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Background: The pathology in Duchenne muscular dystrophy (DMD) is characterized by degenerating muscle fibers, inflammation, fibro-fatty infiltrate, and edema, and these pathological processes replace normal healthy muscle tissue. The mdx mouse model is one of the most commonly used preclinical models to study DMD. Mounting evidence has emerged illustrating that muscle disease progression varies considerably in mdx mice, with inter-animal differences as well as intra-muscular differences in pathology in individual mdx mice. This variation is important to consider when conducting assessments of drug efficacy and in longitudinal studies. We developed a magnetic resonance imaging (MRI) segmentation and analysis pipeline to rapidly and non-invasively measure the severity of muscle disease in mdx mice. Methods: Wildtype and mdx mice were imaged with MRI and T2 maps were obtained axially across the hindlimbs. A neural network was trained to rapidly and semi-automatically segment the muscle tissue, and the distribution of resulting T2 values was analyzed. Interdecile range and Pearson Skew were identified as biomarkers to quickly and accurately estimate muscle disease severity in mice. Results: The semiautomated segmentation tool reduced image processing time approximately tenfold. Measures of Pearson skew and interdecile range based on that segmentation were repeatable and reflected muscle disease severity in healthy wildtype and diseased mdx mice based on both qualitative observation of images and correlation with Evans blue dye uptake. Conclusion: Use of this rapid, non-invasive, semi-automated MR image segmentation and analysis pipeline has the potential to transform preclinical studies, allowing for pre-screening of dystrophic mice prior to study enrollment to ensure more uniform muscle disease pathology across treatment groups, improving study outcomes. [ABSTRACT FROM AUTHOR]

Background Confidence in natural immunity after infection with severe acute respiratory syndrome coronavirus 2 is one reason for vaccine hesitancy. Methods We measured antibody-mediated neutralization of spike protein-ACE2 receptor binding in a large community-based sample of seropositive individuals who differed in severity of infection (N = 790). Results A total of 39.8% of infections were asymptomatic, 46.5% were symptomatic with no clinical care, 13.8% were symptomatic with clinical care, and 3.7% required hospitalization. Moderate/high neutralizing activity was present after 41.3% of clinically managed infections, in comparison with 7.9% of symptomatic and 1.9% of asymptomatic infections. Conclusions Prior coronavirus disease 2019 infection does not guarantee a high level of antibody-mediated protection against reinfection in the general population. [ABSTRACT FROM AUTHOR]

Protecting muscles with antibody administration: Latent TGFβ binding protein 4 (LTBP4) has previously been shown to be a modifier of muscular dystrophy in mice. Here, Demonbreun and colleagues developed a monoclonal antibody to human LTBP4 that binds to the protein's hinge region, preventing proteolytic cleavage. In dystrophic mice expressing the human LTBP4 protein, anti-LTBP4 treatment protected muscle function, with enhanced force production and reduced fibrosis. Twenty-four weeks of treatment also improved diaphragm muscle function. The combination of anti-LTBP4 treatment with prednisone substantially enhanced muscle function and protected against injury in dystrophic mice, suggesting that anti-LTBP4 antibodies may be useful in treating muscular dystrophies in humans. Duchenne muscular dystrophy, like other muscular dystrophies, is a progressive disorder hallmarked by muscle degeneration, inflammation, and fibrosis. Latent transforming growth factor β (TGFβ) binding protein 4 (LTBP4) is an extracellular matrix protein found in muscle. LTBP4 sequesters and inhibits a precursor form of TGFβ. LTBP4 was originally identified from a genome-wide search for genetic modifiers of muscular dystrophy in mice, where there are two different alleles. The protective form of LTBP4, which contains an insertion of 12 amino acids in the protein's hinge region, was linked to increased sequestration of latent TGFβ, enhanced muscle membrane stability, and reduced muscle fibrosis. The deleterious form of LTBP4 protein, lacking 12 amino acids, was more susceptible to proteolysis and promoted release of latent TGF-β, and together, these data underscored the functional role of LTBP4's hinge. Here, we generated a monoclonal human anti-LTBP4 antibody directed toward LTBP4's hinge region. In vitro, anti-LTBP4 bound LTBP4 protein and reduced LTBP4 proteolytic cleavage. In isolated myofibers, the LTBP4 antibody stabilized the sarcolemma from injury. In vivo, anti-LTBP4 treatment of dystrophic mice protected muscle against force loss induced by eccentric contraction. Anti-LTBP4 treatment also reduced muscle fibrosis and enhanced muscle force production, including in the diaphragm muscle, where respiratory function was improved. Moreover, the anti-LTBP4 in combination with prednisone, a standard of care for Duchenne muscular dystrophy, further enhanced muscle function and protected against injury in mdx mice. These data demonstrate the potential of anti-LTBP4 antibodies to treat muscular dystrophy. [ABSTRACT FROM AUTHOR]

