Xenobiotic particles can be considered in two genres: air pollution particulate matter and engineered nanoparticles. Particle exposures can occur in the greater environment, the workplace, and our homes. The majority of research in this field has, justifiably, focused on pulmonary reactions and outcomes. More recent investigations indicate that cardiovascular effects are capable of correlating with established mortality and morbidity epidemiological data following particle exposures. While the preliminary and general cardiovascular toxicology has been defined, the mechanisms behind these effects, specifically within the microcirculation, are largely unexplored. Therefore, the purpose of this review is several fold: first, a historical background on toxicological aspects of particle research is presented. Second, essential definitions, terminology, and techniques that may be unfamiliar to the microvascular scientist will be discussed. Third, the most current concepts and hypotheses driving cardiovascular research in this field will be reviewed. Lastly, potential future directions for the microvascular scientist will be suggested. Collectively speaking, microvascular research in the particle exposure field represents far more than a “niche”. The immediate demand for basic, translational, and clinical studies is high and diverse. Microvascular scientists at all career stages are strongly encouraged to expand their research interests to include investigations associated with particle exposures.

Objective

We examined the effects of exogenously delivered thrombin on cell recruitment in skeletal muscle and the formation of new collateral arterioles in the microvasculature in response to ligation-induced ischemia.

Methods

Thrombin or vehicle was locally applied to both ligated and non-operated Balb/c spinotrapezius muscles which were harvested after three or seven days, imaged using confocal microscopy, and analyzed.

Results

Thrombin treatment resulted in accelerated arterialization of collateral capillaries and accelerated tissue reperfusion in ischemic muscles. Uninjured muscle treated with thrombin displayed increased vascular cell adhesion molecule 1 expression on arteriole and venule endothelium, increased expression of smooth muscle α-actin on capillary-sized vessels, increased infiltration by CD11b+ leukocytes, and mast cell infiltration and degranulation.

Conclusion

Exogenous delivery of thrombin enhances microvascular collateral development in response to ischemic insult and accelerates tissue reperfusion. Elicited responses from multiple cell types likely contribute to these effects.

To study the effect of myoendothelial communication on vascular reactivity, we integrated detailed mathematical models of Ca2+ dynamics and membrane electrophysiology in arteriolar smooth muscle (SMC) and endothelial (EC) cells. Cells are coupled through the exchange of Ca2+, Cl−, K+, and Na+ ions, inositol 1,4,5-triphosphate (IP3), and the paracrine diffusion of nitric oxide (NO). EC stimulation reduces intracellular Ca2+ ([Ca2+]i) in the SMC by transmitting a hyperpolarizing current carried primarily by K+. The NO-independent endothelium-derived hyperpolarization was abolished in a synergistic-like manner by inhibition of EC SKCa and IKCa channels. During NE stimulation, IP3 diffusing from the SMC induces EC Ca2+ release, which, in turn, moderates SMC depolarization and [Ca2+]i elevation. On the contrary, SMC [Ca2+]i was not affected by EC-derived IP3. Myoendothelial Ca2+ fluxes had no effect in either cell. The EC exerts a stabilizing effect on calcium-induced calcium release–dependent SMC Ca2+ oscillations by increasing the norepinephrine concentration window for oscillations. We conclude that a model based on independent data for subcellular components can capture major features of the integrated vessel behavior. This study provides a tissue-specific approach for analyzing complex signaling mechanisms in the vasculature.

Blood is modeled as a suspension of red blood cells using the dissipative particle dynamics method. The red blood cell membrane is coarse-grained for efficient simulations of multiple cells, yet accurately describes its viscoelastic properties. Blood flow in microtubes ranging from 10 to 40 μm in diameter is simulated in three dimensions for values of hematocrit in the range of 0.15–0.45 and carefully compared with available experimental data. Velocity profiles for different hematocrit values show an increase in bluntness with an increase in hematocrit. Red blood cell center-of-mass distributions demonstrate cell migration away from the wall to the tube center. This results in the formation of a cell-free layer next to the tube wall corresponding to the experimentally observed Fahraeus and Fahraeus–Lindqvist effects. The predicted cell-free layer widths are in agreement with those found in in vitro experiments; the results are also in qualitative agreement with in vivo experiments. However, additional features have to be taken into account for simulating microvascular flow, e.g., the endothelial glycocalyx. The developed model is able to capture blood flow properties and provides a computational framework at the mesoscopic level for obtaining realistic predictions of blood flow in microcirculation under normal and pathological conditions.

