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Biomedical subjects

A De Luca

Publications and source records attributed to A De Luca.

At least 217 records · Page 12Linked to original sources
PubMed1995

Molecular cloning, expression, and developmental characterization of the murine retinoblastoma-related gene Rb2/p130.

The product of the retinoblastoma-related human gene Rb2/p130 is highly homologous with the product of the retinoblastoma tumor suppressor gene (pRb) and Rb-related p107. this homology is shared mainly in the pocket domain, a region that seems to play a key role in the functions of these proteins. Here we report the molecular cloning and initial characterization of the cDNA encoding the murine homologue of the human Rb2/p130 gene product. The 4.8-kb cDNA encodes a protein of 1125 amino acids that shows 90% indentity to that of the human protein. The Rb2/p130 mRNA is found to be expressed in all of the adult mouse tissues examined, with the highest level being detected in kidney and skeletal muscle. For the protein characterization, we used a polyclonal antibody raised against the COOH terminus of the human Rb2/p130 protein that also recognizes the mouse protein. In developing mouse embryos, the Rb2/p130 protein is expressed as early as day 10 of gestation and reached a peak of expression around day 13 of gestation, implying a developmental regulation of the Rb2/p130 gene in murine ontogeny.

3T3 Cells↗
PubMed1994

Transcription of histone H4, H3, and H1 cell cycle genes: promoter factor HiNF-D contains CDC2, cyclin A, and an RB-related protein.

Cell cycle-controlled human histone genes are coordinately expressed during S phase, and transcriptional regulation involves a series of trans-acting factors (HiNFs). The proliferation-specific factor HiNF-D interacts with multiple recognition motifs in histone H4, H3, and H1 promoters. Using gel shift immunoassays, we show that CDC2, cyclin A, and an RB-related protein are ubiquitous subunits of HiNF-D binding activity isolated from several cell types. HiNF-D levels in vivo are sensitive to okadaic acid and staurosporine, indicating that HiNF-D activity and/or assembly is influenced by phosphorylation status. Thus, HiNF-D appears to be a multicomponent phosphoprotein that participates in coordinate control of multiple histone H4, H3, and H1 genes during the cell cycle. The presence of cell cycle mediators in the HiNF-D complex suggests linkage between transcriptional control of histones, enzymes involved in DNA synthesis, and the onset of DNA replication during the G1/S phase transition.

Alkaloids↗
PubMed1994

p130/pRb2 has growth suppressive properties similar to yet distinctive from those of retinoblastoma family members pRb and p107.

The retinoblastoma tumor suppressor gene product, as well as its related protein p107, has been shown clearly to exert its growth suppressive effects in a cell cycle dependent manner. In this study we demonstrate that the introduction of our recently cloned Rb family member p130/pRb2 causes growth arrest in three tumor cell lines. In addition, in the nasopharyngeal carcinoma derived cell line HONE-1, we identified a low level of expression of p130/pRb2, possibly due to gene rearrangement, and a drastic reduction in proliferation upon introduction of a constitutive active p130/pRb2 complementary DNA clone. Furthermore, we were able to dissect distinct properties of the Rb family by demonstrating that p130/pRb2 inhibits proliferation of the glioblastoma cell line T98G, which is resistant to the growth suppressive effects of both pRb and p107. Our studies demonstrate that the Rb family proteins identified to date may complement each other but they are not fully functionally redundant.

Cell Division↗
PubMed1994

PITALRE, a nuclear CDC2-related protein kinase that phosphorylates the retinoblastoma protein in vitro.

Members of the cell division cycle 2 (CDC2) family of kinases play a pivotal role in the regulation of the eukaryotic cell cycle. In this communication, we report the isolation of a cDNA that encodes a CDC2-related human protein kinase temporarily designated PITALRE for the characteristic Pro-Ile-Thr-Ala-Leu-Arg-Glu motif. Its deduced amino acid sequence is 47% identical to that of the human cholinesterase-related cell division controller (CHED) kinase, which is required during hematopoiesis, and 42% identical to the Saccharomyces cerevisiae SGV1 gene product, a putative kinase involved in the response to pheromone via its guanine nucleotide-binding protein alpha subunit. PITALRE expression is ubiquitous, but its expression levels are different in various human tissues. PITALRE is an approximately 43-kDa protein that associates with three cellular polypeptides of 80, 95, and 155 kDa. PITALRE is localized primarily to the nucleus. In addition, we have identified a retinoblastoma protein kinase activity associated with PITALRE immunocomplexes that cannot phosphorylate histone H1, suggesting that the target phosphorylation site of PITALRE differs from that of CDC2 kinase. Interestingly, the retinoblastoma kinase activity associated with PITALRE does not oscillate during the cell cycle.

