Jens Walter Fischer

Heart failure with preserved ejection fraction (HFpEF) is a clinical syndrome whose pathophysiological mechanisms are incompletely understood, potentially due to a lack of preclinical models reflecting the broad range of pathophysiological aspects. We describe a diabetic DIO-STZ mouse model showing a pronounced HFpEF with underlying mechanisms that remarkably differ from other HFpEF models, making this model a relevant extension of the range of HFpEF models, especially for investigating molecular mechanisms or therapeutical interventions in diabetes.

Background: Neurodegeneration and dementia are key factors in Alzheimer’s disease (AD). The deposition of amyloid-ß into senile plaques in the brain parenchyma and in cerebral vessels known as cerebral amyloid angiopathy (CAA) are the main clinical parameters of AD. Acute myocardial infarction (AMI) and AD share a comparable pathophysiology. However, the underlying mechanisms and the consequences of AMI in AD patients are unclear to date. Methods: AD transgenic APP23 mice were analyzed in experimental AMI using the closed-chest model. Results: APP23 mice displayed significantly decreased left ventricular function as detected by FS/MPI (fractional shortening/myocardial performance index) after 24 h and 3 weeks after ligation of the LAD compared to WT controls. No differences have been observed in infarct and scar size. The analysis of cardiac remodeling after 3 weeks showed an altered composition of the collagen tissue of the scar with elevated tight but reduced fine collagen in APP23 mice. Altered scar formation was accompanied by elevated degranulation of platelets following activation of the collagen receptor GPVI. Conclusions: These results suggest that AD patients are at higher risk for cardiac damage after AMI. This implies the need for a personalized therapy of AMI in AD patients.

Background: Platelets are major players of thrombosis and inflammation after acute myocardial infarction (AMI). The impact of thrombocytopenia on platelet-induced cellular processes post AMI is not well defined. Methods: The left anterior descending artery was ligated in C57/Bl6 mice and in two thrombocytopenic mouse models to induce AMI. Results: Platelets from STEMI patients and from C57/Bl6 mice displayed enhanced platelet activation after AMI. This allows platelets to migrate into the infarct but not into the remote zone of the left ventricle. Acute thrombocytopenia by antibody-induced platelet depletion resulted in reduced infarct size and improved cardiac function 24 h and 21 days post AMI. This was due to reduced platelet-mediated inflammation after 24 h and reduced scar formation after 21 days post AMI. The collagen composition and interstitial collagen content in the left ventricle were altered due to platelet interaction with cardiac fibroblasts. Acute inflammation was also significantly reduced in Mpl^−/− mice with chronic thrombocytopenia, but cardiac remodeling was unaltered. Consequently, left ventricular function, infarct size and scar formation in Mpl^−/− mice were comparable to controls. Conclusion: This study discovers a novel role for platelets in cardiac remodeling and reveals that acute but not chronic thrombocytopenia protects left ventricular function post AMI.

Background: Vascular injury induces the exposure of subendothelial extracellular matrix (ECM) important to serve as substrate for platelets to adhere to the injured vessel wall to avoid massive blood loss. Different ECM proteins are known to initiate platelet adhesion and activation. In atherosclerotic mice, the small, leucine-rich proteoglycan biglycan is important for the regulation of thrombin activity via heparin cofactor II. However, nothing is known about the role of biglycan for hemostasis and thrombosis under nonatherosclerotic conditions. Methods: The role of biglycan for platelet adhesion and thrombus formation was investigated using a recombinant protein and biglycan knockout mice. Results: The present study identified biglycan as important ECM protein for the adhesion and activation of platelets, and the formation of three-dimensional thrombi under flow conditions. Platelet adhesion to immobilized biglycan induces the reorganization of the platelet cytoskeleton. Mechanistically, biglycan binds and activates the major collagen receptor glycoprotein (GP)VI, because reduced platelet adhesion to recombinant biglycan was observed when GPVI was blocked and enhanced tyrosine phosphorylation in a GPVI-dependent manner was observed when platelets were stimulated with biglycan. In vivo, the deficiency of biglycan resulted in reduced platelet adhesion to the injured carotid artery and prolonged bleeding times. Conclusions: Loss of biglycan in the vessel wall of mice but not in platelets led to reduced platelet adhesion at the injured carotid artery and prolonged bleeding times, suggesting a crucial role for biglycan as ECM protein that binds and activates platelets via GPVI upon vessel injury.

