N-(4-hydroxyphenyl)retinamide increases ceramide and is cytotoxic to acute lymphoblastic leukemia cell lines, but not to non-malignant lymphocytes.

The retinoid, N-(4-hydroxyphenyl)retinamide (4-HPR), mediates p53-independent cytotoxicity and can increase reactive oxygen species and ceramide in solid tumor cell lines. We determined changes in ceramide and cytotoxicity upon treatment with 4-HPR (3-12 microM) in six human acute lymphoblastic leukemia (ALL) cell lines: T cell (MOLT-3, MOLT-4, CEM), pre-B-cell (NALM-6, SMS-SB), and null cell (NALL-1). Exposure to 4-HPR (12 microM) for 96 h caused 4.7 (MOLT-3), 3.5 (MOLT-4), 3.9 (CEM), 2.9 (NALM-6), 4.7 (SMS-SB), AND 4.5 (NALL-1) logs of cell kill. The average 4-HPR concentration that killed 99% of cells (LC(99)) for all six lines was 4.8 microM (range: 1.5-8.9 microM). Treatment with 4-HPR (9 microM) for 24 h resulted in an 8.9 +/- 1.0-fold (range: 4.9-15.7-fold) increase of ceramide. Ceramide increase was time- and dose-dependent and abrogated by inhibitors of de novo ceramide synthesis. Concurrent inhibition of ceramide glycosylation/acylation by d,l-threo-(1-phenyl-2-hexadecanoylamino-3-morpholino-1-propanol) (PPMP) further increased ceramide levels, and synergistically increased 4-HPR cytotoxicity in four of six ALL cell lines. 4-HPR was minimally cytotoxic to peripheral blood mononuclear cells and a lymphoblastoid cell line, and increased ceramide <2-fold. Thus, 4-HPR was cytotoxic and increased ceramide in ALL cell lines, but not in non-malignant lymphoid cell types.

N-(4-hydroxyphenyl)retinamide elevates ceramide in neuroblastoma cell lines by coordinate activation of serine palmitoyltransferase and ceramide synthase.

The retinoid N-(4-hydroxyphenyl)retinamide (4-HPR; fenretinide) is cytotoxic to a variety of cancer cell lines, and we previously showed an association between ceramide generation and 4-HPR cytotoxicity for neuroblastoma cell lines (B. J. Maurer et al., J. Natl. Cancer Inst. (Bethesda), 91: 1138-1146, 1999). Here we determine whether the increased ceramide mediated by 4-HPR in the CHLA-90 human neuroblastoma cell line results from de novo ceramide synthesis. Treatment of CHLA-90 with 4-HPR for 2 h, in the presence of [(3)H]palmitic acid, caused sequential formation of [(3)H]sphinganine (220% over control) and [(3)H]ceramide (160% over control), with sphinganine returning to baseline at 4 h, and ceramide continuing to increase (215% over control). 4-HPR treatment did not accelerate cellular decay of sphingomyelin. Preincubation of cells with either L-cycloserine, an inhibitor of serine palmitoyltransferase (SPT), or fumonisin B(1), an inhibitor of ceramide synthase, retarded ceramide formation in response to 4-HPR treatment, although sphinganine was still generated when 4-HPR and FB(1) were present. Data from in vitro enzyme assays using microsomes showed that preexposure of intact cells to 4-HPR resulted in a time (175% over control; 6 h)- and dose-dependent increase (173% over control; 10 microM) in SPT activity as well as a time (265% over control)- and dose-dependent increase (215% above control; 10 microM) in ceramide synthase activity. Our results show that 4-HPR-mediated ceramide generation is derived from the de novo synthetic pathway by coordinate activation of SPT and ceramide synthase. Knowledge of these biochemical events is of utility when downstream modulators of ceramide metabolism are used to heighten the cytotoxic response to chemotherapy.

Synergistic cytotoxicity in solid tumor cell lines between N-(4-hydroxyphenyl)retinamide and modulators of ceramide metabolism.

