Biology subjects

Friebe, A.

Publications and source records attributed to Friebe, A..

7 recordsLinked to original sources

bioRxiv2026

Arteriolar GMP signaling revealed by FRET-based real-time measurement in vivo

cGMP evokes arteriolar vasorelaxation and is generated in smooth muscle by the soluble guanylyl cyclase (sGC) stimulated by endothelial NO. Phosphodiesterases (PDEs) degrade cGMP and may contribute in diameter regulation. We examined arteriolar cGMP levels in real-time in vivo upon NO and acetylcholine (ACh) and effects of PDE inhibition. Changes in cGMP were measured by FRET using the indicator protein cGi500. Arterioles in the cremaster muscle of anesthetized mice expressing cGi500 were exposed for imaging. They were stimulated with various NO donors (DEA/NO, SNP, SNAP) and ACh with or without PDE inhibition (non-specific: IBMX; PDE-5 specific: sildenafil). Furthermore, dilations induced by ACh and SNP were studied in sGC-deficient mice. DEA/NO and SNAP led to a fast, concentration-dependent rise of fluorescence ratio of cGi500 (by 4.0% at 10{micro}M), that declined on removal slowly. These ratio changes indicated cGMP increases as verified by inhibition of the sGC using ODQ. PDE inhibition increased cGMP levels slowly despite NO synthase inhibition. However, it did not amplify NO-induced ratio increases, but the decline after drug removal was decelerated. Interestingly, ACh did not modify cGMP levels and dilations were not impaired in sGC-deficient mice. FRET-based imaging allows reliable real-time assessment of arteriolar cGMP levels in vivo. PDE activity does not limit the amplitude, but the duration of cGMP signaling after stimulation. Furthermore, we conclude that ACh does not release endothelial NO in murine arterioles. Preliminary experiments demonstrate that simultaneous measurement of cGMP and diameter is feasible using mice with expression of cGi500 in smooth muscle cells.

physiology↗

bioRxiv2026

Using activity data to estimate brown bear den exit and entry dates

BackgroundIn hibernating mammals, the timing of den entry and exit reflects complex interactions among environment, physiology, and energetic constraints, with consequences for fitness. Consequently, shifts in denning phenology can affect population dynamics, particularly under climate change. Reliable estimation of denning timing is therefore critical, yet current methods often rely on GPS-derived movement data, limited by coarse sampling intervals, detection issues, and the inability to distinguish true inactivity from active presence at the den site. In this study, we developed and apply a method to estimate denning phenology in a brown bear population in south-central Sweden using accelerometer-derived activity data. Our approach employs adaptive, individual-specific thresholds to account for variation in baseline activity across bears, focusing on day-to-day changes to identify the start and end of inactivity periods. This method allows flexible and reproducible detection of den entry and exit dates, overcoming limitations associated with fixed thresholds and small sample sizes. ResultsWe compared activity-based estimates with GPS-derived den occupancy and examined variation in denning behavior across demographic groups. Analyzing 388 bear-winters, the method successfully identified inactivity periods in 360 cases. The method failed to identify clear start and end dates of hibernation for 28 (7%) bear-winters, which were characterized by unusually high or low daily activity levels at the boundaries of the inactivity period. Den site occupancy ranged from September 5 to June 2, with durations of 112-260 days, whereas inactivity periods detected from activity data extended from September 6 to May 13, lasting 83-217 days. Our comparison of activity-based and GPS-based methods indicates that bears may arrive at the den site several weeks before the onset of inactivity, with timing varying among demographic groups. ConclusionWe show that activity-based analysis provides a robust framework for estimating denning phenology, distinguishing actual inactivity from site presence, and improving understanding of the timing and variability of bear denning behavior. Applying an individual-level activity-based method improves accuracy in assessing ecological mechanisms underlying hibernation in bears and other hibernators, while also enhancing interpretation of environmental drivers and providing a reliable tool to monitor phenological shifts in response to climate change.

animal behavior and cognition↗

bioRxiv2024

Brown Bears activates a Satiety Hormone Cholecystokinin (CCK) pathway in adipose tissue during hibernation