Objective

Provide regional flow measurement in the hearts of small mammals using a new, higher-resolution technique based on the deposition of a molecular marker.

Methods

We determined the instantaneous extraction and retention of the “molecular microsphere” radiolabeled desmethylimipramine in retrogradely perfused hamster hearts. In a separate series of experiments, autoradiography was used to measure regional myocardial deposition densities in hamster hearts of about 0.5 g with spatial area resolution of 16 × 16 μm.

Results

Radiolabeled desmethylimipramine is almost 100% extracted during a single transcapillary passage and is retained in the tissue for many minutes. Autoradiographic images demonstrated a spatial flow heterogeneity with standard deviations of 31 ± 4% of the mean flow (N = 5) in 16 × 16 × 20-μm3 voxels. This is equivalent to the projections made using fractal relationships from cruder observations obtained with microspheres in the hearts of baboons, sheep, and rabbits.

Conclusion

Autoradiography using a molecular deposition marker provides quantitative information on myocardial flow heterogeneities with resolution at the size of cardiac myocytes. Because the regions resolved are smaller than the volume of regions supplied by single arterioles, the results must slightly exaggerate the true heterogeneity of regional flows.

Objective

Keratin proteins have been utilized as biomaterials for decades and are currently under investigation for a variety of tissue regeneration and trauma applications. It has been suggested that certain keratins may have the capacity to act as a colloid in fluid resuscitation applications, providing viscosity and oncotic properties that may be beneficial during acute ischemic events. Oxidized keratin derivatives, also known as keratoses, show good blood and cardiovascular compatibility and thus are the subject of this study.

Methods

The effects of Keratose compounds will be assessed using a topload intravenous infusion model and observation of changes in the microvasculature of the cremaster muscle of rats.

Results

Keratose resuscitation fluid (KRF) administration resulted in significant vasodilation in the cremaster muscle. This effect was blocked with pretreatment of L-NA to inhibit nitric oxide. Another keratin fraction, alpha keratose, which is the primary viscosic compound, was not found to induce vasodilation.

Conclusions

The apparent mechanism of vasodilation was found to be nitric oxide- mediated and isolated to a particular purified fraction, the keratin associated proteins (KAP).

In vivo video microscopy has been used to study blood flow regulation as a function of varying oxygen concentration in microcirculatory networks. However, previous studies have measured the collective response of stimulating large areas of the microvascular network at the tissue surface.

Objective

We aim to limit the area being stimulated by controlling oxygen availability to highly localized regions of the microvascular bed within intact muscle.

Design and Method

Gas of varying O2 levels was delivered to specific locations on the surface of the Extensor Digitorum Longus muscle of rat through a set of micro-outlets (100 μm diameter) patterned in ultrathin glass using state-of-the-art microfabrication techniques. O2 levels were oscillated and digitized video sequences were processed for changes in capillary hemodynamics and erythrocyte O2 saturation.

Results and Conclusions

Oxygen saturations in capillaries positioned directly above the micro-outlets were closely associated with the controlled local O2 oscillations. Radial diffusion from the micro-outlet is limited to ~75 μm from the center as predicted by computational modelling and as measured in vivo. These results delineate a key step in the design of a novel micro-delivery device for controlled oxygen delivery to the microvasculature to understand fundamental mechanisms of microvascular regulation of O2 supply.

Background

Recent studies have hypothesized that endothelial and microvascular dysfunction may play a role in the development of obesity. Previous studies have shown that retinal microvascular changes are associated with diabetes, hypertension, and cardiovascular disease. In contrast, few prospective studies have examined the association between retinal microvascular changes and the risk of developing obesity.