Question: Is antibody response to COVID-19 mRNA vaccination similar in women and men?Findings: In a community cohort without prior COVID-19 infection, the first vaccine dose produced higher IgG levels and percent inhibition of spike-ACE2 receptor binding, a surrogate measure of virus neutralization, in women compared to men (7.0µg/ml; 51.6% vs 3.3µg/ml; 36.4%). After two doses, IgG levels remained significantly higher for women (30.4µg/ml) compared to men (20.6µg/ml), while percent inhibition was similar (98.4% vs 97.7%).Meaning: Sex-specific antibody response to mRNA vaccination informs future efforts to understand vaccine protection and side effects. [ABSTRACT FROM AUTHOR]

In a community-based sample of seropositive adults (n = 1101), we found that seropositive individuals who lived with a known coronavirus disease 2019 (COVID-19) case exhibited higher blood anti–severe acute respiratory syndrome coronavirus 2 spike receptor-binding domain immunoglobulin G concentrations and greater symptom severity compared to seropositive individuals who did not live with a known COVID-19 case. [ABSTRACT FROM AUTHOR]

Objective: Serological testing is needed to investigate the extent of transmission of SARS-CoV-2 from front-line essential workers to their household members. However, the requirement for serum/plasma limits serological testing to clinical settings where it is feasible to collect and process venous blood. To address this problem we developed a serological test for SARS-CoV-2 IgG antibodies that requires only a single drop of finger stick capillary whole blood, collected in the home and dried on filter paper (dried blood spot, DBS). We describe assay performance and demonstrate its utility for remote sampling with results from a community-based study. Methods: An ELISA to the receptor binding domain of the SARS-CoV-2 spike protein was optimized to quantify IgG antibodies in DBS. Samples were self-collected from a community sample of 232 participants enriched with health care workers, including 30 known COVID-19 cases and their household members. Results: Among 30 individuals sharing a household with a virus-confirmed case of COVID-19, 80% were seropositive. Of 202 community individuals without prior confirmed acute COVID-19 diagnoses, 36% were seropositive. Of documented convalescent COVID-19 cases from the community, 29 of 30 (97%) were seropositive for IgG antibodies to the receptor binding domain. Conclusion: DBS ELISA provides a minimally-invasive alternative to venous blood collection. Early analysis suggests a high rate of transmission among household members. High rates of seroconversion were also noted following recovery from infection. Serological testing for SARS-CoV-2 IgG antibodies in DBS samples can facilitate seroprevalence assessment in community settings to address epidemiological questions, monitor duration of antibody responses, and assess if antibodies against the spike protein correlate with protection from reinfection. [ABSTRACT FROM AUTHOR]

Destruction of clonogenic cells in the crypt following irradiation are thought to cause altered gastrointestinal function. Previously, we found that an amino acid-based oral rehydration solution (AA-ORS) improved gastrointestinal function in irradiated mice. However, the exact mechanisms were unknown. Electrophysiology, immunohistochemistry, qPCR, and Western blot analysis were used to determine that AA-ORS increased proliferation, maturation, and differentiation and improved electrolyte and nutrient absorption in irradiated mice. A single-hit, multi-target crypt survival curve showed a significant increase in crypt progenitors in irradiated mice treated with AA-ORS for six days (8.8 ± 0.4) compared to the saline-treated group (6.1 ± 0.3; P < 0.001) without a change in D0 (4.8 ± 0.1 Gy). The Dq values increased from 8.8 ± 0.4 Gy to 10.5 ± 0.5 Gy with AA-ORS treatment (P < 0.01), indicating an increased radiation tolerance of 1.7 Gy. We also found that AA-ORS treatment (1) increased Lgr5+, without altering Bmi1 positive cells; (2) increased levels of proliferation markers (Ki-67, p-Erk, p-Akt and PCNA); (3) decreased apoptosis markers, such as cleaved caspase-3 and Bcl-2; and (4) increased expression and protein levels of NHE3 and SGLT1 in the brush border membrane. This study shows that AA-ORS increased villus height and improved electrolyte and nutrient absorption. [ABSTRACT FROM AUTHOR]

