The role of pannexin/purinergic signaling in intervascular communication from capillaries during skeletal muscle contraction in male Golden hamsters.

We sought to determine the physiological relevance of pannexin/purinergic-dependent signaling in mediating conducted vasodilation elicited by capillary stimulation through skeletal muscle contraction. Using hamster cremaster muscle and intravital microscopy we stimulated capillaries through local muscle contraction while observing the associated upstream arteriole. Capillaries were stimulated with muscle contraction at low and high contraction (6 and 60CPM) and stimulus frequencies (4 and 40 Hz) in the absence and presence of pannexin blocker mefloquine (MEF; 10-5 M), purinergic receptor antagonist suramin (SUR 10-5 M) and gap-junction uncoupler halothane (HALO, 0.07%) applied between the capillary stimulation site and the upstream arteriolar observation site. Conducted vasodilations elicited at 6CPM were inhibited by HALO while vasodilations at 60CPM were inhibited by MEF and SUR. The conducted response elicited at 4 Hz was inhibited by MEF while the vasodilation at 40 Hz was unaffected by any blocker. Therefore, upstream vasodilations resulting from capillary stimulation via muscle contraction are dependent upon a pannexin/purinergic-dependent pathway that appears to be stimulation parameter-dependent. Our data highlight a physiological importance of the pannexin/purinergic pathway in facilitating communication between capillaries and upstream arteriolar microvasculature and, consequently, indicating that this pathway may play a crucial role in regulating blood flow in response to skeletal muscle contraction.

Quantitative Capillary Refill Time with image-based finger force estimation.

Skin color observation provides a simple and non-invasive method to estimate the health status of patients. Capillary Refill Time (CRT) is widely used as an indicator of pathophysiological conditions, especially in emergency patients. While the measurement of CRT is easy to perform, its evaluation is highly subjective. This study proposes a method to aid quantified CRT measurement using an RGB camera. The procedure consists in applying finger compression to the forearm, and the CRT is calculated based on the skin color change after the pressure release. We estimate compression applied by a finger from its fingernail color change during compression. Our study shows a step towards camera-based quantitative CRT for untrained individuals.

Reliability of power output, maximal rate of capillary blood lactate accumulation, and phosphagen contribution time following 15-s sprint cycling in amateur cyclists.

Based on Mader's mathematical model, the rate of capillary blood lactate concentration (νLamax) following intense exercise is thought to reflect the maximal glycolytic rate. We aimed to investigate the reliability of important variables of Mader's model (i.e. power output, lactate accumulation, predominant phosphagen contribution time frames (tP Cr)) and resulting νLamax values derived during and after a 15-s cycling sprint. Fifty cyclists performed a 15-s all-out sprint test on a Cyclus2 ergometer three times. The first sprint test was considered a familiarization trial. Capillary blood was sampled before and every minute (for 8 min) after the sprint to determine νLamax. Test-retest analysis between T2 and T3 revealed excellent reliability for power output (Pmean and Ppeak; ICC = 0.99, 0.99), ∆La and νLamax with tPCr of 3.5 s (ICC = 0.91, 0.91). νLamax calculated with tPCr = tP peak (ICC = 0.87) and tP Cr = tPpeak-3.5% (ICC = 0.79) revealed good reliability. tPpeak and tPpeak-3.5% revealed only poor and moderate reliability (ICC = 0.41, 0.52). Power output and ∆La are reliable parameters in the context of this test. Depending on tPCr, reliability of νLamax varies considerably with tP Cr of 3.5 s showing excellent reliability. We recommend standardization of this type of testing especially tP Cr.

Impact of capillary and sarcolemmal proximity on mitochondrial structure and energetic function in skeletal muscle.

