Semiquantitative 123I-Metaiodobenzylguanidine Scintigraphy to Distinguish Pheochromocytoma and Paraganglioma from Physiologic Adrenal Uptake and Its Correlation with Genotype-Dependent Expression of Catecholamine Transporters.

UNLABELLED: (123)I-metaiodobenzylguanidine ((123)I-MIBG) scintigraphy plays an important role in the diagnostic evaluation of patients with pheochromocytoma and paraganglioma (PPGL). (123)I-MIBG targets cell membrane and vesicular catecholamine transporters of chromaffin cells and facilitates localization of the primary tumor and metastatic lesions. Its specificity for the diagnosis of adrenomedullary chromaffin cell tumors can be jeopardized by physiologic uptake by the normal adrenal medulla. The aim of this study was to distinguish between PPGLs and normal adrenal glands by evaluating semiquantitative (123)I-MIBG uptake and to examine genotype-specific differences in correlation with expression of catecholamine transporter systems. METHODS: Sixty-two PPGLs collected from 57 patients with hereditary mutations in SDHA (n = 1), SDHB (n = 2), and SDHD (n = 4) (SDH is succinate dehydrogenase); von Hippel-Lindau (VHL; n = 2); RET (n = 12); neurofibromin 1 (NF1; n = 2); and MYC-associated factor X (MAX; n = 1), and with sporadic PPGLs (n = 33) were investigated. Preoperative planar and SPECT images were semiquantitatively analyzed using uptake measurements. Tumor-to-liver and normal adrenal-to-liver ratios were calculated and correlated with clinical characteristics including genotype, tumor size, and plasma metanephrines concentrations. The expression of norepinephrine transporter (NET) and vesicular monoamine transporter (VMAT-1) was evaluated immunohistochemically in paraffin-embedded tumor tissues. RESULTS: Mean tumor-to-liver ratios of PPGL lesions were significantly higher than normal adrenal-to-liver ratios (P < 0.001). Cutoff values to distinguish between physiologic and pathologic adrenal uptake were established at 0.7 (100% sensitivity, 10.3% specificity) and 4.3 (100% specificity, 66.1% sensitivity). No statistically significant differences in (123)I-MIBG uptake were found across PPGLs of different genotypes. Mean NET expression in hereditary cluster 2 (RET, NF1, MAX) and apparently sporadic tumors was significantly higher than for hereditary cluster 1 (SDHx, VHL) PPGLs (P = 0.011 and 0.006, respectively). Mean VMAT-1 expression in hereditary cluster 1 PPGLs was significantly higher than for cluster 2 tumors (P = 0.010). (123)I-MIBG uptake significantly correlated with maximum tumor diameter (P = 0.002). (123)I-MIBG uptake, however, did not correlate with either NET or VMAT-1 expression. CONCLUSION: Liver-normalized semiquantitative (123)I-MIBG uptake may be helpful to distinguish between pheochromocytoma and physiologic adrenal uptake. Genotype-specific differences in the expression of NET and VMAT-1 do not translate into differences in (123)I-MIBG uptake.

Influence of agonist induced internalization on [3H]Ro15-4513 binding-an application to imaging fluctuations in endogenous GABA with positron emission tomography.

Low affinity α1/α2 containing GABAA receptors are significantly less able to bind [(11) C]/[(3) H]Ro15-4513 following translocation to the endosomal environment. The alterations in [(11) C]Ro15-4513 binding observed in vivo following perturbations in endogenous GABA are likely driven by both alterations in receptor binding parameters following agonist induced internalisation and the GABA Shift.

Reduced striatal adenosine A2A receptor levels define a molecular subgroup in schizophrenia.

Schizophrenia (SZ) is a mental disorder of unknown origin. Some scientific evidence seems to indicate that SZ is not a single disease entity, since there are patient groups with clear symptomatic, course and biomarker differences. SZ is characterized by a hyperdopaminergic state related to high dopamine D2 receptor activity. It has also been proposed that there is a hypoadenosynergic state. Adenosine is a nucleoside widely distributed in the organism with neuromodulative and neuroprotective activity in the central nervous system. In the brain, the most abundant adenosine receptors are A1R and A2AR. In the present report, we characterize the presence of both receptors in human postmortem putamens of patients suffering SZ with real time TaqMan PCR, western blotting and radioligand binding assay. We show that A1R levels remain unchanged with respect to age-matched controls, whereas nearly fifty percent of patients have reduced A2AR, at the transcriptional and translational levels. Moreover, we describe how DNA methylation plays a role in the pathological A2AR levels with the bisulfite-sequencing technique. In fact, an increase in 5-methylcytosine percentage in the 5' UTR region of ADORA2A was found in those SZ patients with reduced A2AR levels. Interestingly, there was a relationship between the A2A/ß-actin ratio and motor disturbances as assessed with some items of the PANSS, AIMS and SAS scales. Therefore, there may be a subgroup of SZ patients with reduced striatal A2AR levels accompanied by an altered motor phenotype.

