Plasma IL-17A Research Articles

Plasma IL-17A is increased in patients with critical MIS-C and associated to in-hospital mortality

BackgroundMultisystem inflammatory syndrome in children (MIS-C) is a rare and severe post-COVID-19 complication with multiple phenotypes.ObjectivesThe aim of this study is to study inflammatory biomarkers (cytokines and oxidative stress) in critical MIS-C patients and to observe if there is association between these biomarkers and mortality.MethodsA single-center prospective study enrolled patients with MIS-C (with positive molecular test), aged between 1 month and 18 years of age. Data was collected from 20 pediatric intensive care unit (PICU)’s bed. Inflammatory biomarkers (cytokines and oxidative stress markers) were performed on day 1 and 3 after hospitalization. Survival rate was calculated, and Kaplan-Meier curves were plotted. Univariate and multivariate Cox regression analyses were conducted. The ROC (Receiver Operating Characteristic) curve analysis was performed.Results and conclusionsA total of 41 patients out of 109 patients admitted at PICU with suspected MIS-C during the study period were included, of which 33 (80.5%) were male, 9 (22%) were under one year old, and 30 (73.2%) presented comorbidities. Among them, 16 (39%) did not survive. The mean survival time was shorter in patients with higher levels of IL-17A (≥ 19.71 pg/mL) on day 1 (115 vs 323 days, p = 0.004). Higher levels of IL-17A on day 1 were associated with mortality in both the crude model (HR 1.03, CI95% 1.004-1.057, p = 0.022) and the adjusted model (HR 1.043, CI95% 1.013-1.075, p = 0.012). ROC analysis revealed a cut-off value for the IL-17A of 14.32 pg/ml. The other immunological and inflammatory markers did not demonstrate an association with survival (p>0.05). Our findings suggest that patients with high levels of IL-17A are at greater risk for death.

Astaxanthin protects fludrocortisone acetate-induced cardiac injury by attenuating oxidative stress, fibrosis, and inflammation through TGF-β/Smad signaling pathway

Hypertensive rats serve as a good experimental model for studying the pathophysiology of cardiac hypertrophy and remodeling leading to heart failure. In this study, we aimed to analyze the effect of astaxanthin and possible mechanisms involved in alleviating oxidative stress, fibrosis and inflammation that triggers cardiac remodeling using male uninephrectomized Long Evans rats. Cardiac hypertrophy and hypertension were induced in rats termed as ‘FCA-Salt rats’ by an oral administration of fludrocortisone acetate (FCA) and 1 % NaCl in drinking water. Biochemical assays showed that FCA-Salt rats exhibited an upregulation of oxidative stress markers AOPP, MDA and downregulation of NO in heart and kidney, which was reversed by astaxanthin treatment. Astaxanthin further regularized the reduced activities of antioxidant enzymes GSH, SOD and CAT in these tissues. ELISA revealed that astaxanthin significantly reduced the inflammatory response by reducing the elevated levels of IL-1β, IL-17a, and TNF-α and pro-fibrotic marker TGF-β1 in plasma. Real-time qPCR depicted an upregulation of TNF-α, IL-1β, IL-6, IL-17A as well as signaling molecules TGF-β1, Smad2 and Smad3 in heart of FCA-Salt rats, which was reduced significantly by astaxanthin. Sirius red staining showed that the cardiac and renal fibrosis was significantly improved by astaxanthin treatment. Together, our results suggest that astaxanthin treatment is beneficial in protecting cardio-renal damage in hypertension through TGF-β/Smad signaling pathway, hence, this molecule may be considered for the maintenance of cardio-renal health.

Immune alterations in children with autism spectrum disorders

Autism spectrum disorders (ASD) are a heterogeneous group of mental and nervous system disorders. Patients with ASD are characterized by communication and cognitive impairments and obsessive behavior. The pathogenesis and etiology of ASD are still unclear. According to the literature, patients suffering from ASD have not only mental, but also somatic disorders, including changes in the immune system. The aim of this work was to study the concentration of cytokines in the blood plasma of children with ASD and the level of expression of proinflammatory genes in peripheral blood mononuclear cells. The clinical group included 71 children aged 4-12 years with a diagnosis of ASD (F84.02). Patients scored more than 36 on the Childhood Autism Rating Scale (CARS). The control sample included 27 apparently healthy children of the same age. The following methods were used in this study: isolation of mononuclear cells from heparinized peripheral blood, Ficoll-Verografin density gradient centrifugation, evaluation of cytokine blocks using commercially available enzyme immunoassay kits, isolation of random total RNA, reverse transcription using hexaprimers, and real-time polymerase chain reaction using intercalating dye SYBR Green. The concentration of proinflammatory cytokines IL-1β, IL-8, and IL-17A in the peripheral blood plasma of children with ASD was statistically significantly increased compared to the control sample. Moreover, the concentration of the anti-inflammatory cytokine IL-10 in patients with ASD was 3.6 times lower compared to the control sample (p 0.001). The level of expression of the NF-κB1, IL1β, IL8 and TNFα genes at the RNA level in peripheral blood mononuclear cells was increased by 2.8, 2.5, 4.8 and 4.2 times in patients with ASD compared to the control sample (all p 0.01). The results obtained indicate an increase in the concentration of proinflammatory cytokines (IL-1β, IL-8, IL-17A) in the blood plasma and a decrease in the concentration of anti-inflammatory cytokines (IL-10) in patients with ASD compared to the control sequence.

