Serum Levels Of Interferon-gamma Research Articles

Alterations in CD4+ T Cell Cytokines Profile in Female Patients with Hashimoto's Thyroiditis Following Vitamin D Supplementation: A Double-blind, Randomized Clinical Trial.

Hashimoto's thyroiditis (HT) is an autoimmune disease characterized by the destruction of thyroid cells through immune processes involving T helper (Th)1 cytokines. This clinical trial investigates the impact of vitamin D supplementation on serum cytokine levels and gene expression in CD4+ T cells from HT patients, aiming to understand its effects on Th-1, Th-2, Th-17, and regulatory T (Treg) cell-associated factors. Female patients were randomly assigned in a double-blind design to either a vitamin D-supplemented group, which received cholecalciferol [1, 25(OH)2D3] at a dose of 50,000 IU, or the placebo group, which received a weekly placebo for a duration of three months. Serum cytokine levels were assessed using enzyme-linked immunosorbent assay (ELISA), while genes' expression levels were measured using real-time PCR. Serum 25-hydroxyvitamin D and levels exhibited a significant increase following vitamin D supplementation, in comparison to the placebo group. Additionally, the vitamin D supplementation resulted in a significant elevation of serum calcium (Ca) levels compared to baseline. In the vitamin D group, there was a significant decrease in both serum levels and expression of the interleukin (IL)-17 gene when compared to baseline, although no statistical difference was observed between the placebo and vitamin D groups. The gene expression of transforming growth factor-beta (TGFβ) was significantly increased in the vitamin D group compared to baseline, with no significant difference between the two study groups. Vitamin D treatment had no effect on serum levels of interferon-gamma (IFNϒ) and IL-4. While the gene expression of IL-4 in the vitamin D group did not exhibit a statistically significant increase, the level of GATA3 transcription factor increased significantly when compared to the placebo group. The expression of IFNϒ and transcription factors, T-bet, RORc, and forkhead box protein 3 (FOXP3) in genes did not show significant changes following vitamin D supplementation. The findings suggest that vitamin D supplementation may hold potential benefits for autoimmune diseases, such as HT. However, further longitudinal clinical trials are necessary to gain a more comprehensive understanding of the specific effects of vitamin D on HT. IRCT2016110130644N1.

Sequential immunotherapy and targeted therapy for metastatic BRAF V600 mutated melanoma: 4-year survival and biomarkers evaluation from the phase II SECOMBIT trial

No prospective data were available prior to 2021 to inform selection between combination BRAF and MEK inhibition versus dual blockade of programmed cell death protein-1 (PD-1) and cytotoxic T lymphocyte antigen-4 (CTLA-4) as first-line treatment options for BRAFV600-mutant melanoma. SECOMBIT (NCT02631447) was a randomized, three-arm, noncomparative phase II trial in which patients were randomized to one of two sequences with immunotherapy or targeted therapy first, with a third arm in which an 8-week induction course of targeted therapy followed by a planned switch to immunotherapy was the first treatment. BRAF/MEK inhibitors were encorafenib plus binimetinib and checkpoint inhibitors ipilimumab plus nivolumab. Primary outcome of overall survival was previously reported, demonstrating improved survival with immunotherapy administered until progression and followed by BRAF/MEK inhibition. Here we report 4-year survival outcomes, confirming long-term benefit with first-line immunotherapy. We also describe preliminary results of predefined biomarkers analyses that identify a trend toward improved 4-year overall survival and total progression-free survival in patients with loss-of-function mutations affecting JAK or low baseline levels of serum interferon gamma (IFNy). These long-term survival outcomes confirm immunotherapy as the preferred first-line treatment approach for most patients with BRAFV600-mutant metastatic melanoma, and the biomarker analyses are hypothesis-generating for future investigations of predictors of durable benefit with dual checkpoint blockade and targeted therapy.

IL-34 attenuates acute T cell-mediated rejection following renal transplantation by upregulating M2 macrophages polarization

ObjectiveTo investigate the role of Interleukin-34 (IL-34) in acute T cell-mediated rejection (TCMR) following renal transplantation. MethodsThe mice acute TCMR model of renal transplantation was established and identified by hematoxylin and eosin (HE) and immunohistochemistry (IHC) staining. Then, IHC staining of IL-34 was also performed to determine the expression of IL-34 in allografts. Recipients were infected with IL-34 overexpression adeno-associated virus, infection efficiency of which was estimated by enzyme linked immunosorbent assay (ELISA), Western blot, and immunofluorescence. HE and IHC staining were used to estimate the grades of TCMR. Flow cytometry was performed on lymphocytes in spleens of recipients including regulatory T cells (Tregs) and M2 macrophages. The expression of cytokines in vivo was analyzed by Mouse Cytokine Grp I Panel. Finally, Tregs and M2 macrophages were cultured in vitro and treated with IL-34 to observe the effects of IL-34 on the differentiation of the cells. ResultsThe mouse TCMR model was successfully established by HE, periodic acid shiff (PAS), CD4 and CD8 IHC staining. The expression of IL-34 was significantly decreased in allografts with TCMR. BALB/c mice were successfully infected with IL-34 overexpression adeno-associated virus. Subsequently, the grade of rejection in mice TCMR model was evaluated by HE and IHC staining according to Banff criteria. It is suggested that the grade of TCMR in IL-34 overexpressed mice was significantly decreased. IHC staining and Flow cytometry showed that the proportion of Tregs and M2 macrophages in the spleens and allografts were significantly increased in IL-34 overexpressed mice. Serum levels of interferon-gamma (IFN-γ), IL-17 and tumor necrosis factor-alpha (TNF-α) were downregulated in IL-34 overexpressed mice. Moreover, IL-34 could promote macrophage M2 polarization, while failed to promote differentiation of naïve T cells into Tregs in vitro. ConclusionOverexpression of IL-34 may attenuate the progression of TCMR episodes in allografts by increasing the polarization of M2 macrophages in the spleens and allografts, which may become a potential therapeutic strategy for TCMR.

