Background: Cardiac diastolic function and blood pressure are physiologically tightly interconnected. While diastolic function is regulated by post-translational modifications of sarcomeric genes, including titin (TTN), isolating its genetic determinants from those influencing blood pressure is challenging due to variable in vivo loading conditions. We aimed to genetically map diastolic function to the rat genome to prioritise human gene orthologs for genotype-phenotype studies of cardiac function and clinical outcomes. Methods: High-fidelity cardiac phenotyping using ex vivo Langendorff preparation in an F2 cross of inbred rat strains included evaluation of left ventricular contractility (LV) (dP / dtmax) and relaxation (dP/dtmin), while controlling loading conditions. In UK Biobank, whole exome sequencing was performed and cardiovascular image-derived phenotypes and outcomes were compared in adult rare variant carriers and genotype-negative participants. Results: In rats, quantitative trait locus analysis identified a single locus on chromosome 3 that reached genome-wide significance for LV relaxation (dP/dtmin), which was also associated with variation in LV contractility (dP/dtmax) but not blood pressure. The Ttn gene, which encodes varied non-synonymous single-nucleotide polymorphisms of parental strains, is encoded at the peak of linkage. In humans, the prevalence of rare titin truncating variants (TTNtv) (allele frequency < 0.00004) in 454,756 UK Biobank participants was 0.37% (n = 1,672). Carrying a TTNtv was associated with an increased risk of death or major adverse cardiac events compared to genotype negative controls (hazard ratio: 1.94; 95% confidence interval [CI]: 1.78-2.12; P < 0.001), mainly due to heart failure endpoints (hazard ratio: 3.76; 95% CI: 3.30-4.29; P < 0.001). In 33,988 participants with both cardiac magnetic resonance imaging and whole exome sequencing, TTNtvs were associated with reduced diastolic function, even in participants with normal systolic function and no ventricular dilatation. Conclusions: These studies demonstrate for the first time in rats, that naturally occurring variations in Ttn are associated with diastolic function, independently of blood pressure, and in humans TTNtv cause impaired cardiac relaxation and adverse outcomes before overt cardiomyopathy develops. ### Competing Interest Statement J.S.W. has received research support from BMS, has acted as a paid advisor to Health Lumen, Tenaya Therapeutics, and Solid Biosciences, and is a founder with equity in Saturnus Bio. D.P.OR has consulted for Bayer AG and BMS, and receives grant support from Bayer AG and Calico Labs. S.A.C. is co-founder and shareholder of Enleofen Bio Pte Ltd and VVB Bio Pte Ltd. B.P.H. has consulted for AZ and Eli Lily. The remaining authors have nothing to disclose. ### Funding Statement The study was supported by Medical Research Council (MC\UP\1605/13); National Institute for Health Research (NIHR) Imperial College Biomedical Research Centre; the British Heart Foundation (RG/F/24/110138, RE/24/130023, CH/F/24/90015, FS/IPBSRF/22/27059, FS/ICRF/21/26019, NH/F/23/70013), Engineering and Physical Sciences Research Council (EP/W01842X/1, EP/Z531297/1), Rosetrees Trust and the Fetal Medicine Foundation (495237). D.P.OR., S.A.C and J.S.W. are supported by the British Heart Foundation's Big Beat Challenge award to CureHeart (BBC/F/21/220106). ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: The UK National Research Ethics Service gave ethical approval for the UK Biobank study (11/NW/0382). I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes Data from UK Biobank are available for approved research. Other data are available upon reasonable request to the authors.

