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Biomedical subjects

L Pellegrini

Publications and source records attributed to L Pellegrini.

At least 37 records · Page 2Linked to original sources
PubMed1999

Interaction of Alzheimer's presenilin-1 and presenilin-2 with Bcl-X(L). A potential role in modulating the threshold of cell death.

The familial Alzheimer's disease gene products, presenilin-1 and presenilin-2, have been reported to be functionally involved in amyloid precursor protein processing, notch receptor signaling, and programmed cell death or apoptosis. However, the molecular mechanisms by which presenilins regulate these processes remain unknown. With regard to the latter, we describe a molecular link between presenilins and the apoptotic pathway. Bcl-X(L), an anti-apoptotic member of the Bcl-2 family was shown to interact with the carboxyl-terminal fragments of PS1 and PS2 by the yeast two-hybrid system. In vivo interaction analysis revealed that both PS2 and its naturally occurring carboxyl-terminal products, PS2short and PS2Ccas, associated with Bcl-X(L), whereas the caspase-3-generated amino-terminal PS2Ncas fragment did not. This interaction was corroborated by demonstrating that Bcl-X(L) and PS2 partially co-localized to sites of the vesicular transport system. Functional analysis revealed that presenilins can influence mitochondrial-dependent apoptotic activities, such as cytochrome c release and Bax-mediated apoptosis. Together, these data support a possible role of the Alzheimer's presenilins in modulating the anti-apoptotic effects of Bcl-X(L).

Alzheimer Disease↗
PubMed1999

Alternative, non-secretase processing of Alzheimer's beta-amyloid precursor protein during apoptosis by caspase-6 and -8.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder. Although the pathogenesis of AD is unknown, it is widely accepted that AD is caused by extracellular accumulation of a neurotoxic peptide, known as Abeta. Mutations in the beta-amyloid precursor protein (APP), from which Abeta arises by proteolysis, are associated with some forms of familial AD (FAD) and result in increased Abeta production. Two other FAD genes, presenilin-1 and -2, have also been shown to regulate Abeta production; however, studies examining the biological role of these FAD genes suggest an alternative theory for the pathogenesis of AD. In fact, all three genes have been shown to regulate programmed cell death, hinting at the possibility that dysregulation of apoptosis plays a primary role in causing neuronal loss in AD. In an attempt to reconcile these two hypotheses, we investigated APP processing during apoptosis and found that APP is processed by the cell death proteases caspase-6 and -8. APP is cleaved by caspases in the intracellular portion of the protein, in a site distinct from those processed by secretases. Moreover, it represents a general effect of apoptosis, because it occurs during cell death induced by several stimuli both in T cells and in neuronal cells.

Alzheimer Disease↗
PubMed1999

Cloning of AIP1, a novel protein that associates with the apoptosis-linked gene ALG-2 in a Ca2+-dependent reaction.

ALG-2 is a 22-kDa calcium-binding protein necessary for cell death induced by different stimuli in 3DO T-cell hybridoma. 3DO cell clones depleted of ALG-2 protein exhibit normal caspases activation, suggesting that ALG-2 function is required downstream or is independent of caspase proteases activity for apoptosis to occur. Using the yeast two-hybrid screening system, we have isolated and characterized the mouse cDNA encoding for ALG-2 interacting protein 1 (AIP1), a novel protein that interacts with ALG-2. ALG-2 and AIP1 colocalize in the cytosol and the presence of calcium is an indispensable requisite for their association. Sequence alignment shows that AIP1 is highly similar to BRO1, a yeast protein related to components of the Pkc1p-MAP kinase cascade. Overexpression of a truncated form of AIP1 protects two different cell types from death induced by trophic factors withdrawal; thus, our data indicate that AIP1 cooperates with ALG-2 in executing the calcium-dependent requirements along the cell death pathway.

Amino Acid Sequence↗
PubMed1999

Shock waves (SW) noninvasive extracorporeal thrombolysis treatment (NISWT).

A group of 24 patients were considered for noninvasive shock waves thrombolysis (NISWT). Of these, 15 patients gave their informed consent. NISWT was attempted in eight patients (while seven patients were randomized for follow-up only). NISWT was possible in six of seven patients. In one patient randomized for NISWT, local inguinal scarring, due to previous surgery, made impossible the visualization of the femoral vein, and therefore focusing of shock waves (SWs). No side effects were reported in the days after SWs administration during the 4-month follow-up. In patients treated with NISWT it was possible to observe just after the SWs session the presence of echolucent "acoustic holes" and flow (by color and power Doppler) within the "holes." All "echolucent holes" produced at the first session were still present at 4 months, and color flow imaging also detected new flow channels in echogenic areas of thrombi previously not visible. In one patient thrombolysis was achieved after the first treatment, but at 3 and 4 months the thrombus was completely avascular. In conclusion, thrombolysis using SWs was obtained in selected cases and it was still persisting at 4 months in six of the seven treated patients. NISWT appears feasible and promising. These results should be confirmed by larger, prospective trials.

