Interleukin Research Articles

Human Amniotic Epithelial Stem Cells Promote Functional Recovery After Spinal Cord Injury In Rats By Regulating The Polarization Of Macrophages.

Spinal cord injury (SCI) is a catastrophic nerve injury caused by extremelysevere damage to thespinalcord, for which effective treatments are currently unavailable. Human amniotic epithelial stem cells (hAESCs) are considered promising candidates for transplantation in various clinical and preclinical applications, due to their lack of limitations such as ethical barriers, immune rejection, tumorigenicity, or cell origin. Nevertheless, the effectiveness and mechanism by which hAESCs treat SCI remain elusive.To assess the motor function recovery process following SCI in rats, the Basso Beattie Bresnahan (BBB) behavior test, inclined plate scale and motor evoked potential (MEP) analysis were used in this study after transplantation of hAESCs at different doses. And the underlying mechanism was investigated by histological and molecular methods. The transplantation of hAESCs can significantly promote the recovery of motor function in SCI group, and the higher the dose, the better the effect. Compared with SCI group, hAESCs group had reduced tissue damage, significantly increased the number of neurons, neurofilaments and myelin sheath, and significantly reduced syringomyelia and glial scars. In addition, hAESCs inhibited the Levels of tumor necrosis factor (TNF-α) and interleukin-6 (IL-6) and increased the expression of the interleukin-4 (IL-4), interleukin-10 (IL-10) and interleukin-13 (IL-13), and promoted the shift of M1-polarized macrophages to M2-polarized macrophages. Our results demonstrate that hAESCs promoted the recovery of motor function after SCI by promoting M2 polarization of macrophages and reducing neuroinflammation.These findings may provide novel therapeutic strategies for SCI.

Amelioration of Systemic Amyloidosis by Blocking IL-17A and Not by IL-17F, and Arteriosclerosis by Blocking Both IL-17A and IL-17F in an Inflammatory Skin Mouse Model.

There are comorbidities and complications in atopic dermatitis and psoriasis that often occur after the appearance of skin inflammation. Statistically, data show that patients with psoriasis and atopic dermatitis have a shorter life expectancy than patients without psoriatic dermatitis, due to the occurrence of arteriosclerosis, myocardial infarction, and cerebral infarction. Many types of skin inflammation are treated with various antibody preparations, and marked improvement in patients' quality of life can be achieved. The next theme is to understand the pathogenesis of arteriosclerosis, myocardial infarction, stroke, and other complications associated with dermatitis and to find treatments and drugs to reduce their occurrence. The skin, a crucial immune organ, generates large amounts of inflammatory cytokines in response to various stimuli, leading to systemic inflammation and potential damage to internal organs. The link between inflammatory skin conditions like psoriasis and atopic dermatitis with serious health complications such as vascular disorders and systemic amyloidosis has been increasingly recognized. In psoriasis, biological treatments targeting Interleukin (IL)-17A, a key cytokine, have shown promise in reducing cardiovascular risks. Recent developments include treatments that target both IL-17A and IL-17F in the psoriasis field, though each cytokine's impact on internal organ damage is still under debate. Among visceral complications secondary to dermatitis, systemic amyloidosis and atherosclerosis have been reported to be controlled by suppressing IL-17 in the early stages of dermatitis. Still, it remains unclear whether suppressing IL-17 prevents organ damage in the late stages of persistent severe dermatitis. A study using a long-lasting dermatitis mouse model that overexpressed human caspase-1 in keratinocytes (Kcasp1Tg) investigated the effects of deleting IL-17A and IL-17F on visceral complications. Cross-mating Kcasp1Tg with IL-17A-, IL-17F-, and IL-17AF-deficient mice assessed the skin and visceral organs histologically, and RT-PCR analysis of aortic sclerosis markers was performed. Despite less improvement in dermatitis, deletion of IL-17A in Kcasp1Tg mice showed promising results in reducing multiple organ amyloidosis. On the other hand, the effect was observed in both IL-17A and IL-17F deleted mice for aortic sclerosis. The inhibition of IL-17A and IL-17F was suggested to reduce the risk of developing comorbidities in internal organs. IL-17A and IL-17F were found to act similarly or produce very different results, depending on the organ.

A microglia-containing 3D human brain organoid for studying HSV-1-induced Alzheimer’s disease

Alzheimer’s disease (AD), recognized for its profound incapacitation, presents with notable cognitive decline and memory deficits. A growing body of research points to a link between infection with herpes simplex virus type 1 (HSV-1) and the exacerbation of AD. Our study pioneers the use of a cutting-edge, three-dimensional human brain organoid model enriched with microglia, referred to as MC-HBO, to explore the mechanisms by which HSV-1 influences the pathogenesis of AD. Through this model, we examined the regulatory effects of cytokines on amyloid-β (Aβ) deposition, a cardinal pathological manifestation of AD. Our findings demonstrate that the suppression of pro-inflammatory cytokines, including tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6), leads to a significant reduction in Aβ deposition. In contrast, the inhibition of anti-inflammatory cytokines, such as interleukin-10 (IL-10) and interleukin-4 (IL-4), was correlated with an increase in Aβ accumulation. These insights into the role of cytokine signaling pathways suggest a promising therapeutic strategy for potentially decelerating the progression of AD.

CKIP-1 inhibits M2 macrophage polarization to suppress the progression of gastric cancer by inactivating JAK/STAT3 signaling.

