Lower Cytokine Concentrations Research Articles

TNFR1 Absence Is Not Crucial for Different Types of Cell Reaction to TNF: A Study of the TNFR1-Knockout Cell Model.

One of the mechanisms regulating the biological activity of tumor necrosis factor (TNF) in cells is the co-expression of TNFR1/TNFR2 receptors. A model with a differential level of receptor expression is required to evaluate the contribution of these mechanisms. The development of a cellular model to compare the effects of TNF on cells depending on the presence of both receptors and TNFR2 alone. TNFR1 absence modifications of ZR-75/1 and K-562 cell lines were obtained by TNFR1 knockout. The presence of deletions was confirmed by Sanger sequencing, and the absence of cell membrane receptor expression was confirmed by flow cytometry. The dose-dependent effect of TNF on intact and knockout cells was comparatively evaluated by the effect on the cell cycle, the type of cell death, and the profile of expressed genes. Knockout of TNFR1 resulted in a redistribution of TNFR2 receptors with an increased proportion of TNFR2+ cells in both lines and a multidirectional change in the density of expression in the lines (increased in K562 and decreased in ZR75/1). The presence of a large number of cells with high TNFR2 density in the absence of TNFR1 in the K562 cells was associated with greater sensitivity to TNF-stimulating doses and increased proliferation but did not result in a significant change in cell death parameters. A twofold increase in TNFR2+ cell distribution in this cell line at a reduced expression density in ZR75/1 cells was associated with a change in sensitivity to low cytokine concentrations in terms of proliferation; an overall increase in cell death, most pronounced at standard stimulating concentrations; and increased expression of the lymphocyte-activation gene groups, host-pathogen interaction, and innate immunity. The absence of TNFR1 leads to different variants of compensatory redistribution of TNFR2 in cellular models, which affects the type of cell response and the threshold level of sensitivity. The directionality of cytokine action modulation and sensitivity to TNF levels depends not only on the fraction of cells expressing TNFR2 but also on the density of expression.

Immunological Characterization of Chronic Nonbacterial Osteomyelitis (CNO) in Adults: A Cross-Sectional Exploratory Study.

Chronic nonbacterial osteomyelitis (CNO) is a rare disease spectrum affecting children and adults. Adult CNO may occur as isolated bone inflammation, or with a broad range of extraskeletal features. CNO pathophysiology, including the key drivers of inflammation, remains largely unknown. For pediatric CNO, a role for pro-inflammatory cytokine dysregulation has been proposed, but studies in adults are scarce. We therefore provide immunological characterization of adult CNO. Cross-sectional study in our referral center including adult CNO patients (n = 172) and healthy controls (n = 65). Inflammation parameters and systemic inflammatory based scores(SIBS, including neutrophil/lymphocyte ratio [NLR] and systemic immune inflammation index [SII]) were compared between groups. Cytokine expression was explored with electrochemiluminescent immunoassays in 33 patients, eight healthy controls and 21 osteoporosis patients. Routine inflammation markers were higher in patients than in controls, but generally remained within reference range. Systemic inflammation was more pronounced in patients with additional vertebral involvement as compared to those osteitis in the anterior chest wall alone, in patients with comorbid pustulosis palmoplantaris or psoriasis, and in patients with strongly rather than moderately increased lesional uptake on nuclear imaging. SII was elevated in CNO patients too, but NLR was not. Cytokine expression was generally nondifferential between patients and both control groups, and patients displayed low absolute concentrations of pro-inflammatory cytokines. In this adult CNO cohort, systemic inflammation was generally subtle, but more pronounced in patients with vertebral lesions, associated skin disease, and strongly increased uptake on nuclear imaging. SII was increased in patients compared to healthy controls. Contrasting pediatric studies, we found no increased expression of the pro-inflammatory cytokines that have been proposed to drive the inflammatory cascade, like interleukin-6, -8, and -17 (IL-6, IL-8, and IL-17), and tumor necrosis α (TNF-α). Further studies are needed to evaluate the use of SII in diagnosis and monitoring of CNO, and elucidate the role of cytokine dysregulation in adult disease. © 2023 The Authors. JBMR Plus published by Wiley Periodicals LLC. on behalf of American Society for Bone and Mineral Research.

