International journal of microcirculation, clinical and experimental最新文献

The goal of this study was to determine whether exogenous application of L-arginine could restore impaired agonist-induced increases in arteriolar diameter during diabetes mellitus. We used intravital microscopy to examine reactivity of cheek pouch arterioles (50 microns in diameter) in nondiabetic and diabetic (2 weeks after injection of streptozotocin) hamsters in response to histamine and substance P. In nondiabetic hamsters histamine (1.0 and 5.0 microM) dilated cheek pouch arterioles by 15 +/- 1 and 22 +/- 1%, respectively, and substance P (50 and 100 nM) dilated arterioles by 14 +/- 3 and 21 +/- 4%, respectively. In addition, dilatation of arterioles in response to histamine and substance P in nondiabetic hamsters was abolished by application of an enzymatic inhibitor of nitric oxide synthase (L-NMMA). In contrast, histamine- and substance P-induced increases in arteriolar diameter were markedly reduced in diabetic hamsters. Histamine (1.0 and 5.0 microM) dilated arterioles by only 5 +/- 1 and 4 +/- 2%, respectively, and substance P (50 and 100 nM) dilated arterioles by only 6 +/- 2 and 5 +/- 3%, respectively (p < 0.05 vs. nondiabetic hamsters). Nitroglycerin produced similar vasodilatation in nondiabetic and diabetic hamsters. Next, we examined whether exogenous application of L-arginine (100 microM) could restore impaired histamine- and substance P-induced increases in arteriolar diameter in diabetic hamsters. We found that L-arginine did not restore altered nitric oxide synthase-dependent vasodilatation in diabetic hamsters. These findings suggest that short-term diabetes mellitus alters agonist-induced increases in arteriolar diameter. In addition, the mechanism of altered arteriolar reactivity during diabetes mellitus does not appear to be related to an impaired availability of L-arginine.

Effects of low- and high-intensity endurance training on the capillary luminal diameter and number were studied morphometrically in the rat plantaris muscle. Male Wistar-Imamichi rats were divided into three groups: sedentary control group (Cont, n = 9), low-intensity (running speed of 20 m/min) training group (T-20, n = 8) and high-intensity (running speed of 40 m/min) training group (T-40, n = 7). Rats in both training groups were subjected to each treadmill running program for 60 min/day, 5 days/week for 9 weeks. After 9 weeks of training, citrate synthase activity significantly increased in T-40 compared with Cont, but did not change in T-20. All morphometric parameters with respect to capillary and muscle fiber area were determined in the perfusion-fixed plantaris muscle. The mean muscle fiber areas in both T-20 and T-40 were similar to that in Cont. The capillary-to-fiber ratios were significantly higher in T-20 (2.28 +/- 0.06) and T-40 (2.29 +/- 0.06) than in Cont (2.00 +/- 0.07). The number of capillaries with a small luminal diameter (2-4 microns) was significantly higher in T-20 than in Cont. In contrast, T-40 had a significantly higher number of capillaries with a large luminal diameter (8-10 microns) compared with Cont. This study indicates that endurance training induces changes in the capillary luminal diameter as well as capillary number, and that the adaptive response of the capillary luminal diameter to endurance training depends on the training intensity.

We investigated in vitro whether endothelial cell edema is induced by cellular hypoxia or oxygen radical formation. Measurements of relative cell volume (RCV) were made using microweight analysis, liquid scintillation spectrometry and analysis of cellular protein content. To validate this method of determining cell volume, endothelial cells were incubated in media of different osmolarities. Vascular endothelial cells reacted to osmotic stress with a volume increase or decrease. The addition of xanthine oxidase (XOD; 3 mU/ml) and hypoxanthine (1 mM) for the enzymatic production of O2- caused a reproducible and significant increase in RCV by 29 +/- 8% (from 5.5 to 7.1 microliters/10(6) cells; p < 0.001) after an incubation time of 60 min. Nonenzymatically produced H2O2 (100 microM) caused a similar increase in RCV by 35 +/- 5% (from 5.5 to 7.6 microliters/10(6) cells; p < 0.001) over the same incubation period. The addition of catalase (50 U/ml) diminished the increasing effect of XOD as well as that of H2O2 on cell volume. As assessed by the uptake of the vital dye trypan blue and the release of lactate dehydrogenase into the medium, there was no significant loss of viability during the incubation time. Lower concentrations of H2O2 as well as lower activities of XOD did not induce a significant increase in RCV. Higher H2O2 concentrations and increased XOD activities caused a considerable time- and concentration-dependent injury of endothelial cells. RCV was unchanged even after long exposure (5 h) to two different hypoxic gas mixtures (3% O2:5% CO2:92% N2; 0% O2:5% CO2:95% N2). Cell viability was not impaired under hypoxic conditions. The results suggest that reactive oxygen species play a more important role in the development of endothelial cell edema than cellular hypoxia.

