IFN-gamma Production In Patients Research Articles

NKG2C+NKG2A-CD56dim Subset of NK Cells Show Increased Anti-Leukemia Potential in Presence of CTLA4Ig in-Vitro and Is the Key Determinant of Early Relapse and Long-Term Disease-Free Survival without Gvhd Following CTLA4Ig-DLI Based Haploidentical HCT

<h3>Background & Methods</h3> CTLA4Ig-primed donor lymphocyte infusions (CTLA4Ig-DLI) following haploidentical HCT were earlier shown to be associated with early proliferation of CD56dim NK cells with lower incidence of disease relapse. We prospectively studied the reconstitution of NKG2C+ subset of NK cells in 60 patients with advanced leukemia (myeloid-28/lymphoid-32), along with functional assays. Conditioning was Flu-Bu-Mel followed by CTLA4Ig-DLI on days +7, +21 and +35. GVHD prophylaxis was PTCy followed by cyclosporine for 60 days [BBMT,2019]. All donors and recipients were CMV seropositive. <h3>Results</h3> All patients engrafted with incidences of grade 2-4 acute and chronic GVHD being 12% and 19.8% respectively. NRM was 5% with an overall survival (OS) of 86.5% at a median of 26 months. Disease progression (DP) was 17.6% between days+28 to +144 (median 90 days). This tended to be lower in patients with myeloid leukemia (7.7% vs 26.3% in lymphoid; p=0.06) and NK-KIR B haplotype (10.9% vs 36.6%, p=0.03). Higher CD56dim NK cells at days+30 (135 vs 17 cells/µl), +60 (150 vs 19 cells/µl) and +90 (121 vs 33 cells/µl) (p< 0.01) without any difference in T cell subsets correlated with lack of DP. NKG2C+/NKG2A- subset of CD56dim NK cells were significantly higher at days +30 (5.3% vs 1.0%), +60 (18.5% vs 1.4%) and + 90 (30.6% vs 2.1%) in those without DP (p <0.0001). The surge of NKG2C+ NK cell subset was independent of CMV reactivation and was sustained at a mean value of 27.2%, 32.8%, 34.5% and 28.1% at 6 months, 1, 2 at 3 years, with no DP in survivors beyond 6 months(n = 50). Functionally, NKG2C+ NK cells showed a higher production of IFN-gamma in patients without DP at 30 to 90 days. In normal donors, IFN-gamma production was increased several folds on incubation with CTLA4Ig for 24 hours, supporting the anti-leukemia potential of CTLA4Ig-DLI. On multivariate analysis, NKG2C+/ NKG2A- NK cells had the strongest impact on DP (OR-0.3[0.1-0.6]). Likewise, OS strongly correlated with NKG2C+NK cells (p <0.001). Low levels of NKG2C+NK cells at day+60 and +180 correlated with increased acute and chronic GVHD respectively. They reverted to higher and sustained levels after resolution of chronic GVHD. <h3>Conclusions</h3> In patients with advanced leukemia receiving grafts from CMV seropositive donors, rapid surge of functional NKG2C+NK cell subsets, irrespective of CMV reactivation post-HCT, was observed following CTLA4Ig-DLI. CTLA4Ig-DLI probably promotes an early and sustained NKG2C+NK cell surge with functional anti-leukemia potential demonstrated both in-vitro and in-vivo, an absence of which both numerically and functionally, strongly correlated with early DP. Importantly, NKG2C+NK cells remained persistently elevated, even beyond two years post-transplantation and was the predominant factor associated with disease-free survival without GVHD, on long-term follow-up.

The clinical implication of HAV viral load and HAV proteins-specific IFN-gamma production in patients with acute hepatitis A (P6120)

Abstract The relationship between immune responses to hepatitis A virus (HAV) and the clinical manifestation of HAV infection have been obscured. We investigated the clinical manifestation of acute hepatitis A (AHA) and the relationship between HAV-specific, IFN-gamma-producing T cell response and clinical outcomes in patients with AHA. We analyzed 770 patients diagnosed with AHA for their clinical manifestation. Among them, 128 patients with a prothrombin time (PT) of less than 40% of normal were classified as “severe acute hepatitis (s-AH)”. Others were defined as having “mild acute hepatitis (m-AH)”. In 110 patients, direct ex vivo IFN-gamma ELISpot assays were performed with peripheral blood mononuclear cells using HAV proteins. Mean values of serum HAV RNA at admission were 4.79 log10 copies/ml in s-AH, and 3.30 log10 copies/ml in m-AH (P = 0.003). Viral load showed a significant correlation with ALT (r = 0.417, P = 0.0032) and PT levels (r = -0.453, P = 0.0012). In addition, the mean frequencies of IFN-gamma-producing T cells against VP1-2A (aa 669-782), and 2C(aa 1121-1234) proteins were significantly lower in patients with s-AH, compared to patients with m-AH (3.3 vs. 17.7, P &amp;lt; 0.001; 2.1 vs. 13.3, P &amp;lt; 0.001). In view of viral factors, the load of HAV is closely correlated with liver damage and disease severity in patients with AHA. As a host factor, HAV-specific, IFN-gamma-producing T cells might induce effective viral eradication and result in mild hepatitis

