Gq-activated fibroblasts induce cardiomyocyte action potential prolongation and automaticity in a three-dimensional microtissue environment.

Cardiac fibroblasts (CFs) are known to regulate cardiomyocyte (CM) function in vivo and in two-dimensional in vitro cultures. This study examined the effect of CF activation on the regulation of CM electrical activity in a three-dimensional (3-D) microtissue environment. Using a scaffold-free 3-D platform with interspersed neonatal rat ventricular CMs and CFs, Gq-mediated signaling was selectively enhanced in CFs by Gαq adenoviral infection before coseeding with CMs in nonadhesive hydrogels. After 3 days, the microtissues were analyzed by signaling assay, histological staining, quantitative PCR, Western blots, optical mapping with voltage- or Ca2+-sensitive dyes, and microelectrode recordings of CF resting membrane potential (RMPCF). Enhanced Gq signaling in CFs increased microtissue size and profibrotic and prohypertrophic markers. Expression of constitutively active Gαq in CFs prolonged CM action potential duration (by 33%) and rise time (by 31%), prolonged Ca2+ transient duration (by 98%) and rise time (by 65%), and caused abnormal electrical activity based on depolarization-induced automaticity. Constitutive Gq activation in CFs also depolarized RMPCF from -33 to -20 mV and increased connexin 43 and connexin 45 expression. Computational modeling confers that elevated RMPCF and increased cell-cell coupling between CMs and CFs in a 3-D environment could lead to automaticity. In conclusion, our data demonstrate that CF activation alone is capable of altering action potential and Ca2+ transient characteristics of CMs, leading to proarrhythmic electrical activity. Our results also emphasize the importance of a 3-D environment where cell-cell interactions are prevalent, underscoring that CF activation in 3-D tissue plays a significant role in modulating CM electrophysiology and arrhythmias.NEW & NOTEWORTHY In a three-dimensional microtissue model, which lowers baseline activation of cardiac fibroblasts but enables cell-cell, paracrine, and cell-extracellular matrix interactions, we demonstrate that selective cardiac fibroblast activation by enhanced Gq signaling, a pathophysiological trigger in the diseased heart, modulates cardiomyocyte electrical activity, leading to proarrhythmogenic automaticity.

Functional scaffold-free 3-D cardiac microtissues: a novel model for the investigation of heart cells.

To bridge the gap between two-dimensional cell culture and tissue, various three-dimensional (3-D) cell culture approaches have been developed for the investigation of cardiac myocytes (CMs) and cardiac fibroblasts (CFs). However, several limitations still exist. This study was designed to develop a cardiac 3-D culture model with a scaffold-free technology that can easily and inexpensively generate large numbers of microtissues with cellular distribution and functional behavior similar to cardiac tissue. Using micromolded nonadhesive agarose hydrogels containing 822 concave recesses (800 µm deep × 400 µm wide), we demonstrated that neonatal rat ventricular CMs and CFs alone or in combination self-assembled into viable (Live/Dead stain) spherical-shaped microtissues. Importantly, when seeded simultaneously or sequentially, CMs and CFs self-sorted to be interspersed, reminiscent of their myocardial distribution, as shown by cell type-specific CellTracker or antibody labeling. Microelectrode recordings and optical mapping revealed characteristic triangular action potentials (APs) with a resting membrane potential of -66 ± 7 mV (n = 4) in spontaneously contracting CM microtissues. Under pacing, optically mapped AP duration at 90% repolarization and conduction velocity were 100 ± 30 ms and 18.0 ± 1.9 cm/s, respectively (n = 5 each). The presence of CFs led to a twofold AP prolongation in heterogenous microtissues (CM-to-CF ratio of 1:1). Importantly, Ba(2+)-sensitive inward rectifier K(+) currents and Ca(2+)-handling proteins, including sarco(endo)plasmic reticulum Ca(2+)-ATPase 2a, were detected in CM-containing microtissues. Furthermore, cell type-specific adenoviral gene transfer was achieved, with no impact on microtissue formation or cell viability. In conclusion, we developed a novel scaffold-free cardiac 3-D culture model with several advancements for the investigation of CM and CF function and cross-regulation.

Respiratory enzymes and mitochondrial morphology of HeLa and L cells treated with chloramphenicol and ethidium bromide.