BACKGROUND: We previously reported that N-(4-hydroxyphenyl)retinamide (4-HPR, fenretinide) treatment caused large increases of ceramide levels in neuroblastoma cell lines and induced cell death by a combination of apoptosis and necrosis through p53 (also known as TP53)-independent and caspase-independent pathways. Our goal was to determine if several molecules that inhibit enzymes involved in ceramide metabolism-L-threo-dihydrosphingosine (safingol), d, l-threo-1-phenyl-2-hexadecanoylamino-3-morpholino-1-propanol (PPMP), and tamoxifen-enhanced 4-HPR-mediated cytotoxicity and/or affected ceramide levels. METHODS: Cellular lipids were quantified by radiolabeling and thin-layer chromatography. Cytotoxicity and cytotoxic synergy (expressed as combination index, where combination index <1 indicates synergy and >1 indicates antagonism) were measured in cultured cancer cell lines with the use of a fluorescence-based assay of cell viability employing digital imaging microscopy. Statistical tests were two-sided. RESULTS: 4-HPR increased ceramide levels by de novo synthesis. Safingol (1-4 microM) was incorporated into a stereochemical variant of ceramide and synergized with a 3:1 molar ratio of 4-HPR (3-12 microM), to produce a 100-fold to 10 000-fold (2 to 4 logs) increase in cytotoxicity relative to 4-HPR alone in neuroblastoma (combination index <0.1), lung (combination index <0.1-0.2), melanoma (combination index <0.1-0.2), prostate (combination index <0.1-1.0), colon (combination index 0.1-0.3), breast (combination index = 0.1-0.5), and pancreas (combination index = 0.2) cell lines, including p53 mutant and alkylator-resistant cell lines. The 4-HPR and safingol combination was cytotoxic in low-oxygen conditions and was minimally toxic to normal fibroblasts and bone marrow myeloid progenitor cells. Addition of agents that retard ceramide glucosylation and/or acylation, such as PPMP or tamoxifen, to 4-HPR or to the combination of 4-HPR and safingol further increased cytotoxicity to tumor cells. CONCLUSIONS: Combinations of 4-HPR and modulators of ceramide metabolism may form the basis for a novel chemotherapy that is functional under hypoxic conditions (e.g., such as those within tumors) and is p53 independent and caspase independent.

Retinoic-acid-resistant neuroblastoma cell lines show altered MYC regulation and high sensitivity to fenretinide.

BACKGROUND: High-dose, pulse-13-cis-retinoic acid (13-cis-RA) given after intensive cytotoxic therapy improves event-free survival for high-risk neuroblastoma (NB), but more than 50% of patients have tumor recurrence. PROCEDURE: We conducted multistep selection for resistance to all-trans-retinoic acid (ATRA) in NB cell lines with (SMS-KCNR and LA-N-5) or without (SMS-LHN) MYCN genomic amplification. RESULTS: After 12 exposures to 10 microM ATRA, the two MYCN-amplified cell lines (KCNR 12X RR and LA-N-5 12X RR) showed partial resistance to the cytostatic/differentiation effects of ATRA; complete resistance was seen in LHN 12X RR. ATRA-selected cells showed general RA resistance (cross-resistance to 13-cis-RA). Transient (KCNR 12 X RR, LA-N-5 12X RR) or sustained (LHN 12X RR) novel overexpression of c-myc was associated with RA resistance. RA-insensitive overexpression of MYCN by transduction in SMS-LHN also conferred RA resistance. Both parental and RA-resistant lines showed 2-4 logs of cell kill in response to N-(4-hydroxyphenyl)retinamide (4- HPR, fenretinide). Compared to parental lines, 4-HPR achieved 1-3 log greater cell kills in RA-resistant LHN 12X RR, LA-N-5 12X RR, KCNR 12X RR, and MYCN-transduced SMS-LHN or SK-N-RA. NB cell lines (n = 26) from 21 different patients showed that 16 of 26 (62%) were sensitive to 4-HPR (LC(90) < 10 microM), including lines established at relapse after myeloablative and/or 13-cis-RA therapy. CONCLUSION: Thus, RA-resistant NB cell lines can be sensitive (and in some cases collaterally hypersensitive) to 4-HPR.

Increase of ceramide and induction of mixed apoptosis/necrosis by N-(4-hydroxyphenyl)- retinamide in neuroblastoma cell lines.

