Cardiovascular sex-differences: insights via physiology-based modeling and potential for noninvasive sensing via ballistocardiography.

In this study, anatomical and functional differences between men and women in their cardiovascular systems and how these differences manifest in blood circulation are theoretically and experimentally investigated. A validated mathematical model of the cardiovascular system is used as a virtual laboratory to simulate and compare multiple scenarios where parameters associated with sex differences are varied. Cardiovascular model parameters related with women's faster heart rate, stronger ventricular contractility, and smaller blood vessels are used as inputs to quantify the impact (i) on the distribution of blood volume through the cardiovascular system, (ii) on the cardiovascular indexes describing the coupling between ventricles and arteries, and (iii) on the ballistocardiogram (BCG) signal. The model-predicted outputs are found to be consistent with published clinical data. Model simulations suggest that the balance between the contractile function of the left ventricle and the load opposed by the arterial circulation attains similar levels in females and males, but is achieved through different combinations of factors. Additionally, we examine the potential of using the BCG waveform, which is directly related to cardiovascular volumes, as a noninvasive method for monitoring cardiovascular function. Our findings provide valuable insights into the underlying mechanisms of cardiovascular sex differences and may help facilitate the development of effective noninvasive cardiovascular monitoring methods for early diagnosis and prevention of cardiovascular disease in both women and men.

Multi-focus Image Fusion for Confocal Microscopy Using U-Net Regression Map.

Characterizing the spatial relationship between blood vessel and lymphatic vascular structures, in the mice dura mater tissue, is useful for modeling fluid flows and changes in dynamics in various disease processes. We propose a new deep learning-based approach to fuse a set of multi-channel single-focus microscopy images within each volumetric z-stack into a single fused image that accurately captures as much of the vascular structures as possible. The red spectral channel captures small blood vessels and the green fluorescence channel images lymphatics structures in the intact dura mater attached to bone. The deep architecture Multi-Channel Fusion U-Net (MCFU-Net) combines multi-slice regression likelihood maps of thin linear structures using max pooling for each channel independently to estimate a slice-based focus selection map. We compare MCFU-Net with a widely used derivative-based multi-scale Hessian fusion method [8]. The multi-scale Hessian-based fusion produces dark-halos, non-homogeneous backgrounds and less detailed anatomical structures. Perception based no-reference image quality assessment metrics PIQUE, NIQE, and BRISQUE confirm the effectiveness of the proposed method.

Fluid and protein exchange in microvascular networks: Importance of modelling heterogeneity in geometrical and biophysical properties.

KEY POINTS: Microvascular network architecture defines coupling of fluid and protein exchange. Network arrangements markedly reduce capillary hydrostatic pressures and resting fluid movement at the same time as increasing the capacity for change The presence of vascular remodelling or angiogenesis puts constraints of network behaviour The sites of fluid and protein exchange can be segregated to different portions of the network Although there is a net filtration of fluid from a network of exchange vessels, there are specific areas where fluid moves into the circulation (reabsorption) and, when protein is moving into tissue, the amount is insufficient under basal conditions to result in changes in oncotic pressure. ABSTRACT: Integration of functional results obtained across scales, from chemical signalling to the whole organism, is a daunting task requiring the marriage of experimental data with mathematical modelling. In the present study, a novel coupled computational fluid dynamics model is developed incorporating fluid and protein transport using measurements in an in vivo frog (Rana pipiens) mesenteric microvascular network. The influences of network architecture and exchange are explored systematically under the common assumptions of structurally and functionally identical microvessels (Homogeneous Scenario) or microvessels classified by position in flow (Class Uniform Scenario), which are compared with realistic microvascular network components (Heterogeneous Scenario). The model incorporates ten quantities that vary within a microvessel; pressure boundary conditions are calibrated against experimental measurements. The Homogeneous Scenario standard model showed that assuming a single 'typical' capillary hides the influence of vessels arranged into a network architecture, where capillary hydrostatic pressures (pT ) are reduced, resulting in both a nonuniform distribution of blood flow and reduced volume flow rate (Jf,T ). In the Class Uniform Scenario pT was further attenuated to produce a ∼60% reduction in Jf,T . Finally, the Heterogeneous Scenario, incorporating measures of individual vessel surface area, demonstrates additional lowering of pT from inlet values favouring a >70% reduction of Jf,T in the face of a ∼120% increase in protein movement into the tissues relative to the Homogeneous Scenario. Beyond the impacts of network architecture, an unanticipated finding was the influence of a blind-end microvessel on model convergence, indicating a profound influence of the largely unexplored dynamics of vascular remodelling on tissue perfusion.

