Laser Doppler Perfusion Measurements Research Articles

Myocardial and Skeletal Muscle Microvascular Dysfunction Manifests Prior to Diabetic Cardiomyopathy: Sex-Dependent Differences

Introduction: Microvascular dysfunction is a recognized sign of disease in heart failure progression. Blood vessels exhibit abnormal vasoreactivity in early stage, subsequently deteriorating to rarefaction and reduced perfusion. In managing heart failure patients, where diagnosis is typically possible only when the hypertrophic heart is irreversibly damaged, earlier diagnosis is key to improving management. In this study, we apply a blood-pool MRI method for assessing vasomodulation to investigate if it can sensitively detect abnormal leg muscle vasoreactivity posited to manifest before myocardial mvD. Methods: Male and female Sprague-Dawley rats were maintained on a high-fat, high-sugar diet or a control diet for 6 months after the induction of diabetes. Beginning at month 1 or 2 post-induction and every 2 months thereafter, rats underwent blood-pool MRI to assess vasoreactivity in the heart or skeletal muscle, respectively. Ablavar, a T1-reducing blood-pool contrast agent, was administered and the T1 relaxation time dynamically measured as animals breathed in mild CO2 levels to modulate vessels and elicit a vasodilatory response. CO2 levels were set at 5% for skeletal muscle and 10% for cardiac muscle, and the gas was administered for 10 minutes. At the final timepoint, invasive laser Doppler perfusion measurements in leg muscle were recorded to verify MRI results. Results/Discussion: In this study, we provide the first demonstration that both skeletal muscle and myocardial microvascular vasoreactivity is altered in a non-obese rodent model of type II diabetes, prior to the development of heart failure symptoms. In male rats, the normally unresponsive heart to 10% CO2 reveals a pro-vasoconstriction response beginning at 5 months post-diabetes. Abnormal leg skeletal muscle vasoreactivity appeared even earlier, at 2 months: the usual vasodilatory response to 5% CO2 is interrupted with periods of vasoconstriction in diseased rats. In female rats, differences were observed between healthy and diseased animals only within the first two months post-diabetes and not later. In the heart, vasodilation to 10% CO2 seen in healthy animals was abolished in diabetes. In skeletal muscle, 5% CO2 was suboptimal in inducing reproducible vasoreactivity, but young diabetic females responded by vasodilation only. Conclusion: Abnormal vasoreactivity presents earlier than overt structural and functional cardiac changes in both sexes in HFpEF and can be detected using blood-pool MRI in leg skeletal muscle before the myocardium. Our non-invasive MRI technology sets the foundation for a paradigm shift in diagnosing mvD during the early stages of HFpEF, opening the door for early intervention. This work was supported by the Canadian Institutes of Health Research, Natural Sciences and Engineering Research Council of Canada, Canada Foundation for Innovation/Ontario Research Fund, Dean’s Spark Professorship, Medicine by Design Pivotal Experiment Fund [to H.L.M.C.]; Ted Rogers Centre for Heart Research PhD Education Fund, Scintica Instrumentation Inc [to S.L.]. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Blood-pool MRI assessment of myocardial microvascular reactivity.

The ability to non-invasively image myocardial microvascular dilation and constriction is essential to assessing intact function and dysfunction. Yet, conventional measurements based on blood oxygenation are not specific to changes in blood volume. The purpose of this study was to extend to the heart a blood-pool MRI approach for assessing vasomodulation in the presence of blood gas changes and investigate if sex-related differences exist. Animals [five male and five female healthy Sprague Dawley rats (200-500 g)] were intubated, ventilated, and cycled through room air (normoxia) and hypercapnia (10% CO2) in 10-minute cycles after i.v. injection of blood-pool agent Ablavar (0.3 mmol/kg). Pre-contrast T1 maps and T1-weighted 3D CINE were acquired on a 3 Tesla preclinical MRI scanner, followed by repeated 3D CINE every 5 min until the end of the gas regime. Invasive laser Doppler flowmetry of myocardial perfusion was performed to corroborate MRI results. Myocardial microvascular dilation to hypercapnia and constriction to normoxia were readily visualized on T1 maps. Over 10 min of hypercapnia, female myocardial T1 reduced by 20% (vasodilation), while no significant change was observed in the male myocardium. After return to normoxia, myocardial T1 increased (vasoconstriction) in both sexes (18% in females and 16% in males). Laser Doppler perfusion measurements confirmed vasomodulatory responses observed on MRI. Blood-pool MRI is sensitive and specific to vasomodulation in the myocardial microcirculation. Sex-related differences exist in the healthy myocardium in response to mild hypercapnic stimuli.

