Search

Cardiomyopathy is the predominant defect in Barth syndrome (BTHS) and is caused by a mutation of the X-linked Tafazzin (TAZ) gene, which encodes an enzyme responsible for remodeling mitochondrial cardiolipin. Despite the known importance of mitochondrial dysfunction in BTHS, how specific TAZ mutations cause diverse BTHS heart phenotypes remains poorly understood. We generated a patient-tailored CRISPR/Cas9 knock-in mouse allele ( Taz PM ) that phenocopies BTHS clinical traits. As Taz PM males express a stable mutant protein, we assessed cardiac metabolic dysfunction and mitochondrial changes and identified temporally altered cardioprotective signaling effectors. Specifically, juvenile Taz PM males exhibit mild left ventricular dilation in systole but have unaltered fatty acid/amino acid metabolism and normal adenosine triphosphate (ATP). This occurs in concert with a hyperactive p53 pathway, elevation of cardioprotective antioxidant pathways, and induced autophagy-mediated early senescence in juvenile Taz PM hearts. However, adult Taz PM males exhibit chronic heart failure with reduced growth and ejection fraction, cardiac fibrosis, reduced ATP, and suppressed fatty acid/amino acid metabolism. This biphasic changeover from a mild-to-severe heart phenotype coincides with p53 suppression, downregulation of cardioprotective antioxidant pathways, and the onset of terminal senescence in adult Taz PM hearts. Herein, we report a BTHS genotype/phenotype correlation and reveal that absent Taz acyltransferase function is sufficient to drive progressive cardiomyopathy.

Background: Cardiac fibroblast activation contributes to adverse remodeling, fibrosis, and dysfunction in the pressure-overloaded heart. Although early fibroblast TGF-β (transforming growth factor-β)/Smad (small mother against decapentaplegic)-3 activation protects the pressure-overloaded heart by preserving the matrix, sustained TGF-β activation is deleterious, accentuating fibrosis and dysfunction. Thus, endogenous mechanisms that negatively regulate the TGF-β response in fibroblasts may be required to protect from progressive fibrosis and adverse remodeling. We hypothesized that Smad7, an inhibitory Smad that restrains TGF-β signaling, may be induced in the pressure-overloaded myocardium and may regulate fibrosis, remodeling, and dysfunction., Methods: The effects of myofibroblast-specific Smad7 loss were studied in a mouse model of transverse aortic constriction, using echocardiography, histological analysis, and molecular analysis. Proteomic studies in S7KO (Smad7 knockout) and overexpressing cells were used to identify fibroblast-derived mediators modulated by Smad7. In vitro experiments using cultured cardiac fibroblasts, fibroblasts populating collagen lattices, and isolated macrophages were used to dissect the molecular signals responsible for the effects of Smad7., Results: Following pressure overload, Smad7 was upregulated in cardiac myofibroblasts. TGF-β and angiotensin II stimulated fibroblast Smad7 upregulation via Smad3, whereas GDF15 (growth differentiation factor 15) induced Smad7 through GFRAL (glial cell line-derived neurotrophic factor family receptor α-like). MFS7KO (myofibroblast-specific S7KO) mice had increased mortality, accentuated systolic dysfunction and dilative remodeling, and accelerated diastolic dysfunction in response to transverse aortic constriction. Increased dysfunction in MFS7KO hearts was associated with accentuated fibrosis and increased MMP (matrix metalloproteinase)-2 activity and collagen denaturation. Secretomic analysis showed that Smad7 loss accentuates secretion of structural collagens and matricellular proteins and markedly increases MMP2 secretion. In contrast, Smad7 overexpression reduced MMP2 levels. In fibroblasts populating collagen lattices, the effects of Smad7 on fibroblast-induced collagen denaturation and pad contraction were partly mediated via MMP2 downregulation. Surprisingly, MFS7KO mice also exhibited significant macrophage expansion caused by paracrine actions of Smad7 null fibroblasts that stimulate macrophage proliferation and fibrogenic activation. Macrophage activation involved the combined effects of the fibroblast-derived matricellular proteins CD5L (CD5 antigen-like), SPARC (secreted protein acidic and rich in cysteine), CTGF (connective tissue growth factor), ECM1 (extracellular matrix protein 1), and TGFBI (TGFB induced)., Conclusions: The antifibrotic effects of Smad7 in the pressure-overloaded heart protect from dysfunction and involve not only reduction in collagen deposition but also suppression of MMP2-mediated matrix denaturation and paracrine effects that suppress macrophage activation through inhibition of matricellular proteins., Competing Interests: None.

