Biology subjects

Baehr, A.

Publications and source records attributed to Baehr, A..

8 recordsLinked to original sources

bioRxiv2023

Gene Therapy for Cardiomyopathy associated with Duchenne Muscular Dystrophy in a Pig Model

BackgroundGenetic cardiomyopathies caused by mutations in the dystrophin gene (DMD) are only partially responsive to current pharmacological heart failure treatments, although dilated and arrhythomogenic phenotypes of cardiomyopathy are frequent. ObjectiveIn this study, we tested whether a normalization of Ca2+-handling by forced expression of SERCA2a in cardiomyocytes mitigates heart failure and arrhythmogenesis in a pig model for Duchenne muscular dystrophy (DMD). Methods and resultsMale offspring of pigs lacking DMD exon 52 are characterized by heart failure with reduced ejection fraction (HFrEF, EF 34.5{+/-}1.8% vs. 49.2{+/-}1.0% in control hearts), arrhythmogenesis due to large apical regions of reduced voltage amplitude and sudden cardiac death with a lifespan of usually less than 4 months. Slow antegrade intracoronary infusion of AAV1.SERCA2a (3x1013 virus genomes (vg) per pig) improved left ventricular ejection fraction (EF 47.3{+/-}2.0%, p<0.05) to a similar extent as germline editing of DMD{Delta}52 to DMD{Delta}51-52, inducing a Becker dystrophy (BMD) genotype (EF 46.7{+/-}3.8%). Moreover, AAV.SERCA2a significantly reduced myocardial inflammation and fibrosis and areas of reduced AP amplitude. ConclusionsIn DMD pigs, 3x1013vg/heart of GMP-grade AAV1.SERCA2a sufficed to normalize left ventricular function and improved electrical vulnerability of the heart. Hence, AAV.SERCA2a may serve as a treatment option for DMD cardiomyopathy.

physiology↗

bioRxiv2023

Increased pulmonary monocyte infiltration and attenuated phagocytosis defines perinatal dysfunction of innate immunity in Cystic Fibrosis

In Cystic Fibrosis (CF) patients, cycles of infection and inflammation lead to fatal lung damage. While diminished mucus clearance is restored by highly effective CFTR modulator therapy, inflammation and infection persist in treated patients, suggesting alternative mechanisms may contribute to impaired immunity in CF. Here, we made use of a CF pig model to investigate the innate immune system at birth, before the onset of infection. We observed a substantial change in the composition of tissue resident immune cells towards emergency myelopoiesis, accompanied by increased infiltration of monocytes into CF lungs. A more immature status in the transcriptome profile of classical monocytes in CF pigs and preschool children with CF correlated with reduced phagocytic capacity, confirming a congenital and translationally conserved aberration of the immune system in CF. The lack of CFTR expression in circulating monocytes indicates an indirect etiology of these effects and suggests that additional immunological treatments are necessary for CF patients. One Sentence SummaryIncreased infiltration of lung tissue by monomyeloid cells and their impaired phagocytic potential cause dysfunctional imprinting of mucosal immunity in Cystic Fibrosis airways at birth and suggest early and specific treatment of the innate immune system in patients.

immunology↗

bioRxiv2023

Circuit to target approach defines an autocrine myofibroblast loop that drives cardiac fibrosis

Fibrosis is a broad pathology of excessive scarring with substantial medical implications. The fibrotic scar is produced by myofibroblasts that interact with macrophages. Fibrosis is a complex process involving thousands of factors, therefore, to better understand fibrosis and develop new therapeutic approaches, it is necessary to simplify and clarify the underlying concepts. Recently, we described a mathematical model for a macrophage-myofibroblast cell circuit, predicting two types of fibrosis - hot fibrosis with abundant macrophages and myofibroblasts, and cold fibrosis dominated by myofibroblasts alone. To test these concepts and intervention strategies in a medically relevant system, we use a widely studied in-vivo injury model for fibrosis, myocardial infarction (MI). We show that cold fibrosis is the final outcome of MI in both mice and pigs and demonstrate that fibrosis can shift toward healing in regenerative settings. MI begind with an increase of myofibroblasts and macrophages, followed by macrophage decline leading to persistent cold fibrosis (only myofibroblasts). During this process, fibroblasts, unlike macrophages, acquire distinct fate changes. Using mathematical modeling we predict that targeting of the autocrine signal for myofibroblast division could block cold fibrosis. We identify TIMP1 as an autocrine cardiac myofibroblast growth factor in-vitro. Treatment of adult mice after MI with anti-TIMP1 antibodies reduces fibrosis in-vivo. This study shows the utility of the concepts of hot and cold fibrosis and the feasibility of our circuit-to-target approach to reduce fibrosis after acute cardiac injury by inhibiting the myofibroblast autocrine loop.

systems biology↗

bioRxiv2022

Immature human engineered heart tissues engraft in a guinea pig chronic injury model

AO_SCPLOWBSTRACTC_SCPLOWEngineered heart tissue (EHT) transplantation represents an innovative, regenerative approach for heart failure patients. Late preclinical trials are underway, and the first clinical trial has started in 2021. Preceding studies revealed functional recovery after implantation of in vitro-matured EHT in the subacute stage while transplantation in a chronic injury setting was less efficient. We hypothesized that the use of immature EHT patches (EHTIm) could improve cardiomyocytes (CM) engraftment. Chronic myocardial injury was induced in a guinea pig model (n=14). EHTIm (15x106 cells) were transplanted directly after casting. Functional consequences were assessed by serial echocardiography. Animals were sacrificed four weeks after transplantation and hearts were excised for histological analysis. Cryo-injury lead to large transmural scars amounting to 26% of the left ventricle. Grafts were identified by a positive staining for human Ku80 and dystrophin, remuscularizing 9% of the scar area on average. The CM density in the graft was higher compared to previous studies with in vitro-matured EHTs and showed a greater population of immature CM. Echocardiographic analysis showed a small improvement of left ventricular function after EHTIm transplantation. In a small translational proof-of-concept study human scale EHTIm patches (4.5x108 cells) were epicardially implanted on healthy pig hearts (n=2). In summary, we provide evidence that transplantation of immature EHT patches without pre-cultivation results in better cell engraftment.