Methods

We examined n=2,089 nonobese subjects from a population-based cohort in Beaver Dam, Wisconsin (aged 44-85 years, 49% women). Retinal arteriolar and venular diameters were measured from baseline retinal photographs. The main outcome-of-interest was 15-year incidence of obesity.

Results

Retinal venular widening was positively associated with incident obesity over a 15-year follow-up period. This association was independent of age, gender, smoking, alcohol intake, education, physical activity, body mass index, serum cholesterol, and C-reactive protein levels. Compared to subjects with retinal venular diameter in the lowest tertile (referent), the multivariable RR (95% CI) of obesity among subjects in the highest tertile was 1.68 (1.24-2.28); p-trend=0.0005. In contrast, narrow retinal arterioles were not associated with obesity.

Conclusions

In a population-based cohort, we found that wider retinal venules are positively associated with risk of developing obesity, suggesting a role for microvascular dysfunction in its etiology.

Conduction of arteriolar vasodilation is initiated by activation of nitric oxide (NO) mechanisms, but dependent on conduction of hyperpolarization. Most studies have used brief (<1 sec) activation of the initial vasodilation to evaluate the fast conduction processes. However, most arteriolar mechanisms involving NO production persist for minutes. In this study, fast and slower components of arteriolar conduction in the in vivo rat brain and small intestine were compared using 3 minute stimulation of NO dependent vasodilation and measurement of [NO] at the distal sites. Within 10-15 seconds, both vasculatures had a rapidly conducted vasodilation and dilation at distance had a fast but small [NO] component. A slower but larger distal vasodilation occurred after 60-90 seconds in the intestine, but not the brain, and was associated with a substantial increase in [NO]. This slowly developed dilation appeared to be caused by flow mediated responses of larger arterioles as smaller arterioles dilated to lower downstream resistance. These results indicate while the intestinal and cerebral arterioles have a fast conducted vasodilation system, the intestinal arterioles also have a slower but larger dilation of major arterioles that is NO related and dependent on the conduction of vasodilation between small arterioles.

Elevated blood pressure during hypertension has been associated with microvascular rarefaction defined by loss of microvessels. However, whether rarefaction is a result of impaired angiogenesis remains unclear. The objective of this study was to compare angiogenesis across the time course of mesenteric microvascular network remodeling in adult spontaneously hypertensive versus normotensive rats. Angiogenic responses in 15–16-week-old SHR and Wistar rats at 0, 3, 5, 10 or 25 days post 20 minute exteriorization of the mesentery were quantified. Consistent with the phenomenon of rarefaction, vascularized area in unstimulated SHR was decreased compared to Wistar. By 25 days, SHR vascular area had increased to the Wistar level and vascular length density and capillary sprouting were comparable. At 3 and 5 days, SHR and Wistar tissues displayed an increase in the capillary sprouting and vascular density relative to their unstimulated controls. At 10 days, capillary sprouting in the SHR remained elevated. The percent change in vascular density was elevated in the SHR compared to the Wistar group at 3 and 5 days and by 25 days the rate of change was more negative. Our results suggest that SHR networks undergo an increased rate of growth followed by an increased rate of pruning.

Objective

The equations for transport of hydrophilic solutes through aqueous pores provide a fundamental basis for examining capillary–tissue exchange and water and solute flux through transmembrane channels, but the theory remains incomplete for ratios, α, of sphere diameters to pore diameters greater than 0.4. Values for permeabilities, P, and reflection coefficients, σ, from Lewellen (18), working with Lightfoot et al. (19), at α = 0.5 and 0.95, were combined with earlier values for α < 0.4, and the physically required values at α = 1.0, to provide accurate expressions over the whole range of 0 < α < 1.

Methods

The “data” were the long-accepted theory for α < 0.2 and the computational results from Lewellen and Lightfoot et al. on hard spheres (of 5 different α’s) moving by convection and diffusion through a tight cylindrical pore, accounting for molecular exclusion, viscous forces, pressure drop, torque and rotation of spheres off the center line (averaging across all accessible radial positions), and the asymptotic values at α = 1.0. Coefficients for frictional hindrance to diffusion, F(α), and drag, G(α), and functions for σ(α) and P(α), were represented by power law functions and the parameters optimized to give best fits to the combined “data.”

