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Biomedical subjects

R Lorenzi

Publications and source records attributed to R Lorenzi.

18 recordsLinked to original sources
PubMed2001

Cutting edge: the Wiskott-Aldrich syndrome protein is required for efficient phagocytosis of apoptotic cells.

Phagocytosis of apoptotic cells by macrophages and dendritic cells is necessary for clearance of proinflammatory debris and for presentation of viral, tumor, and self Ags. While a number of receptors involved in the cognate recognition of apoptotic cells by phagocytes have been identified, the signaling events that result in internalization remain poorly understood. Here we demonstrate that clearance of apoptotic cells is accompanied by recruitment of the Wiskott-Aldrich syndrome (WAS) protein to the phagocytic cup and that it's absence results in delayed phagocytosis both in vitro and in vivo. Therefore, we propose that WAS protein plays an important and nonredundant role in the safe removal of apoptotic cells and that deficiency contributes significantly to the immune dysregulation of WAS. The efficiency of apoptotic cell clearance may be a key determinant in the suppression of tissue inflammation and prevention of autoimmunity.

Animals↗
PubMed2000

Wiskott-Aldrich syndrome protein is necessary for efficient IgG-mediated phagocytosis.

Interactions between the Wiskott-Aldrich (WAS) protein (WASp), small GTPases, and the cytoskeletal organizing complex Arp2/3 appear to be critical for the transduction of signals from the cell membrane to the actin cytoskeleton in hematopoietic cells. This study shows that Fcgamma-receptor (FcgammaR)-mediated phagocytosis is impaired in WASp-deficient peripheral blood monocytes, and that in macrophages, formation of the actin cup and local recruitment of tyrosine phosphorylated proteins is markedly attenuated. Results also show that, in normal macrophages, WASp itself is actively recruited to the cup, suggesting that assembly of this specialized cytoskeletal structure is dependent on its expression. (Blood. 2000;95:2943-2946)

Actins↗
PubMed2000

The Wiskott-Aldrich syndrome: disordered actin dynamics in haematopoietic cells.

The Wiskott-Aldrich syndrome protein (WASp) is a member of a unique family whose members share similar domain structures and are responsible for the transduction of signals from the cell membrane to the actin cytoskeleton. For WASp, the interactions with Rho family GTPases and the cytoskeletal organising complex Arp2/3 are critical to these functions, which when disturbed translate into abnormalities of haematopoietic cell signaling, polarisation, migration and phagocytosis. This review discusses the evidence for regulation of highly dynamic cytoskeletal structures by WASp and the consequences of disturbed function on some of these processes.

Actins↗
PubMed1999

Functional Fc epsilonRI engagement by a second secretory IgE isoform detected in humans.

We have recently reported that besides the most abundant form epsilonS1, there exists another human secretory epsilon H chain isoform, epsilonS2, resulting from alternative splicing in the epsilonCH4 exon. Using a specific antibody targeted to the epsilonS2-specific C-terminal tailpiece, we now show that this second secretory IgE isoform (IgE-S2) is constitutively co-expressed with the classical secretory IgE-S1 by human myeloma cells. The epsilonS2 variant was also detected in tonsils and in the serum of three non-atopic donors, but was absent in the vast majority of sera of both atopic and non-atopic individuals tested, indicating rare serum expression. IgE-S2 is capable of binding to cells expressing Fc epsilonRI, the high-affinity receptor for IgE. Analysis of intracellular tyrosine phosphorylation signal, degranulation, and rate of receptor internalization suggest a quantitatively lower response by IgE-S2 compared to IgE-S1. The modest differences observed do not appear to overall affect the degranulation competency of IgE-S2, but suggest that the unique structure of the epsilonS2 tailpiece can exert an effect on the interaction with the alpha chain of Fc epsilonRI.

Humans↗
PubMed1999

Sequence-specific antibodies against human IgE isoforms induced by an epitope display system.