Amino Acid Sequence↗
PubMed1994

Effects of taurine analogues on chloride channel conductance of rat skeletal muscle fibers: a structure-activity relationship investigation.

In rat skeletal muscle, taurine was proposed to interact with a low affinity binding site on sarcolemmal phospholipids near chloride channel, increasing chloride conductance (GCl). In an attempt to evaluate the structure-activity relationship between taurine and its binding site, a series of N-azacycloalkenyl analogues of taurine (A: N-(1'aza-cyclohepten-2'yl)-2-aminoethane sulfonic acid; B: N-(1'-aza-cyclopenten-2'-yl)-2-aminoethane sulfonic acid; C: N-(1'-aza-cyclohepten-2'-yl)-3-amino-propane sulfonic acid; D: N-(1'aza-cyclopenten-2'-yl)-3-aminopropane sulfonic acid) have been synthetized and tested in vitro on rat extensor digitorum longus (EDL) muscle. In spite of the presence of a bulky and lipophilic 5 or 7 membered heterocycle linked to the taurine amino group, analogues A and B determined an increase of GCl, although less potently than taurine. Also 3-amino-propane sulfonic acid (homotaurine), tested in comparison, showed less activity in increasing GCl with respect to taurine, probably for the increased distance between charged groups. Taurine analogues C and D, which differ from compounds A and B for an additional methylene group, showed much lower activity in increasing GCl. It has been reported that guanidinoethane sulfonate (GES) displaces taurine from the low affinity site on sarcolemma by only 7%. This compound, characterized by lower charge density on the guanidinium cationic head, applied in vitro on EDL muscle, show reduced taurine-like activity in increasing GCl.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Aging and chloride channel regulation in rat fast-twitch muscle fibres.

By the use of pharmacological tools, we tested the hypothesis that age-related alterations in the regulatory pathways of chloride channels might contribute to the lowered chloride conductance (GCl) found in skeletal muscle of aged rats. The resting GCl of extensor digitorum longus (EDL) muscles from adult rats either young (3-4 months old) or aged (29 months old) was measured by means of computerized intracellular microelectrode recordings. In EDL muscle from 3 to 4-month-old rats, 4-beta-phorbol 12,13-dibutyrate (4-beta-PDB), a direct activator of protein kinase C (PKC), decreased GCl in a concentration-dependent manner. The same effect was exerted by cholera toxin. The effects of both the phorbol ester and cholera toxin were inhibited by staurosporine, thus indicating that either direct or indirect (via G protein) activation of PKC accounts for the decrease of GCl. An increase of cytosolic Ca2+ by the ionophore A23187 also significantly decreased GCl by 25%. In EDL muscles from aged rats, 4-beta-PDB was 20-fold more potent in blocking GCl than in muscles from younger controls, and the ionophore blocked GCl by 40%. On the other hand, cholera toxin was ineffective. Our findings support the hypothesis that in fast-twitch muscle the regulation of chloride channels by PKC and Ca2+ is a target of the aging process.

Aging↗
PubMed1994

Opiate and opiate antidiarrhoeal drug action on rat isolated intestine.

