Absence of glutathione peroxidase-1 exacerbates cerebral ischemia-reperfusion injury by reducing post-ischemic microvascular perfusion.

Mice deficient in the anti-oxidant enzyme glutathione peroxidase-1 (Gpx1) have a greater susceptibility to cerebral injury following a localized ischemic event. Much of the response to ischemia-reperfusion is caused by aberrant responses within the microvasculature, including inflammation, diminished endothelial barrier function (increased vascular permeability), endothelial activation, and reduced microvascular perfusion. However, the role of Gpx1 in regulating these responses has not been investigated. Wild-type and Gpx1-/- mice underwent focal cerebral ischemia via mid-cerebral artery occlusion followed by measurement of cerebral perfusion via laser Doppler and intravital microscopy. Post-ischemic brains in wild-type mice displayed significant deficit in microvascular perfusion. However, in Gpx1-/- mice, the deficit in cerebral blood flow was significantly greater than that in wild-type mice, and this was associated with significant increase in infarct size and increased vascular permeability. Ischemia-reperfusion also resulted in expression of matrix metalloproteinase-9 (MMP-9) in endothelial cells. The absence of Gpx1 was associated with marked increase in pro-MMP-9 expression as well as potentiated MMP-9 activity. Pre-treatment of Gpx1-/- mice with the anti-oxidant ebselen restored microvascular perfusion, limited the induction and activation of MMP-9, and attenuated the increases in infarct size and vascular permeability. These findings demonstrate that the anti-oxidant function of Gpx1 plays a critical role in protecting the cerebral microvasculature against ischemia-reperfusion injury by preserving microvascular perfusion and inhibiting MMP-9 expression.

[Combined treatment of complicated chlamydial infection in males with ozone therapy].

Urogenital chlamydial monoinfection was diagnosed in 127 males using enzyme immunoassay, polymerase chain reaction, transrectal ultrasound examination of the prostatic gland. Of them, 72 patients had chronic urethroprostatitis. Microhemodynamics of these patients was studied with laser doppler flowmetry of the prostate and urethra. The patients received etiotropic therapy with fromilide, regional transurethral and transrectal ozone therapy. The symptoms relieved in 4-6 weeks. Repeated enzyme immunoassay and polymerase chain reaction stated elimination of the infective agent. Improvement of hemodynamics and urethral, prostatic microcirculation was stated after administration of regional ozone therapy.

[Laser Doppler flowmetry in differential diagnosis of organic erectile dysfunction].

The aim of this study was to investigate penile microcirculation in patients with erectile dysfunction. Laser doppler flowmetry was performed in 67 patients (mean age 43.9 +/- 1.53 years) with organic erectile dysfunction and in 20 men without erectile dysfunction. It was found that in patients with arteriogenic erectile dysfunction blood flow parameters were subnormal including flux motions. The occlusive test revealed reduced postocclusive reactive hyperemia in patients with arteriogenic erectile dysfunction. Patients with neurogenic erectile dysfunction have signs of sympathetic denervation of microcirculation and decreased respiratory response.

Changes of cerebral blood flow during the secondary expansion of a cortical contusion assessed by 14C-iodoantipyrine autoradiography in mice using a non-invasive protocol.

Although changes of cerebral blood flow (CBF) in and around traumatic contusions are well documented, the role of CBF for the delayed death of neuronal cells in the traumatic penumbra ultimately resulting in secondary contusion expansion remains unclear. The aim of the current study was therefore to investigate the relationship between changes of CBF and progressive peri-contusional cell death following traumatic brain injury (TBI). CBF and contusion size were measured in C57Bl6 mice under continuous on-line monitoring of (ETp)CO2 before, and at 15 min and 24 h following controlled cortical impact by 14C-iodoantipyrine autoradiography (IAP-AR; n = 5-6 per group) and by Nissl staining, respectively. Contused and ischemic (CBF < 10%) tissue volumes were calculated and compared over time. Cortical CBF in not injured mice varied between 69 and 93 mL/100mg/min depending on the anatomical location. Fifteen minutes after trauma, CBF decreased in the whole brain by approximately 50% (39 +/- 18 mL/100mg/min; p < 0.05), except in contused tissue where it fell by more than 90% (3 +/- 2 mL/100mg/min; p < 0.001). Within 24 h after TBI, CBF recovered to normal values in all brain areas except the contusion where it remained reduced by more than 90% (p < 0.001). Contusion volume expanded from 24.9 to 35.5 mm3 (p < 0.01) from 15 min to 24 h after trauma (+43%), whereas the area of severe ischemia (CBF < 10%) showed only a minimal (+13%) and not significant increase (22.3 to 25.1 mm3). The current data therefore suggest that the delayed secondary expansion of a cortical contusion following traumatic brain injury may not be caused by a reduction of CBF alone.