Gastric cancer (GC) is a frequently occurring malignancy with poor prognosis. Casein kinase 2 interacting protein-1 (CKIP-1) is a PH domain-containing protein implicated in regulating tumorigenesis and macrophage homeostasis. This study aimed to elucidate the role and potential mechanism of CKIP-1 in the progression of GC. CKIP-1 expression in GC tumor and para-carcinoma tissues was detected using RT-qPCR. Then, human monocyte cell line THP-1 was treated with PMA, interleukin (IL)-4 and IL-13 to induce M2-polarized macrophages. CD206, arginase-1 (Arg-1) and transforming growth factorβ1 (TGFβ1) expression in M2-polarized macrophages with or without CKIP-1 overexpression was evaluated. Moreover, GC cell lines (MKN45 and HGC27 cells) were co-cultured with CKIP-1-overexpressed M2-polarized macrophages, and the viability, migration and invasion of GC cells were measured. Additionally, immunoblotting assessed the expression of JAK/STAT3 signaling-related proteins and STAT3 agonist Colivelin was used to treat GC cells to perform the rescue experiments to analyze the changes of malignant phenotypes of GC cells. Results showed that CKIP-1 was downregulated in GC tissues and M2-polarized macrophages. CKIP-1 overexpression inhibited M2 macrophage polarization and decreased TGFβ1 secretion. Besides, elevated CKIP-1 expression in M2-polarized macrophages inhibited the viability, migration and invasion of GC cells. Furthermore, CKIP-1 overexpression inactivated JAK2/STAT3 signaling in GC cells by inhibiting TGFβ1 level. Specifically, Colivelin treatment abrogated the influences of CKIP-1 upregulation on the malignant phenotypes of GC cells. Collectively, CKIP-1 inhibits M2 macrophage polarization to suppress the progression of GC by inactivating JAK/STAT3 signaling pathway.

Th17 and T Regulatory Cytokines in Serum, Lesional Skin, and Stimulated Peripheral Blood Mononuclear Cell Culture Supernatants from Type 1 Leprosy Reaction Patients

There are conflicting reports regarding the roles of T helper-17 (Th17) and T regulatory (Treg) cells in type 1 leprosy reactions (T1Rs). Also, literature on the correlation of immunological parameters with a validated scoring system and the effect of treatment on cytokines is lacking. Adult patients with untreated T1R and nonreactional spectrum–matched controls were included in the study for comparison of levels of Th17 and Treg pathway cytokines in serum, skin lesions (reactional), and peripheral blood mononuclear cells (PBMCs) culture supernatants. Venous blood samples were collected at baseline and after resolution of reaction (post treatment with nonsteroidal anti-inflammatory drugs [NSAIDs] or steroids) for serum cytokine estimation and PBMC stimulation assays, and lesional (reactional) skin biopsy for cytokine messenger RNA (mRNA) estimation. Thirty-two cases of T1R were recruited (23 patients completed follow-up). Serum levels of cytokines were not significantly different between cases and controls or between pre- and post-treatment samples. Tissue mRNA and Mycobacterium leprae (M. leprae) antigen–stimulated PBMC culture supernatant levels of Interleukin (IL)-17A, IL-17F, IL-6, and IL-23 were significantly higher in T1R than in controls. Levels of IL-10 and Transforming Growth Factor-beta (TGF-β) were comparable among the two groups. The levels of all cytokines were significantly reduced after treatment. There was no significant difference in magnitude of the fall between those treated with steroids versus NSAIDs. This study suggests heightened Th17 response in T1R, with a prominent inability of the regulatory cytokines IL-10 and TGF-β to control the associated inflammation. The dynamics of change after resolution of T1R were comparable between NSAID and oral steroid treatment groups.

Cyclooxygenase-2/prostaglandin E2 pathway orchestrates the replication of infectious bronchitis virus in chicken tracheal explants

ABSTRACT In this study, we investigated the localized pathogenesis of infectious bronchitis virus (IBV) in chicken tracheal organ cultures (TOCs), focusing on the role of inducible cyclooxygenase (COX-2). Two divergent IBV strains, respiratory Connecticut (Conn) A5968 and nephropathogenic Delmarva (DMV)/1639, were studied at 6, 12, 24, and 48 hours post-infection (hpi). Various treatments including exogenous prostaglandin (PGE)2, a selective COX-2 antagonist (SC-236), and inhibitors of PGE2 receptors and Janus kinase (JAK) were administered. IBV genome load and antigen expression were quantified using real-time quantitative PCR and immunohistochemistry. COX-2, interferon (IFN)-α, IFN-β, interleukin (IL)−1β, IL-6, and inducible nitric oxide synthase (iNOS) expressions were measured, along with PGE2 and COX-2 concentrations. IBV genome load and protein expression peaked at 12 and 24 hpi, respectively. Conn A5968-infected TOCs exhibited continuous COX-2 expression for up to 24 hpi, extended PGE2 production up to 48 hpi, and reduced inflammatory cytokine expression. In contrast, DMV/1639-infected TOCs displayed heightened inflammatory cytokine expression, brief COX-2 expression, and PGE2 production. Treatment with IFN-γ, SC-236, PGE2 receptor inhibitors, or JAK inhibitors reduced IBV infection and lesion scores, whereas exogenous PGE2 or IFN-γ pretreatment with a JAK-2 inhibitor augmented infection. These findings shed light on the innate immune regulation of IBV infection in the trachea, highlighting the involvement of the COX-2/PGE2 pathway. IMPORTANCE Understanding the localized pathogenesis of infectious bronchitis virus (IBV) within the trachea of chickens is crucial for developing effective control strategies against this prevalent poultry pathogen. This study sheds light on the role of inducible cyclooxygenase (COX-2) and prostaglandin (PGE)2 in IBV pathogenesis using chicken tracheal organ culture (TOC) models. The findings reveal distinct patterns of COX-2 expression, PGE2 production, and immune responses associated with different IBV strains, highlighting the complexity of host-virus interactions. Furthermore, the identification of specific inhibitors targeting the COX-2/PGE2 pathway and Janus kinase-signal transducer and activator of transcription (JAK-STAT) signaling pathway provides potential therapeutic avenues for mitigating IBV infection in poultry. Overall, this study contributes to our understanding of the innate immune regulation of IBV infection within the trachea, laying the groundwork for the development of targeted interventions to control IBV outbreaks in poultry populations.