For healthspan and lifespan, ERK, AMPK and mTORC1 represent critical pathways and inflammation is a centrally important hallmark1-7. Here we examined whether IL-11, a pro-inflammatory cytokine of the IL-6 family, has a negative effect on age-associated disease and lifespan. As mice age, IL-11 is upregulated across cell types and tissues to regulate an ERK-AMPK-mTORC1 axis to modulate cellular, tissue- and organismal-level ageing pathologies. Deletion of Il11 or Il11ra1 protects against metabolic decline, multi-morbidity and frailty in old age. Administration of anti-IL-11 to 75-week-old mice for 25 weeks improves metabolism and muscle function, and reduces ageing biomarkers and frailty across sexes. In lifespan studies, genetic deletion of Il11 extended the lives of mice of both sexes, by 24.9% on average. Treatment with anti-IL-11 from 75 weeks of age until death extends the median lifespan of male mice by 22.5% and of female mice by 25%. Together, these results demonstrate a role for the pro-inflammatory factor IL-11 in mammalian healthspan and lifespan. We suggest that anti-IL-11 therapy, which is currently in early-stage clinical trials for fibrotic lung disease, may provide a translational opportunity to determine the effects of IL-11 inhibition on ageing pathologies in older people.

Purpose:The critical time between stroke onset and treatment was targeted for reduction by integrating physiological imaging into the angiography suite, potentially improving clinical outcomes. The evaluation was conducted to compare C-Arm cone beam CT perfusion (CBCTP) with multi-detector CT perfusion (MDCTP) in patients with acute ischemic stroke (AIS). Approach:Thirty-nine patients with anterior circulation AIS underwent both MDCTP and CBCTP. Imaging results were compared using an in-house algorithm for CBCTP map generation and RAPID for post-processing. Blinded neuroradiologists assessed images for quality, diagnostic utility, and treatment decision support, with non-inferiority analysis (two one-sided tests for equivalence) and inter-reviewer consistency (Cohen's kappa). Results:The mean time from MDCTP to angiography suite arrival was 50 ± 34    min , and that from arrival to the first CBCTP image was 21 ± 8    min . Stroke diagnosis accuracies were 96% [93%, 97%] with MDCTP and 91% [90%, 93%] with CBCTP. Cohen's kappa between observers was 0.86 for MDCTP and 0.90 for CBCTP, showing excellent inter-reader consistency. CBCTP's scores for diagnostic utility, mismatch pattern detection, and treatment decisions were noninferior to MDCTP scores (alpha = 0.05) within 20% of the range. MDCTP was slightly superior for image quality and artifact score (1.8 versus 2.3, p < 0.01 ). Conclusions:In this small paper, CBCTP was noninferior to MDCTP, potentially saving nearly an hour per patient if they went directly to the angiography suite upon hospital arrival.

BACKGROUND:Untranslated regions (UTRs) are important mediators of post-transcriptional regulation. The length of UTRs and the composition of regulatory elements within them are known to vary substantially across genes, but little is known about the reasons for this variation in humans. Here, we set out to determine whether this variation, specifically in 5'UTRs, correlates with gene dosage sensitivity. RESULTS:We investigate 5'UTR length, the number of alternative transcription start sites, the potential for alternative splicing, the number and type of upstream open reading frames (uORFs) and the propensity of 5'UTRs to form secondary structures. We explore how these elements vary by gene tolerance to loss-of-function (LoF; using the LOEUF metric), and in genes where changes in dosage are known to cause disease. We show that LOEUF correlates with 5'UTR length and complexity. Genes that are most intolerant to LoF have longer 5'UTRs, greater TSS diversity, and more upstream regulatory elements than their LoF tolerant counterparts. We show that these differences are evident in disease gene-sets, but not in recessive developmental disorder genes where LoF of a single allele is tolerated. CONCLUSIONS:Our results confirm the importance of post-transcriptional regulation through 5'UTRs in tight regulation of mRNA and protein levels, particularly for genes where changes in dosage are deleterious and lead to disease. Finally, to support gene-based investigation we release a web-based browser tool, VuTR, that supports exploration of the composition of individual 5'UTRs and the impact of genetic variation within them.

