Transformation of indolent follicular lymphoma into diffuse large B-cell lymphoma - the molecular basis of "cancer aggressiveness".

BACKGROUND: Follicular lymphoma (FL) is the most common indolent non-Hodgkin's lymphoma in the Western world. It is an indolent disease in most patients, but about 20% of patients experience an early relapse after initial treatment, which is associated with shorter overall survival. A histological transformation into an aggressive lymphoma, most frequently diffuse large-cell B-lymphoma, represents another prognostically unfavorable event in the course of the disease. Thanks to recent genomic studies and mouse models, we are able to better understand the molecular nature of the FL onset and evolution of "aggressive" subclones of cells. Recently, deregulation of several molecular pathways associated with the histological transformation has also been described. PURPOSE: This review summarizes the complex molecular mechanisms responsible for FL onset, progression, aggressiveness, and transformation. We believe that the observations in FL have some general implications for understanding the mechanisms leading to the evolution of cancer "aggressiveness," such as divergent evolution, intraclonal variability and tumor plasticity.

Interferon restores replication fork stability and cell viability in BRCA-defective cells via ISG15.

DNA replication and repair defects or genotoxic treatments trigger interferon (IFN)-mediated inflammatory responses. However, whether and how IFN signaling in turn impacts the DNA replication process has remained elusive. Here we show that basal levels of the IFN-stimulated gene 15, ISG15, and its conjugation (ISGylation) are essential to protect nascent DNA from degradation. Moreover, IFNß treatment restores replication fork stability in BRCA1/2-deficient cells, which strictly depends on topoisomerase-1, and rescues lethality of BRCA2-deficient mouse embryonic stem cells. Although IFNß activates hundreds of genes, these effects are specifically mediated by ISG15 and ISGylation, as their inactivation suppresses the impact of IFNß on DNA replication. ISG15 depletion significantly reduces cell proliferation rates in human BRCA1-mutated triple-negative, whereas its upregulation results in increased resistance to the chemotherapeutic drug cisplatin in mouse BRCA2-deficient breast cancer cells, respectively. Accordingly, cells carrying BRCA1/2 defects consistently show increased ISG15 levels, which we propose as an in-built mechanism of drug resistance linked to BRCAness.

Real-time gastric motility monitoring using transcutaneous intraluminal impedance measurements (TIIM).

The stomach plays a critical role in digestion, processing ingested food mechanically and breaking it up into particles, which can be effectively and efficiently processed by the intestines. When the motility of the stomach is compromised, digestion is adversely affected. This can lead to a variety of disorders. Current diagnostic techniques for gastric motility disorders are seriously lacking, and are based more on eliminating other possibilities rather than on specific tests. Presently, gastric motility can be assessed by monitoring gastric emptying, food transit, intragastric pressures, etc. The associated tests are usually stationary and of relatively short duration. The present study proposes a new method of measuring gastric motility, utilizing the attenuation of an oscillator-induced electrical signal across the gastric tissue, which is modulated by gastric contractions. The induced high-frequency oscillator signal is generated within the stomach, and is picked up transluminally by cutaneous electrodes positioned on the abdominal area connected to a custom-designed data acquisition instrument. The proposed method was implemented in two different designs: first a transoral catheter was modified to emit the signal inside the stomach; and second, a gastric retentive pill was designed to emit the signal. Both implementations were applied in vivo on two mongrel dogs (25.50 kg and 25.75 kg). Gastric contractions were registered and quantitatively compared to recordings from force transducers sutured onto the serosa of the stomach. Gastric motility indices were calculated for each minute, with transluminal impedance measurements and the measurements from the force transducers showing statistically significant (p < 0.05) Pearson correlation coefficients (0.65 ± 0.08 for the catheter-based design and 0.77 ± 0.03 for the gastric retentive pill design). These results show that transcutaneous intraluminal impedance measurement has the potential with further research and development to become a useful diagnostic technique.