BACKGROUND: The synthetic retinoid N-(4-hydroxyphenyl)retinamide (4-HPR or fenretinide) is toxic to myeloid leukemia and cervical carcinoma cell lines, probably in part due to its ability to increase levels of reactive oxygen species (ROS). We have studied the effects of 4-HPR on neuroblastoma cell lines. Since neuroblastomas commonly relapse in bone marrow, a hypoxic tissue compartment, and many chemotherapeutic agents are antagonized by hypoxia, our purpose was to study in these cell lines several factors influencing 4-HPR-induced cytotoxicity, including induced levels of ROS, effects of physiologic hypoxia and antioxidants, levels of ceramide, and the mechanism of cell death. METHODS: ROS generation was measured by carboxydichlorofluorescein diacetate fluoresence. Ceramide was quantified by radiolabeling and thin-layer chromatography. Immunoblotting was used to assess p53 protein levels. Apoptosis (programmed cell death) and necrosis were analyzed by nuclear morphology and internucleosomal DNA fragmentation patterns. Cytotoxicity was measured by a fluorescence-based assay employing digital imaging microscopy in the presence or absence of the pancaspase enzyme inhibitor BOC-d-fmk. Statistical tests were two-sided. RESULTS/CONCLUSIONS: In addition to increasing ROS, 4-HPR (2.5-10 microM) statistically significantly increased the level of intracellular ceramide (up to approximately 10-fold; P<.001) in a dose-dependent manner in two neuroblastoma cell lines, one of which is highly resistant to alkylating agents and to etoposide. Cell death induced by 4-HPR was reduced but not abrogated by hypoxia in the presence or absence of an antioxidant, N-acetyl-L-cysteine. Expression of p53 protein was not affected by 4-HPR. Furthermore, the pan-caspase enzyme inhibitor BOC-d-fmk prevented apoptosis, but not necrosis, and only partially decreased cytotoxicity induced by 4-HPR, indicating that 4-HPR induced both apoptosis and necrosis in neuroblastoma cells. IMPLICATIONS: 4-HPR may form the basis for a novel, p53-independent chemotherapy that operates through increased intracellular levels of ceramide and that retains cytotoxicity under reduced oxygen conditions.

Growth of human tumor cells in macroporous microcarriers results in p53-independent, decreased cisplatin sensitivity relative to monolayers.

Multicellular contact has been shown to influence the in vitro sensitivity of cells to drug treatment. We investigated the use of macroporous gelatin microcarriers, CultiSpher-G, as a convenient laboratory system for the molecular analysis of this "contact effect". We determined that human A549 cells can be grown in CultiSphers with growth and cell cycle parameters similar to those of monolayers. In addition, cells in CultiSphers express less p27/kip1, an indicator of cell cycle arrest, than equivalent cells in monolayers. When treated with drugs, A549 cells grown in CultiSphers or monolayers accumulate equivalent amounts of platinum-DNA adducts and similar amounts of doxorubicin. Moreover, A549 and KB-3-1 cells in CultiSphers have significantly decreased sensitivity to cis-platinum(II)diammine dichloride (cisplatin), 4-hydroperoxycyclophosphamide, doxorubicin, and paclitaxel (taxol) compared with cells in monolayers when assayed by clonogenic survival. Cisplatin treatment in monolayers or CultiSphers did not result in apoptotic cell death. In contrast, paclitaxel caused a significant amount of sub-G1 DNA, an indicator of apoptosis, which was diminished when cells were grown in CultiSphers compared with monolayers. When grown in CultiSphers, cells with abrogated p53 function (A549/16E6 and NCI-H1299) were less sensitive to cisplatin than the corresponding monolayer cells, indicating that the decrease in sensitivity is p53 independent. Taken together, the data suggest that CultiSpher-G microcarriers are a useful in vitro system to examine the effects of three-dimensional cell contact on drug sensitivity of human tumor cells.

Growth and radiation response of cells grown in macroporous gelatin microcarriers (CultiSpher-G).

Four cell lines have been cultured in macroporous gelatin microcarrier beads (CultiSpher-G) to test this as a new model of three-dimensional cell growth for use in experimental cancer therapy. A549, KB 3-1, KB 8-5 and V79 cells were successfully grown in CultiSphers, albeit with longer doubling times than observed for the respective cell type in monolayer cultures. MTT staining and histology demonstrated three-dimensional contact of cells in the microcarriers. A549 cells populated the microcarriers more densely than KB 3-1 cells, and with both cell types there is bead-to-bead variation in occupancy by cells. [3H]TdR autoradiography reveals labelled cells throughout A549 and KB 3-1 CultiSphers, with no proliferation gradient from edge to centre. Central necrosis has not been observed. A549 cells but not KB 3-1 cells demonstrated a radiation contact effect (i.e. resistance) when irradiated in CultiSphers, compared with monolayers. We conclude that CultiSphers may be a useful model for three-dimensional growth and cell contact when investigators want to investigate these phenomena without the microenvironmental gradients of spheroids.