Building a reference set of protein-coding open reading frames (ORFs) has revolutionized biological process discovery and understanding. Traditionally, gene models have been confirmed using cDNA sequencing and encoded translated regions inferred using sequence-based detection of start and stop combinations longer than 100 amino-acids to prevent false positives. This has led to small ORFs (smORFs) and their encoded proteins left un-annotated. Ribo-seq allows deciphering translated regions from untranslated irrespective of the length. In this review, we describe the power of Ribo-seq data in detection of smORFs while discussing the major challenge posed by data-quality, -depth and -sparseness in identifying the start and end of smORF translation. In particular, we outline smORF cataloguing efforts in humans and the large differences that have arisen due to variation in data, methods and assumptions. Although current versions of smORF reference sets can already be used as a powerful tool for hypothesis generation, we recommend that future editions should consider these data limitations and adopt unified processing for the community to establish a canonical catalogue of translated smORFs.

Additional file 3: Table S2. Phenotypic variances with standard errors, due to additive and dominance effects (calculated by genome-wide SNP heritability or from accumulated significant QTLs) of 242 physical traits in F2 pigs (Table S2-1), 206 physical traits of HS rats (Table S2-2) and 124 physical traits of HS mice (Table S2-3), together with average values across each trait class.

Additional file 6: Table S5. Summary of novel pleiotropic QTLs detected by AD model within each population. (Different shared QTL regions are separated by graey background and the rows with red numbers indicate novel QTLs where -log10(P)values of ADvsA Model > 5).

Background Dominance and other non-additive genetic effects arise from the interaction between alleles, and historically these phenomena play a major role in quantitative genetics. However, most genome-wide association studies (GWAS) assume alleles act additively. Results We systematically investigate both dominance—here representing any non-additive within-locus interaction—and additivity across 574 physiological and gene expression traits in three mammalian stocks: F2 intercross pigs, rat heterogeneous stock, and mice heterogeneous stock. Dominance accounts for about one quarter of heritable variance across all physiological traits in all species. Hematological and immunological traits exhibit the highest dominance variance, possibly reflecting balancing selection in response to pathogens. Although most quantitative trait loci (QTLs) are detectable as additive QTLs, we identify 154, 64, and 62 novel dominance QTLs in pigs, rats, and mice respectively that are undetectable as additive QTLs. Similarly, even though most cis-acting expression QTLs are additive, gene expression exhibits a large fraction of dominance variance, and trans-acting eQTLs are enriched for dominance. Genes causal for dominance physiological QTLs are less likely to be physically linked to their QTLs but instead act via trans-acting dominance eQTLs. In addition, thousands of eQTLs are associated with alternatively spliced isoforms with complex additive and dominant architectures in heterogeneous stock rats, suggesting a possible mechanism for dominance. Conclusions Although heritability is predominantly additive, many mammalian genetic effects are dominant and likely arise through distinct mechanisms. It is therefore advantageous to consider both additive and dominance effects in GWAS to improve power and uncover causality.

Objective Alcoholic hepatitis (AH) reflects acute exacerbation of alcoholic liver disease (ALD) and is a growing healthcare burden worldwide. Interleukin-11 (IL-11) is a profibrotic, proinflammatory cytokine with increasingly recognised toxicities in parenchymal and epithelial cells. We explored IL-11 serum levels and their prognostic value in patients suffering from AH and cirrhosis of various aetiology and experimental ALD. Design IL-11 serum concentration and tissue expression was determined in a cohort comprising 50 patients with AH, 110 patients with cirrhosis and 19 healthy volunteers. Findings were replicated in an independent patient cohort (n=186). Primary human hepatocytes exposed to ethanol were studied in vitro. Ethanol-fed wildtype mice were treated with a neutralising murine IL-11 receptor-antibody (anti-IL11RA) and examined for severity signs and markers of ALD. Results IL-11 serum concentration and hepatic expression increased with severity of liver disease, mostly pronounced in AH. In a multivariate Cox-regression, a serum level above 6.4 pg/mL was a model of end-stage liver disease independent risk factor for transplant-free survival in patients with compensated and decompensated cirrhosis. In mice, severity of alcohol-induced liver inflammation correlated with enhanced hepatic IL-11 and IL11RA expression. In vitro and in vivo, anti-IL11RA reduced pathogenic signalling pathways (extracellular signal-regulated kinases, c-Jun N-terminal kinase, NADPH oxidase 4) and protected hepatocytes and murine livers from ethanol-induced inflammation and injury. Conclusion Pathogenic IL-11 signalling in hepatocytes plays a crucial role in the pathogenesis of ALD and could serve as an independent prognostic factor for transplant-free survival. Blocking IL-11 signalling might be a therapeutic option in human ALD, particularly AH.