Progressive divergent shifts in natural and induced T-regulatory cells signify the transition from undifferentiated to definitive connective tissue disease

The objectives of the study is to determine clinical signs and distribution of peripheral T-cell subsets, B cells and T regulatory cells in patients with undifferentiated connective tissue disease (UCTD) and during the development toward well-established connective tissue diseases (CTD). The methods include 46 patients with UCTD were followed and investigated for differentiation into defined CTDs for 2 years. Cell subsets were determined on the basis of cell surface markers, intracellular cytokine production by flow cytometry and serum cytokine levels by ELISA. The results are as follows: 45.6% of UCTD patients developed into a defined CTD. The number and percentage of activated T cells, memory T cells and NKT cells were increased in patients compared with controls. In addition, in patients with UCTD, the percentage of CD4+/IFN gamma+ T(h)1 was significantly higher compared with controls and further increased in patients that developed CTDs. The percentage and absolute number of CD4+CD25+Foxp3+ regulatory T cells (Tregs) were diminished in UCTD patients compared with healthy controls, while the number of CD4+/IL-10+ Tregs increased. The conclusions are Overproduction of IFN gamma and the decrease of natural (Foxp3+) Tregs seem to be characteristic features of UCTD patients. The increased IL-10 production of CD4+ T cells might be a compensatory suppressive mechanism; however, it is probably not able to balance the overproduction of IFN gamma and the observed decrease of Foxp3+ Tregs. The shift toward T(h)1 with increased IFN gamma production in patients with UCTD combined with the degree of immunoregulatory disturbances characterized by the progressive divergent shifts in natural and induced T-regulatory cell populations signify the transition from undifferentiated to definitive CTD.

NK cell activity in tuberculosis is associated with impaired CD11a and ICAM‐1 expression: a regulatory role of monocytes in NK activation

Although the role of natural killer (NK) cells in mycobacterial infections is unclear, it has been postulated that they contribute to protective immunity through the production of interferon (IFN)-gamma. In this study, we evaluate the effect of interleukin (IL)-10, IL-15 and IL-18 on NK lytic activity through the expression of CD16, CD11a and CD69 molecules and the induction of IFN-gamma production in patients with tuberculosis (TB) and healthy individuals (N). Our results showed an impairment of NK lytic activity and a gradual down-regulation of costimulatory and adhesion molecules on NK cells which were dependent on the severity of the disease. NK lytic activity was increased by exogenous IL-15 and IL-18 in both TB and N, and by neutralization of endogenous IL-10 only in TB; IL-15 and IL-18 increased CD69 receptor expression, while anti-IL-10 up-regulated CD16 and CD11a expression in TB. Mycobacterium tuberculosis reduced the number of intracellular adhesion molecule (ICAM)-1(+) CD14(+) cells, but in the presence of IL-15, IL-18 and anti-IL-10 its expression was up-regulated. In cells from TB patients, the observed effects of IL-15 and IL-18 on NK function were not dependent on IL-10 modulation of the surface expression of activator/adhesion molecules. In the absence of monocytes, IL-10 activated NK cells, suggesting an indirect effect on their function. Furthermore, in TB patients the depletion of monocytes increased the production of IFN-gamma by NK cells. Therefore, monocytes from TB patients regulated the NK function involving IL-10 which, through an indirect mechanism, led to the down-regulation of costimulatory/adhesion molecules and/or IFN-gamma production.

Production of IFN-gamma and IL-12 by peripheral whole blood is maintained in hepatitis C virus patients with persistently normal alanine transferase activity; A preliminary report.

The current study was designed to investigate the immune status in hepatitis C virus (HCV) patients with persistently normal alanine transferase activity (ALT) (patients with normal alanine transferase). For this purpose, serum levels and lipopolysaccharide (LPS)-induced IFN-gamma, IL12 p70, IL12 p40 and IL-10 as well as NK cell activity were assayed in six patients with normal ALT, 22 HCV-infected individuals with chronic hepatitis (CH), 13 cases of liver cirrhosis (LC) and 26 age-matched controls. Cytokine production was assayed with the whole blood induction method. IFN-gamma levels were significantly lower in patients with HCV-infected chronic hepatitis and liver cirrhosis than in controls ( [Formula: see text], [Formula: see text] and [Formula: see text] pg/ml, respectively, [Formula: see text] ). However, IFN-gamma production in those individuals with normal ALT was not reduced ( [Formula: see text] pg/ml). Although variation was observed, four of the six patients showed moderate to strong IFN-gamma production. No intergroup differences were observed for IL12 p70, IL12 p40 and IL-10 production and NK cell activity. Our results suggest that preserved IFN-gamma production in patients with normal ALT, in contrast to the reduction in chronic hepatitis and liver cirrhosis, may be related to a slow rate of disease progression.