Adult↗
PubMed1998

Efficacy of a new topical gel-spray formulation of ketoprofen lysine salt in the rat: percutaneous permeation in vitro and in vivo and pharmacological activity.

The aim of this study was to evaluate the percutaneous permeation of a new topical Gel-Spray formulation, containing 15% of ketoprofen lysine salt (KLS), both in vitro, using the Franz-type diffusion cells and in vivo, by evaluating urinary recovery after topical administration and to correlate the absorption data with KLS pharmacological activity in the rat. Concentrations of ketoprofen free acid (KFA) were determined by HPLC in the receptor compartment (in vitro), or in urine (in vivo). The permeation of ketoprofen evaluated in vitro after the application of KLS Gel-Spray was higher than that observed with the marketed formulation Profénid gel (containing KFA at 2.5%). The same evidence was found in vivo, except when the ratio between the administered dose and the area treated was higher than 1 mg cm-2. Thus, the difference between the two formulations seems to be the resultant of two opposing components: a positive gradient of concentration that favours the absorption of ketoprofen from KLS Gel-Spray and the presence of the enhancer ethanol that could favour the efficacy of Profénid gel. Under our conditions the former prevailed. As for the efficacy, evaluated in the carrageenan-induced oedema and hyperalgesia model, KLS Gel-Spray confirmed the data obtained for in vivo absorption, being more efficient than the reference standard Profénid gel. The observed inhibitory effects were due only to dermal absorption, oral absorption was excluded by an Elizabethan collar applied around the neck of the rat. In these experimental conditions, no significant damage of the rat stomach mucosa was observed. These results indicate that KLS Gel-Spray, due to its high KLS concentration, allows a very high efficiency in delivering ketoprofen to the inflamed area using a minimal amount of formulation, even in the absence of permeation enhancers.

Animals↗
PubMed1998

Protein interactions implicated in neurotransmitter release.

Biochemical evidence indicates that the exocytotic release of neurotransmitters involves both evolutionary conserved membrane proteins, the SNAREs, as well as ubiquitous cytosolic fusion proteins, NSF and SNAPs. We have analyzed the biochemical properties and the physiological effects of these proteins. Our data suggest models how NSF, SNAPs and SNAREs may function in neurotransmitter exocytosis.

Adenosine Triphosphatases↗
PubMed1998

Differential contribution of R and S isomers in ketoprofen anti-inflammatory activity: role of cytokine modulation.

Among nonsteroidal anti-inflammatory drugs (NSAIDs), 2-arylpropionic acids exist as a racemic mixture of its enantiomeric forms, with S-enantiomers primarily responsible for inhibition of prostaglandin synthesis and of inflammatory events. The aim of this study was to compare the anti-inflammatory effects of R- and S-ketoprofen in vitro and in vivo. S-Ketoprofen efficiently inhibited carrageenan-induced edema formation, but it could also amplify the LPS-induced production of the inflammatory cytokines tumor necrosis factor (TNF) and interleukin-1 (IL-1), in close correlation with its ability to inhibit prostaglandin synthesis. Because these inflammatory cytokines are among the factors involved in carrageenan-induced inflammation and also are possibly involved in gastric damage, enhanced cytokine production could partially mask the analgesic effect of S-ketoprofen, and it can be associated with the clinical evidence of its gastric toxicity. On the other hand, R-ketoprofen contributes to the overall activity of the racemate, by playing the main role in ketoprofen-induced analgesia. Unlike the S-isomer, R-ketoprofen did not induce a significant increase of cytokine production even at cyclooxygenase-blocking concentrations. It is concluded that the R-isomer directly contributes to the anti-inflammatory effects of ketoprofen, being more analgesic, and because it does not amplify inflammatory cytokine production.

Animals↗
PubMed1997

Analysis of the methylation pattern of the maize opaque-2 (O2) promoter and in vitro binding studies indicate that the O2 B-Zip protein and other endosperm factors can bind to methylated target sequences.

