Inventory of landslides triggered by an extreme rainfall event in Marche-Umbria, Italy, on 15 September 2022.

Systematic and timely documentation of triggered (i.e. event) landslides is fundamental to build extensive datasets worldwide that may help define and/or validate trends in response to climate change. More in general, preparation of landslide inventories is a crucial activity since it provides the basic data for any subsequent analysis. In this work we present an event landslide inventory map (E-LIM) that was prepared through a systematic reconnaissance field survey in about 1 month after an extreme rainfall event hit an area of about 5000 km2 in the Marche-Umbria regions (central Italy). The inventory reports evidence of 1687 triggered landslides in an area of ~550 km2. All slope failures were classified according to type of movement and involved material, and documented with field pictures, wherever possible. The database of the inventory described in this paper as well as the collection of selected field pictures associated with each feature is publicly available at figshare.

Antisense oligonucleotide drug design.

Maneuvering single gene expression is not only an optimal way to study gene function but also an ambitious goal, which will lead to the treatment of a variety of human diseases whose main pathogenetic event is a genetic alteration. The recent efforts focusing on the genome project have led to array based, high throughput, gene expression analysis techniques that allow the study of complex molecular networks. Combining these powerful new technologies with modulation of gene expressions is making it possible to unravel complex molecular networks or, vice versa, to find new gene products responsible for pathological conditions on which exogenous modulation can be productive. Efficient and specific modulation of gene expression can be obtained either by producing transgenic or gene knockout organisms or cells (gene targeting), or by treating organisms or cells with short synthetic nucleic acid segments in antisense orientation with respect to the targeted mRNAs (mRNA targeting by antisense strategy). While genome manipulation is a time consuming and expensive approach, requiring invasive intervention, the "antisense strategy" is characterized by high flexibility resulting from safeness, specificity, reversibility, modulability, and low cost. The rationale of the antisense strategy is that, once one gene sequence is known, its expression can be silenced by application of synthetic single-strand nucleic acid segments (oligonucleotides) whose sequence is in antisense orientation compared to the targeted mRNA. Recently, this "informational" strategy has been boosted by the discovery of the RNA interference: a natural mechanism by which cells are thought to fight detrimental exogenous viruses and endogenous transposons. Despite promising futures, antisense-based therapeutics are far from being an established reality. This review analyses the recent improvements in antisense-based gene expression modulation, focuses on the treatment of diseases in the light of the past, and provides our personal findings on this topic.

The mechanisms of apoptosis in biology and medicine: a new focus for ophthalmology.

Defects in apoptosis (programmed cell death) have recently emerged as being closely involved in the pathogenesis of most ocular diseases and, therefore, apoptosis is now a topic of exponential interest in ophthalmology. This review summarizes recent works on mechanisms of apoptosis, from its initiation and modulation to the switching-on of its execution machinery. Interactions of cell death with cell division programs to orchestrate ontogenesis, aging, and adult life and their alterations in human diseases are pointed out. Two main apoptotic signaling pathways are identified: a death receptor-dependent (extrinsic) pathway and a mitochondrion-dependent (intrinsic) pathway. Mitochondrion harbors both antiapoptotic (Bcl-2, Bcl-XL) and apoptotic factors (Smac/Diablo, Apaf-1, cytochrome c). Its permeability transition pore (mPTP) is the main trigger of cell suicide. The process of mPTP opening, in association with extrusion to cytoplasm of a variety of apoptotic factors, is shown. Cytochrome c is one of these apoptotic factors. When expelled to cytoplasm, this double-faced respiratory chain component assembles with two other modules, Apaf-1 and procaspase 9, to form a protein complex--the apoptosome--that starts apoptosis execution. Another respiratory chain component, the CoQ10, is believed to counteract mPTP opening. What makes apoptosis particularly exciting for medicine is that its dysfunctions play a central role in the pathogenesis of several human diseases. For instance, excesses of apoptosis lead to cell loss that accompanies neurodegenerative diseases, whereas genetically determined defects of apoptosis lead to the deregulated cell proliferation typical of cancer. A variety of ophthalmologic diseases, such as post-keratectomy haze, corneal lesions, cataract, glaucoma, senile maculopathies, and genetic ocular pathologies, that underlie apoptosis dysfunctions are treated in detail in the other reviews of this issue.

Apoptosis is associated with modifications of bcl-2 mRNA AU-binding proteins.

The expression of genes requiring finely tuned control is regulated by a posttranscriptional mechanism involving mRNA A + U-rich elements (AREs) cooperating with ARE-binding proteins (AUBPs) in modulation of mRNA stability. We reported previously that an ARE in the bcl-2 mRNA 3'-untranslated region (3'-UTR) had destabilizing activity and was involved in bcl-2 downregulation during apoptosis in vitro. Here we demonstrate that the bcl-2 ARE complexes with a number of specific AUBPs, whose pattern undergoes changes following application of apoptotic stimuli. The caspase inhibitor Z-VAD-fmk strongly attenuates both bcl-2 mRNA decay and bcl-2 AUBP pattern changes elicited by apoptotic stimuli, indicating the involvement of bcl-2 AUBPs in bcl-2 mRNA stability control.