Exposure of HeLa and L cells to chloramphenicol causes a progressive dose-dependent decrease in cytochrome oxidase and succinate-cytochrome c reductase activities, concomitant with an increase in the amount of cytochrome c. At 2-3 days, the specific activities of the enzymes have fallen to about one-half of control values; the mitochondria appear swollen. By day 5, enzyme activities are about one-quarter of control values; the mitochondria are more swollen, with disorientation and disintegration of cristae. By day 6-8, after three generations, growth has stopped, enzyme activities are approximately the same as on day 5, and cytochrome c content has reached 170% of control value. Mitochondria show severe changes, cristae being affected more than peripheral inner membrane. The number of profiles continues to be nearly normal. After 30 days, cytochrome oxidase activity remains low but now there are mitochondria in intermediate and condensed configuration. There is a gradual accumulation in the cytoplasm of smooth membrane elements. If chloramphenicol is removed, cells recover. Ethidium bromide treatment for up to 8 days yields results virtually identical to those obtained with chloramphenicol. Cells treated with 10(-4)M KCN show a decrease in cytochrome oxidase activity to about one-third of control value and an elevated amount of cytochrome c. Only a small number of mitochondria appear damaged. Autochthonous mitochondrial syntheses appear to be essential for the organization of the cristae. When cytochrome oxidase activity is impaired, a regulatory mechanism for cytochrome biosynthesis geared to mitochondrial function may be lacking, resulting in an increase in cytochrome c content.

Molecular beam scattering from liquid surfaces.

By means of controlled collisions of atoms and molecules with liquid surfaces, molecular beam experiments can be used to probe how gases stick to, rebound from, and exchange energy with molecules in the liquid phase. This report describes measurements of energy exchange in collisions between gases (neon, xenon, and sulfur hexafluoride) and polyatomic liquids (squalane and perfluoropolyether). Energy transfer depends critically on liquid composition and is more efficient for the hydrocarbon than for the perfluorinated ether.

Regulation of de novo purine synthesis in human bone marrow mononuclear cells by hypoxanthine.

In previous studies from this laboratory, human bone marrow hypoxanthine concentrations were found to average 7.1 microM, three times higher than plasma hypoxanthine concentrations measured simultaneously. To assess the significance of this finding, the relationship between hypoxanthine concentration and the rate of purine nucleotide synthesis by the de novo pathway was studied in normal human bone marrow mononuclear cells and in the human promyelocytic cell line, HL-60, in vitro. Utilizing a [14C]formate incorporation technique, rates of total cellular de novo purine synthesis as well as rates of de novo adenine, de novo guanine, and thymine synthesis and incorporation into RNA and DNA were measured as a function of hypoxanthine concentration. In normal human marrow cells, the rate of total de novo purine synthesis declined by 81%, while the rate of de novo adenine and de novo guanine synthesis and incorporation into macromolecules declined by 89 and 75%, respectively, when media hypoxanthine was increased from 0 to 10 microM. Similar results were seen in the HL-60 cell line. In contrast, rates of thymine synthesis and incorporation into DNA as well as overall rates of RNA and DNA synthesis did not change with varying media hypoxanthine concentrations. In addition, hypoxanthine salvage and incorporation into RNA and DNA was shown to progressively increase with increasing media hypoxanthine concentrations. These results indicate that physiologic concentrations of hypoxanthine are sufficient to regulate the rate of de novo purine synthesis in human bone marrow in vivo.

Intraperitoneal cytosine arabinoside therapy in ovarian carcinoma.

Previous pharmacokinetic modeling has suggested that cytosine arabinoside (ara-C) may be an ideal agent for peritoneal administration in the treatment of intraabdominal neoplasms. In vitro drug sensitivity testing with clonogenic assays demonstrated sensitivity to the high concentrations of ara-C potentially achievable by intraperitoneal (IP) administration in five of nine patients with ovarian carcinoma. The kinetics of IP ara-C were then studied in three patients, and a 2 to 3 log concentration gradient was observed between the peritoneal compartment and plasma. Subsequently, the clinical efficacy of IP ara-C given for a five-day course every four weeks was evaluated in 10 patients with ovarian carcinoma who had failed systemic chemotherapy. Chemical peritonitis did not occur and systemic toxicity was minimal. Two patients were rendered free of disease and are alive without disease 14+ and 15+ months since therapy was discontinued. These results demonstrate that there is a very large pharmacologic advantage to the IP route of administration for ara-C, that ara-C is active against advanced ovarian carcinoma, and that the IP route of administration permits the cytokinetically rational use of very long durations of exposure for this cell-cycle phase-specific agent.

