Biomedical subjects

P De Luca

Publications and source records attributed to P De Luca.

At least 19 recordsLinked to original sources

PubMed2005

Modulation of the catalytic activity of free and immobilized peroxidase by extremely low frequency electromagnetic fields: dependence on frequency.

A study of the influence of electromagnetic fields (EMF) of various frequencies, from 50 up to 400 Hz, on the catalytic activity of soluble and insoluble horseradish peroxidase (POD) was carried out. To simulate the conditions in which the enzyme operates in vivo, the POD was immobilized by entrapment on a gelatin membrane or by covalent attachment on a nylon graft membrane. The rate of inactivation of the soluble POD was found to exhibit positive and negative interactions with the 1 mT applied magnetic field, with an optimum positive effect at 130 Hz. The immobilized PODs, on the contrary, do not exhibit negative interactions, but show a maximum positive interaction at 150 Hz when entrapped and at 170 Hz when covalently attached. At 50 Hz and at frequencies higher than 250 Hz no effects were observed with insoluble POD. The optimum frequency of positive interaction between the EMF and the catalytic activity of the insoluble enzymes is shifted with respect to that of the soluble enzymes towards higher frequencies, the size of the shifts being dependent on the intensity of the physical forces involved in the immobilization process.

Catalysis↗

PubMed2003

In vitro studies of the influence of ELF electromagnetic fields on the activity of soluble and insoluble peroxidase.

The influence of an extremely low frequency (ELF) magnetic field (50 Hz and 1 mT, EMF) on the activity of a soluble and insoluble horseradish peroxidase (E.C. 1.11.17) has been studied as a function of time. Insoluble derivatives were obtained by enzyme entrapment into two different gelatin membranes or by covalent attachment of the enzyme on two nylon membranes, differently preactivated. Results have shown that the field affects the inactivation rate of the soluble enzyme, while no effects are observed with insoluble derivatives. Since in vivo enzymes are immobilised into the biomembrane bilayer or entrapped into the cytoplasmic mixture, one might speculate that our experimental conditions do not reflect the catalytic activity of the enzymes in vivo.

Benzothiazoles↗

PubMed2002

Advantages of using non-isothermal bioreactors for the enzymatic synthesis of antibiotics: the penicillin G acylase as enzyme model.

A new hydrophobic and catalytic membrane was prepared by immobilizing Penicillin G acylase (PGA, EC.3.5.1.11) from E. coli on a nylon membrane, chemically grafted with butylmethacrylate (BMA). Hexamethylenediamine (HMDA) and glutaraldehyde (Glu) were used as a spacer and coupling agent, respectively. PGA was used for the enzymatic synthesis of cephalexin, using D(-)-phenylglycine methyl ester (PGME) and 7-amino-3-deacetoxycephalosporanic acid (7-ADCA) as substrates. Several factors affecting this reaction, such as pH, temperature, and concentrations of substrates were investigated. The results indicated good enzyme-binding efficiency of the pre-treated membrane, and an increased stability of the immobilized PGA towards pH and temperature. Calculation of the activation energies showed that cephalexin production by the immobilized biocatalyst was limited by diffusion, resulting in a decrease of enzyme activity and substrate affinity. Temperature gradients were employed as a way to reduce the effects of diffusion limitation. Cephalexin was found to linearly increase with the applied temperature gradient. A temperature difference of about 3 degrees C across the catalytic membrane resulted into a cephalexin synthesis increase of 100% with a 50% reduction of the production times. The advantage of using non-isothermal bioreactors in biotechnological processes, including pharmaceutical applications, is also discussed.

Anti-Bacterial Agents↗

PubMed2001

Maternal inheritance of plastids in Encephalartos Lehm. (Zamiaceae, Cycadales).

The mode of inheritance of chloroplast DNA has been determined in Encephalartos by employing a restriction fragment length polymorphism analysis of chloroplast DNA. Artificial F1 hybrids were produced between a female specimen of E. natalensis and a male specimen of E. woodii. The hybridization patterns of all hybrids correspond, in all cases, with that of E. natalensis, and are different from that of E. woodii, thus indicating the maternal inheritance of cpDNA in cycads.

Chloroplasts↗

PubMed2000

Comparison of the ADP-ribosylating thermozyme from Sulfolobus solfataricus and the mesophilic poly(ADP-ribose) polymerases.