Prevention of corneal keratocyte apoptosis after argon fluoride excimer laser irradiation with the free radical scavenger ubiquinone Q10.

PURPOSE: To assess in vitro the potential of the free radical scavenger ubiquinone Q10 in preventing keratocyte apoptosis after argon fluoride (ArF) excimer laser irradiation. METHODS: Cultured rabbit keratocytes were irradiated at very low single-pulse laser fluences. The cumulative effects generated by three total fluence doses between 12 and 45 mJ/cm2, representative of single-pulse subablative doses during photorefractive keratectomy (PRK) in humans, were evaluated. We employed the following parameters to compare pretreated (10 microM ubiquinone Q10) and untreated samples: 1) number and morphology of living cells by Trypan blue test and ultramicroscopy, respectively; 2) level of free-radical formation assessed by malonaldehyde quantitation; 3) cellular energy level evaluated by ATP assay. RESULTS: Excimer laser irradiation kills cultured keratocytes by inducing apoptosis. The effect increases with the cumulative fluence dose. In the samples pretreated with ubiquinone Q10 there were significantly fewer cumulative apoptotic events than in the untreated ones. Quantitative analysis of malonaldehyde cellular levels suggested this protective action of ubiquinone Q10 was connected with its ability to scavenge laser-generated free radicals. ATP assay also confirmed that it raised cellular energy levels. CONCLUSIONS: The treatment of corneal keratocytes with relatively low concentrations of ubiquinone Q10 can prevent apoptosis after ArF excimer laser irradiation. If these findings are confirmed on human keratocytes this treatment could be usefully exploited in the PRK surgical procedure. That might lead to a reduction in the occurrence of haze and curvature regression triggered by programmed cell death.

A conserved AU-rich element in the 3' untranslated region of bcl-2 mRNA is endowed with a destabilizing function that is involved in bcl-2 down-regulation during apoptosis.

The control of mRNA stability is becoming recognized as a crucial point of gene expression regulation. A common element responsible for mRNA decay modulation is the adenine- and uracil-rich element that is found in the 3' untranslated region of numerous mRNAs subjected to fast expression changes in response to various stimuli. Previously we identified a post-transcriptional regulation level for the antiapoptotic bcl-2 gene, which could be involved in t(14;18) lymphoma-associated bcl-2 overexpression. Here we demonstrate that bcl-2 mRNA is endowed with an adenine- and uracil-rich element (ARE) characterized by high evolutionary conservation not only among all chordates examined, but even between chordates and the nematode Caenorhabditis elegans (ced-9 gene). As for other well-established destabilizing AREs, the insertion of the bcl-2 ARE downstream from stable beta-globin mRNA causes an enhanced decay of the beta-globin transcript, which proves its functional role. This possibility is corroborated by the fact that the pathway leading to the modulating activity of bcl-2 ARE is influenced by PKC, since the addition of DAG and TPA markedly attenuated the bcl-2 ARE destabilizing potential. Conversely, it is noteworthy that when C(2)-ceramide is added to the culture medium as the apoptotic agent, the beta-globin transcript harboring the bcl-2 ARE undergoes a dramatic increase in decay. This observation clearly indicates that the destabilizing function of bcl-2 ARE is enhanced by apoptotic stimuli and suggests that this element could be involved in a post-transcriptional mechanism of bcl-2 down-regulation during apoptosis. The half-life of the mRNA of bcl-2 in Jurkat cells is prolonged by PKC stimulation and shortened by C(2)-ceramide addition, strongly supporting the view that bcl-2 mRNA stability plays a physiological role in modulating bcl-2 expression, particularly in its down-regulation during apoptosis. Thus, this element becomes a new candidate for mediating those bcl-2 gene expression changes-from apoptosis-associated down-regulation to tumor-associated overexpression-observed thus far that profoundly influence single cell fate and tissue homeostasis.

Induction of apoptosis and mitosis inhibition by degraded DNA lipotransfection mimicking genotoxic drug effects.