Neuroserpin, IL-33 and IL-17A as potential markers of mild symptoms of depressive syndrome in Toxoplasma gondii-infected pregnant women.

Depressive syndrome (DS) is a common complication during pregnancy and the postpartum period, and is triggered by multiple organic/genetic and environmental factors. Clinical and biochemical follow-up is essential for the early diagnosis and prognosis of DS. The protozoan Toxoplasma gondii causes infectious damage to the fetus during parasite primary-infection. However, in long-term infections, pregnant women develop immune protection to protect the fetus, although they remain susceptible to pathological or inflammatory effects induced by T. gondii. This study aimed to investigate plasma inflammatory biomarkers in pregnant women seropositive and seronegative for T. gondii, with diagnoses of minor and moderate/severe DS. Pregnant women (n=45; age=18-39 years) were recruited during prenatal care at health centers in Ouro Preto, Minas Gerais, Brazil. Participants were asked to complete a socio-demographic questionnaire to be submitted to well-standardized DS scale calculators (Beck Depression Inventory Questionnaire, Edinburgh Postnatal Depression Scale, and Major Depressive Episode Module). Additionally, 4 mL of blood was collected for plasma neuroserpin, CCL2, IL-17A, and IL-33 analysis. Pregnant volunteers with chronic T. gondii contact were all IgG+ (44%; n=21) and exhibited increased plasma IL-33, IL-17A, and neuroserpin levels, but not CCL2, compared to uninfected pregnant women. Using Beck's depression inventory, we observed an increase in plasma IL-17A and IL-33 in women with T. gondii infeCction diagnosed with mild DS, whereas neuroserpin was associated with minor and moderate/severe DS. Our data suggest a close relationship between DS in pregnant women with chronic T. gondii infection and neurological conditions, which may be partially mediated by plasma neuroserpin, IL-33, and IL-17A levels.

IL-23/IL-17 Axis in Chronic Hepatitis C and Non-Alcoholic Steatohepatitis-New Insight into Immunohepatotoxicity of Different Chronic Liver Diseases.

Considering the relevance of the research of pathogenesis of different liver diseases, we investigated the possible activity of the IL-23/IL-17 axis on the immunohepatotoxicity of two etiologically different chronic liver diseases. A total of 36 chronic hepatitis C (CHC) patients, 16 with (CHC-SF) and 20 without significant fibrosis (CHC-NSF), 19 patients with non-alcoholic steatohepatitis (NASH), and 20 healthy controls (CG) were recruited. Anthropometric, biochemical, and immunological cytokines (IL-6, IL-10, IL-17 and IL-23) tests were performed in accordance with standard procedure. Our analysis revealed that a higher concentration of plasma IL-23 was associated with NASH (p = 0.005), and a higher concentration of plasma IL-17A but a lower concentration of plasma IL-10 was associated with CHC in comparison with CG. A lower concentration of plasma IL-10 was specific for CHC-NSF, while a higher concentration of plasma IL-17A was specific for CHC-SF in comparison with CG. CHC-NSF and CHC-SF groups were distinguished from NASH according to a lower concentration of plasma IL-17A. Liver tissue levels of IL-17A and IL-23 in CHC-NSF were significantly lower in comparison with NASH, regardless of the same stage of the liver fibrosis, whereas only IL-17A tissue levels showed a difference between the CHC-NSF and CHC-SF groups, namely, a lower concentration in CHC-NSF in comparison with CHC-SF. In CHC-SF and NASH liver tissue, IL17-A and IL-23 were significantly higher in comparison with plasma. Diagnostic accuracy analysis showed significance only in the concentration of plasma cytokines. Plasma IL-6, IL-17A and IL-23 could be possible markers that could differentiate CHC patients from controls. Plasma IL-23 could be considered a possible biomarker of CHC-NSF patients in comparison with controls, while plasma IL-6 and IL-17-A could be biomarkers of CHC-SF patients in comparison with controls. The most sophisticated difference was between the CHC-SF and CHC-NSF groups in the plasma levels of IL-10, which could make this cytokine a useful biomarker of liver fibrosis.

