IL-6 Revisited: From Rheumatoid Arthritis to CAR T Cell Therapy and COVID-19.

The diverse biological activity of interleukin-6 (IL-6) contributes to the maintenance of homeostasis. Emergent infection or tissue injury induces rapid production of IL-6 and activates host defense through augmentation of acute-phase proteins and immune responses. However, excessive IL-6 production and uncontrolled IL-6 receptor signaling are critical to pathogenesis. Over the years, therapeutic agents targeting IL-6 signaling, such as tocilizumab, a humanized anti-IL-6 receptor antibody, have shown remarkable efficacy for rheumatoid arthritis, Castleman disease, and juvenile idiopathic arthritis, and their efficacy in other diseases is continually being reported. Emerging evidence has demonstrated the benefit of tocilizumab for several types of acute inflammatory diseases, including cytokine storms induced by chimeric antigen receptor T cell therapy and coronavirus disease 2019 (COVID-19). Here, we refocus attention on the biology of IL-6 and summarize the distinct pathological roles of IL-6 signaling in several acute and chronic inflammatory diseases.

Single-Cell Sequencing of Peripheral Mononuclear Cells Reveals Distinct Immune Response Landscapes of COVID-19 and Influenza Patients.

The coronavirus disease 2019 (COVID-19) pandemic poses a current world-wide public health threat. However, little is known about its hallmarks compared to other infectious diseases. Here, we report the single-cell transcriptional landscape of longitudinally collected peripheral blood mononuclear cells (PBMCs) in both COVID-19- and influenza A virus (IAV)-infected patients. We observed increase of plasma cells in both COVID-19 and IAV patients and XIAP associated factor 1 (XAF1)-, tumor necrosis factor (TNF)-, and FAS-induced T cell apoptosis in COVID-19 patients. Further analyses revealed distinct signaling pathways activated in COVID-19 (STAT1 and IRF3) versus IAV (STAT3 and NFκB) patients and substantial differences in the expression of key factors. These factors include relatively increase of interleukin (IL)6R and IL6ST expression in COVID-19 patients but similarly increased IL-6 concentrations compared to IAV patients, supporting the clinical observations of increased proinflammatory cytokines in COVID-19 patients. Thus, we provide the landscape of PBMCs and unveil distinct immune response pathways in COVID-19 and IAV patients.

Berberine-loaded PLGA nanoparticles alleviate ulcerative colitis by targeting IL-6/IL-6R axis.

AIMS: The present study aims to develop a nano-delivery system that encapsulates berberine (BBR) into PLGA-based nanoparticles (BPL-NPs), to treat ulcerative colitis (UC). Furthermore, the therapeutic efficacy and molecular targeting mechanisms of BPL-NPs in the management of UC are thoroughly examined. METHODS: Emulsion solvent-driven methods were used to self-assemble BBR and PLGA into nanoparticles, resulting in the development of the nano-delivery system (BPL-NPs). The therapeutic effectiveness of BPL-NPs was evaluated using a dextran sulfate sodium (DSS)-induced model of ulcerative colitis in mice and a lipopolysaccharide (LPS)-induced model of inflammation in THP-1 macrophages. The interaction between Mφs and NCM-460 cells was investigated using a co-culture system. The molecular targeting ability of BPL-NPs in the treatment of UC was validated through in vitro as well as in vivo experiments. RESULTS: The BPL-NPs demonstrated a particle size of 184 ± 22.4 nm, enhanced dispersibility in deionized water, and a notable encapsulation efficiency of 31.1 ± 0.2%. The use of BPL-NPs clearly improved the clinical symptoms and pathological changes associated with UC in mice while also ensuring minimal toxicity. In addition, BPL-NPs improved intestinal epithelial cell apoptosis and enhanced the function of the intestinal barrier by inhibiting M1 Mφs infiltration and IL-6 signaling pathway in mice with UC. Furthermore, the BPL-NPs were found to selectively target the IL-6/IL-6R axis during the M1 Mφs-induced apoptosis of NCM460 cells. CONCLUSION: The BPL-NPs were confirmed to harbor anti-inflammatory effects both in vitro and in vivo, along with enhanced water solubility and bioactivity. In addition, the precise targeting of the IL-6/IL-6R axis was confirmed as the mechanism by which the BPL-NPs exerted therapeutic effects in UC, as demonstrated in both in vitro as well as in vivo studies.