pharmacology and toxicology↗

bioRxiv2021

The second lineage differentiation of bovine embryos fails in the absence of OCT4/POU5F1

The mammalian blastocyst undergoes two lineage segregations, i.e., formation of the trophectoderm and subsequently differentiation of the hypoblast (HB) from the inner cell mass, leaving the epiblast (EPI) the remaining pluripotent lineage. To clarify expression patterns of markers specific for these lineages in bovine embryos, we analyzed day 7, 9 and 12 blastocysts completely derived ex vivo by staining for OCT4, NANOG, SOX2 (EPI) and GATA6, SOX17 (HB) and identified genes specific for these developmental stages in a global transcriptomics approach. To study the role of OCT4, we generated OCT4-deficient (OCT4 KO) embryos via somatic cell nuclear transfer or in vitro fertilization. OCT4 KO embryos reached the expanded blastocyst stage by day 8 but lost of NANOG and SOX17 expression, while SOX2 and GATA6 were unaffected. Blastocysts transferred to recipient cows from day 6 to 9 expanded, but the OCT4 KO phenotype was not rescued by the uterine environment. Exposure of OCT4 KO embryos to exogenous FGF4 or chimeric complementation with OCT4 intact embryos did not restore NANOG or SOX17 in OCT4-deficient cells. Our data show, that OCT4 is required cell-autonomously for the maintenance of pluripotency of the EPI and differentiation of the HB in bovine embryos.

developmental biology↗

bioRxiv2021

A scalable, clinically severe pig model for Duchenne muscular dystrophy

Large animal models for Duchenne muscular dystrophy (DMD) are crucial for preclinical evaluation of novel diagnostic procedures and treatment strategies. Pigs cloned from male cells lacking DMD exon 52 (DMD{Delta}52) resemble molecular, clinical and pathological hallmarks of DMD, but cannot be propagated by breeding due to death before sexual maturity. Therefore, female DMD+/- carriers were generated. A single founder animal had 11 litters with 29 DMDY/-, 34 DMD+/- as well as 36 male and 29 female wild-type (WT) offspring. Breeding with F1 and F2 DMD+/- carriers resulted in additional 114 DMDY/- piglets. The majority of them survived for 3-4 months, providing large cohorts for experimental studies. Pathological investigations and proteome studies of skeletal muscles and myocardium confirmed the resemblance of human disease mechanisms. Importantly, DMDY/- pigs reveal progressive fibrosis of myocardium and increased expression of connexin-43, associated with significantly reduced left ventricular fractional shortening and ejection fraction already at age 3 months. Furthermore, behavioral tests provided evidence for impaired cognitive ability of DMDY/- pigs. Our breeding cohort of DMD{Delta}52 pigs and standardized tissue repositories from DMDY/- pigs, DMD+/- carriers, and WT littermate controls provide important resources for studying DMD disease mechanisms and for testing novel diagnostic procedures and treatment strategies.

pathology↗

bioRxiv2021

Early Disruption of Photoreceptor Cell Architecture and Loss of Vision in a Humanized Pig Model of Usher Syndrome

Usher syndrome (USH) is the most common form of monogenic deaf-blindness. Loss of vision is untreatable and, so far, there are no suitable animal models for testing therapeutic strategies. By introducing a human mutation into the harmonin-encoding USH1C gene in pigs, we generated the first translational animal model for USH type 1 with characteristic hearing defect, vestibular dysfunction and visual impairment. Changes in photoreceptor architecture, quantitative motion analysis and electroretinography were characteristics of the reduced retinal virtue in USH1C pigs. Primary cells from those animals and USH1C patients showed significantly elongated primary cilia, compared to wild-type, confirming the nature of USH as a true and general ciliopathy and proving the therapeutic capacity of gene supplementation and gene repair approaches.

genetics↗

bioRxiv2019

Agrin promotes coordinated therapeutic processes leading to improved cardiac repair in pigs

Ischemic heart diseases are classified among the leading cause of death and reduced life quality worldwide. Although revascularization strategies significantly reduce mortality after acute myocardial infarction (MI), a significant number of MI patients develop chronic heart failure over time. We have recently reported that a fragment of the extra cellular matrix (ECM) protein Agrin promotes cardiac regeneration following MI in adult mice. Here, we tested the therapeutic potential of Agrin in a preclinical porcine model, comprising either 3 or 28 days (d) reperfusion period. We first demonstrate that local (antegrade) delivery of recombinant human Agrin (rhAgrin) to the infarcted pig heart can target the affected regions in an efficient and clinically-relevant manner. Single dose of rhAgrin resulted in significant improvement in heart function, infarct size, fibrosis and adverse remodeling parameters 28 days post MI. Short-term MI experiment along with complementary murine MI studies revealed myocardial protection, improved angiogenesis, inflammatory suppression and cell cycle re-entry, as Agrins mechanisms of action. We conclude that a single dose of Agrin is capable of reducing ischemia reperfusion injury and improving cardiac function, demonstrating that Agrin could serve as a therapy for patients with acute MI and potentially heart failure.

physiology↗