Rationale: Plaque instability remains poorly understood and new therapeutic approaches to reduce plaque rupture and subsequent clinical events are of great interest. Recent studies revealed an important role of phenotypic switching of smooth muscle cells (SMC) in controlling plaque stability, including ECM (extracellular matrix) deposition. Objective: The aim of this study was to elucidate the role of hyaluronan derived from SMC–hyaluronan synthase 3 ( Has3 ), in phenotypic switching and plaque stability in an animal model of atherosclerosis. Methods and Results: A mouse line with SMC-specific deletion of Has3 and simultaneous SMC-lineage tracing ( e YFP [enhanced yellow fluorescent protein]) on an Apoe ^−/− background was used. Lineage tracing of SMC with e YFP revealed that SMC-specific deletion of Has3 significantly increased the number of LGALS3 ⁺ (galectin-3) transition state SMC and decreased ACTA2 ⁺ (alpha-smooth muscle actin) SMC. Notably, SMC- Has3 deletion led to significantly increased collagen deposition and maturation within the fibrous cap and the whole lesion, as evidenced by picrosirius red staining and LC-PolScope analysis. Single-cell RNA sequencing of brachiocephalic artery lesions demonstrated that the loss of SMC- Has3 enhanced the transition of SMC to a Lgals3 ⁺ , ECM-producing phenotype with elevated acute-phase response gene expression. Experiments using cultured murine aortic SMC revealed that blocking CD44 (cluster of differentiation-44), an important hyaluronan binding receptor, recapitulated the enhanced acute-phase response, and synthesis of fibrous ECM. Conclusions: These studies provide evidence that the deletion of SMC- Has3 results in an ECM-producing transition state SMC phenotype (characterized by LGALS3 ⁺ expression), likely via reduced CD44 signaling, resulting in increased collagen formation and maturation, an index consistent with increased plaque stability.

Introduction: Hyaluronic acid (hyaluronan; HA) is an essential component of the extracellular matrix (ECM) of the skin. The HA-degrading enzyme hyaluronidase (HYAL) is critically involved in the HA-metabolism. Yet, only little information is available regarding the skin’s HA–HYAL interactions on the molecular and cellular levels.Objective: To analyze the dose- and time-dependent molecular and cellular effects of HYAL on structural cells and the HA-metabolism in the skin.Materials and methods: Chip-based, genome-wide expression analyses (Affymetrix® GeneChip PrimeView™ Human Gene Expression Array), quantitative real-time PCR analyses, enzyme-linked immunosorbent assay (ELISA), immunohistochemistry (DAB), and in vitro wound healing assays were performed to assess dose-dependent and time-kinetic effects of HA and HYAL (bovine hyaluronidase, Hylase “Dessau”) on normal human dermal fibroblasts (NHDF), primary human keratinocytes in vitro and human skin samples ex vivo.Results: Genome-wide expression analyses revealed an upregulation of HA synthases (HAS) up to 1.8-fold change in HA- and HYAL-treated NHDF. HA and HYAL significantly accelerated wound closure in an in vitro model for cutaneous wound healing. HYAL induced HAS1 and HAS2 mRNA gene expression in NHDF. Interestingly, low concentrations of HYAL (0.015 U/ml) resulted in a significantly higher induction of HAS compared to moderate (0.15 and 1.5 U/ml) and high concentrations (15 U/ml) of HYAL. This observation corresponded to increased concentrations of HA measured by ELISA in conditioned supernatants of HYAL-treated NHDF with the highest concentrations observed for 0.015 U/ml of HYAL. Finally, immunohistochemical analysis of human skin samples incubated with HYAL for up to 48 h ex vivo demonstrated that low concentrations of HYAL (0.015 U/ml) led to a pronounced accumulation of HA, whereas high concentrations of HYAL (15 U/ml) reduced dermal HA-levels.Conclusion: HYAL is a bioactive enzyme that exerts multiple effects on the HA-metabolism as well as on the structural cells of the skin. Our results indicate that HYAL promotes wound healing and exerts a dose-dependent induction of HA-synthesis in structural cells of the skin. Herein, interestingly the most significant induction of HAS and HA were observed for the lowest concentration of HYAL.