Pentazocine inhibits norepinephrine transporter function by reducing its surface expression in bovine adrenal medullary cells.

(±)-Pentazocine (PTZ), a non-narcotic analgesic, is used for the clinical management of moderate to severe pain. To study the effect of PTZ on the descending noradrenergic inhibitory system, in the present study we examined the effect of [(3)H]norepinephrine (NE) uptake by cultured bovine adrenal medullary cells and human neuroblastoma SK-N-SH cells. (-)-PTZ and (+)-PTZ inhibited [(3)H]NE uptake by adrenal medullary cells in a concentration-dependent (3-100 µM) manner. Eadie-Hofstee analysis of [(3)H]NE uptake showed that both PTZs caused a significant decrease in the V(max) with little change in the apparent K(m), suggesting non-competitive inhibition. Nor-Binaltorphimine and BD-1047, κ-opioid and σ-receptor antagonists, respectively, did not affect the inhibition of [(3)H]NE uptake induced by (-)-PTZ and (+)-PTZ, respectively. PTZs suppressed specific [(3)H]nisoxetine binding to intact SK-N-SH cells, but not directly to the plasma membranes isolated from the bovine adrenal medulla. Scatchard analysis of [(3)H]nisoxetine binding to SK-N-SH cells revealed that PTZs reduced the B(max) without changing the apparent K(d). Western blot analysis showed a decrease in biotinylated cell-surface NE transporter (NET) expression after the treatment with (-)-PTZ. These findings suggest that PTZ inhibits the NET function by reducing the amount of NET in the cell surface membranes through an opioid and σ-receptor-independent pathway.

Unbinding of targeted ultrasound contrast agent microbubbles by secondary acoustic forces.

Targeted molecular imaging with ultrasound contrast agent microbubbles is achieved by incorporating targeting ligands on the bubble coating and allows for specific imaging of tissues affected by diseases. Improved understanding of the interplay between the acoustic forces acting on the bubbles during insonation with ultrasound and other forces (e.g. shear due to blood flow, binding of targeting ligands to receptors on cell membranes) can help improve the efficacy of this technique. This work focuses on the effects of the secondary acoustic radiation force, which causes bubbles to attract each other and may affect the adhesion of targeted bubbles. First, we examine the translational dynamics of ultrasound contrast agent microbubbles in contact with (but not adherent to) a semi-rigid membrane due to the secondary acoustic radiation force. An equation of motion that effectively accounts for the proximity of the membrane is developed, and the predictions of the model are compared with experimental data extracted from optical recordings at 15 million frames per second. A time-averaged model is also proposed and validated. In the second part of the paper, initial results on the translation due to the secondary acoustic radiation force of targeted, adherent bubbles are presented. Adherent bubbles are also found to move due to secondary acoustic radiation force, and a restoring force is observed that brings them back to their initial positions. For increasing magnitude of the secondary acoustic radiation force, a threshold is reached above which the adhesion of targeted microbubbles is disrupted. This points to the fact that secondary acoustic radiation forces can cause adherent bubbles to detach and alter the spatial distribution of targeted contrast agents bound to tissues during activation with ultrasound. While the details of the rupture of intermolecular bonds remain elusive, this work motivates the use of the secondary acoustic radiation force to measure the strength of adhesion of targeted microbubbles.

Imaging and measuring the rituximab-induced changes of mechanical properties in B-lymphoma cells using atomic force microscopy.

The topography and mechanical properties of single B-lymphoma cells have been investigated by atomic force microscopy (AFM). With the assistance of microfabricated patterned pillars, the surface topography and ultrastructure of single living B-lymphoma cell were visualized by AFM. The apoptosis of B-lymphoma cells induced by rituximab alone was observed by acridine orange/ethidium bromide (AO/EB) double fluorescent staining. The rituximab-induced changes of mechanical properties in B-lymphoma cells were measured dynamically and the results showed that B-lymphoma cells became dramatically softer after incubation with rituximab. These results can improve our understanding of rituximab'effect and will facilitate the further investigation of the underlying mechanisms.