Advancements in the study of IL-6 and its receptors in the pathogenesis of gout.

Gout is an autoinflammatory disease characterized by the deposition of monosodium urate crystals in or around the joints, primarily manifesting as inflammatory arthritis that recurs and resolves spontaneously. Interleukin-6 (IL-6) is a versatile cytokine with both anti-inflammatory and pro-inflammatory capabilities, linked to a variety of inflammatory diseases such as gouty arthritis, rheumatoid arthritis, inflammatory bowel disease, vasculitis, and several types of cancer. The rapid production of IL-6 during infections and tissue damage aids in host defense. However, excessive synthesis of IL-6 and dysregulation of its receptor signaling (IL-6R) might contribute to the pathology of diseases. Recent advancements in clinical and basic research, along with developments in animal models, have established the significant role of IL-6 and its receptors in the pathogenesis of gout, although the precise mechanisms remain to be fully elucidated. This review discusses the role of IL-6 and its receptors in gout progression and examines contemporary research on modulating IL-6 and its signaling pathways for treatment. It aims to provide insights into the pathogenesis of gout and to advance the development of targeted therapies for gout-related inflammation.

Small protein blockers of human IL-6 receptor alpha inhibit proliferation and migration of cancer cells.

BACKGROUND: Interleukin-6 (IL-6) is a multifunctional cytokine that controls the immune response, and its role has been described in the development of autoimmune diseases. Signaling via its cognate IL-6 receptor (IL-6R) complex is critical in tumor progression and, therefore, IL-6R represents an important therapeutic target. METHODS: An albumin-binding domain-derived highly complex combinatorial library was used to select IL-6R alpha (IL-6Rα)-targeted small protein binders using ribosome display. Large-scale screening of bacterial lysates of individual clones was performed using ELISA, and their IL-6Rα blocking potential was verified by competition ELISA. The binding of proteins to cells was monitored by flow cytometry and confocal microscopy on HEK293T-transfected cells, and inhibition of signaling function was examined using HEK-Blue IL-6 reporter cells. Protein binding kinetics to living cells was measured by LigandTracer, cell proliferation and toxicity by iCELLigence and Incucyte, cell migration by the scratch wound healing assay, and prediction of binding poses using molecular modeling by docking. RESULTS: We demonstrated a collection of protein variants called NEF ligands, selected from an albumin-binding domain scaffold-derived combinatorial library, and showed their binding specificity to human IL-6Rα and antagonistic effect in HEK-Blue IL-6 reporter cells. The three most promising NEF108, NEF163, and NEF172 variants inhibited cell proliferation of malignant melanoma (G361 and A2058) and pancreatic (PaTu and MiaPaCa) cancer cells, and suppressed migration of malignant melanoma (A2058), pancreatic carcinoma (PaTu), and glioblastoma (GAMG) cells in vitro. The NEF binders also recognized maturation-induced IL-6Rα expression and interfered with IL-6-induced differentiation in primary human B cells. CONCLUSION: We report on the generation of small protein blockers of human IL-6Rα using directed evolution. NEF proteins represent a promising class of non-toxic anti-tumor agents with migrastatic potential.

No Association of Polymorphisms in the Genes Encoding Interleukin-6 and Interleukin-6 Receptor Subunit Alpha with the Risk of Keloids in Polish Patients.