A cytokine-induced spheroid-based in vitro model for studying osteoarthritis pathogenesis.

Given the lack of in vitro models faithfully reproducing the osteoarthritis (OA) disease on-set, this work aimed at manufacturing a reliable and predictive in vitro cytokine-based Articular Cartilage (AC) model to study OA progression. Cell spheroids of primary human fetal chondrocytes (FCs) and h-TERT mesenchymal stem cells differentiated chondrocytes (Y201-C) were analysed in terms of growth kinetics, cells proliferation and apoptosis over 10 days of culture, in healthy condition or in presence of cytokines (interleukin-1ß, -6 and TNF-α). Then, the spheroids were assembled into chondrospheres using a bottom-up strategy, to obtain an in vitro cytokines-induced OA model. The resulting chondrospheres were evaluated for gene expression and anabolic ECM proteins. Compared to the healthy environment, the simulated OA environment induced chondrocyte hyperproliferation and apoptotic pathway, decreased expression of anabolic ECM proteins, and diminished biosynthetic activity, resembling features of early-stage OA. These characteristics were observed for both Y201-C and HC at high and low concentrations of cytokines. Both HC and Y201-C demonstrated the suitability for the manufacturing of a scaffold-free in vitro OA model to facilitate studies into OA pathogenesis and therapeutic strategies. Our approach provides a faithful reproduction of early-stage osteoarthritis, demonstrating the ability of obtaining different disease severity by tuning the concentration of OA-related cytokines. Given the advantages in easy access and more reproducible performance, Y201-C may represent a more favourable source of chondrocytes for establishing more standardized protocols to obtain OA models.

Well Array Microelectrode Electrochemical Sensor for High Sensitivity Cytokine Detection

Introduction The activity of cytokines in the human body can be associated with certain diseases such as cancer, autoimmune, and infectious disease [1]. Recent studies found that it was possible to non-invasively detect certain cytokines such as Il6 through human saliva and sweat [2]. However, detection of cytokine in such bodily fluid poses a challenge due to very low cytokine concentration, ranging from fg/ml to pg/ml. Thus, signal amplification is necessary to achieve high sensitivity in cytokine detection. In this study, we propose a microfabricated electrochemical sensor with array of platinum micro-well as working electrode (Figure 1a and b). The charge transfer current density is compared to commonly used interdigitated electrode configuration (Figure 1c). A COMSOL model was developed to analytically compare charge transfer current for both electrode type. The electrodes were then compared experimentally through square wave voltammetry measurement. Method The microelectrode chips consist of platinum electrode and silicon nitride as insulation layer was fabricated using standard microfabrication techniques. The well array electrode consists of 1024 (32x32) array of 50 μm diameter well (counter electrode) and 2 μm ledge width for ring working electrode. The interdigitated electrode has 30 μm width and 1000 μm length with 10 parallel fingers. Square wave voltammetry measurement was carried using 20 mM Potassium Ferricyanide/ Ferrocyanide as redox probe, dissolved in 1x PBS and Ag/AgCl used as a reference electrode. Scanning range was set between 0 to 0.5 V with 5 mV step size, 25 mV pulse amplitude, and scanning 10 Hz frequency. Results and Conclusions Based on experimental results the well microelectrode configuration has 187% higher current density compared to its interdigitated electrode counterpart. The low current amplitude issue of well microelectrode is solved by having an array of 1024 well microelectrode (32x32) which amplify the signal from nanoampere to microampere scale.