Microvascular Sex- and Age- Dependent Phosphodiesterase Expression.

Objective: The cyclic nucleotide second messengers, cAMP and cGMP, are pivotal regulators of vascular functions; their cellular levels are tightly controlled by the cyclic nucleotide hydrolases, phosphodiesterases (PDE). Biologic sex and age are recognized as independent factors impacting the mechanisms mediating both vascular health and dysfunction. This study focused on microvessels isolated from male and female rats before (juvenile) and after (adult) sexual maturity under resting conditions. We tested the hypothesis that sexual dimorphism in microvascular PDE expression would be absent in juvenile rats, but would manifest in adult rats. Methods: Abdominal skeletal muscle arterioles and venules were isolated from age-matched juvenile and adult male and female rats under resting conditions. Transcripts of five PDE families (1-5) associated with coronary and vascular function with a total of ten genes were measured using TaqMan real-time RT-PCR and protein expression of microvessel PDE4 was assessed using immunoblotting and immunofluorescence. Results: Overall expression levels of PDE5A were highest while PDE3 levels were lowest among the five PDE families (p < 0.05) regardless of age or sex. Contrary to our hypothesis, in juveniles, sexual dimorphism in PDE expression was observed in three genes: arterioles (PDE1A, female > male) and venules (PDE1B and 3A, male > female). In adults, gene expression levels in males were higher than females for five genes in arterioles (PDE1C, 3A, 3B, 4B, 5A) and three genes (PDE3A, 3B, and 5A) in venules. Furthermore, age-related differences were observed in PDE1-5 (in males, adult > juvenile for most genes in arterioles; in females, adult > juvenile for arteriolar PDE3A; juvenile gene expression > adult for two genes in arterioles and three genes in venules). Immunoblotting and immunofluorescence analysis revealed protein expression of microvessel PDE4. Conclusion: This study revealed sexual dimorphism in both juvenile and adult rats, which is inconsistent with our hypothesis. The sex- and age-dependent differences in PDE expression implicate different modulations of cAMP and cGMP pathways for microvessels in health. The implication of these sex- and age-dependent differences, as well as the duration and microdomain of PDE1-5 activities in skeletal muscle microvessels, in both health and disease, require further investigation.

Corrections to "Cardiovascular Function and Ballistocardiogram: A Relationship Interpreted via Mathematical Modeling" [Oct 19 2906-2917].

Presents a missing funding acknowledgment for Dr. Mihail Popescu in the above named paper.

Complex Non-sinus-associated Pachymeningeal Lymphatic Structures: Interrelationship With Blood Microvasculature.

The contribution of cranial dura mater vascular networks, as means for maintaining brain fluid movement and balance, and as the source of significant initiators and/or contributors to neurological disorders, has been overlooked. These networks consist of both blood and lymphatic vessels. The latter were discovered recently and described as sinus-associated structures thus changing the old paradigm that central nervous system lacks lymphatics. In this study, using markers specific to blood and lymphatic endothelia, we demonstrate the existence of the complex non-sinus-associated pachymeningeal lymphatic vasculature. We further show the interrelationship and possible connections between lymphatic vessels and the dural blood circulatory system. Our novel findings reveal the presence of lymphatic-like structures that exist on their own and/or in close proximity to microvessels. Of particular interest are sub-sets of vascular complexes with dual (lymphatic and blood) vessel identity representing a unique microenvironment within the cranial dura. The close association of the systemic blood circulation and meningeal lymphatics achieved in these complexes could facilitate fluid exchange between the two compartments and constitute an alternative route for CSF drainage.

Cardiovascular Function and Ballistocardiogram: A Relationship Interpreted via Mathematical Modeling.