A keloid is a benign fibroproliferative hypertrophy of scar tissue that extends outside the original wound and invades adjacent healthy skin. Keloid formation is thought to be a complex process including overactivity of the interleukin-6 signaling pathway and genetic susceptibility. The aim of the study was to investigate possible associations between rs1800797, rs1800796, and rs1800795 polymorphisms in the promoter of the IL6 gene encoding interleukin-6 and the rs2228145 polymorphism in the IL6R gene encoding the interleukin-6 receptor subunit alpha with the predisposition to keloids in Polish patients. The genetic polymorphisms were identified either using Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) or sequencing of samples of genomic DNA extracted from blood leukocytes of 86 adult patients with keloids and 100 newborns comprising a control group. No significant differences in the distributions of IL6 or IL6R alleles or genotypes were found between keloid patients and newborn controls. There were also no significant differences between both groups in the distribution of IL6 haplotypes. The IL6 rs1800797, rs1800796 and rs1800795 and IL6R rs2228145 polymorphisms were not found to predispose individuals in the study group to keloids. IL6 promoter haplotypes were not found to be associated with a higher risk of keloids in the studied group.

IL-6 upregulates the expression of IL-6R through the JAK2/STAT3 signalling pathway to promote progression of hepatocellular carcinoma.

The progression of hepatocellular carcinoma (HCC) involves multifactor, multistep interactions. High expression of interleukin-6 receptor (IL-6R) plays an important role in the occurrence and development of tumours, but the regulatory mechanism of IL-6R expression and its function in HCC have not been fully defined. Western blot was used to evaluate the phosphorylation of key kinases in the JAK2/STAT3 pathway and the protein expression levels of related proliferation molecules, migration molecules and apoptotic molecules. The antiapoptosis, migration and proliferation of cells of each group were analysed with JC-1 to judge the cell apoptosis rate, the EdU method to determine the proliferation vitality of the cells, clone formation experiments and Transwell experiments. High expression of IL-6R in cell lines, lower protein levels of the apoptotic molecules c-Caspase7 and c-Caspase3 and higher protein levels of the proliferative molecules p-P70S6K and migration molecules MMP9 and MMP2 were consistent with stronger antiapoptosis, proliferation and migration. Interestingly, IL-6 upregulated the expression of IL-6R by activating the JAK2/STAT3 signalling pathway. Also, the expression of IL-6R protein was downregulated after lentivirus knockdown of STAT3. In nude mice bearing subcutaneous tumours, upregulation of IL-6R expression after activation of the JAK2/STAT3 signalling pathway by IL-6 significantly increased tumour growth. Moreover, the expression of IL-6R protein was downregulated, and the terminal tumour volume was significantly downregulated in the lentiviral STAT3 knockdown group. IL-6 regulated the transcription of IL-6R through the activation of the JAK2/STAT3 signalling pathway.

Janus kinase 1 in Megalobrama amblycephala: Identification, phylogenetic analysis and expression profiling after Aeromonas hydrophila infection.

Janus kinase 1 (JAK1), a member of the JAK family, plays an essential and non-redundant role in the mammalian immune system. However, the potential role of JAK1 in fish immune response remains largely unclear. In the present study, the JAK1 gene of Megalobrama amblycephala (MamJAK1) was identified and characterized. The open reading frame (ORF) of MamJAK1 was 3462 bp, encoding 1153 amino acids. MamJAK1 consists of four common domains of the JAK family, including B41, SH2, STyrKc (a pseudo kinase domain), and TyrKc (a kinase domain). Phylogenetic analysis showed that JAK1s are divided into two evolutionary clades, one containing fish JAK1s, and the other containing JAK1s from other vertebrates. The results of quantitative real-time PCR (qPCR) showed that in healthy M. amblycephala, MamJAK1 mRNA was highest expressed in blood, followed by spleen, intestine and mid-kidney, and lowly expressed in other tissues including gill, liver, head kidney, muscle, brain and heart. After Aeromonas hydrophila infection, the expression of MamJAK1 mRNA was significantly induced in four selected tissues including spleen, mid-kidney, liver and intestine, reaching a peak at 24 hpi (hour post infection) in spleen and mid-kidney, at 12 hpi in liver and at 4 hpi in intestine, and then the expression level was restricted to control levels at 72 or 120 hpi. In addition, the results of Western blot showed that the phosphorylation level of MamJAK1 protein in spleen and mid-kidney increased significantly after A. hydrophila infection, although MamJAK1 protein did not change obviously. Further, the JAK1 phosphorylation in Ctenopharyngodon idellus kidney (CIK) cells was found to be significantly induced by LPS stimulation and IL-6R over-expression. The results above suggest that MamJAK1 may play an essential role in the immune response against bacterial infection through the IL-6R mediated JAK1/STAT signaling pathway, which further deepen our understanding of JAK1 and provides a potential target for the treatment and prevention of bacterial diseases in teleost.