Korelasi Kadar Serum Vitamin D terhadap Derajat Keparahan Penyakit Infeksi Covid-19: Sebuah Literatur Review

In recent years, infectious diseases caused by viruses have become one of the biggest global health problems, including coronavirus disease 2019 (Covid-19) infection. Covid-19 is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which can induce immune responses and inflammatory reactions, thereby leading to the damage of tissues. Hyperinflammation due to cytokine storm in infected patients increases the risk of severe infection and death. Vitamin D can also reduce the severity of infections because it acts as an immunomodulator. Therefore, this study aimed to determine the immunomodulatory effect of vitamin D on Covid-19 infections. A total of nine articles were collected using literature searching on several databases, such as PUBMED and Science Direct with the keywords of “Covid-19”, “immunomodulator”, and “vitamin D”. Five of them were observational or retrospective studies, which determined the correlation between serum concentration of vitamin D and Covid-19 infection severity. Meanwhile, another four articles were random clinical trials testing the effect of the drugs’s supplementation on infected people. The results showed that high levels of serum vitamin D can cause low concentrations of pro-inflammatory cytokines, which reduces the incidence of acute respiratory infections and the severity of the viral infection. Furthermore, vitamin D, specifically D3, has the potential to act as an immunomodulator in Covid-19 by reducing pro-inflammatory cytokines, thereby reducing the risk of severity.

Metabolic Reprograming via Deletion of CISH in Human iPSC-Derived NK Cells Promotes InVivo Persistence and Enhances Anti-tumor Activity.

Cytokine-inducible SH2-containing protein (CIS; encoded by the gene CISH) is a key negative regulator of interleukin-15 (IL-15) signaling in natural killer (NK) cells. Here, we develop human CISH-knockout (CISH-/-) NK cells using an induced pluripotent stem cell-derived NK cell (iPSC-NK cell) platform. CISH-/- iPSC-NK cells demonstrate increased IL-15-mediated JAK-STAT signaling activity. Consequently, CISH-/- iPSC-NK cells exhibit improved expansion and increased cytotoxic activity against multiple tumor cell lines when maintained at low cytokine concentrations. CISH-/- iPSC-NK cells display significantly increased invivo persistence and inhibition of tumor progression in a leukemia xenograft model. Mechanistically, CISH-/- iPSC-NK cells display improved metabolic fitness characterized by increased basal glycolysis, glycolytic capacity, maximal mitochondrial respiration, ATP-linked respiration, and spare respiration capacity mediated by mammalian target of rapamycin (mTOR) signaling that directly contributes to enhanced NK cell function. Together, these studies demonstrate that CIS plays a key role to regulate human NK cell metabolic activity and thereby modulate anti-tumor activity.

Metabolic Reprograming Due to Deletion of <i>CISH</i> in Human Natural Killer Cells Promotes <i>in vivo</i> Persistence and Enhances Anti-Tumor Activity

Cytokine-inducible SH2-containing protein (CIS, encoded by the gene CISH) is a key negative regulator of IL-15 signaling in natural killer (NK) cells. Here, we developed human CISH knockout (CISH-/-) NK cells using an induced pluripotent stem cell-derived NK cell (iPSC-NK cell) platform. CISH-/- iPSC-derived NK cells demonstrate increased IL-15 mediated JAK-STAT pathway signaling activity. Consequently CISH-/- iPSC-NK cells exhibit improved expansion and increased cytotoxic activity against multiple tumor cell lines when maintained at low cytokine concentration. CISH-/- iPSC-NK cells display significantly increased in vivo persistence and inhibition of tumor progression in a leukemia xenograft model. Mechanistically, CISH-/- iPSC-NK cells display improved metabolic fitness characterized by a significant increase in basal glycolysis, glycolytic capacity, maximal mitochondrial respiration, ATP-linked respiration and spare capacity respiration which are mediated by mTOR signaling and directly contribute to enhanced NK cell function. In addition, we demonstrate that deletion of CISH in human NK cells increases single-cell polyfunctionality by secreting multiple effector cytokines, a profile has been proven to be positively associated with clinical outcome of CAR-T cells. Together, these studies demonstrate CIS plays a key role to regulate human NK cell metabolic activity and deletion of CIS leads to enhanced anti-tumor activity.