Limits of normal left ventricular dimensions in growth and development: analysis of dimensions and variance in the two-dimensional echocardiograms of 268 normal healthy subjects.

The majority of studies generating normal echocardiographic reference values for left ventricular dimensions have been based on blindly performed M-mode measurements, and there are no previous reports based on two-dimensional echocardiography that provide a comprehensive analysis of the two-dimensional measurements from infancy to old age. This report presents the results of analyzing the left ventricular internal dimensions from cross-sectional echocardiographic studies on 268 normal healthy subjects (none were hospitalized for any reason) whose ages ranged from 6 days to 76 years. The mean data are reported as functions of body surface area and, in addition, the variance is modeled as a function of body surface area to provide an accurate and clinically useful determination of normal limits and to model changes in the cardiac dimensions and in their variance representing normal growth and development. The data fit well to the exponential growth model (r values 0.85 to 0.95). Variance about the central values also depended significantly on body size; that relation is represented effectively by a quadratic function of body surface area (r values 0.82 to 0.98). The model parameters allow calculation of normal limits at any desired level of confidence. Areas determined by hand planimetry have significantly greater variance compared with variance of linear dimensions, and also compared with variance of cross-sectional area using ellipses generated from the anteroposterior and mediolateral dimensions. This implies that either biologic variations in the amount of infolding or errors in freehand planimetry constitute a significant source of variance; this may be remedied by filtering out high frequency oscillations of contour. There is no significant difference in midnormal values and confidence limits for corresponding dimensions measured from orthogonal views. Furthermore, the anteroposterior and mediolateral dimensions of the left ventricle superimpose at each body size, consistent with circular cross section for normal subjects throughout growth and development. The data presented should comprise a useful set of reference standards for interpretation of cross-sectional echocardiograms.

Prevalence of mitral valve prolapse in normal children.

Although echocardiography has become the standard noninvasive method of diagnosing mitral valve prolapse, the diagnostic criteria have been established without clearly defining the range of normal patterns for mitral valve closure. The current study reports the analysis of mitral valve closure patterns in 193 children (aged 5 days to 18 years) making scheduled visits for well child care who were screened by history and physical examination to exclude structural heart disease. Mitral valve systolic leaflet position was analyzed for the appearance of any portion of either leaflet superior to the plane of the anulus. Superior systolic motion was noted in 13% of the overall study group; this pattern was uncommon in infants but more frequent in older children, with a prevalence of 35% in the 10 to 18 year age group. There was no statistically significant difference between male and female children at any age. The prevalence of superior systolic motion decreased markedly if consideration was given to its presence in more than one echocardiographic view (1%) or to displacement of the coaptation point of the mitral valve leaflets (0.5%). Superior systolic motion occurs with such frequency in normal children as to call into question the reliability of this pattern of mitral valve closure as a standard for the diagnosis of mitral valve prolapse. More restrictive diagnostic criteria which consider the degree of leaflet displacement or its presence in multiple echocardiographic views may be necessary to identify those subjects whose mitral valve closure patterns truly fall outside the range of normal.

Left atrial dimensions in growth and development: normal limits for two-dimensional echocardiography.

Reference values for normal left atrial dimensions have been based primarily on blind M-mode measurements, with no reports based on two-dimensional echocardiography to provide a comprehensive analysis of the two-dimensional measurements from infancy to old age. This report analyzes the left atrial dimensions from two-dimensional echocardiographic studies in 268 normal healthy subjects to determine normal limits and relations among linear, area and volume measurements of the left atrium. The group mean values change with body size, fitting well to the exponential growth model (r = 0.78 to 0.92). The variance about the mean (which determines normal limits) is represented effectively by a quadratic function of body surface area (r = 0.84 to 0.99). The variables determined by this modeling simplify evaluation of normal limits for any body size at any desired level of confidence, and the data are useful reference standards for interpretation of two-dimensional echocardiograms.

Pseudoaneurysm of the pulmonary artery after the banding procedure: two-dimensional echocardiographic description.