Realtime photoacoustic microscopy in vivo with a 30-MHz ultrasound array transducer.

We present a novel high-frequency photoacoustic microscopy system capable of imaging the microvasculature of living subjects in realtime to depths of a few mm. The system consists of a high-repetition-rate Q-switched pump laser, a tunable dye laser, a 30-MHz linear ultrasound array transducer, a multichannel high-frequency data acquisition system, and a shared-RAM multi-core-processor computer. Data acquisition, beamforming, scan conversion, and display are implemented in realtime at 50 frames per second. Clearly resolvable images of 6-microm-diameter carbon fibers are experimentally demonstrated at 80 microm separation distances. Realtime imaging performance is demonstrated on phantoms and in vivo with absorbing structures identified to depths of 2.5-3 mm. This work represents the first high-frequency realtime photoacoustic imaging system to our knowledge.

Windkesselness of coronary arteries hampers assessment of human coronary wave speed by single-point technique.

A novel single-point technique to calculate local arterial wave speed (SPc) has recently been presented and applied in healthy human coronary arteries at baseline flow. We investigated its applicability for conditions commonly encountered in the catheterization laboratory. Intracoronary pressure (P(d)) and Doppler velocity (U) were recorded in 29 patients at rest and during adenosine-induced hyperemia in a distal segment of a normal reference vessel and downstream of a single stenosis before and after revascularization. Conduit vessel tone was minimized with nitroglycerin. Microvascular resistance (MR) and SPc were calculated from P(d) and U. In the reference vessel, SPc decreased from 21.5 m/s (SD 8.0) to 10.5 m/s (SD 4.1) after microvascular dilation (P < 0.0001). SPc was substantially higher in the presence of a proximal stenosis and decreased from 34.4 m/s (SD 18.2) at rest to 27.5 m/s (SD 13.4) during hyperemia (P < 0.0001), with a concomitant reduction in P(d) by 20 mmHg and MR by 55.4%. The stent placement further reduced hyperemic MR by 26% and increased P(d) by 26 mmHg but paradoxically decreased SPc to 13.1 m/s (SD 7.7) (P < 0.0001). Changes in SPc correlated strongly with changes in MR (P < 0.001) but were inversely related to changes in P(d) (P < 0.01). In conclusion, the single-point method yielded erroneous predictions of changes in coronary wave speed induced by a proximal stenosis and distal vasodilation and is therefore not appropriate for estimating local wave speed in coronary vessels. Our findings are well described by a lumped reservoir model reflecting the "windkesselness" of the coronary arteries.

Current concepts in assessment of microvascular endothelial function using laser Doppler imaging and iontophoresis.

Effective evaluation of endothelial function is a powerful tool for determining patients at risk of development and progression of cardiovascular disease. As an alternative to invasive tests of endothelial function, several noninvasive methods have been developed, including the use of laser Doppler flowmetry/imaging to measure cutaneous perfusion accompanied by iontophoresis of acetylcholine and sodium nitroprusside. It is clear from previous studies that this technique provides an easy, validated, and reproducible method for investigators to assess and monitor endothelial function in patients with a variety of pathologic conditions, but it may also be used to examine disease progression over time and responsiveness to treatment, thereby facilitating clinical trials. However, a standardization of protocols would help reduce the apparent controversy seen in the literature. With its increasing use by other groups, it is anticipated that further published studies will help to provide a better understanding of the development and progression of cardiovascular disease.

Myocardial perfusion characterization from contrast angiography spectral distribution.

Despite recovering a normal coronary flow after acute myocardial infarction, percutaneous coronary intervention does not guarantee a proper perfusion (irrigation) of the infarcted area. This damage in microcirculation integrity may detrimentally affect the patient survival. Visual assessment of the myocardium opacification in contrast angiography serves to define a subjective score of the microcirculation integrity myocardial blush analysis (MBA). Although MBA correlates with patient prognosis its visual assessment is a very difficult task that requires of a highly expertise training in order to achieve a good intraobserver and interobserver agreement. In this paper, we provide objective descriptors of the myocardium staining pattern by analyzing the spectrum of the image local statistics. The descriptors proposed discriminate among the different phenomena observed in the angiographic sequence and allow defining an objective score of the myocardial perfusion.

Analysis of muscle microcirculation in advanced diabetes mellitus by contrast enhanced ultrasound.

