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Biomedical subjects

M Sala

Publications and source records attributed to M Sala.

At least 91 records · Page 5Linked to original sources
PubMed1996

Ambiguous base pairing of the purine analogue 1-(2-deoxy-beta-D-ribofuranosyl)-imidazole-4-carboxamide during PCR.

In principle the hydrogen bonding capacities of 1-(2-deoxy-beta-D-ribofuranosyl)-imidazole-4-carboxamide (dY), and its N-propyl derivative (dYPr), allow them to pair to all four deoxynucleosides. Their triphosphate derivatives (dYTP and dYPrTP) are preferentially incorporated as dATP analogues in a PCR reaction. However, once incorporated into a DNA template their ambiguous hydrogen bonding potential gave rise to misincorporation at frequencies of approximately 3 x 10(-2) per base per amplification. Most of the substitutions were transitions resulting from rotation about the carboxamide bond when part of the template. Between 11-15% of transversions were noted implying rotation of purine or imidazole moieties about the glycosidic bond. As part of a DNA template, dYPr behaved in the same way as dY, despite its propyl moiety. These deoxyimidazole derivatives are among the most radical departures from the canonical bases used so far as substrates in PCR and could be used to generate mutant gene libraries.

DNA↗
PubMed1996

An inverted U-shaped curve for heptylphysostigmine on radial maze performance in rats: comparison with other cholinesterase inhibitors.

The potential of heptylphysostigmine tartrate (pyrrolo [2,3b] indol-5-ol, 3,3a,8,8a-hexahydro-1,3a,8-trimethylheptylcarbamate [ester, (3aS-cis)]) (MF201), a new second-generation cholinesterase inhibitor, to antagonize scopolamine-induced amnesia in rats was assessed in an 8-arm radial maze. Upon completing the training session, the rats were orally administered increasing doses of MF201 (2, 3, 4, 6 and 8 mg/kg) 60 min prior to a s.c. injection of scopolamine (0.25 mg/kg). 9-Amino-1,2,3,4-tetrahydroamino-acridine hydrochloride hydrate (tacrine) (0.25, 0.37, 0.5, 1 and 2 mg/kg), 1-benzil-4-[(5,6-dimethoxy-1-indanon)-2-yl]-methyl piperidine (E2020) (0.125, 0.18, 0.25 and 0.5 mg/kg) and physostigmine (0.15, 0.25, 0.5 and 1 mg/kg) were orally administered and rats were tested in the same task. As previously described, scopolamine induced an impairment in radial maze performance, measured in terms of total number of errors, total time taken to complete the task and the percentage of amnesic animals. The reversal of scopolamine-induced impairment was characterized by the presence of an inverted U-shaped dose-response curve. A significant antagonistic effect was achieved with a dose (mg/kg) of 0.25 for E2020, 0.5 for tacrine and physostigmine and 3, 4 and 6 for MF201, the latter manifesting a broader spectrum of activity (3-6 mg/kg). While the maximal active doses restored the scopolamine-induced modified pattern of arm entry, they were ineffective in reducing hypermotility, suggesting the drugs have a specific effect on cognitive function.

Amnesia↗
PubMed1995

Human immunodeficiency virus type 1 reverse transcriptase tG:T mispair formation on RNA and DNA templates with mismatched primers: a kinetic and thermodynamic study.

