Idiopathic pulmonary fibrosis (IPF) is a disease with a poor prognosis and no curative therapies. Fibroblast activation by transforming growth factor β1 (TGFβ1) and disrupted metabolic pathways, including the arginine–polyamine pathway, play crucial roles in IPF development. Polyamines are agonists of the calcium/cation-sensing receptor (CaSR), activation of which is detrimental for asthma and pulmonary hypertension, but its role in IPF is unknown. To address this question, we evaluated polyamine abundance using metabolomic analysis of IPF patient saliva. Furthermore, we examined CaSR functional expression in human lung fibroblasts (HLFs), assessed the anti-fibrotic effects of a CaSR antagonist, NPS2143, in TGFβ1-activated normal and IPF HLFs by RNA sequencing and immunofluorescence imaging, respectively; and NPS2143 effects on polyamine synthesis in HLFs by immunoassays. Our results demonstrate that polyamine metabolites are increased in IPF patient saliva. Polyamines activate fibroblast CaSR in vitro, elevating intracellular calcium concentration. CaSR inhibition reduced TGFβ1-induced polyamine and pro-fibrotic factor expression in normal and IPF HLFs. TGFβ1 directly stimulated polyamine release by HLFs, an effect that was blocked by NPS2143. This suggests that TGFβ1 promotes CaSR activation through increased polyamine expression, driving a pro-fibrotic response. By halting some polyamine-induced pro-fibrotic changes, CaSR antagonists exhibit disease-modifying potential in IPF onset and development.

Progressive fibrosing interstitial lung diseases (PFILDs) cause substantial morbidity and mortality. Antifibrotic agents slow progression, but most of the clinical need remains unmet. The archetypal PFILD is idiopathic pulmonary fibrosis (IPF). Chronic progression is driven by transforming growth factor (TGF-)β1 signalling. It is punctuated by inflammatory flares known as acute exacerbations (AE-IPF), which are associated with accelerated decline and high mortality. We hypothesized that acute injury responses underlying exacerbations and the mechanisms of chronic fibrosis overlap at the molecular level, via a cell surface assembly nucleated by galectin-3 that we term the ‘gal-3-fibrosome’. We focused upon a putative pro-inflammatory galectin-3 ligand, the CD98:integrin complex. Our data indicate CD98 and β1-integrin co-localise with galectin-3 within epithelial cells in IPF lung tissue, and within 40 nm in human lung tissue treated with TGF-β1 compared to controls. CD98 is required for interleukin (IL-)6 and IL-8 responses to biochemical and biophysical conditions mimicking stimuli of AE-IPF in vivo , ex vivo and in cells, and for an interstitial neutrophilic response in a mouse model. We demonstrate this pathway progresses via intracellular influx of Ca 2+ mediated by TRPV4, and NF-κB activation, operating in positive feedback. Lastly we show the CD98- and galectin-3-dependence of IL-6 and IL-8 responses to the SARS-CoV-2 spike protein receptor binding domain and the conservation of this response pattern between lung epithelial cells and monocyte-derived macrophages. Taken together our findings identify CD98 as a key mediator of both pro-fibrotic and acute inflammatory responses in the lung with relevance to AE- and chronic progression of IPF, and the priming of fibrotic lungs for acute inflammatory responses. They similarly implicate CD98 and galectin-3 as mediators of COVID pneumonitis and worse outcomes in ILD patients with COVID.

In this review article we present the evidence to date supporting the role of the calcium-sensing receptor (CaSR) as a key, pluripotential molecular trigger for asthma and speculate on the likely benefits of topical therapy of asthma with negative allosteric modulators of the CaSR: calcilytics.

International public health policies increasingly favor mandatory immunization. If its short-term effects on vaccine coverage are well documented, there has been little consideration to its effects on public attitudes towards vaccines. In this paper, we examine Google searches related to vaccines in five countries (Australia, France, Germany, Italy, Serbia) and two American states (California) which experienced at least one vaccine mandate extension in the past decade. We found that the effects of a new mandate implementation heavily depends on the context in each specific country or state. We also observed that there is little indication that the passing of new or extended mandates attenuated public doubt towards vaccines.

