Regulation of Na,K-ATPase Transport Activity by Protein Kinase C

Considerable evidence indicates that the renal Na⁺,K⁺-ATPase is regulated through phosphorylation/dephosphorylation reactions by kinases and phosphatases stimulated by hormones and second messengers. Recently, it has been reported that amino acids close to the NH₂-terminal end of the Na⁺,K⁺-ATPase α-subunit are phosphorylated by protein kinase C (PKC) without apparent effect of this phosphorylation on Na⁺,K⁺-ATPase activity. To determine whether the α-subunit NH₂-terminus is involved in the regulation of Na⁺,K⁺-ATPase activity by PKC, we have expressed the wild-type rodent Na⁺,K⁺-ATPase α-subunit and a mutant of this protein that lacks the first thirty-one amino acids at the NH₂-terminal end in opossum kidney (OK) cells. Transfected cells expressed the ouabain-resistant phenotype characteristic of rodent kidney cells. The presence of the α-subunit NH₂-terminal segment was not necessary to express the maximal Na⁺,K⁺-ATPase activity in cell membranes, and the sensitivity to ouabain and level of ouabain-sensitive Rb⁺-transport in intact cells were the same in cells transfected with the wild-type rodent α1 and the NH₂-deletion mutant cDNAs. Activation of PKC by phorbol 12-myristate 13-acetate increased the Na⁺,K⁺-ATPase mediated Rb⁺-uptake and reduced the intracellular Na⁺ concentration of cells transfected with wild-type α1 cDNA. In contrast, these effects were not observed in cells expressing the NH₂-deletion mutant of the α-subunit. Treatment with phorbol ester appears to affect specifically the Na⁺,K⁺-ATPase activity and no evidence was observed that other proteins involved in Na⁺-transport were affected. These results indicate that amino acid(s) located at the α-subunit NH₂-terminus participate in the regulation of the Na⁺,K⁺-ATPase activity by PKC. Received: 10 July 1996/Revised: 19 September 1996     

Sensitivity to weighting in life cycle impact assessment (LCIA)

The International Journal of Life Cycle Assessment - Weighting in life cycle assessment (LCA) incorporates stakeholder preferences in the decision-making process of comparative LCAs. Research...     

Benthic marine flora in the Tuscan Archipelago. A first contribution: Isles of Capraia,Elba, Formiche di Grosseto,Giglio, Scoglio d'Africa,Montecristo and Giannutri

Abstract

Benthic marine flora of Tuscan Archipelago has been studied. Several samples were collected along five islands and some rocks. This paper is a compendium of new and already published data, in which 267 species, varieties, forms and stadia of algae and seagrasses are listed: 5 Bangiophyceae, 182 Florideophyceae, 45 Phaeophyceae, 6 Chlorophyceae, 28 Bryopsidophyceae, 1 Angiospermae.     

Note floristique et repartition de Rhodophycees rares (Kallymeniacees et Sebdeniacees) de la Mediterranee occidentale

Abstract

Floristic notes and geographical distribution of some rare Rhodophyta (Kallymeniaceae and Sebdeniaceae) of the western Mediterranean. Callophyllis laciniata, Kallymenia requienii and Kallymenia lacerata (Kallymeniaceae) and Sebdenia rodrigueziana (Sebdeniaceae) are recorded for the first time in the bay of Naples. For each species systematic and distribution data are also given.     

Studio preliminare sulla tipologia della vegetazione sommersa del Canale di Sicilia e isole vicine

Abstract

Preliminary studies on the marine phythobenthic communities in the Straits of Sicily and adjacent islands. — The results of algological researches carried out by the Gruppo di Algologia of the S.B.I. in the Straits of Sicily and adjacent islands are reported. 288 species have been collected, 275 of which have been identified: 160 were Rhodophyta, 62 Phaeophyta, 32 Chlorophyta, 16 Cyanophyta and 2 Angiospermae. The vegetational research leads to the following conclusions: 1 - the harbour areas (Pantelleria, Malta: loc. 1 and 11) show a nitrophile vegetation, particulary near La Valletta where an almost pure population of Pterocladia pinnata is found; 2 - there are important biogeographical differences between Pelagie islands (Lampedusa, Linosa) and Pantelleria. In fact the former ones have an algal flora composed mostly of eastern mediterranean species, whilst the latter shows strong affinities with the north-african basin, as is demonstrated by the rich well developed belt of Cystoseira sedoides; 3 - several species of atlantic origin, such as Callophyllis laciniata, Peyssonnelia coriacea, P. inamoena and Castagnea chordariaeformis, have been found on the banks (loc. 3, 6, 7).     

Visual BOLD Response in Late Blind Subjects with Argus II Retinal Prosthesis

Retinal prosthesis technologies require that the visual system downstream of the retinal circuitry be capable of transmitting and elaborating visual signals. We studied the capability of plastic remodeling in late blind subjects implanted with the Argus II Retinal Prosthesis with psychophysics and functional MRI (fMRI). After surgery, six out of seven retinitis pigmentosa (RP) blind subjects were able to detect high-contrast stimuli using the prosthetic implant. However, direction discrimination to contrast modulated stimuli remained at chance level in all of them. No subject showed any improvement of contrast sensitivity in either eye when not using the Argus II. Before the implant, the Blood Oxygenation Level Dependent (BOLD) activity in V1 and the lateral geniculate nucleus (LGN) was very weak or absent. Surprisingly, after prolonged use of Argus II, BOLD responses to visual input were enhanced. This is, to our knowledge, the first study tracking the neural changes of visual areas in patients after retinal implant, revealing a capacity to respond to restored visual input even after years of deprivation.     