AIMS: Contrast enhanced ultrasound (CEUS) was recently established to quantify perfusion deficits in peripheral arterial disease (PAD). However, this approach was not suitable to assess microangiopathy of skeletal muscle, a major contributor to PAD in diabetic patients. We hypothesized that an optimized methodology would detect impaired microcirculation. METHODS: Ten patients with advanced diabetes mellitus (mean diabetes duration 21 years), 10 PAD patients, and 10 control subjects were enrolled consecutively. The arrival times of the contrast agent Sonovue after intravenous injection were assessed selectively in a small artery, muscle tissue and a muscle vein of the calf muscle. Contrast transit times (CTTs) were calculated as the differences between arrival times. RESULTS: The median CTT for artery-vein was significantly higher in the diabetes group (43 s) than in the PAD (22 s, p=0.007) and control groups (11 s, p<0.001, no value overlap). CTTs for artery-muscle and muscle-vein were shorter with highest median values in the diabetes group. CONCLUSIONS: We validated improved CEUS as consistent method to detect changes in the microvascular bed. This method may become a valuable tool to quantify impaired microcirculation in diabetes and help to improve patient care.

Total blush score: a new index for the assessment of microvascular perfusion in idiopathic dilated cardiomyopathy.

BACKGROUND: The aim of this study was to evaluate tissue-level perfusion in patients with idiopathic dilated cardiomyopathy (IDC), using the myocardial blush grade technique. METHOD: The study population consisted of 26 prospectively enrolled IDC patients (15 women and 11 men; mean age, 59+/-8.8 years) and 26 control subjects (11 women and 15 men; mean age, 54.9+/-10.6 years), whose angiographic films were technically adequate for myocardial blush grade analysis. After grading, we measured total blush score (TBS) for both groups. TBS was determined as the sum of the blush grades of each coronary territory. RESULTS: A total of 156 coronary territories in both groups were assessed. Average of TBS was significantly lower in patients with IDC than in control group (7.6+/-1.2 vs. 8.8+/-0.4; P<0.0001). The TBS significantly and inversely correlated with New York Heart Association class, heart rate, left ventricular end-systolic dimension, and left ventricular end-diastolic pressure, and positively correlated with left ventricular ejection fraction (r=-0.76, P<0.001; r=-0.61, P=0.001; r=-0.77, P<0.0001; r=-0.68, P<0.0001; and r=0.67, P<0.0001, respectively). CONCLUSION: In IDC, decreased TBS might be assumed to be a surrogate marker for a diseased microvascular network in the catheterization laboratory. The relationship between reduced TBS and IDC severity suggests that this index might have prognostic significance.

Microcirculation volumetric flow assessment using high-resolution, contrast-assisted images.

To improve the resolution of contrast-assisted imaging systems, we previously developed a 25-MHz microbubbles-destruction/replenishment imaging system with a spatial resolution of 160 X 160 microm. The goal of the present study was to propose a new approach for functionally evaluating the microvascular volumetric blood flow based on this high-frequency, ultrasound imaging system. The approach includes locating the perfusion area and estimating the blood flow velocity therein. Because the correlation changes between before and after microbubble destruction in two adjacent images, a correlated-based approach was introduced to detect the blood perfusion area. We also have derived a new sigmoid-based model for characterizing the microbubbles replenishment process. Two parameters derived from the sigmoid-based model - the rate constant and inflection time - were adopted to evaluate the blood flow velocity. This model was validated using both simulations and in vitro experiments for mean flow velocities ranging from 1 to 10 mm/s, which showed that the model was in good agreement with simulated and measured microbubble-replenishment time-intensity curves. The results indicate that the actual flow velocity is highly correlated with the estimates of the rate constant and the reciprocal of the inflection time. B-mode imaging experiments for mean flow velocities ranging from 0.4 to 2.1 mm/s were used to assess the volumetric flow in the microcirculation. The results indicated the high correlation between the actual volumetric flow rate and the product of the estimated perfusion area and rate constant, and the reciprocal of the inflection time. We also found that the boundary of the microbubble destruction volume significantly affected estimations of the flow velocity. The perfusion area can be located, and the corresponding flow velocity can be estimated simultaneously in a one-stage, microbubble-destruction/replenishment process, which makes the assessment of the volumetric bloo- d flow in the microcirculation feasible using a real-time, high-frequency ultrasound system.

A novel angiographic methodology for the quantification of angiogenesis.

