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PubMed1998

A novel approach for comparative study of periosteum, muscle, subcutis, and skin microcirculation by intravital fluorescence microscopy.

Herein, we report a new model, which allows comparative study of the microcirculation of different peripheral tissues, i.e., periosteum, skeletal muscle, subcutis, and skin. Using dextran-insensitive Wistar rats gracilis and semitendinosus muscles of the left hindlimb were prepared in association with their appertaining tibial fragments, subcutis, and skin. Blood supply was guaranteed by the femoral artery via the saphenous vessels. High-resolution intravital epi-illumination microscopy of the two muscles displayed the typical microvascular architecture with the capillaries running in parallel to each other (capillary density (CD) 128.4 +/- 4.5 cm-1). In subcutis and skin, capillaries were found arranged as interconnecting mesh-like networks with a density, which was significantly higher (P < 0.05) in subcutis (191.0 +/- 5.5 cm-1) compared with skin (108.9 +/- 3.3 cm-1). Analysis of periosteal tissue revealed two distinct types of arrangements of microvascular architecture. Adjacent to the major feeding and draining vessels of the periosteum, capillaries were organized in densely meshed shunt-like networks, revealing the highest capillary density (242.7 +/- 13.2 cm-1; P < 0.05) of all tissues studied. Periosteal capillaries distant from the major feeding and draining vessels were arranged in parallel to the longitudinal axis of the tibial bone and presented with a density similar to that of the skeletal muscle (128. 6 +/- 9.4 cm-1). Topical application of acetylcholine for analysis of physiological reactivity of the microvasculature showed dose-dependent arteriolar dilation. Moreover, a 3-min upstream femoral artery occlusion demonstrated an appropriate hyperemic response in all tissues studied, indicating intact myogenic control. A prolonged period of ischemia (120 min) followed by reperfusion (60 min) caused massive (P < 0.05) leukocyte-endothelial cell interaction in postcapillary venules, similarly as reported in other microvascular tissue preparations. We propose that the model presented provides a good approach to all peripheral tissues for both the analysis of the physiology of tissue-confined microvascular control and the development of novel therapeutic strategies to counteract manifestation of nutritional dysfunction and inflammatory response in disease.

Acetylcholine↗

PubMed1998

In vivo neutralization of P-selectin inhibits leukocyte-endothelial interactions in retinal microcirculation during ocular inflammation.

P-selectin is one of the adhesion molecules involved in leukocyte rolling during an inflammatory reaction. The aim of this study was to examine the role of P-selectin in leukocyte-endothelial interactions in retinal microcirculation during ocular inflammation, known as endotoxin-induced uveitis (EIU), in vivo. EIU was induced in Lewis rats by footpad injection of lipopolysaccharide (LPS). At the time of LPS treatment or 12 h later, anti-rat P-selectin mAb (ARP) was injected intravenously, and its effect on leukocyte behavior in the retina was studied after intravital staining with acridine orange using a scanning laser ophthalmoscope. P-selectin gene expression in the retina was also studied by a semiquantitative polymerase chain reaction (PCR) method. Administration of ARP at the time of LPS treatment significantly reduced the number of rolling leukocytes at 6 and 12 h by 68% (P < 0.05) and 83% (P < 0.01), respectively, and the number of cells infiltrating the vitreous at 48 h by 61% (P < 0.05). Interestingly, ARP significantly inhibited the vasodilation observed during EIU. In contrast, delayed administration of ARP blocked neither cellular infiltration nor vasodilation. P-selectin gene expression was upregulated during the course of EIU. In conclusion, P-selectin may significantly contribute to the development of inflammation in the early stage of endotoxin-induced ocular inflammation.

Animals↗

PubMed1998

Vasomotion and blood flow regulation in hamster skeletal muscle microcirculation: A theoretical and experimental study.