Human ageing is associated with metabolic dysfunction, sarcopenia and frailty that taken together reduce healthspan. For age-associated diseases and lifespan, ERK, AMPK and mTORC1 represent critical pathways, across species 1–7 . Here we examined the hypothesis that IL11, recently shown to regulate ERK/mTORC1, is an inflammaging factor important for healthspan. As mice age, IL11 is progressively upregulated in liver, skeletal muscle, and fat to stimulate an ERK/AMPK/mTORC1 axis of cellular, tissue- and organismal-level ageing pathologies. In old mice, deletion of Il11 or Il11ra1 protects against metabolic multi-morbidity, sarcopenia, and frailty. Administration of anti-IL11 therapy to elderly mice for six months reactivates an age-repressed program of white fat beiging, reverses metabolic dysfunction, restores muscle function, and reduces frailty. Across studies, inhibition of IL11 lowers epigenetic age, reduces telomere attrition, and preserves mitochondrial function. Towards clinical translation, we generated, humanised, and engineered a neutralising, high-affinity IL11 antibody. These studies identify IL11 as a key inflammaging factor and therapeutic target for mammalian healthspan.

Introduction: Marfan Syndrome (MFS) is an inherited connective tissue disorder caused by fibrillin-1 (FBN1) mutations that is associated with aortic disease. Lung abnormalities are common in MFS, but their pathogenesis is poorly understood. Here we studied IL11 in a mouse model of MFS (mMFS). Methods: We studied histological and molecular phenotypes in the lungs of Fbn1 C1041G/+ (mMFS) mice, an established model of MFS. To identify IL11-expressing lung cell types, we studied 4- and 16-week-old Fbn1 C1041G/+ : Il11 EGFP/+ reporter mice lungs by immunohistochemistry. We studied the protective effects of IL11 inhibition in the lung by RT-qPCR, immunoblots and histopathology in two models: 1) 16-week-old IL11 receptor (IL11RA) knockout mMFS mice ( Fbn1 C1041G/+ : Il11ra1 -/- ) and 2) administered IgG isotype control or anti-IL11RA (X209; 20mg/kg) antibodies twice weekly in mMFS mice between 4 to 24 weeks of age. Results: IL11 was upregulated in the lungs of mMFS mice, which exhibited progressive loss and enlargement of distal airspaces and increased extracellular matrix production. In the distal airways of young mMFS mice (4 weeks old), IL11 was specifically expressed in smooth muscle cells. In the older (16 weeks old) mMFS lungs, IL11 was expressed in smooth muscle cells, fibroblasts and type II alveolar epithelial cells. In mMFS mice, genetic ( Fbn1 C1041G/+ : Il11ra1 -/- mice) or pharmacologic (anti-IL11RA) inhibition of IL11 protected against emphysematous changes, pulmonary fibrosis and inflammation and was associated with reduced pulmonary ERK1/2 signaling and matrix metalloproteinase (MMP2 and MMP9) expression. Conclusions: IL11 causes pulmonary disease in mMFS. This adds to the recent finding that IL11 underlies aortic pathology in mMFS and provides a common disease mechanism across affected tissues in MFS. This has potential implications for treating multi-organ disease in MFS.

