Biomedical subjects

G Esposito

Publications and source records attributed to G Esposito.

At least 55 records · Page 3Linked to original sources

PubMed2001

Cardiac overexpression of a G(q) inhibitor blocks induction of extracellular signal-regulated kinase and c-Jun NH(2)-terminal kinase activity in in vivo pressure overload.

BACKGROUND: Understanding the cellular signals that initiate cardiac hypertrophy is of critical importance in identifying the pathways that mediate heart failure. The family of mitogen-activated protein kinases (MAPKs), including the extracellular signal-regulated kinases (ERKs), c-Jun NH(2)-terminal kinase (JNK), and p38 MAPKs, may play specific roles in myocardial growth and function. METHODS AND RESULTS: To determine the mechanism of activation of MAPK pathways during the development of cardiac hypertrophy, we evaluated the induction of MAPK activity after aortic constriction in wild-type and in 2 types of cardiac gene-targeted mice: one overexpressing a carboxyl-terminal peptide of Galpha(q) that inhibits G(q)-mediated signaling (TG GqI mouse) and another overexpressing a carboxyl-terminal peptide of beta-adrenergic receptor kinase-1 that inhibits Gbetagamma signaling (TG betaARKct mouse). Wild-type mice with pressure overload showed an acute induction of JNK, followed by the induction of p38/p38beta at 3 days and ERK at 7 days. Both JNK and p38 activity remained elevated at 7 days after banding. In TG GqI mice, hypertrophy was significantly attenuated, and induction of ERK and JNK activity was abolished, whereas the induction of p38 and p38beta was robust, but delayed. By contrast, all 3 MAPK pathways were activated by aortic constriction in the TG betaARKct hearts, suggesting a role for Galpha(q), but not Gbetagamma. CONCLUSIONS: Taken together, these data show that the induction of ERK and JNK activity in in vivo pressure-overload hypertrophy is mediated through the stimulation of G(q)-coupled receptors and that non-G(q)-mediated pathways are recruited to activate p38 and p38beta.

Animals↗

PubMed2001

P27kip1 expression is associated with tumor response to preoperative chemoradiotherapy in rectal cancer.

BACKGROUND: Our aim was to ascertain whether or not the response to preoperative chemoradiotherapy for rectal cancer is associated with p27kip1 and p53 protein expression. METHODS: Thirty-eight patients (27 male, 11 female) with a mean age of 59 years (age range 33-87) and stage II-III rectal cancer received preoperative chemoradiotherapy (45-50.4 Gy; 5-FU 350 mg/m2/day and leucovorin 10 mg/m2/day). Thirty-one underwent low anterior resection; seven underwent abdominoperineal excision. Endoscopic tumor biopsies, performed before adjuvant therapy, were evaluated for: histologic type, tumor differentiation, mitotic index, and p27kip1 and p53 protein expression which were immunohistochemically determined. p53 expression was graded as: a) absent or present in < or =10% of tumor cells; b) present in 11-25%; c) present in 26-75%; and d) present in >75% of tumor cells. p27kip1 expression was assessed using both light microscopy (percent of stained cells x10 HPF) and cytometry with an image analysis workstation. Tumor response, ascertained with histology, was classified using a scale from 0 (no response) to 6 (complete pathologic response). RESULTS: The mitotic index for the endoscopic biopsies was low in 14 cases, moderate in 17 cases, and high in 7 cases. p53 protein expression was found in 21 (a), 3 (b), 3 (c), and 11 (d) cases. The mean percentage of cells expressing the p27kip1 protein was 34 (range 0-77.14%). A close correlation was found between cytometric and light microscopy findings for p27kip1 (r2 = 0.92, P = .0001). Tumor differentiation was good in 5 cases, poor in 2 cases, and moderate in the remaining 31 cases. While the response to adjuvant therapy was good/complete in 25 (65.78%) cases, it was absent/poor in 13 (34.21%) cases. Univariate analysis associated type of adjuvant therapy (chemoradiotherapy, P = .0428) and p27kip1 protein lower expression (P = .0148) with a poor response to adjuvant treatment. Stepwise linear regression found overexpression of p53 and p27kip1 and young age to be independent variables that were linked to a good response to adjuvant therapy. CONCLUSIONS: Lack of p27kip1 and p53 protein expression in rectal cancer is associated with a poor response to preoperative adjuvant therapy.