Molecular Design of Novel Inhibitor by Targeting IL-6Rα using Combined Pharmacophore and Experimentally Verified Plant Products with Scaffold-Hopping Techniques: A Dual Therapeutic Strategy for COVID-19 and Cancer.

The IL-6/IL-6R/gp130 complex serves as a significant indicator of cytokine release syndrome in COVID-19 and chronic inflammation, increasing the risk of cancer. Therefore, we identified IL-6Rα as a potential target to block gp130 interaction. Notably, there has been no reception of approval for an orally available drug to serve this purpose, to date. In this study, we targeted IL-6Rα to inhibit IL-6Rα/gp130 interaction. The selection of the lead candidate L821 involved the amalgamation of three drug discovery approaches. This library was screened employing tertiary structure-based pharmacophore models followed by molecular docking models, scaffold-hopping, MM/PBSA as well as MM/GBSA analysis, and assessments of pKi and ADMET properties. After evaluating the binding interactions with key amino acids, 15 potential ligands were chosen, with the top ligand undergoing further investigation by means of molecular dynamics simulations. Considering the stability of the complexes, the strong interactions observed between ligand and residues of IL-6Rα/gp130, and the favorable binding free energy calculations, L821 emerged as the prime candidate for inhibiting IL-6Rα. Notably, L821 exhibited a docking-based binding affinity of -9.5 kcal/mol. Our study presents L821 as a promising inhibitor for future in vitro analysis, potentially combatting SARS-CoV-2-related cytokine storms and serving as an oncogenic drug therapy.

Blood PD-1+TFh and CTLA-4+CD4+ T cells predict remission after CTLA-4Ig treatment in early rheumatoid arthritis.

OBJECTIVE: Treatment with CTLA-4Ig blocks T-cell activation and is clinically effective in RA. However, it is unknown if specific CD4+ T-cell subsets in blood at baseline predict remission after CTLA-4Ig, or other biological treatments with different modes of action, and how treatment affects CD4+ T cells in patients with untreated early RA (eRA). METHODS: This study included 60 patients with untreated eRA from a larger randomized trial. They were treated with methotrexate combined with CTLA-4Ig (abatacept, n = 17), anti-IL6 receptor (tocilizumab, n = 21) or anti-TNF (certolizumab-pegol, n = 22). Disease activity was assessed by clinical disease activity index (CDAI), DAS28, swollen joint counts, tender joint counts, CRP and ESR. The primary outcome was CDAI remission (CDAI ≤ 2.8) at week 24. Proportions of 12 CD4+ T-cell subsets were measured by flow cytometry at baseline and after 4, 12 and 24 weeks of treatment. RESULTS: In patients treated with CTLA-4Ig, the proportions of PD-1+TFh and CTLA-4+ conventional CD4+ T cells at baseline predicted CDAI remission at week 24. CD4+ T-cell subset proportions could not predict remission after treatment with anti-IL6R or anti-TNF. The percentage of regulatory T cells (Tregs) expressing CTLA-4 decreased in all treatment arms by 24 weeks, but only CTLA-4Ig treatment significantly reduced the proportions of Tregs and PD-1+T follicular helper (TFh) cells. CONCLUSION: These findings indicate that circulating proportions PD-1+TFh and CTLA-4+ conventional CD4+ T cells at baseline may serve as predictive biomarkers for remission in early RA after CTLA-4Ig treatment.

Eukaryotic translation initiation factor 3 subunit B promotes head and neck cancer via CEBPB translation.

