Background The ChAdOx1 nCoV-19 (AZD1222) vaccine has been approved for emergency use by the UK regulatory authority, Medicines and Healthcare products Regulatory Agency, with a regimen of two standard doses given with an interval of 4–12 weeks. The planned roll-out in the UK will involve vaccinating people in high-risk categories with their first dose immediately, and delivering the second dose 12 weeks later. Here, we provide both a further prespecified pooled analysis of trials of ChAdOx1 nCoV-19 and exploratory analyses of the impact on immunogenicity and efficacy of extending the interval between priming and booster doses. In addition, we show the immunogenicity and protection afforded by the first dose, before a booster dose has been offered. Methods We present data from three single-blind randomised controlled trials—one phase 1/2 study in the UK (COV001), one phase 2/3 study in the UK (COV002), and a phase 3 study in Brazil (COV003)—and one double-blind phase 1/2 study in South Africa (COV005). As previously described, individuals 18 years and older were randomly assigned 1:1 to receive two standard doses of ChAdOx1 nCoV-19 (5 × 10 10 viral particles) or a control vaccine or saline placebo. In the UK trial, a subset of participants received a lower dose (2·2 × 10 10 viral particles) of the ChAdOx1 nCoV-19 for the first dose. The primary outcome was virologically confirmed symptomatic COVID-19 disease, defined as a nucleic acid amplification test (NAAT)-positive swab combined with at least one qualifying symptom (fever ≥37·8°C, cough, shortness of breath, or anosmia or ageusia) more than 14 days after the second dose. Secondary efficacy analyses included cases occuring at least 22 days after the first dose. Antibody responses measured by immunoassay and by pseudovirus neutralisation were exploratory outcomes. All cases of COVID-19 with a NAAT-positive swab were adjudicated for inclusion in the analysis by a masked independent endpoint review committee. The primary analysis included all participants who were SARS-CoV-2 N protein seronegative at baseline, had had at least 14 days of follow-up after the second dose, and had no evidence of previous SARS-CoV-2 infection from NAAT swabs. Safety was assessed in all participants who received at least one dose. The four trials are registered at ISRCTN89951424 (COV003) and ClinicalTrials.gov , NCT04324606 (COV001), NCT04400838 (COV002), and NCT04444674 (COV005). Findings Between April 23 and Dec 6, 2020, 24 422 participants were recruited and vaccinated across the four studies, of whom 17 178 were included in the primary analysis (8597 receiving ChAdOx1 nCoV-19 and 8581 receiving control vaccine). The data cutoff for these analyses was Dec 7, 2020. 332 NAAT-positive infections met the primary endpoint of symptomatic infection more t...

Summary After injury or cytokine stimulation, fibroblasts transdifferentiate into myofibroblasts, contractile cells that secrete extracellular matrix for wound healing and tissue remodeling. Here, a genome-wide screen identified TRPC6, a Ca2+ channel necessary and sufficient for myofibroblast transformation. TRPC6 overexpression fully activated myofibroblast transformation, while fibroblasts lacking Trpc6 were refractory to transforming growth factor-β (TGFβ) and angiotensin II-induced transdifferentiation. Trpc6 gene-deleted mice showed impaired dermal and cardiac wound healing after injury. The pro-fibrotic ligands TGFβ and angiotensin II induced TRPC6 expression through p38 mitogen-activated protein kinase (MAPK) - serum response factor (SRF) signaling via the TRPC6 promoter. Once induced, TRPC6 activates the Ca2+-responsive protein phosphatase calcineurin, which itself induced myofibroblast transdifferentiation. Moreover, inhibition of calcineurin prevented TRPC6-dependent transdifferentiation and dermal wound healing. These results demonstrate an obligate function for TRPC6 and calcineurin in promoting myofibroblast differentiation, suggesting a comprehensive pathway for myofibroblast formation in conjunction with TGFβ, p38 MAPK and SRF.

Cardiac fibrosis is a substantial problem in managing multiple forms of heart disease. Fibrosis results from an unrestrained tissue repair process orchestrated predominantly by the myofibroblast. These are highly specialized cells characterized by their ability to secrete extracellular matrix (ECM) components and remodel tissue due to their contractile properties. This contractile activity of the myofibroblast is ascribed, in part, to the expression of smooth muscle α-actin (αSMA) and other tension-associated structural genes. Myofibroblasts are a newly generated cell type derived largely from residing mesenchymal cells in response to both mechanical and neurohumoral stimuli. Several cytokines, chemokines, and growth factors are induced in the injured heart, and in conjunction with elevated wall tension, specific signaling pathways and downstream effectors are mobilized to initiate myofibroblast differentiation. Here we will review the cell fates that contribute to the myofibroblast as well as nodal molecular signaling effectors that promote their differentiation and activity. We will discuss canonical versus non-canonical transforming growth factor-β (TGFβ), angiotensin II (AngII), endothelin-1 (ET-1), serum response factor (SRF), transient receptor potential (TRP) channels, mitogen-activated protein kinases (MAPKs) and mechanical signaling pathways that are required for myofibroblast transformation and fibrotic disease.

Activation of the NLRP3 inflammasome and subsequent maturation of IL-1β have been implicated in acute lung injury (ALI), resulting in inflammation and fibrosis. We investigated the role of vimentin, a type III intermediate filament, in this process using three well-characterized murine models of ALI known to require NLRP3 inflammasome activation. We demonstrate that central pathophysiologic events in ALI (inflammation, IL-1β levels, endothelial and alveolar epithelial barrier permeability, remodeling, and fibrosis) are attenuated in the lungs of Vim-/- mice challenged with LPS, bleomycin, and asbestos. Bone marrow chimeric mice lacking vimentin have reduced IL-1β levels and attenuated lung injury and fibrosis following bleomycin exposure. Furthermore, decreased active caspase-1 and IL-1β levels are observed in vitro in Vim-/- and vimentin-knockdown macrophages. Importantly, we show direct protein-protein interaction between NLRP3 and vimentin. This study provides insights into lung inflammation and fibrosis and suggests vimentin may be a key regulator of the NLRP3 inflammasome.