Translated small open reading frames (smORFs) can have important regulatory roles and encode microproteins, yet their genome-wide identification has been challenging. We determined the ribosome locations across six primary human cell types and five tissues and detected 7,767 smORFs with translational profiles matching those of known proteins. The human genome was found to contain highly cell-type- and tissue-specific smORFs and a subset that encodes highly conserved amino acid sequences. Changes in the translational efficiency of upstream-encoded smORFs (uORFs) and the corresponding main ORFs predominantly occur in the same direction. Integration with 456 mass-spectrometry datasets confirms the presence of 603 small peptides at the protein level in humans and provides insights into the subcellular localization of these small proteins. This study provides a comprehensive atlas of high-confidence translated smORFs derived from primary human cells and tissues in order to provide a more complete understanding of the translated human genome.

N-acetyl-p-aminophenol (APAP)-induced liver damage is associated with upregulation of Interleukin-11 (IL11), which is thought to stimulate IL6ST (gp130)-mediated STAT3 activity in hepatocytes, as a compensatory response. However, recent studies have found IL11/IL11RA/gp130 signaling to be hepatotoxic. To investigate further the role of IL11 and gp130 in APAP liver injury, we generated two new mouse strains with conditional knockout (CKO) of either Il11 (CKOIl11) or gp130 (CKOgp130) in adult hepatocytes. Following APAP, as compared to controls, CKOgp130 mice had lesser liver damage with lower serum Alanine Transaminase (ALT) and Aspartate Aminotransferase (AST), greatly reduced serum IL11 levels (90% lower), and lesser centrilobular necrosis. Livers from APAP-injured CKOgp130 mice had lesser ERK, JNK, NOX4 activation and increased markers of regeneration (PCNA, Cyclin D1, Ki67). Experiments were repeated in CKOIl11 mice that, as compared to wild-type mice, had lower APAP-induced ALT/AST, reduced centrilobular necrosis and undetectable IL11 in serum. As seen with CKOgp130 mice, APAP-treated CKOIl11 mice had lesser ERK/JNK/NOX4 activation and greater features of regeneration. Both CKOgp130 and CKOIl11 mice had normal APAP metabolism. After APAP, CKOgp130 and CKOIl11 mice had reduced Il6, Ccl2, Ccl5, Il1β, and Tnfα expression. These studies exclude IL11 upregulation as compensatory and establish autocrine, self-amplifying, gp130-dependent IL11 secretion from damaged hepatocytes as toxic and anti-regenerative.

BACKGROUND AND PURPOSE:The availability of cone-beam CT perfusion (CBCTP) in angiography suites may improve large-vessel occlusion (LVO) triage and reduce reperfusion times for patients presenting during extended time window. We aim to evaluate the perfusion maps correlation and agreement between multidetector CT perfusion (MDCTP) and CBCTP when obtained sequentially in patients undergoing endovascular therapy. METHODS:This is a prospective, pilot, single-arm interventional cohort study of consecutive patients with anterior circulation LVO. All patients underwent MDCTP and CBCTP prior to endovascular therapy, generating cerebral blood flow (CBF), cerebral blood volume (CBV), mean transit time (MTT), and time-to-maximum/time to peak contrast concentration maps. We compared the two imaging modalities using three different methods: (1) six regions of interest (ROIs) placed in the anterior circulation territory; (2) ROIs placed in all 10 Alberta Stroke Program Early CT Score regions; and (3) ROI drawn around the entire ischemic area. ROI ratios (unaffected/affected area) were compared for all sequences in each method. We used the intraclass correlation coefficient to calculate the correlation between the studies. Bland-Altman plots were also created to measure the degree of agreement. Finally, a sensitivity analysis was done comparing both modalities in patients with low infarct growth rate. RESULTS:Fourteen patients were included (median age 81 years [74-87], 50% males, median National Institutes of Health Stroke Scale 19 [14-22]). Median time between studies was 42 minutes (interquartile range 29-61). Independently of the method used, we found moderate to excellent correlation in CBF, CBV, and MTT between modalities. CBF correlation further improved in patients with low infarct growth. CONCLUSION:These results demonstrate promising accuracy of CBCTP in evaluating ischemic tissue in patients presenting with LVO ischemic stroke.