The neurofibromatosis type 1 (NF1) RASopathy is associated with persistent fibrotic nonunions (pseudarthrosis) in human and mouse skeletal tissue. Here, we performed spatial transcriptomics to define the molecular signatures occurring during normal endochondral healing following fracture in mice. Within the control fracture callus, we observed spatially restricted activation of morphogenetic pathways, such as TGF-β, WNT, and BMP. To investigate the molecular mechanisms contributing to Nf1-deficient delayed fracture healing, we performed spatial transcriptomic analysis on a Postn-cre;Nf1fl/- (Nf1Postn) fracture callus. Transcriptional analyses, subsequently confirmed through phospho-SMAD1/5/8 immunohistochemistry, demonstrated a lack of BMP pathway induction in Nf1Postn mice. To gain further insight into the human condition, we performed spatial transcriptomic analysis of fracture pseudarthrosis tissue from a patient with NF1. Analyses detected increased MAPK signaling at the fibrocartilaginous-osseus junction. Similar to that in the Nf1Postn fracture, BMP pathway activation was absent within the pseudarthrosis tissue. Our results demonstrate the feasibility of delineating the molecular and tissue-specific heterogeneity inherent in complex regenerative processes, such as fracture healing, and reconstructing phase transitions representing endochondral bone formation in vivo. Furthermore, our results provide in situ molecular evidence of impaired BMP signaling underlying NF1 pseudarthrosis, potentially informing the clinical relevance of off-label BMP2 as a therapeutic intervention.

Context: Acute sinusitis is one of the leading causes of antibiotic prescriptions in children. No recent systematic reviews have examined the efficacy of antibiotics compared with placebo., Objective: We sought to determine if antibiotics are superior to placebo in the treatment of acute sinusitis in children., Data Sources: Medline and Embase were searched from their origin to July 2023., Study Selection: We considered randomized placebo-controlled studies focusing on the treatment of acute sinusitis. In all studies, symptoms were present for <4 weeks and subjects were <18 years of age., Data Extraction: Two authors independently extracted the data. We pooled data primarily using fixed-effects models., Results: Analysis of 6 included studies showed that antibiotic treatment reduced the rate of treatment failure by 41% (with a risk ratio of 0.59; 95% confidence interval 0.49-0.72) compared with placebo. There was substantial heterogeneity between the studies (I2 = 69.7%), which decreased substantially when the 1 study with a high risk of bias was removed (I2 = 26.9%). Children treated with antibiotics were 1.6 times more likely to have diarrhea than those who were not treated with antibiotics (risk ratio = 1.62, 95% confidence interval 1.04-2.51)., Limitations: A small number of studies were eligible for inclusion. Included studies differed in their methodology., Conclusions: In children with clinically diagnosed acute sinusitis, antibiotics significantly reduced the rate of treatment failure compared with placebo. However, given the favorable natural history of sinusitis, our results could also support close observation without immediate antibiotic treatment., (Copyright © 2024 by the American Academy of Pediatrics.)

Importance: Acute otitis media (AOM) is a frequently diagnosed illness in children, yet the accuracy of diagnosis has been consistently low. Multiple neural networks have been developed to recognize the presence of AOM with limited clinical application., Objective: To develop and internally validate an artificial intelligence decision-support tool to interpret videos of the tympanic membrane and enhance accuracy in the diagnosis of AOM., Design, Setting, and Participants: This diagnostic study analyzed otoscopic videos of the tympanic membrane captured using a smartphone during outpatient clinic visits at 2 sites in Pennsylvania between 2018 and 2023. Eligible participants included children who presented for sick visits or wellness visits., Exposure: Otoscopic examination., Main Outcomes and Measures: Using the otoscopic videos that were annotated by validated otoscopists, a deep residual-recurrent neural network was trained to predict both features of the tympanic membrane and the diagnosis of AOM vs no AOM. The accuracy of this network was compared with a second network trained using a decision tree approach. A noise quality filter was also trained to prompt users that the video segment acquired may not be adequate for diagnostic purposes., Results: Using 1151 videos from 635 children (majority younger than 3 years of age), the deep residual-recurrent neural network had almost identical diagnostic accuracy as the decision tree network. The finalized deep residual-recurrent neural network algorithm classified tympanic membrane videos into AOM vs no AOM categories with a sensitivity of 93.8% (95% CI, 92.6%-95.0%) and specificity of 93.5% (95% CI, 92.8%-94.3%) and the decision tree model had a sensitivity of 93.7% (95% CI, 92.4%-94.9%) and specificity of 93.3% (92.5%-94.1%). Of the tympanic membrane features outputted, bulging of the TM most closely aligned with the predicted diagnosis; bulging was present in 230 of 230 cases (100%) in which the diagnosis was predicted to be AOM in the test set., Conclusions and Relevance: These findings suggest that given its high accuracy, the algorithm and medical-grade application that facilitates image acquisition and quality filtering could reasonably be used in primary care or acute care settings to aid with automated diagnosis of AOM and decisions regarding treatment.

