Biology subjects

Thomsen, M. B.

Publications and source records attributed to Thomsen, M. B..

10 recordsLinked to original sources

bioRxiv2026

Revisiting Mouse Cardiac Myocyte Isolation: A Simplified Langendorff-based Method

This protocol describes a Langendorff-based method for isolating intact adult mouse ventricular myocytes using syringe pump-driven perfusion. The approach retains the key physiological advantage of the conventional Langendorff technique, continuous retrograde coronary perfusion, while simplifying the overall setup. By combining retrograde aortic perfusion with widely available laboratory equipment, the method provides an accessible alternative to traditional Langendorff systems. A precision syringe pump connected to an in-line heater is used to deliver temperature-controlled, constant-flow perfusion during enzymatic digestion. In contrast to gravity-driven constant-pressure systems, constant-flow perfusion maintains stable enzyme delivery despite changes in coronary resistance that occur during tissue digestion. Use of an inline heater allows precise, rapid temperature-controlled delivery, avoiding the complexity, leak risk, thermal lag, and contamination susceptibility associated with traditional water-jacketed systems. Our setup reduces variability in perfusion rate and minimizes susceptibility to occlusion, flow interruption, or compliance-related artifacts, enhancing reproducibility. The method consistently yields adult ventricular myocytes with high viability (>70% rod-shaped, calcium-tolerant), enabling a broad range of functional analyses including electrophysiology, contractile performance and calcium handling. Step-by-step instructions, troubleshooting guidance, and anticipated outcomes are provided to facilitate adoption in laboratories without dedicated isolated-heart perfusion infrastructure. Key FeaturesO_LISimplified Langendorff-based mouse cardiomyocyte isolation method that eliminates the need for specialized perfusion rigs. C_LIO_LISyringe pump-driven constant-flow perfusion combined with inline temperature control improves reproducibility by ensuring stable enzyme delivery and precise temperature regulation. C_LIO_LIGenerates high-yield, calcium-tolerant adult mouse ventricular myocytes suitable for functional studies. C_LI Graphical Overview O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=190 SRC="FIGDIR/small/718810v1_ufig1.gif" ALT="Figure 1"> View larger version (63K): org.highwire.dtl.DTLVardef@15061cdorg.highwire.dtl.DTLVardef@44fd48org.highwire.dtl.DTLVardef@1509285org.highwire.dtl.DTLVardef@c362a4_HPS_FORMAT_FIGEXP M_FIG Graphical overview of the simplified Langendorff-based mouse cardiac myocyte isolation protocol. C_FIG

physiology↗

bioRxiv2026

Satellite Glial Cells Control Sensory Neuron Excitability via the Release of Fibulin-2

Pain remains a major clinical challenge, with current therapies often limited by low efficacy and adverse effects. While excitability of sensory neurons in the dorsal root ganglia (DRG) is central to pain signaling, accumulating evidence highlights the importance of non-neuronal cells in modulating neuronal activity. Here, we identify satellite glial cells (SGCs) as a source of Fibulin-2, an extracellular matrix glycoprotein with diverse roles in nervous system development and repair. Using SGC primary cultures and mass spectrometry, we demonstrate that Fibulin-2 is secreted by SGCs in part via extracellular vesicles. Electrophysiological assays reveal that application of recombinant Fibulin-2 to cultured sensory neurons reduces neuronal excitability by modulating Kv4-mediated currents. In vivo, loss of Fibulin-2 leads to reduced Kv4.2 and Kv4.3 expression and heightened mechanical, heat and cold sensitivity in mice. Our findings uncover a SGC-sensory neuron signaling mechanism modulating pain sensitivity, suggesting Fibulin-2 as a potential therapeutic target for pain management.

neuroscience↗

bioRxiv2026

Single-cell transcriptomic atlas of glial cells in adult mouse dorsal root ganglia identifies multipotent progenitors

