Defective IFN-γ Production Research Articles

Severe COVID-19 patients display hyper-activated NK cells and NK cell-platelet aggregates.

COVID-19 is characterised by a broad spectrum of clinical and pathological features. Natural killer (NK) cells play an important role in innate immune responses to viral infections. Here, we analysed the phenotype and activity of NK cells in the blood of COVID-19 patients using flow cytometry, single-cell RNA-sequencing (scRNA-seq), and a cytotoxic killing assay. In the plasma of patients, we quantified the main cytokines and chemokines. Our cohort comprises COVID-19 patients hospitalised in a low-care ward unit (WARD), patients with severe COVID-19 disease symptoms hospitalised in intensive care units (ICU), and post-COVID-19 patients, who were discharged from hospital six weeks earlier. NK cells from hospitalised COVID-19 patients displayed an activated phenotype with substantial differences between WARD and ICU patients and the timing when samples were taken post-onset of symptoms. While NK cells from COVID-19 patients at an early stage of infection showed increased expression of the cytotoxic molecules perforin and granzyme A and B, NK cells from patients at later stages of COVID-19 presented enhanced levels of IFN-γ and TNF-α which were measured ex vivo in the absence of usual in vitro stimulation. These activated NK cells were phenotyped as CD49a+CD69a+CD107a+ cells, and their emergence in patients correlated to the number of neutrophils, and plasma IL-15, a key cytokine in NK cell activation. Despite lower amounts of cytotoxic molecules in NK cells of patients with severe symptoms, majority of COVID-19 patients displayed a normal cytotoxic killing of Raji tumour target cells. In vitro stimulation of patients blood cells by IL-12+IL-18 revealed a defective IFN-γ production in NK cells of ICU patients only, indicative of an exhausted phenotype. ScRNA-seq revealed, predominantly in patients with severe COVID-19 disease symptoms, the emergence of an NK cell subset with a platelet gene signature that we identified by flow and imaging cytometry as aggregates of NK cells with CD42a+CD62P+ activated platelets. Post-COVID-19 patients show slow recovery of NK cell frequencies and phenotype. Our study points to substantial changes in NK cell phenotype during COVID-19 disease and forms a basis to explore the contribution of platelet-NK cell aggregates to antiviral immunity against SARS-CoV-2 and disease pathology.

Chronic upper airway inflammation related to high Th2 cytokines in Mendelian susceptibility to mycobacterial disease case.

In this report, we have described a child suffering from Mendelian susceptibility to mycobacterial disease (MSMD) owing to an autosomal recessive, complete T-bet deficiency, which impairs IFN-γ production by innate and innate-like adaptive, but not mycobacterial-reactive purely adaptive lymphocytes. In this study, we explored the persistent upper airway inflammation (UAI) and blood eosinophilia in this patient. Unlike the wild-type (WT) T-bet, the mutant form of T-bet from this patient did not inhibit the production of T helper 2 (Th2) cytokines, including IL-4, IL-5, IL-9, and IL-13, when overexpressed in Th2 cells. Moreover, Herpesvirus saimiri immortalized T cells from the patient produced abnormally large amounts of Th2 cytokines, and the patient had markedly high plasma IL-5 and IL-13 concentrations. Finally, the patient’s CD4+ αβ T cells produced most of the Th2 cytokines in response to chronic stimulation, regardless of their antigen specificities, a phenotype reversed by the expression of WT T-bet. T-bet deficiency thus underlies the excessive production of Th2 cytokines, particularly IL-5 and IL-13, by CD4+ αβ T cells, causing blood eosinophilia and UAI. The MSMD of this patient results from defective IFN-γ production by innate and innate-like adaptive lymphocytes, whereas the UAI and eosinophilia result from excessive Th2 cytokine production by adaptive CD4+ αβ T lymphocytes.

Defective Interferon-Gamma Production Is Common in Chronic Pulmonary Aspergillosis.