This report describes an infant with double-outlet right ventricle who underwent pulmonary artery banding as palliation for excessive left to right shunting through a ventricular septal defect. Three weeks after this procedure, there was abrupt clinical deterioration, and two-dimensional echocardiography clearly defined a large pseudoaneurysm arising from a breach in the posterior pulmonary artery wall, just proximal to the band. The diagnosis was confirmed at surgery, during which total correction was performed with successful outcome. The two-dimensional echocardiographic features of a pseudoaneurysm of the pulmonary artery are shown and the role of this noninvasive technique in the evaluation of pulmonary artery bands is discussed.

Doppler and two-dimensional echocardiographic diagnosis of Björk-Shiley prosthetic valve malfunction: importance of interventricular septal motion and the timing of onset of valve flow.

In a 69 year old woman with a "sticking" Björk-Shiley mitral prosthesis, the diagnosis was suggested by both the two-dimensional and the Doppler ultrasound examinations. In particular, the findings of early diastolic paradoxic septal motion, intermittent delayed opening of the prosthetic disc and variable timing of the onset of mitral valve inflow were believed to be diagnostic of a sticking tilting disc prosthesis.

Comparison of the echocardiographic and hemodynamic diagnosis of rheumatic tricuspid stenosis.

Two-dimensional echocardiography has proved to be reliable in the diagnosis of mitral, aortic and pulmonary stenosis. Its role in the diagnosis of rheumatic tricuspid stenosis is still being defined; therefore, the tricuspid valve echograms of 147 patients with rheumatic heart disease were examined. Thirty-eight of these patients also underwent hemodynamic evaluation. Tricuspid stenosis was defined echocardiographically as diastolic anterior leaflet doming, thickening and restricted excursion of the other two tricuspid leaflets and decreased separation of the leaflet tips. Using these criteria, the sensitivity and specificity of the echocardiogram in detecting tricuspid stenosis were 69 and 96%, respectively, in the group of 38 patients who had both echocardiographic and hemodynamic evaluations. However, when the smaller group of 17 patients who had simultaneous right atrial and right ventricular pressure recordings were considered separately, there was complete agreement between the echocardiographic and hemodynamic data. Thus, the two-dimensional echocardiogram is a sensitive and specific test for diagnosing rheumatic tricuspid stenosis. In addition, these data provided an opportunity to determine the prevalence of tricuspid stenosis in this group of patients with chronic rheumatic valvular disease. Tricuspid stenosis was present in 14 (9.5%) of the total group of 147 patients who had two-dimensional echocardiograms, and in 10 (26.3%) of the 38 who had both echocardiographic and hemodynamic studies. In patients with rheumatic heart disease about to undergo cardiac catheterization, an echocardiographic study should prove useful in making the diagnosis of tricuspid stenosis.

Closure of patent foramen ovale for paradoxical emboli: intermediate-term risk of recurrent neurological events following transcatheter device placement.

OBJECTIVES: We report the largest and the longest follow-up to date of patients who underwent transcatheter patent foramen ovale (PFO) closure for paradoxical embolism. BACKGROUND: Closure of a PFO has been proposed as an alternative to anticoagulation in patients with presumed paradoxical emboli. METHODS: Data were collected for patients following PFO closure with the Clamshell, CardioSEAL or Buttoned Devices at two institutions. RESULTS: There were 63 patients (46 +/- 18 years) with a follow-up of 2.6 +/- 2.4 years. Fifty-four (86%) had effective closure of the foramen ovale (trivial or no residual shunt by echocardiography) while seven (11%) had mild and two (3%) had moderate residual shunting. There were four deaths (leukemia, pulmonary embolism, sepsis following a hip fracture and lung cancer). There were four recurrent embolic neurological events following device placement: one stroke and three transient events. The stroke occurred in a 56-year-old patient six months following device placement. A follow-up transesophageal echocardiogram showed a well seated device without residual shunting. Two of the four events were associated with suboptimal device performance (one patient had a significant residual shunt and a second patient had a "friction lesion" in the left atrial wall associated with a displaced fractured device arm). The risk of recurrent stroke or transient neurological event following device placement was 3.2% per year for all patients. CONCLUSION: Transcatheter closure of PFO is an alternative therapy for paradoxical emboli in selected patients. Improved device performance may reduce the risk of recurrent neurological events. Further studies are needed to identify patients most likely to benefit from this intervention.

New perspectives in the assessment of cardiac chamber dimensions during development and adulthood.