Genotoxic damage induces cell cycle arrest and/or apoptosis by activation of p53 oncosuppressor protein. A number of anticancer drugs are genotoxic and their damaging effect upon cells is mediated by this mechanism. Microinjection of defined DNA species directly into nucleus has been reported previously to activate p53 and inhibit cell cycle. Here, we demonstrate that simple addition of heterogeneous degraded DNA to cultured cells (Rat-1 fibroblasts) in combination with lipotransfecting agent DOTAP leads to apoptosis induction and mitosis inhibition by a molecular mechanism which mimics that of the cellular response to genotoxic anticancer agents. Indeed, both cellular effects induced by lipotransfected degraded DNA (essentially, heterogeneous small DNA fragments) are associated to p53 activation and modulated by two apoptosis-related genes, such as bcl-2 and c-myc, which also modulate the apoptotic threshold to anticancer agents. Here we raise the hypothesis of exogenous DNA segment lipotransfection as possible new tool for anticancer therapy.

Effects of moclobemide on depressive symptoms and cognitive performance in a geriatric population: a controlled comparative study versus imipramine.

Moclobemide, a novel monoamine oxidase-A reversible inhibitor with demonstrated antidepressive efficacy, was administered double-blind versus imipramine to aged depressive subjects. The two drugs were given for 60 days in increasing doses up to 600 mg for moclobemide and 100 mg for imipramine. Fifteen patients received moclobemide and 15 received imipramine. Psychiatric conditions and symptoms were rated at 0, 7, 14, 30, 45, and 60 days after the beginning of the trial by means of the Scale for the Assessment of Psychoorganic Syndromes, Hamilton Rating Scale for Depression, Rome. Depression Inventory, Hamilton Anxiety Rating Scale, State-Trait Anxiety Inventory-X form, and the Clinical Global Impression Scale. Cognition was tested through the Benton visual retention test at days 0, 30, and 60 and the Digit Substitution Test of the Wechsler Adult Intelligence Scale at days 0 and 60. Side effects were assessed through the Dosage Record Emergent Symptoms at days 0, 7, 14, 30, 45, and 60. The dropout rate was significantly greater in the moclobemide group. Both drugs induced an improvement in depressive and anxious symptomatology, with moclobemide showing a faster onset. Furthermore, moclobemide showed an enhancing effect on cognition, which was not shown by imipramine. Such results indicate that moclobemide could prove to be the drug of choice in geriatric depression, given that cognitive effects are prominent in the aged.

A novel pharmacological approach for paraquat poisoning in rat and A549 cell line using ambroxol, a lung surfactant synthesis inducer.

Paraquat (PQ) is a widely used herbicide that causes acute adult respiratory distress syndrome (ARDS) and chronic lung damage (diffuse fibrosis). One of the earliest biochemical effects induced by PQ is damage to type II pneumocytes with consequent depletion of surfactant. With the aim of counteracting the toxic effects of PQ, a series of investigations were performed into the possible protective effect of the drug ambroxol, which induces the synthesis of surfactant in lung alveolar type II cells. The number of survivors and survival time of rats treated ip with 35 mg PQ/kg was significantly increased by 3 days of ambroxol pretreatment and by ambroxol treatment 30 min or 2 hr after PQ. Total phospholipid content in lung and bronchoalveolar lavage fluid (BALF) was significantly reduced 30 hr after treatment with PQ alone. The association of ambroxol with PQ significantly antagonized this reduction. In BALF the ratio between palmitic acid and stearic acid concentrations was significantly lower in animals treated with PQ alone but was returned to normal by the association with ambroxol. The cell line A549, exposed in vitro to PQ concentrations from 0.5 x 10(-4) to 2 x 10(-3) M, showed a significant dose-dependent loss of viability. Cells pretreated with ambroxol (10 mg/ml) were more resistant to PQ and their viability started to decrease significantly only from a PQ concentration of 0.8 x 10(-3) M. Membrane microviscosity was measured on the same cells. Cells treated with PQ alone showed a reduction of membrane microviscosity, which was significantly counteracted by ambroxol pretreatment. The curves of modification of membrane microviscosity of cells treated with PQ and with ambroxol plus PQ paralleled those of cell viability, indicating that the stimulation of surfactant synthesis in vitro may be a prerequisite for counteracting some of the early effects of PQ.

Some properties of the alkaline proteinase from Aspergillus melleus.

Seaprose is a semi-alkaline proteinase produced by Aspergillus melleus. The aim of our study was to further characterize the properties of this enzyme, particularly looking at its interaction with alpha 1-proteinase inhibitor, the major human plasma proteinase inhibitor. We studied the cleavage of three synthetic peptide substrates induced by seaprose and the inhibitory profile of the enzyme by means of a panel of inhibitors, including alpha 1-proteinase inhibitor. The interaction between seaprose and alpha 1-proteinase inhibitor was also studied with SDS-PAGE. Finally, the elastolytic activity of seaprose was checked by means of bovine elastin solubilization. We found that seaprose cleaves preferentially the substrate containing a Phe residue in the P1 position. The inhibitory profile showed that seaprose is a serine-proteinase that cannot be inhibited by alpha 1-proteinase inhibitor. The SDS-PAGE revealed that alpha 1-proteinase inhibitor, after incubation with seaprose, underwent a limited proteolysis. Finally, seaprose 10(-2) M and 10(-3) M was able to solubilize bovine elastin. We conclude that seaprose is a serine-proteinase able to inactivate human alpha 1-proteinase inhibitor with limited proteolysis at (or near) the active site and that it has mild elastinolytic capacity.