Phase structure and molecular dynamics of liquid-crystalline side-on organosiloxane tetrapodes.

X-ray diffraction and proton NMR relaxation measurements were carried out on two liquid-crystalline organosiloxane tetrapodes with side-on mesogenic groups, exhibiting nematic and smectic- C phases, and on a monomeric analog. Packing models for the mesophases exhibited by these systems are proposed on the basis of x-ray diffraction data. As a consequence of microsegregation, the aromatic cores are packed in between two sublayers formed by a mixture of interdigitated aliphatic and siloxane chains. The mixed sublayers are characteristic for the tetrapodes with side-on mesogenic groups presented in this work and have not been observed in tetrapodes with terminally attached mesogens. The tilt angle in the smectic- C phase is found very large, i.e., approximately 61 degrees -62 degrees . Notably, smectic- C clusters are present also in the whole temperature range of the nematic phase. NMR relaxometry yields T(1)-1 dispersions clearly different from those of conventional calamitics. The influence of molecular tendency to form interdigitated structures is evidenced by frequency-dependent relaxation rate in the isotropic phase-indicating the presence of ordered clusters far above the phase transition-and by the diminished role of molecular self-diffusion in ordered phases. Nematiclike director fluctuations are the dominating relaxation mechanism whereas the translational displacements are strongly hindered by the interdigitation of dendrimer arms.

Biaxial nematic order and phase behavior studies in an organosiloxane tetrapode using complementary deuterium NMR experiments.

The biaxial nematic phase was recently observed in different thermotropic liquid crystals, namely bent-core compounds, side-chain polymers, bent-core dimers, and organosiloxane tetrapodes. In this work, a series of experiments with a nematic organosiloxane tetrapode where nuclear magnetic resonance (NMR) spectra are collected while the sample is continuously rotating around an axis perpendicular to the magnetic field, are discussed in conjunction with the analysis of a deuterium NMR experiment on the same system reported earlier. The sample used is a mixture of a deuterated probe with the tetrapode. The mixture exhibits a nematic range between -40 degrees C and 37 degrees C. The results of the two independent, but complementary deuterium NMR experiments confirm the existence of a biaxial nematic phase for temperatures below 0 degrees C with high values of the asymmetry parameter at low temperatures. The presence of slow movements of the tetrapode mesogenic units in the low-temperature regime could also be detected through the analysis of the NMR spectra. Simulations indicate that these movements are mainly slow molecular reorientations of the mesogenic units associated with the presence of collective modes in the nematic phases of this compound. In the case of tetrapodes, recent investigations attribute the origin of biaxiality to the hindering of reorientations of the laterally attached mesogenic units which constitute the tetrapode. This study relates the molecular movements with the nematic biaxial ordering of the system.

Structure and molecular dynamics of the mesophases exhibited by an organosiloxane tetrapode with strong polar terminal groups.

The polymorphism of a new organosiloxane tetrapode compound with cyano terminal polar groups was characterized by means of polarizing optical microscopy and x-ray diffraction. The compound exhibits smectic- A and smectic- C phases with a partial bilayer arrangement due to a certain degree of head-to-head association of the mesogenic units through their cyano end groups. On the basis of x-ray diffraction results, evidencing the microsegregation of polyphilic molecules, packing models for the smectic- A and smectic- C phases are proposed. A high degree of smectic positional order and a relatively low value of the tilt angle in the smectic- C phase are indicated. Molecular dynamics of the studied compound was investigated by means of proton NMR relaxometry. The frequency dispersions of the spin-lattice relaxation time (T1) show that the relaxation is induced by three rotational modes of individual dendrimer arms with frequencies between 10;{6} and 10;{9}Hz . In the smectic phases, the effect of individual rotations is overwhelmed by a well expressed contribution of layer undulations at Larmor frequencies below approximately 10MHz . The appearance of this relaxation mechanism over the frequency range of three decades is so far unique in the case of thermotropic liquid crystals. The analysis of the layer undulations contribution supports the microsegregation model of the smectic phases by revealing a slowing-down of translational diffusion and the lack of interactions among the sublayers formed by the mesogenic groups.