A mathematical model of a microvasculature was used to study the effects of myogenic and flow-dependent stimuli on the characteristics of vasomotion and microvascular perfusion regulation. The model includes three branching orders of arterioles derived from in vivo observations and incorporates a mechanism for terminal arteriolar closure during vasomotion. Simulations were performed to evaluate the effect of vasodilation and vasoconstriction on vasomotion pattern, and the changes in arteriolar effective diameter and flow in response to arterial blood pressure variations triggering the regulatory mechanisms. Vasomotion patterns were studied in the hamster cutaneous muscle, visualized by fluorescent microscopy, in control conditions and after injection of acetylcholine (Ach) or NG-monomethyl-L-arginine (L-NMMA). We have found that vasomotion may be caused by different combinations of feedback mechanisms, including a strong rate-dependent myogenic response or a strong flow-dependent mechanism with no rate-dependent response. Decreasing the rate-dependent component of the myogenic mechanism and increasing the time constant of the flow-dependent mechanism causes vessel stabilization and disappearance of vasomotion. In hamster microcirculation, Ach decreased vasomotion frequency and increased vasomotion amplitude and arteriolar effective diameter, whereas L-NMMA caused a slight increase in vasomotion frequency and decrease in effective diameter. Model simulations, under dilatory and constrictory stimuli, confirmed these results. Moreover, the model predicted that mean blood flow is maintained closer to normal despite arterial pressure changes (+/-15% flow changes versus +/-50% pressure variations) when the vessels were in nonoscillatory than when they are in oscillatory state. In conclusion, a large variety of vasomotion patterns affect arteriolar resistance and microvessel perfusion in skeletal muscle. Furthermore, in the presence of vasomotion the network exhibits a poorer aptitude for regulating blood flow during arterial pressure changes (i.e., worse autoregulation) than the nonoscillatory network.

Animals↗

PubMed2000

Effects of hydrogen peroxide scavenger Catalase on villous microcirculation in the rat small intestine in a model of inflammatory bowel disease.

This study was conducted to quantify the effect of systemic Catalase, a hydrogen peroxide scavenger, on villous microcirculation in the inflamed small intestine of the rat. Intestinal inflammation was induced with s.c. application of Indomethacin. Intravital fluorescence microscopy and FITC-labeled erythrocytes were used to quantify erythrocyte velocity and arteriolar diameter in the main arteriole of the villi in the terminal ileum following i.v. application of Catalase in the inflamed intestine, and the blood flow was calculated. Control groups were formed for Ringer's lactate, Catalase and Indomethacin, respectively. We found that villous blood flow was significantly increased in the in the inflamed intestine. Application of Catalase led to a significant decrease in villous perfusion, but had no effect in the control group. The increase in villous blood flow was accompanied by changes in the diameter of the main arteriole. This effect on arteriolar diameter was reversed by i.v. Catalase. Our results provide evidence that systemic application of Indomethacin leads to vasodilatation of the main arteriole of the villus in the rat ileum and hyperemia in the mucosa. Hyperemia and the vascular diameter of the main arteriole were significantly reduced by H(2)O(2)-scavenger Catalase, suggesting that endogenous H(2)O(2) may be one of the mediators of hyperemia in the mucosa in this animal model of intestinal inflammation.

Animals↗

PubMed2000

A novel model for the in vivo monitoring of uterine microcirculation and intracellular free calcium changes in rat.

The aim of this work was to develop a model to study the microcirculation and relative levels of intracellular free calcium in the myometrium of pregnant rats. On Day 21 of gestation a lobe of uterus was prepared free, flipped over, and mounted in a superfusion chamber leaving the radix and thereby the innervation and circulation intact. RBC velocity and arteriolar diameters were determined by means of intravital video microscopy before and after stimulation (norepinephrine). To study intracellular free calcium changes, the fluorescent dye Indo-1 AM was added to the superfusate in the chamber. Fluorescence images were recorded and ratios of the images collected at 400 and 506 nm were calculated and changes thereof were assumed to represent intracellular free calcium changes. RBC velocity and arteriolar diameter did not change for at least 1 h, while the response to norepinephrine was similar at the beginning of the experiment and after 120 min. In four separate interventions, the uterus was challenged with 5 x 10(-4) IU/ml oxytocin, 4.5 mM calcium, 5 x 10(-4) IU/ml oxytocin with 4.5 mM calcium, and 5 microM ionomycin, resulting in an increase of the 400/506 nm ratio of 27, 31, 76, and 103%, respectively, representing a relative increase in intracellular free calcium. This novel in vivo model is suitable for monitoring intracellular free calcium changes and to record RBC velocities and blood vessel diameters in the myometrium of pregnant rats.

Animals↗

PubMed2000

Stimulation of NO production and of eNOS phosphorylation in the microcirculation in vivo.