The poly(ADP-ribose) polymerase-like thermozyme purified from Sulfolobus solfataricus was characterised with respect to some physico-chemical properties. The archaeal protein exhibited a scarce electrophoretic mobility at both pH 2.9 and pH 7.5. Determination of the isoelectric point (pI=7.0-7.2) allowed us to understand the reason for the limited migration at pH 7.5, while amino acid composition analysis showed a moderate content of basic residues, which reduced mobility at pH 2.9. With respect to the charge, the archaeal enzyme behaved differently from the eukaryotic thermolabile poly(ADP-ribose) polymerase, described as a basic protein (pI=9.5). Well known inhibitors of the mesophilic polymerase like Zn(2+), nicotinamide and 3-aminobenzamide exerted a smaller effect on the enzyme from S. solfataricus, reducing the activity by at most 50%. Mg(2+) was a positive effector, although in a dose-dependent manner. It influenced the fluorescence spectrum of the archaeal protein, whereas NaCl had no effect.

Amino Acids↗

PubMed2000

Class A helix-loop-helix proteins are positive regulators of several cyclin-dependent kinase inhibitors' promoter activity and negatively affect cell growth.

The class A of basic helix-loop-helix (bHLH) proteins are ubiquitously expressed transcription factors playing a pivotal role in the regulation of cell growth and differentiation. We determined that enforced expression of all four different mammalian members of this family, E12, E47, E2-2, and HEB, suppresses the cell colony-forming efficiency of several cell lines. To gain insights into the mechanisms by which class A bHLH factors affect cell growth, we have investigated their role in the transcriptional regulation of cyclin-dependent kinase inhibitors. We found that p21CIP1/ WAF1, p15INK4B, and p16INK4B promoter sequences contain E-boxes that render these genes competent for class A bHLH-mediated transcriptional activation and Id-mediated repression. The mechanism underlying the class A bHLH-mediated inhibition of cell growth does not involve an arrest of G1 progression in 293T cells. In fact, contrary to what has been found in 3T3 NIH fibroblasts, we found that enhanced expression of class A bHLH proteins led to a decreased proliferation rate by promoting cell death associated with the induction of apoptosis. These findings highlight the role of the class A bHLH proteins as general negative regulators of cell proliferation through a mechanism(s) that involves both enhancement of several cyclin-dependent kinase inhibitor genes expression and promotion of cell death.

Animals↗

PubMed2000

Oligomerization of RAR and AML1 transcription factors as a novel mechanism of oncogenic activation.

RAR and AML1 transcription factors are found in leukemias as fusion proteins with PML and ETO, respectively. Association of PML-RAR and AML1-ETO with the nuclear corepressor (N-CoR)/histone deacetylase (HDAC) complex is required to block hematopoietic differentiation. We show that PML-RAR and AML1-ETO exist in vivo within high molecular weight (HMW) nuclear complexes, reflecting their oligomeric state. Oligomerization requires PML or ETO coiled-coil regions and is responsible for abnormal recruitment of N-CoR, transcriptional repression, and impaired differentiation of primary hematopoietic precursors. Fusion of RAR to a heterologous oligomerization domain recapitulated the properties of PML-RAR, indicating that oligomerization per se is sufficient to achieve transforming potential. These results show that oligomerization of a transcription factor, imposing an altered interaction with transcriptional coregulators, represents a novel mechanism of oncogenic activation.

Cell Transformation, Neoplastic↗

PubMed2000

Burning mouth syndrome (BMS): controlled open trial of the efficacy of alpha-lipoic acid (thioctic acid) on symptomatology.

BACKGROUND: Alpha-lipoic acid (ALA), is a potent antioxidant mitochondrial coenzyme, the trometamol salt of thioctic acid that has been shown in clinical studies to be neuroprotective. This study examined the effect of ALA on the symptomatology of Burning mouth syndrome (BMS). SUBJECTS AND METHODS: Forty-two patients with BMS and no clinical or laboratory evidence of organic oral disease were divided into two groups (Test and Control) each of 21 subjects, matched for age and sex. The Test group were given ALA (thioctic acid; Tiobec) for 30 days, as 600 mg per day orally for 20 days followed by 200 mg per day for 10 days. The Control group were given cellulose starch 100 mg per day as placebo for 30 days. All BMS patients were reviewed at 10-day intervals and scored for changes in symptomatology. RESULTS: Significant improvements were shown in the symptomatology of BMS in up to two-thirds of patients with BMS receiving alpha-lipoic acid, in about 15% of those using placebo and also in up to two-thirds of those who, having tried placebo, were switched to ALA.