Introduction Little is known about the impact of mandatory vaccination on people who are reluctant to be vaccinated, despite the potential importance in terms of public health policy. Objective We aimed to explore the relationship between vaccine hesitancy and onset, severity and characteristics of self-reported adverse events. Methods We used a cross-sectional online survey conducted in 2021 among a representative sample of the French population aged 18 years and older (n = 1593). All reported adverse events were analyzed and categorized by trained experts in drug safety and pharmacovigilance. Multivariate binomial regressions on the onset of self-reported adverse events, vaccine hesitancy categories and predefined responders' characteristics were performed. Results Overall, 590 (37.0%) participants reported at least one adverse event, with 121 (20.5%) considered it to be severe. Proportions of reported adverse events, ranging from 18% (in non-reluctant responders) to 65% (in very reluctant responders), and their severity, ranging from 5% (non-reluctant responders) to 41% (very reluctant responders), depended highly on attitudes toward vaccination. The adverse events profile remained similar between groups. In the multivariate regression model, beyond attitude toward vaccination, younger age and female gender were significantly associated with higher reporting of vaccine adverse events. Conclusions Our results suggest that vaccine hesitancy could be a major driver of patient-reported vaccine-related adverse events and their perceived severity. In this context, vaccinators must pay special attention to reluctant patients and inform them on the possible nocebo nature of these adverse events so as to prevent them.

Idiopathic pulmonary fibrosis (IPF) is a fatal, currently incurable interstitial lung disease. Central to its progression is the activation of fibroblasts by TGFβ, initiating a plethora of fibrotic processes. Here, we investigate a potential role for the extracellular calcium-sensing receptor (CaSR) in mediating TGFβ-induced expression of pro-fibrotic genes in primary human lung fibroblasts (HLFs) using a negative allosteric modulator (NAM) of the CaSR. Primary HLFs were treated with TGFβ in the presence/absence of NAM or NAM alone for 72 hours. RNA sequencing was used to overview their genomic phenotypes. Analysis of the transcriptomic profile of TGFβ-treated HLFs revealed regulation of genes and biological pathways known to be altered in IPF fibroblasts. We identified 4756 genes the expression of which was up- (2757) or down- (1999) regulated by TGFβ compared with vehicle, reduced to 1904 genes up- (1033) or down- (871) regulated in the additional presence of the NAM, which exerted no effect in isolation. TGFβ-activated HLFs up-regulated many genes associated with key metabolic processes such as glycolysis, glutamine metabolism and polyamine homeostasis, as well as a range of key, pro-fibrotic genes implicated in the pathogenesis of IPF including αSMA, Collagens (I-VII), MMPs and TIMPs. Pharmacological blockade of CaSR re-established baseline expression of most of the altered metabolic and pro-fibrotic genes, consistent with the hypothesis that the CaSR plays a key role in TGFβ-induced fibroblast activation and thereby the pathogenesis of IPF.

Removing or extracting the commentary sections from a series of websites is a tedious task, as no standard way to code them is widely adopted. This operation is thus very rarely performed. In this paper, we show that these commentary sections can induce significant biases in the analyses, especially in the case of controversial Highlights $\bullet$ Commentary sections can induce biases in the analysis of websites' contents $\bullet$ Analyzing these sections can be interesting per se. $\bullet$ We illustrate these points using a corpus of anti-vaccine websites. $\bullet$ We provide guidelines to remove or extract these sections.

Background Idiopathic pulmonary fibrosis (IPF) is a disease with very poor prognosis and no curative therapies. The G protein-coupled, calcium/cation-sensing receptor (CaSR) is activated by environmental pollutants and by arginine-derived polyamines, which are thought to play a role in IPF. Whether the CaSR is involved in the pathogenesis of pulmonary fibrosis is unknown. Objective To investigate the CaSR as a novel drug target for the treatment of pulmonary fibrosis (PF). Methods and results CaSR protein expression is found in the airway epithelium in the neuroepithelial bodies of the healthy and IPF human lung. Expression of arginine pathway-linked polyamines is increased in PF patient saliva samples compared to non-PF patients. Arginine pathway metabolites, ornithine and spermine, activate the CaSR in primary normal human lung fibroblasts (NHLF), effects prevented by CaSR antagonism using the calcilytic NPS2143. In NHLF calcilytic also reversed the pro-fibrotic effects of exogenous TGFβ1 administration on Rho kinase and αSMA expression, proliferation, collagen production and IL-8 secretion. Targeted CaSR ablation from fibroblasts and smooth muscle cells protects mice from spontaneously occurring, age-related lung fibrosis. Conclusions Sustained CaSR activation in the lung drives pro-fibrotic processes, which can be reversed by calcilytic. Pharmacological and genetic CaSR blockade reduce both TGFβ1-induced and naturally occurring pro-fibrotic changes. This work provides the scientific rationale for developing inhaled calcilytics as novel therapeutics for IPF. Key question How does the calcium/cation-sensing receptor (CaSR) promote pulmonary fibrosis? Bottom line The CaSR is expressed in human IPF and experimental models of PF where receptor inhibition prevents pro-fibrotic changes and pulmonary remodeling. Why read on CaSR blockers, calcilytics, represent a novel treatment for IPF. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement The authors wish to thank the financial support of the King’s Commercialization Institute (to DR), the Saunders Legacy Lung Research (to BHG and DR), and the Marie Curie ETN “Multifaceted CaSR” (to PJK and DR). No author has received payment or services from third party for any aspect of the submitted work. ### Author Declarations All relevant ethical guidelines have been followed; any necessary IRB and/or ethics committee approvals have been obtained and details of the IRB/oversight body are included in the manuscript. Yes All necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived. 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 and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable. Yes All data relevant to the study are included in the article or uploaded as supplementary information