Plastic-to-elastic transition in aggregated emulsion networks, studied with atomic force microscopy-confocal scanning laser microscopy microrheology.

In this paper, we demonstrate how the simultaneous application of atomic force microscopy (AFM) and confocal scanning laser microscopy (CSLM) can be used to characterize the (local) rheological properties of soft condensed matter at micrometer length scales. Measurement of AFM force curves as a function of the indentation amplitude and speed (magnitude and direction) can produce a "mechanical fingerprint" that contains information about material stiffness, hysteretic losses, and time scales for stress relaxation and/or network recovery. The simultaneous CSLM visualization of changes in the material's structure provides complementary information about how the material accommodates the indentation load. Since these experiments are done on areas of O(100 microm2) on materials having a surface of O(1 cm2), the measurements can be repeated on "fresh" material many times, contrary to traditional rheometers where the whole sample is loaded at once. As a particular example, we consider the case of a network of aggregated water-in-oil (W/O) emulsion droplets, in which the mechanical behavior changes drastically over time. Whereas the freshly prepared material shows a soft plastic behavior, after a time lapse of several weeks, the very same sample shows a much stiffer and elastic response. This drastic change in behavior is clearly reflected both in the signature of the AFM force curves and in (the reversibility of) the structural deformations observed with CSLM. The fact that these drastic mechanical changes take place without significant changes in the structure of the material (before loading) indicates that the stiffening of the droplet network is caused by an increase in the strength of the bonds between droplets. A remarkable finding for the elastic droplet network is that, while the structure recovers completely after the indenter is taken out, there is still an appreciable hysteresis in the force curves, indicating that dissipation also occurs. This hysteresis was not found to depend on the indentation speed.

Surfactant-mediated water transport at gelatin gel/oil interfaces.

We studied spontaneous emulsification (SE) at Water/Oil (W/O) interfaces, using several types of aqueous reservoirs immersed in dodecane plus Span80 surfactant. Above a threshold surfactant concentration C(SE), aqueous satellite droplets are formed at the W/O interface. Varying the aqueous reservoir size, from below 100 microm (droplets) to centimeters (macroscopic phases), allowed investigating SE with complementary techniques. Release (rates) and size distributions for SE droplets were measured with microscopy. For gelled aqueous phases, water expulsion due to SE was quantified. Values for C(SE) were measured and were found to be higher for aqueous phases containing gelatin and/or NaCl. We also studied water exudation during network building and syneresis in aqueous gelatin gels immersed in dodecane/Span80. Below C(SE) (i.e., in the absence of SE) this process is still responsible for significant physico-chemical changes at the W/O interface. To study these in more detail, we performed atomic force microscopy experiments (in force-distance mode) on macroscopic gels. Both changes in the local elastic response and in the wettability of the AFM tip were detected. Together they suggest the formation of "water pockets" after prolonged (gel) setting times, along with a densification of the interfacial gelatin network.

Microrheology of aggregated emulsion droplet networks, studied with AFM-CSLM.

