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M Toni

Publications and source records attributed to M Toni.

At least 19 recordsLinked to original sources
PubMed2006

Characterization of beta-keratins in lizard epidermis: electrophoresis, immunocytochemical and in situ-hybridization study.

Lizard scales are composed of alpha-(cyto-) keratins and beta-keratins. The characterization of the molecular weight and isoelectric point (pI) of alpha- and beta-keratins of lizard epidermis (Podarcis sicula) has been done by using two-dimensional electrophoresis, immunoblotting, and immunocytochemistry. Antibodies against cytokeratins, against a chicken scale beta-keratin or against lizard beta-keratin bands of 15-16kDa, have been used to recognize alpha- and beta-keratins. Acid and basic cytokeratins of 42-67kDa show a pI from 5.0 to 8.9. This indicates the presence of specific keratins for the formation of the stratum corneum. Main protein spots of beta-keratin at 15-17kDa, and pI at 8.5, 8.2, and 6.7, and one spot at 10kDa and pI at 7.3 were recognized. Therefore, beta-keratins are mainly basic proteins, and are used for the formation of the hard corneous layer of the epidermis. Ultrastructural immunocytochemistry confirms that beta-keratin is packed into large and dense bundles of beta-keratin cells of lizard epidermis. The use of a probe against a lizard beta-keratin in situ-hybridization studies confirms that the mRNA for beta-keratins is present in beta-cells and is localized around or even associated with beta-keratin filaments.

Actin Cytoskeleton↗
PubMed2005

Distribution and characterization of proteins associated with cornification in the epidermis of gecko lizard.

The distribution and molecular weight of epidermal proteins of gecko lizards have been studied by ultrastructural, autoradiographic, and immunological methods. Setae of the climbing digital pads are cross-reactive to antibodies directed against a chick scutate scale beta-keratin but not against feather beta-keratin. Cross-reactivity for mammalian loricrin, sciellin, filaggrin, and transglutaminase are present in alpha-keratogenic layers of gecko epidermis. Alpha-keratins have a molecular weight in the range 40-58 kDa. Loricrin cross-reactive bands have molecular weights of 42, 50, and 58 kDa. Bands for filaggrin-like protein are found at 35 and 42 kDa, bands for sciellin are found at 40-45 and 50-55 kDa, and bands for transglutaminase are seen at 48-50 and 60 kDa. The specific role of these proteins remains to be elucidated. After injection of tritiated histidine, the tracer is incorporated into keratin and in setae. Tritiated proline labels the developing setae of the oberhautchen and beta layers, and proline-labeled proteins (beta-keratins) of 10-14, 16-18, 22-24 and 32-35 kDa are extracted from the epidermis. In whole epidermal extract (that includes the epidermis with corneous layer and the setae of digital pads), beta-keratins of low-molecular weight (10, 14-16, and 18-19 kDa) are prevalent over those at higher molecular weight (34 and 38 kDa). In contrast, in shed epidermis of body scales (made of corneous layer only while setae were not collected), higher molecular weight beta-keratins are present (25-27 and 30-34 kDa). This suggests that a proportion of the small beta-keratins present in the epidermis of geckos derive from the differentiating beta layer of scales and from the setae of digital pads. Neither small nor large beta-keratins of gecko epidermis cross-react with an antibody specifically directed against the feather beta-keratin of 10-12 kDa. This result shows that the 10 and 14-16 kDa beta-keratins of gecko (lepidosaurian) have a different composition than the 10-12 kDa beta-keratin of feather (archosaurian). It is suggested that the smaller beta-keratins in both lineages of sauropsids were selected during evolution in order to build elongated bundles of keratin filaments to make elongated cells. Larger beta-keratins in reptilian scales produce keratin aggregations with no orientation, used for mechanical protection.

Animals↗
PubMed2004

Immuno-cross reactivity of transglutaminase and cornification marker proteins in the epidermis of vertebrates suggests common processes of soft cornification across species.