Extracorporeal Photopheresis as a Possible Therapeutic Approach for Adults with Severe and Critical COVID-19 Non-Responsive to Standard Treatment: A Pilot Investigational Study.

Background: The optimal approach for adult patients hospitalized with severe and critical coronavirus disease 2019 (COVID-19), non-responsive to antiviral and immunomodulatory drugs, is not well established. Our aim was to evaluate feasibility and safety of extracorporeal photopheresis (ECP) in this setting. Methods: A prospective, single-center investigational study was performed between 2021 and 2022 at a tertiary referral center for COVID-19. Patients diagnosed with COVID-19 were screened, and cases with severe or critical disease fulfilling pre-defined clinical and biochemical criteria of non-response for >5 days, despite remdesivir, dexamethasone and immunomodulation (tocilizumab, baricitinib, ruxolitinib), were consecutively enrolled. After patient inclusion, two ECP sessions on two consecutive days per week for 2 weeks were applied. Patients were followed-up per protocol from study inclusion, and clinical, virological and radiological outcomes were assessed at the end of treatment (EOT) +28 days. Results: A total of seven patients were enrolled. At inclusion, four out of seven (57.1%) were admitted to the ICU, all patients had ongoing cytokine storm. Additionally, 3/7 (42.9%) had radiological progression on chest CT. At EOT+28 days, 2/7 (28.6%) patients died due to non-ECP-related causes. Among the survivors, no additional requirement for intensive care unit admission or radiological progression was observed, and invasive mechanical ventilation could be weaned off in 1/5 (20.0%). All patients achieved whole-blood SARS-CoV-2 RNAemia clearance, while 3/7 (42.9%) no longer showed detectable respiratory SARS-CoV-2 RNA. According to immune biomarker profiling, ECP mainly facilitated a decrease in plasma IL-6 and IL-17A levels, as well as the physiological regeneration of peripheral blood immunocyte subpopulations, notably CD8+/CD45RO+ memory T-cells. No safety signals were identified. Conclusions: ECP appears to be a safe and feasible option for adults hospitalized with severe or critical COVID-19 who do not respond to pharmacological interventions. Further trial data are warranted to assess its optimal use. Trial registration: ClinicalTrials.gov NCT05882331 (retrospectively registered).

Biomarkers detected in cord blood predict vaccine responses in young infants.

Factors influencing vaccine immune priming in the first year of life involve both innate and adaptive immunity but there are gaps in understanding how these factors sustain vaccine antibody levels in healthy infants. The hypothesis was that bioprofiles associated with B cell survival best predict sustained vaccine IgG levels at one year. Longitudinal study of plasma bioprofiles in 82 term, healthy infants, who received standard recommended immunizations in the United States, with changes in 15 plasma biomarker concentrations and B cell subsets associated with germinal center development monitored at birth, soon after completion of the initial vaccine series at 6 months, and prior to the 12-month vaccinations. Post vaccination antibody IgG levels to Bordetella pertussis, tetanus toxoid, and conjugated Haemophilus influenzae type B (HiB) were outcome measures. Using a least absolute shrinkage and selection operator (lasso) regression model, cord blood (CB) plasma IL-2, IL-17A, IL-31, and soluble CD14 (sCD14) were positively associated with pertussis IgG levels at 12 months, while CB plasma concentrations of APRIL and IL-33 were negatively associated. In contrast, CB concentrations of sCD14 and APRIL were positively associated with sustained tetanus IgG levels. A separate cross-sectional analysis of 18 mother/newborn pairs indicated that CB biomarkers were not due to transplacental transfer, but rather due to immune activation at the fetal/maternal interface. Elevated percentages of cord blood switched memory B cells were positively associated with 12-month HiB IgG levels. BAFF concentrations at 6 and 12 months were positively associated with pertussis and HiB IgG levels respectively. Sustained B cell immunity is highly influenced by early life immune dynamics beginning prior to birth. The findings provide important insights into how germinal center development shapes vaccine responses in healthy infants and provide a foundation for studies of conditions that impair infant immune development.