Adult↗

PubMed2001

DNA fragmentation in mammalian cells exposed to various light ions.

Elucidation of how effects of densely ionizing radiation at cellular level are linked to DNA damage is fundamental for a better understanding of the mechanisms leading to genomic damage (especially chromosome aberrations) and developing biophysical models to predict space radiation effects. We have investigated the DNA fragmentation patterns induced in Chinese hamster V79 cells by 31 keV/micrometer protons, 123 keV/micrometer helium-4 ions and gamma rays in the size range 0.023-5.7 Mbp, using calibrated Pulsed Field Gel Electrophoresis (PFGE). The frequency distributions of fragments induced by the charged particles were shifted towards smaller sizes with respect to that induced by comparable doses of gamma rays. The DSB yields, evaluated from the fragments induced in the size range studied, were higher for protons and helium ions than for gamma rays by a factor of about 1.9 and 1.2, respectively. However, these ratios do not adequately reflect the RBE observed on the same cells for inactivation and mutation induced by these beams. This is a further indication for the lack of correlation between the effects exerted at cellular level and the initial yield of DSB. The dependence on radiation quality of the fragmentation pattern suggests that it may have a role in damage repairability. We have analyzed these patterns with a "random breakage" model generalized in order to consider the initial non-random distribution of the DNA molecules. Our results suggest that a random breakage mechanism can describe with a reasonable approximation the DNA fragmentation induced by gamma rays, while the approximation is not so good for light ions, likely due to the interplay between ion tracks and chromatin organization at the loop level.

Animals↗

PubMed2001

Important role of endogenous norepinephrine and epinephrine in the development of in vivo pressure-overload cardiac hypertrophy.

OBJECTIVES: We sought to define the role of norepinephrine and epinephrine in the development of cardiac hypertrophy and to determine whether the absence of circulating catecholamines alters the activation of downstream myocardial signaling pathways. BACKGROUND: Cardiac hypertrophy is associated with elevated plasma catecholamine levels and an increase in cardiac morbidity and mortality. Although considerable evidence suggests that G-protein-coupled receptors are involved in the hypertrophic response, it remains controversial whether catecholamines are required for the development of in vivo cardiac hypertrophy. METHODS: We performed transverse aortic constriction (TAC) in dopamine beta-hydroxylase knockout mice (Dbh(-/-), genetically altered mice that are completely devoid of endogenous norepinephrine and epinephrine) and littermate control mice. After induction of cardiac hypertrophy, the mitogen-activated protein kinase (MAPK) signaling pathways were measured in pressure-overloaded/wild-type and Dbh(-/-) hearts. RESULTS: Compared with the control animals, cardiac hypertrophy was significantly blunted in Dbh(-/-) mice, which was not associated with altered cardiac function, as assessed by transthoracic echocardiography in conscious mice. The extracellularly regulated kinase (ERK 1/2), c-jun-NH(2)-terminal kinase (JNK) and p38 MAPK pathways were all activated by two- to threefold after TAC in the control animals. In contrast, induction of the three pathways (ERK 1/2, JNK and p38) was completely abolished in Dbh(-/-) mice. CONCLUSIONS: These data demonstrate a nearly complete requirement of endogenous norepinephrine and epinephrine for the induction of in vivo pressure-overload cardiac hypertrophy and for the activation of hypertrophic signaling pathways.

Animals↗

PubMed2001

Analysis of S-nitroso-N-acetylpenicillamine effects on dopamine release in the striatum of freely moving rats: role of endogenous ascorbic acid and oxidative stress.