BACKGROUND: Unlike other immunoglobulin isotypes, the human C epsilon gene generates by alternative splicing two types of secretory and two types of membrane epsilon chains. The two secreted epsilon heavy chains, epsilon(S1) and epsilon(S2), differ only in the sequence of the last eight C-terminal amino acids, being epsilon(S2) six amino acids longer. The two types of membrane isoforms differ in the extracellular membrane proximal domain, with the longer variant, epsilon(mL), containing 52 extra amino acids which are absent in the shorter epsilon(mS) isoform. OBJECTIVES: We wished to produce quality antibody reagents that specifically detect epitopes that are epsilon isoform-specific. STUDY DESIGN: Short sequences of seven or ten amino acids were chosen as target epitopes and expressed as part of the highly immunogenic loops of deletion variants of engineered Flock House Virus capsid protein RNA2. Chimeric proteins were expressed in E. coli, and used to immunize rabbits. Antisera were screened by immunoblotting of purified IgE isoforms expressed by murine transfectomas. RESULTS: Chimeric proteins expressing epsilon isoform-specific epitopes proved to be strong immunogens in vivo and induced highly specific rabbit antisera. Two antisera so obtained recognize specifically the IgE-S2 isoform. A third one recognizes the long membrane variant m(L)IgE and a fourth one detects an epitope specific to m(S)IgE. CONCLUSION: Here we describe a simplified and efficient protocol of immunization which does not require peptide synthesis and conjugation to carrier protein. Our results show that short peptides of unknown immunogenicity, when genetically introduced into the modified Flock House Virus epitope display system, successfully induced IgE isoform-specific polyclonal antisera in rabbits. These are valuable tools to specifically identify secretory and membrane isoforms of human IgE, and the method is potentially applicable to other variant isoforms or mutants of a given protein.

Amino Acid Sequence↗
PubMed1999

Dentin desensitizing effects of Gluma Alternate, Health-Dent Desensitizer and Scotchbond Multi-Purpose.

PURPOSE: To evaluate the effects of topical applications of Gluma Alternate, a Gluma Desensitizer version with reduced glutaraldehyde content, Health-Dent Desensitizer and Scotchbond Multi-Purpose (SMP) on hypersensitive erosion/abrasion lesions. MATERIALS AND METHODS: 55 patients were included in the trial with at least three teeth each presenting severe sensitivity. From a total of 184 teeth, 69 were treated with Gluma Alternate, 58 with Health Dent and 51 treated SMP, serving as a control. Sensitivity was recorded as response to cold air stimulus prior to treatment, immediately after the topical application of the agents, and after 1 week, 1 month and 6 months. RESULTS: Both Gluma Alternate and SMP showed significant reduction in sensitivity between pre- and postoperative pain scores (P < 0.05). The post-treatment sensitivity scores (0 and 1) were no different between 1 week and 6 months. In the Health-Dent group only 10 of the 58 teeth showed sensitivity reduction. For this reason the remaining 48 teeth received a "rescue treatment" with Gluma Desensitizer. At the end of the 6-month observation time, all teeth treated with Gluma Alternate, Gluma Desensitizer, and SMP showed no or very little dentin sensitivity.

Adult↗
PubMed1998

Retinal identification in Pelvetia fastigiata.

Unidirectional blue light directs the rhizoid-thallus axis in the apolar zygote of the brown alga Pelvetia fastigiata. This effect is mediated by an increase in the intracellular concentration of cGMP. Here, we show the extraction, purification and identification of 1 microgram of all-trans retinal from 1.2 x 10(6) Pelvetia zygotes. The number of retinal molecules per cell was about 4 x 10(9). Since retinal, wherever present, is exclusively associated with an opsin to form a light sensitive complex (rhodopsin-like proteins), and since the physiological response originated by this protein produces a variation of cGMP concentration, this new finding suggests that a rhodopsin-like protein could be the photoreceptor in this brown alga.

Chromatography, High Pressure Liquid↗
PubMed1997

A methylation-dependent DNA-binding activity recognising the methylated promoter region of the mouse Xist gene.