Alpha-Pinene-encapsulated lipid nanoparticles diminished inflammatory responses in THP-1 cells and imiquimod-induced psoriasis-like skin injury and splenomegaly in mice.

Psoriasis, a persistent skin condition caused by the disorder of the immune system, impacts approximately 1.25 million individuals globally. Nevertheless, the presence of adverse effects in conventional clinical drugs necessitates further exploration of novel medications or combination therapies to mitigate these reactions and enhance their effectiveness. Hence, our intention here in this paper is to utilize the lipid nanoparticle delivery system for overcoming the volatility and hydrophobic properties of α-pinene, a naturally occurring compound renowned for its anti-inflammatory and antiviral effects, and further explore its potential pharmacological applications both in vitro and in vivo. The production of α-pinene lipid nanoparticles (APLNs) was achieved through the utilization of high pressure homogenization methods. APLNs was successfully fabricated with enhanced stability and water solubility. Meanwhile, the application of APLNs could drastically reduce the expression of lipopolysaccharide (LPS)-induced inflammation-related factors in THP-1 cells. Administration of APLNs to a mouse model of auricular swelling could effectively reduce redness and swelling in the auricles of mice as well. Furthermore, APLNs were also found to alleviate skin damage in mice with Imiquimod (IMQ)-induced psoriasis model, as well as decrease the levels of psoriasis-related protein nuclear factor kappa-B (NF-κB) and interleukin-17 (IL-17), interleukin-23 (IL-23), and other inflammation-related cytokines. More importantly, utilization of APLNs successfully mitigated the systemic inflammatory reactions in mice, resulting in the reduction of spleen-to-body ratio (wt%) and of inflammatory cytokines' expression in the serum. Overall, our results suggest that with the help of lipid nanoparticle encapsulation, APLNs possess a better pharmacological effect in anti-inflammation and could potentially serve as an anti-psoriasis drug.

Legumain deficiency halts atherogenesis by modulating T cell receptor signaling.

Atherosclerosis is an age-related pathological process associated with elevated levels of legumain in plaques and plasma. However, the underlying mechanisms remain unclear. The aim of this study was to investigate the role of legumain in the progression of atherosclerotic plaques, with a particular focus on functional and phenotypic changes in CD4+ T cells. Apolipoprotein E-deficient (Apoe-/-) mice were crossed with legumain-deficient (Lgmn-/-) mice to generate Lgmn-/-Apoe-/- mice. CD4+ T cells accumulated in the atherosclerotic plaques of Apoe-/- mice fed a high-fat diet. Deletion of legumain attenuated the deposition of CD4+ T cells in plaques and reduced the number of atherosclerotic lesions. The levels of CD4+ T cells in the blood, lymph nodes, and spleen were decreased in Lgmn-/- mice. Transcriptomic analysis revealed that the deletion of legumain decreased the differentiation, survival, and function of CD4+ memory T cells by suppressing the T cell receptor (TCR) signaling pathway. These changes are accompanied by the downregulation of the antiapoptotic protein B-cell lymphoma 2 (Bcl-2) and the reduced release of interleukin (IL)-2 and interferon (IFN)-γ. These results suggest that legumain deficiency may play a role in the development of atherosclerosis by impairing the survival, proliferation, and function of CD4+ T cells. Inhibition of legumain activity may be an innovative therapy for the treatment of atherosclerosis.

Biologics in congenital ichthyosis: are they effective?

Congenital ichthyoses (CI) comprise a heterogeneous group of genetic diseases requiring lifelong treatment and having a major effect on quality of life. Conventional treatments reduce scaling and skin discomfort; however, they usually have little or no effect on erythema and pruritus. The identification of cytokine alterations in CI raised the possibility of repurposing available biologics. Several case reports in the literature report successes using different biologics. We aimed to report the effects of biologics in real life. This was a retrospective, observational, international multicenter study of patients with CI treated with at least one biologic for a minimum of 3 months. The effect of the biologics was evaluated using an Investigator Global Assessment-Change (IGA-C) scale. A comprehensive literature search was performed in parallel. A total of 98 patients were included, with a mean age of 19.7 years and both sexes equally represented. Patients with Netherton syndrome (NS) or congenital ichthyosiform erythroderma (CIE) represented the majority of patients (30% and 21.4%, respectively). Most patients (84.7%) had a severe or very severe form of CI. The most frequently used biologics were inhibitors targeting interleukin-17 (IL-17), IL-12/IL-23, or the IL-4 receptor. The mean duration of treatment was 22+20.1 months. There were 45 responders (45.9%), including 18 patients (18.3%) who were good responders; all had an erythrodermic CI subset and received one of the three main biologics. In 2 NS and CIE, IL-12/IL-23 and IL-4 receptor inhibitors tended to be most effective. Review of the literature revealed a shorter mean duration of use of biologics (11.5+8.5 months) and higher percentage of responders (85.7%), suggesting reporter bias. This series identified subsets of CI that may respond to biologics and will aid in designing future clinical trials of biologics for CI.