OBJECTIVE: To develop quantitative methods for the clinical interpretation of the ballistocardiogram (BCG). METHODS: A closed-loop mathematical model of the cardiovascular system is proposed to theoretically simulate the mechanisms generating the BCG signal, which is then compared with the signal acquired via accelerometry on a suspended bed. RESULTS: Simulated arterial pressure waveforms and ventricular functions are in good qualitative and quantitative agreement with those reported in the clinical literature. Simulated BCG signals exhibit the typical I, J, K, L, M, and N peaks and show good qualitative and quantitative agreement with experimental measurements. Simulated BCG signals associated with reduced contractility and increased stiffness of the left ventricle exhibit different changes that are characteristic of the specific pathological condition. CONCLUSION: The proposed closed-loop model captures the predominant features of BCG signals and can predict pathological changes on the basis of fundamental mechanisms in cardiovascular physiology. SIGNIFICANCE: This paper provides a quantitative framework for the clinical interpretation of BCG signals and the optimization of BCG sensing devices. The present paper considers an average human body and can potentially be extended to include variability among individuals.

Sex-Specific Characteristics of the Microcirculation.

The requirements of metabolizing tissue are both continuous and variable; accordingly, the microvasculature serving that tissue must be similarly dynamic. Just as it is recognized that males and females of the same species have differing metabolic requirements, is it not likely that the microvasculature serving these tissues will differ by sex? This section focusing on the constituents of the microcirculation identifies what is known presently about the role sex plays in matching metabolic demand with microvascular function and areas requiring additional study. Many of the identified sex differences are subtle and easily ignored. In the aggregate, though, they can profoundly alter phenotype, especially under stressful conditions including pregnancy, exercise, and disease states ranging from diabetes to heart failure. Although the features presently identified to "have sex" range from differences in growth, morphology, protein expression, and intracellular signaling, males and females alike achieve homeostasis, likely by different means. Studies of microvascular sexual dimorphism are also identifying age as an independent but interacting factor requiring additional attention. Overall, attempting to ignore either sex and/or age is inappropriate and will prevent the design and implementation of appropriate interventions to present, ameliorate, or correct microvascular dysfunction.

Sex differences influencing micro- and macrovascular endothelial phenotype in vitro.

KEY POINTS: Endothelial dysfunction is an early hallmark of multiple disease states that also display sex differences with respect to age of onset, frequency and severity. Results of in vivo studies of basal and stimulated microvascular barrier function revealed sex differences that are difficult to ascribe to specific cells or environmental factors. The present study evaluated endothelial cells (EC) isolated from macro- and/or microvessels of reproductively mature rats under the controlled conditions of low-passage culture aiming to test the assumption that EC phenotype would be sex independent. The primary finding was that EC, regardless of where they are derived, retain a sex-bias in low-passage culture, independent of varying levels of reproductive hormones. The implications of the present study include the fallacy of expecting a universal set of mechanisms derived from study of EC from one sex and/or one vascular origin to apply uniformly to all EC under unstimulated conditions, and no less in disease. ABSTRACT: Vascular endothelial cells (EC) are heterogeneous with respect to phenotype, reflecting at least the organ of origin, location within the vascular network and physical forces. As an independent influence on EC functions in health or aetiology, susceptibility, and progression of dysfunction in numerous disease states, sex has been largely ignored. The present study focussed on EC isolated from aorta (macrovascular) and skeletal muscle vessels (microvascular) of age-matched male and female rats under identical conditions of short-term (passage 4) culture. We tested the hypothesis that genomic sex would not influence endothelial growth, wound healing, morphology, lactate production, or messenger RNA and protein expression of key proteins (sex hormone receptors for androgen and oestrogens α and ß; platelet endothelial cell adhesion molecule-1 and vascular endothelial cadherin mediating barrier function; αv ß3 and N-cadherin influencing matrix interactions; intracellular adhesion molecule-1 and vascular cell adhesion molecule-1 mediating EC/white cell adhesion). The hypothesis was rejected because the EC origin (macro- vs. microvessel) and sex influenced multiple phenotypic characteristics. Statistical model analysis of EC growth demonstrated an hierarchy of variable importance, recapitulated for other phenotypic characteristics, with predictions assuming EC homogeneity < sex < vessel origin < sex and vessel origin. Furthermore, patterns of EC mRNA expression by vessel origin and by sex did not predict protein expression. Overall, the present study demonstrated that accurate assessment of sex-linked EC dysfunction first requires an understanding of EC function by position in the vascular tree and by sex. The results from a single EC tissue source/species/sex cannot provide universal insight into the mechanisms regulating in vivo endothelial function in health, and no less in disease.