The maize opaque-2 locus (o2) has an endosperm-specific expression and is positively autoregulated by its gene product, a b-Zip protein, to a TGACGTTG motif. The genomic sequencing method was used here to describe, in leaf and endosperm, the methylation pattern of a 390-base pair region of the o2 promoter. In leaf, 96% of the C residues are methylated, whereas in endosperm the 5-methylcytosine content is 84%. Comparison of these methylation patterns indicates that the o2 tissue-specific expression does not result from the demethylation of any specific C residue and that, in vivo, O2 interacts with a methylated target sequence. Consistently, gel-shift experiments using a CpG-methylated, partially methylated, and hemimethylated o2 promoter fragments showed that, in vitro, the O2 protein binds to the major groove of a methylated target sequence, although its binding activity decreases at increasing levels of C-methylation and is more effectively reduced by methylation of the lower strand than of the upper strand of the DNA. Using partially purified endosperm cell extracts, we also show that, besides the O2 protein, other proteins specifically bind to a partially methylated o2 promoter fragment, therefore indicating that in plants a subset of different proteins may mediate the expression of a naturally occurring methylated o2 promoter.

Base Sequence↗
PubMed1997

Tartronates: a new generation of drugs affecting bone metabolism.

In the search for a new class of bone-sparing agents for treating osteopenic disorders, we hypothesized that tartronic acid derivatives, sharing the chemical characteristics both of bisphosphonates and of Gla residues contained in matrix proteins such as osteocalcin, could positively affect bone metabolism. A series of tartronates was therefore tested for their ability to affect bone metabolism. In vitro resorption tests were performed examining pit formation by freshly isolated rat and rabbit osteoclasts plated onto bone slices and exposed to the drugs for 48 h. Tartronates bearing a linear side-chain (DF 1222 and DF 1363A) were the most effective in inhibiting pit excavation in the pM-nM range. Tartronates did not affect osteoclast viability, number, adhesion, or tartrate resistant acid phosphatase activity. Transient cell retraction was observed in osteoclasts plated onto glass and exposed to DF 1222. The maximal effect was seen in cells treated for 4 h at a concentration of 1 pM. DF 1222 accelerated mineralization in cultures of periosteal cells without affecting other osteoblast-like functions. This product was therefore tested in vivo in ovariectomized mice. Bone mass in femur was evaluated, by ash gravimetry, 21 days after ovariectomy. Unfortunately, DF 1222, the most active of tartronates in vitro, was inactive in this test because of its high hydrophilicity and the subsequent too short residence time. On the contrary, its tetrahydropyranyl ether derivative, DF 1363A, endowed with a significantly higher lipophilicity, showed a dose-dependent bone-sparing effect when administered subcutaneously at 10, 30, and 100 mg/kg/die, thus confirming the activity seen in in vitro tests. Because of their feasible parallel effect on both bone resorption and formation, tartronate derivatives may be tested to candidate this class of products for clinical studies.

Animals↗
PubMed1997

[Lipoprotein(a), parameters of lipid metabolism and hemostasis in obese patients].

BACKGROUND: The prevalence of obesity is increasing in the population of industrialized countries and this condition is today considered a risk factor for cardiovascular diseases because, at least partly, it represents a prothrombotic state. METHODS: For this reason we have studied plasma lipid concentrations (C, TG, HDL-C), Lp(a) and some parameters of haemostasis (PAI-1, t-PA, D-dimer) in 41 non diabetic obese patients (38 females and 3 males) and in 36 healthy normal weight subjects. RESULTS: Lipid pattern has resulted in overlapping both in the two studied populations and in the two subgroups of pre- and postmenopausal women, while greater concentrations of Lp(a), (p < 0.001) and t-PA (p < 0.05) have been found in obese populations vs controls, but not different between the two subgroups; in particular, Lp(a) has resulted > 30 mg/dl in 34.14% of obese patients and in no case in the control group. Finally, Lp(a) concentrations have been found for the first time to be positively related to C and negatively to HDL-C in the obese population. CONCLUSIONS: As Lp(a) is believed to be a pathogenetic linkage between atherogenesis and thrombosis and can be affected by weight loss, the efficacy of the reduction of high plasma levels of Lp(a) needs to be considered with longitudinal studies.

Adult↗
PubMed1996

[Blood glucose and insulin responses to oral glucose in hyperthyroidism].