Interleukin 6 in asymptomatic patients with nonsevere aortic valve stenosis: a post-hoc study of the SEAS trial

Abstract Background Inflammation plays a pivotal role in the pathogenesis of aortic valve stenosis (AS) through cytokine-mediated valve calcification. Studies of explanted valves found associations of an elevated pro-inflammatory cytokine, Interleukin 6 (IL-6), with increased postoperative mortality. However, limited knowledge exists regarding the prognostic value of elevated circulating IL-6 in asymptomatic patients with nonsevere AS. Purpose To investigate the hypothesis that elevated plasma IL-6 concentration ≥7 pg/mL (above the upper limit of normal 95th percentile) correlates with more severe AS and a higher risk of adverse outcomes in asymptomatic patients with nonsevere AS. Methods We analyzed data of 1574 asymptomatic patients who had mild-to-moderate AS, preserved systolic and kidney function, no overt cardiovascular diseases, and an event-free follow-up one year after enrollment in the randomized Simvastatin and Ezetimibe in Aortic Stenosis (SEAS) trial. We ascertained the correlation of year-0 and year-1 IL-6 levels with echocardiographic AS severity, defined as moderate-severe AS in the presence of ≥2 measurements at year-0: aortic valve area <1 cm², mean pressure gradient ≥40 mmHg, maximal transaortic velocity ≥4 m/s, and velocity ratio <0.25. Multivariable regression examined correlations between IL-6 at year-0 and clinical variables. Cox proportional hazard models and competing risk analyses examined associations of elevated IL-6 at year-1 and 4-year risk of the primary composite endpoint of major cardiovascular events (MACE) comprising the first event of cardiovascular death, aortic valve replacement, hospitalization with heart failure, myocardial infarction, percutaneous coronary intervention, coronary artery bypass graft, or ischemic stroke. Results At year-0, among 1574 patients with a mean (SD) age of 67.5 (9.7) years, 629 (40.0%) were women, 263 (16.7%) had moderate-severe AS, and 194 (12.3%) had elevated IL-6. The median relative IL-6 change from year-0 to year-1 was 1.0 (IQR, 0.8-1.4) across AS-severity groups (P=0.12) (Figure 1). In adjusted models, IL-6 at year-0 did not correlate with AS-severity (P=0.30). In mild AS, an elevated IL-6 at year-1 was associated with an increased 4-year MACE risk (HR 1.40; 95% CI, 1.02-1.92; p=0.04) in adjusted Cox regression analyses (Figure 2A). In sensitivity analyses, the annual IL-6 increase >50% combined with normal IL-6 was associated with an increased 4-year MACE-risk of HR 1.48 (95% CI, 1.10-2.00; P=0.009) in patients with mild AS and 1.82 (95% CI, 0.68-1.74; P=0.73) in patients with moderate-severe AS (all P for interaction>0.25) (Figure 2C-D). Conclusion In asymptomatic patients with nonsevere AS, circulating IL-6 remains stable within the reference range in 7 out of 8 patients during a 1-year follow-up, regardless of AS severity. An elevated IL-6 was associated with a 1.4-fold increased 4-year MACE-risk in patients with mild AS, suggesting its possible role as an early risk marker.Figure 1Figure 2

Peroxisome proliferator-activated receptor gamma regulates interleukin-6 induced lipoprotein(a) gene expression in human HepG2 cells

Abstract Background Lipoprotein(a)[Lp(a)] is an independent risk factor for atherosclerotic cardiovascular disease (ASCVD) and outcomes. The LPA gene promoter contains an interleukin 6 (IL-6) responsive binding site. IL-6 is a pro-inflammatory cytokine and data from patients with inflammatory diseases indicate an impact of the inflammatory milieu on LPA gene regulation and Lp(a) production. In a clinical study, the peroxisome proliferator-activated receptor gamma (PPARγ) agonist pioglitazone was associated with decreased Lp(a) levels. Publicly accessible ChIP-sequencing of human hepatocytes and measures of Lp(a) production by HepG2 cells provide an opportunity to explore PPARγ regulation of hepatic Lp(a) production. Purpose To investigate the role of PPARγ as a regulator of hepatic Lp(a) production. Methods Public ChIP-sequencing of human hepatocytes was analyzed. Commercially available HepG2 cells were incubated with one of the following: (1) scrambled small interfering RNA (siRNA) alone (control); (2) IL-6 (20ng/mL) and scrambled siRNA ; (3) IL-6 and PPARγ siRNA ; and (4) IL-6 and pioglitazone (25uM). LPA transcription was measured using the quantitative polymerase chain reaction. Results Analysis of the ChIP-sequencing data demonstrated that the LPA locus variant rs56393506, which is associated with increased Lp(a) levels and ASCVD risk in the general population, was within the location of the PPARγ binding site in the LPA promoter (Figure 1). IL-6 significantly increased LPA transcription in the HepG2 cells, compared to control (1.65±0.44 vs. 0.66±0.45, p=0.0027). In the presence of siRNA-mediated knock-down of PPARγ (58±17% PPARγ reduction, p<0.0001 vs. scrambled control), the effect of IL-6 on LPA transcription was significantly amplified (6.78±2.4 vs. 1.65±0.44, p=0.0009). IL-6 did not increase LPA transcription in the presence of the PPARγ agonist pioglitazone compared to control (0.16±0.03 vs. 0.66+0.45, p=0.11, Figure 2). Conclusion PPARγ is a negative regulator of IL-6 induced hepatic Lp(a) production and may represent a new therapeutic target in patients with inflammatory conditions.Figure 1Figure 2

Interleukin-6 levels in atrium and onset of spontaneous atrial fibrillation: is there a causal relationship