SUMMARY The heart either hypertrophies or dilates in response to familial mutations in genes encoding sarcomeric proteins, which are responsible for contraction and pumping. These mutations typically alter calcium-dependent tension generation within the sarcomeres, but how this translates into the spectrum of hypertrophic versus dilated cardiomyopathy is unknown. By generating a series of cardiac-specific mouse models that permit the systematic tuning of sarcomeric tension generation and calcium fluxing, we identify a significant relationship between the magnitude of tension developed over time and heart growth. When formulated into a computational model the integral of myofilament tension development predicts hypertrophic and dilated cardiomyopathies in mice associated with essentially any sarcomeric gene mutations, but also accurately predicts human cardiac phenotypes from data generated in induced-pluripotent stem cell-derived myocytes from familial cardiomyopathy patients. This tension-based model also has the potential to inform pharmacologic treatment options in cardiomyopathy patients.

Background In the heart acute injury induces a fibrotic healing response that generates collagen rich scarring that is at first protective but if inappropriately sustained can worsen heart disease. The fibrotic process is initiated by cytokines, neuroendocrine effectors and mechanical strain that promote resident fibroblast differentiation into contractile and extracellular matrix producing myofibroblasts. The mitogen-activated protein kinase (MAPK) p38α (Mapk14 gene) is known to influence the cardiac injury response, but its direct role in orchestrating programmed fibroblast differentiation and fibrosis in vivo is unknown. Methods A conditional Mapk14 allele was used to delete the p38α encoding gene specifically in cardiac fibroblasts or myofibroblasts using 2 different tamoxifen-inducible Cre recombinase expressing gene-targeted mouse lines. Mice were subjected to ischemic injury or chronic neurohumoral stimulation and monitored for survival, cardiac function and fibrotic remodeling. Antithetically, mice with fibroblast-specific transgenic overexpression of activated MAPK kinase 6 (MKK6), a direct inducer of p38, were generated to investigate if this pathway can directly drive myofibroblast formation and the cardiac fibrotic response. Results In mice loss of Mapk14 blocked cardiac fibroblast differentiation into myofibroblasts and ensuing fibrosis in response to ischemic injury or chronic neurohumoral stimulation. A similar inhibition of myofibroblast formation and healing was also observed in a dermal wounding model with deletion of Mapk14. Transgenic mice with fibroblast-specific activation of MKK6-p38 developed interstitial and perivascular fibrosis in the heart, lung and kidney due to enhanced myofibroblast numbers. Mechanistic experiments show that p38 transduces cytokine and mechanical signals into myofibroblast differentiation through the transcription factor serum-response factor (SRF) and the signaling effector calcineurin. Conclusions These findings suggest that signals from diverse modes of injury converge on p38α MAPK within the fibroblast to program the fibrotic response and myofibroblast formation in vivo, suggesting a novel therapeutic approach with p38 inhibitors for future clinical application.

Rationale An increase in cardiac afterload typically produces concentric hypertrophy characterized by an increase in cardiomyocyte width, while volume overload or exercise results in eccentric growth characterized by cellular elongation and addition of sarcomeres in series. The signaling pathways that control eccentric versus concentric heart growth are not well understood. Objective To determine the role of extracellular signal-regulated kinases 1/2 in regulating the cardiac hypertrophic response. Methods and results Here we used mice lacking all ERK1/2 protein in the heart (Erk1−/− Erk2fl/fl-Cre) and mice expressing activated Mek1 in the heart to induce ERK1/2 signaling, as well as mechanistic experiments in cultured myocytes to assess cellular growth characteristics associated with this signaling pathway. While genetic deletion of all ERK1/2 from the mouse heart did not block the cardiac hypertrophic response per se, meaning that the heart still increased in weight with both aging and pathologic stress stimulation, it did dramatically alter how the heart grew. For example, adult myocytes from hearts of Erk1−/− Erk2fl/fl-Cre mice showed preferential eccentric growth (lengthening) while myocytes from Mek1 transgenic hearts showed concentric growth (width increase). Isolated adult myocytes acutely inhibited for ERK1/2 signaling by adenoviral gene transfer showed spontaneous lengthening while infection with an activated Mek1 adenovirus promoted constitutive ERK1/2 signaling and increased myocyte thickness. A similar effect was observed in engineered heart tissue under cyclical stretching, where ERK1/2 inhibition led to preferential lengthening. Conclusions Taken together these data demonstrate that the ERK1/2 signaling pathway uniquely regulates the balance between eccentric and concentric growth of the heart. Summary We studied mice lacking all ERK1/2 protein in the heart and mice expressing activated Mek1 in the heart to evaluate the role of the ERK 1/2 signaling cascade in regulating the cardiac hypertrophic response. While activation of the ERK pathway induced concentric hypertrophy, inhibition of this pathway resulted in eccentric hypertrophy. Using cardiomyocytes isolated from these mouse models, and using ex vivo culture models we show that these effects were mediated directly by ERK signaling. Greater ERK1/2 signaling directly programmed myocyte thickening while inhibition of ERK1/2 signaling promoted myocyte lengthening. Thus, this study sheds light on the molecular mechanisms that are partially responsible for the differential hypertrophic response of the heart to an increase in preload versus afterload.

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