D2 dopamine receptor internalization prolongs the decrease of radioligand binding after amphetamine: a PET study in a receptor internalization-deficient mouse model.

Dopamine released by amphetamine decreases the in vivo binding of PET radioligands to the dopamine D(2) receptor. Although concentrations of extracellular dopamine largely return to baseline within 1 to 2 h after amphetamine treatment, radioligand binding remains decreased for several hours. The purpose of this study was to determine whether the prolonged decrease of radioligand binding after amphetamine administration is caused by receptor internalization. To distinguish dopamine displacement from receptor internalization, we used wild-type and arrestin3 (arr3) knockout mice, which are incapable of internalizing D(2) receptors. In addition, we used both the D(2) selective agonist [(11)C]MNPA (which is thought to bind to the high affinity state of the receptor) and the D(2) selective antagonist [(18)F]fallypride (which does not differentiate between high and low affinity state). After an initial baseline scan, animals were divided in three groups for a second scan: either 30 min or 4 h after amphetamine administration (3 mg/kg, i.p.) or as retest. At 30 min, [(11)C]MNPA showed greater displacement than [(18)F]fallypride, but each radioligand gave similar displacement in knockout and wild-type mice. At 4 h, the binding of both radioligands returned to baseline in arr3 knockout mice, but remained decreased in wild-type mice. Radioligand binding was unaltered on retest scanning. Our results suggest that the prolonged decrease of radioligand binding after amphetamine is mainly due to internalization of the D(2) receptor rather than dopamine displacement. In addition, this study demonstrates the utility of small animal PET to study receptor trafficking in vivo in genetically modified mice.

Application of therapeutic insonation to malignant glioma cells and facilitation by echo-contrast microbubbles of levovist.

BACKGROUND: Malignancies affecting the central nervous system are intractable to conventional therapies thereby requiring an alternative strategy, such as ultrasound irradiation. MATERIALS AND METHODS: We originally designed a transducer for intracranial insonation and investigated the effect of 210.4 kHz ultrasound on malignant glioma cells. RESULTS: The insonation of 2.61 W/cm2 effectively disrupted the malignant cells. This effect was reinforced by the echo-contrast agent, Levovist. The condition was applied to tumor-bearing animals and external insonation inhibited subcutaneous tumor growth. It also repressed the growth of intracranially implanted tumors and prolonged survival of the animals. When Levovist was stereotactically injected into the tumors, the effect of insonation was significantly enhanced. CONCLUSION: A neuronavigation system or stereotactic device has been used commonly for patients with brain tumor. Administration of combination therapy consisting of insonation and a local echo-contrast agent will have a role in improving the treatment for malignant gliomas.

Membrane damage effect of therapeutic ultrasound on Ehrlich ascitic tumor cells.

PURPOSE: The biologic effects and the underlying mechanisms of Ehrlich ascitic tumor (EAT) cells induced by ultrasound were investigated in this study. METHODS: Cells were subjected to ultrasonic irradiation with a frequency of 2.17 MHz and an intensity of 3 W/cm(2) for variable periods of time. Trypan blue exclusion was used to detect the integrity of cellular membrane; the membrane permeability was investigated by the incorporation of fluorescein isothiocyanate dextran during ultrasound exposure; and the cell membrane ultrastructure changes were observed under a scanning electron microscope. The potential mechanism was estimated from the generation of hydroxyl radicals, the lipid peroxidation levels, and intracellular reactive oxygen radicals production. RESULTS: The cell membrane damage effects induced by ultrasound increased with a prolonged exposure time; the fluorescent rates of the cells irradiated with ultrasound for 30 and 60 seconds were 11.46% and 18.50%, respectively; the amount of hydroxyl radicals in 30 (26.10 U/mL) and 60 seconds (28.47 U/mL) were significantly enhanced, compared with the control group (24.44 U/mL); then, the level of lipid peroxidation was also changed from 0.27 to 0.54 (30 seconds) and 1.21 nmol/mL (60 seconds). CONCLUSIONS: Shear forces and free radicals produced by acoustic cavitation may play important roles in these actions.

Coiled sperm from infertile patients: characteristics, associated factors and biological implication.