Administration, Oral↗

PubMed1999

B-MYB transactivates its own promoter through SP1-binding sites.

B-MYB is an ubiquitous protein required for mammalian cell growth. In this report we show that B-MYB transactivates its own promoter through a 120 bp segment proximal to the transcription start site. The B-MYB-responsive element does not contain myb-binding sites and gel-shift analysis shows that SP1, but not B-MYB, protein contained in SAOS2 cell extracts binds to the 120 bp B-myb promoter fragment. B-MYB-dependent transactivation is cooperatively increased in the presence of SP1, but not SP3 overexpression. When the SP1 elements of the B-myb promoter are transferred in front of a heterologous promoter, an increased response to B-MYB results. In contrast, c-MYB, the prototype member of the Myb family, is not able to activate the luciferase construct containing the SP1 elements. With the use of an SP1-GAL4 fusion protein, we have determined that the cooperative activation occurs through the domain A of SP1. These observations suggest that B-MYB functions as a coactivator of SP1, and that diverse combinations of myb and SP1 sites may dictate the responsiveness of myb-target genes to the various members of the myb family.

Adenovirus E1A Proteins↗

PubMed1999

Isolation and characterization of four type-1 ribosome-inactivating proteins, with polynucleotide:adenosine glycosidase activity, from leaves of Phytolacca dioica L.

Four type-1 (single-chain) ribosome-inactivating proteins (RIPs), with isoelectric points between 9.5 and 9.7, were isolated from leaves of Phytolacca dioica L. The purification procedure furnished the four proteins with an overall yield of about 16% and separated them from a protein of 29,407 +/- 2 Da, as determined by electrospray mass spectrometry, whose N-terminal amino acid sequence differed from that of pokeweed (Phytolacca americana L.) leaf chitinase (PLC-B) by only one amino acid (R17I). The four RIPs (PD-L1 to PD-L4) inhibited protein synthesis by a rabbit reticulocyte lysate with 50% inhibition at the picomolar level, and produced the beta-fragment, diagnostic of the specific enzymatic action of RIPs, on yeast ribosomes. Comparison of their N-terminal sequences, up to residue 45, showed that PD-L1 is identical to PD-L2 [designated the isoleucine (Ile) form from the N-terminal residue] and PD-L3 is identical to PD-L4 [designated the valine (Val) form from the N-terminal residue] and that there are 35 identical residues between the two forms. Furthermore, the Val form presents the same number of identical residues as PD-S2, an RIP isolated from the seeds of the same plant. With the exception of PD-L4, the purified RIPs gave a positive reaction when stained for sugars on SDS-PAGE gels and, when analyzed by electrospray mass spectrometry, had M(r) values of 32,715 +/- 1 (PD-L1), 31,542 +/- 1 (PD-L2), 30,356 +/- 1 (PD-L3) and 29,185 +/- 1 Da (PD-L4). The 1171 kDa difference in M(r), within the same RIP form, could be due to glycosylation. Like leaf saporins and many other RIPs, the four RIPs released several adenines from poly(A), herring sperm DNA and rRNA 16S + 23S, thus acting as polynucleotide:adenosine glycosidases. This property was less pronounced in PD-L1 and PD-L3 than in PD-L2 and PD-L4, respectively. The proteins PD-L1 and PD-L4 showed 3.7% reactivity with the antiserum anti-dianthin 32 and no reactivity with antisera to PAP-R saporin-S6, momordin 1 and even PD-S2, an RIP isolated from the seeds of the same plant. Protein PD-L4 showed 12.5% cross-reactivity with anti-PD-L1, while the opposite cross-reactivity was 100%.

Amino Acid Sequence↗

PubMed1999

Recent advances on the pathogenesis of oral lichen planus (OLP). The adhesion molecules.

