Development of a clinically useful mechanical leech device that promotes flap survival in an animal model of venous-congested skin flaps.

The authors describe the design and testing of a mechanical leech device that has shown greater efficacy in alleviating venous congestion and promoting skin flap survival than previously described clinical therapies. Abdominal skin flaps (3 x 6 cm) were raised on Sprague-Dawley rats and were subjected to two ischemic events that simulated clinical venous congestion. The animals received two treatments with the device prototype during 7-day experiment (N = 9). Flap perfusion was monitored with a scanning laser Doppler imager (LDI) and was normalized to adjacent skin perfusion. Photographic images of flaps at 7 days were assessed for areas of nonnecrotic tissue, and LDI data consisted of perfusion measurements at nine time points during the 7-day experiment. The device prototype was able to promote an average survival area in the flap of 60.0 +/- 3.5%, which was not only a significant (p < 0.05) increase over previously published control groups (8.0 +/- 5.0%) that did not receive any treatment, but was a significant increase over the same treatment schedule with clinically used leeches (34.6 +/- 7.3%). At 7 days, LDI data showed flap perfusion to be 65.6 +/- 7% of adjacent skin perfusion, a significant increase over this level at the end of the second ischemic event (21 +/- 1%; p < 0.05). These results suggest that this device could be used clinically to alleviate venous congestion and to promote flap survival. The device could also be useful for prophylactic treatments and to minimize treatment delays because its long shelf-life permits immediate availability when a treatment decision is made.

Selective A(2A) adenosine receptor activation reduces skin pressure ulcer formation and inflammation.

Activation of A(2A) adenosine receptors (A(2A)-AR) by ATL-146e (formerly DWH-146e) prevents inflammatory cell activation and adhesion. Recurrent ischemia-reperfusion (I/R) of the skin results in pressure ulcer formation, a major clinical problem. ATL-146e was evaluated in a novel reproducible rat model of pressure ulcer. A 9-cm(2) region of dorsal rat skin was cyclically compressed at 50 mmHg using a surgically implanted metal plate and an overlying magnet to generate reproducible tissue necrosis. Osmotic minipumps were implanted into 24 rats divided into four equal groups to infuse vehicle (control), ATL-146e (0.004 microg x kg(-1) x min(-1)), ATL-146e plus an equimolar concentration of A(2A) antagonist, ZM-241385, or ZM-241385 alone. Each group received 10 I/R cycles. In non-I/R-treated skin, ATL-146e has no effect on blood flow. I/R-treated skin of the ATL-146e group compared with the vehicle group had 65% less necrotic area, 31% less inhibition of average skin blood flow, and fewer extravasated leukocytes (23 +/- 3 vs. 49 +/- 6 per 500 microm(2)). These data suggest that ATL-146e, acting via an A(2A)-AR, reduces leukocyte infiltration and is a potent prophylactic for I/R injury in skin.

Chronic vasodilation induces matrix metalloproteinase 9 (MMP-9) expression during microvascular remodeling in rat skeletal muscle.

OBJECTIVE: The process of microvessel growth and remodeling depends on the presence of matrix metalloproteinases (MMPs) in a specific spatial pattern of expression. This study characterizes the spatial distribution of metalloproteinase-9 (MMP-9) expression during microvascular remodeling in the rat spinotrapezius muscle. METHODS: Female Sprague-Dawley rats (4 weeks old) were administered the alpha1-adrenergic blocker prazosin for 7 days to induce chronic vasodilation and associated increases in capillary and arteriolar density. MMP-9 expression was analyzed by Western blotting analysis of microdissected regions of muscle and immunolabeling of muscle sections. RESULTS: Capillary density expressed as both capillary-to-fiber ratio (C/F) and capillaries per mm2 increased significantly (p < 0.01) due to prazosin administration. Western blotting on microdissected regions of spinotrapezius muscle showed that there was an increase in MMP-9 expression in prazosin-treated animals. Additionally, the Western blots showed a presence of the activated form of MMP-9 in regions of spinotrapezius muscle containing vessels on the order of capillaries and small arterioles. Immunohistochemistry demonstrated that MMP-9 significantly increased in the muscles of treated animals in areas within the vessel walls of microvessels greater than 20 microm in diameter. However, interstitial MMP-9 expression did not significantly increase with prazosin administration. CONCLUSIONS: These results demonstrate that the expression of MMP-9 increases during in vivo microvascular remodeling in adult skeletal muscle. The MMP-9 expression is limited to larger (>20-microm diameter) microvessels, suggesting a role for MMP-9 in the remodeling of such vessels during prazosin-induced vasodilation and the subsequent capillary proliferation.