The objective is to develop a method to quantify the dynamic information of contrast transport using angiography for investigating angiogenic treatments. In the rabbit hindlimb ischemia model, contrast media transport was examined for both arteries and the microvasculature. Time histories of image intensity were constructed and modeled. The differences in contrast transport quantified by the parameters of the mathematical model were statistically compared between animals treated with an adenoviral vector that expressed vascular endothelial growth factor and untreated animals. The data reveal that after one week of ischemia, treated animals have a statistical increase in the number of large vessels that convect blood more efficiently. This analysis further shows a statistically significant increase in the angiographic blush in the treated animals. A methodology is described that offers the capability of examining the number and geometry of large arteries, the dynamics of contrast transport, and the amount of angiographic blush that is related to microvascular density. In therapeutic angiogenesis, numerous techniques are used to measure variables such as the angiographic score, capillary density, and regional blood flow. The analysis presented herein can offer information of these variables, and is transferable from the laboratory to the clinical arena.

Assessment of coronary blood flow and the reactivity of the microcirculation non-invasively with transthoracic echocardiography.

BACKGROUND: The development in ultrasound technology has allowed the use of non-invasive transthoracic echocardiography (TTE) for the study of coronary artery physiology and pathophysiology. TTE can be used to detect atherosclerotic changes in epicardial coronary arteries and to study the effects of specific interventions on coronary microcirculation. AIM: The purpose of this review was to summarize the development of TTE, and outweigh the strenghts and weaknesses of the method for the evaluation of coronary artery blood flow. Moreover, findings from clinical trials studying microcirculatory reactivity using TTE are presented. CONCLUSIONS: TTE is a feasible and reproducible method for the evaluation of coronary artery blood flow. It can also be used in assessing the vasodilation of the epicardial coronary artery simultaneously with flow velocity measurement during the cold pressor test and coronary flow velocity reserve assessment. It is specifically suitable for repeated measurements in interventional trials.

Molecular imaging and ultrasound-assisted drug delivery.

Ultrasound imaging is undergoing a major revolution, about to bring this modality well beyond its established role as a low-cost noninvasive real-time imaging modality. Particularly important has been the commercial availability of microbubble-based contrast agents. Several new indications for contrast ultrasound have been developed and have entered clinical practice. The characterization of focal liver lesions and the follow-up of antiangiogenic therapy are two applications that will have a major impact in medical practice. Others, such as prostate cancer diagnosis, are still under investigation and need to be clinically validated. Thanks to its outstanding sensitivity, contrast-enhanced ultrasound is also geared to become the preferred modality for molecular imaging of diseases occurring at the vascular level. Angiogenesis, inflammation, and other endothelial dysfunctions are important targets that can be imaged and investigated with targeted microbubbles. In the field of prostate cancer, targeted agents will facilitate detection and provide additional information on tumor size, and hopefully on aggressiveness as well. Ultrasound can also play a role to deliver drugs or genes locally. This is an exciting area that has become an important field of research. Imaging and drug delivery can be performed simultaneously, thereby achieving the release of a drug, at the site where it is needed, in a most efficient way.

The extent of microvascular damage during myocardial contrast echocardiography is superior to other known indexes of post-infarct reperfusion in predicting left ventricular remodeling: results of the multicenter AMICI study.

OBJECTIVES: We sought to evaluate the value of the extent of microvascular damage as assessed with myocardial contrast echocardiography (MCE) in the prediction of left ventricular (LV) remodeling after ST-segment elevation myocardial infarction (STEMI) as compared with established clinical and angiographic parameters of reperfusion. BACKGROUND: Early identification of post-percutaneous coronary intervention microvascular dysfunction may help in tailoring appropriate pharmacological interventions in high-risk patients. The ideal method to establish effective microvascular reperfusion after percutaneous coronary intervention remains to be determined. METHODS: A total of 110 patients with first successfully reperfused STEMI were enrolled in the AMICI (Acute Myocardial Infarction Contrast Imaging) multicenter study. After reperfusion, peak creatine kinase, ST-segment reduction, and Thrombolysis In Myocardial Infarction (TIMI) and myocardial blush grade were calculated. We evaluated perfusion defects with MCE by using continuous infusion of Sonovue (Bracco, Milan, Italy) in real-time imaging. The endocardial length of contrast defect (CD) on day 1 after reperfusion was calculated. Wall motion score index, the extent of wall motion abnormalities, LV end-diastolic volume, and ejection fraction after reperfusion and at follow-up also were calculated. RESULTS: Of 110 patients, 25% evolved in LV remodeling and 75% did not. Although peak creatine kinase, ST-segment reduction >70%, and myocardial blush grade were not different between groups, in patients exhibiting LV remodeling, TIMI flow grade 3 was less frequent (p < 0.001), wall motion score index was greater (p < 0.001), and CD was greater (p < 0.001). At multivariate analysis, only TIMI flow grade <3 and CD with a cutoff of >25% were independently associated with LV remodeling. Among patients with TIMI flow grade 3, CD was the only independent variable associated with LV remodeling. CONCLUSIONS: Among patients with TIMI flow grade 3, the extent of microvascular damage, detected and quantitated by MCE, is the most powerful independent predictor of LV remodeling after STEMI as compared with persistent ST-segment elevation and myocardial blush grade.