BACKGROUND: Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer type worldwide. Deregulation of mRNA translation is a frequent feature of cancer. Eukaryotic translation initiation factor 3 subunit B (EIF3B) has been reported as an oncogene; however, its role in HNSCC has yet to be fully elucidated. METHODS: In this study, the clinical significance of EIF3B expression was analyzed based on TCGA datasets. Then, EIF3B expression was knocked down and its role in HNSCC was revealed. To explore the molecular mechanisms of EIF3B, we applied RNA sequencing and proteomics and acquired deregulated pathways. RNA immunoprecipitation (RIP) sequencing was conducted to reveal the target mRNAs of EIF3B, and TCGA datasets were used to validate potential targets of EIF3B. RESULTS: Elevated expression of EIF3B was observed in the HNSCC cancer samples. The expression of EIF3B was significantly correlated with the patient's sex, age, HPV infection status, T stage, N stage, perineural invasion status and survival status. EIF3B serves as a marker of an unfavorable HNSCC prognosis. EIF3B-silenced Fadu and Cal27 cells exhibited reduced cell numbers, and EIF3B knockdown induced apoptosis in both cell lines. The EIF3B-silenced cells demonstrated decreased invasion and migration capabilities, and the EIF3B knockdown group mice showed significantly decreased tumor volumes. The results show that EIF3B promotes CEBPB translation and activates the MAPK pathway and revealed that IL6R and CCNG2 are targets of EIF3B-regulated CEBPB translation. CONCLUSION: In summary, the results indicated that EIF3B is a novel oncogene in HNSCC that promotes CEBPB translation and IL6R expression, and these findings provide a link between the molecular basis and pathogenesis of HNSCC.

Resolvin D1 suppresses inflammation-associated tumorigenesis in the colon by inhibiting IL-6-induced mitotic spindle abnormality.

While failure in resolution of inflammation is considered to increase the risk of tumorigenesis, there is paucity of experimental as well as clinical evidence supporting this association. Resolvin D1 (RvD1) is a representative pro-resolving lipid mediator that is endogenously generated from docosahexaenoic acid for the resolution of inflammation. Here, we report a decreased level of RvD1 in the blood from colorectal cancer patients and mice having inflammation-induced colon cancer, suggesting plasma RvD1 as a potential biomarker for monitoring colorectal cancer. Administration of RvD1 attenuated dextran sodium sulfate (DSS)-induced colitis and azoxymethane (AOM) plus DSS-induced colorectal carcinogenesis by suppressing the production of interleukin-6 (IL-6) and IL-6-mediated chromosomal instability. The protective effect of RvD1 against chromosomal instability is associated with downregulation of IL-6-induced Cyclin D1 expression, which appears to be mediated by blocking the Janus kinase 2 (JAK2)-signal transducer and activator of transcription 3 (STAT3) axis. RvD1 inhibited the STAT3 signaling pathway by interfering with the binding of IL-6 to its receptor (IL-6R), suggesting the novel function of RvD1 as a putative IL-6R antagonist. Together, our findings suggest that RvD1-mediated blockade of IL-6 signal transmission may contribute to inhibition of chromosomal instability and tumorigenesis.

Phosphorylation of meprin ß controls its cell surface abundance and subsequently diminishes ectodomain shedding.

Meprin ß is a zinc-dependent metalloprotease exhibiting a unique cleavage specificity with strong preference for acidic amino acids at the cleavage site. Proteomic studies revealed a diverse substrate pool of meprin ß including the interleukin-6 receptor (IL-6R) and the amyloid precursor protein (APP). Dysregulation of meprin ß is often associated with pathological conditions such as chronic inflammation, fibrosis, or Alzheimer's disease (AD). The extracellular regulation of meprin ß including interactors, sheddases, and activators has been intensively investigated while intracellular regulation has been barely addressed in the literature. This study aimed to analyze C-terminal phosphorylation of meprin ß with regard to cell surface expression and proteolytic activity. By immunoprecipitation of endogenous meprin ß from the colon cancer cell line Colo320 and subsequent LC-MS analysis, we identified several phosphorylation sites in its C-terminal region. Here, T694 in the C-terminus of meprin ß was the most preferred residue after phorbol 12-myristate 13-acetate (PMA) stimulation. We further demonstrated the role of protein kinase C (PKC) isoforms for meprin ß phosphorylation and identified the involvement of PKC-α and PKC-ß. As a result of phosphorylation, the meprin ß activity at the cell surface is reduced and, consequently, the extent of substrate cleavage is diminished. Our data indicate that this decrease of the surface activity is caused by the internalization and degradation of meprin ß.

miR-451a targeting IL-6R activates JAK2/STAT3 pathway, thus regulates proliferation and apoptosis of multiple myeloma cells.