BACKGROUND: There is no systematic study on coiled sperm in semen, although they are commonly observed. This work characterizes coiled sperm in infertile men to understand the clinical implications and investigate the possible cause by osmotic swelling. METHODS: Coiled sperm in semen from 439 infertile patients were quantified and their ultrastructure examined by electron microscopy. Hypo-osmotic swelling (HOS) and demembranation tests were performed to elucidate the nature of the coiling. RESULTS: Semen from patients contained overall 3% of sperm with head-in-coil (HIC) and 8% other coiled forms, with 12% of patients having 20% or more such sperm. The percentage of coiled sperm (but not HIC) was correlated with age (R = 0.26, P = 0.003) and the epididymal secretory marker neutral alpha-glucosidase (R = 0.16, P < 0.001), and associated with heavy smoking and varicocele. Electron microscopy revealed coiling of tail filaments within the plasma membrane, resembling HOS. Some seminal coiled sperm and most sperm freshly coiled upon HOS could be opened by demembranation, while those that could not be opened were probably fixed in position by oxidation, which occurred more frequently in patients than semen donors. CONCLUSIONS: Sperm coiling in semen is common and independent of sperm quantity or hormonal status. Whereas HIC may have a genetic background, other coiled forms may be associated with a hostile endogenous milieu in the epididymis that causes swelling.

Peritoneal membrane recruitment in rats: a micro-computerized tomography (muCT) study.

The peritoneal contact surface area (PCSA), which represents the area parameter in the mass transfer area coefficient (MTAC), is a crucial marker in the evaluation of peritoneal dialysis effectiveness. However, the capacity to recruit a larger PCSA has only been rarely demonstrated in vivo and, in most cases, changes in MTAC are interpreted as permeability changes and not as surface area variations. Here, we report the use of micro-computerized tomography (muCT) for the measurement of PCSA changes to various fill volumes. Using this three-dimensional imaging method, PCSA was measured in vivo in 26 healthy Wistar rats receiving intraperitoneally increasing fill volumes of peritoneal dialysis solutions: 5 mL (group 1, n = 8), 10 mL (group 2, n = 8) and 15 mL (group 3, n = 10) per 100 g of body weight. A non-ionic iodinated contrast agent was added to the dialysis solution in order to distinguish the intraperitoneal dialysis solutions from soft tissues. The normalized PCSA/weight ratio (cm(2)/g) increased with fill volume: 1.12 +/- 0.10 cm(2)/g (range 0.98-1.25) in group 1; 1.74 +/- 0.08 cm(2)/g (range 1.64-1.87) in group 2; 2.13 +/- 0.09 cm(2)/g(range 1.90-2.30) in group 3. With this muCT method, PCSA recruited in vivo with a 10 mL/100 g fill volume was in the range 94-107%) of ex vivo total peritoneal surface area (evPSA), as calculated with the Kuzlan's formula. With a 15 mL/100 g fill volume, the in vivo-measured PCSA, the exchange surface area, surpassed the evPSA (range 113-139%).

Antibacterial activity of cyclodextrins against Bacillus strains.

Growth of alkaliphilic Bacillus halodurans C-125 both on agar plates and in liquid culture was inhibited by methyl-beta-cyclodextrin (CD). Furthermore, resting cells of the strain were lysed by contact with methyl-beta-CD higher than 10 mM. alpha-CD also showed lysis activity against Bacillus and related strains. The activity was not observed with Gram-negative and Gram-positive bacteria except for Bacillus strains. Fluorescence staining and scanning electron microscopy of cells revealed that methyl-beta-CD disrupted cell membranes, and consequently, the cells were lysed. This is a novel physiological property of CDs.

Anatomical and clinical study on effects of sonography with pulse inversion and microbubble contrast in rabbit kidney.

Aim of the present study was to evaluate by transmission electron microscope (TEM) modifications in rabbit kidney-parenchyma after submission to ultrasound contrast agent (UCA) with Pulse Inversion Harmonic Imaging (PIHI). Seven inbred male albino rabbits were divided into three groups: 1) control group (n = 1 animal); 2) sonicated group (n = 3 animals); 3) sonicated group with UCA injection (CEUS) (n = 3 animals). The first group was not exposed to ultrasonography (US) and/or UCA. The second and third groups underwent baseline US and later to US with PIHI with a high mechanical index; in the third group UCA was simultaneously administered. Ultrastructural studies and image analysis were blindly performed on 50 samples (2mm3), including cortex and medulla, by two experienced pathologists with TEM. By TEM observations of the first and second groups showed no structural modifications of renal cortex and medulla. TEM observations of the third group showed ultrastructural changes of renal corpuscle, proximal and distal convoluted tubules and collecting tubules; further in the most of observed sections the filtration membrane had an alteration of typical trilaminar pattern and vacuolisation of glomerular endothelial cells with irregular edges. Therefore in rabbit kidney submitted to CEUS some ultrastructural modifications were observed.