Differential methylation of CpG sites in the promoter region of the mouse Xist gene is correlated with Xist expression and X-chromosome inactivation in the female. Using oligonucleotides encompassing the differentially methylated sites as probes in band-shift assays, we have identified a nuclear protein which binds to a specific region of the promoter (between base pairs -45 and -30 upstream from the transcription start site) only when CpG sites within the CG rich region (GCGCCGCGG, -44 to -36) are methylated. Competition experiments with methylated or unmethylated heterologous oligonucleotides demonstrate that the activity is sequence-specific as well as methylation-dependent. Analysis by Southwestern blot identifies a protein of approximately 100 kDa molecular weight and confirms strong binding to the methylated Xist promoter oligonucleotide. Using a 233bp Xist-promoter luciferase construct in which the cytosines in the three CpG sites in the -44 to -36 region are mutated to thymine, we have established that this region is required for transcription from the mouse Xist promoter. Therefore, we suggest that the binding of the 100kDa protein to the methylated sequence leads to repression of transcription from the methylated Xist allele, thus suggesting a role in the regulation of both imprinted and random Xist transcription and X-chromosome inactivation.

Animals↗
PubMed1997

Laboratory evaluation of compomers in Class V restorations.

PURPOSE: To evaluate the in vitro microleakage, marginal morphology and resin tags configuration of a compomer restorative material (Dyract) alternatively used with two different bonding systems (Dyract-PSA and Prime & Bond 2.0). A hybrid resin composite (Prisma TPH) used with two different bonding systems (Universal Bond 3 and ProBond) was evaluated as control material. MATERIALS AND METHODS: Class V non-retentive restorations were made in vitro at CEJ and divided at random in four groups: Dyract/PSA, Dyract/P&B 2.0, Prisma TPH/UB3, and Prisma TPH/ProBond. The latter two groups were as controls. After finishing the restorations, an impression material was used to obtain an epoxy resin replica for SEM analysis of margin alterations. Percentage of perfect margin was evaluated under SEM and calculated comparing it with the width/length of restoration margin. The presence of gaps, enamel fractures and other marginal alterations was also recorded inspecting under SEM replicas. Each tooth was then immersed for 24 hours in erythrosin B dye solution at room temperature and evaluated at superficial margins as marginal leakage (ML) for dye penetration. Each sample was then longitudinally sectioned, and leakage was expressed as % dye penetration of the total cavity wall, longitudinal leakage (LL). Finally, each tooth was dissolved (10%H3PO4 for 48 hours and NaOCl solution for additional 24 hours) to leave only the resin restoration. The internal walls of each restoration were then inspected by SEM at x1,000-15,000 to evaluate the presence of resin-dentin infiltrated layer (hybrid layer) and to calculate the ability of primer/bonding to infiltrate peritubular dentin and to form resin tags. RESULTS: Gap widths were 2-6 microns at dentin and lower at enamel. Several enamel fractures along the margin were observed. Resin tags were observed only in deeper dentin. Only Prime & Bond 2.0/Dyract formed resin tags at medium-superficial dentin. The length of resin tags ranged from 1-6 microns for Dyract-PSA primer to 100 microns for Prime & Bond 2.0. Marginal and longitudinal leakage was observed both at dentin and enamel levels. The best dentin seal was obtained with Prime & Bond 2.0/Dyract, while enamel seals were equivalent with both bonding systems (P > 0.05). No correlations were demonstrated between leakage and SEM observations.

Adult↗
PubMed1990

Concerted evolution of class I genes in the major histocompatibility complex of murine rodents.

Full-length cDNA sequences of two class I major histocompatibility complex molecules from the DA strain of Rattus norvegicus are reported. One codes for the classical class I restriction element RT1.Aa, which maps to the locus in the rat major histocompatibility complex homologous to H-2K in the mouse. The other probably codes for a soluble nonclassical class I molecule present in DA rat serum; a short deletion in the fifth exon implies that the translated product will terminate in the membrane-spanning region. These sequences have been compared with mouse classical class I sequences as well as with three published rat class I cDNA partial sequences. The results show, first, that "locus-specific" substitutions from the H-2K, H-2D, and H-2L data set are scrambled in the RT1.Aa molecule; a majority of these substitutions have H-2D/L-specific features. Second, the data show that the four rat sequences are strikingly similar to one another regardless of locus or haplotype of origin; they share a number of apparently species-specific features that distinguish them all from mouse classical class I sequences, which likewise share distinctive features of their own. The results suggest that segmental sequence exchange plays a major role in determining the evolution of sequence in class I major histocompatibility complex molecules.

Amino Acid Sequence↗
PubMed1990

In Vivo Gibberellin Biosynthesis in Endosperm of Sechium edule Sw. Seeds.