1. We showed previously that interaction between NO and iron(II), both released following decomposition of sodium nitroprusside (SNP), accounted for the late SNP-induced dopamine (DA) increase in dialysates from the striatum of freely moving rats. 2. In this study, intrastriatal infusion of the NO-donor S-nitroso-N-acetylpenicillamine (SNAP) (0.2 mM for 180 min) induced a moderate increase in dialysate DA and decreases in ascorbic acid dialysate concentrations; in contrast, SNAP 1 mM infusion induced a long-lasting decrease in both DA and ascorbic acid dialysate concentrations. 3-Methoxy-tyramine (3-MT), dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), and uric acid levels were unaffected. 3. Co-infusion of ferrous sulphate [iron(II), 1 mM for 40 min] with SNAP either 1 or 0.2 mM (for 180 min), produced a significant increase in both DA and 3-MT dialysate concentrations, but it did not affect decreases in dialysate ascorbic acid levels. All other dialysate neurochemicals were unaffected. 4. Co-infusion of ascorbic acid (0.1 mM) with SNAP (1 mM) for 180 min did not modify SNAP-induced decreases in dialysate DA levels. In contrast, co-infusion of uric acid (1 mM) reversed SNAP-induced decreases in dialysate DA; co-infusion of a superoxide dismutase mimetic delayed SNAP-induced DA decreases for a short period, while co-infusion of the antioxidant N-acetylcysteine (NAC, 0.1 mM) significantly increased dialysate DA. 5. The results of this study show that SNAP induces concentration-related changes in DA dialysate levels. At higher concentrations, SNAP induces non-enzymatic DA oxidation, which is inhibited by uric acid and NAC; ascorbic acid failed to protect dialysate DA from oxidation, probably owing to its promoting effect on SNAP decomposition; exogenous iron(II) may react with NO generated from SNAP decomposition, with a consequent increase in dialysate DA and 3-MT, therefore mimicking SNP effects on striatal DA release.

Acetylcysteine↗

PubMed2001

A study on the role of nitric oxide and iron in 3-morpholino-sydnonimine-induced increases in dopamine release in the striatum of freely moving rats.

1. We showed previously that interaction between NO and iron (II), both released following the decomposition of sodium nitroprusside (SNP), accounted for the late SNP-induced dopamine (DA) increase in dialysates from the striatum of freely moving rats; in addition, we showed that co-infusion of iron (II) with the NO-donor S-nitroso-N-acetylpenicillamine mimicked SNP effects on striatal DA release. 2. In the present study, intrastriatal co-infusion of iron (II) (given as FeSO(4), 1 mM for 40 min) with the NO-donor and potential peroxynitrite generator 3-morpholinosydnonimine (SIN-1) (0.2, 0.5, 1.0 or 5.0 mM for 180 min), potentiated the SIN-1-induced increase in DA concentration in dialysates from the striatum of freely moving rats. Neither alone nor associated with iron (II) did SIN-1 induce changes in dialysate ascorbic acid or uric acid concentrations. 3. Neither co-infusion of a superoxide dismutase mimetic nor uric acid affected SIN-1-induced increases in dialysate DA concentration. 4. Infusion of the iron chelator deferoxamine (0.2 mM for 180 min) decreased dialysate DA and attenuated SIN-1-induced increases in dialysate DA concentrations. 5. These results suggest that iron plays a key role in SIN-1-induced release of striatal DA and do not support any role for either peroxynitrite or superoxide anion in SIN-1-induced release of striatal DA.

3,4-Dihydroxyphenylacetic Acid↗

PubMed2001

Cannabinoid CB1-receptor mediated regulation of gastrointestinal motility in mice in a model of intestinal inflammation.

