Inflammation in rats born to mothers treated with dexamethasone 15cH during pregnancy: an immunohistochemical study

In previous studies, we observed that rats born to mothers treated with dexamethasone 15CH (10-33M) had a higher level of mast cell degranulation and greater arteriolar dilation after the exposure of an inflammatory stimulus, suggesting the possibility of vertical transmission of the effects of ultra-diluted substances between mother and offspring. In this study, a more detailed assessment of the cellular events in acute inflammation was made using techniques of immunohistochemistry. The identification of adhesion molecules expression was made by the markers: anti-CD54 (ICAM-1) and anti-CD18 (β2-Integrin). The identification of inflammatory cells was performed by the markers anti-MAC387 (mononuclear cells) and anti-CD163 (active macrophages). Polymorphonuclear cells were identified by hematoxylin-eosin staining. The number of labeled cells per field was recorded, except for the anti-CD54 marker, whose intensity of staining on the endothelial cells was defined by scores assigned by two independent observers. The results point toward to an up regulation of the whole inflammatory process in rats born to mothers treated with dexamethasone 15CH during pregnancy. This conclusion is justified by the following statistically significant (p≤0.05) findings: a) bigger mast cell degranulation and increased of arteriolar diameter; b) increased migration of polymorphonuclear cells in relation to the mononuclear cells; c) earlier expression of CD163 in monocytes, d) higher level of adhesion molecules expression.

Intravital Assessment of Pre-capillary Pulmonary Arterioles of Type-1 Diabetic Mice Shows Oxidative Damage and Increased Tone in Response to NOS Inhibition

Microvascular dilation, important for peripheral tissue glucose distribution, also modulates alveolar perfusion and is inhibited by loss of bioavailable nitric oxide (NO) in diabetes mellitus (DM). We hypothesized that DM-induced oxidative stress decreases bioavailable NO and pulmonary pre-capillary arteriolar diameter, causing endothelial injury. We examined sub-pleural pulmonary arterioles after acute NO synthase (NOS) inhibition with L-NAME in streptozotocin (STZ) and saline (CTRL)-treated C57BL/6J mice. Microvascular changes were assessed by intravital microscopy in the right lung of anesthetized mice with open-chest and ventilated lungs. Arteriolar tone in pulmonary arterioles (27.2 to 48.7 µm diameter), increased in CTRL mice (18.0 ± 11% constriction p=0.034, n=5) but decreased in STZ (13.6 ± 7.5% dilation p= 0.009, n=5), after L-NAME. Lung tissue DHE fluorescence (superoxide), inducible NOS expression, and protein nitrosylation (3-nitrotyrosine) increased in STZ mice and correlated with increased glucose levels (103.8 ± 8.8 mg/dL). Fluorescently-labeled fibrinogen administration and fibrinogen immunostaining showed fibrinogen adhesion, indicating endothelial injury in STZ mice. In CTRL mice, vasoconstriction to L-NAME was likely due to the loss of bioavailable NO. Vasodilation in STZ mice may be due to decreased formation of a vasoconstrictor or emergence of a vasodilator. These findings provide novel evidence that DM targets the pulmonary microcirculation and that decreased NO bioavailability and increased precapillary arteriolar tone could potentially lead to ventilation-perfusion abnormalities, exacerbating systemic DM complications.

Air pollution and retinal vessel diameter and blood pressure in school-aged children in a region impacted by residential biomass burning

Little is known about the early-life cardiovascular health impacts of fine particulate air pollution (PM2.5) and oxidant gases. A repeated-measures panel study was used to evaluate associations between outdoor PM2.5 and the combined oxidant capacity of O3 and NO2 (using a redox-weighted average, Ox) and retinal vessel diameter and blood pressure in children living in a region impacted by residential biomass burning. A median of 6 retinal vessel and blood pressure measurements were collected from 64 children (ages 4–12 years), for a total of 344 retinal measurements and 432 blood pressure measurements. Linear mixed-effect models were used to estimate associations between PM2.5 or Ox (same-day, 3-day, 7-day, and 21-day means) and retinal vessel diameter and blood pressure. Interactions between PM2.5 and Ox were also examined. Ox was inversely associated with retinal arteriolar diameter; the strongest association was observed for 7-day mean exposures, where each 10 ppb increase in Ox was associated with a 2.63 μm (95% CI − 4.63, − 0.63) decrease in arteriolar diameter. Moreover, Ox modified associations between PM2.5 and arteriolar diameter, with weak inverse associations observed between PM2.5 and arteriolar diameter only at higher concentrations of Ox. Our results suggest that outdoor air pollution impacts the retinal microvasculature of children and interactions between PM2.5 and Ox may play an important role in determining the magnitude and direction of these associations.