The use of body surface area to assess the normalcy of cardiac dimensions has several limitations. To determine whether cardiac dimensions can be assessed by other indexes of body size and growth, this study evaluated the relations between cardiac dimensions assessed by two-dimensional echocardiography and age, height, weight and body surface area. The study group included 268 normal persons aged 6 days to 76 years of age. The dimensions examined included the aortic anulus, left atrium and left ventricular end-diastolic diameter, each measured in the parasternal long-axis plane, and left ventricular length measured from the apical two-chamber view. The analysis confirmed that the heart and great vessels grow in unison and at a predictable rate after birth, reaching 50% of their adult dimensions at birth, 75% by 5 years and 90% by 12 years. Although each cardiac dimension related linearly with height (aortic anulus, r = 0.96; left atrium, r = 0.91; left ventricular diameter, r = 0.94; left ventricular length, r = 0.93), the relations among age, weight and body surface area were best expressed by quadratic equations. Multiple regression confirmed that after adjustment for height, other indexes including age, gender, weight and body surface area had no independent effect on the prediction of each dimension. Therefore, because height is a nonderived variable that relates linearly with cardiac dimensions independent of age, it offers a simple yet accurate means of assessing the normalcy of cardiac dimensions in children and adults.

Three-dimensional echocardiography: in vivo validation for right ventricular free wall mass as an index of hypertrophy.

OBJECTIVES: This study tested the ability of three-dimensional echocardiography to reconstruct the right ventricular free wall and determine its mass in vivo using a system that automatically combines two-dimensional images with their spatial locations. BACKGROUND: Right ventricular free wall thickness is limited as an index of right ventricular hypertrophy because right ventricular mass may increase by dilation without increased thickness and because trabeculations and oblique views can exaggerate thickness in individual M-mode and two-dimensional scans. Three-dimensional echocardiography may have potential advantages because it can integrate the entire free wall mass, uninfluenced by oblique views or geometric assumptions. METHODS: The three-dimensional system was applied to 12 beating canine hearts to reconstruct the right ventricular free wall in intersecting views. The corresponding mass was compared with actual weights of the excised right ventricular free wall (15.5 to 78 g). For comparison, right ventricular sinus and outflow tract thickness were also measured by two-dimensional echocardiography, and the ability to predict mass from these values was determined. RESULTS: The three-dimensional algorithm successfully reproduced right ventricular free wall mass, which agreed well with actual values: y = 1.04x + 0.02, r = 0.985, SEE = 2.7 g (5.7% of the mean value). The two-dimensional predictions showed increased scatter: The variance of mass estimation, based on thickness, was 9.5 to 12.5 (average 11) times higher than the three-dimensional method (p < 0.02). CONCLUSIONS: Despite the irregular crescentic shape of the right ventricle, its free wall mass can be accurately measured by three-dimensional echocardiography in vivo, providing closer agreement with actual mass than predictions based on wall thickness. This method, with the increased efficiency of the three-dimensional system, can potentially improve our ability to evaluate the presence and progression of right ventricular hypertrophy.

A new echocardiographic model for quantifying three-dimensional endocardial surface area.

A new technique for quantitatively mapping the three-dimensional left ventricular endocardial surface was developed, using measurements from standard cross-sectional echocardiographic images. To validate the accuracy of this echocardiographic mapping technique in an animal model, the endocardial areas of 15 excised canine ventricles were calculated using measurements made from echocardiographic studies of the hearts and compared with areas determined with latex casts of the same ventricles. Close correlation (r = 0.87, p less than 0.001) between these two measures of endocardial area provided preliminary confirmation of the accuracy of the maps. To further characterize the mapping algorithm, it was translated into computer format and used to map the surfaces of idealized hemiellipsoids. Areas measured with this mapping technique closely approximated the actual areas of idealized surfaces with a wide spectrum of shapes; maps were particularly accurate for ellipsoids with shapes similar to those of undistorted human ventricles. Also, the accuracies of area calculations were relatively insensitive to deviation from the assumed positions of the echocardiographic short-axis planes. Finally, although the accuracy of the mapping technique improved as data from more transverse planes were added, the procedure proved reliable for estimating surface areas when data from only three planes were used. These studies confirm the accuracy of the echocardiographic mapping technique, and they suggest that the resulting planar plots might be useful as templates for localizing and quantifying the overall extent of abnormal wall motion.

Three-dimensional reconstruction of ventricular septal defects: validation studies and in vivo feasibility.