In vivo chemotactic properties and spatial expression of PDGF in developing mesenteric microvascular networks.

The recruitment of perivascular cells to developing microvessels is a key component of microvessel assembly. Whereas platelet-derived growth factor (PDGF) signaling is critical for this process during embryonic development, its role from the postnatal stages through adulthood remains unclear. We investigated the potential role of PDGF signaling during microvessel assembly by measuring in vivo the migration of labeled fibroblasts to PDGF in mesenteric connective tissue and by examining PDGF-B and PDGF receptor-beta (PGDFR-beta) expression in microvascular networks during normal maturation. PDGF-B homodimer (PDGF-BB; 30 ng/ml) application elicited a significant (P < 0.05) increase (7.8 +/- 4.1 cells) in labeled fibroblasts within 100 microm of the source micropipette after 2 h. PDGF-A homodimer (30 ng/ml) application and control solution did not elicit directed migration. PDGF-B was expressed in microvessel endothelium and smooth muscle, whereas PDGFR-beta was expressed in endothelium, smooth muscle, and interstitial fibroblasts. Given that PDGF-BB elicits fibroblast migration in the mesentery and that PDGF-B and PDGFR-beta are expressed in a pattern that indicates paracrine signaling from microvessels to the interstitium, the results are consistent with a role for PDGF-B in perivascular cell recruitment to microvessels.

Ischemia-reperfusion injury in chronic pressure ulcer formation: a skin model in the rat.

Most animal models of chronic pressure ulcers were designed to study only the role of ischemic injury in wound formation, often using single applications of constant pressure. The purpose of this study was to develop and characterize a reproducible model of cyclic ischemia-reperfusion injury in the skin of small un-anesthetized animals using clinically relevant pressures and durations. Ischemia-reperfusion injury was created in a 9 cm2 region of dorsal skin in male rats by periodically compressing skin under a pressure of 50 mm Hg using an implanted metal plate and an overlying magnet. We varied the total number of ischemia-reperfusion cycles, examined the effect of varying the frequency and duration of ischemic insult, and compared ischemia-induced injury to ischemia-reperfusion-induced injury with this model. Tissue injury increased with an increasing number of total ischemia-reperfusion cycles, duration of ischemia, and frequency of ischemia-reperfusion cycles. This model generates reproducible ischemia-reperfusion skin injury as characterized by tissue necrosis, wound thickness, leukocyte infiltration, transcutaneous oxygen tension, and wound blood flow. Using this model, the biological markers of ischemia-reperfusion-induced wound development can be studied and therapeutic interventions can be evaluated in a cost-effective manner.

Role of leukocytes and tissue-derived oxidants in short-term skeletal muscle ischemia-reperfusion injury.

The relative contribution of xanthine oxidase (XO) and leukocytes to tissue injury after short-term ischemia is unknown. In this study, we subjected three groups of rat spinotrapezius muscles to 30-min ischemia and 1-h reperfusion: 1) ischemia-reperfusion (I/R) + 0.9% saline, 2) I/R + superoxide dismutase, and 3) I/R + oxypurinol. A fourth group served as nonischemic control. We quantified the increase in resistance (%DeltaR) caused by leukocyte-capillary plugging concurrently with myocyte uptake of propidium iodide (PI) [expressed as no. of PI spots per total volume of perfused tissue (N(PI)/V)] and performed assays to quantify XO activity, thiobarbituric acid-reactive substances (TBARS), and myeloperoxidase (MPO). Groups 2 and 3 exhibited significant decreases in N(PI)/V relative to group 1. MPO levels and TBARS were similar among all groups, and mean %DeltaR was significantly reduced in groups 2 and 3 relative to group 1. However, elevated XO was observed in groups 1 and 2 relative to group 3 and nonischemic controls. These data are consistent with the hypothesis that XO, rather than toxic species produced by plugging or venule-adherent leukocytes, is responsible for postischemic damage in this model.

Lysophosphatidic acid enhances healing of acute cutaneous wounds in the mouse.

