Exposure to 900 MHz radiofrequency radiation induces caspase 3 activation in proliferating human lymphocytes

Palumbo, R., Brescia, F., Capasso, D., Sannino, A., Sarti, M., Capri, M., Grassilli, E. and Scarfì, M. R. Exposure to 900 MHz Radiofrequency Radiation Induces Caspase 3 Activation in Proliferating Human Lymphocytes. Radiat. Res. 170, 327- 334 (2008).In this study, the induction of apoptosis after exposure to 900 MHz radiofrequency radiation (GSM signal) was investigated by assessing caspase 3 activation in exponentially growing Jurkat cells and in quiescent and proliferating human peripheral blood lymphocytes (PBLs). The exposure was carried out at an average specific absorption rate of 1.35 W/kg in a dual wire patch cell exposure system where the temperature of cell cultures was accurately controlled. After 1 h exposure to the radiofrequency field, a slight but statistically significant increase in caspase 3 activity, measured 6 h after exposure, was observed in Jurkat cells (32.4%) and in proliferating human PBLs (22%). In contrast, no effect was detected in quiescent human PBLs. In the same experimental conditions, apoptosis was also evaluated in Jurkat cells by Western blot analysis and in both cell types by flow cytometry. To evaluate late effects due to caspase 3 activity, flow cytometry was also employed to assess apoptosis and viability 24 h after radiofrequency-radiation exposure in both cell types. Neither the former nor the latter was affected. Since in recent years it has been reported that caspases are also involved in processes other than apoptosis, additional cell cycle studies were carried out on proliferating T cells exposed to radiofrequency radiation; however, we found no differences between sham-exposed and exposed cultures. Further studies are warranted to investigate the biological significance of our findings of a dose-response increase in caspase 3 activity after exposure to radiofrequency radiation.     

Population structure and phylogeography reveal pathways of colonization by a migratory marine reptile (Chelonia mydas) in the central and eastern Pacific

Climate, behavior, ecology, and oceanography shape patterns of biodiversity in marine faunas in the absence of obvious geographic barriers. Marine turtles are an example of highly migratory creatures with deep evolutionary lineages and complex life histories that span both terrestrial and marine environments. Previous studies have focused on the deep isolation of evolutionary lineages (>3 mya) through vicariance; however, little attention has been given to the pathways of colonization of the eastern Pacific and the processes that have shaped diversity within the most recent evolutionary time. We sequenced 770 bp of the mtDNA control region to examine the stock structure and phylogeography of 545 green turtles from eight different rookeries in the central and eastern Pacific. We found significant differentiation between the geographically separated nesting populations and identified five distinct stocks (F_ST = 0.08–0.44, P < 0.005). Central and eastern Pacific Chelonia mydas form a monophyletic group containing 3 subclades, with Hawaii more closely related to the eastern Pacific than western Pacific populations. The split between sampled central/eastern and western Pacific haplotypes was estimated at around 0.34 mya, suggesting that the Pacific region west of Hawaii has been a more formidable barrier to gene flow in C. mydas than the East Pacific Barrier. Our results suggest that the eastern Pacific was colonized from the western Pacific via the Central North Pacific and that the Revillagigedos Islands provided a stepping‐stone for radiation of green turtles from the Hawaiian Archipelago to the eastern Pacific. Our results fit with a broader paradigm that has been described for marine biodiversity, where oceanic islands, such as Hawaii and Revillagigedo, rather than being peripheral evolutionary “graveyards”, serve as sources and recipients of diversity and provide a mechanism for further radiation.     

Binding and segmentation of multiple objects through neural oscillators inhibited by contour information

 Temporal correlation of neuronal activity has been suggested as a criterion for multiple object recognition. In this work, a two-dimensional network of simplified Wilson-Cowan oscillators is used to manage the binding and segmentation problem of a visual scene according to the connectedness Gestalt criterion. Binding is achieved via original coupling terms that link excitatory units to both excitatory and inhibitory units of adjacent neurons. These local coupling terms are time independent, i.e., they do not require Hebbian learning during the simulations. Segmentation is realized by a two-layer processing of the visual image. The first layer extracts all object contours from the image by means of “retinal cells” with an “on-center” receptive field. Information on contour is used to selectively inhibit Wilson-Cowan oscillators in the second layer, thus realizing a strong separation among neurons in different objects. Accidental synchronism between oscillations in different objects is prevented with the use of a global inhibitor, i.e., a global neuron that computes the overall activity in the Wilson-Cowan network and sends back an inhibitory signal. Simulations performed in a 50×50 neural grid with 21 different visual scenes (containing up to eight objects + background) with random initial conditions demonstrate that the network can correctly segment objects in almost 100% of cases using a single set of parameters, i.e., without the need to adjust parameters from one visual scene to the next. The network is robust with reference to dynamical noise superimposed on oscillatory neurons. Moreover, the network can segment both black objects on white background and vice versa and is able to deal with the problem of “fragmentation.” The main limitation of the network is its sensitivity to static noise superimposed on the objects. Overcoming this problem requires implementation of more robust mechanisms for contour enhancement in the first layer in agreement with mechanisms actually realized in the visual cortex. Received: 25 October 2001 / Accepted: 26 February 2003 / Published online: 20 May 2003 Correspondence to: Mauro Ursino (e-mail: mursino@deis.unibo.it, Tel.: +39-051-2093008, Fax: +39-051-2093073)     

Mechanism of control of cytochrome oxidase activity by the electrochemical-potential gradient.