OBJECTIVES: To investigate the effects of miR-451a targeting interleukin-6 (IL-6) on the proliferation and apoptosis of multiple myeloma (MM) cells and its potential mechanism via JAK2/STAT3 pathway. METHODS: mRNA expression of miR-451a and IL-6R in the plasma of patients with MM and normal controls were determined by RT-qPCR. U266 cells were cultured, transfected with miR-451a mimics, the proliferative ability of U266 cells was determined by CCK-8. Potential targets of miR-451a were predicted with the biological software TargetScan, and the direct relationship between miR-451a and the target IL-6R was analyzed by a dual-luciferase reporter assay. U266 cells were stimulated with IL-6R (100 ng/ml), and the proliferative ability and apoptosis rate were determined by CCK-8 and flow cytometry after 48h. RESULTS: In the plasma of patients with MM, miR-451a expression was low and IL-6R expression was high. miR-451a targeted and negatively regulated IL-6R. Overexpressing miR-451a inhibited the proliferation and promoted the apoptosis of U266 cells. IL-6R acting on U266 cells promoted the proliferation and inhibited the apoptosis of U266 cells. Overexpressing miR-451a inhibited the activation of JAK2/STAT3 pathway and down-regulating miR-451a promoted the activation of JAK2/STAT3 pathway. CONCLUSIONS: miR-451a targeting IL-6R activates JAK2/STAT3 pathway, thus regulates the proliferation and apoptosis in MM cells.

Deficiency of interleukin-6 receptor ameliorates PM2.5 exposure-induced pulmonary dysfunction and inflammation but not abnormalities in glucose homeostasis.

BACKGROUND: Ambient fine particulate matter (PM2.5) exposure increases local and systemic interleukin-6 (IL-6). However, the pathogenic role of IL-6 signalling following PM2.5 exposure, particularly in the development of pulmonary dysfunction and abnormal glucose homeostasis, has hardly been investigated. RESULTS: In the study, IL-6 receptor (IL-6R)-deficient (IL-6R-/-) and wildtype littermate (IL-6R+/+) mice were exposed to concentrated ambient PM2.5 (CAP) or filtered air (FA), and their pulmonary and metabolic responses to these exposures were analyzed. Our results demonstrated that IL-6R deficiency markedly alleviated PM2.5 exposure-induced increases in lung inflammatory markers including the inflammation score of histological analysis, the number of macrophages in bronchoalveolar lavage fluid (BALF), and mRNA expressions of TNFα, IL-1ß and IL-6 and abnormalities in lung function test. However, IL-6R deficiency did not reduce the hepatic insulin resistance nor systemic glucose intolerance and insulin resistance induced by PM2.5 exposure. CONCLUSION: Our findings support the crucial role of IL-6 signalling in the development of pulmonary inflammation and dysfunction due to PM2.5 exposure but question the putative central role of pulmonary inflammation for the extra-pulmonary dysfunctions following PM2.5 exposure, providing a deep mechanistic insight into the pathogenesis caused by PM2.5 exposure.

Classical Signaling and Trans-Signaling Pathways Stimulated by Megalobrama amblycephala IL-6 and IL-6R.