Patch-Based Semantic Segmentation for Detecting Arterioles and Venules in Epifluorescence Imagery.

Segmentation and quantification of microvasculature structures are the main steps toward studying microvasculature remodeling. The proposed patch based semantic architecture enables accurate segmentation for the challenging epifluorescence microscopy images. Our pixel-based fast semantic network trained on random patches from different epifluorescence images to learn how to discriminate between vessels versus nonvessels pixels. The proposed semantic vessel network (SVNet) relies on understanding the morphological structure of the thin vessels in the patches rather than considering the whole image as input to speed up the training process and to maintain the clarity of thin structures. Experimental results on our ovariectomized - ovary removed (OVX) - mice dura mater epifluorescence microscopy images shows promising results in both arteriole and venule part. We compared our results with different segmentation methods such as local, global thresholding, matched based filter approaches and related state of the art deep learning networks. Our overall accuracy (> 98%) outperforms all the methods including our previous work (VNet). [1].

Estrogen-Dependent Changes in Dura Mater Microvasculature Add New Insights to the Pathogenesis of Headache.

The pathogenesis of headaches is a matter of ongoing discussion of two major theories describing it either as a vascular phenomenon resulting from vasodilation or primarily as a neurogenic process accompanied by secondary vasodilation associated with sterile neurogenic inflammation. While summarizing current views on neurogenic and vascular origins of headache, this mini review adds new insights regarding how smooth muscle-free microvascular networks, discovered within dura mater connective tissue stroma (previously thought to be "avascular"), may become a site of initial insult generating the background for the development of headache. Deficiencies in estrogen-dependent control of microvascular integrity leading to plasma protein extravasation, potential activation of perivascular and connective tissue stroma nociceptive neurons, and triggering of inflammatory responses are described. Finally, possible avenues for controlling and preventing these pathophysiological changes are discussed.

Confocal Vessel Structure Segmentation with Optimized Feature Bank and Random Forests.

In this paper, we consider confocal microscopy based vessel segmentation with optimized features and random forest classification. By utilizing multi-scale vessel-specific features tuned to capture curvilinear structures such as Frobenius norm of the Hessian eigenvalues, Laplacian of Gaussians (LoG), oriented second derivative, line detector and intensity masked with LoG scale map. we obtain better segmentation results in challenging imaging conditions. We obtain binary segmentations using random forest classifier trained on physiologists marked ground-truth. Experimental results on mice dura mater confocal microscopy vessel segmentations indicate that we obtain better results compared to global segmentation approaches.

Random Forests for Dura Mater Microvasculature Segmentation Using Epifluorescence Images.

Automatic segmentation of microvascular structures is a critical step in quantitatively characterizing vessel remodeling and other physiological changes in the dura mater or other tissues. We developed a supervised random forest (RF) classifier for segmenting thin vessel structures using multiscale features based on Hessian, oriented second derivatives, Laplacian of Gaussian and line features. The latter multiscale line detector feature helps in detecting and connecting faint vessel structures that would otherwise be missed. Experimental results on epifluorescence imagery show that the RF approach produces foreground vessel regions that are almost 20 and 25 percent better than Niblack and Otsu threshold-based segmentations respectively.

Multiquadric Spline-Based Interactive Segmentation of Vascular Networks.

Commonly used drawing tools for interactive image segmentation and labeling include active contours or boundaries, scribbles, rectangles and other shapes. Thin vessel shapes in images of vascular networks are difficult to segment using automatic or interactive methods. This paper introduces the novel use of a sparse set of user-defined seed points (supervised labels) for precisely, quickly and robustly segmenting complex biomedical images. A multiquadric spline-based binary classifier is proposed as a unique approach for interactive segmentation using as features color values and the location of seed points. Epifluorescence imagery of the dura mater microvasculature are difficult to segment for quantitative applications due to challenging tissue preparation, imaging conditions, and thin, faint structures. Experimental results based on twenty epifluorescence images is used to illustrate the benefits of using a set of seed points to obtain fast and accurate interactive segmentation compared to four interactive and automatic segmentation approaches.