ENVIRONMENTAL OPTIMIZATION ENABLES MAINTENANCE OF QUIESCENT HEMATOPOIETIC STEM CELLS EX VIVO

Hematopoietic stem cells (HSCs) maintain lifelong hematopoiesis by remaining quiescent in the bone marrow niche. Recapitulation of a quiescent state in culture has not been achieved, as cells rapidly proliferate and differentiate in vitro. After exhaustive analysis of different environmental factor combinations and concentrations as a way to mimic physiological conditions, we were able to maintain engraftable quiescent HSCs for 1 month in culture under very low cytokine concentrations, hypoxia, and very high fatty acid levels. Exogenous fatty acids were required likely due to suppression of intrinsic fatty acid synthesis by hypoxia and low cytokine conditions. By contrast, high cytokine concentrations or normoxia induced HSC proliferation and differentiation. Our novel culture system provides a means to evaluate properties of steady state HSCs and test effects of defined factors in vitro under near-physiological conditions.

Environmental Optimization Enables Maintenance of Quiescent Hematopoietic Stem Cells Ex Vivo

Hematopoietic stem cells (HSCs) maintain lifelong hematopoiesis by remaining quiescent in the bone marrow niche. Recapitulation of a quiescent state in culture has not been achieved, as cells rapidly proliferate and differentiate invitro. After exhaustive analysis of different environmental factor combinations and concentrations as a way to mimic physiological conditions, we were able to maintain engraftable quiescent HSCs for 1month in culture under very low cytokine concentrations, hypoxia, and very high fatty acid levels. Exogenous fatty acids were required likely due to suppression of intrinsic fatty acid synthesis by hypoxia and low cytokine conditions. By contrast, high cytokine concentrations or normoxia induced HSC proliferation and differentiation. Our culture system provides a means to evaluate properties of steady-state HSCs and test effects of defined factors invitro under near-physiological conditions.

Inflammatory Cytokine Profiles of Semen Influence Cytokine Responses of Cervicovaginal Epithelial Cells.

Genital inflammatory cytokine responses increase HIV risk. Since male partner semen is a complex mixture of immune-modulatory prostaglandins and cytokines, we hypothesized that exposure to semen may influence genital inflammation in women. Here, we investigated cytokine response kinetics of cervical cells following stimulation with seminal plasma from HIV-negative and HIV-positive men characterized as having low or high concentrations of inflammatory cytokines. Irrespective of the HIV status or semen cytokine profile, in vitro stimulation of cervical cells with seminal plasma resulted in significantly elevated concentrations of secreted IL-6, IL-8, TNF-β, MCP-1, GM-CSF, and VEGF within 8 h of stimulation, which tended to decline by 24 h, although this was only significant for TNF-β. Consistent with this, cervical cells responded to seminal plasma with increases in IL-8 and IL-1β mRNA expression of 10-fold. These findings suggest that the impact of semen on local female genital cytokines is likely transient. Although these findings suggest that the impact of semen on local female genital cytokines may not be sustained long-term, this heightened genital inflammation may have implications for HIV risk in women.

Robust erythroid differentiation system for rhesus hematopoietic progenitor cells allowing preclinical screening of genetic treatment strategies for the hemoglobinopathies

γ-globin expression can be induced by various gene modification strategies, which could be beneficial for hemoglobin (Hb) disorders. To translate promising ideas into clinics, large animal models have proven valuable to evaluate safety and efficacy of the approaches; however, in vitro erythroid differentiation methods have not been established to determine whether they can be modeled in nonhuman primates. We optimized erythroid differentiation culture to produce high-level adult Hb from rhesus hematopoietic progenitor cells by using low (LC) or high cytokine concentration (HC) protocols with or without feeder cells. In addition, we established rhesus globin protein analysis using reverse-phase high performance liquid chromatography and mass spectrometry. Robust adult Hb production at protein levels was observed in the LC protocol when feeder cells were used, whereas the HC protocol resulted in higher baseline fetal Hb levels (P < 0.01). We then compared lentiviral transduction of rhesus cells between serum-containing LC media and serum-free StemSpan-based differentiation media, revealing 100-fold more efficient transduction in serum-free differentiation media (P < 0.01). Finally, rhesus CD34+ cells were transduced with lentiviral vectors encoding artificial zinc finger proteins (ZF-Ldb1), which can reactivate γ-globin expression via tethering the transcriptional co-regulator Ldb1 to γ-globin promoters, and were differentiated in the optimized erythroid differentiation method. This resulted in marked increases of γ-globin levels compared with control groups (P < 0.01). In conclusion, we developed an efficient rhesus erythroid differentiation protocol from hematopoietic progenitor cells with low fetal and high adult Hb production. Further studies are warranted to optimize gene modification and transplantation of rhesus hematopoietic progenitor cells.