Interleukin-6 (IL-6) is a multipotent cytokine. IL-6 plays a dual role in inflammation through both classical signaling (IL-6 binds membrane IL-6 receptor/IL-6R) and trans-signaling (IL-6 binds soluble IL-6R). However, the regulation of IL-6 activity, especially the regulation of signaling pathways and downstream genes mediated by IL-6 trans-signaling, remains largely unclear in teleost. Grass carp (Ctenopharyngodon idellus) hepatic (L8824) cells, kidney (CIK) cells, and primary hepatocytes were used as test models in this study. First, the biological activity of recombinant blunt snout bream (Megalobrama amblycephala) IL-6 (rmaIL-6) and sIL-6R (rmasIL-6R) was verified by quantitative PCR (qPCR) and western blot. The western blot results showed that rmaIL-6 significantly upregulated signal transducer and activator of transcription 3 (STAT3) phosphorylation in L8824 cells and primary hepatocytes, while rmaIL-6 in combination with rmasIL-6R (rmaIL-6+rmasIL-6R) significantly upregulated STAT3 phosphorylation in all types of cells. Furthermore, maIL-6 and maIL-6+rmasIL-6R could only induce extracellular-signal-regulated kinase 1/2 (ERK1/2) phosphorylation in L8824 cells and CIK cells, respectively. Therefore, IL-6 mainly acts by activating the janus kinase (JAK)/STAT3 pathway rather than the mitogen-activated protein kinase (MEK)/ERK pathway. Finally, the activation of the JAK2/STAT3 pathway was shown to be essential for the generation of socs3a and socs3b induced by IL-6 trans-signaling after treatment by JAK2/STAT3 pathway inhibitors (c188-9 and TG101348). These findings provide functional insights into IL-6 classical signaling and trans-signaling regulatory mechanisms in teleost, enriching our knowledge of fish immunology.

The informative value of CD3+CD4+ and CD3+CD8+ T-cell count and cHIS scale as predictors of severe COVID-19 when using interleukin-6 receptor blockers in the in-hospital setting.

BACKGROUND: Clinical and laboratory signs of hyperinflammatory response in COVID-19 may serve as prognostic markers of the disease scenario. In real-world practice, there is an unmet need to determine the optimal timing of identifying predictors of SARS-CoV-2 adverse outcomes in the context of patient stratification to improve the effectiveness of anti-IL-6R therapy. Lymphopenia has a high informative value for the adverse prognosis of the COVID-19 course; however, the informative value of CD3+CD4+, CD3+CD8+ T-cell count remains questionable. In addition to lymphocyte phenotyping, a six-criterion additive scale (cHIS) was used in the study. AIM: To study the informative value of CD3+CD4+, CD3+CD8+ T-cell phenotyping and cHIS scale as predictors of severe COVID-19 when using IL-6R blockers. MATERIALS AND METHODS: A single-center, bi-directional study included 179 patients with SARS-CoV-2-induced community-acquired pneumonia with severe acute inflammation and progressing respiratory failure. Data were obtained from electronic patient records. Anti-IL-6R was administered in addition to standard therapy in the cohorts. The following disease outcomes were used to determine the informative value of the studied parameters: mortality and hospital discharge. Inflammatory markers were measured before and after administering anti-IL-6R, followed by monitoring. Statistical analysis was performed using SPSS (version 25.0). The quantitative indices were described using the median and interquartile range. Quantitative indices were compared using nonparametric methods: Mann-Whitney U-test, Kruskal-Wallis test. The groups were compared by qualitative characteristics using Pearson's chi-square test. Correlation analysis of quantitative indicators was performed using Spearman rank correlation. For additional analysis of the cHIS scale, odds ratio and decision tree methods were used. Differences were considered statistically significant at р≤0,05. RESULTS: Immunophenotyping of lymphocytes as a predictor of the severe SARS-CoV-2 requires further research. The cHIS scale may be implemented in routine clinical practice due to its high predictive value. A cHIS score of ≥2 on the first day of admission is a critical threshold for intensification and revision of therapy. The prognosis with cHIS is logically relevant in the first three days of hospitalization. CONCLUSION: The main result of the study is the definition of target groups of patients with community-acquired SARS-CoV-2 pneumonia for the IL-6R-blockers, considering the timing of their effective use in real clinical practice.

Circ_0023404 sponges miR-136 to induce HK-2 cells injury triggered by hypoxia/reoxygenation via up-regulating IL-6R.