Elevated Interleukin-37 Associated with Dengue Viral Load in Patients with Dengue Fever

Dengue remains a public health issue worldwide. Similar to chronic infectious diseases, stimulation of cytokine production is not enough to drive immune effector cells for effective virus clearance. One possible mechanism is the virus induces a large number of negative stimulatory cytokines inhibiting immune response. Interleukin 37 (IL-37) plays a crucial regulatory role in infection and immunity, inhibits innate and adaptive immunity as an anti-inflammatory cytokine by inhibiting proinflammatory mediators and pathways. To date, there are few studies reporting correlations between dengue fever (DF) and IL-37. In this study we found that the serum IL-37b and IL-37b-producing monocytes in patients were significantly increased in DF patients. A majority of the IL-37b produced by DF patients was produced by monocytes, not lymphocytes. Increased levels of IL-6, IL-10, and IFN-α were also found in DF patients. However, we failed to detect IL-1β, IL-17A and TNF-α in plasma, because of off-target. In our study, there was no relation between IL-6, IL-10, and IFN-α expressions and IL-37b in serum (P > 0.05). The IL-37b-producing monocytes were negatively correlated with the level of IFN-α in serum and platelet count, and positively correlated with lymphocytes percentage (P < 0.05, respectively). Additionally, serum DENV nonstructural protein 1 levels were positively correlated with monocytes percentages (P < 0.05). Our data represents findings for IL-37b expression and its potential mechanisms in DF patients’ immune response.

Nitric Oxide, Neutrophil/Lymphocyte, and Platelet/Lymphocyte Ratios as Promising Inflammatory Biomarkers in Complicated Crohn’s Disease: Outcomes of Corticosteroids and Anti-TNF-α Therapies

Crohn's disease (CD) is a relapsing-remitting inflammatory bowel disease with a progressive course. The aim of our study was to evaluate the relationship between nitric oxide (NO), pro-inflammatory cytokines, and blood count-based ratios in patients with complicated Crohn's disease as well as the outcome of corticosteroid or anti-TNF-α therapy. In this context, we evaluated the NLR as the ratio of neutrophils count to lymphocytes count, PLR as the ratio of platelets count to lymphocytes count, and MLR as the ratio of monocytes count to lymphocytes count in patients and controls. Furthermore, we assessed NO production by the Griess method in plasma along with iNOS and NF-κB expression by immunofluorescence method in intestinal tissues of patients and controls. In the same way, we evaluated plasma TNF-α, IL-17A, and IL-10 levels using ELISA. Our results indicate that blood count-based ratios NLR, PLR, and MLR were significantly higher in patients compared to controls. In addition, increased systemic levels of NO, TNF-α, and IL-17A and colonic expression of iNOS and NF-κB were observed in the same patients. Interestingly, the high ratio of NLR and MLR as well as NO production were significantly decreased in treated patients. Collectively, our findings suggest that nitric oxide as well as the blood count-based ratios (NLR, PLR, MLR) could constitute useful biomarkers in complicated Crohn's disease, predicting the response to treatments.

Inducible co-stimulatory molecules participate in mesenteric vascular endothelial-mesenchymal transition and sclerosis of mesenteric vessels in spontaneously hypertensive rats

To investigate the correlation of inducible co-stimulatory molecules (ICOS) with mesenteric vascular endothelial- mesenchymal transition (EndMT) and sclerosis in spontaneously hypertensive rats (SHR). Twenty 4-week-old WKY rats and 20 SHRs of the same strain were both randomly divided into 4 groups for observation at 4, 6, 10 and 30 weeks of age. ICOS expression frequency in rat spleen CD4+T cells was analyzed using flow cytometry, and the expressions of ICOS, VE-cad, α-SMA and Col3 mRNA in rat mesentery were detected by RT-PCR. The distributions of ICOS, IL-17A and TGF-β in rat mesentery were detected by immunohistochemistry. The levels of IL-17A and TGF-β in rat plasma were measured using ELISA. The morphological changes of rat mesenteric vessels were observed with Masson staining. Spearman or Pearson correlation analyses were used to evaluate the correlation between ICOS expression and the expressions of the markers of vascular EndMT and sclerosis. Compared with the control WKY rats, the SHRs began to show significantly increased systolic blood pressure and ICOS expression frequency on CD4+T cells at 6 weeks of age (P < 0.05). In the SHRs, the mRNA and protein expressions of ICOS, α-SMA, Col3, IL-17A and TGF-β in the mesentery were significantly higher than those in control group (P < 0.05), while the mRNA and protein expressions of VE-cad started to reduce significantly at 10 weeks of age (P < 0.05). The plasma levels of IL-17A and TGF-β were significantly increased in SHRs since 6 weeks of age (P < 0.05) with progressive worsening of mesenteric vascular sclerosis (P < 0.05). ICOS mRNA and protein expression levels in the mesenteric tissues of SHRs began to show positive correlations with α-SMA and Col3 expression levels and the severity of vascular sclerosis at 6 weeks of age (P < 0.05) and a negative correlation with VE-cad expression level at 10 weeks (P < 0.05). ICOS play an important pathogenic role in EndMT and sclerosis of mesenteric vessels in essential hypertension by mediating related immune responses.