Artificial ion transporters have been explored both as tools for studying fundamental ion transport processes and as potential therapeutics for cancer and channelopathies. Here we demonstrate that synthetic transporters may also be used to regulate the transport of catalytic metal ions across lipid membranes and thus control chemical reactivity inside lipid-bound compartments. We show that acyclic lipophilic pyridyltriazoles enable Pd(II) cations to be transported from the external aqueous phase across the lipid bilayer and into the interior of large unilamellar vesicles. In situ reduction generates Pd(0) species, which catalyze the generation of a fluorescent product. Photocaging the Pd(II) transporter allows for photoactivation of the transport process and hence photocontrol over the internal catalysis process. This work demonstrates that artificial transporters enable control over catalysis inside artificial cell-like systems, which could form the basis of biocompatible nanoreactors for applications such as drug synthesis and delivery or to mediate phototargeted catalyst delivery into cells.

Aims: Transforming growth factor β (TGF-β) signalling is one of the critical pathways in fibroblast activation, and several drugs targeting the TGF-β/Smad signalling pathway in heart failure with cardiac fibrosis are being tested in clinical trials. Some caveolins and cavins, which are components of caveolae on the plasma membrane, are known for their association with the regulation of TGF-β signalling. Cavin-2 is particularly abundant in fibroblasts; however, the detailed association between Cavin-2 and cardiac fibrosis is still unclear. We tried to clarify the involvement and role of Cavin-2 in fibroblasts and cardiac fibrosis., Methods and Results: To clarify the role of Cavin-2 in cardiac fibrosis, we performed transverse aortic constriction (TAC) operations on four types of mice: wild-type (WT), Cavin-2 null (Cavin-2 KO), Cavin-2 flox/flox , and activated fibroblast-specific Cavin-2 conditional knockout (Postn-Cre/Cavin-2 flox/flox , Cavin-2 cKO) mice. We collected mouse embryonic fibroblasts (MEFs) from WT and Cavin-2 KO mice and investigated the effect of Cavin-2 in fibroblast trans-differentiation into myofibroblasts and associated TGF-β signalling. Four weeks after TAC, cardiac fibrotic areas in both the Cavin-2 KO and the Cavin-2 cKO mice were significantly decreased compared with each control group (WT 8.04 ± 1.58% vs. Cavin-2 KO 0.40 ± 0.03%, P < 0.01; Cavin-2 flox/flox , 7.19 ± 0.50% vs. Cavin-2 cKO 0.88 ± 0.44%, P < 0.01). Fibrosis-associated mRNA expression (Col1a1, Ctgf, and Col3) was significantly attenuated in the Cavin-2 KO mice after TAC. α1 type I collagen deposition and non-vascular αSMA-positive cells (WT 43.5 ± 2.4% vs. Cavin-2 KO 25.4 ± 3.2%, P < 0.01) were reduced in the heart of the Cavin-2 cKO mice after TAC operation. The levels of αSMA protein (0.36-fold, P < 0.05) and fibrosis-associated mRNA expression (Col1a1, 0.69-fold, P < 0.01; Ctgf, 0.27-fold, P < 0.01; Col3, 0.60-fold, P < 0.01) were decreased in the Cavin-2 KO MEFs compared with the WT MEFs. On the other hand, αSMA protein levels were higher in the Cavin-2 overexpressed MEFs compared with the control MEFs (2.40-fold, P < 0.01). TGF-β1-induced Smad2 phosphorylation was attenuated in the Cavin-2 KO MEFs compared with WT MEFs (0.60-fold, P < 0.01). Heat shock protein 90 protein levels were significantly reduced in the Cavin-2 KO MEFs compared with the WT MEFs (0.69-fold, P < 0.01)., Conclusions: Cavin-2 loss suppressed fibroblast trans-differentiation into myofibroblasts through the TGF-β/Smad signalling. The loss of Cavin-2 in cardiac fibroblasts suppresses cardiac fibrosis and may maintain cardiac function., (© 2023 The Authors. ESC Heart Failure published by John Wiley & Sons Ltd on behalf of European Society of Cardiology.)