Chondrocyte and mesenchymal stem cell derived engineered cartilage exhibits differential sensitivity to pro-inflammatory cytokines.

Tissue engineering is a promising approach for the repair of articular cartilage defects, with engineered constructs emerging that match native tissue properties. However, the inflammatory environment of the damaged joint might compromise outcomes, and this may be impacted by the choice of cell source in terms of their ability to operate anabolically in an inflamed environment. Here, we compared the response of engineered cartilage derived from native chondrocytes and mesenchymal stem cells (MSCs) to challenge by TNFα and IL-1β in order to determine if either cell type possessed an inherent advantage. Compositional (extracellular matrix) and functional (mechanical) characteristics, as well as the release of catabolic mediators (matrix metalloproteinases [MMPs], nitric oxide [NO]) were assessed to determine cell- and tissue-level changes following exposure to IL-1β or TNF-α. Results demonstrated that MSC-derived constructs were more sensitive to inflammatory mediators than chondrocyte-derived constructs, exhibiting a greater loss of proteoglycans and functional properties at lower cytokine concentrations. While MSCs and chondrocytes both have the capacity to form functional engineered cartilage in vitro, this study suggests that the presence of an inflammatory environment is more likely to impair the in vivo success of MSC-derived cartilage repair. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:2901-2910, 2018.

Neural pathways involved in infection-induced inflammation: recent insights and clinical implications.

Although the immune and nervous systems have long been considered independent biological systems, they turn out to mingle and interact extensively. The present review summarizes recent insights into the neural pathways activated by and involved in infection-induced inflammation and discusses potential clinical applications. The simplest activation concerns a reflex action within C-fibers leading to neurogenic inflammation. Low concentrations of pro-inflammatory cytokines or bacterial fragments may also act on these afferent nerve fibers to signal the central nervous system and bring about early fever, hyperalgesia and sickness behavior. In the brain, the preoptic area and the paraventricular hypothalamus are part of a neuronal network mediating sympathetic activation underlying fever while brainstem circuits play a role in the reduction of food intake after systemic exposure to bacterial fragments. A vagally-mediated anti-inflammatory reflex mechanism has been proposed and, in turn, questioned because the major immune organs driving inflammation, such as the spleen, are not innervated by vagal efferent fibers. On the contrary, sympathetic nerves do innervate these organs and modulate immune cell responses, production of inflammatory mediators and bacterial dissemination. Noradrenaline, which is both released by these fibers and often administered during sepsis, along with adrenaline, may exert pro-inflammatory actions through the stimulation of β1 adrenergic receptors, as antagonists of this receptor have been shown to exert anti-inflammatory effects in experimental sepsis.

Synovial Fluid Profile at the Time of Anterior Cruciate Ligament Reconstruction and Its Association With Cartilage Matrix Composition 3 Years After Surgery