CD1a promotes systemic manifestations of skin inflammation

Inflammatory skin conditions are increasingly recognised as being associated with systemic inflammation. The mechanisms connecting the cutaneous and systemic disease are not well understood. CD1a is a virtually monomorphic major histocompatibility complex (MHC) class I-like molecule, highly expressed by skin and mucosal Langerhans cells, and presents lipid antigens to T-cells. Here we show an important role for CD1a in linking cutaneous and systemic inflammation in two experimental disease models. In human CD1a transgenic mice, the toll-like receptor (TLR)7 agonist imiquimod induces more pronounced splenomegaly, expansion of the peripheral blood and spleen T cell compartments, and enhanced neutrophil and eosinophil responses compared to the wild-type, accompanied by elevated skin and plasma cytokine levels, including IL-23, IL-1α, IL-1β, MCP-1 and IL-17A. Similar systemic escalation is shown in MC903-induced skin inflammation. The exacerbated inflammation could be counter-acted by CD1a-blocking antibodies, developed and screened in our laboratories. The beneficial effect is epitope dependent, and we further characterise the five best-performing antibodies for their capacity to modulate CD1a-expressing cells and ameliorate CD1a-dependent systemic inflammatory responses. In summary, we show that a therapeutically targetable CD1a-dependent pathway may play a role in the systemic spread of cutaneous inflammation.

Combination Therapy of Carnosic Acid and Methotrexate Effectively Suppressed the Inflammatory Markers and Oxidative Stress in Experimental Arthritis.

Background: Combination therapy with methotrexate (MTX) is the most common therapeutic strategy used for the treatment of patients with rheumatoid arthritis (RA). In this study, we combined the natural compound carnosic acid (CA) with MTX to reduce inflammation and oxidative stress in adjuvant arthritis (AA). Methods: AA was induced in 6–8 rats per group. MTX was administrated twice a week at a dose of 0.3 mg/kg b.w., while CA was administered daily at a dose of 100 mg/kg both in monotherapy and in combination with MTX. Plasma samples were collected on the 14th, 21st, and 28th day. Body weight and hind paw volume were measured once a week. Results: We found that, mainly, the CA + MTX combination significantly reduced the hind paw swelling, the levels of IL-17A, MMP-9, and MCP-1 in plasma, and GGT activity in joint homogenates. The mRNA expression of HO-1, catalase, and IL-1β in the liver were significantly improved by CA + MTX only. Our results indicate that adding CA to MTX treatment could be a good therapeutic option for patients suffering from RA. Conclusions: The addition of CA to methotrexate treatment significantly improved its efficacy in decreasing the development of AA by inhibiting the markers of inflammation and oxidative stress.

Type 3 innate lymphoid cells as an indicator of renal dysfunction and serum uric acid in hyperuricemia.

Type 3 innate lymphoid cells (ILC3s) are a newly identified group of innate immune cells that participate in the progression of several metabolic diseases by secreting interleukin (IL)-17 and IL-22. These cytokines are associated with hyperuricemia (HUA) severity and development; however, the relationship between ILC3s and HUA remains unclear. To determine the characteristics of circulating ILC3s in patients with HUA. Type 3 innate lymphoid cells and their subsets were detected using flow cytometry in peripheral blood mononuclear cells (PBMCs) of 80 HUA patients and 30 healthy controls (HC). Plasma levels of IL-17A and IL-22 were measured with enzyme-linked immunosorbent assay (ELISA). Clinical data of enrolled subjects were collected from electronic medical records. In patients with HUA, the frequency of circulating ILC3s was elevated and positively correlated with levels of serum uric acid and serum creatinine (Scr). Although there was no significant difference in the plasma concentration of IL-17A between the patients with HUA and healthy controls, positive correlations between plasma IL-17A and the concentration of serum uric acid and frequency of circulating ILC3s were observed in the patients with HUA. In patients with HUA, positive correlations were detected between circulating ILC3 levels, plasma IL-17A and serum uric acid. Therefore, ILC3s and IL-17A may be useful indicators of disease severity, and are potential new therapeutic targets in HUA.

Protective effect of Secukinumab on severe sepsis model rats by neutralizing IL-17A to inhibit IKBα/NFκB inflammatory signal pathway

Secukinumab is a specific neutralizing antibody for IL-17A. At present, numerous studies have confirmed the important role of IL-17A in sepsis, but the role of secukinumab in sepsis has not been studied. The present study explored the protective effect and underlying mechanism of secukinumab in severe sepsis model rats. We established a severe sepsis rat model using cecal ligation and puncture (CLP). The optimal dose of secukinumab was determined by observing the 7-day survival rate of severe sepsis model rats. The expression levels of TNF-α, IL-6, and IL-17A in plasma and lung tissue were determined by enzyme-linked immunosorbent assay. The degree of pathological damage to lung tissue was evaluated by hematoxylin–eosin (H–E) staining and pathological damage scale. The expressions of IKBα/NFκB pathway proteins and downstream-related inflammatory factors were detected by western blotting and real-time quantitative polymerase chain reaction (RT-qPCR). Our results show that high-dose secukinumab can inhibit the activation of the IKBα/NFκB inflammatory pathway by neutralizing IL-17A and reducing the gene expression of pathway-related inflammatory cytokines, thereby reducing the levels of inflammatory cytokines in lung tissue and plasma, thereby reducing the damage of lung tissue in severe sepsis model rats and improving the systemic inflammatory response.