Results

The reflection coefficient σ = {1 − [1 − (1 − ϕ)2]G′(α)} + 2α2ϕF′(α), and the relative permeability P/Pmax = ϕF′(α)[1+9α5.5 · (1.0−α5)0.02], where ϕ is the partition coefficient or volume fraction of the pore available to solute. The new expression for the diffusive hindrance is F′(α) = (1−α2)3/2ϕ/[1 + 0.2·α2·(1−α2)16], and for the drag factor is G′(α) = (1−2α2/3 − 0.20217α5)/(1 − 0.75851α5) − 0.0431[1 − (1 − α10)]. All of these converge monotonically to the correct limits at α = 1.

Conclusions

These are the first expressions providing hydrodynamically based estimates of σ(α) and P(α) over 0 < α < 1 They should be accurate to within 1–2%.

Objective

Changes in vasomotion may precede other global indices of autonomic dysfunction that track the onset and progression of diabetes. Recently we showed that baseline spectral properties of vasomotion can discriminate among normoglycemic (N), prediabetic (PreDM), and diabetic (T2DM) nonhuman primates. In this study, our aims were 1. To determine the time-dependence and complexity of the spectral properties of vasomotion in three metabolic groups of monkeys; 2. To examine the effects of heat-provoked vasodilatation on the power spectrum; and 3. To compare the effects of exogenous insulin on the vasomotion.

Materials and Methods

Laser Doppler flow rates were measured from the foot in 9 N, 11 PreDM, and 7 T2DM monkeys. Baseline flow was measured at 34 °C, and under heat stimulation at 44 °C. Euglycemic, hyperinsulinemic clamps were performed to produce acute hyperinsulinemia. The Lempel-Ziv complexity, prediction error, and covariance complexity of 5-dimensional embeddings were calculated as measures of randomness.

Results and Conclusions

With progression of diabetes, measures of randomness of the vasomotion progressively decreased, suggesting a progressive loss of the homeostatic capacity of the peripheral circulation to respond to environmental changes. Power spectral density among T2DM animals resided mostly in the 0 – 1.45 Hz range, which excluded the cardiac component, suggesting that with progression of the disease, regulation of flow shifts toward local rather than central (autonomic) mechanisms. Heating increased all components of the spectral power in all groups. In N, insulin increased the vasomotion contributed by endothelial, neurogenic, vascular myogenic and respiratory processes but diminished that due to heart rate. By contrast, in T2DM, insulin failed to stimulate the vascular myogenic and respiratory activities, but increased the neural/endothelial and heart rate components. Interestingly, acute hyperinsulinemia resulted in no significant vasomotion changes in the chronically hyperinsulinemic PreDM, suggesting yet another form of “insulin resistance” during this stage of the disease.

Objective

To evaluate the age-related changes in pumping of mesenteric lymphatic vessels (MLV) in male 9-mo and 24-mo old Fisher-344 rats.

Methods

Lymphatic diameters, contraction amplitude, contraction frequency and fractional pump flow were determined in isolated MLV before and after L-NAME application.

Results

The data demonstrate a severe weakening of the lymphatic pump in aged MLV including diminished lymphatic contraction amplitude, contraction frequency and as a result, - lymphatic pump activity. The data also suggest that the imposed flow gradient-generated shear-dependent relaxation does not exist in aged rat MLV, and the sensitivity of both adult and aged MLV to such shear cannot be eliminated by nitric oxide synthases blockade.

Conclusions

These data provide new evidence of lymphatic regional heterogeneity for both adult and aged MLV. In MLV, a constant interplay between the tonic and phasic components of the myogenic response and the shear-dependent release of nitric oxide predominantly determines the level of contractile activity; the existence of another shear-dependent but NO-independent regulatory mechanism is likely present. Aging remarkably weakens MLV contractility, which would predispose this lymphatic network to lower total lymph flow in resting conditions and limit the ability it’s to respond to an edemagenic challenge in the elderly.