Plasma atrial natriuretic peptide is elevated in patients with hypertrophic cardiomyopathy.

STUDY OBJECTIVES: To determine if plasma levels of atrial natriuretic peptide are elevated in patients with hypertrophic cardiomyopathy and to determine the relationship of atrial natriuretic peptide to symptoms and echocardiographic indices of left ventricular structure and diastolic function in these patients. DESIGN: A prospective study in which atrial natriuretic peptide was measured in peripheral venous plasma in 14 patients (age 44 +/- 14 years) with hypertrophic cardiomyopathy and 17 healthy controls. Echocardiography was performed in all cases and 30 controls to examine indices of left heart structure and function. All patients underwent clinical evaluation. RESULTS: The concentration of atrial natriuretic peptide was significantly higher in patients with hypertrophic cardiomyopathy than controls, (17.86 +/- 8.72 vs. 6.22 +/- 3.26 pmol/l, P = 0.0001). Diastolic dysfunction was observed in 11 of 14 patients with hypertrophic cardiomyopathy. No correlation was demonstrated between atrial natriuretic peptide levels and the degree of diastolic dysfunction, septal or free wall thickness, left atrial size, degree of mitral regurgitation or New York Heart Association functional class. CONCLUSIONS: Plasma levels of atrial natriuretic peptide are elevated in patients with hypertrophic cardiomyopathy but do not correlate with symptoms or echocardiographically-derived indices of left ventricular structure or diastolic function.

Doppler echocardiographic detection of left ventricular diastolic dysfunction in patients with pulmonary sarcoidosis.

STUDY OBJECTIVE: To determine the prevalence of left ventricular diastolic dysfunction in patients with biopsy specimen-proved pulmonary sarcoidosis without clinical evidence of cardiac disease. DESIGN: A cross-sectional study. SETTING: A large tertiary care university teaching hospital. PATIENTS AND CONTROL SUBJECTS: Fifty consecutive subjects had biopsy specimen-proved pulmonary sarcoidosis without suspected cardiac involvement. Those with other conditions known to affect diastolic function were excluded. The control group comprised 30 healthy hospital workers. INTERVENTIONS: Clinical examination, 12-lead ECG, and combined echocardiographic/phonocardiographic examination. MEASUREMENTS: Indexes of left ventricular diastolic function, including isovolumic relaxation time, peak velocity of early (E) and late (A) ventricular filling, deceleration rate of early diastolic flow, and the sum of the time velocity integrals of E and A were obtained in each patient and control subject. Systolic function was determined using a modification of Simpson's rule. RESULTS: Diastolic dysfunction was present in 7 (14%) patients, 6 of whom had normal systolic function and normal two-dimensional echocardiographic examination. Those with diastolic dysfunction had a longer duration of illness (15 +/- 7 vs 6 +/- 5 years; p = 0.0004), were significantly older (52 +/- 11 vs 38 +/- 9 years; p = 0.0009), and had higher systolic BP (130 +/- 13 vs 117 +/- 12 mm Hg; p = 0.01) than the sarcoid patients with normal diastolic function. CONCLUSIONS: These results demonstrate a significant prevalence of left ventricular diastolic dysfunction in patients with pulmonary sarcoidosis. The cause of this abnormality may be a subclinical sarcoid cardiomyopathy.

Doppler echocardiographic evidence of left ventricular diastolic dysfunction in ankylosing spondylitis.