Skin trauma rapidly induces thermoregulatory plexus hyperemia, while an increased nutritive papillary capillary function can be detected after 24h.

Clinical assessments and laser Doppler perfusion measurements (LDPM) of skin microcirculation have limited value, as they fail to capture events regulated by local metabolic needs at a papillary capillary level. This study aimed to examine the ability of computer-assisted video microscopy (CAVM) and diffuse reflectance spectroscopy (DRS) to assess skin nutritive perfusion-compared to LDPM. Healthy volunteers (n=10) were examined after (≈1 and ≈24h) an incision (5×1mm) on the forearm, at 0.1mm (only with CAVM), 2-3mm, and 30mm from the trauma. No changes were detected by CAVM after ≈1h. After ≈24h, 0-1mm from the trauma, both CAVM parameters were increased: functional capillary density (capillary crossings/mm, 11.8±1.4 vs. 7.3±1.2, p<.01) and capillary flow velocities (CFV, %capillaries with brisk flow, 10±6.8 vs. 1±1, p<.01). At a distance of 2-3mm, only CFV was increased (6.2±6.1 vs. 1±1, p<.05). DRS and LDPM measurements increased 2-3mm from the trauma line in relation to baseline after both ≈1 and ≈24h, that is, with DRS (%microvascular oxygen saturation): 45.8±7.4% (baseline), 70.0±12.5% (≈1h), and 73.1±10.4% (≈24h), p<.01 and with LDPM (a.u.): 7.2±2.5 (baseline), 28.3±18.7 (≈1h), and 45.9±16.3 (≈24h), p<.01. ≈24h after skin trauma, an increased function of the nutritive papillary capillaries can be detected by CAVM.

A novel MRI analysis for assessment of microvascular vasomodulation in low-perfusion skeletal muscle

Compromised microvascular reactivity underlies many conditions and injuries, but its assessment remains difficult, particularly in low perfusion tissues. In this paper, we develop a new mathematical model for the assessment of vasomodulation in low perfusion settings. A first-order model was developed to approximate changes in T1 relaxation times as a result of vasomodulation. Healthy adult rats (N = 6) were imaged on a 3-Tesla clinical MRI scanner, and vasoactive response was probed on gadofosveset using hypercapnic gases at 20% and 5% CO2 to induce vasoconstriction and vasodilation, respectively. MRI included dynamic 3D T1 mapping and T1-weighted images during gas challenge; heart rate was continuously monitored. Laser Doppler perfusion measurements were performed to corroborate MRI findings. The model was able to identify hypercapnia-mediated vasoconstriction and vasodilation through the partial derivative frac{partial {T}_{1}}{partial t}. MRI on animals revealed gradual vasoconstriction in the skeletal muscle bed in response to 20% CO2 followed by gradual vasodilation on transitioning to 5% CO2. These trends were confirmed on laser Doppler perfusion measurements. Our new mathematical model has the potential for detecting microvascular dysfunction that manifests in the early stages across multiple metabolic and ischemic pathologies.

A non-invasive magnetic resonance imaging approach for assessment of real-time microcirculation dynamics

We present a novel, non-invasive magnetic resonance imaging (MRI) technique to assess real-time dynamic vasomodulation of the microvascular bed. Unlike existing perfusion imaging techniques, our method is sensitive only to blood volume and not flow velocity. Using graded gas challenges and a long-life, blood-pool T1-reducing agent gadofosveset, we can sensitively assess microvascular volume response in the liver, kidney cortex, and paraspinal muscle to vasoactive stimuli (i.e. hypercapnia, hypoxia, and hypercapnic hypoxia). Healthy adult rats were imaged on a 3 Tesla scanner and cycled through 10-minute gas intervals to elicit vasoconstriction followed by vasodilatation. Quantitative T1 relaxation time mapping was performed dynamically; heart rate and blood oxygen saturation were continuously monitored. Laser Doppler perfusion measurements confirmed MRI findings: dynamic changes in T1 corresponded with perfusion changes to graded gas challenges. Our new technique uncovered differential microvascular response to gas stimuli in different organs: for example, mild hypercapnia vasodilates the kidney cortex but constricts muscle vasculature. Finally, we present a gas challenge protocol that produces a consistent vasoactive response and can be used to assess vasomodulatory capacity. Our imaging approach to monitor real-time vasomodulation may be extended to other imaging modalities and is valuable for investigating diseases where microvascular health is compromised.