Abstract Background Cumulative evidence suggests that atrial interleukin-6 (IL-6) may play a significant role in the development of spontaneous postoperative atrial fibrillation (sPOAF) in coronary artery bypass grafting (CABG) patients. Our previous studies found that the IL-6 produced by the atrium is positively related to the development of sPOAF. However, the causal relationship between IL-6 and sPOAF is not established. Purpose This study aims to explore the potential causality between atrial IL-6 and spontaneous AF(sAF), alongside assessing the impact of IL-6 on the electrophysiological properties of atrial myocytes (AM). Methods To determine the causal link between IL-6 and sAF, mice were randomly divided into eight groups: seven receiving injections of varying doses of recombinant IL-6 protein groups (15ng/kg to180ng/kg) into the left atrial wall, and one control group (vehicle). Mice were monitored for sAF for 7 days using implanted telemetry device. At 48 hrs post-injection, the left atrial appendages of another set of mice were collected for analyzing levels of IL-6, Myeloperoxidase (MPO), Transforming Growth Factor-β1 (TGF-β1), and Tumor Necrosis Factor-α (TNF-α). To assess the effect of IL-6 on the electrophysiological characteristics of AM, AMs were isolated from the hearts of 12-week-old mice and divided into experimental and control groups for electrophysiological testing. The experimental group was perfused with extracellular solutions containing 100 ng/mL of IL-6 while the control group was perfused with standard extracellular solutions. The measurements focused on action potential (AP), transient outward potassium current (Ito), and inward rectifier potassium current (Ik1) after a 30-minute perfusion. Results As depicted in Fig. 1A, a dose-response relationship between IL-6 levels and sAF occurrence was observed, with incidence increasing from 0% at 15 ng/kg to 75% at 180 ng/kg (χ²= 16.691, P = 0.005). The injection of IL-6 also induced the production of MPO, TGF-β1, and TNF-α with a dose-dependent effects (Fig. 2). In comparison to control, the AP duration at 90% (APD90) and APD50 of repolarization of atrial myocytes were significantly shortened in IL-6 group (Fig. 1B). For voltages from -10mV to +70mV, atrial myocytes in the IL-6 group showed significantly lower Ito current density than those in the control (Fig. 1C). No significant alterations were observed in the impact of IL-6 on Ik1 current density in atrial myocytes (Fig.1D). Conclusion To the best of our knowledge, this is the first study that experimentally tested the causal relation between IL-6 and sAF. The injection of IL-6 could cause the onset of sAF and produce MPO, TGF-β1, and TNF-α with a significant dose-dependent effect. The effect of IL-6 on inducing sAF might be through its effect on reducing APD and Ito current density in mouse atrial myocytes. This suggests that inhibition of atrial IL-6 might be a potential novel sPOAF prevention strategy.

The role of IL15 in angiotensin II-mediated cardiac remodelling

Abstract Hypertension (HTN) mediated target organ damage, such as cardiac remodelling, is a major cause of death and disability. Cytokines are important regulators of cardiac function, inflammation, and remodelling in cardiovascular disease. Increased levels of interleukin 15 (IL15) have been noted in HTN, but its role in HTN heart disease remains unknown. This study investigates the role of the IL15 axis in the pathogenesis of cardiac remodelling in HTN. Male 12-week-old IL15 receptor α knock-out (IL15RAKO), IL15 knock-out (IL15KO) and matching wild type (WT) mice were administered vehicle or angiotensin II (AngII) (490 ng/min/kg) for 2 weeks by osmotic minipump. Blood pressure was measured by tail cuff (n=5-8). Cardiac function was analysed by echocardiography (n=3-4). Cardiac infiltrating immune cells were analysed by flow cytometry (n=2-3 for IL15RAKO, n=6-10 for IL15KO). Cardiac remodelling was assessed by picrosirius red, WGA staining (n=5-8) and expression of markers of remodelling using RT-qPCR and western blots (n=5-8). Data are presented as mean±SEM and analysed using two-way ANOVA. AngII infusion caused a significant increase in systolic blood pressure in WT and IL15KO animals. In contrast, in IL15RAKO, the response to AngII was blunted. This was in line with a decrease in ejection fraction, however only WT mice had an increase in global longitudinal strain (-20.3±2.0 vs -10.9±1.2, p=0.0039 in WT and 19.9±0.8 vs -16.0±2.1, p=0.1878 in IL15RAKO). HTN was associated with increased cardiac IL15 protein levels in mice. The heart-to-body ratio, along with cardiomyocyte cross-sectional, were significantly increased in WT, but not in IL15RAKO HTN mice. Furthermore, histological analysis showed significantly higher cardiac collagen accumulation in WT but not IL15RAKO upon AngII infusion (5.8±1.0% in WTvvs. 2.1±0.5% in IL15RAKO, p=0.0021). This was in line with a higher expression of col1a1 in WT but not IL15RAKO HTN hearts (20.0±6.0 vs 4.5±1.7, p=0.0173). Similarly, the FN1 protein level was significantly increased in WT but not IL15RAKO AngII-treated mice. These changes were associated with a higher number of Cd11b+ macrophages accumulated in WT hypertensive hearts only. In contrast to IL15RAKO, mice lacking IL15 showed similar increases in global longitudinal strain, heart hypertrophy, and cardiomyocyte cross-sectional area as WT mice upon AngII infusion. Similarly, IL15KO hypertensive hearts showed a significant increase of col1a1 and FN1 in comparison to vehicle-treated mice. A similar level of Cd11b+ macrophages was found in hypertensive IL15KO and WT mice's hearts. In conclusion, IL15RA, but not IL15, plays a critical role in cardiac remodelling mediated by AngII infusion. This suggests that the IL15-IL15RA axis may play an important yet complex role in hypertensive heart disease.