Although cardiac involvement in the form of conduction abnormalities or aortic regurgitation occurs in 5 to 10% of patients with ankylosing spondylitis, few studies have assessed left ventricular (LV) function. This study assesses the prevalence of both systolic and diastolic LV dysfunction and other cardiac abnormalities in patients with ankylosing spondylitis who have no clinical cardiac manifestations. Fifty-nine patients (49 men and 10 women, mean age 42 +/- 10 years) underwent full clinical examination, electrocardiography, 24-hour Holter monitoring and 2-dimensional, M-mode and Doppler echocardiography. Mean disease duration was 17 +/- 9 years (range 1 to 42). Seventeen patients had evidence of noncardiac extraarticular manifestations. Precordial examination was normal in all. An age- and sex-matched control group of 44 healthy subjects was also studied. On echocardiography, abnormal LV diastolic function was detected in 12 patients (20%). Prolonged isovolumic relaxation time, prolonged deceleration time, reduced rate of descent of flow velocity in early diastole (EF slope) and reversal of the early and late peak transmitral diastolic flow velocities (E/A ratio) were noted in 9 patients. In 3 patients there was an increased E/A ratio, reduced deceleration time and increased EF slope. Mild aortic regurgitation and mitral regurgitation was seen in 1 and 3 patients, respectively. No abnormalities of left atrial size, LV systolic or diastolic dimensions or wall thicknesses were noted. There was no correlation between the presence of LV diastolic dysfunction and age, disease severity, disease duration, or the presence of extraarticular manifestations.(ABSTRACT TRUNCATED AT 250 WORDS)

Core and peripheral temperature response to exercise in patients with impaired left ventricular function.

OBJECTIVE: Exercise induced hypotension is a specific but insensitive indicator of severe coronary artery disease. Skin blood flow is subject to control by baroreceptor mediated reflexes as well as thermoregulatory reflexes. Monitoring skin temperature or the skin to central temperature gradient may be a more sensitive indicator of impaired cardiac output response to exercise than hypotension. DESIGN AND PARTICIPANTS: Central and skin temperature changes associated with exercise were studied in 10 normal volunteers and eight patients with impaired resting ventricular function due to ischaemic heart disease. Patients exercised according to a modified Bruce protocol. The two sample independent t test was applied to compare the central and peripheral temperatures in the two groups at three minute intervals during exercise and at two minute intervals after exercise. RESULTS: A significant decrease was found in mean (1 SEM) central temperature on exercise in our patient group (98.2(0.2) degrees F to 97.2(0.3) degrees F), compared with the normal increase in central temperature (97.7(0.2) degrees F to 98.3(0.3) degrees F). Mean (1 SEM) skin temperature changes reflected the expected skin blood flow changes with exercise in normal subjects. In the patient group skin temperature declined during exercise (89.7(2.1) degrees F to 86.6(1.7) degrees F) and was significantly lower than normal from six minutes onwards. CONCLUSIONS: The abnormal peripheral temperature changes of patients with impaired resting ventricular function is an early and sensitive indicator of an abnormal haemodynamic response to exercise. It is possible that skin temperature measurement during exercise could help detect exercise induced ventricular dysfunction due to ischaemia or impaired cardiac output due to valvar heart disease.

Silent ischaemia in diabetic men with autonomic neuropathy.

Autonomic neuropathy is associated with an increased incidence of silent myocardial infarction and sudden death. The purpose of this study was to investigate the prevalence of silent myocardial ischaemia in diabetic patients with autonomic neuropathy and without. Five standard autonomic function tests were performed on 41 men with diabetes: postural change in blood pressure, postural change in heart rate, heart rate response to deep breathing, heart rate response to Valsalva's manoeuvre, and blood pressure response to sustained handgrip. There were 17 patients with autonomic neuropathy (group A) and 24 with normal autonomic function (group B). All patients underwent 24 hour ambulatory electrocardiographic monitoring to detect silent ischaemia. There was no significant difference in risk factors for coronary artery disease or history of angina pectoris between these groups. The prevalence of silent ischaemia was 64.7% in group A (95% confidence interval (95% CI) 38.33 to 85.79%) and 4.1% in group B (95% CI 0.11 to 21.12%). This represents a relative risk of 42.2 (95% CI 4.5 to 39.4, p less than 0.001). These results are consistent with the concept that autonomic neuropathy may prevent the development of anginal pain and thus obscure the presence of ischaemic heart disease. Twenty four hour ambulatory electrocardiographic monitoring may identify a subgroup of diabetic patients with autonomic neuropathy who have myocardial ischaemia and to whom treatment may be offered.

Cardiac catheterisation with 5 French catheters.