Arginase inhibition attenuates arteriogenesis and interferes with M2 macrophage accumulation

l-Arginine is the common substrate for nitric oxide synthases (NOS) and arginase. Whereas the contribution of NOS to collateral artery growth (arteriogenesis) has been demonstrated, the functional role of arginase remains to be elucidated and was topic of the present study. Arteriogenesis was induced in mice by ligation of the femoral artery. Laser Doppler perfusion measurements demonstrated a significant reduction in arteriogenesis in mice treated with the arginase inhibitor nor-NOHA (Nω-hydroxy-nor-arginine). Accompanying in vitro results on murine primary arterial endothelial cells and smooth muscle cells revealed that nor-NOHA treatment interfered with cell proliferation and resulted in increased nitrate/nitrite levels, indicative for increased NO production. Immuno-histological analyses on tissue samples demonstrated that nor-NOHA administration caused a significant reduction in M2 macrophage accumulation around growing collateral arteries. Gene expression studies on isolated growing collaterals evidenced that nor-NOHA treatment abolished the differential expression of Icam1 (intercellular adhesion molecule 1). From our data we conclude that arginase activity is essential for arteriogenesis by promoting perivascular M2 macrophage accumulation as well as arterial cell proliferation.

Cooling Effect on Skin Microcirculation in Asphyxiated Newborn Infants with Increased C-Reactive Protein

Background: Therapeutic hypothermia is presumed to suppress inflammatory processes after perinatal asphyxia. In a previous study of neonatal hypoxic-ischemic encephalopathy (HIE) we found altered skin microcirculation in about a third of the infants after rewarming. We speculated whether this could be linked to increased inflammatory responses, such as high C-reactive protein (CRP). The present study further explored this question. Objective: The aim of this study was to explore the differences in skin microcirculation and its oxygen delivery ability during cooling and after rewarming in HIE infants with or without high CRP. Methods: A previously studied population of 28 HIE infants was divided into two subgroups depending on low or high CRP (repeated values above 30 mg/l for more than 24 h). The differences between the two groups regarding laser Doppler perfusion measurements (LDPMs), computer-assisted video microscopy and diffuse reflectance spectroscopies during cooling on days 1 and 3 and after rewarming on day 4 were assessed. Results: After rewarming, infants with high CRP showed significantly higher skin LDPM perfusion, lower functional vessel density and larger heterogeneity of capillary flow velocities as compared to infants with low CRP, while no such differences were found during cooling. Conclusion: Skin microcirculatory responses differed significantly after rewarming, but not during cooling, between asphyxiated neonates with or without high CRP. We speculate whether cooling influences the inflammatory skin microcirculatory response and the ability of oxygen delivery to the cells. Further studies are needed to investigate this as well as its applicability to other vascular beds in the body.

Noninvasive assessments of oxygen delivery from the microcirculation to skin in hypothermia-treated asphyxiated newborn infants.

Therapeutic hypothermia (TH) has become standard treatment for severe and moderate hypoxic-ischemic neonatal encephalopathy (HIE). Our group has developed an optically based, noninvasive concept of assessing the capacity for oxygen delivery from the microcirculation to the cells of a tissue under investigation. The hypothesis was that mechanisms of reduced oxygen delivery due to reduced metabolism in cooled asphyxiated neonates could be characterized with this concept. The skin of 28 asphyxiated newborn infants was studied on days 1 and 3 during TH and on day 4 following rewarming with laser Doppler perfusion measurements (LDPM), computer-assisted video microscopy (CAVM), and diffuse reflectance spectroscopy (DRS). Twenty-five healthy neonates served as a control group. The LDPM decreased during cooling (P < 0.01). Functional capillary density was higher both during and following TH compared with control infants (P < 0.01). Capillary flow velocities were reduced during TH (P < 0.05). The heterogeneity of the flow velocities was larger in the HIE infants than in the control infants. Tissue oxygen extraction was higher during TH (P < 0.01). This study indicates that assessments of skin microvascular density, capillary flow velocity, and oxygen extraction can be used to characterize reduced oxygen delivery to cells during TH.

Enhanced angiogenesis in ischemic skeletal muscle after transplantation of cell sheets from baculovirus-transduced adipose-derived stromal cells expressing VEGF165.