Interleukin-6 and suPAR improves prediction of all-cause mortality in type 1 diabetes - the Thousand & 1 Study

Abstract Background Cardiovascular diseases (CVD) are the leading cause of mortality in type 1 diabetes (T1D). Inflammation frequently accompanies T1D and is suggested as an important contributor to the pathogenesis and progression of CVD. However, current risk prediction models for T1D lack biomarkers of inflammation. Methods A prospective study of individuals with T1D without overt heart disease was evaluated. Interleukin-6 (IL-6), soluble urokinase-type plasminogen activator receptor (suPAR) and high-sensitivity C-reactive protein (hsCRP) were analysed. In Cox proportional hazards model, adjustments were made for variables in the Steno T1 Risk Engine: age, sex, duration of diabetes, HbA1c, systolic blood pressure, glomerular filtration rate, albuminuria status, use of statins, tobacco use and physical activity. The model included the biomarkers as continuous variables, and a "combination" category was defined as simultaneously elevated suPAR > 4 ng/mL and IL-6 > 3 pg/mL. The net reclassification improvement (NRI) and C-statistics were calculated. Results For included individuals (n=1,014, 52% male, mean age 50±15), follow-up was 100% after median of 12.2 years (interquartile range: 11.8 to 12.9).In the fully adjusted model, IL-6 and suPAR were both associated with all-cause mortality (IL-6 hazard ratio (HR): 1.6 (95% confidence interval: 1.04 to 2.4), suPAR HR: 1.9 (1.2 to 2.9). HsCRP was not associated with the outcome. The combination category showed even stronger association: HR 2.7 (1.4 to 5.1). When added to the Steno T1 Risk Engine, IL-6, suPAR, and their combination all added discriminatory power in predicting all-cause mortality assessed by NRI; IL-6: 38% (19 to 58), suPAR: 45% (25 to 65), combination 56% (36 to 76). C-statistics showed similar results for only suPAR. Area under the curve for suPAR: from 0.84 to 0.86 (P=0.049), IL-6: 0.84 to 0.85 (P=0.338), combination: 0.84 to 0.86 (P=0.066). Conclusions IL-6 and suPAR are independently associated with all-cause mortality in T1D without known heart disease. Assessment of IL-6 and suPAR in T1D may enhance risk prediction.Figure 1Figure 2

Interleukin 21, plasminogen activator inhibitor 1 and fractalkine improve risk stratification in elderly patients with pulmonary embolism

Abstract Background The importance of clinical parameters for predicting short-term mortality in hemodynamically stable patients with pulmonary embolism (PE) is acknowledged within the Pulmonary Embolism Severity Index (PESI) Score. Little is known about the value of novel biomarkers in this context. Given the pathophysiology of PE, markers related to inflammation and thrombosis, such as interleukin 21 (IL-21), fractalkine and plasminogen activator inhibitor 1 (PAI1) are of particular interest. Purpose To identify new protein biomarkers that improve 90d mortality prediction compared to the PESI score in elderly PE patients. Methods 1003 patients with venous thromboembolism aged ≥65 years were recruited between 2009 and 2011 from 9 hospitals in Switzerland (SWITCO65+ cohort). Among these, 778 patients were excluded due to missing data, deep venous thrombosis only, blood sample taken >1 day after diagnosis, no initial anticoagulation, systolic blood pressure <90 mmHg, withdrawal within 1 day or denying further use of data; 225 patients were prospectively included. Outcome was independently adjudicated up to 1 year after the index event. To identify candidate proteins, untargeted proteomic analyses (OLINK) were conducted in cases with 90d all-cause mortality (n=20) matched 1:2 (age, sex) to controls without events (n=40). 26 candidate proteins were then validated in all 225 PE patients using ELISA and Mesoscale analyses. Using ROC analyses, the most promising proteins to predict death within 90d were identified within this cohort. Optimal biomarker cut-off levels were derived using the Youden Index and survival analyses were computed. To evaluate the predictive value of these biomarkers, hazard ratios (HR) were calculated after adjusting for age, sex, and BMI. AUC-ROC analyses were performed to assess the biomarkers’ value for risk prediction compared to the PESI score. Results Baseline characteristics (Figure 1) did not differ between patients who died within 90d and those who did not, except for median PESI score (108 [95 – 125] vs. 87 [79 – 101], p<0.001). Optimal biomarker cut-off values were: IL-21: 56.17pg/ml, PAI1: 36.82pg/ml, fractalkine: 66’515pg/ml. Increased levels of IL-21 and PAI1 were associated with increased mortality (HR IL-21 = 1.5 [95% CI: 0.99 - 2.79], HR PAI1 = 2.6 [95% CI: 1.7 – 4.8]), whereas patients with elevated fractalkine levels were at lower risk to die within 90d (HR fractalkine = 0.37 [95% CI: 0.16 - 0.60]). For prediction of 90d mortality, the three assessed biomarkers showed a higher AUC than the PESI score. Combining the PESI score with the biomarkers did not further improve the AUC (Figure 2). Conclusions The protein biomarkers IL-21, PAI1 and fractalkine improve prediction of 90d mortality in elderly, hemodynamically stable PE patients compared to the PESI score. These biomarkers highlight the contribution of inflammation and thrombosis in the pathophysiology of PE and warrant further investigation.Baseline CharacteristicsROC Curves for Biomarkers and PESI Score

Evaluation of aortic inflammation in marfan syndrome patients with 18-fluorodeoxyglucose positron emission and computed tomography