1. We have studied the effect of cannabinoid agonists (CP 55,940 and cannabinol) on intestinal motility in a model of intestinal inflammation (induced by oral croton oil in mice) and measured cannabinoid receptor expression, endocannabinoids (anandamide and 2-arachidonylglycerol) and anandamide amidohydrolase activity both in physiological and pathophysiological states. 2. CP 55,940 (0.03 - 10 nmol mouse(-1)) and cannabinol (10 - 3000 nmol mouse(-1)) were more active in delaying intestinal motility in croton oil-treated mice than in control mice. These inhibitory effects were counteracted by the selective cannabinoid CB(1) receptor antagonist SR141716A (16 nmol mouse(-1)). SR141716A (1 - 300 nmol mouse(-1)), administered alone, increased intestinal motility to the same extent in both control and croton oil-treated mice. 3. Croton oil-induced intestinal inflammation was associated with an increased expression of CB(1) receptor, an unprecedented example of up-regulation of cannabinoid receptors during inflammation. 4. High levels of anandamide and 2-arachidonylglycerol were detected in the small intestine, although no differences were observed between control and croton oil-treated mice; by contrast anandamide amidohydrolase activity increased 2 fold in the inflamed small intestine. 5. It is concluded that inflammation of the gut increases the potency of cannabinoid agonists possibly by 'up-regulating' CB(1) receptor expression; in addition, endocannabinoids, whose turnover is increased in inflamed gut, might tonically inhibit intestinal motility.

Analgesics↗

PubMed2001

G protein-coupled receptor signalling in in vivo cardiac overload.

Cardiac myocytes respond to biomechanical stress by initiating cellular processes that lead to hypertrophy. Although cardiac hypertrophy is a response to increased stress on the heart, it is associated with elevated plasma catecholamine levels and an increase in cardiac morbidity and mortality. Understanding the cellular signals that initiate the hypertrophic response will be of critical importance to identify pathways that mediate the maladaptive deterioration of the hypertrophic heart to one of cardiac failure. This review will focus on the role of G protein-coupled receptors in the activation of signalling pathways in the heart, such as the mitogen activated protein kinase and phosphoinositide-3 kinase pathways.

Animals↗

PubMed2001

Selective cannabinoid CB1 receptor-mediated inhibition of inducible nitric oxide synthase protein expression in C6 rat glioma cells.

We have studied the effects of two cannabinoid receptor agonists, WIN 55,212-2 and cannabinol, on nitric oxide (NO) production and inducible nitric oxide synthase (iNOS) expression in the C6 glioma cell line. After 24 h of lipopolysaccharide (LPS) (1 microg/mL) and interferon-gamma (IFN-gamma) (300 U/mL) stimulation, a significant increase in NO production, evaluated as nitrite, was observed in the culture medium. WIN 55,212-2 (0.1-10000 nM) and cannabinol (0.3-30000 nM), dose-dependently inhibited nitrite production showing a different potency (WIN 55,212-2 EC(50): 4.2 nM; cannabinol EC(50): 700 nM). WIN 55,212-2 (100 nM), given concomitantly to the stimulus also inhibited iNOS expression but had no effect when added to the cells 2 h after LPS/IFN-gamma, indicating a possible interference at the protein synthesis level or at an earlier step, as gene transcription. The cannabinoid CB1 receptor antagonist, SR141716A (0.1-100 nM), but not the cannabinoid CB2 receptor antagonist, SR144528 (0.1-100 nM), reduced in a dose-related manner WIN 55,212-2-and cannabinol-induced inhibition of nitrite production. SR141161A also reversed the WIN 55,212-2-induced inhibition of iNOS expression. These data suggest that selective cannabinoid CB1 receptor activation, by inhibiting iNOS expression and NO overproduction in glial cells, might be helpful in NO-mediated inflammation leading to neurodegeneration.

Analgesics↗

PubMed2001

Transglutaminase 1 gene mutations in Italian patients with autosomal recessive lamellar ichthyosis.

We analyzed the transglutaminase 1 gene locus in patients from six unrelated Italian families affected by autosomal recessive lamellar ichthyosis. In two families we identified a novel mutation (E520G) in the gene coding region, a previously reported splicing mutation (A3447G), and the mis-sense mutations S272P and V518M. The latter mutation, hitherto considered disease causing, was found to be a simple polymorphism. Linkage to transglutaminase 1 gene was excluded in two of the other four families examined. Single strand conformational polymorphism analysis of the transglutaminase 1 gene in the remaining two families did not reveal any alteration in the coding region. This finding confirms the genetic heterogeneity of the disease.