The role of nitric oxide (NO) in microvascular permeability is controversial, in part because the regulation of its endothelial constitutive synthase, eNOS, has been studied in vitro but not in vivo. Our study was designed to detect the morphologic and functional presence of eNOS and to test whether eNOS could be phosphorylated by platelet-activating factor (PAF), an agent that induces hyperpermeability. Immunocytochemistry was applied using human anti-eNOS antibodies in the hamster cheek pouch (hcp). The hcp microvessels demonstrated positive reaction products in the endothelium. The functional presence of eNOS in hcp was investigated by topical application of 10(-7) M PAF to the hcp and by measuring NO production by chemiluminescence. The mean baseline value of NO release was 63.3 +/- 6.9 pmol/ml (mean +/- SE). Application of PAF led to an increase in mean NO release to 120.8 +/- 31.2 pmol/ml (P < 0.05). In another series of experiments, 10(-7) M PAF was applied topically to hcp preincubated with [(32)P]orthophosphoric acid. Immunoprecipitation and Western blots detected (32)P-labeled bands that migrated with the mobility of positive eNOS indicating phosphorylated eNOS protein. The intensity of the radioactive bands was evaluated by computer-assisted image analysis. Comparison of the net band intensities yielded a mean PAF-treated/control ratio of 1.6 +/- 0.1. Our data demonstrate the morphologic and functional presence of eNOS in the microcirculation. The data also provide evidence that the function of microvascular eNOS is subject to regulation by phosphorylation.

Animals↗

PubMed2000

Influence of a new monomeric nonionic radiographic contrast medium (iobitridol-350 versus NaCl) on cutaneous microcirculation: single-center, prospective, randomized, double-blind phase IV study in parallel group design.

This single-center, prospective, randomized, double-blind phase IV study with parallel-group design was performed to test whether injection of a newly designed non-ionic monomeric radiographic contrast medium in the axillary artery influences the ipsilateral cutaneous microcirculation. Twenty patients with coronary heart disease or suspected coronary heart disease were investigated. Ten patients received iobitridol and 10 normal saline solution. The confirmatory target variable was the capillary erythrocyte velocity (v(RBC)). The v(RBC) was continuously recorded before and up to 3 min after the injection of iobitridol or normal saline solution and subsequently evaluated off-line. Thirty seconds after injection of 20 ml iobitridol, v(RBC) was reduced by 32% (P < 0.05). This was the strongest reduction, but v(RBC) was reduced at all time points measured up to 180 s following the injection (P < 0.05 up to 180 s). The overall reduction in v(RBC) was also statistically significant (P 0.0446). In contrast, there was a tendency toward an increase in v(RBC) following injection of 20 ml normal saline (P 0.0512). There was also a tendency toward a difference in the time course of changes in v(RBC) following injection of iobitridol compared to normal saline (P 0.0607). The decrease in v(RBC) following iobitridol injection (standardized difference (SD) according to Cohen, 0.67) can be regarded as biometrically weak, and the increase following normal saline injection (SD, 0.24) as very weak. The microcirculatory disturbance induced by iobitridol injection is less severe than that due to comparable high-viscosity contrast media.

Aged↗

PubMed2001

Impact of inhibition of complement by sCR1 on hepatic microcirculation after warm ischemia.

Recent observations provide evidence that complement is implicated as an important factor in the pathophysiology of ischemia/reperfusion injury (IRI). Here, we assessed the effects of complement inhibition on hepatic microcirculation by in vivo microscopy (IVM) using a rat model of warm hepatic ischemia clamping the left pedicle for 70 min. Ten animals received the physiological complement regulator soluble complement receptor type 1 (sCR1) intravenously 1 min prior to reperfusion. Controls were given an equal amount of Ringer's solution (n = 10). Microvascular perfusion and leukocyte adhesion were studied 30 to 100 min after reperfusion by IVM. Microvascular perfusion in hepatic sinusoids was significantly improved in the sCR1 group (80.6 +/- 0.6% of all observed sinusoids were perfused [sCR1] vs 67.3 +/- 1.2% [controls]). The number of adherent leukocytes was reduced in sinusoids (49.9 +/- 3.4 [sCR1] vs 312.3 +/- 14.2 in controls [adherent leukocytes per square millimeter of liver surface]; P < 0.001) as well as in postsinusoidal venules after sCR1 treatment (230.9 +/- 21.7 [sCR1] vs 1906.5 +/- 93.5 [controls] [adherent leukocytes per square millimeter of endothelial surface]; P < 0.001). Reflecting reduced hepatocyte injury, liver transaminases were decreased significantly upon sCR1 treatment compared to controls. Our results provide further evidence that complement plays a decisive role in warm hepatic IRI. Therefore, we conclude that complement inhibition by sCR1 is effective as a therapeutical approach to reduce microcirculatory disorders after reperfusion following warm organ ischemia.

