the cytokine profile of iNKT-keratinocyte cocultures showed inhibition of IFN-γ, IL-5, IL-10, IL-13, and IL-17 secretion but upregulation of IL-4 and TNF-α after the infection. Moreover, cell-to-cell contact between infected keratinocytes and iNKT cells was required for the inhibition of activation, as the cell-free supernatants containing virus did not affect activation. Productive infection of iNKT cells was however not required for the inhibitory effect. After coculture with infected cells, iNKT cells were no longer responsive to further stimulation with α-GalCer-loaded CD1d-expressing cells. We found that exposure to HSV-1-infected cells resulted in impaired TCR signaling downstream of ZAP70. Additionally, infected cells upregulated the expression of the negative T cell regulator, galectin-9; however, blocking experiments indicated that the impairment of iNKT cell responses was independent of galectin-9. Thus, interference with activation of human iNKT cells by HSV-1 may represent a novel immunoevasive strategy used by the virus to avoid immune clearance.     

Human dendritic cells are activated by dengue virus infection: enhancement by gamma interferon and implications for disease pathogenesis

The ability of dendritic cells (DCs) to shape the adaptive immune response to viral infection is mediated largely by their maturation and activation state as determined by the surface expression of HLA molecules, costimulatory molecules, and cytokine production. Dengue is an emerging arboviral disease where the severity of illness is influenced by the adaptive immune response to the virus. In this report, we have demonstrated that dengue virus infects and replicates in immature human myeloid DCs. Exposure to live dengue virus led to maturation and activation of both the infected and surrounding, uninfected DCs and stimulated production of tumor necrosis factor alpha (TNF-alpha) and alpha interferon (IFN-alpha). Activation of the dengue virus-infected DCs was blunted compared to the surrounding, uninfected DCs, and dengue virus infection induced low-level release of interleukin-12 p70 (IL-12 p70), a key cytokine in the development of cell-mediated immunity (CMI). Upon the addition of IFN-gamma, there was enhanced activation of dengue virus-infected DCs and enhanced dengue virus-induced IL-12 p70 release. The data suggest a model whereby DCs are the early, primary target of dengue virus in natural infection and the vigor of CMI is modulated by the relative presence or absence of IFN-gamma in the microenvironment surrounding the virus-infected DCs. These findings are relevant to understanding the pathogenesis of dengue hemorrhagic fever and the design of new vaccination and therapeutic strategies.     

IFN-gamma-induced L-arginine-dependent toxoplasmastatic activity in murine peritoneal macrophages is mediated by endogenous tumor necrosis factor-alpha.

Activated murine peritoneal macrophages inhibit the intracellular proliferation of Toxoplasma gondii and produce a number of cytokines, such as TNF-alpha and IL-1. Both TNF-alpha and IL-1 have been reported to be involved in the immune response against various microorganisms, but the mechanisms responsible for these effects are not known. In the present study it was investigated whether endogenously produced TNF-alpha and IL-1 are involved in the activation of peritoneal macrophages by rIFN-gamma leading to toxoplasmastatic activity and the production of reactive nitrogen intermediates. The rIFN-gamma-induced toxoplasmastatic activity was inhibited by neutralizing antibodies against mouse TNF-alpha in a dose-dependent and time-dependent way, but neutralizing antibodies against mouse IL-1 alpha and IL-1 beta did not affect this activity. Involvement of TNF-alpha in the induction of toxoplasmastatic activity was confirmed by our finding that rTNF-alpha in combination with a nonactivating concentration of rIFN-gamma inhibited the intracellular proliferation of T. gondii. No synergistic activity of rIL-1 and rIFN-gamma on the inhibition of T. gondii proliferation was found. Both rTNF-alpha and rIL-1 alpha alone inhibited the intracellular proliferation of T. gondii only slightly. Because it has been reported recently that activated macrophages produce reactive nitrogen intermediates that are essential in the induction of toxoplasmastatic activity, we investigated whether these intermediates are involved in the TNF-dependent induction of toxoplasmastatic activity. Neutralizing antibodies against mouse TNF-alpha inhibited also the release of NO2- by rIFN-gamma-activated macrophages almost completely. Macrophages incubated with rTNF-alpha in combination with a nonactivating concentration of rIFN-gamma released substantial amounts of NO2-, but rTNF-alpha and rIL-1 alpha alone, and the combination of rIL-1 alpha and a nonactivating concentration of rIFN-gamma induced only little NO2(-)-release by macrophages. To assess whether reactive nitrogen intermediates act directly or indirectly on the intracellular proliferation of T. gondii, macrophages were incubated with the L-arginine analog NG-monomethyl-L-arginine or the NADPH-inhibitor diphenylene iodonium, both inhibitors of the generation of reactive nitrogen intermediates. Good correlation was found between toxoplasmastatic activity and the release of NO2- during the 24-h activation period before infection of the macrophages with T. gondii, but no correlation was found between toxoplasmastatic activity and the release of NO2- during infection of the macrophages.(ABSTRACT TRUNCATED AT 400 WORDS)     

TLR9-signaling pathways are involved in Kilham rat virus-induced autoimmune diabetes in the biobreeding diabetes-resistant rat