IntroductionCell therapy using adipose-derived stromal cells (ADSC) is an intensively developing approach to promote angiogenesis and regeneration. Administration technique is crucial and among others minimal constructs - cell sheets (CS) have certain advantages. Delivery of CS allows transplantation of cells along with matrix proteins to facilitate engraftment. Cells’ therapeutic potential can be also increased by expression of proangiogenic factors by viral transduction. In this work we report on therapeutic efficacy of CS from mouse ADSC transduced to express human vascular endothelial growth factor 165 a/a isoform (VEGF165), which showed potency to restore perfusion and protect tissue in a model of limb ischemia.MethodsMouse ADSC (mADSC) isolated from C57 male mice were expanded for CS formation (106cells per CS). Constructs were transduced to express human VEGF165 by baculoviral (BV) system. CS were transplanted subcutaneously to mice with surgically induced limb ischemia and followed by laser Doppler perfusion measurements. At endpoint animals were sacrificed and skeletal muscle was evaluated for necrosis and vessel density; CS with underlying muscle was stained for apoptosis, proliferation, monocytes and blood vessels.ResultsUsing BV system and sodium butyrate treatment we expressed human VEGF165 in mADSC (production of VEGF165 reached ≈ 25-27 ng/ml/105 cells) and optimized conditions to ensure cells’ viability after transduction. Implantation of mock-transduced CS resulted in significant improvement of limb perfusion, increased capillary density and necrosis reduction at 2 weeks post-surgery compared to untreated animals. Additional improvement of blood flow and angiogenesis was observed after transplantation of VEGF165-expressing CS indicating enhanced therapeutic potential of genetically modified constructs. Moreover, we found delivery of mADSC as CS to be superior to equivalent dose of suspended cells in terms of perfusion and angiogenesis. Histology analysis of extracted CS detected limited proliferation and approximately 10 % prevalence of apoptosis in transplanted mADSC. Significant vascularization of CS and infiltration by monocytes were found in both – BV-transduced and control CS indicating graft and host interaction after transplantation.ConclusionsDelivery of ADSC by subcutaneous transplantation of CS is effective for stimulation of angiogenesis and tissue protection in limb ischemia with a potential for efficacy improvement by BV transduction to express VEGF165.Electronic supplementary materialThe online version of this article (doi:10.1186/s13287-015-0199-6) contains supplementary material, which is available to authorized users.

Skin microcirculation in healthy term newborn infants – assessment of morphology, perfusion and oxygenation

Despite microcirculation's fundamental role, assessments of its function are limited. We explored the applicability of Computer Assisted Video Microscope (CAVM), Laser Doppler Perfusion Measurements (LDPM) and Diffuse Reflectance Spectroscopy (DRS) to study skin microvascular morphology, perfusion and oxygen saturation in twenty-five healthy newborns day 1-3 of life. Day 1-3 (mean (SD)): Microvascular density (CAVM; number of microvessels crossing a grid of lines/mm line, c/mm): Chest: 11.3 (1.5), 11.0 (1.7), 10.7 (1.6). Hand: 13.2 (2.0), 13.2 (1.9), 12.4 (1.6). Capillary density was significantly higher in the hand than in the chest each day (p < 0.001). Perfusion (LDPM; arbitrary units): Chest: 109.1 (26.0), 101.4 (24.6), 100.8 (25.3). Hand: 58.9 (17.5), 54.3 (15.8), 46.9 (14.8). Perfusion was significantly higher in the chest than in the hand each day (p < 0.01). Microvascular oxygen saturation (DRS; %): Chest: 88.1 (5.2), 87.8 (10.0), 86.7 (9.0). Hand: 79.9 (15.2), 82.7 (11.8), 82.2 (12.1) (p < 0.05). Capillary flow velocities (CAVM) were similar in the chest and hand: 60-70% capillaries had "continuous high flow" and 30-40% "continuous low flow".Multimodal skin microvascular assessments with CAVM, LDPM and DRS are feasible with reproducible data in newborns. The hand has lower perfusion, higher capillary density and higher oxygen extraction than the chest.