The impact of interferon-gamma level on the health status of patients with sickle cell disease in Basrah

Sickle cell disease (SCD) is one of the most prevalent autosomal recessive diseases, characterized by the generation of abnormal hemoglobin S. Our study aimed to assess how the serum level of interferon-gamma affects the health status of patients with SCD in Basrah. A total of 90 participants were enrolled in this study and divided into two main groups: a SCD group and a control group. The SCD group included 30 patients with SCD in steady state and 30 patients with SCD in vasoocclusive crisis; the control group included 30 ageand sexmatched apparently healthy individuals. Approval was obtained from the Research Ethics Committee of the College of Medicine, University of Basrah before conducting the study. Two milliliters of venous blood were drawn from all the participants, and ELISA tests were utilized to determine the levels of serum interferon-gamma. There was a statistically significant increase in the serum level of interferon-gamma among SCD patients (both in steady state and in crisis) compared to the control group (p = 0.05). There were no significant differences in the levels of interferon-gamma between the patients in steady state and during vaso-occlusive crisis (p > 0.05). Interferon-gamma may influence the general health of sickle cell patients and contribute to the cause of inflammation, no matter whether the patient is in stable condition or is experiencing a crisis.

The impact of ionizing radiation compared to drug-induced immunological changes.

Ionizing radiation (IR) activates several signaling pathways. This study shows the impact of acute low-dose IR on crucial cytokines involved in cell-mediated immunity. The immunomodulatory effects of 0.25 and 0.5 Gray (Gy) gamma rays and standard immunomodulatory drugs (cyclophosphamide) on blood counts and significant pro-inflammatory cytokines implicated in various inflammatory conditions were tested in 20 rats. Examined was the effect of acute low doses on critical cytokines, which could be utilized as an alternative to current immunosuppressive drugs. One day post-irradiation, serum levels of interferon-gamma (INF-γ), tumor necrosis factor-alpha, and interleukin-2/1-beta were measured. A 0.25Gy exposure did not affect the detected cytokines or blood cell count compared to the nonirradiated group. On the other hand, 0.5Gy raises the majority of the immunologically examined cytokines except for INF-γ. Except for INF-γ, cyclophosphamide reduces all of the cytokines examined. As a result, low-dose IR has a less negative influence on essential inflammatory cytokines, permitting its use. More research is needed to determine how low amounts could be used in different immunological disorders.

Serum Interferon Gamma (IFN-ɣ) Levels and Hematological Indices in Patients with HIV-MTB Co-Infection in North-Eastern Nigeria

Introduction: The dual epidemic of human immunodeficiency virus (HIV) infection and Mycobacterium tuberculosis (MTB) poses significant health challenges, particularly in sub-Saharan Africa. Understanding the immune response and hematological changes in HIV-MTB co-infection is crucial for better management of affected individuals. Objectives: This study aimed to evaluate the serum levels of IFN-ɣ and hematological indices in patients with HIV-MTB co-infection in North-Eastern Nigeria, as well as explore any potential relationships between these factors. Methods: A total of 88 participants were enrolled in the study, including 44 antiretroviral therapy-naive patients with HIV-MTB co-infection (study group) and 44 HIV mono-infected individuals as controls. Data on personal biodata and clinical details were collected using an interviewer-administered questionnaire. Blood samples were obtained from each participant and analyzed for IFN-ɣ levels using ELISA and hematological indices using an automated hematology analyzer. Statistical analysis, including Mann-Whitney U test, independent samples t-test, and Pearson's correlation analysis, was conducted to compare the study and control groups and assess the relationship between IFN-ɣ levels and hematological parameters. Results: Serum IFN-ɣ was insignificantly increased in the study group compared to the control group (p=0.093). The WBC count was also significantly reduced in the study group compared to the control group (p=0.038). The HGB, HCT, MCV and MCH were significantly reduced in the study group compared to the control group (p=0.001, 0.001, 0.002 and 0.001) respectively. Participants with HIV-TB co-infection have insignificantly increased serum IFN-ɣ levels, low total WBC, lymphocyte and monocyte counts compared to those with HIV mono-infection. In conclusion, participants with HIV-TB co-infection have insignificant increased serum IFN-ɣ levels, low total WBC, and lymphocyte and monocyte counts compared to those with HIV mono-infection. There was no correlation of IFN-ɣ with any of the haematological indices.