1. Stimulation of segments of rat jejunum (2.5, 5, 10, 20 and 40 Hz for 8 s) and ileum (10 Hz) resulted in a fast atropine-sensitive contraction during stimulation and a non-cholinergic after-contraction. Stimulation of segments with single pulses at 0.1 Hz had no effect. The colon (10 Hz) usually responded only with single large atropine-sensitive contractions. 2. The responses of the jejunum (2.5-40 Hz, 8 s) were unaffected by the mu-receptor ligands morphine (0.3 microM) and RX 783006 (0.3 microM) or the kappa-receptor ligand, ethylketocyclazocine (0.3 microM). The cholinergic contraction of the colon was unaffected by ethylketocyclazocine (0.3 microM), but was reduced slightly by morphine at high concentrations only. 3. The prototype delta-opiate receptor ligand D-Ala2-D-Leu5-enkephalin (DADLE; 30 nM) reduced the contraction during stimulation (2.5, 5, 10 and 20 Hz for 8 s) of the rat jejunum. The IC50 value for inhibition of the contraction elicited by stimulation at 10 Hz was 3.2 in the jejunum and was 12 nM in the ileum. Contractions of the colon were only slightly inhibited at a high concentration of DADLE (1 microM). 4. DADLE had no effect on contractions elicited by acetylcholine, at a concentration of up to 3 microM in segments of jejunum. Naloxone (1 microM) abolished the inhibitory effect of DADLE on the cholinergic contraction of the electrically stimulated jejunum. In addition, naloxone caused a large enhancement of control responses to stimulation. 5. Only high concentrations of the antidiarrhoeals loperamide and diphenoxylate inhibited the contraction elicited at 10 Hz in the jejunum (IC50, 1.3 and 7.1 microM, respectively) and ileum (loperamide IC50, 0.5 microM). The same concentrations of these drugs also inhibited the effect of exogenously added acetylcholine. A similar pattern of findings was made in the colon. 6. The results show marked differences in responses to transmural stimulation and to opiate drugs compared to those previously obtained from guinea-pig ileum.(ABSTRACT TRUNCATED AT 400 WORDS)

Acetylcholine↗
PubMed1994

Effects of opiates on blood rheology.

To evaluate the possible effect of opiates on blood rheology, the plasma fibrinogen, prothrombin time and leukocyte aggregation were measured in 75 heroin addicts categorized by the time of abstention from heroin and the administration of naltrexone (25 active heroin abusers, 25 abstaining for 1 week, 11 abstaining for at least 5 months and 14 abstaining for 1 month and taking naltrexone during this period). No difference was detected in prothrombin time, but the leukocyte aggregation and fibrinogen were significantly different among the four groups (p = 0.028 and p = 0.0001, respectively). In particular, fibrinogen was 318 +/- 10.9 mg/dL in heroin abusers, significantly higher than that of the remaining three groups; the percentage of aggregated leukocytes was 5.01 +/- 0.77 in heroin users, significantly higher than that of subjects abstaining for at least 5 months. The fibrinogen levels declined sharply with abstention and an additive effect was noted with the administration of naltrexone, but leukocyte aggregation changed more slowly, and the effect of naltrexone (if any) was weaker. These data indicate an adverse effect of opiates on blood rheology and suggest that further studies should be performed to evaluate whether naltrexone may be useful in the prevention of major ischemic syndromes in patients with hyperfibrinogenemia and, perhaps, in those with high levels of leukocyte aggregation.

Adult↗
PubMed1994

Growth hormone administration to aged rats improves membrane electrical properties of skeletal muscle fibers.

Decreased secretion of growth hormone (GH) during aging may play a role in the impairment of muscle performance. To test this hypothesis we evaluated the effects of a chronic treatment with 150 micrograms/kg of GH on membrane electrical properties of extensor digitorum longus muscle of 23-month-old Wistar Kyoto female rats. We found that such treatment could partially but significantly reduce the age-related changes of these parameters. Indeed, with respect to untreated age-matched control rats we found a complete restoration of fiber diameter to the adult value, and a partial improvement of membrane resistance and capacitance values. Furthermore, the GH treatment significantly increased the low total membrane conductance characterizing aged muscle fibers. The increase of total membrane conductance was due to a significant increase toward the adult value of chloride conductance. In addition, the muscles from GH-treated aged rats had values of potassium conductance completely restored down to the adult ones. Accordingly, membrane excitability was also significantly improved by the treatment. The same treatment in adult female rats produced a slight but significant increase of both chloride conductance and potassium conductance, whereas in adult male rats we observed only a significant increase of fiber diameter. Our data show that impairment of GH secretion may contribute to the age-related changes of membrane electrical properties of skeletal muscle and support the potential benefit of GH administration to improve muscle performance in aged subjects.

Aging↗
PubMed1994

[Chronobiological standards of 24-hour arterial blood pressure validated by bayesian criteria].