Selective insufficiency of IFN-gamma secretion in patients with hyper-IgE syndrome.

Hyper-immunoglobulin E (IgE) syndrome is a complex immune deficiency characterized by chronic eczematous dermatitis, recurrent staphylococcal infections, pneumatoceles, reduced neutrophil chemotaxis, and variably impaired T cell function. Although decreased interferon-gamma (IFN-gamma) production in patients with hyper-IgE syndrome is pointed out and known as a cause of reduced neutrophil chemotaxis, precise mechanism of their inadequate production of IFN-gamma remains unknown. To elucidate the pathogenesis of the defective production of IFN-gamma in patients with hyper-IgE syndrome, we assessed the in vitro production and secretion of IFN-gamma by peripheral blood mononuclear cells (PBMCs) from patients with hyper-IgE syndrome. Chemotaxis of neutrophils, mRNA levels of several cytokines, intracellular production and extracellular secretion of IFN-gamma, interleukin-2 (IL-2), and IL-4 by PBMCs from three patients with hyper-IgE syndrome were determined. The transcription of IFN-gamma mRNA and the production of its protein molecules progressed normally. However, selective insufficiency in the secretion of IFN-gamma molecules was found in patients with hyper-IgE syndrome. Confocal laser scanning microscopy clearly demonstrated the accumulation of IFN-gamma in patients with hyper-IgE syndrome. We demonstrated that there was a selective insufficiency in the secretion of IFN-gamma in patients with hyper-IgE syndrome. We hope that this fact would offer a new paradigm for understanding this disease.

Profiles of IFN-gamma and its regulatory cytokines (IL-12, IL-18 and IL-10) in peripheral blood mononuclear cells from patients with multidrug-resistant tuberculosis.

This study investigated the profiles of IFN-gamma and its regulatory cytokines (IL-12, IL-18 and IL-10) in response to a purified protein derivative (PPD) antigen in peripheral blood mononuclear cells (PBMC) from 18 HIV-negative patients with multidrug-resistant tuberculosis (MDRTB), and compared them with those from 19 healthy tuberculin reactors (HTR). ELISA results showed that following stimulation with PPD, IFN-gamma production was significantly reduced, whereas production of both IL-18 and IL-10 was significantly elevated in MDRTB patients compared with HTR. Three out of 18 patients with MDRTB of greater than 4 years duration showed significantly elevated IL-12 p70 production, induced by in vitro PPD stimulation of their PBMC, when compared with data from HTR. However, when taken as a group, MDRTB patients were similar to HTR in their IL-12 p70-producing capacity. IL-12 p70 protein paralleled IL-12 p40 protein expression. In addition, the production of IL-12 p40 was significantly correlated with IL-10 in all patients, but was not correlated with IFN-gamma. Neutralization of IL-10 increased IL-12 p40 about twofold, but did not significantly alter IFN-gamma induction in MDRTB. IFN-gamma in MDRTB was highly correlated with lymphoproliferation and CD4 counts, but was not correlated with IL-12, IL-18 or IL-10 production. Our findings suggest that patients with MDRTB have dysregulated IL-12, IL-18 and IL-10 production during Mycobacterium tuberculosis infection, and the cytokine profiles are similar to those in patients with drug-sensitive advanced TB previously reported in the literature. In addition, IL-10 may not have a dominant role in defective IFN-gamma production in patients with MDRTB.

Production of IFN-gamma is impaired in patients with paracoccidioidomycosis during active disease and is restored after clinical remission.

Cellular immunity is usually suppressed during paracoccidioidomycosis (PCM) and is restored after treatment. In this study we evaluated the induction of a type 1 (interferon gamma (IFN-gamma)), a type 2 (interleukin (IL)-10) and a primarily macrophage derived cytokine (tumor necrosis factor (TNF)-alpha) in peripheral blood mononuclear cells (PBMC) from patients with PCM. Eight male patients with active PCM, nine male patients with clinical remission of the disease and 10 healthy control subjects were enrolled in the study. Cytokines were induced with non-specific stimuli --phytohaemagglutin (PHA) (induces IL-10 and IFN-gamma), Lipopolysaccharide (induces TNF-alpha)--and Paracoccidioides brasiliensis antigen (PbAg) (induces IL-10, IFN-gamma and TNF-alpha). Induction of IFN-gamma with PHA differed among the three groups (P < 0.01; Kruskal-Wallis test) and with PbAg was lower in patients with active disease compared to those in clinical remission (P = 0.05; Mann-Whitney). Induction of IL-10 and of TNF-alpha was similar in the three groups. The suppressed production of IFN-gamma in patients with active disease may underscore the cellular immune deficiency seen in these patients.

