The Golgi apparatus is a primary site of intracellular damage after photosensitization with Rose Bengal acetate

The aim of the present investigation was to elucidate whether the Golgi apparatus undergoes photodamage following administration of the fluorogenic substrates Rose Bengal acetate (RBAc) and irradiation at the appropriate wavelength. Human HeLa cells were treated in culture and the changes in the organization of the Golgi apparatus were studied using fluorescence confocal microscopy and electron microscopy, after immunocytochemical labeling. To see whether the cytoskeletal components primarily involved in vescicle traffic (i.e., microtubules) might also be affected, experiments of tubulin immunolabeling were performed. After treatment with RBAc and irradiation, cells were allowed to grow in drug-free medium for different times. 24hr after irradiation, the cisternae of the Golgi apparatus became packed, and after 48-72 hr they appeared more fragmented and scattered throughout the cytoplasm; these changes in the organization of the Golgi cisternae were confirmed at electron microscopy. Interestingly enough, apoptosis was found to occur especially 48-72h after irradiation, and apoptotic cells exhibited a dramatic fragmentation of the Golgi membranes. The immunolabeling with anti-tubulin antibody showed that microtubules were also affected by irradiation in RBAc-treated cells

Intracellular distribution of Tankyrases as detected by multicolor immunofluorescence techniques

Poly(ADP-ribose) polymerases are a family of enzymes that catalyze the conversion of NAD+ into ADP-ribose. Among them, Tankyrases have been found to bind to centrosome, mitotic spindle and microsome proteins, in the cytoplasm, and to telomeres in the nucleus, where they play a relevant role in telomere metabolism. However, their precise intracellular localization during interphase has not been so far fully elucidated. We investigated this aspect in situ by double immunofluorescence experiments using antibodies recognizing Tankyrases 1–2 or other proteins residing in specific organelles (Golgi apparatus, mitochondria, lysosomes, endoplasmic reticulum). We used HeLa cells as a model system in vitro, before and after treatment with either actinomycin D or etoposide, to also investigate the possible relocation of Tankyrases during apoptosis. We observed that Tankyrases are distributed both in the nucleus and in the cytoplasm; in this latter compartment, they were found to colocate with the Golgi apparatus but never with the mitochondria; a pool of Tankyrases also colocates with the endoplasmic reticulum and lysosomes. Interestingly, in cells with clear signs of apoptosis, Tankyrases were detectable in the cytoplasmic blebs: this suggests that they are not massively cleaved during apoptosis and persist in the largely heterogeneous apoptotic remnants which are known to contain components of cytoplasmic and nuclear origin

Harbour water managment for port structures and sea bottom design, coast proximity navigation managment, water quality control.

Tide is a sea level up-down water motion basically depending on three different phenomena: the Earth-Moon-Sun gravitational relationship, the water surface fluid reaction to atmospheric meteorological dynamic action and the Newtonian vertical adjustment of the sea surface due to atmospheric pressure variations. The first tide component (astro-tide) is periodic and well known in all points of the Earth surface; the second one is directly related to the meteorological phenomenon and then it is foreseeable; the Newtonian component, on the contrary, is not readily predictable by a general hydrostatic law, because the factor “J” that represents the Newtonian transfer (from the atmospheric weight to the consequent sea level) is variable in each harbour area. A statistical study and the related numerical data interpretation of the measurements performed in the Ports of Genoa, La Spezia, Marina di Carrara, Livorno, Piombino, Civitavecchia and Ravenna (belonging to the Italian Newtonian Meteotide Network) show port values of Jph (from 1.4-1.6 cm/hPa to > 2 cm/hPa, on the contrary of the off-shore areas where Jph is about 1 cm/hPa). This phenomenon (hydrobarometric tide wave) produces even double values of harbour sea level fluctuations amplitude in comparison to astronomic tide sea level oscillations, and is characterized by a wavelength from 8-12 h to same days and a 10-25 days/year as mean temporal occurrence in the Northern Tyrrhenian Sea. This is the most relevant ordinary risk factor for our ports activity and structures design. The present note shows a quantitative method to define the values of Jph factor for ports and its use in the Harbour WaterSide Management (HWSM) based on the joined use of barometers, hydrometers and clocks, the preliminary results related to the use of the gravimeters as hydrobarometric predictor in La Spezia Port and two examples of use of Jph factor in the port management: refloating of a landing ship and optimisation of a dock performance as pleasure boats mooring

Nuclear localization of phosphorylated c-Myc protein in human tumor cells.