Asthma is still an incurable disease, and there is a recognized need for novel small-molecule therapies for people with asthma, especially those poorly controlled by current treatments. We previously demonstrated that calcium-sensing receptor (CaSR) negative allosteric modulators (NAMs), calcilytics, uniquely suppress both airway hyperresponsiveness (AHR) and inflammation in human cells and murine asthma surrogates. Here we assess the feasibility of repurposing four CaSR NAMs, which were originally developed for oral therapy for osteoporosis and previously tested in the clinic as a novel, single, and comprehensive topical antiasthma therapy. We address the hypotheses, using murine asthma surrogates, that topically delivered CaSR NAMs 1) abolish AHR; 2) are unlikely to cause unwanted systemic effects; 3) are suitable for topical application; and 4) inhibit airway inflammation to the same degree as the current standard of care, inhaled corticosteroids, and, furthermore, inhibit airway remodeling. All four CaSR NAMs inhibited poly-L-arginine–induced AHR in naïve mice and suppressed both AHR and airway inflammation in a murine surrogate of acute asthma, confirming class specificity. Repeated exposure to inhaled CaSR NAMs did not alter blood pressure, heart rate, or serum calcium concentrations. Optimal candidates for repurposing were identified based on anti-AHR/inflammatory activities, pharmacokinetics/pharmacodynamics, formulation, and micronization studies. Whereas both inhaled CaSR NAMs and inhaled corticosteroids reduced airways inflammation, only the former prevented goblet cell hyperplasia in a chronic asthma model. We conclude that inhaled CaSR NAMs are likely a single, safe, and effective topical therapy for human asthma, abolishing AHR, suppressing airways inflammation, and abrogating some features of airway remodeling. SIGNIFICANCE STATEMENT Calcium-sensing receptor (CaSR) negative allosteric modulators (NAMs) reduce airway smooth muscle hyperresponsiveness, reverse airway inflammation as efficiently as topical corticosteroids, and suppress airway remodeling in asthma surrogates. CaSR NAMs, which were initially developed for oral therapy of osteoporosis proved inefficacious for this indication despite being safe and well tolerated. Here we show that structurally unrelated CaSR NAMs are suitable for inhaled delivery and represent a one-stop, steroid-free approach to asthma control and prophylaxis.

Background: Idiopathic pulmonary fibrosis (IPF) is a disease with very poor prognosis and no curative therapies. The G protein-coupled, calcium/cation-sensing receptor (CaSR) is activated by environmental pollutants and by arginine-derived polyamines, which are thought to play a role in IPF. Whether the CaSR is involved in the pathogenesis of pulmonary fibrosis is unknown. Objective: To investigate the CaSR as a novel drug target for the treatment of pulmonary fibrosis (PF). Methods and results: CaSR protein expression is found in the airway epithelium in the neuroepithelial bodies of the healthy and IPF human lung. Expression of arginine pathway-linked polyamines is increased in PF patient saliva samples compared to non-PF patients. Arginine pathway metabolites, ornithine and spermine, activate the CaSR in primary normal human lung fibroblasts (NHLF), effects prevented by CaSR antagonism using the calcilytic NPS2143. In NHLF calcilytic also reversed the pro-fibrotic effects of exogenous TGF{beta}1 administration on Rho kinase and SMA expression, proliferation, collagen production and IL-8 secretion. Targeted CaSR ablation from fibroblasts and smooth muscle cells protects mice from spontaneously occurring, age-related lung fibrosis. Conclusions: Sustained CaSR activation in the lung drives pro-fibrotic processes, which can be reversed by calcilytic. Pharmacological and genetic CaSR blockade reduce both TGF{beta}1-induced and naturally occurring pro-fibrotic changes. This work provides the scientific rationale for developing inhaled calcilytics as novel therapeutics for IPF.