The relationship between human immunodeficiency virus (HIV) type 1 reverse transcriptase tG:T mispair formation and base pair stability was investigated using DNA and RNA templates with 15 bp matched or mismatched DNA primers. tG:T mispair formation during primer elongation was undetectable on tDNA-DNA duplexes but occurred with a frequency of 10(-4) on matched tRNA-DNA duplexes. The frequency increased to 7.0 x 10(-4) and 1.3 x 10(-3) on tRNA-DNA duplexes with tG:T mismatches located 6 and 9 bp beyond the polymerization site. From Km values at 37 degrees C, the free energy change upon dissociation (delta G degrees 37) of the tG:T mispair increased from matched to mismatched tRNA-DNA duplexes by 0.36-1.21 kcal/mol. delta G degrees 37 for a correct tG:C pair decreased by 0.06-1.00 kcal/mol. In comparison with DNA-DNA duplexes, thermal melting measurements on RNA-DNA duplexes demonstrated smaller enthalpy (delta delta H degrees = -17.7 to -28.1 kcal/mol) and entropy (delta delta S degrees = -59.3 to -83.4 cal/mol/K) components. A strong entropy-enthalpy compensation resulted in small free energy differences (delta delta G degrees 37 = 0.8 to -2.2 kcal/mol). Thus, although DNA-DNA and RNA-DNA duplexes are of comparable stability in solution, the RNA-DNA duplex presents more facile base pair opening and higher conformational flexibility. The release of helical strain at constant helix stability in RNA-DNA duplexes may facilitate base mispairing during reverse transcription, particularly in the context of lentiviral G-->A hypermutation.

Base Composition↗
PubMed1995

Reverse transcriptase and substrate dependence of the RNA hypermutagenesis reaction.

G-->A hypermutation is a remarkable phenomenon resulting from retroviral reverse transcription in the presence of highly biased dNTP concentrations. Of the three reverse transcriptases (RTases) available, those of human immunodeficiency virus type 1 (HIV-1), avian myeloblastosis virus (AMV) and Moloney murine leukemia virus (MoMLV), the HIV-1 enzyme showed the greatest sensitivity to biased [dCTP]/[dTTP] ratios. The HIV-1 RTase was able to discriminate between dUTP, dITP and the four DNA precursors and was insensitive to pH. There was little preference for nucleotide contexts. A few exceptionally modified sequences were found presumably resulting from G-->A hypermutation and multiple strand transfer. This particular predilection of the HIV-1 and, by extrapolation, the lentiviral RTases towards G-->A hypermutation suggests that the phenomenon may have contributed to the remarkably elevated A content of these retroviral genomes.

Avian Myeloblastosis Virus↗
PubMed1995

Different kinetics of tolerance to behavioral and electroencephalographic effects of chlordiazepoxide in the rat.

The daily oral administration of chlordiazepoxide (40 mg/kg) over 9 weeks in rats elicited full tolerance to muscle relaxant effects within 7 weeks, as revealed by twice weekly evaluations of abdominal tone myorelaxation and decreased grip strength. No full tolerance was achieved, however, during the 9 weeks of treatment in terms of ataxia. Electroencephalographic (EEG) studies showed that this tolerance to the behavioural effects was accompanied by a progressive decrease in mean power spectra, associated with a progressive decrease in the beta band, but in this case, full tolerance was reached within 4 weeks. Once weekly evaluations of the ability of chlordiazepoxide to protect the animals against pentylenetetrazole seizures revealed a similar pattern. Treatment with flumazenil (50 mg/kg p.o.) 24 h after the last chlordiazepoxide administration induced a clear withdrawal syndrome associated with EEG changes which consisted of an increase in total power spectra associated with an increase in the delta band (in comparison with chlordiazepoxide-dependent rats not given the antagonist). These findings suggest that the different kinetics of the tolerance to anticonvulsant and EEG effects in comparison to myorelaxant effects can be attributed to a different involvement of benzodiazepine receptor subtypes.

Animals↗
PubMed1995

Behavioral and biochemical evidence of opioidergic involvement in cocaine sensitization.