Cellular and transcriptional impacts of Janus kinase and/or IFN-gamma inhibition in a mouse model of primary hemophagocytic lymphohistiocytosis.

Primary hemophagocytic lymphohistiocytosis (pHLH) is an inherited inflammatory syndrome driven by the exuberant activation of interferon-gamma (IFNg)-producing CD8 T cells. Towards this end, ruxolitinib treatment or IFNg neutralization (aIFNg) lessens immunopathology in a model of pHLH in which perforin-deficient mice (Prf1-/-) are infected with Lymphocytic Choriomeningitis virus (LCMV). However, neither agent completely eradicates inflammation. Two studies combining ruxolitinib with aIFNg report conflicting results with one demonstrating improvement and the other worsening of disease manifestations. As these studies used differing doses of drugs and varying LCMV strains, it remained unclear whether combination therapy is safe and effective. We previously showed that a ruxolitinib dose of 90 mg/kg lessens inflammation in Prf1-/- mice infected with LCMV-Armstrong. To determine whether this dose controls inflammation induced by a different LCMV strain, we administered ruxolitinib at 90mg/kg to Prf1-/- mice infected with LCMV-WE. To elucidate the impacts of single agent versus combination therapy, Prf1-/- animals were infected with LCMV, treated or not with ruxolitinib, aIFNg or both agents, and analyzed for disease features and the transcriptional impacts of therapy within purified CD8 T cells. Ruxolitinib is well-tolerated and controls disease regardless of the viral strain used. aIFNg, administered alone or with ruxolitinib, is most effective at reversing anemia and reducing serum IFNg levels. In contrast, ruxolitinib appears better than aIFNg, and equally or more effective than combination therapy, at lessening immune cell expansion and cytokine production. Each treatment targets distinct gene expression pathways with aIFNg downregulating IFNg, IFNa, and IL-6-STAT3 pathways, and ruxolitinib downregulating IL-6-STAT3, glycolysis, and reactive oxygen species pathways. Unexpectedly, combination therapy is associated with upregulation of genes driving cell survival and proliferation. Ruxolitinib is tolerated and curtails inflammation regardless of the inciting viral strain and whether it is given alone or in combination with aIFNg. When administered at the doses used in this study, the combination of ruxolitinb and aIFNg appears no better than treatment with either drug alone in lessening inflammation. Further studies are warranted to elucidate the optimal doses, schedules, and combinations of these agents for the treatment of patients with pHLH.

Anti-inflammatory effect of hydrogen sulfide donor sodium-sulfide in an experimental mouse model of contact hypersensitivity

Background: Hydrogen sulfide (H2S), synthesized by most immune cells, has been shown to exert anti-inflammatory and antipruritic effects. The effect of H2S on allergic contact dermatitis (ACD), an inflammatory skin disease that negatively affects the quality of life, is unknown. Objectives: We planned to investigate the antipruritic and anti-inflammatory effects of the H2S donor sodium sulfide (Na2S) in the experimental mouse model of contact hypersensitivity (CHS), which is widely used for ACD research. Methods: CHS was created in Balb/c mice using 1-fluoro-2, 4-dinitrobenzene. Na2S was administered systemically (0.2-2-20 mg/kg/i.p.) and locally (1-3-10 nmol/both ear/i.d.) at 3 h and 25 h after the challenge. Ear thickness and the number of scratches were determined at 24 h and 48 h following the challenge. Ear tissue and serum interferon-gamma, interleukin (IL)-2, IL-4, and IL-5 cytokine levels were evaluated by enzyme-linked immunosorbent assay (ELISA). H and E staining was performed for histopathological studies. CD4+ and CD8+ T cells located in the skin were examined by immunohistochemical staining. Results: Locally (1-3-10 nmol/ear/i.d., P < 0.001, P < 0.0001, P < 0.0001, respectively) and systemically (2–20 mg/kg/i.p., P < 0.01, P < 0.0001, respectively), Na2S administration decreased ear thickness dose dependently. Local (1-3-10 nmol/ear/i.d.) Na2S treatment decreased serum IL-2 levels (P < 0.01, P < 0.05, and P < 0.01, respectively). Na2S administered locally (3–10 nmol/ear/i.d., P < 0.05) and systemically (20 mg/kg/i. p., P < 0.05) decreased the number of CD4+ T lymphocytes. Conclusion: Locally and systemically administered Na2S reduces ear thickness, which is one of the symptoms of CHS, probably by preventing CD4+ T lymphocyte infiltration and proliferation and decreasing IL-2 synthesis.

Effect of exogenous galectin-9, a natural TIM-3 ligand, on the severity of TNBS- and DSS-induced colitis in mice.