Optical coherence tomography of arteriolar diameter and capillary perfusion during spreading depolarizations

Spreading depolarization (SD) is associated with profound oligemia and reduced oxygen availability in the mouse cortex during the depolarization phase. Coincident pial arteriolar constriction has been implicated as the primary mechanism for the oligemia. However, where in the vascular bed the hemodynamic response starts has been unclear. To resolve the origin of the hemodynamic response, we used optical coherence tomography (OCT) to simultaneously monitor changes in the vascular tree from capillary bed to pial arteries in mice during two consecutive SDs 15 minutes apart. We found that capillary flow dropped several seconds before pial arteriolar constriction. Moreover, penetrating arterioles constricted before pial arteries suggesting upstream propagation of constriction. Smaller caliber distal pial arteries constricted stronger than larger caliber proximal arterioles, suggesting that the farther the constriction propagates, the weaker it gets. Altogether, our data indicate that the hemodynamic response to cortical SD originates in the capillary bed.

Relation between retinal vessel diameter and posterior segment optical coherence tomography variables in middle-aged Caucasians: the Northern Finland Birth Cohort Eye Study

AimsStudying the relationship between retinal vessel diameter (RVD) with (1) macular thickness and volume, (2) retinal nerve fibre layer (RNFL), (3) ganglion cell-inner plexiform layer (GC-IPL) and (4) optic nerve head (ONH) in a population cohort of middle-aged Caucasians.MethodsWe collected data from 3070 individuals. We used a semiautomated computer-assisted programme to measure central retinal arteriolar equivalent and central retinal venular equivalent. Macular and ONH parameters were assessed by optical coherence tomography.ResultsData from 2155 persons were analysed. A larger RVD was associated with a thicker macula and increased macular volume; each SD increase in average macular thickness and volume was associated with a 3.28 µm and a 3.19 µm increase in arteriolar diameter and a 5.10 µm and a 5.08 µm increase in venular diameter, respectively (p<0.001 for all). A larger rim area, greater GC-IPL and RNFL thicknesses were associated with larger RVD; each SD increase in rim area, GC-IPL thickness and RNFL thickness was associated with a 1.21 µm, 2.68 µm and a 3.29 µm increase in arteriolar diameter and a 2.13 µm, 4.02 µm and 5.04 µm increase in venular diameter, respectively (p<0.001 for all).ConclusionsIncreased macular thickness, macular volume, GC-IPL thickness, RNFL thickness and optic nerve rim area were associated with larger RVDs in all subjects. This study clarified the anatomical correlations between both macular and ONH parameters with RVD for middle-aged Caucasians; these can represent a basis for further studies investigating the vascular aetiology of eye diseases.

Acid-Sensing Ion Channels

Rationale: Precise regulation of cerebral blood flow is critical for normal brain function. Insufficient cerebral blood flow contributes to brain dysfunction and neurodegeneration. Carbon dioxide (CO 2 ), via effects on local acidosis, is one of the most potent regulators of cerebral blood flow. Although a role for nitric oxide in intermediate signaling has been implicated, mechanisms that initiate CO 2 -induced vasodilation remain unclear. Objective: Acid-sensing ion channel-1A (ASIC1A) is a proton-gated cation channel that is activated by extracellular acidosis. Based on work that implicated ASIC1A in the amygdala and bed nucleus of the stria terminalis in CO 2 -evoked and acid-evoked behaviors, we hypothesized that ASIC1A might also mediate microvascular responses to CO 2 . Methods and Results: To test this hypothesis, we genetically and pharmacologically manipulated ASIC1A and assessed effects on CO 2 -induced dilation of cerebral arterioles in vivo. Effects of inhalation of 5% or 10% CO 2 on arteriolar diameter were greatly attenuated in mice with global deficiency in ASIC1A ( Asic1a −/− ) or by local treatment with the ASIC inhibitor, psalmotoxin. Vasodilator effects of acetylcholine, which acts via endothelial nitric oxide synthase were unaffected, suggesting a nonvascular source of nitric oxide may be key for CO 2 responses. Thus, we tested whether neurons may be the cell type through which ASIC1A influences microvessels. Using mice in which Asic1a was specifically disrupted in neurons, we found effects of CO 2 on arteriolar diameter were also attenuated. Conclusions: Together, these data are consistent with a model wherein activation of ASIC1A, particularly in neurons, is critical for CO 2 -induced nitric oxide production and vasodilation. With these findings, ASIC1A emerges as major regulator of microvascular tone.