Animals↗

PubMed2002

Systemic effects of low-intensity laser irradiation on skin microcirculation in patients with diabetic microangiopathy.

Low-intensity laser irradiation has been shown to induce wound healing in conditions of reduced microcirculation, which is in part explained by systemic effects. We therefore investigated such a potential systemic effect of low-intensity laser irradiation on skin blood circulation in patients with diabetic microangiopathy. Patients with diabetic microangiopathy were randomized to receive either a single helium-neon (HeNe, 632.8 nm) low-intensity laser irradiation with a dose of 30 J/cm(2) or a sham irradiation over the forefoot region in a double-blind, placebo-controlled clinical study. Skin blood circulation by means of temperature recordings over forefoot regions was detected by infrared thermography. Following a single transcutaneous low-intensity laser irradiation, a rise in skin temperature in both feet of the subjects in the laser group was noted, whereas in both feet of the subjects in the placebo group a drop in skin temperature occurred. The baseline-adjusted skin temperature 15 min after the end of the irradiation was significantly higher in the laser-treated forefeet compared to the placebo-"treated" forefeet (p < 0.0001); the baseline-adjusted difference in the temperature was 1.94 +/- 0.35 degrees C. Simultaneously, the baseline-adjusted skin temperature was significantly higher in the laser-untreated forefeet compared to the placebo-"untreated" forefeet (P < 0.0001); the baseline-adjusted difference was 1.70 +/- 0.33 degrees C. Our data show a significant increase in skin circulation due to athermic laser irradiation in patients with diabetic microangiopathy and point to the possibility of inducing systemic effects.

Adult↗

PubMed1998

An evaluation of linear model analysis techniques for processing images of microcirculation activity.

Sequences of images of the cortical surface can be processed to reveal information about the cortical microcirculation, regional cerebral blood flow (rCBF), and changes induced by neuronal activity. This study examined the use of different analysis methodologies on intrinsic optical images taken from rat sensory motor cortex and testes. Generalized linear model (GLM) analysis was used and compared with standard signal processing methods including principal component analysis. The GLM method has been used by Friston et al. (1994, Hum. Brain Map., 1: 214-220) in the analysis of functional magnetic resonance imagery to identify regions of focal activity. We investigated the use of this method to analyze video image data of the modulation of rCBF from rat cortex. The results revealed spatiotemporal variations in rCBF in response to stimulation within local regions of cortex. The advantage of the GLM method is that it augments ordinary signal processing methods with an estimate of statistical reliability. The use of different wavelengths of illumination reveals spatial structures with different temporal relationships. In image time series data collected under green and red illumination a phase difference was found in the low frequency approximately 0.1 Hz vasomotion oscillation. This phase difference occurred in data from both cortex and testes. A possible explanation of these differences is that the spectral absorption characteristics of the tissue reflect changes in the volume proportions of the different hemoglobin derivatives in interacting with the modulation of the volume of blood. It is suggested that the combination of these effects produces the phase differences we detect.

Animals↗

PubMed1995

Myocardial microcirculation as evaluated with CT.

This study explores use of whole-body CT imaging to separately quantitate the physiological behavior of the nonrecruitable (e.g., arterioles) and recruitable (mostly capillary) components of the intramyocardial microcirculation. In two groups of dogs, one with microembolization and one with epicardial artery stenosis, we demonstrate that intramyocardial blood volume (rho) and blood flow (F) follow the relationship rho = AF + BF1/2 where A represents the transit time of the recruitable and BF-1/2 is the transit time of the nonrecruitable components. These transit times are shown to change in different, but characteristic, patterns.

Animals↗

PubMed1997

Behavior of stimulated leukocytes in the pulmonary microcirculation of perfused rat lungs.