Apoptosis: Molecular regulation of cell death and hematologic malignancies

We describe the molecular mechanisms of apoptosis and its relationships with hematologic malignancies, stressing the concept that, both positive and negative deregulation of apoptosis, may be involved in hematologic human diseases. So, this fundamental process must be balanced by so far unknown mechanisms, involving caspases (cysteine proteases, cleaving the protein substrate after an aspartate residue). These, so far known, ten proteases, are interconnected in a molecular cascade, initiated by the release of cytochrome C from mitochondrial membranes and its interaction with APAF-1 (the homolog of the Caenorhabditis e. CED-4) and with caspase 9, that initiates the proteolitic cascade (1,2). The conclusion is that apoptosis is a very important process, but yet poorly known in molecular details, in spite of the efforts of many scientists. Even the role of bcl-2, the main gene protecting from apoptosis, is still unknown. We close this chapter with a list of ten different technical approaches that can be useful tools to study apoptosis, and tracing the molecular principles on which they are based.     

High-definition mapping of neural activity using voltage-sensitive dyes

The distribution of patterns of activity in different brain structures has been related to the encoding and processing of sensory information. Consequently, it is important to be able to image the distribution of these patterns to understand basic brain functions. The spatial resolution of voltage-sensitive dye (VSD) methods has recently been enhanced considerably by the use of video imaging techniques. The main factor that now hampers the resolution of VSD patterns is the inherent limitation of the optical systems. Unfortunately, the intrinsic characteristics of VSD images impose important limitations that restrict the use of general deconvolution techniques. To overcomes this problem, in this study an image restoration procedure has been implemented that takes into consideration the limiting characteristics of VSD signals. This technique is based on applying a set of imaging processing steps. First, the signal-to-noise (S/N) ratio of the images was improved to avoid an increase in the noise levels during the deconvolution procedures. For this purpose, a new filter technique was implemented that yielded better results than other methods currently used in optical imaging. Second, focal plane images were deconvolved using a modification of the well-known nearest-neighbor deconvolution algorithm. But to reduce the light exposure of the preparation and simplify image acquisition procedures, adjacent image planes were modeled according to the in-focus image planes and the empirical point spread function (PSF) profiles. Third, resulting focal plane responses were processed to reduce the contribution of optical responses that originate in distant image planes. This method was found to be satisfactory under simulated and real experimental conditions. By comparing the restored and unprocessed images, it was clearly demonstrated that this method can effectively remove the out-of-focus artifacts and produce focal plane images of better quality. Evaluations of the tissue optical properties allowed assessment of the maximum practical optical section thickness using this deconvolution technique in the optical system tested. Determination of the three-dimensional PSF permitted the correct application of deconvolution algorithms and the removal of the contaminating light arising from adjacent as well as distant optical planes. The implementation of this deconvolution approach in salamander olfactory bulb allowed the detailed study of the laminar distribution of voltage-sensitive changes across the bulb layer. It is concluded that (1) this deconvolution procedure is well suited to deconvolved low-contrast images and offers important advantages over other alternatives; (2) this method can be properly used only when the tissue optical properties are first determined; (3) high levels of light scattering in the tissue reduce the optical section capabilities of this technique as well as other deconvolution procedures; and (4) use of the highest numerical aperture in the objectives is advisable because this improves not only the light-collecting efficiency to detect poor-contrast images, but also the spatial frequency differences between adjacent image planes. Under this condition it is possible to overcome some of the limitations imposed by the light scattering/birefringence of the tissue.     

Raloxifene reduces urokinase-type plasminogen activator-dependent proliferation of synoviocytes from patients with rheumatoid arthritis

Extracellular fibrinolysis, controlled by the membrane-bound fibrinolytic system, is involved in cartilage damage and rheumatoid arthritis (RA) synovitis. Estrogen status and metabolism seem to be impaired in RA, and synoviocytes show receptors for estrogens. Our aims in this study were to evaluate in healthy and RA synoviocytes the effects of Raloxifene (RAL), a selective estrogen receptor modulator (SERM), on: proliferation; the components of the fibrinolytic system; and chemoinvasion. The effects of RAL were studied in vitro on synoviocytes from four RA patients and four controls. Proliferation was evaluated as cell number increase, and synoviocytes were treated with 0.5 microM and 1 microM RAL with and without urokinase-plasminogen activator (u-PA) and anti-u-PA/anti-u-PA receptor (u-PAR) antibodies. Fibrinolytic system components (u-PA, u-PAR and plasminogen activator inhibitor (PAI)-1) were assayed by ELISA with cells treated with 0.5 microM and 1 microM RAL for 48 h. u-PA activity was evaluated by zymography and a direct fibrinolytic assay. U-PAR/cell and its saturation were studied by radioiodination of u-PA and a u-PA binding assay. Chemoinvasion was measured using the Boyden chamber invasion assay. u-PA induced proliferation of RA synoviocytes was blocked by RAL (p < 0.05) and antagonized by antibodies alone. The inhibitory effect of RAL was not additive with u-PA/u-PAR antagonism. RA synoviocytes treated with RAL showed, compared to basal, higher levels of PAI-1 (10.75 +/- 0.26 versus 5.5 +/- 0.1 microg/10(6) cells, respectively; p < 0.01), lower levels of u-PA (1.04 +/- 0.05 versus 3.1 +/- 0.4 ng/10(6) cells, respectively; p < 0.001), and lower levels of u-PAR (11.28 +/- 0.22 versus 23.6 +/- 0.1 ng/10(6) cells, respectively; p < 0.001). RAL also significantly inhibited u-PA-induced migration. Similar effects were also shown, at least partially, in controls. RAL exerts anti-proliferative and anti-invasive effects on synoviocytes, mainly modulating u-PAR and, to a lesser extent, u-PA and PAI-1 levels, and inhibiting cell migration and proliferation.