Using immunocytochemical techniques at light and electron microscopy, we analysed the distribution of phosphorylated c-Myc in actively proliferating human HeLa cells. The distribution pattern of c-Myc was also compared with those of other ribonucleoprotein (RNP)-containing components (PANA, hnRNP-core proteins, fibrillarin) or RNP-associated nuclear proteins (SC-35 splicing factor). Our results provide the first evidence that phosphorylated c-Myc accumulates in the nucleus of tumor cells, where it colocalizes with fibrillarin, both in the nucleolus and in extranucleolar structures

The newtonian approach in the meteorological tide waves forecasting: preliminary observations in the East Ligurian harbours

Sea level oscillations are the superposition of many contributions, among which the main are astronomic and meteorological low-frequency tides. In Ligurian Sea meteo-tide components, being most ample than astronomic fluctuations, drive water exchange in harbours. The present note shows first results about port of Genoa concerning a coherency study between atmospheric variation and corresponding sea level adjustment (meteorological tide). The newtonian forecasting method of meteorological tides is based on measurements of time elapsing between barometric sea level unbalance (Δg) and its meteorological tide compensation (inverse barometer component). Meteorological tide component is independent on the Earth-Moon-Sun gravitational relationships, moreover parameters related to the shifted water mass are too many to describe the phenomenon analytically (basin topography, barometric strength position and time, chemical water quality, off-shore sea circulation, etc.); then, meteorological tide can’t be accurately foreseen by atmospheric pressure measurements only. A gravimeter can detect the geodetic unbalance starting time and a tide-gauge can detect the newtonian compensation (tide wave) coming time. The difference between these two times is the meteorological tide delay. An opportune statistic of this delay provides an experimental law typical for each harbour to forecast the meteo-tide compensation wave delay. This paper describes the methodological procedure adopted and first evidences of the phenomenon in Genoa harbour

Sistema per l'acquisizione e la trasmissione dei dati della stazione mareografica MENFOR

Il presente documento descrive i componenti e le funzionalità del sistema realizzato per l’acquisizione e la trasmissione dei dati della stazione mareografica MENFOR sviluppata nell’ambito del progetto “Sviluppo di una stazione portuale per la previsione dei flussi di marea meteorologica, finalizzata a costituire un servizio per la sicurezza della navigazione e per la protezione dei natanti nel Golfo della Spezia” supportato dal programma PRAI-FESR della Regione Liguria. Il sistema qui descritto è stato realizzato con il contributo di tutti gli Enti coinvolti

Effect of platelet lysate on human cells involved in different phases of wound healing

Background Platelets are rich in mediators able to positively affect cell activity in wound healing. Aim of this study was to characterize the effect of different concentrations of human pooled allogeneic platelet lysate on human cells involved in the different phases of wound healing (inflammatory phase, angiogenesis, extracellular matrix secretion and epithelialization). Methodology/Principal Findings Platelet lysate effect was studied on endothelial cells, monocytes, fibroblasts and keratinocytes, in terms of viability and proliferation, migration, angiogenesis, tissue repair pathway activation (ERK1/2) and inflammatory response evaluation (NFκB). Results were compared both with basal medium and with a positive control containing serum and growth factors. Platelet lysate induced viability and proliferation at the highest concentrations tested (10% and 20% v/v). Whereas both platelet lysate concentrations increased cell migration, only 20% platelet lysate was able to significantly promote angiogenic activity (p<0.05 vs. control), comparably to the positive control. Both platelet lysate concentrations activated important inflammatory pathways such as ERK1/2 and NFκB with the same early kinetics, whereas the effect was different for later time-points. Conclusion/Significance These data suggest the possibility of using allogeneic platelet lysate as both an alternative to growth factors commonly used for cell culture and as a tool for clinical regenerative application for wound healing

Two-color fluorescence detection of Poly (ADP-Ribose) Polymerase-1 (PARP-1) cleavage and DNA strand breaks in etoposide-induced apoptotic cells

During apoptosis, the nuclear enzyme Poly(ADPRibose) Polymerase-1 (PARP-1) catalyzes the rapid and transient synthesis of poly(ADP-ribose) from NAD+ and becomes inactive when cleaved by caspases. The regulation of these two opposite roles of PARP-1 is still unknown. We have recently investigated PARP-1 activation/degradation in Hep-2 cells driven to apoptosis by actinomycin D. In the present work, we have extended our analysis to the effect of the DNA damaging agent etoposide, and paid attention to the relationship between PARP-1 cleavage and DNA fragmentation. An original fluorescent procedure was developed to simultaneously identify in situ the p89 proteolytic fragment of PARP-1 (by immunolabeling) and DNA degradation (by the TUNEL assay). The presence of p89 was observed both in cells with advanced signs of apoptosis (where the PARP-1 fragment is extruded from the nucleus into the cytoplasm) and in TUNEL-negative cells, with only incipient signs of chromatin condensation; this evidence indicates that PARP-1 degradation in etoposide-treated apoptotic cells may precede DNA cleavage