To investigate the dynamics of activated leukocytes and the roles of CD18-ICAM-1 pathway, we examined the effects of rat IL-8 and monoclonal antibodies (mAbs) against CD18 and ICAM-1 on the behavior of leukocytes in microvessels of perfused rat lungs. Specific pathogen free male Sprague-Dawley rats were used. Perfused rat lungs were prepared so as to obtain stable physiological shear rates. We used a confocal laser scanning microscope equipped with a high speed video analysis system to visualize pulmonary microcirculation. Rat leukocytes were activated with rat IL-8. No rolling leukocytes were observed in either pulmonary arterioles or venules, and leukocytes were sequestered in capillaries. The majority of unstimulated capillary leukocytes moved smoothly. About 50% of stimulated leukocytes, however, showed a transient cessation of movement in pulmonary capillaries. Rat IL-8 decreased the relative leukocyte velocities against mean blood velocities in capillaries (45%) and venules (65%), and increased intracapillary neutrophils. Anti-CD18 and anti-ICAM-1 mAbs attenuated these changes. These results suggest that unique features exist in the interaction between activated leukocytes and pulmonary microvessels, and that CD18-ICAM-1-dependent capillary sequestration is one of the major mechanisms by which activated leukocytes accumulate in lungs.

Analysis of Variance↗

PubMed1986

Influence of PaO2 on cerebral macro- and microcirculation as observed by light reflection: time course of changes.

A silicon intensified target camera was used to study cerebral cortical vessels of the cat through a skull window implant, and red cell content changes were measured by light reflectance. Red cell content changes were observed in cerebral arterioles, capillaries, and venules when the PaO2 was decreased by lowering the PiO2. The time course of change in the diameter of the arterioles and venules was measured by selecting a cross section of the vessel plus some surrounding tissue. From the averaged cross-sectional reflectance signal, the change in vessel diameter was followed as a function of time following the PiO2 change. All vessels of greater than 10 microns were observed in focus. Substantial areas where no vessels could be discriminated would contain only capillaries, and changes in light reflectance from such areas would indicate changes in capillary red cell content. The time course of these changes following a step decrease in PiO2 was recorded. Results show that the sequence of red cell content increase in cerebral microcirculation during hypoxia is capillary before venule and arteriole. The times of initial red cell content increase are 37.9 +/- 7 s, 59.7 +/- 7.9 s, and 60.8 +/- 9.1 s, respectively. These results suggest an increase in the capillary bed red cell content as the initial response to hypoxia, but venules and arterioles change only on longer exposure to hypoxia. The sequence of the increase in red cell content suggests the capillaries rather than the arterioles are the vessels which respond to the oxygen autoregulation signal.

Animals↗

PubMed2000

Influence of the bradykinin B1/B2-receptor-antagonist B 9430 on the cerebral microcirculation and outcome of gerbils from global cerebral ischemia.

The influence of the bradykinin B1/B2 antagonist B 9430 on the cerebral microcirculation following global cerebral ischemia was investigated in a closed cranial window preparation in Mongolian gerbils by intravital fluorescence microscopy. Global cerebral ischemia (GCI) was induced by occlusion of both common carotid arteries for 15 min. Leukocyte-endothelium interactions, vessel diameters, and the segmental microvascular blood flow were observed by intravital microscopy before and up to three hours after global cerebral ischemia. Following the early reperfusion period the animals survived up to 4 days after ischemia. The neurological deficit and the body weight were assessed daily. On day 4 animals were subjected to perfusion fixation and the brain was removed. Nerve cell damage from ischemia was quantified histologically in cortex, hippocampus, and striatum. Animals with treatment received the bradykinin B1/B2 receptor antagonist B 9430 before (i.v.), during, and after ischemia (s.c.) until the end of the experiment. The frequency of leukocytes (cells/100 microns x min) rolling along the venular endothelium post ischemia was significantly decreased (p < 0.05) in treated animals as compared to untreated controls (33.0 +/- 6.2 vs. 8.5 +/- 2.3) as well as the number of leukocytes attached to the endothelial surface (7.2 +/- 3.0 vs. 2.0 +/- 1.0, n.s.). The neuroscore on day 4 (pre-ischemic control: 22 points) was reduced to 13.4 +/- 3.2 in untreated animals, while to 4.7 +/- 3.2 points in the treatment group. No differences between animals with and without treatment were found as to the number of viable neurons. Although bradykinin is released in the brain during global cerebral ischemia, its antagonisation does not improve outcome despite the effective inhibition of leukocyte-endothelium interactions.