Academic Medical Centers (AMCs) and Federally Qualified Health Centers (FQHCs) are similarly tasked with managing the health of their local community, yet they each face unique challenges in their ability to do so. Integrating AMCs and FQHCs into novel care delivery models can leverage both organizations strengths, providing care in a comprehensive and sustainable fashion. Johns Hopkins Medicine (JHM) implemented this model with a large East Baltimore medical center, creating an AMC-FQHC collaboration focused on providing care to the East Baltimore patient population. This system provided various improvements in care delivery, including increased staffing, new wraparound services, improved access to funding dollars, and decreased out of pocket costs for patients qualifying for financial assistance. The academic missions of research and training were preserved, serving as the primary continuity clinic for several residency programs and as a community site for research. These changes resulted in more robust care for patients while improving the financial standing of the clinic. Through AMC and FQHC partnership, progress can be made toward providing holistic and financially sustainable primary care services in underserved areas while preserving the tripartite mission of academic medicine, with significant pedagogical and research opportunities., Competing Interests: Declaration of Conflicting InterestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Schistosomiasis is a disease affecting >200 million people worldwide, but its treatment relies on a single agent, praziquantel. To investigate new avenues for schistosomiasis control, we have conducted the first systematic analysis of bromodomain-containing proteins (BCPs) in a causative species, Schistosoma mansoni . Having identified 29 putative bromodomains (BRDs) in 22 S. mansoni proteins, we selected Sm BRD3, a tandem BRD-containing BCP that shows high similarity to the human bromodomain and extra terminal domain (BET) family, for further studies. Screening 697 small molecules identified the human BET BRD inhibitor I-BET726 as a ligand for Sm BRD3. An X-ray crystal structure of I-BET726 bound to the second BRD of Sm BRD3 [ Sm BRD3(2)] enabled rational design of a quinoline-based ligand ( 15 ) with an ITC K d = 364 ± 26.3 nM for Sm BRD3(2). The ethyl ester pro-drug of compound 15 (compound 22 ) shows substantial effects on sexually immature larval schistosomula, sexually mature adult worms, and snail-infective miracidia in ex vivo assays.

Target validation remains a challenge in drug discovery, which leads to a high attrition rate in the drug discovery process, particularly in Phase II clinical trials. Consequently, new approaches to enhance target validation are valuable tools to improve the drug discovery process. Here, we report the combination of site-directed mutagenesis and electrophilic fragments to enable the rapid identification of small molecules that selectively inhibit the mutant protein. Using the bromodomain-containing protein BRD4 as an example, we employed a structure-based approach to identify the L94C mutation in the first bromodomain of BRD4 [BRD4(1)] as having a minimal effect on BRD4(1) function. We then screened a focused, KAc mimic-containing fragment set and a diverse fragment library against the mutant and wild-type proteins and identified a series of fragments that showed high selectivity for the mutant protein. These compounds were elaborated to include an alkyne click tag to enable the attachment of a fluorescent dye. These clickable compounds were then assessed in HEK293T cells, transiently expressing BRD4(1) WT or BRD4(1) L94C , to determine their selectivity for BRD4(1) L94C over other possible cellular targets. One compound was identified that shows very high selectivity for BRD4(1) L94C over all other proteins. This work provides a proof-of-concept that the combination of site-directed mutagenesis and electrophilic fragments, in a mutate and conjugate approach, can enable rapid identification of small molecule inhibitors for an appropriately mutated protein of interest. This technology can be used to assess the cellular phenotype of inhibiting the protein of interest, and the electrophilic ligand provides a starting point for noncovalent ligand development.

Leishmaniases are a collection of neglected tropical diseases caused by kinetoplastid parasites in the genus Leishmania . Current chemotherapies are severely limited, and the need for new antileishmanials is of pressing international importance. Bromodomains are epigenetic reader domains that have shown promising therapeutic potential for cancer therapy and may also present an attractive target to treat parasitic diseases. Here, we investigate Leishmania donovani bromodomain factor 5 ( Ld BDF5) as a target for antileishmanial drug discovery. Ld BDF5 contains a pair of bromodomains (BD5.1 and BD5.2) in an N-terminal tandem repeat. We purified recombinant bromodomains of L. donovani BDF5 and determined the structure of BD5.2 by X-ray crystallography. Using a histone peptide microarray and fluorescence polarization assay, we identified binding interactions of Ld BDF5 bromodomains with acetylated peptides derived from histones H2B and H4. In orthogonal biophysical assays including thermal shift assays, fluorescence polarization, and NMR, we showed that BDF5 bromodomains bind to human bromodomain inhibitors SGC-CBP30, bromosporine, and I-BRD9; moreover, SGC-CBP30 exhibited activity against Leishmania promastigotes in cell viability assays. These findings exemplify the potential BDF5 holds as a possible drug target in Leishmania and provide a foundation for the future development of optimized antileishmanial compounds targeting this epigenetic reader protein.