Oral Lichen Planus is a chronic inflammatory pathology with not well defined etiology, characterized by an immunoreactivity directed against the keratinocytes of basal layer and mediated by a cellular infiltrate of T-lymphocytes. This immunoreactivity is caused by a modification of basal keratinocytes surface antigens, with activation of antigen-presenting cells allowing the recognition on non-self antigens by CD4 lymphocytes. The migration of T-cells at the sites of the damage and their interactions with leucocytes and keratinocytes represent a key effect of pathogenesis of OLP, and are mediated by specific cell surface adhesion molecules. Recent studies in molecular biology, with the discovery and the definition of cell adhesion receptors, have greatly contributed to clarify the pathogenic mechanism of OLP; these molecules regulate immune functions such as adhesion to endothelial cells, lymphocytes migration into extravasal tissues at the site of immune response and T-cell interactions with target cell antigens. The adhesion molecules have been classified into various families, differing on the basis of their molecular structure: the Immunoglobulin Superfamily (ICAM 1-3, PECAM-1, VCAM-1), the Selectins (ELAM-1, LECAM-1, GMP-140), the Integrins (LEA-1) and others. In this paper, the principal classes of adhesion molecules which are involved in the immune response are described and their importance in the pathogenesis of Lichen Planus is underlined.

Cell Adhesion Molecules↗

PubMed1999

Stevens-Johnson syndrome. Description of an unusual clinical case due to glucocorticoid therapy for oral lichen planus.

Erythema multiforme (EM) is an acute inflammatory disease with an autoimmune pathogenesis clinically expressing in a wide variety of mucocutaneous illnesses. It is usually described in a minor form (Von Hebra) characterized by classical cutaneous lesions, and in major form (Stevens-Johnson), involving mucosal damage, while a clinical type restricted to the oral mucosa is described in oral pathology. A considerable number of factors of different nature have been reported as etiologic agents of EM, but most of them are not well documented; however, a certain relationship with EM is recognized for different classes of systemic drugs. This paper describes a case of Stevens-Johnson syndrome with initial oral involvement, in which the precipitating factor was due to the administration of systemic glucocorticoids, prescribed for the therapeutic treatment of an erosive form of oral lichen planus.

Aged↗

PubMed1998

Retinoblastoma protein tethered to promoter DNA represses TBP-mediated transcription.

The retinoblastoma (RB) tumour suppressor protein negatively regulates cell proliferation by modulating transcription of growth-regulatory genes. Recruitment of Rb to promoters, by association with E2F complex or by fusion with heterologous DNA-binding domains, demonstrated that Rb represses directly transcription. Recent studies also suggest that the RB protein is able to repress gene transcription mediated by the RNA polymerase I and III. Since the TATA-binding protein (TBP) is an important component for transcription mediated by all three RNA polymerases, we have analysed the functional interaction between Rb and TBP in vivo in the context of RNA pol II-driven transcription. We demonstrated that in mammalian cells Rb tethered to promoter represses TBP-mediated activation in vivo, and Rb-mediated repression is reversed in the presence of the inhibition of histone deacetylase activity by trichostatin A (TSA).

Cell Line↗

PubMed1998

Recruitment of human TBP selectively activates RNA polymerase II TATA-dependent promoters.

An increasing body of evidence suggests that eukaryotic activators stimulate polymerase II transcription by facilitating the assembly of the functional basal machinery at the promoter. Here we describe experiments that provide added support for the idea that recruitment of TATA-binding protein (TBP) is a rate-limiting step for transcription activation in mammalian cells. We found that, in human cell lines, recruitment of TBP to a promoter, as a GAL4-TBP fusion protein, can provide a substantial activation of transcription. Activation mediated by the hTBP, tethered to promoter DNA, is strictly dependent upon the presence of a functional TATA element, and it directs faithful transcription initiation. Interestingly, GAL4-hTBP activation was not observed from initiator (Inr) -dependent TATA-less promoters. These results suggest that TBP binding to DNA is not a rate-limiting step for the initial stages of TFIID recruitment to initiator-dependent TATA-less promoters. Finally, we provide evidence that synergy between GAL4-hTBP and defined transcription domains is restricted to activators, such as VP16 and Tat, which are likely to function at steps subsequent to the TFIID recruitment. These findings strengthen the idea that recruitment of TBP represents an important mechanism of activation of TATA-dependent promoters, and on the other hand, they suggest that TBP-DNA interactions are largely dispensable for specific transcription of initiator dependent TATA-less promoters.