Biosynthesis of gibberellins (GAs) was studied in vivo in endosperms of Sechium edule Sw. Exogenous ent-[(14)C]kaurene was metabolized into four major products: GA(12), GA(4), GA(7) and 16, 17-dihydro-16-hydroxy-GA(15) alcohol glucoside. Other minor metabolites were also observed including ent-kaurenol and ent-kaurenal. Conversion of ent-[(14)C]kaurene to ent-kaurenol glucoside by endosperm cell-free preparations in the presence of UDPG was observed. However, the finding was not confirmed in in vivo studies and is probably artifactual. Overall evidence coming from the analysis of endogenous GAs and in vitro and in vivo biosynthetic studies are discussed in relation to the possible existence in the Sechium seeds of a different route, along with the known pathway, branching from ent-kaurene or ent-7-alpha-hydroxykaurenoic acid and this also leading to biologically active GAs.

Journal Article↗
PubMed1989

Gibberellins in Embryo-Suspensor of Phaseolus coccineus Seeds at the Heart Stage of Embryo Development.

Gibberellins (GAs) in suspensors and embryos of Phaseolus coccineus seeds at the heart stage of embryo development were analyzed by combined gas chromatography-mass spectrometry (GC-MS). From the suspensor four C(19)-GAs, GA(1), GA(4), GA(5), GA(6), and one C(20) GA, GA(44), were identified. From the embryo, five C(19)-GAs GA(1), GA(4), GA(5), GA(6), GA(60) and two C(20) GAs, GA(19) and GA(44) were identified. The data, in relation to previous results, suggest a dependence of the embryo on the suspensor during early stages of development.

Journal Article↗
PubMed1989

Abscisic Acid Levels during Early Seed Development in Sechium edule Sw.

The time-course growth of single tissues in pollinated and unpollinated ovules of Sechium edule Sw. is described in relation to the endogenous levels of abscisic acid. Quantitation of abscisic acid (ABA) in the minute amounts of material obtained after ovule dissection has been performed by using a highly specific and sensitive solid-phase radioimmunoassay based on a monoclonal antibody raised against free (S)-ABA. While the absolute amount of ABA rises in both types of ovules, only in unpollinated ones does this leads to an increase in the hormone concentration. Infact in pollinated ovules the rapid growth following pollination prevents, through a dilution effect, the increase in ABA concentration. Growth patterns and endogenous ABA levels are similar for integuments and nucellus tissues either in pollinated or unpollinated ovules. It is suggested that the growth inhibition induced by the increase in ABA concentration after anthesis could be counteracted by the pollination triggered fast ovule growth.

Journal Article↗
PubMedDate not supplied

Serum enzymes of lysosomal origin as indicators of the metabolic control in diabetes: comparison with glycated hemoglobin and albumin.

Several lysosomal enzymes (beta-N-D-acetylglucosaminidase, beta-D-glucuronidase, alpha-D-galactosidase, beta-D-galactosidase, alpha-L-fucosidase, alpha-D-glucosidase, alpha-D-mannosidase, beta-D-glucosidase), glycated albumin and glycated hemoglobin (HbA1c) were determined in the serum of 81 insulin-dependent diabetics with different degrees of metabolic control (optimal, 21 patients; good, 39 patients; poor, 21 patients) and without signs of complications, and in 42 control subjects. All parameters examined increased in serum in inverse proportion to the degree of metabolic control. A highly significant correlation (p less than 0.01) was found between lysosomal enzymes and both glycated albumin and HbA1c. All parameters correlated with hyperglycemia, glycated albumin having the highest r-value (0.586) and lysosomal enzymes the lowest one. Unlike glycated albumin and HbA1c, serum levels of lysosomal enzymes in patients with optimal metabolic control were undistinguishable or even lower than those of controls. A 2-month longitudinal monitoring of a patient who was hospitalized in conditions of poor metabolic control and adequately treated, proved that lysosomal enzymes diminished in serum parallel to glycated albumin and HbA1c in relation to improvement of the metabolic situation. The conclusion is drawn that serum lysosomal enzymes are good indicators of the metabolic control of diabetic patients probably reflecting the overall metabolic state connected with insulin action rather than hyperglycemia.

Adolescent↗