Sensory ganglia in the peripheral nervous system contain diverse glial populations that modulate sensory signaling, respond to injury and contribute to tissue homeostasis. Satellite glial cells (SGCs) surrounding neuronal soma in dorsal root ganglia (DRG) were suggested to retain developmental potential, but the identity of progenitor cells remains undefined. To capture glial diversity, we assembled a comprehensive single-cell transcriptomic atlas by integrating over 200,000 DRG and sciatic nerve transcriptomes across multiple studies and injury paradigms. High-resolution clustering resolved 28 cell types and demonstrated significant transcriptional heterogeneity within SGCs and Schwann cells, including repair and reactive sub-states. We identified two distinct populations of progenitor cells that reflect different states in the progenitor trajectory. Functionally, progenitor cell numbers increase after injury, and endothelin signaling regulates glial cell proliferation early in development. This integrated DRG and peripheral nerve atlas represents an essential resource for exploring new features of the peripheral nervous system.

neuroscience↗

bioRxiv2025

Slow Calcium Removal Prolongs Ventricular Relaxation in Mice with HFpEF

BackgroundHeart failure with preserved ejection fraction (HFpEF) accounts for over half of heart failure cases, yet its underlying mechanisms incompletely understood and effective therapies are lacking. Diastolic dysfunction, the hallmark of HFpEF, may arise from impaired active myocardial relaxation, but the contribution of intracellular calcium (Ca2+) handling remains unclear. MethodsWe used a validated "two-hit" murine model of HFpEF, induced by diet-driven obesity and hypertension, to investigate ventricular Ca2+ dynamics. Cardiac function was assessed in vivo by echocardiography, ex vivo in isolated working hearts, and at the cellular level using Fura-2-based Ca2+ imaging of isolated ventricular myocytes. ResultsHFpEF mice developed obesity, diastolic dysfunction, hypertrophy, reduced cardiac index, and exercise intolerance despite preserved ejection fraction. Impaired lusitropy was evident in vivo, ex vivo, and at the cellular level, where ventricular myocytes from HFpEF hearts displayed elevated diastolic [Ca2+]i, increased Ca2+ transient amplitudes, and frequency-dependent slowing of Ca2+ clearance ({tau}), most pronounced at 4 Hz (33% slower vs. controls, p < 0.05). HFpEF myocytes also exhibited an attenuated {beta}-adrenergic response to isoprenaline, further limiting diastolic reserve. ConclusionsHFpEF is characterised by a distinct ventricular myocyte Ca2+ handling phenotype, diverging from heart failure with reduced ejection fraction (HFrEF), with elevated diastolic Ca2+, exaggerated and prolonged Ca2+ transients, and blunted {beta}-adrenergic modulation. These abnormalities converge to impair lusitropy and exercise tolerance, highlighting altered Ca2+ dynamics as a central mechanism in HFpEF. Targeting these specific Ca2+ handling defects may represent a novel therapeutic strategy.

physiology↗

bioRxiv2025

Targeted delivery of RNA-based therapeutics enables functional analysis of macrophage subpopulations

Macrophages infiltrate all human tissues where they play key roles in innate immunity, homeostasis, and tissue function. However, extensive clinical and experimental evidence indicates that macrophages also contribute significantly to the progression of several diseases such as cancer, cardiometabolic disorders, and inflammatory and neurodegenerative conditions. Advances in single-cell omics have revealed diverse macrophage populations in both healthy and diseased tissues. However, studying their functions is challenging due to limitations in tools for targeting specific populations. The Cre-lox system, involving Cre recombinase expression driven by macrophage-specific promoters, is widely used for gene manipulation. Despite its utility, this method has drawbacks like leaky expression, variable efficiency, and potential toxicity. Moreover, genetic models are costly and can have unintended effects on immune cells, hindering comprehensive studies on macrophage function. To address this challenge, we developed an advanced lipid nanoparticle-based system for precise RNA therapeutic delivery to macrophages, either broadly or via antibody-mediated targeting of specific subsets. This versatile platform enables the administration of various RNA molecules, such as mRNA, siRNA, and sgRNA for CRISPR/Cas9 applications, in both in vitro and in vivo settings. It allows for targeted cell depletion or gene knockout, facilitating detailed functional analysis. Furthermore, the systems flexibility and precision are enhanced by its compatibility with Cre-specific Cas9 expression, enabling comprehensive genomic and proteomic targeting of specific macrophage subsets.