The brown bear (Ursus arctos) hibernates to survive cold winters without access to food. It builds enormous subcutaneous fat stores during summer and relies on it for energy during winter. Remarkably, the weight loss during winter occurs without muscle loss despite inactivity. Studying brown bear biology can therefore provide insights for improving human health in obesity and weight loss treatments. We here investigate subcutaneous adipose tissue biopsies obtained during summer and winter from free-living brown bears. During winter, a signature of genes involved in food intake and digestion is upregulated. Among these are several regulators of satiety, substrate transport and lipid metabolism. Interestingly, in humans these genes are enriched in distinct metabolic organs including, brain, intestine, stomach, liver and even salivary glands. We focused on the satiety brain/intestinal hormone cholecystokinin (CCK), which we demonstrate is produced in adipocytes, accompanied by an upregulation of the CCK receptor CCKBR. Importantly, CCK was undetectable in the circulation during winter and presence of sensory neurons suggest a neuronal feedback mechanism within the adipose tissue. Using RNA sequencing, we predict additionally 537 secreted proteins to be seasonally regulated, 37 of which could be confirmed with plasma proteomics. In conclusion, we propose that brown bears have developed a strategy of healthy fat burning and satiety regulation through an adipose tissue-contained mechanism which includes digestion factors and satiety mediators to provide safe energy turnover during hibernation-dependent weight loss.

molecular biology↗

bioRxiv2023

Role of the NO-GC/cGMP signaling pathway in platelet biomechanics

Cyclic guanosine monophosphate (cGMP) is a second messenger produced by the NO-sensitive guanylyl cyclase (NO-GC) enzyme. In platelets, the NO-GC/cGMP pathway inhibits aggregation. One aspect of the inhibitory mechanism involves changes in the cytoskeleton; however, the molecular mechanisms underlying platelet inhibition and its correlation with cytoskeletal cellular stiffness are poorly understood. We measured the cellular stiffness of individual platelets after treatment with the NO-GC stimulator riociguat or the NO-GC activator cinaciguat, using scanning ion conductance microscopy (SICM). We quantified changes in platelet shape using deep learning-based platelet morphometry. Cytoskeletal actin polymerization and platelet activation were measured by co-immunostaining F-actin and P-selectin, respectively. To test for clinical applicability of NO-GC stimulators in the context of increased thrombogenicity risk, we investigated the effect of riociguat on platelets from human immunodeficiency virus (HIV)-positive patients taking abacavir sulphate (ABC)-containing regimens, compared with HIV-negative volunteers. Stimulation of human and murine platelets with the NO-GC stimulator riociguat or with the NO-GC activator cinaciguat downregulated P-selectin expression, decreased F-actin polymerization, and decreased cellular stiffness by {approx}50%, compared to vehicle control. In addition, platelets became more circular, indicating decreased activation. Riociguat did not cause any change in platelet aggregation or circularity in HIV-positive patients taking ABC-containing regimens. These results corroborate a functional role of the NO-GC enzyme in platelet biomechanics (cellular stiffness) in correlation with the inhibition of platelet activation and morphological changes. The observed changes in stiffness and platelet shape therefore demonstrate the possibility of pharmacologically targeting the NO-GC/cGMP pathway.

biochemistry↗

bioRxiv2021

Loss of soluble guanylyl cyclase in platelets contributes to atherosclerotic plaque formation and vascular inflammation