Regions of hypoxia occur in most tumors and are a predictor of poor patient prognosis. Hypoxia-activated prodrugs (HAPs) provide an ideal strategy to target the aggressive, hypoxic, fraction of a tumor, while protecting the normal tissue from toxicity. A key challenge associated with the development of novel HAPs, however, is the ability to visualize the delivery of the prodrug to hypoxic regions and determine where it has been activated. Here, we report a modified version of the commonly used nitroimidazole bioreductive group that incorporates the fluoroethyl epitope of the antibody-based hypoxia imaging agent, EF5. Attachment of this group to the red fluorescent dye, dicyanomethylene (DCM), enabled us to correlate the release of the DCM dye with imaging of the reduced bioreductive group using the EF5 antibody. This study confirmed that the antibody was imaging reduction and fragmentation of the pro-fluorophore. We next employed the modified bioreductive group to synthesize a new prodrug of the KDAC inhibitor Panobinostat, EF5-Pano. Release of EF5-Pano in hypoxic multiple myeloma cells was imaged using the EF5 antibody, and the presence of an imaging signal correlated with apoptosis and a reduction in cell viability. Therefore, EF5-Pano is an imageable HAP with a proven cytotoxic effect in multiple myeloma, which could be utilized in future in vivo experiments., Competing Interests: The authors declare no competing financial interest., (© 2023 The Authors. Published by American Chemical Society.)

United States health systems face unique challenges in transitioning from volume-based to value-based care, particularly for academic institutions. Providing complex specialty and tertiary care dependent on servicing large geographic areas, and concomitantly meeting education and research academic missions may limit the time and resources available for focusing on the care coordination needs of complex local populations. Despite these challenges, academic medicine is well situated to capitalize on the promise of value-based care and to lead broad improvements in both teaching and nonteaching hospitals. If properly executed, value-based care and complex specialty care can be complementary and synergistic. We postulate that the transition from volume to value in population health requires all health care organizations to advance and formalize infrastructure in 3 core areas: organizational capabilities; provider engagement; and engagement of the patient, family, and community. Although these apply to all organizations, for academic health systems, this transition must also be interwoven with the other domains of the tripartite mission., (Copyright © 2023 Elsevier Inc. All rights reserved.)

MicroRNA-150 (miR-150) is conserved between rodents and humans, is significantly downregulated during heart failure (HF), and correlates with patient outcomes. We previously reported that miR-150 is protective during myocardial infarction (MI) in part by decreasing cardiomyocyte (CM) apoptosis and that proapoptotic small proline-rich protein 1a (Sprr1a) is a direct CM target of miR-150. We also showed that Sprr1a knockdown in mice improves cardiac dysfunction and fibrosis post-MI and that Sprr1a is upregulated in pathological mouse cardiac fibroblasts (CFs) from ischemic myocardium. However, the direct functional relationship between miR-150 and SPRR1A during both post-MI remodeling in mice and human CF (HCF) activation was not established. Here, using a novel miR-150 knockout;Sprr1a-hypomorphic (Sprr1a hypo/hypo ) mouse model, we demonstrate that Sprr1a knockdown blunts adverse post-MI effects caused by miR-150 loss. Moreover, HCF studies reveal that SPRR1A is upregulated in hypoxia/reoxygenation-treated HCFs and is downregulated in HCFs exposed to the cardioprotective β-blocker carvedilol, which is inversely associated with miR-150 expression. Significantly, we show that the protective roles of miR-150 in HCFs are directly mediated by functional repression of profibrotic SPRR1A. These findings delineate a pivotal functional interaction between miR-150 and SPRR1A as a novel regulatory mechanism pertinent to CF activation and ischemic HF., (© 2023. The Author(s).)