We studied the mechanical behavior of densely packed (up to approximately 30% v/v), sedimented layers of (1 microm) water-in-oil W/O emulsion droplets, upon indentation with a (10 microm) large spherical probe. In the presence of attractive forces, the droplets form solid like networks which can resist deformation. Adding a polymer to the oil phase was used to control droplet attraction. The droplet layers were assembled via normal gravity settling. Considering that both the network structure and the droplet interactions play a key role, we used a combination of atomic force microscopy (AFM) and confocal scanning laser microscopy (CSLM) to characterize the mechanical behavior. Here the AFM was used both as indentation tool and as force sensor. Indentation experiments were performed via a protocol consisting of approach, waiting, and retract stages. CSLM was used to observe the network structure at micron resolution in real time. Use of refractive index matched fluorescent droplets allowed the visualization of the entire layer. Upon compression with the probe, a markedly nonhomogeneous deformation occurred, evidenced by the formation of a dense corona (containing practically all of the displaced droplets) in the direct vicinity of the probe, as well as more subtle deformations of force-chains at larger distances. Upon decompression, both the imprint of the indenter and the corona remained, even long after the load was released. The force-distance curves recorded with the AFM correspond well to these observations. For each deformation cycle performed on fresh material, the retract curve was much steeper than the approach curve, thus corroborating the occurrence of irreversible compaction. Contrary to classic linear viscoelastic materials, this hysteresis did not show any dependence on the deformation speed. Our force-indentation approach curves were seen to scale roughly as F approximately delta(3/2). The pre-factor was found to increase with the polymer concentration and with the density of the network. These findings suggest that this new AFM-CSLM method could be used for rheological characterization of small volumes of "granular networks" in liquid. Our hypothesis that the mechanical resistance of the networks originates from interdroplet friction forces, which in turn are set by the interdroplet potential forces, is supported by the predictions from a new mechanical model in which the interdroplet bonds are represented by stick-slip elements.

NMR relaxation study of molecular dynamics in columnar and smectic phases of a PAMAM liquid-crystalline co-dendrimer.

We present the first results obtained by proton ((1)H) nuclear magnetic relaxation studies of molecular dynamics in a supermolecular liquid-crystal dendrimer exhibiting columnar rectangular and smectic-A phases. The (1)H spin-lattice relaxation time (T(1)) dispersions are interpreted using two relaxation mechanisms associated with collective motions and local molecular reorientations of the dendritic segments in the low- and high-frequency ranges, respectively. The T(1) values show a drop around 2.3 MHz that is attributed to a contribution coming from cross-relaxation between (1)H and nitrogen nuclear spins. In the high-frequency range the motions appear to be of similar nature in both mesophases and are ascribed to reorientations of dendritic segments (belonging to the core and/or to the mesogenic units) characterized by two correlation times. Notable differences in the dynamics between the columnar and layered phases are observed in the low-frequency range. Depending on the mesophase they are discussed in terms of elastic deformations of the columns and layer undulations. In this study we find that the dendritic core influences the dynamics of the mesogenic units both for local and collective motions. These results can be understood in terms of spatial constraints imposed by the dendritic architecture and by the supermolecular arrangement in the mesophases.

Deuterium NMR investigation of the biaxial nematic phase in an organosiloxane tetrapode.

Deuterium NMR is used to examine the molecular order exhibited by an organosiloxane tetrapode giving the first experimental evidence, using a bulk sample, for the existence of a biaxial nematic phase in this type of compounds. The temperature dependence of the averaged quadrupolar coupling constant and asymmetry parameter was determined in the compound's nematic phase. Two distinct regimes could be identified, one with a vanishing asymmetry parameter corresponding to a uniaxial nematic phase and another with a significant temperature dependent asymmetry parameter, corresponding to a biaxial nematic phase. The high values obtained for the asymmetry parameter at the lower end of the nematic range are well above experimental error and constitute a definite proof of the biaxial nature of the nematic phase exhibited by the studied compound for those temperatures.

Influence of bulk elasticity and interfacial tension on the deformation of gelled water-in-oil emulsion droplets: an AFM study.