Mitochondria within skeletal muscle cells are located either between the muscle contractile apparatus (interfibrillar mitochondria, IFM) or beneath the cell membrane (subsarcolemmal mitochondria, SSM), with several structural and functional differences reported between IFM and SSM. However, recent 3D imaging studies demonstrate that mitochondria are particularly concentrated in the proximity of capillaries embedded in sarcolemmal grooves rather than in proximity to the sarcolemma itself (paravascular mitochondria, PVM). To evaluate the impact of capillary vs. sarcolemmal proximity, we compared the structure and function of skeletal muscle mitochondria located either lateral to embedded capillaries (PVM), adjacent to the sarcolemma but not in PVM pools (SSM) or interspersed between sarcomeres (IFM). Mitochondrial morphology and interactions were assessed by 3D electron microscopy coupled with machine learning segmentation, whereas mitochondrial energy conversion was assessed by two-photon microscopy of mitochondrial membrane potential, content, calcium, NADH redox and flux in live, intact cells. Structurally, although PVM and SSM were similarly larger than IFM, PVM were larger, rounder and had more physical connections to neighbouring mitochondria compared to both IFM and SSM. Functionally, PVM had similar or greater basal NADH flux compared to SSM and IFM, respectively, despite a more oxidized NADH pool and a greater membrane potential, signifying a greater activation of the electron transport chain in PVM. Together, these data indicate that proximity to capillaries has a greater impact on resting mitochondrial energy conversion and distribution in skeletal muscle than the sarcolemma alone. KEY POINTS: Capillaries have a greater impact on mitochondrial energy conversion in skeletal muscle than the sarcolemma. Paravascular mitochondria are larger, and the outer mitochondrial membrane is more connected with neighbouring mitochondria. Interfibrillar mitochondria are longer and have greater contact sites with other organelles (i.e. sarcoplasmic reticulum and lipid droplets). Paravascular mitochondria have greater activation of oxidative phosphorylation than interfibrillar mitochondria at rest, although this is not regulated by calcium.

Self-Organization of Long-Lasting Human Endothelial Capillary-Like Networks Guided by DLP Bioprinting.

Tissue engineering holds great promise for regenerative medicine, drug discovery, and as an alternative to animal models. However, as soon as the dimensions of engineered tissue exceed the diffusion limit of oxygen and nutriments, a necrotic core forms leading to irreversible damage. To overcome this constraint, the establishment of a functional perfusion network is essential. In this work, digital light processing bioprinting is used to encapsulate endothelial progenitor cells (EPCs) in 3D light-cured hydrogel scaffolds to guide them toward vascular network formation. In these scaffolds, EPCs proliferate and self-organize within a few days into branched tubular structures with predefined geometry, forming capillary-like vascular tubes or trees of diameters in the range of 10 to 100 µm. Presenting a confluent monolayer wall of cells strongly connect by tight junctions around a central lumen-like space, these structures can be microinjected with a fluorescent dye and are stable for several weeks in vitro. These endothelial structures can be recovered and manipulated in an alginate patch without altering their shape or viability. This approach opens new opportunities for future applications, such as stacking with other cell sheets or multicellular constructs to yield bioengineered tissue with higher complexity and functionality.

Capillary oxygen regulates demand-supply coupling by triggering connexin40-mediated conduction: Rethinking the metabolic hypothesis.

Coupling red blood cell (RBC) supply to O2 demand is an intricate process requiring O2 sensing, generation of a stimulus, and signal transduction that alters upstream arteriolar tone. Although actively debated, this process has been theorized to be induced by hypoxia and to involve activation of endothelial inwardly rectifying K+ channels (KIR) 2.1 by elevated extracellular K+ to trigger conducted hyperpolarization via connexin40 (Cx40) gap junctions to upstream resistors. This concept was tested in resting healthy skeletal muscle of Cx40-/- and endothelial KIR2.1-/- mice using state-of-the-art live animal imaging where the local tissue O2 environment was manipulated using a custom gas chamber. Second-by-second capillary RBC flow responses were recorded as O2 was altered. A stepwise drop in PO2 at the muscle surface increased RBC supply in capillaries of control animals while elevated O2 elicited the opposite response; capillaries were confirmed to express Cx40. The RBC flow responses were rapid and tightly coupled to O2; computer simulations did not support hypoxia as a driving factor. In contrast, RBC flow responses were significantly diminished in Cx40-/- mice. Endothelial KIR2.1-/- mice, on the other hand, reacted normally to O2 changes, even when the O2 challenge was targeted to a smaller area of tissue with fewer capillaries. Conclusively, microvascular O2 responses depend on coordinated electrical signaling via Cx40 gap junctions, and endothelial KIR2.1 channels do not initiate the event. These findings reconceptualize the paradigm of blood flow regulation in skeletal muscle and how O2 triggers this process in capillaries independent of extracellular K+.