Background: Anterior cruciate ligament tears can lead to posttraumatic osteoarthritis. In addition to biomechanical factors, changes in biochemical profiles within the knee joint after injury and anterior cruciate ligament reconstruction (ACLR) may play a role in accelerating joint degeneration. Hypothesis/Purpose: It was hypothesized that cartilage matrix composition after ACLR is associated with the degree of inflammatory response after initial injury. This study evaluated the association between the inflammatory response after injury—as indicated by cytokine, metalloproteinase, and cartilage degradation marker concentrations in synovial fluid—and articular cartilage degeneration, measured by T1ρ and T2 quantitative magnetic resonance imaging up to 3 years after ACLR. Study Design: Cohort study; Level of evidence, 2. Methods: Twenty-six subjects from a longitudinal cohort study who underwent ACLR at a mean 8.5 weeks after injury (range, 4-19 weeks) had synovial fluid aspirated at the time of surgery. Immunoassays quantified biomarkers in synovial fluid. T1ρ and T2 values of articular cartilage were calculated with magnetic resonance scans acquired prior to surgery and at 6 months and 1, 2, and 3 years after surgery. Pearson correlation coefficients were calculated among the various biomarkers. K-means clustering was used to group subjects with similar biomarker profiles. Generalized estimating equations were used to find the overall differences in T1ρ and T2 values throughout these first 3 years after surgery between the clusters while controlling for other factors. Results: Significant and strong correlations were observed between several cytokines (interleukin 6 [IL-6], IL-8, IL-10, and tumor necrosis factor α) and 2 matrix metalloproteinases (MMP-1 and MMP-3) (P < .05). Moderate correlations were found among combinations of C-terminal crosslinked telopeptide type II collagen, N-terminal telopeptide, cartilage oligomeric matrix protein, and sulfated glycosaminoglycan (P < .05). Two clusters were generated, 1 of which was characterized by lower concentrations of cytokines (IL-6, IL-8, IL-10, tumor necrosis factor α) and MMP-1 and MMP-3 and higher sulfated glycosaminoglycan. This cluster was associated with significantly higher T1ρ and T2 values in the medial tibial and patellar cartilage over the first 3 years after ACLR. Conclusion: At the time of ACLR surgery, profiles of synovial fluid inflammatory cytokines, degradative enzymes, and cartilage breakdown products show promise as predictors of abnormal cartilage tissue integrity (increased T1ρ and T2 values) throughout the first 3 years after surgery. Clinical Relevance: The results suggest an intricate relationship between inflammation and cartilage turnover, which can in turn be influenced by timing after injury and patient factors.

Spheroid Coculture of Hematopoietic Stem/Progenitor Cells and Monolayer Expanded Mesenchymal Stem/Stromal Cells in Polydimethylsiloxane Microwells Modestly Improves In Vitro Hematopoietic Stem/Progenitor Cell Expansion.

While two-dimensional (2D) monolayers of mesenchymal stem/stromal cells (MSCs) have been shown to enhance hematopoietic stem/progenitor cell (HSPC) expansion in vitro, expanded cells do not engraft long term in human recipients. This outcome is attributed to the failure of 2D culture to recapitulate the bone marrow (BM) niche signal milieu. Herein, we evaluated the capacity of a novel three-dimensional (3D) coculture system to support HSPC expansion in vitro. A high-throughput polydimethylsiloxane (PDMS) microwell platform was used to manufacture thousands of uniform 3D multicellular coculture spheroids. Relative gene expression in 3D spheroid versus 2D adherent BM-derived MSC cultures was characterized and compared with literature reports. We evaluated coculture spheroids, each containing 25–400 MSCs and 10 umbilical cord blood (CB)-derived CD34+ progenitor cells. At low exogenous cytokine concentrations, 2D and 3D MSC coculture modestly improved overall hematopoietic cell and CD34+ cell expansion outcomes. By contrast, a substantial increase in CD34+CD38− cell yield was observed in PDMS microwell cultures, regardless of the presence or absence of MSCs. This outcome indicated that CD34+CD38− cell culture yield could be increased using the microwell platform alone, even without MSC coculture support. We found that the increase in CD34+CD38− cell yield observed in PDMS microwell cultures did not translate to enhanced engraftment in NOD/SCID gamma (NSG) mice or a modification in the relative human hematopoietic lineages established in engrafted mice. In summary, there was no statistical difference in CD34+ cell yield from 2D or 3D cocultures, and MSC coculture support provided only modest benefit in either geometry. While the high-throughput 3D microwell platform may provide a useful model system for studying cells in coculture, further optimization will be required to generate HSPC yields suitable for use in clinical applications.