DNA, Recombinant↗

PubMed2001

Results and complications of laparoscopic surgery for pediatric varicocele.

BACKGROUND: The aim of this study was to evaluate the results and complications of laparoscopic varicocelectomy in children. METHODS: Over a 36-month period, 211 children underwent laparoscopic treatment of varicocele. Their ages ranged between 6 and 17 years; the varicocele was located on the left side in 209 cases (99.1%) and was bilateral in 2 (0.9%). In 195 patients the laparoscopic transperitoneal approach was used and in 16 retroperitoneoscopy was used. Thirty children (14.2%) underwent ligation of the veins alone, and 181 (85.8%) underwent ligation of testicular veins and artery. In 15 (7.1%) cases an additional procedure was applied during the same operation. RESULTS: Average operating time was 30 minutes and hospitalization about 24 hours. At an average follow-up of 26 months, there were 19 (9%) postoperative complications: 14 children had a left hydrocele, 3 children a scrotal emphysema, and 2 an umbilical granuloma. There were 5 recurrences of varicocele in our series: 2 (2 of 30, 6.6%) after the Ivanissevitch procedure, and 3 (3 of 181, 1.6%) after Palomo's. Testicular atrophy did not occur in any patient of this series. CONCLUSIONS: This preliminary experience shows that the results of the laparoscopic approach are comparable to those of the open approach. The ligation of testicular veins and artery is preferable to the ligation of the testicular veins alone. Hydrocele seems to be the most frequent postoperative complication and a potential problem, especially in children operated on with the Palomo procedure.

Adolescent↗

PubMed2001

Amyloid fibrils derived from the apolipoprotein A1 Leu174Ser variant contain elements of ordered helical structure.

We recently described a new apolipoprotein A1 variant presenting a Leu174Ser replacement mutation that is associated with a familial form of systemic amyloidosis displaying predominant heart involvement. We have now identified a second unrelated patient with very similar clinical presentation and carrying the identical apolipoprotein A1 mutation. In this new patient the main protein constituent of the amyloid fibrils is the polypeptide derived from the first 93 residues of the protein, the identical fragment to that found in the patient previously described to carry this mutation. The X-ray fiber diffraction pattern obtained from preparations of partially aligned fibrils displays the cross-beta reflections characteristic of all amyloid fibrils. In addition to these cross-beta reflections, other reflections suggest the presence of well-defined coiled-coil helical structure arranged with a defined orientation within the fibrils. In both cases the fibrils contain a trace amount of full-length apolipoprotein A1 with an apparent prevalence of the wild-type species over the variant protein. We have found a ratio of full-length wild-type to mutant protein in plasma HDL of three to one. The polypeptide 1--93 purified from natural fibrils can be solubilized in aqueous solutions containing denaturants, and after removal of denaturants it acquires a monomeric state that, based on CD and NMR studies, has a predominantly random coil structure. The addition of phospholipids to the monomeric form induces the formation of some helical structure, thought most likely to occur at the C-terminal end of the polypeptide.

Amino Acid Substitution↗

PubMed2001

Pressure-independent enhancement of cardiac hypertrophy in natriuretic peptide receptor A-deficient mice.