Self-assembled and perfusable microvasculature-on-chip for modeling leukocyte trafficking.

Leukocyte recruitment from blood to tissue is a process that occurs at the level of capillary vessels during both physiological and pathological conditions. This process is also relevant for evaluating novel adoptive cell therapies, in which the trafficking of therapeutic cells such as chimeric antigen receptor (CAR)-T cells throughout the capillaries of solid tumors is important. Local variations in blood flow, mural cell concentration, and tissue stiffness contribute to the regulation of capillary vascular permeability and leukocyte trafficking throughout the capillary microvasculature. We developed a platform to mimic a biologically functional human arteriole-venule microcirculation system consisting of pericytes (PCs) and arterial and venous primary endothelial cells (ECs) embedded within a hydrogel, which self-assembles into a perfusable, heterogeneous microvasculature. Our device shows a preferential association of PCs with arterial ECs that drives the flow-dependent formation of microvasculature networks. We show that PCs stimulate basement membrane matrix synthesis, which affects both vessel diameter and permeability in a manner correlating with the ratio of ECs to PCs. Moreover, we demonstrate that hydrogel concentration can affect capillary morphology but has no observed effect on vascular permeability. The biological function of our capillary network was demonstrated using an inflammation model, where significantly higher expression of cytokines, chemokines, and adhesion molecules was observed after tumor necrosis factor-alpha (TNF-α) treatment. Accordingly, T cell adherence and transendothelial migration were significantly increased in the immune-activated state. Taken together, our platform allows the generation of a perfusable microvasculature that recapitulates the structure and function of an in vivo capillary bed that can be used as a model for developing potential immunotherapies.

Mechanical study of blood flow through a permeable capillary with slippery wall.

This research presents the mechanical behavior of blood flow through capillary having smooth inner surface. In this study modelling of blood flow via permeable and lubricated capillary caused by nutrients re-absorption has been done by the help of laws of momentum and mass. The nutrients re-absorption is assumed to be constant and inner walls of the capillary are smooth and slippery therefore slip condition on the velocity and constant rate in vertical direction at the wall has considered. The Kelvin Voigt model is employed to simulate blood flow via capillaries, and results for pressure, blood flow pattern, and shear force necessary for blood flow are discovered by recursive approach. Numerical results for nutrient re-absorption from the blood and impact of smooth and slippery surfaces on blood flow are shown through graphs. The novelty of the research invents that the smoothness and slickness of capillary wall is a crucial presumption to examine the blood as non-Newtonian fluid via capillary.

Estimation of shear stress heterogeneity along capillary segments in angiogenic rat mesenteric microvascular networks.

OBJECTIVE: Fluid shear stress is thought to be a regulator of endothelial cell behavior during angiogenesis. The link, however, requires an understanding of stress values at the capillary level in angiogenic microvascular networks. Critical questions remain. What are the stresses? Do capillaries experience similar stress magnitudes? Can variations explain vessel-specific behavior? The objective of this study was to estimate segment-specific shear stresses in angiogenic networks. METHODS: Images of angiogenic networks characterized by increased vascular density were obtained from rat mesenteric tissues stimulated by compound 48/80-induced mast cell degranulation. Vessels were identified by perfusion of a 40 kDa fixable dextran prior to harvesting and immunolabeling for PECAM. Using a network flow-based segment model with physiologically relevant parameters, stresses were computed per vessel for regions across multiple networks. RESULTS: Stresses ranged from 0.003 to 2328.1 dyne/cm2 and varied dramatically at the capillary level. For all regions, the maximum segmental shear stresses were for capillary segments. Stresses along proximal capillaries branching from arteriole inlets were increased compared to stresses along capillaries in more distal regions. CONCLUSIONS: The results highlight the variability of shear stresses along angiogenic capillaries and motivate new discussions on how endothelial cells may respond in vivo to segment-specific microenvironment during angiogenesis.