In order to compare the extent of the elicited vasoconstrictive and vasodilative response at the microcirculatory level in essential hypertension (EH), we measured the skin blood flow by means of a laser Doppler flowmeter (LDF). Thirty-four mild-to-moderate EH patients were enrolled. Twenty-two sex- and age-matched healthy subjects were selected as a control group. The LDF measurements were carried out with the probe over the fingertip of the distal phalanx at baseline conditions (Rest flow, RF), after an ischaemic test (post-ischaemic peak flow) and during an arithmetic stress test (AT). The flow was expressed in arbitrary units. The data were processed using the Perisoft computer program. The relative flows after the ischaemic test (Rel F1) and during the AT (Rel F2) were expressed as a percentage of the previous RF values (RF1 and RF2, respectively). During the AT, the lag time was calculated (in seconds). As compared to the control subjects, RF was significantly lower in the EH group (p < 0.01). During the AT, the EH patients showed a statistically lower mean Rel F2 decrease compared to the control subjects (p < 0.01). No statistically significant difference occurred in the Rel F1 and lag time. These data suggest that the vasoconstrictive capacity of the precapillary vessels is impaired in patients with hypertension.

When skin is exposed to cold, cutaneous blood flow is initially restricted due to sympathetic vasoconstriction. Prolonged exposure to cold has a secondary protective vasodilator effect. In the present study, the effects of severe and prolonged local cold on the cutaneous microcirculation were assessed. In 10 young healthy subjects, laser Doppler skin flux and flux motion were measured at the calf during 20 min of local ice cooling and 15 min subsequent recovery. In the 6th minute of cooling, mean skin flux decreased to 58 +/- 6% of the resting value (p < 0.05), then increased and reached 129 +/- 10% of the resting value at the end of the cooling period, followed by a phase of reactive hyperemia with a maximum of 225 +/- 24% (p < 0.05). Mean flux motion frequency, spontaneously present at rest with a frequency of 2.6 +/- 0.2 cycles/ min decreased rapidly during the first minutes of cold exposure, was absent during the 6th to the 10th minute, reappeared and started to increase from the 11th minute and reached 4.4 +/- 0.3 cycles/min during the recovery period (p < 0.05). This study seems to indicate that the phenomenon of protective increase of cutaneous microcirculatory blood flow consists of an initial phase of vasodilation, followed by a phase of active and enhanced microvessel vasomotor activity.

The chorioallantoic membrane (CAM) of the chick embryo represents an in vivo model to evaluate microvascular function during the sequential phases of endothelial proliferation (angiogenesis), cytodifferentiation, and senescence. The principal focus of this study was to characterize microvascular barrier functions of the CAM endothelium during its nonproliferating, aging phase (senescence). A graded series of fluorescein isothiocyanate (FITC) dextrans served to index macromolecular selectivity of the senescent capillary endothelium. Extravasation of FITC dextrans 40 and 150 was restricted, while FITC dextran 10 progressively accumulated within the intercapillary interstitia during 15-min perfusion periods. Endothelial vesicle densities were greater in the first- and second-order pre- and first- order postcapillary endothelia (28 +/- 8/micron2) than those recorded for the capillary endothelium (8 +/- 3/micron2). Junctional cleft lengths (luminal to abluminal distances), on the other hand, were greater in the capillary endothelium (1.08 +/- 0.50 microns) than those recorded for the second-order pre- and postcapillaries (0.46 +/- 0.03 micron). Junctional cleft widths were segmentally uniform (20 nm) in the microvascular units. That permselectivity of the segmental microvascular endothelia was homogeneous is ultrastructurally consistent with the uniform junctional cleft widths rather than the heterogeneous cleft lengths and vesicle densities. The CAM serves as the principal respiratory exchange surface of the embryo. Thus, maintenance of colloid oncotic balance across the microvascular endothelium is likely critical to gaseous exchange. The retention of a selective barrier during the senescent phase of the CAM endothelium serves to support this concept.