DNA↗

PubMed1997

Sp3 is a bifunctional transcription regulator with modular independent activation and repression domains.

Sp3 is a member of the Sp family of transcription factors and binds to DNA with affinity and specificity comparable to that of Sp1. We demonstrate that Sp3 is a bifunctional transcription factor that can both activate and repress transcription. Gene fusion experiments in mammalian cells demonstrate that the Sp3 activation potential is distributed over an extensive glutamine-rich N-terminal region, whereas the repressor activity has been mapped in a 72-amino acid region located at the 5' of the zinc finger DNA-binding domain. We demonstrated that the repression activity is strictly dependent on the context of the DNA-binding sites bound by Sp3. We found that Sp3 represses transcription of promoters bearing multiple GAL4 DNA-binding sites, whereas it activates isogenic reporters containing a single GAL4-binding site. Transfection experiments in Drosophila cells that lack endogenous Sp activity demonstrated that Sp3 does not possess an active repression domain that can function in insect cells, rather it is a weak transcriptional activator of the c-myc promoter. Our results strongly suggest that Sp3 is a dual-function regulator whose activity is dependent upon both the promoter and the cellular context.

Animals↗

PubMed1997

Transcriptional regulation by the Sp family proteins.

Sp1 is one of the very first cellular transcription factors to be identified and cloned in virtue of its binding to a G-rich motif in the SV40 early promoter. Sp1 protein binds to the G-rich sequences present in a variety of cellular and viral promoters and stimulates their transcriptional activity. Recently, a number of other GC and/or GT box-binding factors homologous to Sp1 have been isolated, namely Sp2, Sp3 and Sp4, and the two more distantly related factors, BTEB and BTEB2. The discovery of this family highlights a previously unknown level of complexity of transcriptional regulation of promoters containing GC and/or GT box motifs. This review focuses primarily on strategies aimed to elucidate the transcription properties of the Sp1-like factors and discusses the experimental problems inherent in the attempt to define their respective functions.

Amino Acid Sequence↗

PubMed1996

Sp3 represses transcription when tethered to promoter DNA or targeted to promoter proximal RNA.

Sp3 is a member of the Sp family of transcription factors, and it binds to the GC box with an affinity and specificity comparable with that of Sp1. Previous studies have shown that Sp3 repressed Sp1-mediated transcriptional activation, suggesting that Sp3 is an inhibitory member of the Sp family. The experiments described here demonstrate that Sp3 contains a portable repression domain that can function independently from the zinc finger DNA-binding domain. We found that the amino-terminal region of Sp3 tethered to a promoter DNA by connecting to a heterologous DNA-binding protein domain represses transcriptional activation by different positive regulators. Moreover, we determined that Sp3 targeted to a promoter-proximal RNA sequence acts as a transcriptional repressor. Taken together, our results suggest that Sp3 functions as a repressor by protein-protein interaction with components of the general transcription complex.

Base Sequence↗

PubMed1996

Prevalence of Helicobacter pylori infection in elderly inpatients and in institutionalized old people: correlation with nutritional status.

Helicobater pylori plays an important role in the aetiology and development of peptic ulcer disease. The prevalence of H. pylori infection increases with age, and is influenced by low socioeconomic status and poor hygiene owing to person-to-person transmission of the organism by the oral-faecal route. The aim of this study was to investigate the prevalence of H. pylori infection, detected serologically, in elderly patients admitted to a geriatric rehabilitation ward and in a sample of institutionalized old subjects. Nutritional status was also evaluated in order to examine its relation to H. pylori infection. The overall prevalence of H. pylori infection was 70.8%, the prevalence in hospitalized patients being 72.9% and in institutionalized subjects 68.7%. No significant correlation was observed between anti-H. pylori IgG levels and either age or length of stay in the institution. We found no difference between H. pylori positive and negative patients as regards their self-sufficiency and cognitive functions. The prevalence of anti-H. pylori antibodies in the serum was not related to blood variables (including nutritional indices), history of drug consumption (in particular nonsteroidal anti-inflammatory drugs), dyspeptic symptoms, or alcohol and smoking habits.

Aged↗