Lysophosphatidic acid is a phospholipid growth factor and intercellular signaling molecule released by activated platelets and injured fibroblasts that affects keratinocytes, fibroblasts, neutrophils,and monocytes. We therefore hypothesized that lysophosphatidic acid could influence the inflammation and reepithelialization phases of wound healing. Lysophosphatidic acid (100 microM) was applied daily for 5 days to 2 mm-diameter excisional mouse ear skin wounds and re-epithelialization was measured. We also evaluated whether the bioactivity of lysophosphatidic acid could be increased by zinc (Zn2+, 1 mM). Inflammatory cells were counted in wound sections after 1, 3, or 5 days of healing. Reepithelialization was accelerated significantly by either lysophosphatidic acid or lysophosphatidic acid + Zn2+ (p < 0.01 and p < 0.0001, respectively). Both lysophosphatidic acid solutions significantly increased the amount of new epithelium in the wounds on days 1, 2, and 3 (p < 0.05). Little wound area remained on day 4, and all wounds were fully reepithelialized by day 5. Lysophosphatidic acid did not affect the number of neutrophils or macrophages present in the wound area. Our findings show that lysophosphatidic acid increased the amount of reepithelialization in the early stages of cutaneous wound healing in excisional ear wounds, without affecting inflammatory function.

Evaluation of clinically applicable exsanguination treatments to alleviate venous congestion in an animal skin flap model.

This study compares the effectiveness of alleviating venous congestion with mechanically-made outlets or leech therapy in promoting skin flap survival. Free flaps of abdominal skin (3 x 6 cm) were raised on Sprague-Dawley rats and subjected to ischemic events, simulating venous congestion. Animals received 1) no treatment; 2) two treatments involving two 18-gauge needle-puncture outlets; or 3) two sessions of leech therapy. Flap perfusion was monitored with a scanning laser Doppler flowmeter. Photographic images of flaps at 7 days were assessed for areas of normal tissue (n = 15), and laser Doppler flowmeter data consisted of control (n = 6), outlet (n = 6), and leech (n = 7). Both the needle-puncture outlet (40.0% +/- 9.24%) and leech treated (34.6% +/- 7.34%) groups had a significantly greater surviving skin area than untreated control flaps (8.0% +/- 5.0%), with 2 of 15 flaps receiving mechanical outlets exhibiting > 90% surviving area. After 7 days, laser Doppler flowmeter data showed greater mean perfusion in the outlet (71.7% +/- 16.8%) and leech (92.6% +/- 17.2%) treated groups, compared to controls (15.2% +/- 10.2%). There was a significant increase in perfusion in the outlet (13.3% +/- 6.2%) and leech (9.1% +/- 1.1%) treated groups from the end of secondary ischemia to day 7 (p < 0.05) compared to controls. The results suggest that two spatially separated outlets are as effective as one leech in increasing the area of surviving skin in venous congested flaps.

A dedication in memoriam of Dr. Richard Skalak.

Richard Skalak (1923-1997) played a leadership role in the formative decades of the discipline of biomedical engineering through his technical contributions in biomechanics, his educational influence on students, and his service to many developing societies and journals. But always, the distinguishing marks of his involvement with any activity or person were his generosity, respect and tolerance for others, integrity, and curiosity. These very qualities are what first brought him as a traditional engineering trained in engineering mechanics into the young field of biomedical engineering in the 1960s, and they are what led him to new approaches to cellular and molecular engineering, tissue engineering, and orthopedic biomechanics. His technical papers and lectures on blood cell mechanics, pulmonary circulation, dental implants, and tissue growth were models of clarity and often pointed the way to new areas of exploration, while his personal writings offer advice on life, academic organizations, and the pursuit of significant work. He would be deeply appreciative that this first volume of the Annual Review of Biomedical Engineering is dedicated to his memory.

The Microcirculation Physiome Project.

Presented is a discussion of steps towards the creation of a database of the microcirculation encompassing anatomical and functional experimental data, and conceptual and computational models. The discussion includes issues of database utility, organization, data deposition, and linkage to other databases. The database will span levels from gene to tissue and will serve both research and educational purposes.

Spontaneous redistribution after reperfusion: a unique property of AIP 201, an ultrasound contrast agent.