Characterization of membrane potential-dependent uptake of the novel PET tracer 18F-fluorobenzyl triphenylphosphonium cation.

PURPOSE: Mitochondrial dysfunction has been attributed a critical role in the etiology and pathogenesis of numerous diseases, and is manifested by alterations of the organelle's membrane potential (Deltapsi(m)). This suggests that Deltapsi(m) measurement can be highly useful for diagnostic purposes. In the current study, we characterized the capability of the novel PET agent (18)F-fluorobenzyl triphenylphosphonium ((18)F-FBnTP) to assess Deltapsi(m), compared with the well-established voltage sensor (3)H-tetraphenylphosphonium ((3)H-TPP). METHODS: (18)F-FBnTP and (3)H-TPP uptake under conditions known to alter Deltapsi(m) and plasma membrane potential (Deltapsi(p)) was assayed in the H345 lung carcinoma cell line. (18)F-FBnTP biodistribution was assessed in CD1 mice using dynamic PET and ex vivo gamma well counting. RESULTS: (18)F-FBnTP and (3)H-TPP demonstrated similar uptake kinetics and plateau concentrations in H345 cells. Stepwise membrane depolarization resulted in a linear decrease in (18)F-FBnTP cellular uptake, with a slope (-0.58+/-0.06) and correlation coefficient (0.94+/-0.07) similar (p>0.17) to those measured for (3)H-TPP (-0.63+/-0.06 and 0.96+/-0.05, respectively). Selective collapse of Deltapsi(m) caused a substantial decrease in cellular uptake for (18)F-FBnTP (81.6+/-8.1%) and (3)H-TPP (85.4+/-6.7%), compared with control. Exposure to the proapoptotic staurosporine, known to collapse Deltapsi(m), resulted in a decrease of 68.7+/-10.1% and 71.5+/-8.4% in (18)F-FBnTP and (3)H-TPP cellular uptake, respectively. (18)F-FBnTP accumulated mainly in kidney, heart and liver. CONCLUSION: (18)F-FBnTP is a mitochondria-targeting PET radiopharmaceutical responsive to alterations in membrane potential with voltage-dependent performance similar to that of (3)H-TPP. (18)F-FBnTP is a promising new voltage sensor for detection of physiological and pathological processes associated with mitochondrial dysfunction, such as apoptosis, using PET.

In vitro and in vivo binding of neuroactive steroids to the sigma-1 receptor as measured with the positron emission tomography radioligand [18F]FPS.

Sigma-1 receptors are widely expressed in the mammalian brain and also in organs of the immune, endocrine and reproductive systems. Based on behavioral and pharmacological assessments, sigma-1 receptors are important in memory and cognitive processes, and are thought to be involved in specific psychiatric illnesses, including schizophrenia, depression, and drug addiction. It is thought that specific neuroactive steroids are endogenous ligands for these sites. In addition, several sigma-1 receptor binding steroids including progesterone, dihydroepiandrosterone (DHEA), and testosterone are being examined clinically for specific therapeutic purposes; however, their mechanisms of action have not been clearly defined. We previously described the high affinity sigma-1 receptor selective PET tracer [(18)F]FPS. This study examines the effect of neuroactive steroids on [(18)F]FPS binding in vitro and in vivo. Inhibition constants were determined in vitro for progesterone, testosterone, DHEA, estradiol, and estriol binding to the [(18)F]FPS labeled receptor. The affinity order (K(i) values) for these steroids ranged from 36 nM for progesterone to >10,000 nM for estrodiol and estriol. Biodistribution studies revealed that i.v. coadministration of progesterone (10 mg/kg), testosterone (20 mg/kg), or DHEA (20 mg/kg) significantly decreased [(18)F]FPS uptake (%ID/g) by up to 50% in nearly all of eight brain regions examined. [(18)F]FPS uptake in several peripheral organs that express sigma-1 receptors (heart, spleen, muscle, lung) was also reduced (54-85%). These studies clearly demonstrate that exogenously administered steroids can occupy sigma-1 receptors in vivo, and that [(18)F]FPS may provide an effective tool for monitoring sigma-1 receptor occupancy of specific therapeutic steroids during clinical trials.