Objective

Cytomegalovirus (CMV) has been implicated in cardiovascular disease, possibly through the induction of inflammatory processes. P-selectin and L-selectin are adhesion molecules that mediate early microvascular responses to inflammatory stimuli. This study examined the role of these selectins in the microvascular dysfunction that occurs during persistent CMV infection.

Methods

C57Bl/6, P- or L-selectin-deficient mice were mock-inoculated or infected with murine CMV (mCMV), and 5 wk later placed on normal diet (ND) or high cholesterol diet (HC) for 6 wk. P-selectin expression was measured, or intravital microscopy was performed to determine arteriolar vasodilation, and venular blood cell recruitment.

Results

P-selectin expression was significantly increased in the heart, lung and spleen of mCMV-ND, but not mCMV-HC C57Bl/6. mCMV-ND and mCMV-HC exhibited impaired arteriolar function, which was reversed by treatment with an anti-P-selectin antibody, but not L-selectin deficiency. mCMV-HC also showed elevated leukocyte and platelet recruitment. P-selectin inhibition abrogated, whereas L-selectin deficiency partially reduced, these responses.

Conclusions

We provide the first evidence for P-selectin upregulation by persistent mCMV infection, and implicate this adhesion molecule in the associated arteriolar dysfunction. P-selectin, and to a lesser extent L-selectin, mediates the leukocyte and platelet recruitment induced by CMV infection combined with hypercholesterolemia.

Objective

The endothelium-dependent dilation of skeletal muscle arterioles is mediated by factors that have not been identified in young rats, and partly mediated by an unidentified hyperpolarizing factor in maturing rats. This study was designed to determine if endogenous hydrogen peroxide (H2O2) contributes to this arteriolar dilation at either of these growth stages.

Methods

Gracilis muscle arterioles were isolated from rats at ages 24-26 days (“weanlings”) and 46-48 days (“juveniles”). We investigated the effects of catalase treatment on the endothelium-dependent dilation of these vessels to simvastatin and ACh. Catalase-sensitive 2′,7′-dichlorofluorescein (DCF) fluorescence also was measured as an index of H2O2 formation, and arteriolar dilation to exogenous H2O2 was pharmacologically probed in each age group.

Results

Responses to simvastatin and ACh were attenuated by catalase in juvenile, but not weanling, arterioles. Juvenile, but not weanling, arterioles also displayed catalase-sensitive DCF fluorescence that was increased by ACh. Exogenous H2O2 could induce dilation in juvenile, but not weanling, arterioles. In juvenile arterioles, this dilation was abolished by the K+ channel inhibitors TEA and glibenclamide, and attenuated by NOS inhibition or endothelial removal.

Conclusion

These findings suggest that endogenous H2O2 contributes to endothelium-dependent arteriolar dilation in juvenile rats but not in younger rats, and that H2O2 acts in juvenile rats by stimulating endothelial NO release and activating smooth muscle K+ channels.

Endothelial dysfunction can develop at an early age in children with risk factors for cardiovascular disease. A clear understanding of the nature of this dysfunction and how it can worsen over time requires detailed information on the normal growth-related changes in endothelial function on which the pathological changes are superimposed. This review summarizes our current understanding of these normal changes, as derived from studies in four different mammalian species. Although the endothelium plays an important role in controlling vascular tone from birth onward, the vasoactive molecules that mediate this control often change during postnatal or juvenile growth. The specifics of this transition to an adult endothelial cell phenotype can vary depending on the vascular bed. During growth, the contribution of nitric oxide to endothelium-dependent dilation generally increases in the lung, cerebral cortex and skeletal muscle, but decreases in the intestine. Endothelial capacity for release of other vasoactive factors (e.g., cyclooxygenase products, hydrogen peroxide, carbon monoxide) can also increase or decrease during growth. Although these changes have been well documented, there is less information on their underlying cellular or molecular events. Further research is required to clarify these mechanisms, and to evaluate the functional significance of such shifts in endothelial phenotype.