In differentiating mammalian keratinocytes proteins are linked to the plasma membrane by epidermal transglutaminases through N-epsilon-(gamma-glutamyl)-lysine isopeptide bonds to form the cornified cell envelope. The presence of transglutaminases and their protein substrates in the epidermis of nonmammalian vertebrates is not known. The present study analyses the presence and localization of the above proteins in the epidermis using immuno-cross reactivity across different classes of amniotes. After immunoblotting, some protein bands appear labelled for loricrin, sciellin, and transglutaminase in most species. These proteins are scarce to absent in the epidermis of aquatic species (goldfish and newt) where a stratum corneum is absent or very thin. The molecular weight of transglutaminase immunoreactive bands generally varies between 40 to 62 kDa, with the most represented bands at 52-57 kDa in most species. The more intense loricrin- and sciellin-immunoreactive bands are seen at 50-55-62 kDa, but are weak or absent in aquatic vertebrates. Loricrine-like immunoreactivity is present in the epidermis where alpha-(soft)-keratinization occurs. Isopeptide bonds are mainly associated to bands in the range of 50-62 kDa. In vertebrates where hard-keratin is expressed (the beta-keratin corneous layer of sauropsids and in feathers) or in hair cortex of mammals, no loricrin-like, transglutaminase-, and isopeptide-bond-immunoreactivities are seen. Immunoblotting however shows loricrin-, sciellin-, and trasnsglutaminase-positive bands in the corneous layers containing beta-keratin. Histologically, the epidermis of most amniotes shows variable transglutaminase immunoreactivity, but isopeptide-bond and sciellin immunoreactivities are weak or undetactable in most species. The limitations of immunohistochemical methods are discussed and compared with results from immunoblotting. In reptilian epidermis transglutaminase is mainly localized in 0.15-0.3 microm dense granules or diffuse in transitional alpha-keratogenic cells. In beta-keratogenic cells few small dense granules show a weak immunolabeling. Transglutaminase is present in nuclei of terminal differentiating alpha- and beta-keratinocytes, as in those of mature inner and outer root sheath. The present study suggests that keratinization based on loricrin, sciellin and transglutaminase was probably present in the stratum corneoum of basic amniotes in the Carboniferous. These proteins were mainly maintained in alpha-keratogenic layers of amniotes but decreased in beta-keratogenic layers of sauropsids (reptiles and birds). The study suggests that similar proteins for the formation of the cornified cell envelope are present in alpha-keratinocytes across vertebrates but not in beta-keratinocytes.

Animals↗
PubMed2004

Characterization of beta-keratins and associated proteins in adult and regenerating epidermis of lizards.

Reptilian epidermis contains two types of keratin, soft (alpha) and hard (beta). The biosynthesis and molecular weight of beta-keratin during differentiation of lizard epidermis have been studied by autoradiography, immunocytochemistry and immunoblotting. Tritiated proline is mainly incorporated into differentiating and maturing beta-keratin cells with a pattern similar to that observed after immunostaining with a chicken beta-keratin antibody. While the antibody labels a mature form of beta-keratin incorporated in large filaments, the autoradiographic analysis shows that beta-keratin is produced within the first 30 min in ribosomes, and is later packed into large filaments. Also the dermis incorporates high amount of proline for the synthesis of collagen. The skin was separated into epidermis and dermis, which were analyzed separately by protein extraction and electrophoresis. In the epidermal extract proline-labeled proteic bands at 10, 15, 18-20, 42-45, 52-56, 85-90 and 120 kDa appear at 1, 3 and 5 h post-injection. The comparison with the dermal extract shows only the 85-90 and 120 kDa bands, which correspond to collagen. Probably the glycine-rich sequences of collagen present also in beta-keratins are weakly recognized by the beta-1 antibody. Immunoblotting with the beta-keratin antibody identifies proteic bands according to the isolation method. After-saline or urea-thiol extraction bands at 10-15, 18-20, 40, 55 and 62 kDa appear. After extraction and carboxymethylation, weak bands at 10-15, 18-20 and 30-32 kDa are present in some preparations, while in others also bands at 55 and 62 kDa are present. It appears that the lowermost bands at 10-20 kDa are simple beta-keratins, while those at 42-56 kDa are complex or polymeric forms of beta-keratins. The smallest beta-keratins (10-20 kDa) may be early synthesized proteins that are polymerized into larger beta-keratins which are then packed to form larger filaments. Some proline-labeled bands differ from those produced after injection of tritiated histidine. The latter treatment does not show 10-20 kDa labeled proteins, but tends to show bands at 27, 30-33, 40-42 and 50-62 kDa. Histidine-labeled proteins mainly localize in keratohyalin-like granules and dark keratin bundles of clear-oberhautchen layers of lizard epidermis, and their composition is probably different from that of beta-keratin.