Inflammatory bowel disease (IBD) have a complex pathogenesis that is yet to be completely understood. However, a strong correlation between Toll-like receptor 4 (TLR4)/nuclear factor-κB (NF-κB) signaling and IBD has been observed. T-cell immunoglobulin and mucin domain-containing-3 (Tim-3) has been reported to regulate TLR4/NF-κB by interacting with Galectin-9 (Gal-9), and recombinant Gal-9 can activate Tim-3; however, its potential properties in IBD and the underlying mechanism remain unclear. This study aimed to determine how Gal-9 affects experimental colitis in mice. Dextran sodium sulfate (DSS) and 2,4,6-trinitrobenzene sulfonic acid (TNBS) were used to establish colitis in mice, and the severity of the illness was assessed based on body weight, colon length, and histology. Therefore, we explored the effects of Gal-9 treatment on colitis. Furthermore, we analyzed the effect of Gal-9 on the expression of Tim-3 and TLR4/NF-κB pathway in colonic tissues and the serum levels of interferon-gamma (IFN-γ), interleukin (IL)-1β, and IL-6. Tim-3 expression in the colon was notably decreased in mice with TNBS-induced colitis, whereas TLR4/NF-kB expression was significantly increased. Intraperitoneal injection of Gal-9 dramatically decreased the disease activity index and attenuated the level of intestinal mucosal inflammation in TNBS-induced colitis mice (p<0.05). Intraperitoneal administration of Gal-9 significantly increased Tim-3 expression in the colon and decreased the serum concentrations of IFN-γ, IL-1β, and IL-6. Additionally, Gal-9 treatment significantly downregulated the expression of TLR4 signaling pathway-related proteins. In contrast, Gal-9 did not reduce the severity of DSS-induced colitis. In summary, exogenous Gal-9 increased Tim-3 expression, inhibited the TLR4/NF-κB pathway, and alleviated TNBS-induced colitis in mice but not DSS-induced colitis in mice, revealing its potential therapeutic ramifications for IBD.

The effects of L-citrulline adjunctive treatment of Toxoplasma gondii RH strain infection in a mouse model

Toxoplasma gondii is a zoonotic intracellular protozoan parasite and its therapeutic limitations are one of its major problems. L-citrulline is an organic compound that has beneficial effects on many diseases. The purpose of this study was to assess the impact of L-citrulline, alone or in combination with sulfamethoxazole-trimethoprim (SMZ-TMP) on acute toxoplasmosis caused by Toxoplasma gondii RH virulent strain. In our study, 60 Swiss albino mice were divided into two main groups; the control group and the infected treated group, which was subdivided into group IIa: infected treated with L-citrulline, group IIb: infected treated with SMZ-TMP, and group IIc: infected treated with L-citrulline combined with SMZ-TMP. The effects of treatment were assessed by parasitological study, electron microscopic study of tachyzoites, and histopathological study of the liver. Moreover, ELISA measurement of the serum level of Interferon-gamma, Interleukin 10, Nitric oxide, and apoptotic markers was used. It was noticed that L-citrulline combined with SMZ-TMP significantly increased the survival time of infected mice with a significant decrease in the number of tachyzoites compared to the other groups. Moreover, it increased the levels of measured cytokines and serum anti-apoptotic proteins Bcl-2 and improved the extent of liver cell damage associated with a decrease in inflammatory infiltration. In conclusion, L-citrulline supplementation was found to be effective against acute toxoplasmosis, especially when combined with SMZ-TMP as it has multifactorial mechanisms; nitric oxide production, anti-inflammatory, anti-apoptotic, and immune stimulator.

Subsets of natural killer cells in chronic myeloid leukemia and their relation with some inflammatory cytokines

BACKGROUND: As in other malignancies, different subsets of natural killer (NK) cells play a crucial role in the recognition and lysing of malignant cells in chronic myeloid leukemia (CML). OBJECTIVES: This study aims to identify two subsets of NK, cytotoxic (cluster of differentiation [CD] 16+bright) and cytokine-producing NK (CD56+bright) in newly diagnosed CML patients. MATERIALS AND METHODS: This study is conducted on 20 newly diagnosed Iraqi patients (12 males and 8 females) with CML, in chronic phase, at the age range of 17–55 years. Along with patients, 20 healthy subjects (with matched age and gender) were enrolled to act as a control group. To identify NK cells and their subsets in peripheral blood samples, the expression of CD45, CD3, CD56, and CD16 markers was evaluated by flow cytometry technique. Furthermore, the serum level of interferon gamma (IFN-γ) and interleukin (IL)-18 was determined by the enzyme-linked immunosorbent assay technique. RESULTS: The age of patients at the diagnosis of disease is (35.6 ± 12.2 years) and the male: female ratio was 1.5:1. The serum level of IL-18 in newly diagnosed CML patients (30.3 ± 6.5 pg/mL) was significantly (P &lt; 0.0001) higher than those in control group (18.3 ± 7.8 pg/mL), while the serum levels of IFN-γ in newly diagnosed patients are significantly (P = 0.006) dropped down to (89.1 ± 7.2 pg/mL from that in control group (109.4 ± 30.3 pg/mL). The percentage of NK cells in newly diagnosed CML patients is significantly lower than in control group. There is a significant elevation in the cytotoxic NK cells (CD16+bright) subset, and a significant decrease in the cytokine-producing NK subset (CD56+bright) in newly diagnosed patients when compared to those in control group. CONCLUSION: Although there is an elevation in the percentage of cytotoxic NK cells (CD16+bright) subset of CML patients at the first diagnosis, these cells are not able to recognize and attack malignant cells, which may be due to low expression of their activating receptors and needs more investigation. Furthermore, present results found a low percentage of cytokine-producing NK cells (CD56+bright) and a low level of IFN-γ in CML patients, although there is an elevation in IL-18, which indicates that IL-18 may be not the main stimulator to these cells, so activation pathway of this subset of NK cells needs further investigation.