Abstract Background The role of inflammation in Marfan Syndrome (MS) has been well studied in current research. These studies have investigated the role of serum Interleukin 6 (IL-6), Transforming Growth Factor-b (TGF-b) and Macrophage Colony Stimulating Factor (MCSF) as biomarkers of disease severity and progression. Surgical repair is the recommended treatment of aortic aneurysms when aortic diameter reaches the indication threshold. It has been observed however that dissection may occur even before this threshold is reached. 18-fluorodeoxyglucose (18 F-FDG) positron emission tomography / computed tomography (PET/CT) is the gold standard imaging modality to noninvasively assess vascular inflammation. Purpose Our aim was to investigate whether MS patients have increased aortic wall inflammation as assessed by 18F-FDG PET/CT, and the relationship between aortic wall inflammation with serum levels of IL-6, TGF-b and MCSF. Methods The study population consisted of fifteen MS patients and seventeen sex and age matched controls. In all subjects, serum levels of circulating IL-6, TGF-b and MCSF were measured. In PET/CT, the arterial target-to-background ratio (TBR) was calculated for aortic root (AR), ascending aorta(As), aortic arch (AA) and descending aorta (DA). Also, all patients were assessed with transthoracic echocardiography where the diameters of AR, As, AA, and DA were estimated at baseline and were reexamined at the end of the follow up period. Results TBR for MS vs. controls was 2.17±0.75 vs 2.01±0.36, p=0.75 at AR; 2.15±0.67 vs 1.84±0.35, p=0.040 at As; 2.16±0.64 vs 1.70±0.25, p=0.03 at AA and 1.97±0.52 vs 1.83±0.42, p=0.23 at DA respectively. Additionally, Marfan patients had increased levels of serum biomarkers. However, this difference was statistically significant (SS) only for IL-6 serum levels; 2.17±0.46 vs 1.08±0.12 p=0.011 and IL-6 was SS associated with AR TBR (rho=0.572, p=0.033), As TBR (rho=0.604, p=0.022) and DA TBR (rho=0.714, p=0.004). Two of our patients were operated due to severe dilatation of the aortic root and the surgical specimens showed aortic wall inflammation of moderate severity along with fibrosis. Conclusion MS patients have increased of aortic wall inflammation as assessed by 18F-FDG PET/CT, a finding that highlights the role of inflammation in the pathway of disturbed TGF signaling. Further research is needed to determine its prognostic role and value in identifying patients with an indication for early intervention.

Interleukin-22 inhibits right ventricular remodelling by a unique mechanism in a pulmonary artery constriction model

Abstract Background Fibrosis and remodeling of right ventricle (RV) are associated with prognosis in patients with RV failure. Interleukin-22 (IL-22) is a member of the IL-10 cytokine family, play roles in tissue protection and wound repair. We have previously shown that IL-22 plays an important role in cardiac remodeling after acute myocardial infarction; however, the role of IL-22 in RV fibrosis and remodeling remains elusive. Purpose We investigated the role of IL-22 in the pathogenesis of RV remodeling during pressure overload. Methods Pulmonary artery banding (PAB) is performed by placing a 6-0 suture around the pulmonary artery over a 24 G needle in wild type mice (C57BL/6J) and IL-22 knockout mice (IL-22 KO). Only mice with moderate pulmonary artery stenosis (peak pressure gradient across the pulmonary band: 25–40 mmHg at 1 week after surgery by echo Doppler) were included in the study protocol. Four weeks after PAB, RV function was measured by echocardiography and real-time PCR analysis was performed to measure Col1a1, Col3a1, BNP, and transcriptome analysis was performed. Survival confirmation of mice with moderate pulmonary artery stenosis was also performed up to 56 days. Results IL-22 KO mice had significantly higher 56-day mortality rate compared with WT mice after PAC (p<0.01). Four weeks after PAB, IL-22 KO mice showed markedly increased RV weight (IL-22 KO mice vs. wild-type mice; RV/LV+IVS: 0.48±0.05 vs 0.40±0.02, P=0.002). IL-22 KO mice exhibited significant RV enlargement and RV dysfunction 4 weeks after PAB. (IL-22 KO mice vs wild type mice; RVEDV: 10±2.3mm2 vs 12.9±3.1mm2, P=0.005, TAPSE: 10.8±1.8mm vs 9.5±1.7mm, P=0.04, FAC: 35.2±7.8% vs 23±6.0%, P<0.001). The expressions of Col1a1 and Col3a1 were no difference between in IL22-KO mice and Wild mice. However, BNP, a cardioprotective marker, tended to be lower in IL-22KO mice. The transcriptome analysis also showed that the group of fibrosis-related genes that are elevated in PAC in WT mice were less elevated in IL-22 KO mice. Conclusion These results suggest that IL-22 is involved in RV remodelling during RV pressure loading without a general mechanism.

Interleukin-6 and suPAR are associated with diastolic function in type 1 diabetes: The Thousand & 1 Study

Abstract Background Heart failure (HF) is a common complication in individuals with type 1 diabetes (T1D). Inflammation is an important contributor to the pathogenesis and progression of both T1D and HF – especially HF with preserved ejection fraction (HFpEF). Diastolic dysfunction is a hallmark of HFpEF. Despite this, the association between diastolic function and inflammatory biomarkers is not well-known in individuals with T1D. Methods The Thousand & 1 study of individuals with T1D without known heart disease was studied. Diastolic parameters, including E/e’, was obtained by echocardiography and associated with Interleukin-6 (IL-6), soluble urokinase-type plasminogen activator receptor (suPAR) and high-sensitivity C-reactive protein (hsCRP). These inflammatory biomarkers were analysed as continuous variables and dichotomized in above/below 3 pg/mL (IL-6), 4 ng/mL (suPAR), and 4 mg/L (hsCRP). In multivariable regression analysis, adjustments were made for age, gender, body mass index, duration of diabetes, HbA1c, systolic blood pressure, left ventricular ejection fraction, glomerular filtration rate, albuminuria status, use of statins, tobacco use and physical activity levels. In an additional analyse, we also adjusted for N-terminal pro–B-type natriuretic peptide (NT-proBNP). Results We included 1,014 individuals (52% male, mean age 50±15). IL-6 was elevated in 207 individuals, suPAR in 182 individuals, and hsCRP in 863 individuals. Mean E/e’ was 7.6 and mean left ventricular ejection fraction was 58±5%. In fully adjusted models, inflammatory biomarkers were significantly associated with diastolic function. Thus, individuals with IL-6 > 3 pg/mL had 0.6 (95% confidence intervals: 0.2 to 1.0), P=0.001) higher E/e’ compared to individuals with IL-6 < 3 pg/mL. For every doubling of suPAR levels, E/e’ increased by 0.5 (0.2 to 0.9, P=0.003). For every doubling of hsCRP levels, E/e’ increased by 0.1 (0.02 to 0.2, P=0.022). Individuals with elevated levels of all three biomarkers had 1.9 (0.5 to 3.2, P=0.006) higher E/e’ compared to low levels of all three biomarkers. Including NTproBNP to the model did essentially not change the estimates. Conclusions Firstly, in T1D, elevated levels of IL-6, suPAR and hsCRP are associated with diastolic function assessed by E/e’, independent of traditional risk factors including NT-proBNP. Secondly, IL-6, suPAR and hsCRP may contain information about diastolic function in T1D which is not detected by NT-proBNP.Figure 1Figure 2