Chronic administration of cocaine produces sensitization to its behavioral effects in humans and experimental animals. In the present study, rats treated with cocaine (10 mg/kg, i.p.) once daily for 10 days showed an enhancement in the acute drug stimulation of locomotor activity and stereotypy. Biochemical analysis in the nucleus accumbens of chronic cocaine-treated animals indicated that sensitization of D1 dopamine (DA) receptors had also developed. In fact, stimulation of adenylyl cyclase activity by DA was increased in nucleus accumbens membranes from sensitized rats. Our findings suggest that a novel postsynaptic mechanism, i.e., an increased DA-D1 receptor function, may play a role in the sensitization. A causal relationship between the two events is supported by the observation that neither motor behavioral sensitization nor DA-dependent adenylyl cyclase hyperactivity developed when the opiate antagonist naltrexone (2 mg/kg, s.c.) was given daily for 10 days before cocaine. When given alone, naltrexone was inactive in all respects, which rules out any unspecific action and suggests that its effects may be due to competition at receptors with endogenous opioids mobilized by cocaine. This was indirectly supported by the finding that desensitization of opioid inhibition of adenylyl cyclase developed in nucleus accumbens membranes of cocaine-sensitized rats. Chronic blockade of opioid receptors by naltrexone also counteracted the reinforcing properties of cocaine; conditioned place preference, clearly displayed by cocaine-treated animals, was antagonized in a dose-related manner. Overall, these results confirm that endogenous opioid peptides play an important role in cocaine addiction.

Adenylyl Cyclases↗
PubMed1995

Interleukin-6 production by human liver (myo)fibroblasts in culture. Evidence for a regulatory role of LPS, IL-1 beta and TNF alpha.

BACKGROUND/AIMS: Interleukin-6 is a major trigger for the synthesis of acute phase proteins by liver parenchymal cells. Acute phase proteins may contribute to the regulation of liver fibrosis by inhibition of proteases (e.g. collagenase) and by binding of cytokines. Since liver (myo)fibroblasts play an important role in the production of extracellular matrix in fibrotic livers, a study was undertaken into whether these cells are able to synthesize interleukin-6, which would give them the opportunity to contribute to regulation of synthesis of acute phase proteins by neighbouring parenchymal cells. METHODS: In the present study we investigated interleukin-6 production by two cell types obtained from human liver tissue: human fat-storing cells obtained from 5-15% Percoll fractions, which transformed in culture into myofibroblasts co-expressing alpha-smooth muscle actin and vimentin (VA cells) and fibroblasts obtained from 30-40% Percoll fractions which express vimentin only (V vells). Interleukin-6 production was measured in culture media of these cells using an enzyme-linked immunosorbent assay after incubation with lipopolysaccharide, and mediators like interleukin-1 beta, tumor necrosis factor-alpha, transforming growth factor-beta and interferon-gamma, known to be present in elevated concentrations in fibrotic livers. RESULTS: Unstimulated human liver (myo)fibroblasts produced considerable amounts of interleukin-6 (287 ng/mg cellular protein (VA cells), and 54 ng/mg cellular protein (V cells), within 48 h). Biological activity of these high concentrations of interleukin-6 measured in the enzyme-linked immuno-sorbent assay was confirmed in the B9-bioassay for interleukin-6 and by stimulation of alpha 2-macroglobulin production in rat liver parenchymal cell cultures. Lipopolysaccharide, interleukin-1 beta and tumor necrosis factor-alpha were potent stimulators of basal interleukin-6 production by VA and V cells, 1 microgram/ml lipopolysaccharide enhanced basal interleukin-6 production 3-fold within 48 h. 100 U/ml interleukin-1 beta and 1000 U/ml tumor necrosis factor-alpha each stimulated basal interleukin-6 production by VA cells 2-5 fold, whereas V cells were stimulated 10-25 fold. These effects were specific since the stimulation by lipopolysaccharide was completely inhibited by polymyxin B and the enhancing effects of interleukin-1 beta and tumor necrosis factor-alpha were neutralized by specific antibodies. Transforming growth factor-beta and interferon gamma did not influence interleukin-6 synthesis by either cell type in culture. CONCLUSIONS: These results indicate that transformed fat-storing cells (VA cells) and fibroblasts (V cells) may function as a local source of interleukin-6 in the human liver. Since interleukin-6 plays a key role in the regulation of the production of acute phase proteins by liver parenchymal cells, we hypothesize that human liver (myo)fibroblasts may stimulate local production of acute phase proteins in the fibrotic liver, thus contributing to local regulation of inflammatory and fibrogenic reactions.