Contrasting roles of E2F2 and E2F3 in endothelial cell growth and ischemic angiogenesis

The growth of new blood vessels after ischemic injury requires endothelial cells (ECs) to divide and proliferate, and the E2F transcription factors are key regulators of the genes responsible for cell-cycle progression; however, the specific roles of individual E2Fs in ECs are largely unknown. To determine the roles of E2F2 and E2F3 in EC proliferation and the angiogenic response to ischemic injury, hind–limb ischemia was surgically induced in E2F2−/− mice, endothelial-specific E2F3-knockout (EndoE2F3∆/∆) mice, and their littermates with wild-type E2F2 and E2F3 expression. Two weeks later, Laser-Doppler perfusion measurements, capillary density, and endothelial proliferation were significantly greater in E2F2−/− mice and significantly lower in EndoE2F3∆/∆ mice than in their littermates, and EndoE2F3∆/∆ mice also developed toe and limb necrosis. The loss of E2F2 expression was associated with increases in the proliferation and G1/S-phase gene expression of isolated ECs, while the loss of E2F3 expression led to declines in these parameters. Thus E2F2 impairs, and endothelial E2F3 promotes, the angiogenic response to peripheral ischemic injury through corresponding changes in EC cell-cycle progression.

Evaluation of algorithms for microperfusion assessment by fast simulations of laser Doppler power spectral density

In classical laser Doppler (LD) perfusion measurements, zeroth- and first-order moments of the power spectral density of the LD signal are utilized for the calculation of a signal corresponding to the concentration, speed and flow of red blood cells (RBCs). We have analysed the nonlinearities of the moments in relation to RBC speed distributions, parameters of filters utilized in LD instruments and the signal-to-noise ratio. We have developed a new method for fast simulation of the spectrum of the LD signal. The method is based on a superposition of analytically calculated Doppler shift probability distributions derived for the assumed light scattering phase function. We have validated the method by a comparison of the analytically calculated spectra with results of Monte Carlo (MC) simulations. For the semi-infinite, homogeneous medium and the single Doppler scattering regime, the analytical calculation describes LD spectra with the same accuracy as the MC simulation. The method allows for simulating the LD signal in time domain and furthermore analysing the index of perfusion for the assumed wavelength of the light, optical properties of the tissue and concentration of RBCs. Fast simulations of the LD signal in time domain and its frequency spectrum can be utilized in applications where knowledge of the LD photocurrent is required, e.g. in the development of detectors for tissue microperfusion monitoring or in measurements of the LD autocorrelation function for perfusion measurements. The presented fast method for LD spectra calculation can be used as a tool for evaluation of signal processing algorithms used in the LD method and/or for the development of new algorithms of the LD flowmetry and imaging. We analysed LD spectra obtained by analytical calculations using a classical algorithm applied in classical LD perfusion measurements. We observed nonlinearity of the first moment M1 for low and high speeds of particles (v < 2 mm s−1, v > 10 mm s−1). It was also noted that the first moment M1 is less sensitive to the change of the mean RBC speed for flat speed distributions. The low-pass filter frequency f2 implemented in the LD instrument has a significant influence on the first moment of the spectrum. In particular, for a cut-off frequency lower than 10 kHz the M1 value is strongly underestimated.

In vivo optical path lengths and path length resolved doppler shifts of multiply scattered light

In laser Doppler measurements, perfusion values averaged over different and basically unknown path lengths are recorded. To facilitate quantitative path length resolved perfusion measurements, we developed a phase modulated Mach-Zehnder interferometer with spatially separated fibers for illumination and detection. The goal of this study is to measure in vivo optical path lengths and path length resolved Doppler shifts and to compare these with conventional laser Doppler perfusion measurements. With a phase modulated Mach-Zehnder interferometer, we performed path length resolved perfusion measurements on human skin and its variations to external stimuli and compared these with conventional laser Doppler perfusion measurements. The method was evaluated in three human subjects on the dorsal side of the forearm to establish inter-individual within-site variations. Measurements were performed at three different locations of one individual for observing intra-individual inter-site variations resulting from the heterogeneity of the tissue, both in the static matrix and in the microvascular architecture of the skin. In all measurements, perfusion was simultaneously measured with a conventional laser Doppler perfusion monitor. In this study, we show the first results of path length resolved perfusion measurements in skin and its variations to occlusion and Capsicum cream provocation. From our data, we deduced the Doppler shifted fraction of photons, which is related to the blood volume, and the path length dependent average Doppler shift, which is related to the mean velocities of red blood cells. The Doppler shifted fraction of photons is decreased from 28% to 18% during occlusion and increases to 41% when capsicum cream was applied to the skin. Inter- and intra-individual inter-site measurements demonstrated variations in optical path length distributions and path length resolved Doppler shifts. The Doppler shifted fraction of photons measured on the fingertip is about 38% and that measured on the dorsal and palmar sides of the forearm are 32% and 17%, respectively. The path length distributions depend on the skin site that is being probed and the intra-individual inter-site variability is higher than the inter-individual within-site variability measured on comparable sites between different individuals. In this study, we demonstrated, for the first time to our knowledge, that in vivo path length resolved perfusion measurements are feasible. Optical path length distributions of multiply scattered light, spanning a range of 0-6 mm, and their response to external stimuli such as occlusion and capsicum cream provocation have been measured. This method will enable better interpretation of inter- and intra-individual inter-site variations in the LDF readings that are introduced by the variance in tissue optical properties. The inter- and intra-individual inter-site variations measured with our setup results indicate that that these variations should be taken into account while comparing the perfusion readings from comparable sites between individuals and from different sites of the same individual. Furthermore, the observed inter- and intra-individual inter-site variations in path length resolved Doppler measurements indicate the inherent limitation of conventional LDPM that restrict its clinical usefulness, due to its dependence on the unknown photon path length. Consequently, this method will enable to correctly interpret or counter-act the inter- and intra-individual inter-site variations in the LDF readings introduced by the variance in tissue optical properties. This approach also enables to discriminate between the Doppler-shifted photons resulting from interaction with the moving red blood cells and the non-shifted light scattered only by the surrounding static tissue matrices.