Objective

Impaired endothelium-dependent arteriolar dilation in mice fed high salt is due to local oxidation of nitric oxide (NO) by superoxide anion (O2-). We explored the possibility that “uncoupled” endothelial nitric oxide synthase (eNOS) is the source of this O2-.

Methods

Levels of L-arginine (L-Arg), tetrahydrobiopterin (BH4) and O2- (hydroethidine oxidation) were measured in spinotrapezius muscle arterioles of mice fed normal salt (0.45%, NS) or high salt (4%, HS) diets for 4 weeks, with or without dietary L-Arg supplementation. The contribution of NO to endothelium-dependent dilation was determined from the effect of Nω-nitro-L-arginine methyl ester (L-NAME) on responses to acetylcholine (ACh).

Results

Arterioles in HS mice had lower [BH4] and higher O2- levels than those in NS mice. ACh further increased arteriolar O2- in HS mice only. L-Arg supplementation prevented the reduction in [BH4] in arterioles of HS mice, and O2- was not elevated in these vessels. Compared to NS mice, arteriolar ACh responses were diminished and insensitive to L-NAME in HS mice, but not in HS mice supplemented with L-Arg.

Conclusions

These findings suggest that eNOS uncoupling due to low [BH4] is responsible for O2- generation and reduced NO-dependent dilation in arterioles of mice fed a high salt diet.

Chronic wounds represent a rising health and economic burden to our society. Emerging studies indicate that miRNAs play a key role in regulating several hubs that orchestrate the wound inflammation and angiogenesis processes. Of interest to wound inflammation are the regulatory loops where inflammatory mediators elicited following injury, are regulated by miRNAs as well as regulate miRNA expression. Adequate angiogenesis is a key determinant of success in ischemic wound repair. Hypoxia and cellular redox state are among the key factors that drive wound angiogenesis. We provided first evidence demonstrating that miRNAs regulate cellular redox environment via a NADPH oxidase dependent mechanism in human microvascular endothelial cells (HMECs). We further demonstrated that hypoxia-sensitive miR-200b is involved in induction of angiogenesis by directly targeting Ets-1 in HMECs. These studies points towards a potential role of miRNA in wound angiogenesis. miRNA-based therapeutics represents one of the major commercial hot spots in today’s biotechnology market space. Understanding the significance of miRs in wound inflammation and angiogenesis may help design therapeutic strategies for management of chronic non-healing wounds.

MicroRNAs (miRs) are small non-coding RNAs implicated mainly in post-transcriptional gene silencing by interacting with the unstranslated region of the transcript. miR-210 represents a major hypoxia-inducible miRs, also known as hypoxamirs, which is ubiquitously expressed in a wide range of cells, serving versatile functions. This review article summarizes the current progress on biogenesis of miR-210 and its physiological roles including arrest of cell proliferation, repression of mitochondrial respiration, arrest of DNA repair, vascular biology, and angiogenesis. Given the fact that miR-210 is aberrantly expressed in a number of diseases such as tumor progression, myocardial infarction and cutaneous ischemic wounds, miR-210 could serve as an excellent candidate for prognostic purposes and therapeutic intervention. With the advancement of computational prediction, high-throughput target validation methodology, sequencing, proteomic analysis and microarray, it is anticipated that more down-stream targets of miR-210 and its-associated biological consequences under hypoxia will be unveiled establishing miR-210 as a major hub in the biology of hypoxia-response.

Objective

Neutrophil recruitment to sites of inflammation involves P-selectin dependent rolling. Quantitative dynamic footprinting is a useful tool to visualize the topography of the neutrophil footprint as it interacts with the substrate. However, elucidating the role of specific proteins in addition to topography requires simultaneous visualization of two fluorochromes.