Potential Histopathological and Immune Biomarkers in Malignant and Non-Malignant Oral Lesions.

The presented case-control study was developed to characterize the clinical, histopathological and immunological profile of patients with traumatic injuries, benign neoplasms, potentially malignant oral disorders and malignant neoplasms of the oral cavity, in order to identify biomarkers of malignancy. Clinical information was collected from dental records and several techniques were performed, including histopathological evaluation in sections stained with haematoxylin and eosin, immunohistochemistry for programmed death ligand-1 and measurement of serum levels of interferon-gamma, interleukin-6, -10 and -12. Statistical analysis was performed using IBM SPSS® Statistics software. This study included 8 patients with traumatic injuries, 8 with benign neoplasms, 6 with potentially malignant oral disorders and 11 with malignant neoplasms. An association was observed between the classification of the lesion and smoking (P < 0.05), the size of the lesion (P < 0.05), the density of the inflammatory infiltrate (P < 0.001), the degree of dysplasia (P < 0.01) and programmed death ligand-1 expression (P < 0.01). Therefore, it is suggested that smoking, the size of the lesion, the inflammatory infiltrate and the programmed death ligand-1 expression can be considered potential biomarkers of oral malignancy.

The prevalence and the effect of interferon -γ in the comorbidity of rheumatoid arthritis and depression

BackgroundDepression is the most common comorbidities associated with rheumatoid arthritis (RA). We aimed to explore the mechanism of association between RA and depression. Methods120 subjects were enrolled and depression was diagnosed and assessed using DSM-5 and 24-item version of Hamilton Depression Scale. Pain intensity and joint function in patients with RA were assessed using the visual analog scale (VAS) and health assessment questionnaire (HAQ). Serum levels of interferon-gamma (IFN-γ), indoleamine 2,3-dioxygenase (IDO), kynurenine (KYN), tryptophan (TRP), and quinolinic acid (QUIN)were detected. In animal experiments, K/BxN mice with RA-like phenotype was used and depressive behavior was observed. The protein expression level of N-methyl -D- aspartate receptor 2B (NR2B) in the hippocampus was detected. ResultsIn this study, 36.67 % of patients with RA also had depression. The working status, month family income, tender joint count, the VAS and HAQ score were the main factors influencing the depression in RA patients. HAQ score was found to be an independent risk factor for depression in RA. Serum IDO, IFN-γ, KYN were increased and TRP contents were decreased in RA group. K/BxN mice with RA-like phenotype showed depressive behavior. However, injection of IFN-γ neutralizing antibody could inhibit kynurenine pathway and reverse the depressive behavior in mice. The levels of QUIN in the neurotoxic metabolic pathway were increased and N-methyl -D- aspartate receptors (NMDAR) were activated, which may be the mechanism behind the onset of depression. ConclusionsFrom clinical and preclinical aspects, the occurrence of depression in RA was explored and the related mechanism was revealed.

Kynurenine pathway can be a potential biomarker of fatigue in primary Sjögren's syndrome.

Primary Sjögren's syndrome (pSS) is a chronic autoimmune disease with low quality of life caused by various constitutional symptoms and glandular dysfunction. Although fatigue is one of the most frequent symptoms in pSS, its aetiology or biomarkers are poorly elucidated. We investigated potential relationship between severity of fatigue and the kynurenine pathway in pSS. Clinical data and blood samples of 81 patients were obtained from a prospective cohort for pSS and compared with age- and sex-matched healthy controls (HC). Severity of fatigue was defined according to the fatigue domain scores in the ESSPRI. Potential biomarkers related to the kynurenine pathway were determined using ELISA. Of the total, 44 patients were defined as the "severe fatigue (ESSPRI fatigue ≥ 5)" group, whereas 37 as the "less fatigue (ESSPRI fatigue < 5)". Serum tryptophan levels in the severe fatigue group were significantly lower while those of kynurenine were higher. Serum interferon gamma, IDO1, and quinolinic acid levels were mostly higher in the less fatigue group. Kynurenine/tryptophan ratios were distinctly higher in the severe fatigue group than both HC and the less fatigue group (p < 0.001). This ratio showed a strong degree of positive correlation (r = 0.624, p < 0.001) with severity of fatigue in pSS while the other markers showed fair degrees of correlation. Serum markers related to the kynurenine pathway, especially the kynurenine/tryptophan ratio, may be associated with severity of fatigue in pSS. These results can provide guidance for further investigations on fatigue in pSS.