Laser-Doppler spectrum decomposition applied for the estimation of speed distribution of particles moving in a multiple scattering medium

Recently, a method for the estimation of speed distribution of particles moving in an optically turbid medium has been proposed. The method allows potentially absolute measurement of speed of the particles and can be applied in laser-Doppler perfusion measurements. However, the decomposition technique was limited to short source–detector separations for which the assumption that one photon is Doppler scattered not more than once is fulfilled. In the present paper we show a generalized decomposition technique in which photons can be scattered more than once. We show the theoretical background for decomposition in such a case. We apply a decomposition method for the analysis of laser-Doppler spectra obtained by Monte Carlo simulations. This analysis allows showing noise limits in which the technique can be effectively applied in analysis of measured spectra. We propose an approximated scattering model based on the assumption that for one photon consecutive Doppler scattering events occur on particles moving with the same speed, and we show that this approximation does not influence significantly the uncertainty of the resulting speed distribution. The proposed decomposition procedure is validated in measurements on a physical flow model. The decomposition procedure is also validated by analysis of spectra measured on a physical phantom using laser-Doppler flow meter (Oxford Optronix, UK). A diluted solution of milk was pumped through a tube fixed in an optically turbid material with speed varying from 0 mm s−1 to 4 mm s−1. We observed a linear relation between actual speed of milk solution and speed estimated from results of spectra decomposition.

Non-invasive laser Doppler perfusion measurements of large tissue volumes and human skeletal muscle blood RMS velocity

This study proposes the implementation of an algorithm allowing one to derive absolute blood root-mean-square (RMS) velocity values from laser Doppler perfusion meter (LDP) data. The algorithm is based on the quasi-elastic light scattering theory and holds for multiple scattering. While standard LDP measurements are normally applicable to a small region of interest (∼1 mm2), the present method allows the analysis of both small and large tissue volumes with small and large interoptode spacings (e.g., 1.5 cm). The applicability and the limits of the method are demonstrated with measurements on human skeletal muscle using a custom-built near-infrared LDP meter. Human brachioradialis muscle RMS velocity values of 9.99 ± 0.01 and 5.58 ± 0.03 mm s−1 at 1.5 cm and of 5.18 ± 0.01 and 2.54 ± 0.09 mm s−1 at 2 cm were found when the arm was (a) at rest and (b) occluded, respectively. At very large optode spacings or very high moving particle densities, the theory developed here would need to be amended to take into account second-order effects.

Effects of pico-tesla electromagnetic field treatment on wound healing in rats.