Immune defects in chronic pulmonary aspergillosis (CPA) are poorly characterized. We compared peripheral blood cytokine profiles in patients with CPA versus healthy controls and explored the relationship with disease severity. Interferon-gamma (IFNγ), interleukin (IL)-17, tumor necrosis factor-α, IL-6, IL-12, and IL-10 were measured after in vitro stimulation of whole blood with lipopolysaccharide (LPS), phytohemagglutinin, β-glucan, zymosan (ZYM), IL-12 or IL-18, and combinations. Clinical parameters and mortality were correlated with cytokine production. Cytokine profiles were evaluated in 133 patients (57.1% male, mean age 61 years). In comparison to controls, patients with CPA had significantly reduced production of IFNγ in response to stimulation with β-glucan + IL-12 (312 vs 988 pg/mL), LPS + IL-12 (252 vs 1033 pg/mL), ZYM + IL-12 (996 vs 2347 pg/mL), and IL-18 + IL-12 (7193 vs 12330 pg/mL). Age >60 (hazard ratio [HR], 1.71; 95% confidence interval [CI], 1.00-2.91; P = .05) and chronic obstructive pulmonary disease (HR, 1.69; 95% CI, 1.03-2.78; P = .039) were associated with worse survival, whereas high IFNγ production in response to beta-glucan + IL-12 stimulation (HR, 0.48; 95% CI, .25-0.92; P = .026) was associated with reduced mortality. Patients with CPA show impaired IFNγ production in peripheral blood in response to stimuli. Defective IFNγ production ability correlates with worse outcomes. Immunotherapy with IFNγ could be beneficial for patients showing impaired IFNγ production in CPA.

High Th2 cytokine levels and upper airway inflammation in human inherited T-bet deficiency.

We have described a child suffering from Mendelian susceptibility to mycobacterial disease (MSMD) due to autosomal recessive, complete T-bet deficiency, which impairs IFN-γ production by innate and innate-like adaptive, but not mycobacterial-reactive purely adaptive, lymphocytes. Here, we explore the persistent upper airway inflammation (UAI) and blood eosinophilia of this patient. Unlike wild-type (WT) T-bet, the mutant form of T-bet from this patient did not inhibit the production of Th2 cytokines, including IL-4, IL-5, IL-9, and IL-13, when overexpressed in T helper 2 (Th2) cells. Moreover, Herpesvirus saimiri-immortalized T cells from the patient produced abnormally large amounts of Th2 cytokines, and the patient had markedly high plasma IL-5 and IL-13 concentrations. Finally, the patient's CD4+ αβ T cells produced most of the Th2 cytokines in response to chronic stimulation, regardless of their antigen specificities, a phenotype reversed by the expression of WT T-bet. T-bet deficiency thus underlies the excessive production of Th2 cytokines, particularly IL-5 and IL-13, by CD4+ αβ T cells, causing blood eosinophilia and UAI. The MSMD of this patient results from defective IFN-γ production by innate and innate-like adaptive lymphocytes, whereas the UAI and eosinophilia result from excessive Th2 cytokine production by adaptive CD4+ αβ T lymphocytes.

IMMUNODEFICIENCIES. Impairment of immunity to Candida and Mycobacterium in humans with bi-allelic RORC mutations.

Human inborn errors of immunity mediated by the cytokines interleukin-17A and interleukin-17F (IL-17A/F) underlie mucocutaneous candidiasis, whereas inborn errors of interferon-γ (IFN-γ) immunity underlie mycobacterial disease. We report the discovery of bi-allelic RORC loss-of-function mutations in seven individuals from three kindreds of different ethnic origins with both candidiasis and mycobacteriosis. The lack of functional RORγ and RORγT isoforms resulted in the absence of IL-17A/F-producing T cells in these individuals, probably accounting for their chronic candidiasis. Unexpectedly, leukocytes from RORγ- and RORγT-deficient individuals also displayed an impaired IFN-γ response to Mycobacterium. This principally reflected profoundly defective IFN-γ production by circulating γδ T cells and CD4(+)CCR6(+)CXCR3(+) αβ T cells. In humans, both mucocutaneous immunity to Candida and systemic immunity to Mycobacterium require RORγ, RORγT, or both.

Specific dysregulation of IFNγ production by natural killer cells confers susceptibility to viral infection.

Natural Killer (NK) cells contribute to the control of viral infection by directly killing target cells and mediating cytokine release. In C57BL/6 mice, the Ly49H activating NK cell receptor plays a key role in early resistance to mouse cytomegalovirus (MCMV) infection through specific recognition of the MCMV-encoded MHC class I-like molecule m157 expressed on infected cells. Here we show that transgenic expression of Ly49H failed to provide protection against MCMV infection in the naturally susceptible A/J mouse strain. Characterization of Ly49H+ NK cells from Ly49h-A transgenic animals showed that they were able to mount a robust cytotoxic response and proliferate to high numbers during the course of infection. However, compared to NK cells from C57BL/6 mice, we observed an intrinsic defect in their ability to produce IFNγ when challenged by either m157-expressing target cells, exogenous cytokines or chemical stimulants. This effect was limited to NK cells as T cells from C57BL/6 and Ly49h-A mice produced comparable cytokine levels. Using a panel of recombinant congenic strains derived from A/J and C57BL/6 progenitors, we mapped the genetic basis of defective IFNγ production to a single 6.6 Mb genetic interval overlapping the Ifng gene on chromosome 10. Inspection of the genetic interval failed to reveal molecular differences between A/J and several mouse strains showing normal IFNγ production. The chromosome 10 locus is independent of MAPK signalling or decreased mRNA stability and linked to MCMV susceptibility. This study highlights the existence of a previously uncovered NK cell-specific cis-regulatory mechanism of Ifnγ transcript expression potentially relevant to NK cell function in health and disease.