Influence of gender on coronary atherosclerosis and inflammatory biomarker profile: a CT angiographic study

Abstract Background and aims The burden of coronary atherosclerosis differs between men and women. Beyond traditional cardiovascular risk factors, inflammatory biomarkers can influence plaque formation and progression. We analyzed the influence of gender difference on coronary atherosclerosis and inflammatory cytokines. Methods Coronary CT-Angiography was performed in 301 patients and plaque morphology was quantitatively and qualitatively assessed using semi-automated software (Autoplaque version 2.5, Cedars-Sinai Medical Center, Los Angeles, CA). We analyzed total (TPV), non-calcified (NCPV), calcified (CPV) and low-density plaque volume in mm3 (LDPV, defined as plaque attenuation &amp;lt;30HU). Serum was analyzed for various cytokines including Interleukin (IL)-1a, IL-1b, IL-2, IL-4, IL-6, IL-7, IL-8, IL-10, IL-13, Il-15, IL-17, TNF-a, IFNg, CRP, MCP-1, MIP-1a, Eotaxin, G_CSF and GM-CSF using Luminex assay. Results Out of 301 patients, 94 (31%) were female and 207 (69%) were male. As compared to women, men were significantly younger (62 vs. 57 years, p&amp;lt;0.001) and showed a higher BMI (28 vs. 26 kg/m2, p=0.005). No differences were seen for hypertension, hyperlipidemia, diabetes, smoking habits and family history of CAD. All plaque characteristics showed significant higher values in men as compared to women (all p&amp;lt;0.05). In men, cytokine profile showed significantly lower serum levels for IL-2 (3.20 [3.20; 7.01] vs. 4.31 [3.20; 9.68]; p=0.01) and Interferon-gamma (3.20 [3.20; 14.72] vs. 8.76 [3.20; 26.26]; p&amp;lt;0.001) and significantly higher levels for MCP-1 (224 [117; 327] vs. 155 [49; 260]; p&amp;lt;0.001). In multivariable logistic regression analysis, Interferon-gamma showed significant inverse association to male gender (OR 0.35; 95% CI: 0.17–0.72; p=0.004), but IL-2 and MCP-1 did not. The association of Interferon-gamma to gender was independent of age (OR 0.57; 95% CI: 0.43–0.76; p&amp;lt;0.001) and BMI (OR 2.12; 95% CI 1.25–3.62; p=0.006) Conclusion Coronary plaque volume and morphology significantly differs between men and women and so does the inflammatory cytokine profile. Albeit significantly lower coronary plaque burden in women, serum levels of interferon-gamma - a known pro-inflammatory cytokine - was significantly higher in women with independent association to gender. Whether interferon-gamma plays contradicting roles in the process of coronary atherosclerosis in men and women needs to be explored in future studies to identify potential gender-specific targets for therapeutic interventions. Funding Acknowledgement Type of funding sources: None.

Efficacy and safety of apremilast and its impact on serum cytokine levels in patients with Behçet's disease.

To evaluate the long-term clinical efficacy of apremilast in Behçet's disease (BD) and its effect on serum cytokine levels. This study included 15 BD patients who were treated with apremilast. The rates of change in oral and genital ulcers, skin lesions, arthritis, and arthralgia were evaluated every 3months for 12 months. The efficacy of apremilast was compared between patients with and without oral ulcer remission. Changes in the serum levels of interferon-gamma (IFN-γ), tumor necrosis factor alpha (TNF-α), interleukin (IL)-10, IL-17A, IL-6, IL-8, and IL-23 between baseline and 3months after apremilast initiation were compared. After 3months, oral and genital ulcers disappeared in most cases. The skin and joint lesions tended to improve for up to 6months; however, recurrence was observed after 9months. The improvement of genital ulcers was earlier in the oral ulcer remission group than the oral ulcer non-remission group, with the genital ulcers disappearing within the first 3months. The baseline levels of serum cytokines, analyzed in seven patients, did not exhibit significant associations with specific organ lesions. After administration of apremilast, the TNF-α and IL-23 levels significantly decreased; however, the IFN-γ, IL-6, IL-8, and IL-10 levels did not show significant changes. The rates of decrease in the serum IL-6, IFN-γ, and IL-10 levels were greater in patients with improved oral ulcers. Modulation of serum cytokine levels with apremilast might underlie the efficacy of apremilast in oral ulcers in BD patients.

Targeting the Axl and mTOR Pathway Synergizes Immunotherapy and Chemotherapy to Butylidenephthalide in a Recurrent GBM.