Acute-Phase Proteins↗
PubMed1995

Collagen synthesis by human liver (myo)fibroblasts in culture: evidence for a regulatory role of IL-1 beta, IL-4, TGF beta and IFN gamma.

BACKGROUND/AIMS: Different cytokines have been described in fibrotic livers, including interleukin-1, interleukin-4 and interferon gamma, which are capable of regulating collagen production in human skin and lung fibroblasts. METHODS: To investigate possible involvement of interleukin-1, interleukin-4 and interferon gamma in the regulation of collagen production in human liver fibrosis, we studied the effects of these cytokines on collagen synthesis by nonparenchymal human liver cells in vitro. The effects of interleukin-1, interleukin-4 and interferon gamma were compared with the effect of transforming growth factor-beta, a well-known stimulator of collagen synthesis in liver fibrosis. Using a Percoll gradient we isolated two types of fibroblast-like cells from human liver tissue: fat-storing cells, which transformed in culture into myofibroblasts co-expressing vimentin and alpha-smooth muscle actin (VA-cells), and fibroblasts expressing vimentin only (V-cells). Production of collagen was measured in confluent cell cultures by incorporation of 3H-proline into collagenase degradable proteins. RESULTS: The cytokines studied had comparable effects on collagen synthesis in confluent cultures of VA-cells obtained from three different human livers and in confluent cultures of V-cells. Interleukin-1 beta and interleukin-4 enhanced collagen synthesis dose-dependently. 100 U/ml interleukin-1 beta stimulated collagen synthesis up to 174 +/- 25% (mean +/- sd, VA-cells) and 140 +/- 7% (V-cells) of control values. 1000 U/ml interleukin-4 enhanced collagen formation up to 195 +/- 58% (mean +/- sd, VA-cells) and 153 +/- 4% (V-cells) of control values after 48 h. These values were comparable to the stimulatory effects induced by transforming growth factor-beta (235 +/- 33% (mean +/- sd, VA-cells) and 150 +/- 18% of control values (V-cells) after incubation with 10 ng/ml transforming growth factor-beta for 48 h). Interferon gamma reduced both basal (36 +/- 29% (mean +/- sd) of control values in VA-cells, and 59 +/- 9% in V-cells) and transforming growth factor-beta induced collagen synthesis. CONCLUSIONS: These results indicate that in addition to the well-known role of transformed fat-storing cells (VA-cells) in collagen synthesis, fibroblasts (V-cells) may contribute to collagen production in human liver tissue. Moreover, these data demonstrate that in addition to the extensively documented collagen-inducing mediator transforming growth factor-beta, other cytokines present in fibrotic liver tissue like interleukin-1 beta and interleukin-4 may contribute to the enhanced synthesis of collagen, whereas interferon gamma may reduce collagen formation during liver fibrosis in man.

Cells, Cultured↗
PubMed1994

Relationship between morphine and etonitazene-induced working memory impairment and analgesia.

An 8-arm radial maze task was used to assess the possible role of the opiate system in the spatial memory of the rat. Increasing doses of etonitazene (0.005-0.06 mg/kg i.p.) and morphine (2.5-100 mg/kg i.p.) significantly impaired performance in the working memory components of the task. For both drugs this impairment was linearly related to the log of the administered dose, and the log-dose relationships were parallel. The regression lines calculated for each parameter for both drugs were parallel thus allowing us to calculate the potency: etonitazene proved to about 1000 times more potent than morphine in terms of correct arm entries, the number of errors and the total time taken to complete the task. Moreover, the progressive cognitive impairment produced by both opiates was closely related to an increase in analgesic effect. Pretreatment with naloxone (5 mg/kg i.p.) completely antagonised the disruptive effect of the opiates on working memory. The importance of the mu subtype opiate receptor in cognitive processes is discussed.