To evaluate the effects of a pico-tesla electromagnetic field (PTEF) on healing of sutured and open skin wounds and clinicopathologic variables in rats. 64 male Fischer-344 rats. An incision made in the dorsal aspect of the neck was sutured (n = 32) or left open to heal (32). In each group, 16 rats were not PTEF-treated (controls). Wound treatment consisted of exposure to a PTEF once daily. Rats in each group were euthanatized at days 2, 4, 7, and 14. Wounds were evaluated via tensiometry (sutured wounds), digital planimetry (open wounds), laser Doppler perfusion imaging, bacteriologic culture, and histologic examination. Blood samples were collected from all rats for analysis. At day 14, sutured wounds in PTEF-treated rats were stronger (ultimate stress) and tougher (strain energy) than were sutured wounds in control rats. Open wounds in PTEF-treated rats contracted more quickly at days 2 and 4 than did those in control rats. Compared with control wounds, histologic changes (indicative of improved healing) in sutured and open wounds in PTEF-treated rats were detected as early as day 4. Laser Doppler perfusion measurements, results of CBCs, serum biochemical analyses, and bacteriologic cultures were not different between groups. Exposure to the PTEF caused no adverse effects on clinicopathologic, histologic, or bacteriologic variables tested in this study. It appears that PTEF is a safe form of adjuvant treatment for wounds and improves strength of sutured wounds and speeds contraction of open wounds.

Suppressed cutaneous endothelial vascular control and hemodynamic changes in paretic extremities with edema in the extremities of patients with hemiplegia

Wang J-S, Yang CF, Liaw M-Y, Wong M-K. Suppressed cutaneous endothelial vascular control and hemodynamic changes in paretic extremities with edema in the extremities of patients with hemiplegia. Arch Phys Med Rehabil 2002;83:1017-23. Objective: To investigate peripheral circulatory function and its underlying mechanisms in the paretic upper extremity after a stroke. Design: Case-control study. Setting: A department of physical medicine and rehabilitation in Taiwan. Participants: A total of 53 hemiplegic patients (28 men, 25 women; mean age ± standard deviation; 58.2±3.8y) were studied. Subjects were divided into edema and nonedema groups. The edema group included 29 hemiplegic patients with edematous paretic upper extremities. Twenty-four hemiplegic patients in the nonedema group did not suffer from limb edema in the paretic upper extremity. Interventions: Not applicable. Main Outcome Measures: Cutaneous microvascular perfusion responses to 3 grade levels of iontophoretically applied 1% acetylcholine (ACh), 1% ACh plus 1% NG-monomethyl-L-arginine (L-NMMA), and 1% sodium nitroprusside (SNP) in the skin of subjects' forearms were determined by laser Doppler perfusion measurements. Moreover, hemodynamic characteristics in the arterial and venous vessels were measured by impedance plethysmography. Results: Resting arterial inflow and venous capacity, tone, and outflow in paretic extremities did not significantly differ from nonparetic extremities, but the hyperemic arterial inflow was lower in paretic extremities than in nonparetic extremities, and paretic extremities were associated with lower ACh- and ACh plus L-NMMA–induced cutaneous perfusions than nonparetic extremities. ACh-induced cutaneous perfusions also decreased much more significantly in edematous paretic extremities than in nonedematous paretic extremities, and skin vascular responses to SNP do not differ significantly between paretic and nonparetic extremities. Conclusion: Cutaneous microcirculatory function in the paretic upper extremity after stroke may be impaired. The impairment may occur because of decreased endothelium-dependent dilation in skin vasculature. Dysfunction in cutaneous microcirculation tends to be more pronounced in the edematous than in the nonedematous extremities. © 2002 by the American Congress of Rehabilitation Medicine and the American Academy of physical Medicine and Rehabilitation

Impaired Neurogenic Control of Skin Perfusion in Erythromelalgia

Erythromelalgia is a clinical diagnosis characterized by erythema, increased temperature and burning pain in acral skin. The pain is relieved by cooling and aggravated by warming. The symptoms have been hypothesized to be caused by skin hypoxia due to increased arteriovenous shunting. We examined skin microvascular perfusion in response to vasoconstrictory and vasodilatory stimuli, to characterize local and central neurogenic reflexes as well as vascular smooth muscle and vascular endothelial function, using laser Doppler perfusion measurements in 14 patients with primary erythromelalgia and healthy control persons. Skin perfusion preceding provocative stimuli was significantly reduced in patients with erythromelalgia (p < 0.01). The laser Doppler flowmetry signal after sympathetic stimulation of reflexes mediated through the central nervous system, was significantly diminished in patients with erythromelalgia as compared with healthy controls (Valsalva's maneuver p < 0.01; contralateral cooling test p < 0.05). Local neurogenic vasoconstrictor (venous cuff occlusion and dependency of the extremity) and vasodilator reflexes (local heating of the skin), as well as tests for vascular smooth muscle and vascular endothelial function (postocclusive hyperemic response) were maintained. These results indicate that postganglionic sympathetic dysfunction and denervation hypersensitivity may play a pathogenetic role in primary erythromelalgia, whereas local neurogenic as well as endothelial function is unaffected.