Interleukin-11 (IL11) is important for fibrosis and inflammation, but its role in the pancreas is unclear. In pancreatitis, fibrosis, inflammation and organ dysfunction are associated with pancreatic stellate cell (PSC)-to-myofibroblast transformation. Here, we show that IL11 stimulation of PSCs, which specifically express IL11RA in the pancreas, results in transient STAT3 phosphorylation, sustained ERK activation and PSC activation. In contrast, IL6 stimulation of PSCs caused sustained STAT3 phosphorylation but did not result in ERK activation or PSC transformation. Pancreatitis factors, including TGFβ, CTGF and PDGF, induced IL11 secretion from PSCs and a neutralising IL11RA antibody prevented PSC activation by these stimuli. This revealed an important ERK-dependent role for autocrine IL11 activity in PSCs. In mice, IL11 was increased in the pancreas after pancreatic duct ligation, and in humans, IL11 and IL11RA levels were elevated in chronic pancreatitis. Following pancreatic duct ligation, administration of anti-IL11RA to mice reduced pathologic (ERK, STAT, NF-κB) signalling, pancreatic atrophy, fibrosis and pro-inflammatory cytokine (TNFα, IL6 and IL1β) levels. This is the first description of IL11-mediated activation of PSCs, and the data suggest IL11 as a stromal therapeutic target in pancreatitis.

Motivation The creation and analysis of gene regulatory networks have been the focus of bioinformatic research and underpins much of what is known about gene regulation. However, as a result of a bias in the availability of data-types that are collected, the vast majority of gene regulatory network resources and tools have focused on either transcriptional regulation or protein-protein interactions. This has left other areas of regulation, for instance translational regulation, vastly underrepresented despite them having been shown to play a critical role in both health and disease. Results In order to address this we have developed CLIPreg, a package that integrates RNA, Ribo and CLIP-sequencing data in order to construct translational regulatory networks coordinated by RNA-binding proteins. This is the first tool of its type to be created, allowing for detailed investigation into a previously unseen layer of regulation. Availability and implementation CLIPreg is available at https://github.com/SGDDNB/CLIPreg. Contact owen.rackham@duke-nus.edu.sg or sonia.chothani@u.duke.nus.edu

The kidney has large regenerative capacity, but this is compromised when kidney damage is excessive and renal tubular epithelial cells (TECs) undergo SNAI1-driven growth arrest. Here we investigate the role of IL11 in TECs, kidney injury and renal repair. IL11 stimulation of TECs induces ERK- and p90RSK-mediated GSK3β inactivation, SNAI1 upregulation and pro-inflammatory gene expression. Mice with acute kidney injury upregulate IL11 in TECs leading to SNAI1 expression and kidney dysfunction, which is not seen in Il11 deleted mice or in mice administered a neutralizing IL11 antibody in either preemptive or treatment modes. In acute kidney injury, anti-TGFβ reduces renal fibrosis but exacerbates inflammation and tubule damage whereas anti-IL11 reduces all pathologies. Mice with TEC-specific deletion of Il11ra1 have reduced pathogenic signaling and are protected from renal injury-induced inflammation, fibrosis, and failure. In a model of chronic kidney disease, anti-IL11 therapy promotes TEC proliferation and parenchymal regeneration, reverses fibroinflammation and restores renal mass and function. These data highlight IL11-induced mesenchymal transition of injured TECs as an important renal pathology and suggest IL11 as a therapeutic target for restoring stalled endogenous regeneration in the diseased kidney.