Analgesia↗
PubMed1994

G-->A hypermutation of the human immunodeficiency virus type 1 genome: evidence for dCTP pool imbalance during reverse transcription.

The quasispecies model for RNA viruses predicts the existence of a replication error threshold beyond which there is a melting or total loss of sequence information. Retroviral G-->A hypermutation is probably an example. Here it is shown that G-->A transitions may occur in both GpG and GpA dinucleotide contexts. Transitions in GpG preferentially occur via base mispairing at the ends of runs of G residues, whereas G-->A transitions within GpA may result from temporary dislocation of the primer and template strands by a single base. The two circumstances may be related by the local dCTP substrate concentration. An in vitro elongation assay shows that primer/template dislocation is more frequent for the human immunodeficiency virus type 1 reverse transcriptase than for murine or avian retroviral enzymes. Taken together these data suggest that G-->A hypermutation is an example of induced mutation whereby the viral reverse transcriptase is forced into making errors by imbalances in the intracellular dCTP concentration.

Base Composition↗
PubMed1994

Risk excess of soft-tissue sarcoma and thyroid cancer in a community exposed to airborne organochlorinated compound mixtures with a high hexachlorobenzene content.

Unusually high levels of hexachlorobenzene (HCB) were detected in the air and in sera of volunteers of a village located in the vicinity of an organochlorinated-compounds factory (Flix, Catalonia, Spain). A significant increase for specific causes of death was obtained only for neoplasms of unknown origin. However, an excess of incident cases was observed for thyroid neoplasms, soft-tissue sarcoma and brain neoplasms in men. These descriptive findings are in agreement with previously reported associations between soft-tissue sarcoma and human exposure to organochlorinated compounds, as well as with animal experiments relating HCB and thyroid cancer, and add new information on the possible relation between organochlorinated compounds, and particularly HCB, and cancer.

Air Pollutants↗
PubMed1994

PKU-related dysgammaglobulinaemia: the effect of diet therapy on IgE and allergic sensitization.

The effect of diet on the development of immunoallergic signs and symptoms in children with phenylketonuria (PKU) was evaluated. Immunological indices of 58 children with PKU treated with diets were compared to the immunological indices of 58 healthy (non-PKU) children. In the PKU group, 39 children had been placed on diet therapy within the first month of life; 19 children had been placed on diet therapy after 6 months of age. Total circulating lymphocytes; CD3+, CD4+, CD8+ circulating lymphocytes; and serum IgA, IgM, IgG and total IgE levels were measured for each child. Skin prick tests were performed for common inhalant and food allergens. Every 3 months over the 2-year period of this study, the signs and symptoms of eczema, allergic rhinitis and asthma were recorded. The PKU group had lower IgG levels (p = 0.004) and higher total IgE levels (p = 0.0001) than the control group. Significantly lower IgE levels were found in children started on diet therapy within the first month of life compared with those started on diet therapy after 6 months of age (p = 0.0001). Allergic sensitization was significantly more frequent in the PKU group (24/58 vs 13/58, z = 2.00, p < 0.05), but no significant difference in the incidence of eczema and allergic rhinitis was found. Asthma was less frequent in the PKU group than in the control group (5/58 vs 14/58, z = 2.09, p < 0.05). Thus, diet appeared to prevent the development of immunoallergic signs and symptoms.

Adolescent↗
PubMed1994

Changes in the level of blood suppressor CD8+CD57+ lymphocytes, when HIV-1 p24 antigen reappears in the blood.

Using the marker Leu2 + Leu7 + to detect the CD8 suppressor cells (CD88+CD57+), we have observed that their blood level, which increases after primoinfection, during the latent period, increases still further just before and when the p24 antigen returns and is present in the blood. Its ratio over cytotoxic CD8+ cells remains high during the AIDS period, though the absolute number decreases.