Hypoxia (low oxygen levels) occurs in a range of biological contexts, including plants, bacterial biofilms, and solid tumors; it elicits responses from these biological systems that impact their survival. For example, conditions of low oxygen make treating tumors more difficult and have a negative impact on patient prognosis. Therefore, chemical probes that enable the study of biological hypoxia are valuable tools to increase the understanding of disease-related conditions that involve low oxygen levels, ultimately leading to improved diagnosis and treatment. While small-molecule hypoxia-sensing probes exist, the majority of these image only very severe hypoxia (<1% O 2 ) and therefore do not give a full picture of heterogeneous biological hypoxia. Commonly used antibody-based imaging tools for hypoxia are less convenient than small molecules, as secondary detection steps involving immunostaining are required. Here, we report the synthesis, electrochemical properties, photophysical analysis, and biological validation of a range of indolequinone-based bioreductive fluorescent probes. We show that these compounds image different levels of hypoxia in 2D and 3D cell cultures. The resorufin-based probe 2 was activated in conditions of 4% O 2 and lower, while the Me-Tokyo Green-based probe 4 was only activated in severe hypoxia─0.5% O 2 and less. Simultaneous application of these compounds in spheroids revealed that compound 2 images similar levels of hypoxia to pimonidazole, while compound 4 images more extreme hypoxia in a manner analogous to EF5. Compounds 2 and 4 are therefore useful tools to study hypoxia in a cellular setting and represent convenient alternatives to antibody-based imaging approaches.

Atopic dermatitis (AD) is a chronic relapsing skin disease accompanied by recurrent itching. Although type 2 inflammation is dominant in allergic skin inflammation, it is not fully understood how non-type 2 inflammation co-exists with type 2 inflammation or how type 2 inflammation causes itching. We have recently established the FADS mouse, a mouse model of AD. In FADS mice, either genetic disruption or pharmacological inhibition of periostin, a downstream molecule of type 2 inflammation, inhibits NF-κB activation in keratinocytes, leading to downregulating eczema, epidermal hyperplasia, and infiltration of neutrophils, without regulating the enhanced type 2 inflammation. Moreover, inhibition of periostin blocks spontaneous firing of superficial dorsal horn neurons followed by a decrease in scratching behaviors due to itching. Taken together, periostin links NF-κB-mediated inflammation with type 2 inflammation and promotes itching in allergic skin inflammation, suggesting that periostin is a promising therapeutic target for AD., Competing Interests: Declaration of interests The authors declare no competing financial interest., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

The β 1 -adrenergic receptor (β 1 AR) is found primarily in hearts (mainly in cardiomyocytes [CMs]) and β-arrestin-mediated β 1 AR signaling elicits cardioprotection through CM survival. We showed that microRNA-150 (miR-150) is upregulated by β-arrestin-mediated β 1 AR signaling and that CM miR-150 inhibits maladaptive remodeling post-myocardial infarction. Here, we investigate whether miR-150 rescues cardiac dysfunction in mice bearing CM-specific abrogation of β-arrestin-mediated β 1 AR signaling. Using CM-specific transgenic (TG) mice expressing a mutant β 1 AR (G protein-coupled receptor kinase [GRK] - β 1 AR that exhibits impairment in β-arrestin-mediated β 1 AR signaling), we first generate a novel double TG mouse line overexpressing miR-150. We demonstrate that miR-150 is sufficient to improve cardiac dysfunction in CM-specific GRK - β 1 AR TG mice following chronic catecholamine stimulation. Our genome-wide circular RNA, long noncoding RNA (lncRNA), and mRNA profiling analyses unveil a subset of cardiac ncRNAs and genes as heretofore unrecognized mechanisms for beneficial actions of β 1 AR/β-arrestin signaling or miR-150. We further show that lncRNA Gm41664 and GDAP1L1 are direct novel upstream and downstream regulators of miR-150. Lastly, CM protective actions of miR-150 are attributed to repressing pro-apoptotic GDAP1L1 and are mitigated by pro-apoptotic Gm41664. Our findings support the idea that miR-150 contributes significantly to β 1 AR/β-arrestin-mediated cardioprotection by regulating unique ncRNA and gene signatures in CMs., (© 2022. The Author(s).)