We used atomic force microscopy (AFM) to study the deformation and wetting behavior of large (50-250 microm) emulsion droplets upon mechanical loading with a colloidal glass probe. Our droplets were obtained from water-in-oil emulsions. By adding gelatin to the water prior to emulsification, also droplets with a bulk elasticity were prepared. Systematic variations of surfactant and gelatin concentrations were made, to investigate their effect on the deformation and wetting behavior of the droplets and to identify the contributions of interfacial tension, bulk elasticity, and expelled water. The AFM experiments were performed in force--distance mode and showed on approach a repulsive regime which in many cases was terminated by a jump-in of the probe. In the case of pure water (i.e. gelatin-free) droplets, the repulsive part of the curve showed a good linearity, thus allowing the extraction of an effective droplet spring constant. This quantity was found to decrease on raising the surfactant concentration from below the critical micelle concentration (cmc) to well above the cmc, and its numerical values were found to correspond remarkably well to literature values for the interfacial tension. Our findings indicate that, on gelatin increase inside the droplets, the bulk elasticity gradually becomes dominant and the droplets' stiffness does not depend anymore on surfactant concentration. Also the stability of the droplet interface against wetting, as measured by the force at which the jump-in instability occurs, was enhanced by gelatin. For gelatin concentrations of > or =15 wt %, the droplets were found to behave like purely elastic bodies. Both gelatin and surfactant contribute positively to the stability against interface breakup.

The effects of an interactive instructional strategy for enhancing reading comprehension and content area learning for students with learning disabilities.

This study investigated the effectiveness of an interactive vocabulary instructional strategy, semantic-feature analysis (SFA), on the content area text comprehension of adolescents with learning disabilities. Prior to reading a social studies text, students in resource classes either completed a relationship chart as part of the SFA condition or used the dictionary to write definitions and sentences as part of the contrast condition. Passage comprehension was measured on a multiple-choice test consisting of two types of items, vocabulary and conceptual. Comprehension was measured immediately following teaching and again 6 months after teaching. Prior knowledge for the content of the passage served as a covariate. Results indicated that students in the SFA instructional condition had significantly greater measured comprehension immediately following and 6 months after initial teaching. These results are discussed in relation to concept-driven, interactive strategies for teaching content and facilitating text comprehension.

The hyperlipidemic hamster as a model of experimental atherosclerosis.

Male hamsters were fed a hyperlipidemic diet consisting of standard chow supplemented with 3% cholesterol and 15% commercial butter for 12 months. In about 3 weeks serum total cholesterol doubled, raised 4-fold after the 4th week and after 10 months attained a 17-fold value. Low density lipoproteins (LDL)-cholesterol increased 4-fold after 4 weeks and about 13-fold after 10 months compared to control animals. In the first 2 weeks mononuclear cells began to adhere to the endothelium and a very intense stromal reaction appeared in the intima of the aortic arch. At the end of the 4th week of diet, Oil Red O stainable deposits were visible on the thoracic aorta, mostly on the arch, some of them as isolated, lipid-laden cells and others distributed on focal areas. Smooth muscle cells (SMC) appeared also in the intima of hyperlipidemic hamsters, compared to normal animals which had no macrophages or smooth muscle cells in the intima of the aortic specimens examined. Up to 6 months, smooth muscle cells in the intima and media began to load with lipids, as well as endothelial cells. After 10 months the affected zones looked like human atherosclerotic plaque with huge cholesterol crystal deposits, calcium deposits and necrosis. The endothelium, though very thinned and loaded with lipids, was morphologically intact.

Cellular events in the development of valvular atherosclerotic lesions induced by experimental hypercholesterolemia.

The onset and evolution of ultrastructural changes in the cardiac valves induced by a cholesterol-rich diet were investigated in rabbit and hamster. In both animal models, the atrioventricular and sigmoid valves were comparably affected by lesions intermediary between fatty streak and fibrous plaque. The earliest detectable modification was the progressive accumulation in the subendothelium of extracellular liposome-like structures rich in unesterified cholesterol, associated with the proliferation of a basal lamina-like material. This was followed by the diapedesis of blood monocytes in the same location, which became macrophages increasingly loaded with lipid deposits. Resident interstitial cells accumulate lipids, as well. In advanced stages, the macrophage-derived foam cells clustered, deforming the valve leaflets. The resident macrophages accumulated lipids later and more slowly, while partly preserving their ultrastructure. The advanced lesions are characterized by marked stromal proliferation, massive intra- and extracellular deposition of lipids and cholesterol crystals and the appearance of a necrotic core. The salient findings of these studies were: (1) the appearance of extracellular liposomes as the earliest event in atherogenesis; (2) the capability of the valvular interstitial cells to accumulate lipids; and (3) the slow response of resident macrophages to the cholesterol-rich diet. The results revealed that hypercholesterolemia produces in the cardiac valves atherosclerotic lesions of an intermediate type, which can deform the leaflets thus altering their normal function.