AimThe role of platelets in atherosclerosis remains incompletely understood. Variants in genes encoding the soluble guanylyl cyclase (sGC) in platelets are associated with coronary artery disease (CAD) risk. Here we sought to investigate the contribution of platelet sGC to atherosclerosis and the therapeutic potential of targeting sGC in atherosclerosis. Methods and ResultsWe genetically deleted sGC in platelets of atherosclerosis-prone Ldlr-/- mice. By intravital fluorescence microscopy such Pf4-Cre+Gucy1b1flox/floxLdlr-/- mice displayed enhanced leukocyte adhesion to atherosclerotic plaques in comparison with their litter mates. Moreover, histological and flow cytometry analyses revealed more numerous inflammatory leukocytes and larger plaque sizes in aortic tissue of Ldlr-/- mice lacking sGC in platelets. In vitro, supernatant from activated platelets lacking sGC promoted leukocyte adhesion to endothelial cells (EC) via enhanced EC activation. Using cytokine profiling, we identified reduced angiopoietin-1 release by Pf4-Cre+Gucy1b1flox/flox and human GUCY1A1 risk allele carrier platelets to be responsible for enhanced activation of EC and subsequent leukocyte adhesion. Pharmacological sGC stimulation increased platelet angiopoietin-1 release in vitro and reduced recruitment of adoptively transferred leukocytes in Ldlr-/- mice fed a Western diet. Pharmacological sGC stimulation further reduced atherosclerotic plaque formation and vascular inflammation. ConclusionLoss of sGC in platelets contributes to atherosclerotic plaque formation via reduced release of the soluble factor angiopoietin-1 and, subsequently, enhanced leukocyte recruitment. Pharmacological sGC stimulation might represent a novel therapeutic strategy to prevent and treat CAD. Translational perspectiveReduced platelet soluble guanylyl cyclase activity contributes to atherosclerotic plaque formation and vascular inflammation. Stimulators of the soluble guanylyl cyclase, an emerging class of drugs already used in pulmonary hypertension and heart failure, are able to reduce atherosclerosis and inflammation in this preclinical model. Together with evidence from human genetics, our findings suggest a promising role of soluble guanylyl cyclase stimulation to prevent coronary artery disease.

physiology↗

bioRxiv2021

cGMP signaling: probing antagonistic cyclic nucleotide platelet signals by modeling and experiment

BackgroundThe cyclic nucleotides cAMP and cGMP inhibit platelet activation. ResultsWe extended an older model and systematically integrated drugs as external stimuli. Data driven modeling allowed us to design models that provide a quantitative output for quantitative input information. This relies on condensed information about involved regulation and modeling of pharmacological interventions by systematic optimization methods. By multi-experiment fitting, we validated our model optimizing the parameters of the model. In addition, we show how the output of the developed cGMP model can be used as input for a modular model of VASP phosphorylation and for the activity of cAMP and cGMP pathways in platelets. ConclusionsWe present a model for cGMP signaling and VASP phosphorylation, that allows to estimate drug action on any of the inhibitory cyclic nucleotide pathways (cGMP, cAMP) and has been validated by experimental data.

systems biology↗

bioRxiv2019

Room without a view - den construction in relation to body size in brown bears.

Hibernation is an adaptive strategy to survive harsh winter conditions and food shortage. The use of well-insulated winter dens helps animals minimize energy loss during hibernation. Brown bears (Ursus arctos) commonly use excavated dens for hibernation. Physical properties of excavated dens, such as the amount of space between a bear and the inner wall, wall/roof thickness, and bedding materials, are expected to impact heat retention and energy conservation of bears. The objective of this study was to examine the impact of physical properties of excavated dens on energy conservation in hibernating bears. Our hypothesis was that bears excavate dens in a way to minimize heat loss and optimize energy conservation during hibernation. We predicted that physical properties of excavated dens would significantly affect the bears post-hibernation body condition. To test our hypothesis and prediction, we analyzed data collected from brown bears in Sweden with linear mixed effects models, examining (i) what factors affect den-excavation behavior and (ii) if physical properties of excavated dens affect post-hibernation body condition. We found that bears excavated a den cavity in relation to their body size, that older bears tended to excavate better-fitting den cavities compared to young bears, and that the physical properties of excavated dens did not significantly affect a bears post-hibernation body condition. Older bears excavated better-fitting den cavities, suggesting a potentially experience-based shift with age in den-excavation behavior and an optimum cavity size relative to a bears body size. The strong year effect shown by the most parsimonious model for post-hibernation body condition suggests that variations in physical properties of excavated dens are possibly negligible, compared to the large annual variations in biotic and abiotic factors affecting pre-hibernation body condition and heat loss during hibernation.

ecology↗