TRIM33 is a member of the tripartite motif (TRIM) family of proteins, some of which possess E3 ligase activity and are involved in the ubiquitin-dependent degradation of proteins. Four of the TRIM family proteins, TRIM24 (TIF1α), TRIM28 (TIF1β), TRIM33 (TIF1γ) and TRIM66, contain C-terminal plant homeodomain (PHD) and bromodomain (BRD) modules, which bind to methylated lysine (KMe n ) and acetylated lysine (KAc), respectively. Here we investigate the differences between the two isoforms of TRIM33, TRIM33α and TRIM33β, using structural and biophysical approaches. We show that the N1039 residue, which is equivalent to N140 in BRD4(1) and which is conserved in most BRDs, has a different orientation in each isoform. In TRIM33β, this residue coordinates KAc, but this is not the case in TRIM33α. Despite these differences, both isoforms show similar affinities for H3 1-27 K18Ac, and bind preferentially to H3 1-27 K9Me 3 K18Ac. We used this information to develop an AlphaScreen assay, with which we have identified four new ligands for the TRIM33 PHD-BRD cassette. These findings provide fundamental new information regarding which histone marks are recognized by both isoforms of TRIM33 and suggest starting points for the development of chemical probes to investigate the cellular function of TRIM33.

The lack of evidence for large-scale glacial landscapes on Mars has led to the belief that ancient glaciations had to be frozen to the ground. Here we propose that the fingerprints of Martian wet-based glaciation should be the remnants of the ice sheet drainage system instead of landforms generally associated with terrestrial ice sheets. We use the terrestrial glacial hydrology framework to interrogate how the Martian surface gravity affects glacial hydrology, ice sliding, and glacial erosion. Taking as reference the ancient southern circumpolar ice sheet that deposited the Dorsa Argentea formation, we compare the theoretical behavior of identical ice sheets on Mars and Earth and show that, whereas on Earth glacial drainage is predominantly inefficient, enhancing ice sliding and erosion, on Mars the lower gravity favors the formation of efficient subglacial drainage. The apparent lack of large-scale glacial fingerprints on Mars, such as drumlins or lineations, is to be expected., Competing Interests: The authors declare no conflicts of interest relevant to this study., (© 2022 The Authors.)

Background and Objective: Remodelling of pulmonary arteries (PA) contributes to the progression of pulmonary hypertension (PH). Periostin, a matricellular protein, has been reported to be involved in the development of PH. We examined the role of periostin in the pathogenesis of PH using different types of experimental PH., Methods: PH was induced by vascular endothelial growth factor receptor antagonist (Sugen5416) plus hypoxic exposure (SuHx) and venous injection of monocrotaline-pyrrole (MCT-P) in wild-type (WT) and periostin -/- mice. Pulmonary haemodynamics, PA remodelling, expression of chemokines and fibroblast growth factor (FGF)-2, accumulation of macrophages to small PA and the right ventricle (RV) were examined in PH-induced WT and periostin -/- mice. Additionally, the role of periostin in the migration of macrophages, human PA smooth muscle (HPASMCs) and endothelial cells (HPMVECs) was investigated., Results: In PH induced by SuHx and MCT-P, PH and accumulation of M2 macrophage to small PA were attenuated in periostin -/- mice. PA remodelling post-SuHx treatment was also mild in periostin -/- mice compared to WT mice. Expression of macrophage-associated chemokines and FGF-2 in lung tissue, and accumulation of CD68-positive cells in the RV were less in SuHx periostin -/- than in SuHx WT mice. Periostin secretion in HPASMCs and HPMVECs was enhanced by transforming growth factor-β. Periostin also augmented macrophage, HPASMCs and HPMVECs migration. Separately, serum periostin levels were significantly elevated in patients with PH compared to healthy controls., Conclusion: Periostin is involved in the development of different types of experimental PH, and may also contribute to the pathogenesis of human PH., (© 2022 The Authors. Respirology published by John Wiley & Sons Australia, Ltd on behalf of Asian Pacific Society of Respirology.)

In this study, we present a detailed protocol for live imaging and quantitative analysis of floral meristem development in Aquilegia coerulea , a member of the buttercup family (Ranunculaceae). Using confocal microscopy and the image analysis software MorphoGraphX, we were able to examine the cellular growth dynamics during floral organ primordia initiation, and the transition from floral meristem proliferation to termination. This protocol provides a powerful tool to study the development of the meristem and floral organ primordia, and should be easily adaptable to many plant lineages, including other emerging model systems. It will allow researchers to explore questions outside the scope of common model systems., (Copyright © 2022 The Authors; exclusive licensee Bio-protocol LLC.)