Methods

To validate dual-color quantitative dynamic footprinting, mouse neutrophils were labeled with the membrane dyes DiO and DiI and perfused into microchannels coated with P-selectin-Fc. Footprints of rolling neutrophils were recorded as two separate images, one for each fluorochrome. To assess the localization of the cytoskeletal protein paxillin, we applied dual-color quantitative dynamic footprinting to DiO stained neutrophils of mice expressing an mCherry-paxillin fusion protein.

Results

The footprint topographies obtained from DiO and DiI in the plasma membrane were identical. The z-coordinates of the microvilli tips obtained with the two fluorochromes in the footprint were also identical. Paxillin was found to be localized to some, but not all ridges in the neutrophil footprint.

Conclusions

Our data suggest that the spectral properties of the fluorochrome do not affect the results. Dual-color quantitative dynamic footprinting will be useful for simultaneous visualization of two fluorochromes in the footprint of rolling cells.

OBJECTIVE

We examined the impact of estradiol and progesterone on skin local heating and reactive hyperemia in 25healthy women.

METHODS

Subjects were studied 3 times over 10–12 days. Endogenous sex hormones were suppressed with agonadotropin-releasing hormone antagonist. Subjects were studied on day 4 of suppression (study-day 1),3–4 days later following treatment with either 17β-estradiol or progesterone (study-day 2), and another 3–4 days later, following treatment with both estradiol and progesterone (study-day 3). Subjects underwent identical local heating and reactive hyperemia protocols on all study days. Local heating is characterized by an initial peak in blood flow, followed by a prolonged plateau. A brief nadir is seen between the phases.

RESULTS

Blood flow values are expressed as percent maximum cutaneous vascular conductance (CVC). Estradiol alone increased initial peak CVC from 71±2 to 79±2% (p=0.001). Progesterone alone increased initial peak CVC from 72±2 to 78±2% (p=0.046). Neither estradiol nor progesterone increased plateau CVC. No significant changes were seen between study days 2 and 3 for either group. No differences were observed in reactive hyperemia.

CONCLUSION

Both estradiol and progesterone increased initial peak CVC during local heating, without altering plateau CVC. There was no additive effect of estradiol and progesterone.

Objective

Particle adhesion in vivo is dependent on microcirculation environment which features unique anatomical (bifurcations, tortuosity, cross-sectional changes) and physiological (complex hemodynamics) characteristics. The mechanisms behind these complex phenomena are not well understood. In this study, we used a recently developed in vitro model of microvascular networks, called Synthetic Microvascular Network, for characterizing particle adhesion patterns in the microcirculation.

Methods

Synthetic microvascular networks were fabricated using soft lithography processes followed by particle adhesion studies using avidin and biotin-conjugated microspheres. Particle adhesion patterns were subsequently analyzed using CFD based modeling.

Results

Experimental and modeling studies highlighted the complex and heterogeneous fluid flow patterns encountered by particles in microvascular networks resulting in significantly higher propensity of adhesion (>1.5X) near bifurcations compared to the branches of the microvascular networks.

Conclusion

Bifurcations are the focal points of particle adhesion in microvascular networks. Changing flow patterns and morphology near bifurcations are the primary factors controlling the preferential adhesion of functionalized particles in microvascular networks. Synthetic microvascular networks provide an in vitro framework for understanding particle adhesion.

Objective

To assess the role of adenosine receptors in the regulation of coronary microvascular permeability to porcine serum albumin (PsPSA).

Methods

Solute flux was measured in single perfused arterioles and venules isolated from pig hearts using fluorescent dye-labeled probes by microspectro-fluorometry. Messenger RNA, protein, and cellular distribution of adenosine receptors in arterioles and venules were analyzed by RT-PCR, immunoblot, and immunofluorescence.

Results

Control venule PsPSA (10.7 ± 4.8 × 10−7 cm s−1) was greater than that of arterioles (6.4 ± 2.8 × 10−7 cm · s−1; p <.05). Arteriolar PsPSA decreased ( p <.05) with adenosine suffusion over the range from 10−8 to 10−5 M, while venular PsPSA did not change. The nonselective A1 and A2 receptor antagonist, 8-(p-sulfophenyl) theophylline, blocked the adenosine-induced decrease in arteriolar PsPSA. Messenger RNA for adenosine A1,A2A,A2B, and A3 receptors was expressed in arterioles and venules. Protein for A1, A2A, and A2B, but not A3, was detected in both microvessel types and was further demonstrated on vascular endothelial cells.