Cytokine production assays reveal discriminatory immune defects in adults with recurrent infections and noninfectious inflammation.

Cytokine production assays have been primarily used in research settings studying novel immunodeficiencies. We sought to determine the diagnostic value of cytokine production assays in patients with recurrent and/or severe infectious diseases (IDs) without known immunodeficiencies and unclassified noninfectious inflammatory disorders (NIIDs). We retrospectively examined cytokine production in whole-blood and peripheral blood mononuclear cell samples from 157 adult patients. A cytokine production rate of <5% of that of healthy controls was considered defective. While monocyte-derived cytokine (tumor necrosis factor alpha [TNF-α], interleukin-1β [IL-1β], and IL-6) production was rarely affected, 30% of all included patients had deficient production of interferon gamma (IFN-γ), IL-17A, or IL-22. Twenty-five percent of the NIID patients displayed defective IFN-γ production, whereas IL-17A production was generally unaffected. In the group of ID patients, defective IFN-γ production was found in 19% and 14% of the patients with viral and bacterial infections, respectively, and in 38%, 24%, and 50% of patients with mycobacterial, mucocutaneous, and invasive fungal infections, respectively. Defective IL-17A and IL-22 production was mainly confined to ID patients with mucocutaneous fungal infections. In conclusion, cytokine production assays frequently detect defective Th1 responses in patients with mycobacterial or fungal infections, in contrast to patients with respiratory tract infections or isolated bacterial infections. Defective IL-17A and IL-22 production was primarily found in patients with fungal infections, while monocyte-derived cytokine production was unaffected. Thus, lymphocyte-derived cytokine production assays are helpful in the diagnostic workup of patients with recurrent infections and suspected immunodeficiencies and have the potential to reveal immune defects that might guide adjunctive immunomodulatory therapy.

Soypeptide lunasin in cytokine immunotherapy for lymphoma

Immunostimulatory cytokines can enhance anti-tumor immunity and are part of the therapeutic armamentarium for cancer treatment. We have previously reported that post-transplant lymphoma patients have an acquired deficiency of signal transducer and activator of transcription 4, which results in defective IFNγ production during clinical immunotherapy. With the goal of further improving cytokine-based immunotherapy, we examined the effects of a soybean peptide called lunasin that synergistically works with cytokines on natural killer (NK) cells. Peripheral blood mononuclear cells of healthy donors and post-transplant lymphoma patients were stimulated with or without lunasin in the presence of IL-12 or IL-2. NK activation was evaluated, and its tumoricidal activity was assessed using in vitro and in vivo tumor models. Chromatin immunoprecipitation assay was performed to evaluate the histone modification of gene loci that are regulated by lunasin and cytokine. Adding lunasin to IL-12- or IL-2-stimulated NK cells demonstrated synergistic effects in the induction of IFNG and GZMB involved in cytotoxicity. The combination of lunasin and cytokines (IL-12 plus IL-2) was capable of restoring IFNγ production by NK cells from post-transplant lymphoma patients. In addition, NK cells stimulated with lunasin plus cytokines displayed higher tumoricidal activity than those stimulated with cytokines alone using in vitro and in vivo tumor models. The underlying mechanism responsible for the effects of lunasin on NK cells is likely due to epigenetic modulation on target gene loci. Lunasin represents a different class of immune modulating agent that may augment the therapeutic responses mediated by cytokine-based immunotherapy.