Leukotriene A4 Hydrolase Genotype and HIV Infection Influence Intracerebral Inflammation and Survival From Tuberculous Meningitis.

Background.Tuberculous meningitis (TBM) is the most devastating form of tuberculosis, yet very little is known about the pathophysiology. We hypothesized that the genotype of leukotriene A4 hydrolase (encoded by LTA4H), which determines inflammatory eicosanoid expression, influences intracerebral inflammation, and predicts survival from TBM.Methods.We characterized the pretreatment clinical and intracerebral inflammatory phenotype and 9-month survival of 764 adults with TBM. All were genotyped for single-nucleotide polymorphism rs17525495, and inflammatory phenotype was defined by cerebrospinal fluid (CSF) leukocyte and cytokine concentrations.Results. LTA4H genotype predicted survival of human immunodeficiency virus (HIV)–uninfected patients, with TT-genotype patients significantly more likely to survive TBM than CC-genotype patients, according to Cox regression analysis (univariate P = .040 and multivariable P = .037). HIV-uninfected, TT-genotype patients had high CSF proinflammatory cytokine concentrations, with intermediate and lower concentrations in those with CT and CC genotypes. Increased CSF cytokine concentrations correlated with more-severe disease, but patients with low CSF leukocytes and cytokine concentrations were more likely to die from TBM. HIV infection independently predicted death due to TBM (hazard ratio, 3.94; 95% confidence interval, 2.79–5.56) and was associated with globally increased CSF cytokine concentrations, independent of LTA4H genotype.Conclusions. LTA4H genotype and HIV infection influence pretreatment inflammatory phenotype and survival from TBM. LTA4H genotype may predict adjunctive corticosteroid responsiveness in HIV-uninfected individuals.

High IFN-γ Release and Impaired Capacity of Multi-Cytokine Secretion in IGRA Supernatants Are Associated with Active Tuberculosis.

Interferon gamma (IFN-γ) release assays (IGRAs) detect Mycobacterium tuberculosis (Mtb) infection regardless of the active (ATB) or latent (LTBI) forms of tuberculosis (TB). In this study, Mtb-specific T cell response against region of deletion 1 (RD1) antigens were explored by a microbead multiplex assay performed in T-SPOT TB assay (T-SPOT) supernatants from 35 patients with ATB and 115 patients with LTBI. T-SPOT is positive when over 7 IFN-γ secreting cells (SC)/250 000 peripheral blood mononuclear cells (PBMC) are enumerated. However, over 100 IFN-γ SC /250 000 PBMC were more frequently observed in the ATB group compared to the LTBI group. By contrast, lower cytokine concentrations and lower cytokine productions relative to IFN-γ secretion were observed for IL 4, IL-12, TNF-α, GM-CSF, Eotaxin and IFN-α when compared to LTBI. Thus, high IFN-γ release and low cytokine secretions in relation with IFN-γ production appeared as signatures of ATB, corroborating that multicytokine Mtb-specific response against RD1 antigens reflects host capacity to contain TB reactivation. In this way, testing cytokine profile in IGRA supernatants would be helpful to improve ATB screening strategy including immunologic tests.

A novel intrinsically disordered outer membrane lipoprotein of Aggregatibacter actinomycetemcomitans binds various cytokines and plays a role in biofilm response to interleukin-1β and interleukin-8