The significance of circular RNAs (circRNAs) is reported in various kidney diseases including acute kidney injury (AKI). Specific circRNAs have the capacity to function as novel indicators of AKI. Circ_0023404 exhibits an important role in several diseases. Nevertheless, the detailed biological role of circ_0023404 in AKI remains poorly known. The present study aimed to investigate the effect of circ_0023404 on renal ischaemia/reperfusion (I/R) injury in vitro. Here, we evaluated the function of circ_0023404 in HK-2 cells in response to hypoxia/reoxygenation (H/R). We established a cell AKI model induced by H/R in HK-2 cells. We found circ_0023404 was significantly increased in AKI. Then, we found loss of circ_0023404 increased cell growth, repressed apoptosis, reduced inflammatory factors secretion and oxidative stress generation in vitro. Besides, circ_0023404 sponged miR-136. miR-136 overturned the effects of circ_0023404 on HK-2 cell injury. We assumed IL-6 receptor (IL-6R) as a target of miR-136 and IL-6R was activated by circ_0023404 via sponging miR-136. In conclusion, we revealed circ_0023404 contributed to HK-2 cells injury stimulated by H/R via sponging miR-136 and activating IL-6R.

Molecular dynamics analysis of the binding of human interleukin-6 with interleukin-6 α-receptor.

Human interleukin-6 (hIL-6) is a multifunctional cytokine that regulates immune and inflammatory responses in addition to metabolic and regenerative processes and cancer. hIL-6 binding to the IL-6 receptor (IL-6Rα) induces homodimerization and recruitment of the glycoprotein (gp130) to form a hexameric signaling complex. Anti-IL-6 and IL-6R antibodies are clinically approved inhibitors of IL-6 signaling pathway for treating rheumatoid arthritis and Castleman's disease, respectively. There is a potential to develop novel small molecule IL-6 antagonists derived from understanding the structural basis for IL-6/IL-6Rα interactions. Here, we combine homology modeling with extensive molecular dynamics (MD) simulations to examine the association of hIL-6 with IL-6Rα. A comparison with MD of apo hIL-6 reveals that the binding of hIL-6 to IL-6Rα induces structural and dynamic rearrangements in the AB loop region of hIL-6, disrupting intraprotein contacts and increasing the flexibility of residues 48 to 58 of the AB loop. In contrast, due to the involvement of residues 59 to 78 in forming contacts with the receptor, these residues of the AB loop are observed to rigidify in the presence of the receptor. The binary complex is primarily stabilized by two pairs of salt bridges, Arg181 (hIL-6)- Glu182 (IL-6Rα) and Arg184 (hIL-6)- Glu183 (IL-6Rα) as well as hydrophobic and aromatic stacking interactions mediated essentially by Phe residues in both proteins. An interplay of electrostatic, hydrophobic, hydrogen bonding, and aromatic stacking interactions facilitates the formation of the hIL-6/IL-6Rα complex.

Effectiveness and safety of intravenous tocilizumab to treat COVID-19-associated hyperinflammatory syndrome: Covizumab-6 observational cohort.

Although the starting event in COVID-19 is a viral infection some patients present with an over-exuberant inflammatory response, leading to acute lung injury (ALI) and adult respiratory distress syndrome (ARDS). Since IL-6 plays a critical role in the inflammatory response, we assessed the efficacy and safety of tocilizumab (TCZ) in this single-centre, observational study in all Covid-19 in-patient with a proven SARS-CoV-2 rapidly progressing infection to prevent ALI and ARDS. 104 patients with COVID-19 treated with TCZ had a lower mortality rate (5·8%) compared with the regional mortality rate (11%), hospitalized patient's mortality (10%), and slightly lower than hospitalized patients treated with our standard of care alone (6%). We found that TCZ rapidly decreased acute phase reactants, ferritin and liver release of proteins. D-Dimer decreased slowly. We did not observe specific safety concerns. Early administration of IL6-R antagonists in COVID-19 patients with impending hyperinflammatory response, may be safe and effective treatment to prevent, ICU admission and further complications.