Decreased interferon gamma and increased interleukin-4 production in atopic dermatitis promotes IgE synthesis

The mechanism(s) responsible for increased IgE synthesis in atopic dermatitis (AD) are unknown, but they may be related to either decreased interferon gamma (IFN-gamma) and/or increased interleukin (IL)-4 production. In this study we examined peripheral blood mononuclear cells (PBMCs) from 21 patients with AD, six patients with psoriasis, and 22 nonatopic healthy controls for IFN-gamma and IL-4 production after stimulation with concanavalin A (Con A). The Con A-induced proliferative response of AD PBMCs was similar to the response of healthy controls (p = 0.9). After mitogen stimulation, however, AD culture supernatants contained significantly less IFN-gamma (p = 0.001) but increased IL-4 (p = 0.001) compared with supernatants from nonatopic controls. In contrast, PBMCs from patients with psoriasis produced normal levels of IFN-gamma and IL-4 in vitro. Since IL-4 is known to decrease IFN-gamma synthesis, we examined the effect of neutralizing anti-IL-4 on IFN-gamma production. Anti-IL-4 significantly increased IFN-gamma production in patients with AD (p = 0.008) and nonatopic controls (p = 0.02) but did not normalize IFN-gamma production by AD PBMCs. Supernatants from AD PBMCs, but not supernatants from nonatopic PBMCs, induced IgE synthesis in PBMCs from nonatopic donors (p = 0.02). When an anti-IFN-gamma receptor antibody, which blocks cellular binding of IFN-gamma, was added to supernatants from nonatopic controls their capacity to induce IgE synthesis was significantly greater (p = 0.03). These results demonstrate an imbalance of IL-4 and IFN-gamma production, which may contribute to increased IgE synthesis in AD.

Decrease in interferon-? production by peripheral blood mononuclear cells in patients with uterine cervical cancer

The interferon-gamma (IFN-gamma) productivity of peripheral blood mononuclear cells (PBMCs) was examined in 30 patients with uterine cervical cancer. The patients under 50 years of age had decreased IFN-gamma production compared with the age-matched controls. The IFN-gamma productivity in the patients over 50 years of age was decreased as well as in the age-matched controls. The proportion of monocytes in PBMCs did not correlate with the IFN-gamma productivity. The prostaglandin E2 (PGE2) productivity of PBMCs increased with the progress of cancer. PGE2 inhibited the IFN-gamma production by PBMCs, and the sensitivity of PBMCs to PGE2 was increased in the patients and controls over 60 years of age. The addition of indomethacin resulted in an increase in IFN-gamma production by PBMCs. These results suggest that the increased production of PGE2 and/or increased sensitivity to PGE2 are responsible for the decreased IFN-gamma production in patients with cervical cancer.

Gamma-Interferon Production in Response to Hepatitis B Core Protein and Its Synthetic Peptides in Patients with Chronic Hepatitis B Virus Infection

The nucleocapsid of hepatitis B virus (HBV) is an efficient immunogen in activating T cells to produce interferon-gamma (IFN-gamma) in patients with chronic HBV infection. We investigated hepatitis B core antigen (HBcAg)-specific T cell recognition, which seems to be implicated in the pathogenesis of chronic liver disease. IFN-gamma production by peripheral-blood mononuclear cells of patients with chronic HBV infection [25 patients with chronic active hepatitis (CAH) and 14 asymptomatic carriers of HBV (ASCs)] was measured by an enzyme-linked immunosorbent assay. P19 polypeptide, which is derived from recombinant HBcAg particle (rHBcAg), increased IFN-gamma production in patients with CAH, but its effect was weaker than that of rHBcAg. P19 had no stimulating effect on T cells from ASCs. The fine specificity of T cell recognition of HBcAg was examined using 8 kinds of synthetic peptides. T cells from the patients who responded against P19 polypeptide recognized the sites within the common sequences of HBcAg and HBeAg (p72-90, P90-99, P108-122 and P126-146). These results suggest that HBcAg and P19 are cross-reactive at the T cell level, and that these T cells recognize the sites within the common sequences of HBcAg and HBeAg in HBV-infected patients.