The pathology in Duchenne muscular dystrophy (DMD) is characterized by degenerating muscle fibers, inflammation, fibro-fatty infiltrate, and edema, and these pathological processes replace normal healthy muscle tissue. The mdx mouse model is one of the most commonly used preclinical models to study DMD. Mounting evidence has emerged illustrating that muscle disease progression varies considerably in mdx mice, with inter-animal differences as well as intra-muscular differences in pathology in individual mdx mice. This variation is important to consider when conducting assessments of drug efficacy and in longitudinal studies. Magnetic resonance imaging (MRI) is a non-invasive method that can be used qualitatively or quantitatively to measure muscle disease progression in the clinic and in preclinical models. Although MR imaging is highly sensitive, image acquisition and analysis can be time intensive. The purpose of this study was to develop a semi-automated muscle segmentation and quantitation pipeline that can quickly and accurately estimate muscle disease severity in mice. Herein, we show that the newly developed segmentation tool accurately divides muscle. We show that measures of skew and interdecile range based on segmentation sufficiently estimate muscle disease severity in healthy wildtype and diseased mdx mice. Moreover, the semi-automated pipeline reduced analysis time by nearly 10-fold. Use of this rapid, non-invasive, semi-automated MR imaging and analysis pipeline has the potential to transform preclinical studies, allowing for pre-screening of dystrophic mice prior to study enrollment to ensure more uniform muscle disease pathology across treatment groups, improving study outcomes., Competing Interests: CONFLICT OF INTEREST EMM has consulted for Amgen, AstraZeneca, Cytokinetics, Pfizer, Tenaya Therapeutics and is the founder of Ikaika Therapeutics. ARD is the Chief Scientific Officer at Ikaika Therapeutics. These activities are unrelated to the content of this manuscript. The other authors have no conflicts of interest to declare.

Membrane instability and disruption underlie myriad acute and chronic disorders. Anxa6 encodes the membrane-associated protein annexin A6 and was identified as a genetic modifier of muscle repair and muscular dystrophy. To evaluate annexin A6's role in membrane repair in vivo, we inserted sequences encoding green fluorescent protein (GFP) into the last coding exon of Anxa6. Heterozygous Anxa6gfp mice expressed a normal pattern of annexin A6 with reduced annexin A6GFP mRNA and protein. High-resolution imaging of wounded muscle fibers showed annexin A6GFP rapidly formed a repair cap at the site of injury. Injured cardiomyocytes and neurons also displayed repair caps after wounding, highlighting annexin A6-mediated repair caps as a feature in multiple cell types. Using surface plasmon resonance, we showed recombinant annexin A6 bound phosphatidylserine-containing lipids in a Ca2+- and dose-dependent fashion with appreciable binding at approximately 50 μM Ca2+. Exogenously added recombinant annexin A6 localized to repair caps and improved muscle membrane repair capacity in a dose-dependent fashion without disrupting endogenous annexin A6 localization, indicating annexin A6 promotes repair from both intracellular and extracellular compartments. Thus, annexin A6 orchestrates repair in multiple cell types, and recombinant annexin A6 may be useful in additional chronic disorders beyond skeletal muscle myopathies.

BACKGROUNDEstimates of seroprevalence to SARS-CoV-2 vary widely and may influence vaccination response. We ascertained IgG levels across a single US metropolitan site, Chicago, from June 2020 through December 2020.METHODSParticipants (n = 7935) were recruited through electronic advertising and received materials for a self-sampled dried-blood spot assay through the mail or a minimal contact in-person method. IgG against the receptor-binding domain of SARS-CoV-2 was measured using an established highly sensitive and highly specific assay.RESULTSOverall seroprevalence was 17.9%, with no significant difference between method of contact. Only 2.5% of participants reported having had a diagnosis of COVID-19 based on virus detection, consistent with a 7-fold greater exposure to SARS-CoV-2 measured by serology than that detected by viral testing. The range of IgG level observed in seropositive participants from this community survey overlapped with the range of IgG levels associated with COVID-19 cases having a documented positive PCR test. From a subset of those who participated in repeat testing, half of seropositive individuals retained detectable antibodies for 3 to 4 months.CONCLUSIONQuantitative IgG measurements with a highly specific and sensitive assay indicated more widespread exposure to SARS-CoV-2 than observed by viral testing. The range of IgG concentrations produced from these asymptomatic exposures was similar to IgG levels occurring after documented nonhospitalized COVID-19, which were considerably lower than those produced from hospitalized COVID-19 cases. The differing ranges of IgG response, coupled with the rate of decay of antibodies, may influence response to subsequent viral exposure and vaccine.FundingNational Science Foundation grant 2035114, NIH grant 3UL1TR001422-06S4, NIH National Center for Advancing Translational Sciences grants UL1 TR001422 and UL1 TR002389, Dixon Family Foundation, Northwestern University Cancer Center (NIH grant P30 CA060553), and Walder Foundation's Chicago Coronavirus Assessment Network.

Magnitude of SARS-CoV-2 virus exposure may contribute to symptom severity. In a sample of seropositive adults (n=1101), we found that individuals who lived with a known COVID-19 case exhibited greater symptom severity and IgG concentrations compared to individuals who were seropositive but did not live with a known case (P<0.0001).