The monodiagnosis of hypertension via ambulatory (A) blood pressure (BP) monitoring (M) is erroneous by definition. Its use has to be substituted by the chronodiagnosis. Because of the lack of an international standardization, the BP chronobiologic reference limits have to be computed on a small sample size. For this compilation, it is, however, unavoidable the recruitment of normotensive subjects using the WHO's fixed limits. These chronobiologic standards are by principle contaminated by false negatives and need a bayesian validation. Such a depuration has been made by reclassifying each subject using the BP time-qualified standards. A secondary sample was constituted on the true normotensive reclassified. This sample provided the a posteriori reference standards for BP. The bayesian chronobiologic reference standards seem to be the limits with which the chronodiagnosis of hypertension can be suitably done via ABPM.

Adult↗
PubMed1994

An E2F binding sequence negatively regulates the response of the insulin-like growth factor 1 (IGF-I) promoter to simian virus 40T antigen and to serum.

The promoter of the Insulin-like growth factor I (IGF-I) gene is activated by the Simian Virus 40 large T antigen (SVLT), and one of the elements responding to SVLT activation has been localized to a short 124 bp immediately upstream of the first initiation of transcription site. This short promoter contains an E2F binding site, that, in gel shifts, binds a protein complex, but only when the promoter activity is reduced or absent. A mutation in the E2F binding site deregulates the activity of the promoter, which becomes active even in those conditions in which the wild type promoter is inactive. By using antibodies in gel retardation analyses, we can show that the different protein complexes include, at least, the following proteins: E2F, cyclin A and p107. We conclude that the short IGF-I promoter is negatively regulated by an E2F binding site that complexes with several proteins. Our data suggest that disaggregation of these complexes by the action of SVLT (or other activators) increases expression from the promoter, thus establishing a link between the regulation of cell proliferation by growth factors and the E2F-associated proteins.

3T3 Cells↗
PubMed1994

PISSLRE, a human novel CDC2-related protein kinase.

The cell division cycle have been shown to be regulated by a closely-related family of protein kinases named CDKs (by cyclin-dependent kinases). Using a PCR-based cloning technique, we have isolated cDNAs encoding a human CDC2-related protein kinase. The full-length cDNA accommodates an open reading frame that does not contain any ATG initiation codon upstream of the sequence encoding the catalytic domain of this putative kinase. Three putative non-ATG initiation codons have been detected. Starting at the most 5' non-ATG initiation site, the encoded product is 316 amino acids long with a predicted molecular weight of 35.8 kDa. Analysis of the deduced amino acid sequence showed it to contain the XI subdomains present in all known protein kinases and a PSTAIRE-like motive, PISSLRE, which temporarily names this kinase. PISSLRE is most related to p58/GTA (55% identity in the catalytic domain), the galactosyl transferase associated protein, which has been shown to inhibit entry into S-phase when over-expressed in CHO cells. PISSLRE shares 38-45% identity with all CDKs and contains the regulatory Tyr and Thr residues present in most of the members of the CDK family of protein kinases, which suggests similar modes of regulation. PISSLRE is expressed in all human tissues tested, including those which contain high proportion of terminally differentiated cells. However, the levels of the PISSLRE transcripts are dissimilar among different tissues.

Amino Acid Sequence↗
PubMed1993

[Spontaneous hematoma of the rectus muscle].

The possibility of heart surgery and vascular surgery and the increased number of dialyzed patients receiving dicourmarin and heparin treatment raises the question of the probable rupture of the abdominal muscles, in particular the rectus muscles, following the onset of lancing abdominal pain. The authors report a clinical case referred to them in emergency conditions. They examine the possible etiopathogenetic causes and underline the symptoms of acute abdomen in patients being treated with anti-coagulating drugs.

Hematoma↗
PubMed1993

Blood pressure 24-hour pattern in two industrialized countries (Italy and Japan) with a different culture in salt intake.

This study investigates the blood pressure (BP) 24-hour pattern in representative samples of 2 industrialized countries, Italy and Japan, showing different cultures in salt intake. BP was monitored by means of a noninvasive ambulatory device whose readings were analyzed by means of chronobiometric procedures. The results show that the 24-hour BP pattern is not substantially different in Italian and Japanese subjects. In particular, the expected lower BP in the Italians was not detected despite their lower salt intake. Because the 24-hour mean BP value was seen not to be proportional to salt intake, the hypothesis is formulated that maintenance of the pressure regimen within a given range of variability is a principle of human physiology. To comply with this rule the Japanese people are supposed to have ethnically developed a certain resistance to dietary salt for which their cardiovascular apparatus is protected (phyletic escape to dietary sodium excess).

Adult↗