Animals↗

PubMed2002

Can hyperventilation improve cerebral microcirculation in patients with high ICP?

Gosling's pulsatility index (PI) is generally thought to reflect cerebrovascular resistance. Hyperventilation and increased intracranial pressure (ICP) usually increase PI. In this study, the effect of hyperventilation on PI was assessed in head injured patients with and without elevated ICP. A total of 73 hyperventilation studies were conducted on 20 intubated TBI patients. EtCO2, ICP, MCA flow velocity and MCA PI bilaterally were recorded simultaneously. Hemispheric CO2-Reactivity (hCO2R) was calculated. Only studies with a baseline ICP < 15 or > 30 mm Hg were included for analysis. With hyperventilation pCO2 decreased on average from 33.54.2 to 26.84.8 mmHg. PI increased significantly in patients with baseline ICP < 15 mmHg (11.8%; p < 0.0001) but decreased significantly if baseline ICP was > 30 mm Hg (12.21%; p = 0.0013). High baseline ICP, low initial GCS and impaired hCO2R were associated with the decrease of PI. Hyperventilation unexpectedly reduced PI in patients with high ICP. Because decreased PI suggests decreased CVR, it is postulated that hyperventilation in the setting of raised ICP improves cerebral microcirculation.

Adult↗

PubMed1990

Effect of stimulation of leukocyte chemotaxis by fMLP on white blood cell behaviour in the microcirculation of rat brain.

The role of white blood cells in acute cerebral disorders such as ischemia or stroke is still unclear. Therefore, in the present study we investigated the effects of the leukotaxin n-formyl-methionyl-leucyl-phenylalanine (fMLP) on white blood cell endothelial-cell interactions in the rat brain surface microcirculation. An improved closed cranial window technique was applied. Superfusion of fMLP in rising concentrations (10(-8) - 10(-5) M) was seen to induce rolling and adherence of leukocytes to teh endothelium of small venules. Rolling was more effectively stimulated than firm attachment. fMLP-induced vasodilation was more pronounced in arterioles than in venules. In this study it has been shown that the hydrophilic fMLP is effectively stimulating neutrophil chemotaxis across the blood-brain barrier. Further, the closed cranial window preparation is useful to analyze quantitatively properties of activated leukocytes, which may be pertinent in injury to the blood-brain barrier and induction of microcirculatory disturbances.

Animals↗

PubMed1984

Functional characterization of the microcirculation in tumors.

This review describes some aspects of tumor vessels and the influence of vasoactive agents on tumor blood flow, particularly the characteristic microcirculation of tumors with regard to its selective increase in blood flow. Elevation of blood pressure by infusion of angiotensin II produced a severalfold increase in tumor blood flow. The increase was selective and specific to the tumor vessels as long as the mean arterial blood pressure was kept under 150 mm Hg. Pressure elevation by angiotensin II also selectively increased tumor oxygen tension and influx of lymph flow from the primary transplanted lesion to the lymph node metastatic lesion. Newly devised techniques for analyzing microhemodynamics of tumor vessels showed that the velocity of tumor blood flow, the vascular area in tumor tissue, and the hydrostatic pressure difference between the tumor vessel and extravascular tissue were markedly enhanced. Thus, the extravasation of material into tumor tissues can be increased by the enhancement of blood flow. This demonstration allowed the development of a new approach to cancer chemotherapy, in which the delivery to tumor tissue of systemically administered anticancer drugs can be selectively enhanced.

Angiotensin II↗

PubMed1993

Coronary microcirculation in hypertensive heart disease: functional significance and therapeutic implications.

In arterial hypertension left ventricular hypertrophy comprises myocyte hypertrophy, interstitial fibrosis and structural alterations of the coronary microcirculation. This leads to an impairment of diastolic function of the left ventricle and coronary flow reserve despite normal epicardial arteries. Consequently, antihypertensive treatment should aim at [13] reversing myocyte hypertrophy, [14] restoring myocardial structure and [8] improving coronary flow reserve along with blood pressure normalization.

Antihypertensive Agents↗