Contrast harmonic ultrasound and indocyanine-green fluorescence video angiography for evaluation of dermal and subdermal microcirculation in free parascapular flaps.

PURPOSE: Contrast harmonic ultrasound (CHI) with a linear transducer is a new diagnostic approach that allows dynamic and quantitative flow detection of tissue perfusion in microsurgery. The aim of the study was the evaluation of perfusion of the dermal and subdermal layers of microvascular tissue transplants with CHI in comparison to ICG-fluorescence angiography. MATERIAL AND METHOD: In a prospective clinical study Indocyanine-Green Fluorescence Video Angiography and Contrast Enhanced High Resolution Ultrasound (5-10 MHz; linear transducer; Logiq 9; GE) were used for evaluation of the microcirculation in 10 transplanted free parascapular flaps. Two regions were analysed, the centre of the flap and the region of the anastomosis. The perfusion patterns of both methods were compared. RESULTS: The perfusion indexes measured by ICG-fluorescence angiography correlated very precisely in all patients with the quantitative perfusion curves of contrast-enhanced US with CHI. Two flaps with slow filling and low dye intensity showed low contrast enhancement in CHI with modified perfusion curves with slow increase. In two cases a reduced perfusion and filling were found. There were no statistical differences between the two diagnostic methods (p>0.01). CONCLUSION: CHI improves US detections of dermal and subdermal microcirculation in comparison to ICG fluorescence angiography. CHI is a new diagnostic method for postoperative monitoring of free flaps.

Clinical importance of microvascular obstruction on contrast-enhanced MRI in reperfused acute myocardial infarction.

BACKGROUND: The aim of the present study was to investigate the clinical importance of microvascular obstruction (MO) on contrast-enhanced magnetic resonance imaging (MRI), comparing it with the myocardial perfusion index (MPI) assessed using first-pass MRI. METHODS AND RESULTS: Cardiac MRI was performed in 33 patients within 7 days after reperfusion of a myocardial infarction (MI). Using a bolus injection of Gd-DPTA, first-pass images were obtained with the Turbo-FLASH sequence. Time-intensity curves in the left ventricular cavity and in myocardial sections were generated and then the MPI was assessed by the maximum slope method. Late enhancement (LE) was assessed using the true-FISP sequence. According to the transmurality of LE, the patients were classified into 3 groups: Group 1 included patients with localized endocardial enhancement; Group 2, patients with transmural enhancement; Group 3, patients having LE with MO. In cases of anterior infarction, the MPI for the anterior wall and parts of the inferior wall in Group 3 was significantly lower than that for Group 1 and 2. For inferior infarction, the MPI for parts of the inferior wall in Group 3 was significantly lower than that for Group 1 and 2. CONCLUSION: MO is related to lower MPI, indicating severe microvascular damage. LE with or without MO is an important marker of perfusion status after reperfused MI.

Effect of systemic blood pressure on microcollateral circulation evaluated by real-time contrast echocardiography.

BACKGROUND: In acute myocardial infarction, residual collateral-derived myocardial blood flow (CBF) within the ischemic area is one of the major determinants of infarct size. Management of systemic blood pressure (sBP) related to maintain collateral circulation is still difficult. The aim of this study was to reveal the influence of sBP on the rescue of area at risk by collateral circulation. METHODS: Real-time myocardial contrast echocardiography just after the onset of complete occlusion of the left circumflex coronary artery was performed in collateral-rich open-chest dogs. The video intensity of the ischemic area was evaluated during the occlusion and the CBF (A x beta) was calculated from a replenishment curve: y = A (1 - e(-beta t)). To analyze the effect of sBP on the collateral circulation, sBP was altered by infusion of nitroglycerin or etilefrine hydrochloride. To evaluate the defect size (%DS), every end-systolic myocardial contrast echocardiography image after left circumflex coronary artery occlusion was converted into binary images using custom offline software. RESULTS: The %DS increased and CBF slightly decreased at low sBP. The %DS decreased and CBF increased at high sBP. At excessively high sBP, %DS increased and CBF decreased again. CONCLUSION: Real-time myocardial contrast echocardiography, which is a useful noninvasive method to evaluate the collateral perfusion quantitatively, has a crucial role in the decision of patient treatment and management strategy of acute myocardial infarction.