Acquired STAT4 deficiency as a consequence of cancer chemotherapy

AbstractSignal Transducer and Activator of Transcription 4 (STAT4) is a transcription factor that is activated by IL-12 signaling and promotes Th1-cell differentiation and IFN-γ production. Defective IFN-γ production because of STAT4 mRNA and protein deficiency occurs after autologous stem cell transplantation for lymphoma. In the present study, we investigated the mechanisms of STAT4 deficiency in lymphoma patients. The tumor-bearing state is not responsible, because STAT4 levels were not significantly different in PBMCs obtained from healthy control subjects compared with those from lymphoma patients before treatment. STAT4 protein levels were significantly decreased in PBMCs and T cells obtained from lymphoma patients after standard-dose chemotherapy. Furthermore, treatment of control PBMC cultures or a natural killer cell line with chemotherapy drugs in vitro also resulted in reduced STAT4 protein and diminished, IL-12–induced IFN-γ production. Translation of STAT4 protein was not impaired in chemotherapy-treated cells, whereas the STAT4 protein half-life was significantly reduced. Chemotherapy drugs promoted the ubiquitination and proteasomal degradation of STAT4. Treatment with the proteasome inhibitor bortezomib reversed chemotherapy-induced STAT4 deficiency and defective IFN-γ production. We conclude that acquired STAT4 deficiency in lymphoma patients is a consequence of treatment with chemotherapy, results that have important implications for the design of optimal immunotherapy for lymphoma.

CD8+Foxp3+ T cells share developmental and phenotypic features with classical CD4+Foxp3+ regulatory T cells but lack potent suppressive activity

"Suppressor T cells" were historically defined within the CD8(+) T-cell compartment and recent studies have highlighted several naturally occurring CD8(+) Foxp3(-) Treg populations. However, the relevance of CD8(+) Foxp3(+) T cells, which represent a minor population in both thymi and secondary lymphoid organs of nonmanipulated mice, remains unclear. We here demonstrate that de novo Foxp3 induction in peripheral CD8(+) Foxp3(-) T cells is counter-regulated by DC-mediated co-stimulation via CD80/CD86. CD8(+) Foxp3(+) T cells fail to develop in TCR-transgenic mice with Rag1(-/-) background, similar to classical CD4(+) Foxp3(+) Tregs. Notably, both naturally occurring and induced CD8(+) Foxp3(+) T cells express bona fide Treg markers including CD25, GITR, CTLA4 and CD103, and show defective IFN-γ production upon restimulation when compared with their CD8(+) Foxp3(-) counterparts. However, utilizing DEREG transgenic mice for the isolation of Foxp3(+) cells by eGFP reporter expression, we demonstrate that induced CD8(+) Foxp3(+) T cells similar to activated CD8(+) Foxp3(-) T cells only mildly suppress T-cell proliferation and IFN-γ production. We therefore categorize CD8(+) Foxp3(+) T cells as a tightly controlled population sharing certain developmental and phenotypic properties with classical CD4(+) Foxp3(+) Tregs, but lacking potent suppressive activity.

Carry-over effect on IFN-gamma production induced by allergen-specific immunotherapy

Background Allergic rhinitis is characterized by Th2 polarization and defective IFN-γ production. Specific immunotherapy determines an allergen-specific clinical improvement. Aim of the study It was to evaluate whether a course of pre-seasonal SLIT with pollen allergen extract might affect the in vitro IFN-γ production using Dermatophagoides farinae (Df) allergen as stimulus, in patients presenting with rhinitis due to pollen allergy. Methods Thirty-nine AR patients with pollen allergy were included in the study. They assumed pre-seasonal SLIT for 3 months. IFN-γ-specific producing cells were assessed by cytokine ELISPOT before and 3 months after the SLIT course end. Results SLIT provided a significant increase of both pollen-induced and Df-induced IFN-gamma production ( p < 0.001). Conclusions The present study provides evidence that though the defective IFN-γ production is allergen-specific in allergic subjects, the SLIT increasing effects on IFN-γ may be non-specific.

Patients with Allergic Rhinitis Show an Allergen-Specific Interferon-Gamma Defect

Allergic rhinitis (AR) is characterized by Th2 polarized immune response. Consequently, allergic patients have a defect in IFN-γ production. So far, however, all the experimental studies have investigated only the IFN-γ production induced by the causal allergen. The aim of this study is to investigate whether there is a difference in the in vitro IFN-γ production, using different allergens as stimuli, in patients presenting with rhinitis due to pollen allergy. Forty-one AR patients with pollen allergy were enrolled. IFN-γ-specific producing cells were stimulated with PHA, causal pollen, and House Dust Mite (HDM). IFN-γ production was assessed by cytokine ELISPOT. IFN-γ production of peripheral blood mononuclear cells (PBMC) stimulated by specific pollen was significantly lower than IFN-γ production of PBMC stimulated by HDM (p&lt;0.001). IFN-γ production of PBMC stimulated by specific pollen was significantly lower than IFN-γ production of PBMC stimulated by PHA (p&lt;0.001). Moreover, in the HDM-sensitized patients, there seems to be a trend for a different defect. The present study highlights that the defective IFN-γ production is allergen specific and that HDM sensitization may also affects this parameter.