From the beginning of November 1987 to the end of January 1989, 526 coronary arteriograms and left ventricular angiograms were performed with 5 French coronary catheters. In 448 (85%) patients diagnostic pictures were obtained with three standard types of 5 French catheters (No 4 Judkins): that is, left coronary, right coronary, and pigtail catheters. In 60 patients (11.4%) various other 5 French catheters were required to complete the study. In nine patients (1.7%), a 7 or 8 French catheter was used. Major complications causing cardiac arrest or requiring urgent operation developed in five patients. Sixty two patients (11.77%) had minor complications that required sublingual nitrates or a single bolus of atropine, or developed a haematoma that did not need intervention or had a mild reaction to the contrast material. Complications of moderate severity developed in 17 patients (3.2%): severe chest pain, arrhythmia requiring a temporary pacemaker, contrast reaction associated with hypotension, haematoma requiring blood transfusion, or a transient ischaemic episode. There were no deaths. 5 French catheters were used for routine coronary angiography and left ventriculography in 98.3% of patients. There were no major complications related to femoral artery puncture. The routine use of 5 French coronary catheters should increase the feasibility of safe coronary angiography in outpatients and should reduce the cost of this investigation.

Acute and long-term hemodynamic response to low-dose enoximone in refractory heart failure.

Enoximone possesses both positive inotropic and vasodilatory properties. In heart failure, doses varying between 3 mg/kg and 6 mg/kg produce a beneficial acute hemodynamic response but have been associated with significant side effects. Little is known about the long-term hemodynamic efficacy of this agent. To assess whether a lower dose of enoximone could produce both acute and long-term hemodynamic benefits and be better tolerated, 15 patients with refractory heart failure were given enoximone 100 mg every 8 hours (mean dose, 1.7 mg/kg). The cardiac index, pulmonary capillary wedge pressure, pulmonary artery pressure, right atrial pressure, systemic vascular resistance, pulmonary vascular resistance, and stroke volume index all improved significantly during the first 24 hours. The systemic blood pressure and heart rate did not alter appreciably during this period. Five of six patients remaining on therapy at 6 months had a follow-up hemodynamic study. Sustained improvement was seen in the cardiac index, pulmonary capillary wedge pressure, and pulmonary artery pressure when compared to baseline (all p less than 0.05). A satisfactory trend, which did not reach statistical significance, was noted in the right atrial pressure (p = 0.09) and stroke volume index (p = 0.06). Diarrhea occurred in one patient. These findings indicate that enoximone has a beneficial acute and long-term hemodynamic effect at a low dose that is clinically well tolerated.

Familial aortic valve disease: evidence for a genetic influence?

It has been suggested that genetic factors may play a role in the aetiology of congenital bicuspid aortic valve disease. This report describes a family in which three male siblings with bicuspid aortic valves developed critical aortic stenosis. This provides further evidence that an hereditary mechanism may play a role in some patients with this condition.

Novel submicroscopic extrachromosomal elements containing amplified genes in human cells.

In previous work, several methotrexate (MTX)-resistant variants were isolated frm the human cell line HeLa BU25, which exhibited a high degree of dihydrofolate (DHFR) gene amplification (estimated to be 250- to 300-fold). These variants did not contain any chromosome with a homogeneously staining region (HSR) and exhibited only a small average number of minute chromosomes per cell: these two types of karyotypic abnormalities generally accompany selective gene amplification. We now report that structures containing amplified DHFR genes in one of these variants (HeLa BU25-10B3) can be isolated by pulsed-field gradient or field-inversion gel electrophoresis as homogeneous DNA molecules of approximately 650 kilobases (kb). Electron microscopy of metaphase spreads from these cells reveals chromatin fibres with a similar DNA content, which are probably related to the above elements. These represent a novel type of extrachromosomal structures in mammalian cells.

Percutaneous transluminal coronary angioplasty in unstable angina: comparison with stable angina.

Percutaneous transluminal coronary angioplasty was performed in 25 patients with unstable angina and in a similar group of 25 patients with stable angina. The frequency of single, double, and triple vessel disease was identical in each group. Technical success was achieved in 22 (81%) out of 27 attempts in those with unstable angina and in 14 (52%) out of 27 attempts in those with stable angina. Vessel occlusion occurred in nine patients, necessitating emergency bypass surgery in four. There was evidence of myocardial infarction in three patients in each group and one patient in the unstable group subsequently died. Twenty eight of 32 successfully treated patients were followed up by means of repeat coronary arteriography, exercise electrocardiography, and clinical assessment after a mean (SD) interval of 14 (7) months. There was angiographic evidence of restenosis in 32% (seven of 22) of lesions in the unstable group and 44% (four of nine) of lesions in the stable group. There were no late infarctions or deaths during the follow up period. These results support the growing evidence that angioplasty can be carried out safely and effectively in patients with unstable angina.