Tumor Differentiation is Correlated with Estrogen Receptor Beta (ERβ) Expression but Not with Interleukin-6 (IL-6) Expression in Colorectal Carcinoma

BACKGROUND: Colorectal carcinoma (CRC), the third most common malignant disease worldwide, is associated with estrogen receptor β (ERβ) and interleukin-6 (IL-6). ERβ is known to down-regulate IL-6 in prostate cancer, lung carcinoma, and CRC cell lines; however, its effect on human with CRC remains unclear. Therefore, this study was conducted to investigate the association between ERβ and IL-6 expressions with the clinicopathological features of CRC.METHODS: This was an analytic observational study using 40 paraffin blocks of CRC patients. ERβ and IL-6 expression was measured by immunohistochemistry (IHC) staining. The percentage of immunoreactive tumor cell per 1000 cells was manually recorded and tumor differentiation as well as tumor infiltration were determined. Tumor differentiation was graded according to the World Health Organization (WHO) 2010 criteria, while tumor infiltration was defined based on the American Joint Committee on Cancer (AJCC) 8th edition.RESULTS: Fifty percent of samples were well-differentiated CRC, and 57.5% samples were T3 infiltration tumors. IHC staining showed 35.5% of samples were positive for ERβ expression, while 70.86% were positive for IL-6 expression. There were negative correlation of ERβ expression with tumor differentiation (p=0.018; r=-0.371), but no correlation with tumor infiltration (p=0.836) were found. There was no correlation between ERβ expression with IL-6 expression (p=0.154).CONCLUSION: There is statistically significant correlation between tumor differentiation and ERβ expression, wherein improved tumor differentiation is linked to higher levels of positive ERβ expression. However, there is no discernible relationship between IL-6 and tumor differentiation. These findings suggest that while IL-6 was involved in the growth of the tumor, ERβ expression might have an impact on tumor differentiation.KEYWORDS: colorectal carcinoma, estrogen receptor beta, interleukin-6, cell differentiation

Interleukin-6 signalling, lipoprotein (a) lowering and cardiovascular disease: a mendelian randomisation study

Abstract Background Elevated lipoprotein(a) (Lp[a]) levels and chronic low-grade inflammation play causal roles in cardiovascular disease (CVD) development, representing promising targets for managing residual risk. CVD prevention trials are currently assessing the efficacy of IL-6 pathway inhibition in ameliorating chronic pro-atherogenic inflammation. Lp(a) increases in response to interleukin 6 (IL-6) and acute inflammatory states, and Lp(a) itself induces an inflammatory response that accelerates atherosclerosis. Recent studies indicate that IL-6-receptor inhibitors may lower Lp(a) levels and reduce inflammation, potentially decreasing CVD risk in an additive manner. Purpose This drug-target Mendelian Randomisation (MR) study aims to elucidate the on-target effects of pharmacological IL-6R and LPA inhibition on CVD and explore the relationship between downregulated IL6R signalling and Lp(a) levels. Methods Independent genetic variants associated with C-reactive protein levels from around the IL6R gene locus, and Lp(a) lowering variants from around the LPA gene locus were utilised to assess the effect of genetically proxied IL6R and LPA inhibition on CVD risk in large genomic consortia. Two-step cis-MR was then employed to estimate the effect of lower genetically predicted IL6R signalling on CVD, after adjusting for its influence on Lp(a) levels. Results Genetically proxied LPA inhibition was associated with reduced odds of coronary artery disease (CAD) (OR 0.88, 95% CI 0.86-0.89), ischaemic stroke (IS) (OR 0.98, 95% CI 0.96-1.00), cardioembolic stroke (OR 0.95, 95% CI 0.92-0.97), heart failure (HF) (OR 0.96, 95% CI 0.92-1.00), and aortic stenosis (OR 0.88, 95% CI 0.85-0.91). IL6R inhibition was associated with reduced odds of CAD (OR 0.76, 95% CI 0.68-0.86), IS (OR 0.89, 95% CI 0.80-0.98), small vessel stroke (SVS) (OR 0.72, 95% CI 0.59-0.89), large artery stroke (LAS) (0.61, 95% CI 0.48-0.78) and atrial fibrillation (OR 0.73, 95% CI 0.65-0.83). Furthermore, downregulated IL6R signalling was associated with lower Lp(a) levels (SD change -0.04, 95% CI -0.08,-0.01). In two-step cis-MR, Lp(a) levels partially mediated the total effect of IL6R inhibition on CAD (proportion-mediated = -11.33%, 95% CI -11.35, -11.31%) and LAS (proportion mediated = -4.19%, 95% CI -4.21, -4.17%), but had little effect on IS (proportion-mediated = -1.78%, 95% CI -1.80, -1.76%), SVS (proportion-mediated = -2.50%, 95% CI -2.52, -2.48%), or atrial fibrillation (proportion-mediated = -0.20%, 95% CI -0.22, -0.18%). Conclusion IL6R and Lp(a) inhibition are independently associated with reduced risk of CVD. Furthermore, IL6R inhibition is associated with lower Lp(a) and its effect on CVD is independent of its Lp(a)-lowering effects. Targeting the IL6 signalling pathway offers a promising approach to mitigate residual CVD risk in individuals with elevated Lp(a) levels. Future trials should investigate the additive effects of combined IL-6 inhibition and Lp(a) lowering therapies.