The effect of intravenously administered Ginkbo biloba extract (EGb 761) on the vasospastic response to platelet activation has been assessed using a cutaneous flap preparation in anaesthetized mice. Arterioles of the axillary artery were observed by intravital microscopy, and platelets were activated by topical application of ADP under two steady state conditions: normothermia (37 degrees C) and hypothermia (24 degrees C). Responses of the cutaneous arterioles to stimulation by topical application of a thromboxane agonist (U46619) were also compared in animals treated intravenously with EGb 761 or with a thromboxane synthesis inhibitor (U63557). ADP induced a 34% constriction of the arterioles in control animals. However, no arteriolar constriction occurred in response to ADP in platelet-depleted animals (collagen-induced thrombocytopenia) or in animals treated with EGb 761 (60 mg/kg, i.v.). Exposure of the arterioles to hypothermia (24 degrees C) for 10 min induced constriction of 7-12% in all experimental groups of animals. Under these hypothermic conditions, either EGb 761 or thrombocytopenia abolished ADP-induced arteriolar constriction which was substituted by arteriolar dilation, indicating that EGb 761 can inhibit the vasospasm that is produced by platelet activation. As topically applied U46619 (10(-5) M) induced arterioles constriction (about 22%) that was abolished by intravenous treatment with EGb 761, the extract appears to act directly rather than as a thromboxane synthase inhibitor. Collectively, these findings indicate that platelet factors can play a significant role in cutaneous vasospasm, and that EGb 761, via an action on the thromboxane pathway, could be useful in treating Raynaud's phenomenon and other vascular disorders which involve increased thromboxane production.

Arteriovenous anastomoses (AVA) in skin microcirculation are mediated by the sympathetic stimuli. The inspiratory gasp test (IG test) triggers the sympathetic nervous system, resulting in a decrease in AVA skin blood flow, as measured by laser Doppler fluxmetry (LDF). We studied the reproducibility of the IG test under carefully standardized respiratory conditions. In each of 19 healthy (young) volunteers with a mean skin temperature during the experiment above 28 degrees C 13 IG tests were performed, either under spirometric control or uncontrolled and by using a negative pressure transducer. Starting the IG test end-inspiratory results in the most pronounced absolute LDF decrease [140 PU (70-490)], median (minimum-maximum) as compared to starting end-expiratory [100 PU (40-260)] and during inspiration [110 PU (50-350)], respectively, p < 0.001 and p < 0.001. Inspiration as fast as possible results in a larger absolute LDF decrease [150 PU (40-450)], compared to inspiration in 5 s [120 PU (60-340); p < 0.02] and in 10 s [130 PU (40-350); p < 0.05]. Continuously sucking negative mouth pressure results in a larger LDF decrease [140 PU (30-420)] in comparison with taking one deep breath and holding it for 10 s [110 PU (30-270); p < 0.01]. However, standardization of the IG test did not improve its reproducibility.

We have recently shown that the responsiveness of rat intestinal arterioles to increased sympathetic nerve activity is modulated by the actions of endothelial-derived nitric oxide. Because the microvascular endothelium can also produce vasodilator prostaglandins, the purpose of this study was to determine if endogenous cyclooxygenase products also limit sympathetic arteriolar constriction in this vascular bed. Intravital microscopy was used to study the responses of small feed arteries, first-order arterioles and second-order arterioles to perivascular sympathetic nerve stimulation in the superfused rat small intestine. Stimulation at 3, 8 and 16 Hz caused frequency-dependent constrictions of each vessel type that are abolished by the alpha-adrenoceptor antagonist phentolamine (10(-6) M superfusate concentration). The cyclooxygenase inhibitor meclofenamate (3 x 10(-5) M superfusate concentration) completely abolished the dilator responses to topically applied arachidonic acid, but had no effect on the magnitude or rate of sympathetic constriction in any vessel type. These results suggest that endogenous cyclooxygenase activity does not influence sympathetic tone in the intestinal microvasculature.