OBJECTIVES: We sought to determine the mechanism of spontaneous redistribution of AIP 201 microbubbles after reperfusion from a single left heart injection performed during coronary occlusion. BACKGROUND: AIP 201, an ultrasound contrast agent consisting of 10-microm sized microbubbles, has demonstrated spontaneous myocardial redistribution in preliminary studies. METHODS: Myocardial video intensity (VI) and radiolabeled microsphere-derived myocardial blood flow (MBF) were measured serially after reperfusion in seven dogs undergoing an AIP 201 injection during coronary occlusion. The behavior of these bubbles was also assessed in the rat spinotrapezius muscle using intravital microscopy (IM), both with and without ultrasound. The effect of ultrasound on these bubbles was also determined in vitro. RESULTS: A spontaneous and gradual increase in myocardial VI was noted after reperfusion, which was related to the magnitude of increase in MBF to that region (r=0.82, p < 0.001). On IM, most of the microbubbles were seen entrapped in small arterioles. Some larger arterioles had aggregates of microbubbles that periodically became dislodged and moved downstream. This behavior was not affected in vivo by ultrasound. In vitro, however, microbubble aggregation was noted only during ultrasound exposure. CONCLUSIONS: The magnitude of redistribution of AIP 201 microbubbles to the reperfused myocardium is related to changes in MBF and occurs from their dislodgement from microbubble aggregates entrapped in large arterioles. In vitro microbubble aggregation seen during ultrasound exposure was not reproduced in vivo. These results may have important implications for studying the effects of interventions in acute coronary syndromes and after coronary artery bypass graft surgery.

Delivery of colloidal particles and red blood cells to tissue through microvessel ruptures created by targeted microbubble destruction with ultrasound.

BACKGROUND: We have previously shown that the application of ultrasound to thin-shelled microbubbles flowing through small microvessels (<7 microm in diameter) produces vessel wall ruptures in vivo. Because many intravascular drug- and gene-delivery vehicles are limited by the endothelial barrier, we hypothesized that this phenomenon could be used to deliver drug-bearing vehicles to tissue. METHODS AND RESULTS: An exteriorized rat spinotrapezius muscle preparation was used. Intravascular fluorescent red blood cells and polymer microspheres (PM) (205 and 503 nm in diameter) were delivered to the interstitium of rat skeletal muscle through microvessel ruptures created by insonifying microbubbles in vivo. On intravital microscopy, mean dispersion areas per rupture for red blood cells, 503-nm PM, and 205-nm PM were 14.5x10(3) microm2, 24. 2x10(3) microm2, and 27.2x10(3) microm2, respectively. PM dispersion areas were significantly larger than the mean dispersion area for red blood cells (P<0.05). CONCLUSIONS: Microvessel ruptures caused by insonification of microbubbles in vivo may provide a minimally invasive means for delivering colloidal particles and engineered red blood cells across the endothelial lining of a targeted tissue region.

Arteriolar remodeling in skeletal muscle of rats exposed to chronic hypoxia.

The topological structure of the arteriolar network is an important determinant of microvascular resistance. In several normal and pathological conditions, this network topology is remodeled such that adequate tissue perfusion and oxygenation are maintained. The objective of this study was to quantify the extent of arteriolar remodeling in skeletal muscle of rats chronically exposed to 10% oxygen for 18 +/- 3.6 days. Arcade arteriolar (AA) and transverse arteriolar networks that had been immunofluorescently labeled for smooth muscle alpha-actin and smooth muscle myosin heavy chain were observed in whole-mount gracilis muscles from hypoxic and weight-matched control rats. Eight muscles from 4 animals were used for analysis in each group. The percentage (+/- SD) of terminal arteriolar endings that were positive for smooth muscle alpha-actin, but negative for smooth muscle myosin heavy chain increased significantly (p < 0.05) from 29.2 +/- 8.0 in controls to 70.7 +/- 9.0 in hypoxia, indicating that the rate of terminal arteriolar development was elevated in the hypoxic animals. The number of AA loops/muscle (+/- SD) increased significantly from 9.6 +/- 2.3 in controls to 14.4 +/- 4.6 in hypoxia. This increase in AA loops/muscle was due primarily to a large increase in the number of small-diameter (<15 microm) AA segments/muscle, suggesting that new AA loops were formed via the recent anastomosing of developing small-diameter terminal arterioles. The results demonstrate that exposure to chronic hypoxia stimulates significant remodeling of both the arcade and transverse arteriolar networks in skeletal muscle.