immunology↗

bioRxiv2025

Macrophage depletion restores the DRG microenvironment and prevents axon degeneration in bortezomib-induced neuropathy

Peripheral neuropathy is a common and debilitating side effect of the chemotherapeutic bortezomib (BTZ). To explore the mechanisms underlying BTZ-induced neuropathy (BIPN), we developed a mouse model that replicates the route of administration and approximates the prolonged BTZ exposure experienced by patients. We find that male mice treated with BTZ experience more severe sensorimotor dysfunction and axon loss compared to females and observed similar results when analyzing human data. Using single cell RNA-sequencing, we reveal that BTZ significantly alters the dorsal root ganglia (DRG) microenvironment in mice, producing pronounced sex-specific changes in satellite glial cells (SGCs) in males and females and dysregulation of the extracellular matrix (ECM), particularly in males. These changes are accompanied by expansion of macrophages, which is more pronounced in males. We identify four macrophage subtypes in the DRG, including a pro-fibrotic population that is exclusively associated with BIPN. Depletion of macrophages via anti-CSF1R treatment in male mice prevents BTZ-induced SGC activation and aberrant collagen deposition in DRGs, potently preserves peripheral axons, and improves functional outcomes. These findings highlight SGCs, neuroinflammation and dysregulation of the ECM as drivers of sex-specific differences in BIPN and suggest that targeting neuroinflammation is a promising therapeutic strategy to treat this disease. One Sentence SummaryInflammation and neurofibrosis in DRGs underlie sex-specific differences in bortezomib-induced neuropathy and are promising treatment targets.

neuroscience↗

bioRxiv2024

Low-dose colchicine treatment improves vasorelaxation, reduces arterial remodeling and attenuates blood pressure increases in spontaneously hypertensive rats

Colchicine, a microtubule depolymerizing agent, is an effective therapy for the secondary prevention of cardiovascular disease and has been approved recently as a novel treatment for atherosclerosis associated with coronary artery disease. Hypertension is a leading cause of cardiovascular disease, yet the impact of colchicine on hypertension has not been studied. We hypothesized that low-dose colchicine could be used to treat hypertension to reduce cardiovascular disease risk. The aim of this study was to administer daily, low dose (0.05 mg/kg/day) oral colchicine for 4 weeks to spontaneously hypertensive rats (SHR) and normotensive controls (WKY) and determine the effect on blood pressure, vascular reactivity, remodeling and inflammation, and left ventricular hypertrophy. Daily blood pressure measurements recorded by telemetry in conscious rats showed colchicine prevented increases in mean arterial pressure observed in the SHRs receiving vehicle over the 4-week treatment period. After the 4-weeks of treatment, 3rd order mesenteric artery vasorelaxations to isoproterenol, sodium nitroprusside and the Kv7.2-5 channel activator, ML213, were enhanced in the SHRs receiving colchicine compared to vehicle. The improved isoproterenol-mediated relaxation was also observed in WKY rats receiving colchicine, and in both the SHR and WKY, this improved effect was attenuated by the {beta}2 adrenoceptor antagonist, ICI118,551. Proteomic analysis of the mesenteric arteries by mass spectrometry revealed that colchicine treatment prevented changes observed when comparing the SHR vehicle group with the WKY vehicle group in proteins associated with extracellular matrix pathways. Immunostaining of 3rd order mesenteric arteries with Sirius red found that colchicine treatment attenuated the increased media thickness of the artery wall observed in SHRs receiving vehicle. Multiplex immunoassay and Western blots revealed colchicine reduced certain inflammatory mediators in the wall of the SHR mesenteric arteries, particularly the nucleotide-binding domain and leucine-rich repeat pyrin containing protein-3 (NLRP3), IL-18, CXCL10, and CXCL2, as well as reducing phosphorylated STAT3. Finally, in the left ventricle of the SHR, colchicine treatment attenuated a number of inflammatory mediators, including NLRP3, IL-1{beta}, and IL-18, and reduced fibrosis and cell size, which are indicative of left ventricular hypertrophy. Overall, we show colchicine has the potential to elicit cardiovascular protective effects in hypertension by targeting multiple cell types.