Background The role of inherent tumor heterogeneity and an immunosuppressive microenvironment in therapeutic resistance has been determined to be of importance for the better management of glioblastoma multiforme (GBM). Some studies have suggested that combined drugs with divergent mechanisms may be promising in treating recurrent GBM. Methods Intracranial sustained (Z)-n-butylidenephthalide [(Z)-BP] delivery through Cerebraca Wafers (CWs) to eliminate unresectable brain tumors was combined with the administration of temozolomide (TMZ), pembrolizumab, and cytokine-induced killer (CIK) cells for treating a patient with recurrent glioblastoma. Neurological adverse events and wound healing delay were monitored for estimating tolerance and efficacy. Response Assessment in Neuro-Oncology criteria were applied to evaluate progression-free survival (PFS); further, the molecular characteristics of GBM tissues were analyzed, and the underlying mechanism was investigated using primary culture. Results Intracerebral (Z)-BP in residual tumors could not only inhibit cancer stem cells but also increase interferon gamma levels in serum, which then led to the regression of GBM and an immune-responsive microenvironment. Targeting receptor tyrosine kinases, including Axl and epidermal growth factor receptor (EGFR), and inhibiting the mechanistic target of rapamycin (mTOR) through (Z)-BP were determined to synergize CIK cells in the presence of pembrolizumab and TMZ in recurrent GBM. Therefore, this well-tolerated regimen could simultaneously block multiple cancer pathways, which allowed extended PFS and improved quality of life for 22 months. Conclusion Given the several unique functions of (Z)-BP, greater sensitivity of chemotherapy and the synergism of pembrolizumab and CIK cells could have affected the excellent prognosis seen in this patient with recurrent GBM.

3112 – IMPACT OF IMMUNOPROTEASOME INHIBITION IN MOUSE MODEL OF HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS

Hemophagocytic lymphohistiocytosis (HLH) comprises an expanding group of disorders associated with overactive immune cells that mediate significant tissue damage. Prior studies reveal that persistence of activated antigen presenting cells drives hyperinflammation in HLH. The immunoproteasome is known to play a role in antigen presentation on class I major histocompatibility complex (MHC) and immune effector cell function. Accordingly, we hypothesized that immunoproteasome inhibition might decrease the cellular and molecular hyperinflammatory response in HLH. Supporting this notion, we find that immunoproteasome inhibition significantly reduces in vitro antigen-specific CD8 T cell proliferation and interferon-gamma (IFNg) production. To examine the effects of immunoproteasome inhibition in an in vivo model of primary (genetic) HLH, lymphocytic choriomeningitis virus (LCMV)-infected perforin-deficient (Prf1-/-) mice were treated with the selective immunoproteasome inhibitor zetomipzomib on varying days post-infection. On days 8-10, immune cell subsets, cytokine production, and lymphocytic tissue infiltration were assessed. Untreated LCMV-infected animals developed marked organomegaly with increased effector T cells, neutrophils, and elevated levels of serum IFNg. Each of these parameters was significantly diminished in zetomipzomib-treated animals. Upon ex vivo antigen stimulation, CD8 T cells from zetomipzomib-treated mice exhibited significantly decreased IFNg production. Together, these data highlight the role of persistent antigen presentation in disease development and suggest that immunoproteasome inhibition shows promise as a potential therapeutic strategy for HLH. Further studies are ongoing to optimize the effects and decipher underlying mechanisms. Hemophagocytic lymphohistiocytosis (HLH) comprises an expanding group of disorders associated with overactive immune cells that mediate significant tissue damage. Prior studies reveal that persistence of activated antigen presenting cells drives hyperinflammation in HLH. The immunoproteasome is known to play a role in antigen presentation on class I major histocompatibility complex (MHC) and immune effector cell function. Accordingly, we hypothesized that immunoproteasome inhibition might decrease the cellular and molecular hyperinflammatory response in HLH. Supporting this notion, we find that immunoproteasome inhibition significantly reduces in vitro antigen-specific CD8 T cell proliferation and interferon-gamma (IFNg) production. To examine the effects of immunoproteasome inhibition in an in vivo model of primary (genetic) HLH, lymphocytic choriomeningitis virus (LCMV)-infected perforin-deficient (Prf1-/-) mice were treated with the selective immunoproteasome inhibitor zetomipzomib on varying days post-infection. On days 8-10, immune cell subsets, cytokine production, and lymphocytic tissue infiltration were assessed. Untreated LCMV-infected animals developed marked organomegaly with increased effector T cells, neutrophils, and elevated levels of serum IFNg. Each of these parameters was significantly diminished in zetomipzomib-treated animals. Upon ex vivo antigen stimulation, CD8 T cells from zetomipzomib-treated mice exhibited significantly decreased IFNg production. Together, these data highlight the role of persistent antigen presentation in disease development and suggest that immunoproteasome inhibition shows promise as a potential therapeutic strategy for HLH. Further studies are ongoing to optimize the effects and decipher underlying mechanisms.

Tasquinimod Targets Immunosuppressive Myeloid Cells, Increases Osteogenesis and Has Direct Anti-Myeloma Effects By Inhibiting c-Myc Expression in Vitro and In Vivo