Because thyroid hyperfunction "per sè" can induce impaired glucose tolerance in 2-57% of patients, we have studied glycaemic and insulinaemic response to oral glucose load (OGTT) in this condition. Glycaemic and insulinaemic curves after oral glucose (100 g), the respective secretory areas (AUC-G and AUC-IRI) have been determined in 13 hyperthyroid patients (8 Graves disease, 4 multinodular toxic goiters and 1 Plummer's adenoma) and in 16 healthy control subjects. AUC-G, AUC-IRI and median values of G and IRI in most of considered times have resulted significantly greater in hyperthyroid patients. This study has confirmed the impact of thyroid hyperfunction on carbohydrate metabolism, but has not proved an impaired insulin secretion as previously reported in Literature. In conclusion, we believe that: a) hyperthyroidism is associated with greater circulating levels of G and IRI, b) hyperinsulinaemia in addition to hyperglycaemia suggests that insulin-resistance is underlying in this condition.

Administration, Oral↗
PubMed1995

Mapping of receptor binding sites on IL-1 beta by reconstruction of IL-1ra-like domains.

Upon structure comparison between IL-1 beta and its antagonist IL-1ra, single or multiple residues along the IL-1 beta sequence were replaced with the corresponding amino acids present in the IL-1ra protein, in the attempt to identify sites important for receptor binding and for biologic activity on the two molecules. Ten of fifteen mutant proteins had activity comparable to that of wild-type IL-1 beta in three different biologic assays and in receptor binding, indicating that the introduced changes did not influence the functional structure of the protein. Conversely, three mutants (SMIL-9: 127/263 R/T-->W/Y; SMIL-10: 125/127/263/265 T/R/T/Q-->R/W/Y/E; SMIL-15:222/227 I/E-->S/S) showed an increased binding capacity for IL-1RI, not paralleled by increased agonist activity, indicating that the introduced IL-1ra residues could be involved in the nonagonist IL-1RI binding site. On the other hand, two mutants showed diminished binding capacity with concomitant decrease in biologic activity. Both mutants (SMIL-1, five substitutions in the loop 202-214; and SMIL-3, total replacement of the loop 164-173 with the IL-1ra stretch 52-55) included substitutions of residues allegedly important for agonist binding to IL-1RI. Mutant SMIL-3 showed the most profound reduction in binding capacity for IL-1RI (CDw121a) and a more than 1,000-fold reduced biologic activity both in vitro and in vivo, but it retained full capacity of binding to IL-1RII (CDw121b) and acted as a selective antagonist of IL-1RII. From these results the following conclusions can be drawn. IL-1 beta binds to IL-1RI and to IL-1RII through different sites, and the loop 164-173 appears as one of the areas involved in the selective interaction with IL-1RI. Agonist (IL-1 beta) and nonagonist (IL-1ra) binding to IL-1RI occur through distinct sites, with loops 164-173 and 202-214 of IL-1 beta identified as two of the sites selectively involved in agonist binding to the activating receptor.

Animals↗
PubMed1995

Structure of serum response factor core bound to DNA.

The human serum response factor is a transcription factor belonging to the MADS domain protein family with members characterized from the plant and animal kingdoms. The X-ray crystal structure of the serum response factor core in a specific-recognition DNA complex shows that the functions of DNA binding, dimerization and accessory-factor interaction are compactly integrated into a novel protein unit. The intrinsic and induced conformation of the serum response element DNA is the principal DNA feature recognized in the specific complex.

Amino Acid Sequence↗
PubMed1994

Phenylalanine ammonia-lyase in tobacco. Molecular cloning and gene expression during the hypersensitive reaction to tobacco mosaic virus and the response to a fungal elicitor.

A tobacco (Nicotiana tabacum L. cv Samsun NN) cDNA clone coding the enzyme phenylalanine ammonia-lyase (PAL) was isolated from a cDNA library made from polyadenylated RNA purified from tobacco mosaic virus (TMV)-infected leaves. Southern analysis indicated that, in tobacco, PAL is encoded by a small family of two to four unclustered genes. Northern analysis showed that PAL genes are weakly expressed under normal physiological conditions, they are moderately and transiently expressed after wounding, but they are strongly induced during the hypersensitive reaction to TMV or to a fungal elicitor. Ribonuclease protection experiments confirmed this evidence and showed the occurrence of two highly homologous PAL messengers originating from a single gene or from two tightly co-regulated genes. By in situ RNA-RNA hybridization PAL transcripts were shown to accumulate in a narrow zone of leaf tissue surrounding necrotic lesions caused by TMV infection or treatment with the fungal elicitor. In this zone, no cell specificity was observed and there was a decreasing gradient of labeling from the edge of necrosis. Some labeling was also found in various cell types of young, healthy stems and was shown to accumulate in large amounts in the same cell types after the deposition of an elicitor solution at the top of the decapitated plant.

Amino Acid Sequence↗