Protection against acute paraquat toxicity by ambroxol.

One of the earliest biochemical effects induced by the herbicide paraquat (PQ) is damage to type II pneumocytes with consequent depletion of surfactant (Skillrud and Martin, 1984). We made a series of studies on the possible protective effect of drug ambroxol, which induces surfactant synthesis from alveolar type II cells (Post et al. 1983). The cell line A-549, exposedin vitro to PQ concentrations ranging from 0.5×10(-4) to 2×10(-3) M, showed a significant dose-dependent loss of viability. Ambroxol (10 mg/ml) pretreated cells were more resistant to PQ, their viability starting to decrease from a PQ concentration of 0.8×10(-3) M. Membrane microviscosity was measured on the same cells. Cells treated with PQ alone showed a reduction of membrane microviscosity which was significantly counteracted by ambroxol pretreatment. The curves for membrane microviscosity of PQ and ambroxol-plus-PQ-treated cells overlapped those for cell viability, indicating that the stimulation of surfactant synthesisin vitro may be a prerequisite for counteracting some of the precocious effects of PQ. Partial protection from PQ- induced mortality was also obtainedin vivo.

Alpha 1-antitrypsin variants produced by recombinant DNA: differences in elastase inhibitory activity and resistance to oxidant agents.

Inherited or "acquired" deficiency of alpha 1-antitrypsin (believed to be the cause of pulmonary emphysema) will probably be treated in the future by replacement with alpha 1-antitrypsin purified from human plasma or produced by recombinant DNA, which seems promising because it permits site-specific mutagenesis in the oxidizable active site of the normal human alpha 1-antitrypsin. The aim of this in-vitro study was to investigate the elastase inhibitory activity and the resistance to oxidizing agents of normal human alpha 1-antitrypsin, a recombinant yeast-produced variant (VAL 358) and a recombinant E. coli-produced variant (LEU 358). The inhibitors were exposed to chemical oxidants (NCS, H2O2, xanthine/xanthine oxidase, chloramine-T) and to PMA-activated neutrophils. The elastase inhibitory activity was assayed on porcine pancreatic elastase and neutrophil elastase. Normal alpha 1-antitrypsin and VAL 358 variant were good inhibitors of both elastases. LEU 358 variant was the best inhibitor for neutrophil elastase, but it poorly inhibited the porcine pancreatic elastase. Normal alpha 1-antitrypsin was affected by all oxidants; both variants were almost totally resistant to chemical oxidants and to activated neutrophils. We conclude that recombinant alpha 1-antitrypsin variants differ in their elastase inhibitory activity and offer increased resistance to oxidant agents.

Studies of MR 889, a new synthetic proteinase inhibitor.

We investigated the proteinase inhibitory activity of MR 889, a thiolactic acid derivative. It is able to in vitro inhibit at low concentration (10(-5),10(-6)M) the activity of porcine pancreatic elastase, human neutrophil elastase and bovine chymotrypsin. In addition, MR 889 is able to inhibit the residual activity of alpha 2-macroglobulin-trapped human neutrophil elastase, paralleling the efficacy of phenylmethylsufonylfluoride. Finally, MR 889 has been shown to in vitro reduce the burden of elastase- and chymotrypsin-like activity found in sputum sol-phases of patients admitted for chronic bronchitis exacerbation.

Relationship between changes in alveolar surfactant levels and lung defence mechanisms.

Pulmonary surfactant, besides its mechanical properties, is thought to be involved in lung defence mechanisms. We previously described that: (1) in healthy animals, surfactant synthesis stimulation with ambroxol was accompanied by alveolar macrophage activation and a shift of the alveolar elastase/antielastase balance towards increased antielastase activity, and (2) in bleomycin-treated rats alveolar phospholipidosis was obvious 14 days after drug administration, ambroxol protection reduced the phospholipid peak and the morphological apperance of lung fibrosis at the 14th day of the experiment. The present study found that: (1) in healthy rats, the ambroxol-induced increase of alveolar antielastase activity did not appear due to reactivation of alpha 1-antitrypsin normally oxidized in the alveolar milieu; (2) in bleomycin-induced pulmonary fibrosis, ambroxol protection reduced total long collagen content at day 28, and (3) in paraquat-induced pulmonary fibrosis, alveolar phospholipids were markedly reduced throughout the 21 days of the experiment. On increasing the dose of paraquat, ambroxol protection significantly reduced the animals' death rate.