Abstract P345: Inhibition Of Nuclear Receptor Retinoid-related Orphan Receptor γt Attenuates Interleukin-17A Production And Ameliorates Angiotensin II-induced Neuroinflammation And Hypertension In Rats

Interleukin (IL)-17A, a key inflammatory mediator produced primarily by T helper (Th) 17 cells, has been implicated in the pathogenesis of cardiovascular diseases including hypertension and heart failure. IL-17A can access the brain by disrupting blood-brain barrier (BBB) integrity to induce neuroinflammation. We previously reported that IL-17A contributes to angiotensin (ANG) II-induced hypertension via promoting neuroinflammation and sympathetic excitation. The nuclear receptor retinoid-related orphan receptor γt (RORγt) is a master transcription factor regulating Th17 cell differentiation. Here, we sought to determine whether systemic inhibition of RORγt attenuates IL-17A production and diminishes ANG II-induced neuroinflammation and hypertension. Sprague-Dawley rats received a 2-week subcutaneous (sc) infusion of ANG II (150 ng/kg/min) combined with daily sc injection of a RORγt inhibitor digoxin or vehicle (VEH). Compared with the control animals, blood pressure (162±5* vs 118±3 mmHg, *P&lt;0.05) and sympathetic tone as indicated by blood pressure change in response to ganglionic blockade (55±5* vs 23±3 mmHg) were increased in ANG II+VEH rats, which were reduced (38-41%*) in ANG II+digoxin rats. ANG II+VEH rats also had elevated RORγt mRNA in peripheral blood mononuclear cells (PBMCs: 3.38±0.70* vs 1.12±0.22) and higher IL-17A levels in plasma (23.22±5.85 * vs 4.30±0.66 pg/mL), along with increased IL-17A levels in cerebrospinal fluid (CSF: 11.89±1.91* vs 2.01±0.55 pg/mL), mRNA of IL-17A (3.02±0.48* vs 1.07±0.17), cytokines tumor necrosis factor (TNF)-α (2.78±0.45* vs 1.04±0.12) and IL-1β (2.57±0.53* vs 1.03±0.14) in the hypothalamic paraventricular nucleus (PVN), a key cardiovascular-related center in the brain. Although RORγt mRNA was unchanged in PBMCs, levels of IL-17A in plasma and CSF, mRNA of IL-17A, TNF-α and IL-1β in the PVN was reduced (41-62 %*) in ANG II+digoxin rats. Additionally, mRNA of caveolin-1 (2.26±0.39* vs 1.06±0.17), a marker of the BBB permeability, was increased in the PVN in ANG II+VEH rats and decreased (43%*) in ANG II+digoxin rats. These data suggest that RORγt inhibition improves ANG II-induced hypertension and sympathetic activation probably by reducing IL-17A production and neuroinflammation.

Helminth species dependent effects on Th1 and Th17 cytokines in active tuberculosis patients and healthy community controls.

Despite that the impact of different helminth species is not well explored, the current dogma states that helminths affect the Th1/Th2 balance which in turn affects the risk of tuberculosis (TB) reactivation and severity of disease. We investigated the influence of helminth species on cytokine profiles including IL-17A in TB patients and healthy community controls (CCs). In total, 104 newly diagnosed pulmonary TB patients and 70 HIV negative and QuantiFERON negative CCs in Gondar, Ethiopia were included following helminth screening by stool microscopy. Plasma samples and ex vivo stimulation of peripheral blood mononuclear cells (PBMCs) with purified protein derivative (PPD) and Staphylococcus enterotoxin B (SEB) was used to determine cytokine profiles by cytometric bead array. In CCs, Ascaris lumbricoides or Schistosoma mansoni infections were associated with an impaired Th1-type response (IFN-gamma, IL-6 and TNF-alpha) in PBMCs mainly with SEB stimulations, whereas in TB patients only hookworm infection showed a similar pattern. Among CCs, the IL-17A response in PBMCs stimulated with SEB was higher only for S. mansoni, whereas in TB patients, the elevated systemic IL-17A plasma level was significantly suppressed in hookworm infected TB patients compared to patients without helminth coinfection. Following treatment of TB and helminth infection there was a general decrease in ex vivio IL-10 and TNF-alpha production in unstimulated, PPD or SEB stimulated PBMCs that was the most pronounced and significant in TB patients infected with S. mansoni, whereas the follow-up levels of IFN-gamma and IL-17A was significantly increased only in TB patients without helminth coinfection from PBMCs stimulated mainly with SEB. In summary, in addition to confirming helminth specific effects on the Th1/Th2 response before and after TB treatment, our novel finding is that IL-17A was impaired in helminth infected TB patients especially for hookworm, indicating a helminth species-specific immunoregulatory effect on IL-17A which needs to be further investigated.