Impaired interferon-γ production as a consequence of STAT4 deficiency after autologous hematopoietic stem cell transplantation for lymphoma

AbstractProduction of interferon γ (IFN-γ) is critical for optimal antitumor immunotherapy in several preclinical animal models. Interleukin-12 (IL-12)–induced IFN-γ production is markedly defective after autologous stem cell transplantation. Quantitative deficiency in CD4 T cells, relative increase in CD25+CD4+ T cells, and bias toward T helper 2 (Th2) differentiation are not the primary mechanisms of defective IFN-γ production. IL-12 receptor β1 (IL-12Rβ1) and IL-12Rβ2 are expressed at equivalent or higher levels on posttransplantation patient peripheral blood mononuclear cells (PBMCs) as compared with control PBMCs. IL-12–induced tyrosine phosphorylation of signal transducer and activator of transcription 4 (STAT4) was undetectable or barely detectable in posttransplantation patient PBMCs, whereas IL-4–induced tyrosine phosphorylation of STAT6 did not differ in posttransplantation patient and control PBMCs. Levels of STAT4 protein were decreased by 97% in posttransplantation patient PBMCs. Levels of STAT4 mRNA were also significantly decreased in posttransplantation patient PBMCs. Incubation with IL-12 and IL-18 in combination partially reversed the defective IFN-γ production by posttransplantation patient PBMCs. IFN-γ production in response to IL-12 plus IL-18 did not require increased expression of STAT4 but was dependent on the activity of p38 mitogen-activated protein kinase (MAPK). These results indicate that defective IFN-γ production is due to an intrinsic deficiency in STAT4 expression by posttransplantation patient lymphocytes and suggest strategies for circumventing this deficiency in cancer immunotherapy.

Cytokine production of stimulated whole blood cultures in rheumatoid arthritis patients receiving short-term infliximab therapy

Patients with rheumatoid arthritis (RA) treated with anti-tumor necrosis factor (TNF) strategies have an increased susceptibility to infections, especially those caused by intracellular pathogens. In this study we assessed the cytokine production capacity in patients with RA and we further investigated whether anti-TNF therapy modulates the production of pro-inflammatory cytokines involved in the resistance against infections. Whole blood cultures from 10 RA patients and 10 healthy controls were stimulated with heat-killed Candida albicans, Salmonella typhimurium, Staphyloccocus aureus, Aspergillus fumigatus or Mycobacterium tuberculosis and production of interleukin (IL)-1β, IL-6, IL-10, interferon (IFN)-γ and TNF-α was measured. Before anti-TNF therapy, whole blood cultures from RA patients released significantly less IFN-γ than healthy controls after stimulation with all tested microorganisms. Short-term anti-TNF therapy did not have an inhibitory effect on the release of the cytokines tested. We conclude that cells of patients with RA have a strongly reduced production capacity of IFN-γ after bacterial challenge. Although short-term therapy with anti-TNF agents did not further decrease the release of other proinflammatory cytokines, the combination of defective IFN-γ production in basal conditions and TNF neutralization during anti-TNF therapy is likely to be responsible for the higher susceptibility to infections in patients with RA.

Changes in intracellular cytokine levels in newborn and adult lymphocytes induced by HSV‐1

Changes in the expression of intracellular interleukin-2 (IL-2), interleukin-4 (IL-4), interferon (IFN)-γ, and tumor necrosis factor (TNF)-α in newborn and adult lymphocytes induced by herpes simplex virus (HSV)-1 were examined. Cord blood mononuclear cells (CBMC) or adult peripheral blood mononuclear cells (PBMC) were infected with HSV-1 and cultured with phorbol 2-myristate 13-acetate (PMA) plus ionomycin in the presence of monensin for 4 hr. Surface antigen and intracellular cytokines were stained simultaneously and analyzed by flow cytometry. The percentage of cells that expressed IL-2, IFN-γ, and TNF-α was significantly increased in HSV-1-infected CD3+, CD4+, CD8+, CD45RA+, and CD45R0+ lymphocytes compared with uninfected lymphocytes from adult PBMC. The percentage of cells that expressed IL-2 and TNF-α was increased significantly in HSV-1-infected CD3+, CD4+, CD8+, and CD45RA+ lymphocytes compared with uninfected lymphocytes from CBMC. IFN-γ was under the detectable level in HSV-1-infected and uninfected lymphocytes from CBMC. Intracellular IL-4 was not detected in HSV-1 or in uninfected lymphocytes from PBMC and CBMC. These results demonstrate that HSV-1 enhances intracellular levels of IL-2, IFN-γ, and TNF-α in adult lymphocytes and defective IFN-γ production in cord blood. J. Med. Virol. 56:145–150, 1998. © 1998 Wiley-Liss, Inc.