Animals↗
PubMed2003

Putative histidin-rich proteins in the epidermis of lizards.

In the stratum granulosum of mammalian epidermis, histidin-rich proteins (filaggrins) determine keratin clumping and matrix formation into terminal keratinocytes of the stratum corneum. The nature of matrix, interkeratin proteins in the epidermis of nonmammalian vertebrates, and in particular in that of reptilian, mammalian progenitors are unknown. The present biochemical study is the first to address this problem. During a specific period of the renewal phase of the epidermis of lizards and during epidermal regeneration, keratohyalin-like granules are formed, at which time they take up tritiated histidine. The latter also accumulate in cells of the alpha-keratin layer (soft keratin). This pattern of histidine incorporation resembles that seen in keratohyalin granules of the stratum granulosum of mammalian epidermis. After injection of tritiated histidine, we have analysed the distribution of the radioactivity by histoautoradiography and electrophoretic gel autoradiography of epidermal proteins. Extraction and electrophoretic separation of interfilamentous matrix proteins from regenerating epidermis 3-48 hours post-injection reveals the appearance of protein bands at 65-70, 55-58, 40-43, 30-33, 25-27, and 20-22 kDa. Much weaker bands were seen at 100, 140-160, and 200 kDa. A weak band at 20-22 kDa or no bands at all are seen in the normal epidermis in resting phase and in the dermis. In regenerating epidermis at 22 and 48 hours post-injection, little variation in bands is detectable, but low molecular weight bands tend to increase slightly, suggesting metabolic turnover. Using anti-filaggrin antibodies against rat, human, or mouse filaggrins, some cross-reactivity was seen with more reactive bands at 40-42 and 33 kDa, but it was reduced or absent at 140, 95-100, 65-70, 50-55, and 25 kDa. This suggests that different intermediate degradative proteins of lizard epidermis may share some epitopes with mammalian filaggrins and are different from keratins with molecular weight ranging from 40 to 65-68 kDa. The immunocytochemical observation confirms that a weak filaggrin-like immunoreactivity characterizes differentiating alpha-keratogenic layers in normal and regenerating tail. A weak filaggrin labeling is discernable in small keratohyalin-like granules but is absent from the larger granules and from mature keratinocytes. The present results indicate, for the first time, that histidine-rich proteins are involved in the process of alpha-keratinization in reptilian epidermis. The cationic, interkeratin matrix proteins implicated may be fundamentally similar in both theropsid-derived and sauropsid amniotes.

Animals↗
PubMed2001

Colocalization prostacyclin (PGI2) synthase--caveolin-1 in endothelial cells and new roles for PGI2 in angiogenesis.

In vascular cells, prostacyclin (PGI2) synthase (PGI2s) has been localized in the endoplasmic reticulum of endothelial cells and in the nuclear and plasma membrane of smooth muscle cells. In human umbilical vein endothelial (HUVE) cells, we detected the enzyme in abundant cytoplasmic vesicles apparently originating from the plasma membrane and similar to those stained by gold-albumin, which interacts with a caveolar receptor. This prompted us to try a direct confocal microscopy approach aimed at colocalizing gold-albumin, caveolin-1, and PGI2 synthase. Moreover, the staining of HUVE cells with an anti-BiP7Grp78 antibody (a marker of endoplasmic reticulum) shows a perinuclear localization, sharply separated from PGI2 synthase localization. The results indicate that more than 80% of the enzyme resides in cellular sites costaining with caveolin-1 antibody and gold-albumin. This evidence was confirmed by the demonstration that PGI2 synthase and caveolin-1 coimmunoprecipitate in HUVE cell lysates and that they are associated to detergent-insoluble membrane domains in the same low-density fractions of a sucrose gradient. In addition, depletion of cellular cholesterol by mevalonate and methyl-beta-cyclodextrin leads to the shift of PGI2 synthase and caveolin-1 to higher density fractions of the gradient. Biochemical evidence about colocalization was supported by the use of a fusion protein glutathione S-transferase (GST)/caveolin-1, which retained either PGI2s purified from ram seminal vesicles or PGI2s present in HUVE cell lysates. Binding of PGI2s to caveolin "scaffolding domain" and to C-terminal region was deduced by using full-length GST--Cav-1, GST--Cav 61--101, and GST C- and N-terminal fusion proteins. A double approach based on the usage of filipin as a specific caveolae-disrupting agent and antisense oligonucleotides targeting PGI2 synthase mRNA suggests that the production of PGI2 in caveolae is likely to be connected to the regulation of angiogenesis, at least in vitro.