Background: Marfan syndrome (MFS) is associated with TGF (transforming growth factor) β–stimulated ERK (extracellular signal-regulated kinase) activity in vascular smooth muscle cells (VSMCs), which adopt a mixed synthetic/contractile phenotype. In VSMCs, TGFβ induces IL (interleukin) 11) that stimulates ERK-dependent secretion of collagens and MMPs (matrix metalloproteinases). Here, we examined the role of IL11 in the MFS aorta. Methods: We used echocardiography, histology, immunostaining, and biochemical methods to study aortic anatomy, physiology, and molecular endophenotypes in Fbn1 C1041G/+ mice, an established murine model of MFS (mMFS). mMFS mice were crossed to an IL11-tagged EGFP (enhanced green fluorescent protein; Il11 EGFP/+ ) reporter strain or to a strain deleted for the IL11 receptor ( Il11ra1 −/− ). In therapeutic studies, mMFS were administered an X209 (neutralizing antibody against IL11RA [IL11 receptor subunit alpha]) or IgG for 20 weeks and imaged longitudinally. Results: IL11 mRNA and protein were elevated in the aortas of mMFS mice, as compared to controls. mMFS mice crossed to Il11 EGFP/+ mice had increased IL11 expression in VSMCs, notably in the aortic root and ascending aorta. As compared to the mMFS parental strain, double mutant mMFS: Il11ra1 −/ − mice had reduced aortic dilatation and exhibited lesser fibrosis, inflammation, elastin breaks, and VSMC loss, which was associated with reduced aortic COL1A1 (collagen type I alpha 1 chain), IL11, MMP2/9, and phospho-ERK expression. To explore therapeutic targeting of IL11 signaling in MFS, we administered either a neutralizing antibody against IL11RA (X209) or an IgG control. After 20 weeks of antibody administration, as compared to IgG, mMFS mice receiving X209 had reduced thoracic and abdominal aortic dilation as well as lesser fibrosis, inflammation, elastin breaks, and VSMC loss. By immunoblotting, X209 was shown to reduce aortic COL1A1, IL11, MMP2/9, and phospho-ERK expression. Conclusions: In MFS, IL11 is upregulated in aortic VSMCs to cause ERK-related thoracic aortic dilatation, inflammation, and fibrosis. Therapeutic inhibition of IL11, imminent in clinical trials, might be considered as a new approach in MFS.

In fibroblasts, TGFβ1 stimulates IL11 upregulation that leads to an autocrine loop of IL11-dependent pro-fibrotic protein translation. The signalling pathways downstream of IL11 are contentious and both STAT3 and ERK have been implicated. Here we show that TGFβ1- or IL11-induced ERK activation is consistently associated with fibrogenesis whereas STAT3 phosphorylation (pSTAT3) is unrelated to fibroblast activation. Surprisingly, recombinant human IL11, which has been used extensively in mouse experiments to infer STAT3 activity downstream of IL11, non-specifically increases pSTAT3 in Il11ra1 null mouse fibroblasts. Pharmacologic inhibition of STAT3 prevents TGFβ1-induced fibrogenesis but this effect was found to reflect fibroblast dysfunction due to severe proteotoxic ER stress. In contrast, inhibition of MEK/ERK prevented fibrosis in the absence of ER stress. TGFβ1-stimulated ERK/mTOR/P70RSK-driven protein translation was IL11-dependent and selectivity for pro-fibrotic protein synthesis was ascribed to an EPRS-related mechanism. In TGFβ1-stimulated fibroblasts, the anti-fibrotic drug nintedanib caused dose-dependent ER stress, reduced pSTAT/pERK and inhibited pro-fibrotic protein translation, similarly to generic STAT3 inhibitors or ER stressors. Pirfenidone, while anti-fibrotic, had no effect on ER stress whereas anti-IL11 inhibited the ERK/mTOR axis while reducing ER stress. These studies discount a specific role for STAT3 in pro-fibrotic signaling, suggest a novel mechanism of action for nintedanib and prioritise further the IL11 pathway as a therapeutic target for fibrosis.