T cell-dependent immune response in C1q deficient mice: Defective IFN-γ production by antigen-specific T cells

T cell-dependent immune response in C1q deficient mice: Defective IFN-γ production by antigen-specific T cells

Is deficiency of interferon gamma production by allergen triggered cord blood cells a predictor of atopic eczema?

Peripheral blood mononuclear cell proliferative responses and interferon-gamma production to anti-CD3, cat fur extract, betalactoglobulin and ovalbumin were determined at birth in a group of 34 babies born to families where at least one parent was atopic. The development of atopic eczema with positive allergy skin-prick tests to cows' milk and egg at 1 year of age, where the symptoms improved on an egg and milk-free diet, was significantly associated with raised proliferative responses and defective IFN-gamma production to stimulation with betalactoglobulin with a trend for similar responses to ovalbumin. This was not observed in those who did not develop atopic eczema or those with atopic eczema not associated with foods. Responses to cat fur extract were not significantly different between those with and without atopic eczema. This has important implications for the prediction at birth, not only of the probability of being allergic, but also of the specific allergens which will cause problems. The implementation of specific targeted allergen avoidance during the critical period in infancy, therefore, should be more easily applied and should facilitate attempts to prevent disease.

Relationship between the production of interferon-α/β and interferon-γ during acute toxoplasmosis

Acute toxoplasmosis was induced in mice, and interferon (IFN) production in serum and by spleen cells was evaluated during the infection period. Interferon was characterized by acid-treatment and anti-IFN-alpha/beta neutralization. In order to verify the correlation between the unusual aspects of the IFN production and the induction of immunosuppression, splenocyte mitogen responsiveness was investigated concomitantly to IFN synthesis. The activity of Toxoplasma-induced serum IFN-alpha/beta increased gradually throughout all post-infection days, but IFN-gamma was not detected in the systemic circulation at any time during the infection. It was also observed that IFN-alpha/beta production and the capacity to produce IFN-gamma by spleen cells were closely and inversely correlated. As the infection progressed, more IFN-alpha/beta was produced, and the ability of spleen cells to produce IFN-gamma decreased. The observation that Toxoplasma-infected mice were concomitantly immunosuppressed (as documented by mitogen unresponsiveness and defective IFN-gamma production) in direct correlation to IFN-alpha/beta production, suggests that such IFN-alpha/beta production is an important factor associated with acute toxoplasmosis-induced immunosuppression.

Enhanced interferon- [formula omitted] (IFN- [formula omitted]) and defective IFN-γ production in chronic graft versus host disease: A potential mechanism for immunosuppression

Immunosuppression is a well-characterized consequence of chronic graft-versus-host disease (GVHD). We have previously shown that interferon (IFN) is produced in high levels during acute GVHD. Our objective in this study was to determine if IFN, as a cytokine with known immunosuppressive qualities, could be detected in mice experiencing chronic GVHD-induced immunosuppression. Two different experimental models were used to induce chronic GVHD. The first model involved the injection of parental strain spleen cells into adult F 1 hybrids (AJ → B6AF 1), while the second model utilized GVHD induced across minor histocompatibility barriers (B10·D2 → BALB/c). Results indicated that significant levels of serum IFN- α β are present in mice undergoing chronic GVHD. Spleen cells from chronic GVHD mice were also shown to produce significant levels of IFN- α β upon in vitro culture in medium only. This IFN- α β production was greatly increased when GVHD spleen cells were cultured with either concanavalin A (Con A) or IL-2. In contrast, IFN-γ production was undetectable in these Con A- or IL-2-containing cultures. Additionally, these same spleen cells which produced high levels of IFN- α β were immunosuppressed as measured by mitogen-induced cell proliferation. These results suggest that IFN-γ production is defective in GVHD spleen cells, and that the presence of high IFN- α β production by GVHD mice may contribute to the immunosuppression associated with chronic GVHD.