Pathogens and Pathogenesis in Wheezing Diseases in Children Under 6.

Few studies have comprehensively assessed the roles of cytokine production in wheezing pathogenesis. Therefore, we undertook this study to determine the association between wheezing episodes and cytokines, and to provide further information on this topic. Firstly, we retrospectively collected I176 children, including 122 subjects with first wheezing and 54 subjects with recurrent wheezing, to analyze the etiology and clinical characteristics of children with wheezing diseases. Then, we collected 52 children with wheezing diseases and 25 normal controls to detect the expression of interferon-γ (IFN-γ), interleukin-4 (IL-4), IFN-γ/IL-4, IL-17A, IL-17E, IgE, matrix metalloproteinase-3 (MMP-3), and MMP-9 in serum or plasma. The results showed that boys under 3 years old with history of allergies were more likely to develop wheezing diseases. In our cohort, M. pneumoniae caused a greater proportion of wheezing in children than expected. The expression of IgE [18.80 (13.65-31.00) vs. 17.9 (10.15-21.60)], IL-4 [24.00 (24.00-48.00) vs. 23.00 (9.50-27.00)], IFN-γ [70.59 (41.63-116.46) vs. 49.83 (29.58-81.74)], MMP3 [53.40 (20.02-128.2) vs. 30.90 (13.80-50.95)], MMP9 [148.10 (99.30-276.10) vs. 122.10 (82.20-162.35)], IL-17A [80.55 (54.46-113.08) vs. 61.11 (29.43-93.87)], and IL-17E [1.75 (0.66-2.77) vs. 1.19 (0.488-2.1615)] were significantly increased in the wheezing group (p<0.05) compared to normal controls, while the level of IFN-γ/IL-4 had no significant difference between the two groups (1.24 ± 1.88 vs 0.68 ± 0.74, p>0.05). There was altered cytokine production in children with wheezing diseases which was quite similar to asthma pathogenesis. Sex, age, pathogen infection, and inflammation in our study were also risk factors for wheezing diseases.

Myeloid cell interleukin-10 is essential for atherosclerosis protection in Apoe−/− mice.

Abstract Atherosclerosis, the disease underlying stroke, myocardial infarction and ischemic heart failure, is initiated by the subendothelial retention of lipoproteins and cholesterol, which then triggers a non-resolving inflammatory process driving plaque progression in the artery wall. Myeloid cells and in particular macrophages are the primary driver of this process. IL10 is a prototypic anti-inflammatory cytokine made by several immune cells such as macrophages. IL10 is also essential for the protection against atherosclerosis. However, it is not known which cells are responsible for the IL10 production mediating protection from atherosclerosis. Here, we crossed the VertX mouse model, in which an IRES-enhanced green fluorescent protein (eGFP) fusion protein was placed downstream of exon 5 of the IL-10 gene, to the atherosclerosis-prone Apoe−/− background. We found that myeloid cells express high levels of IL10 GFP in VertX-Apoe−/− mice. By scRNAseq we found that macrophages especially inflammatory macrophages in plaques are the main producers of IL10 in atherosclerosis. To address whether IL10 secreted by myeloid cells is essential for the protection, we developed a LyzMCreIL-10flox/flox mouse crossed in the Apoe−/− background and confirmed that macrophages were unable to secrete IL-10. We discovered that the LyzMCreIL-10flox/floxApoe−/− mice developed significantly more atherosclerosis than LyzMCreIL-10+/+Apoe−/−, both on chow and lipid-rich diet. Flow cytometry data and cytokine measurements suggest that the depletion of IL-10 in myeloid cells increases Th17 cells with elevated IL-17a, IL-17f and CCL20 in blood plasma. These data underscore the critical roles of macrophage-derived IL-10 in limiting atherosclerosis. Supported by grants NIH HL 115232, 145241, and HL088093.