Distribution of cellular proliferation in skeletal muscle transverse arterioles during maturation.

OBJECTIVE: To investigate the spatial and phenotypic origin of the new smooth muscle (SM) cells that are necessary for transverse arteriolar (TA) remodeling by establishing the distribution of cellular proliferation in TA trees during maturation. METHODS: Whole-mount gracilis muscles from rats at 4 and 9 weeks of age were immunolabeled for SM myosin heavy chain to denote arterioles and for bromodeoxyuridine to denote S-phase (DNA synthesizing) nuclei. The dimensions of each clearly visible segment in TA trees were measured. S-phase cells in the wall of, or within 5 microns of, TA segments were identified as (1) endothelium or intimal fibroblasts, (2) SM, or (3) interstitial cells. The relative percentages of each cell type in S-phase, the distribution of arteriolar diameters containing S-phase SM, and the density of S-phase interstitial cells (per unit length and per unit surface area of TA) were determined. Alcian blue counterstaining was used to discern the percentage of interstitial cells that were mast cells. RESULTS: At 4 and 9 weeks, respectively, 3.7% and 2.1% of S-phase cells were endothelium or intimal fibroblasts, 3.0% and 4.2% were SM, and 93.3% and 93.7% were interstitial cells. No S-phase interstitial cells within 5 microns of TAs were mast cells. The mean diameter of TA segments containing as S-phase SM nucleus was 15.22 +/- 1.2 microns at 4 weeks of age, with the minimum diameter being 8.9 microns. From 4 to 9 weeks of age, the number of interstitial cells per unit length of TA decreased 10-fold from 15.2 (n = 115) to 1.5 (n = 182) cells/min. At 4 weeks, the density of S-phase interstitial cells was greatest surrounding the most terminal arterioles. CONCLUSIONS: When coupled with the result that S-phase SM is absent in the most terminal segments, the relatively high density of S-phase interstitial cells surroundings the smallest diameter terminal segments at 4 weeks of age is consistent with the hypothesis that fibroblast hyperplasia is a component of terminal arteriolar development.

Direct in vivo visualization of intravascular destruction of microbubbles by ultrasound and its local effects on tissue.

BACKGROUND: Our aim was to observe ultrasound-induced intravascular microbubble destruction in vivo and to characterize any resultant bioeffects. METHODS AND RESULTS: Intravital microscopy was used to visualize the spinotrapezius muscle in 15 rats during ultrasound delivery. Microbubble destruction during ultrasound exposure caused rupture of < or = 7-microm microvessels (mostly capillaries) and the production of nonviable cells in adjacent tissue. The number of microvessels ruptured and cells damaged correlated linearly (P<0.001) with the amount of ultrasound energy delivered. CONCLUSIONS: Microbubbles can be destroyed by ultrasound, resulting in a bioeffect that could be used for local drug delivery, angiogenesis, and vascular remodeling, or for tumor destruction.

Prazosin administration enhances proliferation of arteriolar adventitial fibroblasts.

Chronic vasodilation stimulates the formation of new arterioles in skeletal muscle, a process that requires the differentiation of mesenchymally derived precursor cells on the abluminal surface of capillaries. Fibroblast proliferation and migration to the arterializing capillary likely precede this differentiation process. In the current study, we investigated the effects of chronic vasodilation with the alpha1 adrenergic blocker prazosin, a treatment that produces enhanced terminal arteriolar development, on the proliferation of fibroblasts present in the adventitia of transverse arterioles. Dual-immunofluorescence labeling for the smooth muscle contractile protein SM-myosin heavy chain (MHC) and for bromodeoxyuridine (BRDU) uptake revealed that prazosin treatment for 4 days stimulated a threefold increase in the density of proliferating fibroblasts surrounding transverse arteriolar trees. This increase was primarily due to an eightfold increase in the density of S-phase fibroblasts surrounding <8 micron m diameter terminal arterioles and a 280% increase in the density of S-phase fibroblasts surrounding 8- to 12-micron m terminal arterioles. Alcian blue counterstaining indicated that no proliferating cells were mast cells. An in vitro study demonstrated that prazosin, at concentrations of 0.5 and 0.05 mg/liter, has no direct effect on fibroblast proliferation. It is concluded that chronic vasodilation with prazosin, a treatment that elicits elevated levels of hemodynamic stress, stimulates the proliferation of adventitial fibroblasts, particularly at the terminal endings of transverse arteriolar trees.