[Vascular biology and hemophilia].

By carrying a systemic circulation, hematopoietic and vascular systems coordinately govern the functional organ connections in the body. Blood vessels play an important role in the development, regeneration, and maintenance of organs by acting as conduits for environmental factors in the blood to tissues and secreting organ-specific cytokines as angiocrine signals. Recently, it has become clear that vascular endothelial cells, which are the main constituent cells of the blood vessels and play a role in homeostasis, are diverse. It has also been established that the cells of stem cell fraction exist in endothelial cells. The vascular endothelial cells in various organs are functionally different. For example, it has been discovered that sinusoidal blood vessels in the liver produce coagulation factor VIII as an organ-specific vascular function. Determining how such tissue-/organ-specific function of the endothelial cells is induced is a topic of interest in the vascular field of study.

Flow Heterogeneity and Factors Contributing to the Variability in Retinal Capillary Blood Flow.

Purpose: Capillary flow plays an important role in the nourishment and maintenance of healthy neural tissue and can be observed directly and non-invasively in the living human retina. Despite their importance, patterns of normal capillary flow are not well understood due to limitations in spatial and temporal resolution of imaging data. Methods: Capillary flow characteristics were studied in the retina of three healthy young individuals using a high-resolution adaptive optics ophthalmoscope. Imaging with frame rates of 200 to 300 frames per second was sufficient to capture details of the single-file flow of red blood cells in capillaries over the course of about 3 seconds. Results: Erythrocyte velocities were measured from 72 neighboring vessels of the parafoveal capillary network for each subject. We observed strong variability among vessels within a given subject, and even within a given imaged field, across a range of capillary flow parameters including maximum and minimum velocities, pulsatility, abruptness of the systolic peak, and phase of the cardiac cycle. The observed variability was not well explained by "local" factors such as the vessel diameter, tortuosity, length, linear cell density, or hematocrit of the vessel. Within a vessel, a moderate relation between the velocities and hematocrit was noted, suggesting a redistribution of plasma between cells with changes in flow. Conclusions: These observations advance our fundamental understanding of normal capillary physiology and raise questions regarding the potential role of network-level effects in explaining the observed flow heterogeneity.

Blood flow characterization in nailfold capillary using optical flow-assisted two-stream network and spatial-temporal image.

The blood flow velocity in the nailfold capillary is an important indicator of the status of microcirculation. The conventional manual processing method is both laborious and prone to human artifacts. A feasible way to solve this problem is to use machine learning to assist in image processing and diagnosis. Inspired by the Two-Stream Convolutional Networks, this study proposes an optical flow-assisted two-stream network to segment nailfold blood vessels. Firstly, we use U-Net as the spatial flow network and the dense optical flow as the temporal stream. The results show that the optical flow information can effectively improve the integrity of the segmentation of blood vessels. The overall accuracy is 94.01 %, the Dice score is 0.8099, the IoU score is 0.6806, and the VOE score is 0.3194. Secondly, The flow velocity of the segmented blood vessel is determined by constructing the spatial-temporal (ST) image. The blood flow velocity evaluated is consistent with the typical blood flow speed reported. This study proposes a novel two-stream network for blood vessel segmentation of nailfold capillary images. Combined with ST image and line detection method, it provides an effective workflow for measuring the blood flow velocity of nailfold capillaries.

Hematoloechus sp. attachment shifts endothelium in vivo from pro- to anti-inflammatory profile in Rana pipiens: evidence from systemic and capillary physiology.