The P2Y2 Receptor Interacts with VE-Cadherin and VEGF Receptor-2 to Regulate Rac1 Activity in Endothelial Cells.

Vascular endothelial cadherin (VE-cadherin) mediates homophylic adhesion between endothelial cells and is an important regulator of angiogenesis, blood vessel permeability and leukocyte trafficking. Rac1, a member of the Rho family of GTPases, controls VE-cadherin adhesion by acting downstream of several growth factors, including angiopoietin-1 and vascular endothelial growth factor (VEGF). Here we show that UTP-induced activation of the Gq protein-coupled P2Y2 nucleotide receptor (P2Y2R) in human coronary artery endothelial cells (HCAECs) activated Rac1 and caused a transient complex to form between P2Y2R, VE-cadherin and VEGF receptor-2 (VEGFR-2). Knockdown of VE-cadherin expression with siRNA did not affect UTP-induced activation of extracellular signal-regulated kinases 1/2 (ERK1/2) but led to a loss of UTP-induced Rac1 activation and tyrosine phosphorylation of p120 catenin, a cytoplasmic protein known to interact with VE-cadherin. Activation of the P2Y2R by UTP also caused a prolonged interaction between p120 catenin and vav2 (a guanine nucleotide exchange factor for Rac) that correlated with the kinetics of UTP-induced tyrosine phosphorylation of p120 catenin and VE-cadherin. Inhibitors of VEGFR-2 (SU1498) or Src (PP2) significantly diminished UTP-induced Rac1 activation, tyrosine phosphorylation of p120 catenin and VE-cadherin, and association of the P2Y2R with VE-cadherin and p120 catenin with vav2. These findings suggest that the P2Y2R uses Src and VEGFR-2 to mediate association of the P2Y2R with VE-cadherin complexes in endothelial adherens junctions to activate Rac1.

Multi-focus image fusion using epifluorescence microscopy for robust vascular segmentation.

Automatic segmentation of three-dimensional mi-crovascular structures is needed for quantifying morphological changes to blood vessels during development, disease and treatment processes. Single focus two-dimensional epifluorescent imagery lead to unsatisfactory segmentations due to multiple out of focus vessel regions that have blurred edge structures and lack of detail. Additional segmentation challenges include varying contrast levels due to diffusivity of the lectin stain, leakage out of vessels and fine morphological vessel structure. We propose an approach for vessel segmentation that combines multi-focus image fusion with robust adaptive filtering. The robust adaptive filtering scheme handles noise without destroying small structures, while multi-focus image fusion considerably improves segmentation quality by deblurring out-of-focus regions through incorporating 3D structure information from multiple focus steps. Experiments using epifluorescence images of mice dura mater show an average of 30.4% improvement compared to single focus microvasculature segmentation.

Pulsed estrogen therapy prevents post-OVX porcine dura mater microvascular network weakening via a PDGF-BB-dependent mechanism.

In postmenopausal women, estrogen (E2) deficiencies are frequently associated with higher risk of intracranial hemorrhage, increased incidence of stroke, cerebral aneurysm, and decline in cognitive abilities. In younger postpartum women and those using oral contraceptives, perturbations in E2 are associated with higher risk of cerebral venous thrombosis. A number of serious intracranial pathologic conditions linked to E2 deficiencies, such as dural sinus thrombosis, dural fistulae, non-parenchymal intracranial hemorrhages, migraines, and spontaneous cerebrospinal fluid leaks, involve the vessels not of the brain itself, but of the outer fibrous membrane of the brain, the dura mater (DM). The pathogenesis of these disorders remains mysterious and how estrogen regulates structural and functional integrity of DM vasculature is largely unknown. Here, we demonstrate that post ovariectomy (OVX) DM vascular remodeling is manifested by microvessel destabilization, capillary rarefaction, increased vascular permeability, and aberrant angio-architecture, and is the result of disrupted E2-regulated PDGF-BB signaling within dura microvasculature. These changes, associated with the reduction in systemic PDGF-BB levels, are not corrected by a flat-dose E2 hormone replacement therapy (HRT), but are largely prevented using HRT schedules mimicking physiological E2 fluctuations. We demonstrate that 1) E2 regulates PDGF-BB production by endothelial cells in a dose-dependent manner and 2) optimization of PDGF-BB levels and induction of robust PDGF-mediated endothelial cell-vascular pericyte interactions require high (estrous) E2 concentrations. We conclude that high (estrous) levels of E2 are important in controlling PDGF-mediated crosstalk between endothelial cells and pericytes, a fundamental mechanism governing microvessel stability and essential for preserving intracranial homeostasis.