Propentdyopents as Heme Degradation Intermediates Constrict Mouse Cerebral Arterioles and Are Present in the Cerebrospinal Fluid of Patients With Subarachnoid Hemorrhage

Rationale: Delayed ischemic neurological deficit is the most common cause of neurological impairment and unfavorable prognosis in patients with subarachnoid hemorrhage (SAH). Despite the existence of neuroimaging modalities that depict the onset of the accompanying cerebral vasospasm, preventive and therapeutic options are limited and fail to improve outcome owing to an insufficient pathomechanistic understanding of the delayed perfusion deficit. Previous studies have suggested that BOXes (bilirubin oxidation end products), originating from released heme surrounding ruptured blood vessels, are involved in arterial vasoconstriction. Recently, isolated intermediates of oxidative bilirubin degradation, known as PDPs (propentdyopents), have been considered as potential additional effectors in the development of arterial vasoconstriction. Objective: To investigate whether PDPs and BOXes are present in hemorrhagic cerebrospinal fluid and involved in the vasoconstriction of cerebral arterioles. Methods and Results: Via liquid chromatography/mass spectrometry, we measured increased PDP and BOX concentrations in cerebrospinal fluid of SAH patients compared with control subjects. Using differential interference contrast microscopy, we analyzed the vasoactivity of PDP isomers in vitro by monitoring the arteriolar diameter in mouse acute brain slices. We found an arteriolar constriction on application of PDPs in the concentration range that occurs in the cerebrospinal fluid of patients with SAH. By imaging arteriolar diameter changes using 2-photon microscopy in vivo, we demonstrated a short-onset vasoconstriction after intrathecal injection of either PDPs or BOXes. Using magnetic resonance imaging, we observed a long-term PDP-induced delay in cerebral perfusion. For all conditions, the arteriolar narrowing was dependent on functional big conductance potassium channels and was absent in big conductance potassium channels knockout mice. Conclusions: For the first time, we have quantified significantly higher concentrations of PDP and BOX isomers in the cerebrospinal fluid of patients with SAH compared to controls. The vasoconstrictive effect caused by PDPs in vitro and in vivo suggests a hitherto unrecognized pathway contributing to the pathogenesis of delayed ischemic deficit in patients with SAH.

Myoglobin facilitates angiotensin II-induced constriction of renal afferent arterioles

Vasoconstriction plays an important role in the development of acute kidney injury in rhabdomyolysis. We hypothesized that myoglobin enhances the angiotensin II (ANG II) response in afferent arterioles by increasing superoxide and reducing nitric oxide (NO) bioavailability. Afferent arterioles of C57Bl6 mice were isolated perfused, and vasoreactivity was analyzed using video microscopy. NO bioavailability, superoxide concentration in the vessel wall, and changes in cytosolic calcium were measured using fluorescence techniques. Myoglobin treatment (10−5 M) did not change the basal arteriolar diameter during a 20-min period compared with control conditions. NG-nitro-l-arginine methyl ester (l-NAME, 10−4 M) and l-NAME + myoglobin reduced diameters to 94.7 and 97.9% of the initial diameter, respectively. Myoglobin or l-NAME enhanced the ANG II-induced constriction of arterioles compared with control (36.6 and 34.2%, respectively, vs. 65.9%). Norepinephrine responses were not influenced by myoglobin. Combined application of myoglobin and l-NAME further facilitated the ANG II response (7.0%). Myoglobin or l-NAME decreased the NO-related fluorescence in arterioles similarly. Myoglobin enhanced the superoxide-related fluorescence, and tempol prevented this enhancement. Tempol also partly prevented the myoglobin effect on the ANG II response. Myoglobin increased the fura 2 fluorescence ratio (cytosolic calcium) during ANG II application (10−12 to 10−6 M). The results suggest that the enhanced afferent arteriolar reactivity to ANG II is mainly due to a myoglobin-induced increase in superoxide and associated reduction in the NO bioavailability. Signaling pathways for the augmented ANG II response include enhanced cytosolic calcium transients. In conclusion, myoglobin may contribute to the afferent arteriolar vasoconstriction in this rhabdomyolysis model.