Acquired Immunodeficiency Syndrome↗
PubMed1994

Spatial discontinuities in human immunodeficiency virus type 1 quasispecies derived from epidermal Langerhans cells of a patient with AIDS and evidence for double infection.

A nonhomogeneous spatial distribution of human immunodeficiency virus type 1 quasispecies was observed for epidermal Langerhans cells purified from skin patches taken from a patient with AIDS soon after death. Each patch presented a unique collection of sequences, distinct from those of juxtaposed patches or those derived from the other leg. Infection of Langerhans cells by virus from underlying T cells in the dermis might explain this partition. The analysis revealed the presence of two distinct cocirculating viral strains, indicating double infection.

Acquired Immunodeficiency Syndrome↗
PubMed1994

Compliance with pharmacologic prophylaxis and therapy in bronchial asthma.

Between January 1 and December 31, 1988, 288 children [185 boys and 103 girls, mean age 8.75 +/- 4.98 years (range 2.50 to 16.83)], followed at the Outpatient Clinic for Lung Diseases of the University of Milan 5th Pediatric Department, were interviewed blindly in order to assess their compliance with pharmacologic therapy. All children were suffering from episodic, frequent, or chronic asthma requiring therapy as needed and preventive drugs for at least 30 days. Prophylaxis (including cromolyn, beclomethasone, theophylline retard, ketotifen, oxatomide, albuterol, and prednisone, alone or in combinations) and therapy in case of symptoms (albuterol, with or without theophylline prompt or beclomethasone) were prescribed. A study questionnaire was completed 30 to 45 days after the prescription with no advance warning by a physician unaware of the prophylactic and therapeutic prescriptions. Out of the 288 patients, 31 (10.8%) failed to return for the second visit. Understanding of prophylaxis was full in 61.1% of cases, partial in 23.3% and nil in 4.9%. Therapy as needed was fully remembered by 77.1% of parents, partially by 9.4% and totally forgotten by 2.8%. Compliance with single-drug prophylaxis was full in 60.1% of cases, partial in 22.2% and nil in 6.4%. Statistical analysis showed compliance was significantly better for ketotifen than for disodium cromoglycate (chi squared 9.85, P < .02), for ketotifen than for theophylline (chi squared 9.98, P < .02), and for beclomethasone than for theophylline (chi squared 8.77, P < .05).(ABSTRACT TRUNCATED AT 250 WORDS)

Administration, Inhalation↗
PubMed1994

[Andrological complications after pancystoprostatovesiculectomy surgery: utility of the bulbocavernosus reflex].

The technique for radical cystoprostatectomy was modified to avoid injury to the branches of pelvic plexus that innervate the corpora cavernosa (monolateral neurovascular bundle preservation or "Nerve sparing technique"). The studies of Walsh and coll. demonstrated that the branches of pelvic plexus that innervate the corpora cavernosa are situated between the rectum and urethra and penetrate the urogenital diaphragm near to the muscular wall of the urethra. Injuries to the pelvic plexus can occur during 1) division of posterior pedicle of bladder (the seminal vesicle can be used as a landmark intraoperatively to avoid injury to pelvic plexus), 2) during apical dissection of prostate with transection of the urethra. The return of sexual function postoperatively is related to preservation of autonomic innervation; the excision of the neurovascular bundle on one side may prevent impotence in 68% patients. Our study was undertaken to identify the cause of impotence in men undergoing radical cystoprostatectomy with "Nerve sparing technique" using bulbo cavernous reflex. Our results suggest that bulbo cavernosus reflex may not be a sensitive clinical tool to establish a diagnosis of neurogenic erectile dysfunction after pelvic surgery. The Authors examine the recent neuro-uro-physiological diagnostic methods for the study of neurogenic erectile dysfunction.

Aged↗