Functional differences between NPFF1 and NPFF2 receptor coupling: high intrinsic activities of RFamide-related peptides on stimulation of [35S]GTPgammaS binding.

By using an optimized [(35)S]GTPgammaS binding assay, the functional activities (potency and efficacy) of peptides belonging to three members of the RFamide family; Neuropeptide FF (NPFF), prolactin-releasing peptide (PrRP) and 26RFamide, were investigated on NPFF(1) and NPFF(2) receptors stably expressed in Chinese Hamster Ovary (CHO) cells. Despite their large differences in affinity and selectivity, all analogues tested behaved as agonists toward NPFF(1) and NPFF(2) receptors. High NaCl concentration in the assay strongly increased the efficacy toward NPFF(2) receptors and augmented differences among agonists. In low sodium conditions, whereas the potencies of agonists correlated with their affinities for NPFF(1) receptors, NPFF(2) receptors exhibited an extraordinary activity since all compounds tested displayed EC(50) values of GTPgammaS binding lower than their K(I) values. Comparisons of functional values between NPFF(1) and NPFF(2) receptors revealed unexpected potent selective NPFF(2) agonists especially for the PLRFamide and the VGRFamide sequences. By using blocker peptides, we also show that Galpha(i3) and Galpha(s) are the main transducers of NPFF(1) receptors while NPFF(2) are probably coupled with Galpha(i2), Galpha(i3), Galpha(o) and Galpha(s) proteins. Our data indicate that NPPF(1) and NPFF(2) receptors are differently coupled to G proteins in CHO cells.

Redistribution of nucleoside transporters to the cell membrane provides a novel approach for imaging thymidylate synthase inhibition by positron emission tomography.

Thymidylate synthase (EC 2.1.1.45) is a key enzyme for the de novo synthesis of DNA and as such a target for anticancer drug development. There is a need to develop noninvasive methods for assessing thymidylate synthase inhibition in tumors. The aim of this study was to assess the potential of 3'-deoxy-3'-[(18)F]fluorothymidine ([(18)F]FLT) positron emission tomography (PET) for early measurement of thymidylate synthase inhibition and to elucidate the cellular mechanisms involved. Radiation-induced fibrosarcoma-1 tumor-bearing mice were injected with a single i.p. dose of the thymidylate synthase inhibitor 5-fluorouracil (5-FU; 165 mg/kg) and imaged by [(18)F]FLT-PET at 1 to 2 hours after treatment. Deoxyuridine, thymidine kinase 1 (cytoplasmic thymidine kinase; EC2.7.1.21), and ATP levels in excised tumors were measured. Cellular assays for membrane transport were also done. There was a 1.8-fold increase in the 60-minute [(18)F]FLT tumor/heart radioactivity ratio in drug-treated mice compared with vehicle controls (P = 0.0016). Plasma and tumor deoxyuridine levels increased significantly but thymidine kinase and ATP levels were unchanged. Whole-cell assays implicated a (low level) functional role for the type-1 equilibrative nucleoside transporter (ENT). There was an increase in type-1 ENT-binding sites per cell from 49,110 in untreated cells to 73,142 (P = 0.03) in cells treated with 10 microg/mL 5-FU for 2 hours, without a change in transporter affinity (P = 0.41). We conclude that [(18)F]FLT-PET can be used to measure thymidylate synthase inhibition as early as 1 to 2 hours after treatment with 5-FU by a mechanism involving redistribution of nucleoside transporters to the plasma membrane.

Membrane blebbing in cancer cells treated with various apoptotic inducers.

The dynamics of cell morphology, in particular membrane blebbing, was studied after induction of apoptosis by etoposide or camptothecin in four human stabilized cell lines (Hep2, Bowes, HT-29 and HL-60). Time lapse videomicroscopy and F-actin staining revealed various dynamic parameters of this process including its duration, maximal extent, stages and final endpoints in individual cells. Although generally occurring in predictable patterns, our results indicate a relatively significant variability both in appearance and temporal organization of blebbing not only between different cell lines but also within them.