6-Ketoprostaglandin F1 alpha↗
PubMed2001

Prevalence of antimicrobial resistance in Streptococcus pneumoniae circulating in Italy: results of the Italian Epidemiological Observatory Survey (1997-1999).

The Italian Epidemiological Observatory (IEO), a surveillance program supported by the SmithKline Foundation, analyzed the susceptibility of 2,664 community-acquired respiratory Streptococcus pneumoniae derived from over 50 clinical microbiology laboratories during 1997-1999, against 21 antibiotics adopting a quantitative methodology. Throughout these years, total penicillin resistance varied from 14.3% to 10.2%. High-level resistance has remained stable, ranging from 3.8% to 4.1%, while a decrease in low-level resistance (from 10.3% to 6.1%) has been recorded. Lack of susceptibility to macrolides ranged from 29.1% in 1997 to 25.5% in 1999. Similar figures have also been observed with tetracycline and co-trimoxazole (rates of resistance around 30%). As expected, large geographical variations in resistance rates were found for all drugs. Amoxicillin and amoxicillin-clavulanate were 100% active on penicillin-intermediate isolates. Injectable third-generation cephalosporins and carbapenems were also capable of inhibiting a large proportion of these microorganisms. Rifampin was the most potent non-beta-lactam compound tested. In contrast to the situation prevailing elsewhere, in Italian children (aged 0-5 years) presenting with respiratory conditions, the total rate of penicillin resistance (3%) was lower than that shown by the adult population (10.9%). However, lack of susceptibility to macrolides, tetracycline, and cotrimoxazole (35%, 41%, 44%) was more incident in pediatric than in adult patients (25%, 26%, 28% respectively). Strains recovered from blood in 1999 (67) were much more susceptible to penicillin (98.5%) than respiratory pneumococci (89.8%), whereas macrolides, tetracycline, and cotrimoxazole were consistently less active (75%, 67%, 64%).

Anti-Bacterial Agents↗
PubMed1992

Antibiotic resistance of clinical isolates of Enterococcus in Italy.

Alarming reports from various parts of the world of an increase in multiple-drug resistance among enterococci have prompted the Italian Association of Clinical Microbiologists to sponsor a nationwide study aimed at assessing the magnitude of this problem in Italy. During the last 3 months of 1988, technicians from a total of 162 general and teaching hospitals, in 16 of the 20 Italian regions and covering 89,061 beds, isolated 58,886 bacterial strains from inpatients and outpatients. Enterococci (4,879) represented 8.3% of all microorganisms identified, with Enterococcus faecalis accounting for 4,554 strains and Enterococcus faecium for the remaining 316 (6.5% of all enterococci analyzed). Enterococci were recovered most frequently from urine (62.5%) and genitourinary samples (13.7%). Only 58 strains (1.2%) were associated with bacteremia. The in vitro susceptibility of these organisms to 21 commonly used antibiotics is reported and compared with the susceptibility of isolates from a smaller epidemiologic study conducted in Italy in 1986 as well as with more recent findings from individual laboratories.

Drug Resistance, Microbial↗
PubMed1991

Acyclovir resistance/susceptibility in herpes simplex virus type 2 sequential isolates from an AIDS patient.