Viral infections are associated epidemiologically with the expression of type 1 diabetes in humans, but the mechanisms underlying this putative association are unknown. To investigate the role of viruses in diabetes, we used a model of viral induction of autoimmune diabetes in genetically susceptible biobreeding diabetes-resistant (BBDR) rats. BBDR rats do not develop diabetes in viral-Ab-free environments, but approximately 25% of animals infected with the parvovirus Kilham rat virus (KRV) develop autoimmune diabetes via a mechanism that does not involve beta cell infection. Using this model, we recently documented that TLR agonists synergize with KRV infection and increase disease penetrance. We now report that KRV itself activates innate immunity through TLR ligation. We show that KRV infection strongly stimulates BBDR splenocytes to produce the proinflammatory cytokines IL-6 and IL-12p40 but not TNF-alpha. KRV infection induces high levels of IL-12p40 by splenic B cells and Flt-3-ligand-induced bone marrow-derived dendritic cells (DCs) but only low levels of IL-12p40 production by thioglycolate-elicited peritoneal macrophages or GM-CSF plus IL-4-induced bone marrow-derived DCs. KRV-induced cytokine production is blocked by pharmacological inhibitors of protein kinase R and NF-kappaB. Genomic KRV DNA also induces BBDR splenocytes and Flt-3L-induced DCs from wild-type but not TLR9-deficient mice to produce IL-12p40; KRV-induced up-regulation of B lymphocytes can be blocked by TLR9 antagonists including inhibitory CpG and chloroquine. Administration of chloroquine to virus-infected BBDR rats decreases the incidence of diabetes and decreases blood levels of IL-12p40. Our data implicate the TLR9-signaling pathway in KRV-induced innate immune activation and autoimmune diabetes in the BBDR rat.     

Differential contribution of herpes simplex virus type 1 gene products and cellular factors to the activation of human immunodeficiency virus type 1 provirus.

We have previously reported that infection with herpes simplex virus type 1 (HSV-1) activates expression of the human immunodeficiency virus type 1 (HIV-1) provirus in T cells. Activation of the HIV-1 provirus correlated with the activation of binding of 55- and 85-kDa proteins to the kappa B enhancer and binding of the 50-kDa HLP-1 protein to the LBP-1 sequences of the HIV-1 long terminal repeat. Further examination of this system has shown that the inhibition of HSV-1 replication by the antiviral drug acyclovir does not inhibit HSV-1-mediated induction of HIV-1 provirus. Surprisingly, the NF-kappa B and HLP-1 binding activities were substantially inhibited in acyclovir-treated cells. In the transient-transfection assay, ICP0, but not ICP4, activated the HIV-1 long terminal repeat promoter region and the effect of ICP0 was greatly enhanced in the presence of the NF-kappa B binding proteins, suggesting that induction of the HIV-1 provirus involves cooperation between the HSV-1-activated cellular factor, NF-kappa B, and the virus-encoded transactivator, ICP0.     

Interleukin-21 Receptor Signalling Is Important for Innate Immune Protection against HSV-2 Infections

Interleukin (IL) -21 is produced by Natural Killer T (NKT) cells and CD4⁺ T cells and is produced in response to virus infections, where IL-21 has been shown to be essential in adaptive immune responses. Cells from the innate immune system such as Natural Killer (NK) cells and macrophages are also important in immune protection against virus. These cells express the IL-21 receptor (IL-21R) and respond to IL-21 with increased cytotoxicity and cytokine production. Currently, however it is not known whether IL-21 plays a significant role in innate immune responses to virus infections. The purpose of this study was to investigate the role of IL-21 and IL-21R in the innate immune response to a virus infection. We used C57BL/6 wild type (WT) and IL-21R knock out (KO) mice in a murine vaginal Herpes Simplex Virus type 2 (HSV-2) infection model to show that IL-21 – IL-21R signalling is indeed important in innate immune responses against HSV-2. We found that the IL-21R was expressed in the vaginal epithelium in uninfected (u.i) WT mice, and expression increased early after HSV-2 infection. IL-21R KO mice exhibited increased vaginal viral titers on day 2 and 3 post infection (p.i.) and subsequently developed significantly higher disease scores and a lower survival rate compared to WT mice. In addition, WT mice infected with HSV-2 receiving intra-vaginal pre-treatment with murine recombinant IL-21 (mIL-21) had decreased vaginal viral titers on day 2 p.i., significantly lower disease scores, and a higher survival rate compared to infected untreated WT controls. Collectively our data demonstrate the novel finding that the IL-21R plays a critical role in regulating innate immune responses against HSV-2 infection.     

Increased interleukin-10 mRNA expression in tumor-bearing or persistently lymphocytotic animals infected with bovine leukemia virus.

Interleukin-10 (IL-10), produced by Th2 helper T cells, B cells, and macrophages, can inhibit cytokine production by Th1 cells and enhance B-cell proliferation and differentiation. Here, we show that peripheral blood mononuclear cells (PBMCs) from bovine leukemia virus-infected animals with late-stage disease express considerably more IL-10 mRNA than animals that are not infected or that are in the early stages of disease. In contrast, the quantities of type 1 cytokines, IL-2 and gamma interferon, decrease with disease progression. In addition, we observed that IL-10 is expressed principally by monocytes/macrophages, not B lymphocytes, in persistently lymphocytotic animals. This observation supports a role for monocytes/macrophages in progression of bovine leukemia virus infection and, of importance, indicates that proliferating B cells are not the source of IL-10 expression. These findings suggest that IL-10 produced by monocytes/macrophages may influence the progression of bovine leukosis in animals that develop persistent lymphocytosis of B cells or B-cell lymphosarcoma.