Introduction: Previously we have shown that blockers of the calcium-sensing receptor (CaSR, calcilytics), reduce BHR, inflammation and remodelling in Th2/IgE- and alarmin-driven asthma models. Exposure to urban particulate matter (UPM) exacerbates asthma symptoms. It is unclear whether UPM exerts its effects via the CaSR. Aim: To determine which airway cells express the CaSR and whether UPM exerts its effects by acting directly at the CaSR. Methods: Immunohistochemistry (expression of CaSR immunoreactivity); intracellular calcium sensing (CaSR activity); flow cytometry (cellular maturation and cytokine production); qPCR (TSLP mRNA expression). Results: CaSR immunoreactivity was detected on human monocytes, myeloid dendritic cells, immature and mature monocyte-derived dendritic cells (MDDC), neutrophils, alveolar macrophages and bronchial epithelial cells (HBEC). UPM mobilised intracellular calcium in HEK-293 cells stably transfected with the human CaSR but not empty vector, an effect inhibited by calcilytics (p<0.001). Calcilytics attenuated UPM-induced MDDC maturation (p<0.0001), IL-10 release (p<0.05) and UPM-upregulated TSLP mRNA expression by HBEC (p<0.05). Conclusions: The CaSR is present in inflammatory and epithelial cells involved in asthma pathogenesis. UPM acts on the CaSR on MDDC and airway epithelial cells to induce inflammatory cytokines including alarmins, an effect likely relevant to asthma exacerbation and inhibitable by calcilytics.

Introduction: We previously implicated the calcium-sensing receptor (CaSR) in the development of airways inflammation and hyperresponsiveness in human asthma, and that topical CaSR antagonists (calcilytics) reduce both in animal surrogates. Aim: Extend these studies to alarmin-driven airways inflammation by comparing the efficacy of topical (intranasal) calcilytic in a head-to-head comparison in Th2/IgE mediated (OVA) and alarmin (interleukin (IL)-33) driven asthma in a murine surrogate. Methods: IL-33 (naïve mice) or OVA (mice IgE-sensitised to OVA) was delivered once daily intranasally for 6 consecutive days followed by the calcilytic NPSP795 or vehicle control twice daily. On day 7, airways resistance was measured (Flexivent) under anaesthesia then the animals euthanised, after which bronchoalveolar lavage fluid (BALF) was collected, one lung paraffin-embedded for histomorphology and the other homogenised to measure cytokines. Results: Topical calcilytic significantly reduced (1) airways hyperresponsiveness (both stimuli); (2) BALF total and differential eosinophil, neutrophil and lymphocyte counts and lung homogenate concentrations of IL-5, IL-13 and IL-6 (variable according to stimulus); (3) peribronchial inflammatory cellular infiltration in the OVA, but not the IL-33 driven model; (4) airways collagen deposition in the IL-33, but not the OVA driven model. Conclusions: Calcilytics show therapeutic benefit in both IgE/Th2 and alarmin-driven airways inflammation and hyperresponsiveness in animal asthma surrogates in vivo, suggesting that they will be effective against any stimulus potentially exacerbating human asthma by inducing an alarmin response in the airways.

Introduction Exposure to urban particulate matter (UPM) exacerbates the development of asthma and COPD. UPM exposure triggers the release of ‘alarmins’ by the airway epithelium, which causes an inflammatory response such as acceleration of the activation and maturation of dendritic cells (DC). Previously, we have demonstrated that, in surrogate models of allergic asthma, certain environmental stimuli activate the airway calcium-sensing receptor (CaSR), which drives bronchial hyperresponsiveness, inflammation and remodelling. We have also shown that CaSR antagonists, calcilytics, can abrogate these changes. Whether UPM exerts its effects acting at the CaSR is unknown. Aims To test the ability of calcilytics to: Prevent the effects of UPM in recombinant systems expressing the CaSR and in DC; Suppress airways hyperresponsiveness, inflammation and remodelling in a murine model of alarmin–driven (IL–33) asthma. Methods [Ca2+]i responses to UPM were investigated in HEK293 cells stably transfected with human CaSR (HEK-CaSR) or empty vector (HEK-0), ± calcilytic NPS2143. FACS and cytometric bead array were used to evaluate maturation (%CD83) and cytokine release by human monocyte-derived DC following 24h exposure to UPM ± calcilytics. IL-33 was delivered intranasally to naïve mice once daily for 6 consecutive days followed by the calcilytic NPSP795 or vehicle control twice daily from day 2. On day 7, airways resistance was measured (Flexivent) under terminal anaesthesia, after which bronchoalveolar lavage fluid (BALF) analysis was performed, and lungs collected for histomorphology and for measurements of cytokine release. Results UPM increased [Ca2+]i in the HEK-CaSR but not the HEK-0 cells, an effect inhibited by calcilytics. Calcilytics attenuated UPM-induced maturation, and release of the cytokines IL-10 and IL-23p40, but not IL-6 by DC. In vivo, inhaled calcilytics significantly reduced (1) bronchial hyperresponsiveness; (2) BALF inflammatory cell infiltration and lung concentrations of IL-5, IL-13 and IL-6; (3) airways collagen deposition. Conclusions UPM activates the CaSR and induces maturation and activation of DC, an effect inhibited by calcilytics. Furthermore, calcilytics show benefit in alarmin-driven airways inflammation and hyperresponsiveness in an animal asthma surrogate, suggesting that they will be effective against exacerbating stimuli such as UPM.