Permeability and contractile responses of collecting lymphatic vessels elicited by atrial and brain natriuretic peptides.

Atrial and brain natriuretic peptides (ANP and BNP, respectively) are cardiac hormones released into the bloodstream in response to hypervolaemia or fluid shifts to the central circulation. The actions of both peptides include natriuresis and diuresis, a decrease in systemic blood pressure, and inhibition of the renin-angiotensin-aldosterone system. Further, ANP and BNP elicit increases in blood microvessel permeability sufficient to cause protein and fluid extravasation into the interstitium to reduce the vascular volume. Given the importance of the lymphatic vasculature in maintaining fluid balance, we tested the hypothesis that ANP or BNP (100 nM) would likewise elevate lymphatic permeability (Ps) to serum albumin. Using a microfluorometric technique adapted to in vivo lymphatic vessels, we determined that rat mesenteric collecting lymphatic Ps to rat serum albumin increased by 2.0 ± 0.4-fold (P = 0.01, n = 7) and 2.7 ± 0.8-fold (P = 0.07, n = 7) with ANP and BNP, respectively. In addition to measuring Ps responses, we observed changes in spontaneous contraction amplitude and frequency from the albumin flux tracings in vivo. Notably, ANP abolished spontaneous contraction amplitude (P = 0.005) and frequency (P = 0.006), while BNP augmented both parameters by ∼2-fold (P < 0.01 each). These effects of ANP and BNP on contractile function were examined further by using an in vitro assay. In aggregate, these data support the theory that an increase in collecting lymphatic permeability opposes the absorptive function of the lymphatic capillaries, and aids in the retention of protein and fluid in the interstitial space to counteract volume expansion.

Endothelial barrier dysfunction in diabetic conduit arteries: a novel method to quantify filtration.

The endothelial barrier plays an important role in atherosclerosis, hyperglycemia, and hypercholesterolemia. In the present study, an accurate, reproducible, and user-friendly method was used to further understand endothelial barrier function of conduit arteries. An isovolumic method was used to measure the hydraulic conductivity (L(p)) of the intact vessel wall and medial-adventitial layer. Normal arterial segments with diameters from 0.2 to 5.5 mm were used to validate the method, and femoral arteries of diabetic rats were studied as an example of pathological specimens. Various arterial segments confirmed that the volume flux of water per unit surface area was linearly related to intraluminal pressure, as confirmed in microvessels. L(p) of the intact wall varied from 3.5 to 22.1 × 10(-7) cm·s(-1)·cmH(2)O(-1) over the pressure range of 7-180 mmHg. Over the same pressure range, L(p) of the endothelial barrier changed from 4.4 to 25.1 × 10(-7) cm·s(-1)·cmH(2)O(-1). During perfusion with albumin-free solution, L(p) of rat femoral arteries increased from 6.1 to 13.2 × 10(-7) cm·s(-1)·cmH(2)O(-1) over the pressure range of 10-180 mmHg. Hyperglycemia increased L(p) of the femoral artery in diabetic rats from 2.9 to 5.5 × 10(-7) cm·s(-1)·cmH(2)O(-1) over the pressure range of 20-135 mmHg. In conclusion, the L(p) of a conduit artery can be accurately and reproducibly measured using a novel isovolumic method, which in diabetic rats is hyperpermeable. This is likely due to disruption of the endothelial glycocalyx.