ABSTRACTIntrinsically disordered proteins (IDPs) do not have a well-defined and stable 3-dimensional fold. Some IDPs can function as either transient or permanent binders of other proteins and may interact with an array of ligands by adopting different conformations. A novel outer membrane lipoprotein, bacterial interleukin receptor I (BilRI) of the opportunistic oral pathogen Aggregatibacter actinomycetemcomitans binds a key gatekeeper proinflammatory cytokine interleukin (IL)-1β. Because the amino acid sequence of the novel lipoprotein resembles that of fibrinogen binder A of Haemophilus ducreyi, BilRI could have the potential to bind other proteins, such as host matrix proteins. However, from the tested host matrix proteins, BilRI interacted with neither collagen nor fibrinogen. Instead, the recombinant non-lipidated BilRI, which was intrinsically disordered, bound various pro/anti-inflammatory cytokines, such as IL-8, tumor necrosis factor (TNF)-α, interferon (IFN)-γ and IL-10. Moreover, BilRI played a role in the in vitro sensing of IL-1β and IL-8 because low concentrations of cytokines did not decrease the amount of extracellular DNA in the matrix of bilRI− mutant biofilm as they did in the matrix of wild-type biofilm when the biofilms were exposed to recombinant cytokines for 22 hours. BilRI played a role in the internalization of IL-1β in the gingival model system but did not affect either IL-8 or IL-6 uptake. However, bilRI deletion did not entirely prevent IL-1β internalization, and the binding of cytokines to BilRI was relatively weak. Thus, BilRI might sequester cytokines on the surface of A. actinomycetemcomitans to facilitate the internalization process in low local cytokine concentrations.

Synergy between Common γ Chain Family Cytokines and IL-18 Potentiates Innate and Adaptive Pathways of NK Cell Activation.

Studies to develop cell-based therapies for cancer and other diseases have consistently shown that purified human natural killer (NK) cells secrete cytokines and kill target cells after in vitro culture with high concentrations of cytokines. However, these assays poorly reflect the conditions that are likely to prevail in vivo in the early stages of an infection and have been carried out in a wide variety of experimental systems, which has led to contradictions within the literature. We have conducted a detailed kinetic and dose–response analysis of human NK cell responses to low concentrations of IL-12, IL-15, IL-18, IL-21, and IFN-α, alone and in combination, and their potential to synergize with IL-2. We find that very low concentrations of both innate and adaptive common γ chain cytokines synergize with equally low concentrations of IL-18 to drive rapid and potent NK cell CD25 and IFN-γ expression; IL-18 and IL-2 reciprocally sustain CD25 and IL-18Rα expression in a positive feedback loop; and IL-18 synergizes with FcγRIII (CD16) signaling to augment antibody-dependent cellular cytotoxicity. These data indicate that NK cells can be rapidly activated by very low doses of innate cytokines and that the common γ chain cytokines have overlapping but distinct functions in combination with IL-18. Importantly, synergy between multiple signaling pathways leading to rapid NK cell activation at very low cytokine concentrations has been overlooked in prior studies focusing on single cytokines or simple combinations. Moreover, although the precise common γ chain cytokines available during primary and secondary infections may differ, their synergy with both IL-18 and antigen–antibody immune complexes underscores their contribution to NK cell activation during innate and adaptive responses. IL-18 signaling potentiates NK cell effector function during innate and adaptive immune responses by synergy with IL-2, IL-15, and IL-21 and immune complexes.

The natural product phyllanthusmin C enhances IFN-γ production by human NK cells through upregulation of TLR-mediated NF-κB signaling.

Natural products are a major source for cancer drug development. NK cells are a critical component of innate immunity with the capacity to destroy cancer cells, cancer-initiating cells, and clear viral infections. However, few reports describe a natural product that stimulates NK cell IFN-γ production and unravel a mechanism of action. In this study, through screening, we found that a natural product, phyllanthusmin C (PL-C), alone enhanced IFN-γ production by human NK cells. PL-C also synergized with IL-12, even at the low cytokine concentration of 0.1 ng/ml, and stimulated IFN-γ production in both human CD56(bright) and CD56(dim) NK cell subsets. Mechanistically, TLR1 and/or TLR6 mediated PL-C's activation of the NF-κB p65 subunit that in turn bound to the proximal promoter of IFNG and subsequently resulted in increased IFN-γ production in NK cells. However, IL-12 and IL-15Rs and their related STAT signaling pathways were not responsible for the enhanced IFN-γ secretion by PL-C. PL-C induced little or no T cell IFN-γ production or NK cell cytotoxicity. Collectively, we identify a natural product with the capacity to selectively enhance human NK cell IFN-γ production. Given the role of IFN-γ in immune surveillance, additional studies to understand the role of this natural product in prevention of cancer or infection in select populations are warranted.