IntroductionImmunotherapy has revolutionized cancer treatment and significantly affected the management of Multiple Myeloma (MM) patients. Unfortunately, these immunotherapeutic approaches are hampered by the presence of a suppressive bone marrow microenvironment including myeloid derived suppressor cells and tumor associated macrophages. Tasquinimod (TasQ), an immunomodulatory compound, is currently in phase Ib/IIa for relapsed/refractory MM patients (NCT04405167). TasQ blocks the interaction between S100A9 and its receptors, which is associated with reduced MDSC accumulation. In this study, we investigated TasQ-mediated direct and indirect effects on MM cell growth, bone disease and immunomodulation in vitro and in vivo using human myeloma cell lines and the immunocompetent 5TMM models.Material and methodsIn vitro, murine (5T33vt, 5TGM1) and human (JJN3, LP1, OPM2, and RPMI8226) MM cell lines were cultured at different concentrations of TasQ. Cell proliferation was assessed by BrdU staining using flow cytometry. C-Myc and pSTAT3 expression were analyzed by western blot. In vitro T cell proliferation experiments were performed using MACS-sorted CD11b + cells and CFSE-labeled T cells from naïve mice. Cells were cocultured for 72h in the presence of MM conditioned medium (5T33MMvt CM) with CD3/CD28 microbeads, followed by flow cytometry to assess T cell proliferation. For in vivo experiments, we used the 5T33 (aggressive) and 5TGM1 (moderate) MM models. On the second day after tumor cell injection, the mice were randomly assigned to the treatment group and the control group. The treatment group received 30 mg/kg of TasQ in drinking water for 35 days (5TGM1) and 21 days (5T33). Anti-tumor and immunomodulating effects were analyzed by flow cytometry (e.g. tumor cells, myeloid subsets, CD4/CD8 + T cells), qRT-PCR, western blot and serum ELISA (interferon-gamma). Effects on osteogenesis in the 5TGM1 model was investigated by Micro-CT. Statistical differences were assessed by Mann-Whitney U test and One-way ANOVA with p<0.05 considered as statistically significant.ResultsTasQ-treatment of murine and human myeloma cell lines (HMCL), at concentrations of 10-25uM, significantly reduced MM cell proliferation after 24h and 48h in vitro (n=3, p<0.05). In addition, a downregulation in c-Myc expression could be observed 6h after treatment of human MM cell lines (n=3). In vitro, TasQ significantly increased T cell proliferation in co-culture experiments with T cells and myeloid cells in 5T33MMvt CM (n=3, p<0.05).Using the immunocompetent 5TGM1 and 5T33MM model, we investigated direct and indirect anti-tumor effects of TasQ. We found that TasQ significantly reduced tumor load in the bone marrow of 5TGM1 (n=10/group, p=0.0012) and 5T33MM mice (n=10/group, p=0.0106) compared to vehicle-treated control mice. Using flow cytometry, we could not observe a difference in the percentage of CD4 + and CD8 + T cells. However, a significant upregulation in serum interferon-gamma could be observed in the 5T33MM mice (p=0.0284). While the percentage of CD11b + cells in the TasQ-treated group was significantly increased (p<0.05), the percentage of monocytic myeloid cells (CD11b +Ly6G -) was significantly reduced in both models (p<0.05). qRT-PCR results showed that the expression of IL-10 was downregulated in purified CD11b + myeloid cells (p<0.05). Consistent with the in vitro data, we observed a decrease in the protein expression of c-Myc in purified MM cells obtained from TasQ-treated mice compared to control mice. Micro-CT analysis of femurs demonstrated a significant increase in the percentage BV/TV (ratio of bone material volume over tissue volume) and trabeculae number (p<0.0001) in TasQ-treated 5TGM1 mice compared to untreated mice.ConclusionTasQ has pleiotropic effects on the MM cells and its surrounding bone marrow microenvironment. It affects MM cell growth by decreasing c-Myc expression. In addition, TasQ targets the immunosuppressive monocytic myeloid cell population and increases serum interferon-gamma levels, indicative for immune cell activation. Moreover, it stimulates osteogenesis in vivo. Taken together, all these data provide evidence for the therapeutic benefits of TasQ as an anti-MM therapy for patients. DisclosuresTörngren: Active Biotech: Current Employment. Eriksson: Active Biotech: Current Employment. De Veirman: Active Biotech AB: Research Funding.

Synthesis and evaluation of metallic nanoparticles-based vaccines against Candida albicans infections

Candida albicans infections have been particularly critical among nosocomial infections in immune-compromised and hospitalized patients. The interaction of microbes with various types of nanoparticles is a crucial stage in determining the biological consequences. The present study aimed to synthesize and evaluate nanomaterial-based vaccines using C. albicans ghosts. The silver nanoparticles (Ag-NPs) and gold nanoparticles (Au-NPs) were environmentally prepared using green biological chemistry using C. albicans ghosts prepared using a modified sponge-like protocol. The interaction of Ag-NPs or Au-NPs with the cell membrane of C. albicans ghost was confirmed by utilizing Transmission Electron Microscope (TEM), Scanning Electron Microscope (SEM), and X-Ray Diffraction (XRD) tools. Experimental rats were injected with different doses of C. albicans ghost mixed with Ag-NPs or Au-NPs to evaluate their effects on the immune system. The ions of Ag-NPs or Au-NPs are entirely converted to zero-valent atoms due to the reduction and stabilization effect of the C. albicans ghost. The prepared nanoparticles exhibited small nanosized with good monodispersity and were stable without noticeable agglomeration at room temperature for two months. Enhancement of both humoral and cellular immune responses was noticed. Au-NPs or Ag-NPs to C. albicans ghost resulted in activation of the immune system and increased serum levels of Interferon Gamma (IFN-γ), a lymphocyte-derived pro-inflammatory cytokine.