Cytochrome c oxidation by bovine cytochrome oxidase embedded into liposomal vesicles with high respiratory control ratio (RCR = 6-10) has been studied by rapid-mixing experiments in the presence and absence of different ionophores. Kinetic analysis of the reaction indicates a linkage between the intrinsic activity of the enzyme, the efficiency of coupling and the electrochemical potential across the membrane. A simple model, based on two allosteric states with different catalytic properties in rapid equilibrium, is presented and successfully applied in the simulation of the observed time-course.     

Evaluation of P-glycoprotein expression in soft tissue sarcomas of the extremities

Soft tissue sarcomas comprise a heterogeneous group of mesenchymal tumors accounting for less than one-percent of adult neoplasms. In the last few years, the use of adjuvant chemotorapy has been proposed for the treatment of these lesions in order to obain a better systemic control, but its usefulness is still controversial. In this study, we evaluated whether P-glycoprotein, a membrane protein strictly associated with multidrug resistance, is overexpressed in soft tissue sarcomas. By using human multidrug resistant sarcoma cell lines as controls, we analyzed P-glycoprotein expression in 34 primary and in 23 relapsed soft tissue sarcomas of the extremities. Overexpression of P-glycoprotein was found in 6 out of 34 primaries (18%) and in 8 out of 23 relapses (35%). In particular, in malignant fibrous histiocytoma, the most frequent soft tissue sarcoma of adults, P-glycoprotein overexpression was found in 23% of primary untreated cases, in agreement with the reported relapse rate of this tumor after surgery and chemotherapy. These data suggest that, in soft tissue sarcomas, overexpression of P-glycoprotein may be of prognostic value and that the assessment of P-glycoprotein expression may be useful for the design of chemotherapy protocols.Abbreviations MDR multidrug-resistance - STS soft tissue sarcomas     

S-Adenosyl-l-methionine Modulates CO and NO? Binding to the Human H2S-generating Enzyme Cystathionine β-Synthase

Cystathionine β-synthase (CBS) is a key enzyme in human (patho)physiology with a central role in hydrogen sulfide metabolism. The enzyme is composed of a pyridoxal 5′-phosphate-binding catalytic domain, flanked by the following two domains: a heme-binding N-terminal domain and a regulatory C-terminal domain binding S-adenosyl-l-methionine (AdoMet). CO or NO^• binding at the ferrous heme negatively modulates the enzyme activity. Conversely, AdoMet binding stimulates CBS activity. Here, we provide experimental evidence for a functional communication between the two domains. We report that AdoMet binding significantly enhances CBS inhibition by CO. Consistently, we observed increased affinity (∼5-fold) and faster association (∼10-fold) of CO to the ferrous heme at physiological AdoMet concentrations. NO^• binding to reduced CBS was also enhanced by AdoMet, although to a lesser extent (∼2-fold higher affinity) as compared with CO. Importantly, CO and NO^• binding was unchanged by AdoMet in a truncated form of CBS lacking the C-terminal regulatory domain. These unprecedented observations demonstrate that CBS activation by AdoMet puzzlingly sensitizes the enzyme toward inhibition by exogenous ligands, like CO and NO^•. This further supports the notion that CBS regulation is a complex process, involving the concerted action of multiple physiologically relevant effectors.     

Effect of gangliosides on membrane permeability studied by enzymic and fluorescence-spectroscopy techniques.

The effect of gangliosides on membrane permeability was investigated by studying the kinetic properties of cytochrome c oxidase, the activity of which, when the enzyme is reconstituted in phospholipid vesicles, is dependent on membrane permeability to H+ and K+. The experiments indicate that three different gangliosides (GM1, DD1a, GT1b) incorporated into cytochrome c oxidase-containing phospholipid vesicles stimulate enzymic activity, in the absence of ionophores, most probably by disorganizing the bilayer lipid assembly and increasing its permeability to ions. This interpretation was confirmed by fluorescence-spectroscopy experiments in which the rate of passive leakage of carboxyfluorescein entrapped in the vesicles was measured. Cholera toxin, or its isolated B-subunit, added to GM1-containing proteoliposomes inhibited cytochrome c oxidase activity, indicating the lack of formation, under these experimental conditions, of channels freely permeable to H+ or K+.     

Transient kinetics of subunit-III-depleted cytochrome c oxidase.

Cytochrome c oxidase from ox heart was depleted of subunit III and its transient kinetic properties studied by stopped-flow and flash photolysis. It was found that the overall mechanism of electron transfer is very similar for subunit-III-depleted and native oxidase, although significant differences in some kinetic parameters have been detected. These include the second-order rate constant for cytochrome c oxidation and the rate-limiting step of the overall process. Moreover, at low cytochrome c/oxidase ratios (where the number of reducing equivalents is insufficient), the rate of reoxidation of cytochrome a was found to be very slow, even in air, and in fact for the subunit-III-depleted enzyme is even slower than for the native oxidase. The stability of reduced cytochrome a excludes the likelihood that removal of subunit III leads to a new O2-binding site, and the result may be relevant to the lowered vectorial H+/e- stoichiometry. The subunit-III-depleted oxidase can be pulsed under appropriate conditions and its combination with CO is unchanged, as shown by kinetic experiments and difference spectroscopy.