Peripheral endothelial dysfunction in heart transplant recipients: possible role of proinflammatory cytokines.

Endothelium-dependent vasodilation in the peripheral circulation may be impaired in heart transplant recipients (HTx rec). Conflicting results have been obtained and the mechanisms involved have not been examined. In the present study, we examined whether long-time survivors of heart transplantation (Tx) show signs of endothelial dysfunction in the peripheral microcirculation, and further investigated the possible role of endothelium-related markers and proinflammatory cytokines in this process. The vasodilatory responses to acetylcholine (Ach) (endothelium-dependent) and sodium nitroprusside (SNP) (endothelium-independent) were evaluated by skin laser-Doppler perfusion measurements in 63 clinically stable HTx rec 6 yr (range 1-13 yr) after Tx, and compared with 20 healthy controls. Ten HTx rec were also followed prospectively with three repeated measurements during the first year after Tx. Plasma von Willebrand factor, big-endothelin (b-ET), and proinflammatory cytokines were measured by enzyme immunoassays. Vascular responses to both Ach and SNP were significantly attenuated in the HTx rec compared with controls. In longitudinal testing, there was a significant reduction in endothelium-dependent vasodilation, but not independent vasodilation from 1 to 12 months after Tx. Plasma levels of vWF and b-ET, as well as levels of proinflammatory cytokines, tumor necrosis factor (TNF)-alpha, interleukin (IL)-6 and IL-1beta, were all markedly increased in HTx rec. HTx rec responses to Ach were negatively correlated to TNF-alpha levels in plasma (r = -0.39, p < 0.01). Moreover, there was also a significant positive correlation between plasma b-ET and TNF-alpha (r = 0.34, p < 0.01). In the long-term follow-up of HTx rec, endothelial dysfunction is demonstrated by both regulation of blood flow in the skin microcirculation and by raised markers of endothelial activation in plasma. This endothelial dysfunction may be related to enhanced levels of proinflammatory cytokines in these patients.

Prediction of imminent amputation in patients with non-reconstructible leg ischemia by means of microcirculatory investigations

Purpose: We investigated the usefulness of skin microcirculatory investigations to predict imminent major amputation in patients with non-reconstructible critical limb ischemia. Methods: One hundred eleven patients with non-reconstructible chronic rest pain or small ulcers and an ankle blood pressure of 50 mm Hg or less or an ankle-to-brachial pressure index of 0.35 or less were included. Nailfold capillary microscopy (CM; big toe, sitting), transcutaneous oxygen pressure (TcpO 2; forefoot, supine; 44°C), and laser Doppler perfusion measurements (LD; pulp of big toe, supine) were performed at rest and during reactive hyperemia. Patients were classified according to their skin microcirculatory status just before the start of the treatment in three groups: those with a “good,” “intermediate,” or “poor” microcirculation, according to a combination of predefined cutoff values (Poor: capillary density less than 20/mm 2, absent reactive hyperemia in CM and LD, TcpO 2 less than 10 mm Hg; good: capillary density of 20/mm 2 or more, present reactive hyperemia in CM and LD, TcpO 2 of 30 mm Hg or more). Subsequently, patients received maximum conservative therapy from the surgeon, who was unaware of the microcirculatory results. After a follow-up period of as long as 36 months, limb survival and disposing factors were analyzed and compared with the initial microcirculatory status. Results: Cox regression analysis showed a significant prognostic value of the microcirculatory classification (hazard ratio = 0.28, P < .0001), but not of the Fontaine stage, ankle blood pressure, or the presence of diabetes mellitus for the occurrence of an amputation. Positive and negative predictive values were 73% and 67%, respectively. The cumulative limb survival at 6 and 12 months was 42% and 17% in the poor microcirculatory group, 80% and 63% in the intermediate microcirculatory group, and 88% and 88% in the good microcirculatory group ( P < .0001, log-rank). Conclusion: Microcirculatory screening and classification is useful in detecting non-reconstructible critical ischemia that requires amputation, which is not detectable by means of the clinical stage or blood pressure parameters. Most of the poor patient group will require amputation. In the intermediate and good groups, nonsurgical treatment appears sufficient for limb salvage. (J Vasc Surg 1999;30:114-21.)