Introduction Idiopathic pulmonary fibrosis (IPF) is a disease with very poor prognosis and no curative therapies. The extracellular calcium-sensing receptor (CaSR) is a chemosensor which is activated by several agonists/modulators including polyvalent cations, polyamines, and basic polypeptides. Previously we and others have shown that CaSR activation drives pulmonary inflammation and remodelling in preclinical models of asthma, COPD and pulmonary hypertension. The aims of the study are to investigate the role of the CaSR in pulmonary fibrosis and evaluate the scientific rationale for repurposing CaSR antagonists (calcilytics) as potential novel therapeutics for IPF. Methods Immunostaining was used to assess lung CaSR expression in IPF patients and control. CaSR-related metabolites were assessed in patient saliva samples (IPF and control) using high-resolution mass spectrometry. In primary human lung fibroblasts (HLF), the polyamine, spermine, was used to assess the functional activation of the CaSR via calcium imaging. HLF were also treated with transforming growth factor-β1 (TGF-β1) in the presence or absence of calcilytics to assess expression of known fibrotic markers and CaSR. Histology was carried out in 15 month old mice with targeted CaSR deletion from sm22α-positive cells to assess age-induced lung remodelling. Results CaSR expression was increased in specific cells of the IPF lungs compared to controls. The expression of several CaSR activators (amines and polyamines) was significantly increased in IPF patients compared to control (p<0.05). In human lung fibroblasts, calcilytics prevented spermine-induced increase in intracellular calcium concentration. Calcilytics also suppressed the effects of exogenous TGF-β1 supplementation on α-smooth muscle actin expression, proliferation, collagen deposition, and inflammation (p<0.01). Selective CaSR deletion from fibroblasts and smooth muscle cells protected mice from age-induced fibrosis (p<0.05). Conclusions This study supports the role of the CaSR in PF, as receptor deletion significantly attenuates fibrosis. Since CaSR activators are elevated in IPF patient saliva, increased levels of these metabolites suggest a role for the CaSR in the pathophysiology of IPF. The efficacy of calcilytics in reducing pro-fibrotic changes seen in activated lung fibroblasts further supports the development of calcilytics as a novel treatment for IPF.

Introduction: Chronic obstructive pulmonary disease (COPD) is predicted to become the third leading cause of mortality worldwide. Exposure to urban particulate matter (UPM) underpins COPD development and exacerbation. We have shown, using in vivo models of inflammatory lung diseases, that airway inflammatory and remodelling changes are at least partly mediated through the CaSR, and that calcilytics abrogate them. Dendritic cells (DC) are involved in the development of COPD and their activation is accelerated by exposure to UPM, but whether the CaSR plays a role in this process is unknown. Aims: To determine whether UPM directly activates the CaSR and if calcilytics attenuate UPM-induced DC maturation and activation. Methods: Intracellular calcium responses to UPM were investigated in HEK293 cells stably transfected with the human CaSR (HEK-CaSR) or the empty vector (HEK-0), in the presence or absence of the calcilytic NPS2143. Flow cytometry and cytometric bead array were employed to evaluate maturation (%CD83) and cytokine release by human monocyte-derived DC following 24h exposure to UPM ± calcilytics. Results: UPM induced an acute, transient intracellular calcium response in the HEK-CaSR but not the HEK-0 cells, an effect inhibited by NPS2143 (p<0.001). NPS2143 significantly attenuated UPM-induced maturation (p<0.005) and release of the immunoregulatory cytokines IL-10 and IL-23p40 (p<0.001), but not IL-6 by DC. Conclusions: This study demonstrates that UPM induces maturation and activation of DC, which is inhibited experimentally by a calcilytic. These data suggest that calcilytics may alter the natural history and progression of COPD.