Interstitial cells of the heart valves possess characteristics similar to smooth muscle cells.

Interstitial cells of heart atrioventricular and sigmoid valves were examined in several laboratory animals (rabbit, hamster, rat, and mouse) and in humans. These cells constitute a large fraction of the total cell population of the valve; in mouse atrioventricular valves, they amount to approximately 30% of the volumetric density. By their ultrastructural features and functional properties, valvular interstitial cells are intermediate between fibroblasts and vascular smooth muscle cells. Like fibroblasts, valvular interstitial cells lack a basal lamina establishing direct and extensive contacts with collagen fibers, elastin microfibrils, and proteoglycans of the matrix. The cells have numerous slender and long processes, connected to one another, forming a complex cellular framework spanning the entire valve. Similar to smooth muscle cells, valvular interstitial cells are extensively coupled by communicating junctions as shown by thin sections, freeze-fracture, lanthanum staining, and carboxyfluorescein microinjection. The cells contain numerous bundles of actin filaments, which are decorated by the S1 fragment of heavy meromyosin. Valvular interstitial cells also express cyclic guanosine-monophosphate-dependent protein kinase, as detected by immunofluorescence and immunoperoxidase histochemistry. Motor nerve endings are located closely apposed to valvular interstitial cells: structurally most of them appear to be of the adrenergic type. Valvular interstitial cells contract on epinephrine or angiotensin II stimulation as shown both in culture and in situ (valvular strips). Taken together these observations suggest that VIC may have contractile properties, which can account for a controlled tonus, actively correlated with the cyclically changing forces acting on valves during diastole and systole.

Mouse atrio-ventricular valve ultrastructure morphometrical correlations.

Morphometric data on mouse atrio-ventricular valve ultrastructure are reported. The statistical analysis of the volumetric density percentage for cellular and extracellular valve components and of endothelial plasmalemmal vesicle density for the different endocardial domains (atrial, valvular and ventricular) showed: the bicuspid compared to the tricuspid valve has a more important lymphatic drainage, less vascularization, higher endothelial plasmalemmal vesicle density and more macrophages, striated muscle cells and collagen along with fewer interstitial cells, nervous terminals and elastin in the leaflet; the valvular endothelium as compared to other endocardial domains has a higher density of plasmalemmal vesicles, considering the results for both endothelial fronts (luminal and abluminal).

The Ranvier node as a chemo-electric pulsatory unit: a study of its structure-functions relations.

The Ranvier node of (Rana temporaria) frog nerve fibres is investigated by electron microscopy, Particular attention is given to the paranodal septate structures and to the extranodal junction of two Schwann cells. An interpretation of the functional meaning of these structures along with a quantitative analysis of the Schwann extranodal junction as regards the diffusion from/to the node is attempted. A 73 per cent reduction of the diffusion coefficient is obtained if the extranodal Schwann cell processes are considered impermeable to the diffusing vectors which indicates a protective role. Only 1 per cent reduction is obtained in the case of excitation-involved cations to which the Schwann cell membrane is considered to be permeable. This indicates the active role of the Schwann cell in the extranodal area ion diffusion, by minimizing the variations in ion concentration near the nodal membrane. Thus the nervous fibre-Schwann cell assembly may be regarded as a balanced pulsatory chemo-electric unit.