Pax3 and Pax7 transcription factors are paralogs within the Pax gene family that that are expressed in early embryos in partially overlapping expression domains and have distinct functions. Significantly, mammalian development is largely unaffected by Pax7 systemic deletion but systemic Pax3 deletion results in defects in neural tube closure, neural crest emigration, cardiac outflow tract septation, muscle hypoplasia and in utero lethality by E14. However, we previously demonstrated that Pax3 hypomorphs expressing only 20% functional Pax3 protein levels exhibit normal neural tube and heart development, but myogenesis is selectively impaired. To determine why only some Pax3-expressing cell lineages are affected and to further titrate Pax3 threshold levels required for neural tube and heart development, we generated hypomorphs containing both a hypomorphic and a null Pax3 allele. This resulted in mutants only expressing 10% functional Pax3 protein with exacerbated neural tube, neural crest and muscle defects, but still a normal heart. To examine why the cardiac neural crest appears resistant to very low Pax3 levels, we examined its paralog Pax7 . Significantly, Pax7 expression is both ectopically expressed in Pax3-expressing dorsal neural tube cells and is also upregulated in the Pax3-expressing lineages. To test whether this compensatory Pax7 expression is functional, we deleted Pax7 both systemically and lineage-specifically in hypomorphs expressing only 10% Pax3. Removal of one Pax7 allele resulted in partial outflow tract defects, and complete loss of Pax7 resulted in full penetrance outflow tract defects and in utero lethality. Moreover, combinatorial loss of Pax3 and Pax7 resulted in severe craniofacial defects and a total block of neural crest cell emigration from the neural tube. Pax7 Cre lineage mapping revealed ectopic labeling of Pax3-derived neural crest tissues and within the outflow tract of the heart, experimentally confirming the observation of ectopic activation of Pax7 in 10% Pax3 hypomorphs. Finally, genetic cell ablation of Pax7 Cr e -marked cells is sufficient to cause outflow tract defects in hypomorphs expressing only 10% Pax3, confirming that ectopic and induced Pax7 can play an overlapping functional genetic compensational role in both cardiac neural crest lineage and during craniofacial development, which is normally masked by the dominant role of Pax3.

In-depth investigation of any developmental process in plants requires knowledge of both the underpinning molecular networks and how they directly determine patterns of cell division and expansion over time. Floral meristems (FMs) produce floral organs, after which they undergo floral meristem termination (FMT); precise control of organ initiation and FMT is crucial to the reproductive success of any flowering plant. Using live confocal imaging, we characterized developmental dynamics during floral organ primordia initiation and FMT in Aquilegia coerulea (Ranunculaceae). Our results uncover distinct patterns of primordium initiation between stamens and staminodes compared with carpels, and provide insight into the process of FMT, which is discernable based on cell division dynamics that precede carpel initiation. To our knowledge, this is the first quantitative live imaging of meristem development in a system with numerous whorls of floral organs, as well as an apocarpous gynoecium. This study provides crucial information for our understanding of how the spatial-temporal regulation of floral meristem behavior is achieved in both evolutionary and developmental contexts. This article has an associated 'The people behind the papers' interview., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2022. Published by The Company of Biologists Ltd.)

Repair of the infarcted heart requires TGF-β/Smad3 signaling in cardiac myofibroblasts. However, TGF-β-driven myofibroblast activation needs to be tightly regulated in order to prevent excessive fibrosis and adverse remodeling that may precipitate heart failure. We hypothesized that induction of the inhibitory Smad, Smad7, may restrain infarct myofibroblast activation, and we examined the molecular mechanisms of Smad7 actions. In a mouse model of nonreperfused infarction, Smad3 activation triggered Smad7 synthesis in α-SMA+ infarct myofibroblasts, but not in α-SMA-PDGFRα+ fibroblasts. Myofibroblast-specific Smad7 loss increased heart failure-related mortality, worsened dysfunction, and accentuated fibrosis in the infarct border zone and in the papillary muscles. Smad7 attenuated myofibroblast activation and reduced synthesis of structural and matricellular extracellular matrix proteins. Smad7 effects on TGF-β cascades involved deactivation of Smad2/3 and non-Smad pathways, without any effects on TGF-β receptor activity. Unbiased transcriptomic and proteomic analysis identified receptor tyrosine kinase signaling as a major target of Smad7. Smad7 interacted with ErbB2 in a TGF-β-independent manner and restrained ErbB1/ErbB2 activation, suppressing fibroblast expression of fibrogenic proteases, integrins, and CD44. Smad7 induction in myofibroblasts serves as an endogenous TGF-β-induced negative feedback mechanism that inhibits postinfarction fibrosis by restraining Smad-dependent and Smad-independent TGF-β responses, and by suppressing TGF-β-independent fibrogenic actions of ErbB2.