This prospective, descriptive study focused on lung flukes (Hematoloechus sp., H) and their impact on systemic and individual capillary variables measured in pithed Rana pipiens, a long-standing model for studies of capillary physiology. Three groups were identified based on Hematoloechus attachment: no Hematoloechus (No H), Hematoloechus not attached (H Not Att), and Hematoloechus attached (H Att). Among 38 descriptive, cardiovascular, and immunological variables, 18 changed significantly with H. Symptoms of H included weight loss, elevated immune cells, heart rate variability, faster coagulation, lower hematocrit, and fluid accumulation. Important capillary function discoveries included median baselines for hydraulic conductivity (Lp) of 7.0 (No H), 12.4 (H Not Att), and 4.2 (H Att) × 10-7 cm·s-1·cmH2O-1 (P < 0.0001) plus seasonal adaptation of sigma delta pi [σ(πc-πi), P = 0.03]. Pro- and anti-inflammatory phases were revealed for Lp and plasma nitrite/nitrate concentration ([NOx]) in both H Not Att and H Att, whereas capillary wall tensile strength increased in the H Att. H attachment was advantageous for the host due to lower edema and for the parasite via a sustained food source illustrating an excellent example of natural symbiosis. However, H attachment also resulted in host weight loss: in time, a conundrum for the highly dependent parasite. The study increases overall knowledge of Rana pipiens by revealing intriguing effects of H and previously unknown, naturally occurring seasonal changes in many variables. The data improve Rana pipiens as a general scientific and capillary physiology model. Diseases of inflammation and stroke are among the clinical applications.

Tissue Oxygenation Around Capillaries: Effects of Hematocrit and Arteriole Oxygen Condition.

Oxygen transfer in the microvasculature is a complex phenomenon that involves multiple physical and chemical processes and multiple media. Hematocrit, the volume fraction of red blood cells (RBCs) in blood, has direct influences on the blood flow as well as the oxygen supply in the microcirculation. On the one hand, a higher hematocrit means that more RBCs present in capillaries, and thus, more oxygen is available at the source end. On the other hand, the flow resistance increases with hematocrit, and therefore, the RBC motion becomes slower, which in turn reduces the influx of oxygen-rich RBCs entering capillaries. Such double roles of hematocrit have not been investigated adequately. Moreover, the oxygen-hemoglobin dissociation rate depends on the oxygen tension and hemoglobin saturation of the cytoplasm inside RBCs, and the dissociation kinetics exhibits a nonlinear fashion at different oxygen tensions. To understand how these factors and mechanisms interplay in the oxygen transport process, computational modeling and simulations are favorite since we have a good control of the system parameters and also we can access to the detailed information during the transport process. In this study, we conduct numerical simulations for the blood flow and RBC deformation along a capillary and the oxygen transfer from RBCs to the surrounding tissue. Different values for the hematocrit, arteriole oxygen tension, tissue metabolism rate and hemoglobin concentration and affinity are considered, and the simulated spatial and temporal variations of oxygen concentration are analyzed in conjunction with the nonlinear oxygen-hemoglobin reaction kinetics. Our results show that there are two competing mechanisms for the tissue oxygenation response to a hematocrit increases: the favorite effect of the higher RBC density and the negative effect of the slower RBC motion. Moreover, in the low oxygen situations with RBC oxygen tension less than 50 mmHg at capillary inlet, the reduced RBC velocity effect dominates, resulting in a decrease in tissue oxygenation at higher hematocrit. On the opposite, for RBC oxygen tension higher than 50 mmHg when entering the capillary, a higher hematocrit is beneficial to the tissue oxygenation. More interestingly, the pivoting arteriole oxygen tension at which the two competing mechanisms switch dominance on tissue oxygenation becomes lower for higher oxygen-hemoglobin affinity and lower hemoglobin concentration. This observation has also been analyzed based on the oxygen supply from RBCs and the oxygen-hemoglobin reaction kinetics. The results and discussions presented in this article could be helpful for a better understanding of oxygen transport in microcirculation.