OBJECTIVES: The purpose of this study was to demonstrate the feasibility of in vivo three-dimensional reconstruction of ventricular septal defects and to validate its quantitative accuracy for defect localization in excised hearts (used to permit comparison of three-dimensional and direct measurements without cardiac contraction). BACKGROUND: Appreciating the three-dimensional spatial relations of ventricular septal defects could be useful in planning surgical and catheter approaches. Currently, however, echocardiography provides only two-dimensional views, requiring mental integration. A recently developed system automatically combines two-dimensional echocardiographic images with their spatial locations to produce a three-dimensional construct. METHODS: Surgically created ventricular septal defects of varying size and location were imaged and reconstructed, along with the left and right ventricles, in the beating heart of six dogs to demonstrate the in vivo feasibility of producing a coherent image of the defect that portrays its relation to surrounding structures. Two additional gel-filled excised hearts with defects were completely reconstructed. Quantitative localization of the defects relative to other structures (ventricular apexes and valve insertions) was then validated for seven defects in excised hearts. The right septal margins of the exposed defects were also traced and compared with their reconstructed areas and circumferences. RESULTS: The three-dimensional images provided coherent images and correct spatial appreciation of the defects (two inlet, two trabecular, one outlet and one membranous Gerbode in vivo; one inlet and one apical in excised hearts). The distances between defects and other structures in the excised hearts agreed well with direct measures (y = 1.05x-0.18, r = 0.98, SEE = 0.30 cm), as did reconstructed areas (y = 1.0x-0.23, r = 0.98, SEE = 0.21 cm2) and circumferences (y = 0.97x + 0.13, r = 0.97, SEE = 0.3 cm). CONCLUSIONS: Three-dimensional reconstruction of ventricular septal defects can be achieved in the beating heart and provides an accurate appreciation of defect size and location that could be of value in planning interventions.

Relationship between urinary dopamine production and natriuresis after acute intravascular volume expansion with sodium chloride in dogs.

The role of renal production of dopamine in mediating the natriuretic response to acute vascular volume expansion was investigated. The effect of infusion of 0.9% saline (30 ml/kg X h) over 2 h on urine excretion of sodium and catecholamines, as well as other hemodynamic and renal function parameters, was examined in seven dogs during control and carbidopa (1 mg/kg every 8 h for 24 h before saline infusion) treatment periods. Acute vascular volume expansion with saline resulted in a rise (P less than 0.01) in the renal excretion of dopamine and a depression (P less than 0.01) in renal excretion of norepinephrine which paralleled the natriuretic response to saline infusion. Epinephrine excretion was not altered by saline infusion. Carbidopa treatment was not associated with changes in left ventricular filling pressure, arterial blood pressure, glomerular filtration rate, renal blood flow, renal excretion of norepinephrine or epinephrine. However, carbidopa eliminated the increase in renal production of dopamine and markedly attenuated the natriuretic response to saline infusion. Since carbidopa blocks tissue conversion of dopa to dopamine, it appears that renal production of dopamine is an important mechanism mediating the natriuretic response to acute volume expansion.

The prevalence of mitral valve prolapse in patients with panic disorders.

Of 131 patients with panic disorder or agoraphobia, 44 (34%) had definite mitral valve prolapse on the basis of clinical findings of an apical mid- or late systolic click and a murmur and/or a two-dimensional echocardiogram showing prolapse of one leaflet in two views or of two leaflets in a single view. Seven other patients (5%) had a probable diagnosis of mitral prolapse on the basis of a typical murmur alone or an intermittent apical mid- or late systolic click and a prolapsing leaflet in a single echocardiographic view. This finding confirms previous reports of the association between panic disorder and mitral valve prolapse.

Myxoid leiomyosarcoma of the uterus. A report of six cases.

Six myxoid leiomyosarcomas of the uterus are described. Four were located within the myometrium and two lay predominantly within the broad ligament but appeared to have arisen in the outer myometrium. These tumors were characterized grossly by a gelatinous appearance and an apparently circumscribed border. On microscopical examination, however, they were observed to invade adjacent tissue and in two cases the lumens of veins. All the tumors contained copious amounts of myxomatous stroma. Although it was difficult to establish the nature of the neoplastic cells in some areas of these tumors, both light- and electron-microscopical examination showed characteristic features of smooth muscle cells in other areas. The mitotic count varied from 0 to 2/10 high-power fields. Follow-up investigation for periods ranging from 1 to 11 years revealed that four patients died and two are living with disease.