Contribution of individual structural components in determining the zero-stress state in small arteries.

Arterial-wall stress distributions are important determinants of arterial function and are determined by the mechanical properties of the vessel wall, the physiological loading conditions, and the zero stress state of the artery. In this study, the relationship between this zero-stress state, the contractile state of the vessel, and the extracellular matrix components was investigated. Rat saphenous arteries were excised, and cross-sectional slices were cut. Wall thickness, intimal circumference, medial to adventitial border circumference, and the outer adventitial circumference were measured. Each slice was cut once longitudinally to relieve the residual stresses, and after a period of 30 min, vessel dimensions were again measured, and the angle to which the arterial section opened, a measure of the residual stress present in the intact artery, was recorded in this new zero-stress state. Control 'opening angle' was 112+/-10 degrees (mean+/-SD, n= 8) as compared to 138+/-6 degrees (n = 18) and 127+/-6 degrees (n = 11) for slices placed in 10(-4) M adenosine at 25 and 37 degrees C, respectively. Fluorescein isothiocyanate dextran was also injected intravenously in 8 animals to measure the homeostatic lumen diameter and wall thickness using intravital microscopy. Comparison of homeostatic strains calculated from these in situ dimensions to residual strains indicated that a given opening angle produced a radially uniform circumferential strain only for a specific level of tone. These results suggest that smooth muscle tone can acutely modify residual strain, possibly through interconnections with matrix components. Furthermore, selective elimination of matrix components by post-treatment of zero-stress slices with elastase or collagenase in 6 animals each increased the opening angle by 53 and 70%, respectively, suggesting that the extracellular matrix structure may regulate arterial strains in the long term.

Flow characteristics of peripherally inserted central catheters.

PURPOSE: Clinical applications of peripherally inserted central catheters (PICCs) are limited by the relatively small lumina and long lengths of these devices. Quantitative analysis of the flow capabilities of a variety of PICCs was performed to aid in deciding which patients should have a PICC and in selecting the appropriate catheter. MATERIALS AND METHODS: Sixteen different PICCs from six manufacturers were infused at flow rates of 25-270 mL/h. Infusions were performed with distilled water, normal saline, total parenteral nutrition solution, intralipids, and blood. Flow versus pressure curves were generated for each PICC and infusate. Additional catheter data recorded included the working length, outer diameter (OD), and inner diameter (ID) of the PICCs. RESULTS: Because of the thin wall construction of polyurethane catheters, PICCs made from polyurethane showed much better flow rates than silicone PICCs of a comparable OD. The measured ODs of the PICCs were 4-6 F, whereas the IDs ranged from 0.012 to 0.032 inch. Because of the small ID of some PICCs, infusing blood or intralipids is not practical. CONCLUSION: There is significant variability in the flow capabilities of available PICCs. Many of the PICCs require pressures greater than those that can be generated by commercially available infusion pumps. Matching PICC characteristics to the desired application will avoid many of the clinical problems currently encountered with PICCs.

Accuracy of the conductance catheter for measurement of ventricular volumes seen clinically: effects of electric field homogeneity and parallel conductance.

The conductance-volume method is an important clinical tool which allows the assessment of left ventricular function in vivo. However, the accuracy of this method is limited by the homogeneity of electric field the conductance catheter produces and the parallel conductance of surrounding structures. This paper examines these sources of error in volumes seen clinically. The characteristics of electric field within a chamber were examined using computer simulation. Nonconductive and conductive models were constructed and experimental measurements obtained using both single-field (SF) and dual-field (DF) excitation. Results from computer simulations and in vitro measurements were compared to validate the purposed theoretical model of conductance-volume method. The effects of field homogeneity and significance of parallel conductance in volume measurement were then determined. The results of this study show that DF provide a more accurate measure of intraventricular volume than SF, especially at larger volumes. However, both significantly underestimate true volume at larger volumes. In addition, the parallel conductance due to the chamber wall is significant at small volumes, but diminishes at larger volumes. Furthermore, the effect of parallel conductance beyond the chamber wall may be negligible. This study demonstrates the limitations in applying current conductance technology to patients with dilated hearts.