Conclusion

Arteriolar PsPSA decreases with adenosine suffusion but not venular PsPSA. Adenosine A1, A2A, and A2B receptors are expressed in both arterioles and venules. Selective receptor-linked cellular signaling mechanisms underlying the regulation of permeability remain to be determined.

Objective

Calcium signaling is integral to endothelium-dependent vasodilation. Our goal was to develop methods enabling automated analyses for accurately and objectively determining the dynamic relationship between endothelial cell (EC) Ca2+ responses and arteriolar diameter in vivo.

Methods

User-friendly software (DiaFluor) written in LabView was applied to images acquired at 15 frames s-1 with a custom macrozoom intravital microscope to evaluate changes in EC Ca2+ concomitant with arteriolar diameter. Transgenic Cx40BAC-GCaMP2 mice expressing a fluorescent Ca2+ indicator molecule in arteriolar ECs enabled resolution of EC Ca2+ signaling in response to acetylcholine (ACh) microiontophoresis (500 nA, 100-1000 ms pulse) from a micropipette (1 μm tip) positioned adjacent to an arteriole in the superfused cremaster muscle preparation.

Results

A 100-ms pulse of ACh (1M) had little effect on EC Ca2+ or arteriolar diameter. As pulse duration increased, vasodilation increased with fluorescence intensity (P<0.01). Based upon fluorescence responses (F/Fo), the effective diffusion distance of ACh along arterioles increased from ~100 μm (250 ms pulse) to ~200 μm (1000 ms pulse) with a peak velocity of ~150 μm s-1.

Conclusions

The novel imaging and software presented here are the first to enable automated simultaneous evaluation of EC Ca2+ signaling and endothelium-dependent vasodilation in vivo.

Object

The obese Zucker rat (OZR) model of the metabolic syndrome is partly characterized by moderate hypercholesterolemia in addition to other contributing co-morbidities. Previous results suggest that vascular dysfunction in OZR is associated with chronic reduction in vascular nitric oxide (NO) bioavailability and chronic inflammation, both frequently associated with hypercholesterolemia. As such, we evaluated the impact of chronic cholesterol reducing therapy on the development of impaired skeletal muscle arteriolar reactivity and microvessel density in OZR and its impact on chronic inflammation and NO bioavailability.

Materials and Methods

Beginning at 7 weeks of age, male OZR were treated with gemfibrozil, probucol, atorvastatin or simvastatin (in chow) for 10 weeks. Subsequently, plasma and vascular samples were collected for biochemical/molecular analyses, while arteriolar reactivity and microvessel network structure were assessed using established methodologies after 3, 6 and 10 weeks of drug therapy

Results

All interventions were equally effective at reducing total cholesterol, although only the statins also blunted the progressive reductions to vascular NO bioavailability, evidenced by greater maintenance of acetylcholine-induced dilator responses, an attenuation of adrenergic constrictor reactivity, and an improvement in agonist-induced NO production. Comparably, while minimal improvements to arteriolar wall mechanics were identified with any of the interventions, chronic statin treatment reduced the rate of microvessel rarefaction in OZR. Associated with these improvements was a striking statin-induced reduction in inflammation in OZR, such that numerous markers of inflammation were correlated with improved microvascular reactivity and density. However, using multivariate discriminant analyses, plasma RANTES, IL-10, MCP-1 and TNF-α were determined to be the strongest contributors to differences between groups, although their relative importance varied with time.

Conclusions

While the positive impact of chronic statin treatment on vascular outcomes in the metabolic syndrome are independent of changes to total cholesterol, and are more strongly associated with improvements to vascular NO bioavailability and attenuated inflammation, these results provide both a spatial and temporal framework for targeted investigation into mechanistic determinants of vasculopathy in the metabolic syndrome.