physiology↗

bioRxiv2024

Serotonin-2B receptor (5-HT2BR) expression and binding in the brain of ageing APPswe/PS1de9 transgenic mice and in human Alzheimer's disease brain tissue

Although evidence of dysregulation in central serotonergic signaling is widespread in Alzheimers disease (AD), relatively little is known about the specific involvement of the serotonin-2B receptor (5-HT2BR) subtype. Here, we assessed 5-HT2BR expression and binding in brain tissue from APPswe/PS1dE9 transgenic (TG) mice and AD patients. 5-HT2BR mRNA was measured by RT-qPCR in 3-to >18-month-old TG and wild-type (WT) littermate mice (n=3-8), and in middle frontal gyrus samples from female, AD and control subjects (n=7-10). The density of 5-HT2BRs was measured by autoradiography using 1 nM [3H]RS127445 and 1 M LY266097. In both mouse and human samples, 5-HT2BR mRNA was detectable after 33 amplification cycles. Levels in WT mice, not TG mice, increased with age, and were higher in AD patients compared to control subjects. [3H]RS127445 binding was low in the mouse brain, detected after 3 months of age. 5-HT2BR density was overall lower in the hippocampus of TG compared to WT mice. Specific binding was too low to be reliably quantified in the human samples. These data provide evidence of a different 5-HT2BR expression and binding in the APPswe/PS1dE9 TG model of AD and AD patients. Studies investigating the functional involvement of the 5-HT2BR in AD are warranted.

neuroscience↗

bioRxiv2023

Neurexin 3 is required for the specific S-cone to S-cone bipolar cell synapse in the mammalian retina

Precise wiring within sensory systems is critical for the accurate transmission of information. In the visual system, S-cone photoreceptors specialize in detecting short-wavelength light, crucial to color perception and environmental cue detection. S-cones form specific synapses with S-cone bipolar cells (SCBCs), a connection that is remarkably consistent across species. Yet, the molecular mechanisms guiding this specificity remain unexplored. To address this, we used the cone-dominant ground squirrel for deep-sequencing of cone subtype transcriptomes and identified Nrxn3 as an essential molecule for the S-cone to SCBC synapse. Using transgenic mouse models, we further examined the role of Nrxn3 in S-cones and discovered a significant reduction of SCBC connections in the absence of Nrxn3. This finding extends the known functions of neurexins, typically associated with synapse regulation, by highlighting their essential role in a specific synaptic connection for the first time. Moreover, the differentially expressed genes identified here pave the way for further investigations into the unique functions of cone subtypes.

neuroscience↗

bioRxiv2020

Proteomics dissection of cardiac protein profiles of humans and model organisms

The study of human cardiac pathologies often relies on research conducted in model organisms to gain molecular insight into disease and to develop novel treatment strategies; however, translating findings from model organisms back to human can present a significant challenge, in part due to a lack of knowledge about the differences across species in cardiac protein abundances and their interactions. Here we set out to bridge this knowledge gap by presenting a global analysis of cardiac protein expression profiles in humans and commonly used model organisms. Using quantitative mass spectrometry-based proteomics, we measured the abundance of ~7,000 proteins in samples from the separate chambers of human, pig, horse, rat, mouse and zebrafish hearts. This knowledgebase of cardiac protein signatures is accessible through an online database at: atlas.cardiacproteomics.com. Quantitative comparison of the protein profiles support the pig as model organism of choice for arrhythmogenic right ventricular cardiomyopathy whereas comparison of profiles from the two-chambered zebrafish heart suggests a better resemblance to the right side of mammalian hearts. This proteomics resource facilitates translational prospect of cardiac studies from model organisms to humans by enabling direct comparison of disease-linked protein networks across species.

systems biology↗