Abstract 217: PCSK6 Is A Key Regulator Of Immune Status In Mice And Its Ablation Increases Atherosclerotic Plaque Burden In A Bone Marrow Transplant Model

Introduction: Proprotein convertase subtilisins/kexins (PCSKs) activate cytokines and growth factors and have been implicated in various cancers. We have previously shown that PCSK6 is a key protease modulating smooth muscle cell response in atherosclerosis, but its expression also correlated positively with typical markers of T lymphocytes and macrophages in plaques and it was localised in the proximity of these cells. Here, we hypothesized that PCSK6 may be involved in modulating inflammatory responses and aimed to elucidate its role in a hyperlipidemic mice model. Methods: Detailed immunophenotyping using histology, FACS-, OLINK- and ELISA-based analyses and primary cell cultures was used to compare Pcsk6 -/- and littermate controls. Atherosclerosis was evaluated in Ldlr -/- mice upon bone marrow transplant with Pcsk6 -/- or wild-type bone-marrow. Results: At baseline Pcks6 -/- mice showed an enrichment of pro-inflammatory cytokines Ccl2, Ccl3, Ccl20, Cxcl1 and in particular Il17a and Il17f in plasma compared controls. Spleens of Pcsk6 -/- mice had an increased number of germinal centres and FACS analysis showed that they contained significantly more CD8+ T cells. Microarray analysis of spleens from Pcsk6 -/- vs. controls confirmed that T cell markers CD4, CD3E and CD3G were upregulated. In vitro , splenocytes isolated from Pcsk6 -/- mice secreted more IFN-γ, IL-2 and IL-10 than controls upon stimulation with α-CD3 and α-CD28 antibodies. Moreover, peritoneal macrophages from Pcsk6 -/- mice secreted more TNF-α, MCP-1, IL-6 and IL-10 compared to control mice upon LPS stimulation. Interestingly, bone marrow derived macrophages from Pcsk6 -/- mice were also more prone to lipid uptake. Finally, in vivo transplantation of Pcsk6 -/- bone marrow to Ldlr -/- mice led to increased atherosclerotic plaque burden compared to controls, as quantified in the aortic root. Conclusions: PCSK6 ablation led to increased number of CD8+ T cells, as well as macrophage and cytokine activation. Transplantation of Pcsk6 -/- bone marrow resulted in an increase in atherosclerotic plaque burden compared to controls. Taken together, these results indicate that PCSK6 is a key regulator of the immune system, though the exact mechanisms involved require further investigation.

High level of plasma TILRR protein is associated with faster HIV seroconversion.

SummaryBackgroundTILRR (Toll-like Interleukin-1 Receptor Regulator) is a modulator of many genes in NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) signaling. It promotes the production of inflammatory mediators and the migration of immune cells. Recently, we showed that TILRR protein circulates in human blood. Thus, it could influence systemic inflammation. Systemic and mucosal inflammations increase the susceptibility to HIV infection. In this study, we analyzed the TILRR protein levels of the archived plasma samples of women enrolled in the Pumwani cohort to determine whether the plasma TILRR protein levels before seroconversion are correlated with differential risk of HIV seroconversion.MethodsTILRR protein of 941 archived HIV negative plasma samples from 390 women who were HIV negative at the cohort enrollment was quantified with an in-house developed multiplex bead array method. Proinflammatory cytokines/chemokines were measured using a 14-plex bead array method. Spearman rank correlation analysis was used to determine the correlation between plasma TILRR protein and proinflammatory cytokines/chemokines. Kaplan-Meier survival analysis was conducted to evaluate whether the median plasma TILRR protein levels correlate with increased risk of HIV seroconversion.FindingsThe level of plasma TILRR protein was positively correlated with plasma IL-1β (rho: 0.2593, p<0.0001), MCP-1 (rho: 0.2377, p<0.0001), and IL-17A (rho: 0.1225, p=0.0216). Women with median plasma TILRR protein levels ≥100 ng/ml seroconverted significantly faster than women with plasma TILRR protein levels <100 ng/ml (log-rank= 100.124, p<0.0001; relative risk= 3.72 and odds ratio= 15.29). Furthermore, the factors causing genital inflammation, such as STIs (sexually transmitted infections), vaginal discharge, and genital ulcers were not statistically significantly different among women with different median plasma TILRR protein levels.InterpretationThe high plasma TILRR protein levels are highly correlated with several plasma proinflammatory cytokines/chemokines. High median plasma TILRR protein (≥100 ng/ml) strongly predicted an increased risk of HIV seroconversion. Reducing plasma TILRR protein levels may reduce the risk of HIV acquisition.FundingThe study was funded by an operating grant from the Canadian Institutes of Health Research (CIHR), operating grant-PA: CHVI Vaccine Discovery and Social Research (http://www.cihr-irsc.gc.ca/e/193.html), and National Microbiology Laboratory of Canada.