The biological characterization of a number of sequential herpes simplex virus type 2 (HSV-2) isolates obtained from an AIDS patient undergoing sequential courses of antiviral treatment due to an extended mucocutaneous genital lesion is reported. Resistance to acyclovir (ACV) and related compounds was linked to a thymidine kinase-deficient (TK-) phenotype. After ACV discontinuation and a course of treatment with foscarnet, a new isolate was recovered, characterized by loss of the ACV-resistant trait and production of a functional TK enzyme. Data presented stress the need for monitoring chemosensitivity of HSV isolates in AIDS patients while suggesting that for better control of the infection, these patients should benefit from alternative treatments with drugs aimed at different viral targets.

Acquired Immunodeficiency Syndrome↗
PubMed1991

[Prosthetic valve endocarditis caused by Gemella haemolysans].

Reported herein is the case of a 47 year old man with mitral and aortic bioprosthetic valves who developed endocarditis. Blood cultures turned out to be positive for Gemella haemolysans, a commensal of the upper respiratory tract. Proper antibiotic therapy was immediately started according to antibiogram, and continued after the acute stage of the disease had been overcome. The aortic prosthesis appeared much more to be incompetent at Doppler and aortography (from 2/4 to 3/4). At surgery, the gross appearance of the explanted xenograft was consistent with infective endocarditis. The case is described for the rarity of the etiology: only 6 cases of infective endocarditis from Gemella haemolysans are reported in the literature, but this is the first involving a prosthetic valve.

Anti-Bacterial Agents↗
PubMed1989

Serum sensitivity of Escherichia coli strains present in the clinical environment.

It is well known that the growth of some Gram-negative microorganisms is inhibited by normal human serum; but there are no reports about the percentage of serum-sensitive Escherichia coli strains present in clinics. The aim of the present paper is to obtain data useful for elucidating this point. The study was carried out using 38 strains of E. coli isolated in clinics. Different concentrations of serum (from 2.5% to 20% in BHI broth) were used for evaluating its bactericidal activity. The bactericidal activity was measured both as colony forming units (CFU) and as a delay in the growth curve recorded by an Avantage-Abbott instrument. The concentration of serum inducing a killing effect greater than 99.9% was considered bactericidal. The results indicate that in the presence of 2.5%, 5%, 10% and 20% of serum in BHI broth, the percentage of serum-sensitive strains was 2.6%, 10.5%, 18.4% and 31.6%, respectively. These data are discussed in view of the possible application of the synergistic effect of serum with sub-MICs of different antibiotics on serum-sensitive E. coli strains present in the clinical environment.

Blood Bactericidal Activity↗
PubMed1985

Isolation of Chlamydia trachomatis from the prostatic cells in patients affected by nonacute abacterial prostatitis.

To assess the presence of Chlamydia trachomatis in nonacute abacterial prostatitis 30 patients with urethral cultures positive for Chlamydia trachomatis underwent microbiological studies, including cultures of transrectal aspiration biopsies of the prostate. Chlamydia trachomatis was isolated from 10 of the prostatic specimens (33 per cent). In 3 cases a nonspecific cytopathogenic effect caused the destruction of the tissue cultures. Our findings demonstrate that Chlamydia trachomatis may cause ascending infections of the prostate and that this microorganism may have an etiological role in the pathogenesis of nonacute abacterial prostatitis.

Adult↗
PubMed1981

[Epidemiology of multiresistant S. aureus in a burn center].

Multi-resistant S. aureus, have been isolated from wounds of patients in the burn unit of the University of Parma. The strains were resistant to penicillin (85%), methicillin (30%), gentamycin (63%), rifampicin (41%), cotrimoxazole (38%), lincomycin (39%), tetracycline (73%), erythromycin (66%) and cephaloridine (11%). The spread of these strains has been monitored by the analysis of the antibiograms and the lysotypes and in few cases some strains were also found in other wards of the Hospital. All of these had some correlation with the burns unit. Medical and technical staff of this ward was found to be carrier of multi-resistant S. aureus in 46% of the cases and that feature is supposed to be the most important in the epidemiology of S. aureus in this Hospital.

Anti-Bacterial Agents↗