Mice lacking natriuretic peptide receptor A (NPRA) have marked cardiac hypertrophy and chamber dilatation disproportionate to their increased blood pressure (BP), suggesting, in support of previous in vitro data, that the NPRA system moderates the cardiac response to hypertrophic stimuli. Here, we have followed the changes in cardiac function in response to altered mechanical load on the heart of NPRA-null mice (Npr1-/-). Chronic treatment with either enalapril, furosemide, hydralazine, or losartan were all effective in reducing and maintaining BP at normal levels without affecting heart weight/body weight. In the reverse direction, we used transverse aortic constriction (TAC) to induce pressure overload. In the Npr1-/- mice, TAC resulted in a 15-fold increase in atrial natriuretic peptide (ANP) expression, a 55% increase in left ventricular weight/body weight (LV/BW), dilatation of the LV, and significant decline in cardiac function. In contrast, banded Npr1+/+ mice showed only a threefold increase in ANP expression, an 11% increase in LV/BW, a 0.2 mm decrease in LV end diastolic dimension, and no change in fractional shortening. The activation of mitogen-activated protein kinases that occurs in response to TAC did not differ in the Npr1+/+ and Npr1-/- mice. Taken together, these results suggest that the NPRA system has direct antihypertrophic actions in the heart, independent of its role in BP control.

Animals↗

PubMed2001

Persistence of delayed adrenarche in boys with thalassemia.

5 years after a previous study, we followed up a group of thalassemic patients, determining DHEA-S levels in peripubertal age, with the aim of evaluating whether adrenarche maturation occurred in boys and advanced in girls. Furthermore, we evaluated the degree of bone mineral density (BMD SDS(BA)) and analyzed growth parameters calculating standard deviation score with respect to bone age (BA) of height (Ht SDS(BA)), sitting height (SH SDSBA), and subischial leg length (SLL SDSBA), body mass index (BMI) and the difference between the values of the previous and the present study (deltaBMI), thyroid function and serum markers of bone metabolism. Our results showed persistent lack of adrenarche (DHEA-S 25+/-9.5 microg/dl) in all 6 boys and the absence of pubertal signs at chronological age (CA) of 12.4+/-1.4 yr and BA of 11.1+/-1.1 yr. Only one boy, 6 months later, showed a testicular volume of 4 ml (Tanner stage G2) with an increase of DHEA-S value (181 microg/dl) at BA 12.8 yr. Body disproportion and severe degree of osteopenia (BMD SDSBA -2.41+/-0.5) were observed in all boys, even though Ht SDSBA (0.14+/-0.8) and markers of bone metabolism were within the normal range. No change in nutritional status was observed (deltaBMI 0.09+/-0.4 kg/m2). In contrast, all the thalassemic girls had DHEA-S values (172.7+/-97.7 microg/dl) within the normal range at BA 12.7 +/-0.6 yr that was similar to CA. Furthermore, the appearance of Tanner stage B2 occurred in each of them at BA, near to CA, of 10.4+/-0.9 yr, and menarche was observed in three of them at mean BA, near to CA, of 11.4+/-0.9 yr. Ht SDSBA was below normal range (-1.11+/-0.8), but SLL SDSBA and SH SDS(BA) values were reduced homogeneously, so that proportional body growth was observed. A significant change in nutritional status was observed (deltaBMI 2.69+/-0.9 kg/m2). Bone density value (BMD SDS(BA) -0.25+/-0.4) was in the normal range. There were no statistically significant differences between boys and girls for ferritin serum levels, blood consumption and desferrioxamine dosage. In conclusion, lack of change in nutritional status, measurable in the form of deltaBMI, but not BMI alone, considered an important physiological regulator of adrenarche, regardless of individual adrenal androgen secretion, could have a key role in the lack of adrenarche persisting in thalassemic boys during peripubertal age. Further follow up is necessary, in particular when boys reach puberty, because delayed adrenarche represents the most intriguing aspect in these patients.

Adolescent↗

PubMed2001

Influence of primary sedimentation on pre-denitrification system performances.

The effects of suppressing primary sedimentation on nitrogen removal efficiency of a pre-denitrification system have been evaluated for a large municipal wastewater treatment plant. Simulations have been carried out using the STOAT model. For both the process schemes with and without primary sedimentation, nitrification efficiencies are calculated for increasing influent loads of COD, total N and suspended solids. The sensitivity analysis shows that for the usual carbon to nitrogen ratios in the raw influent both the process schemes allow the requested removal efficiencies, whereas for significantly high C/N ratios the scheme with primary sedimentation is preferable.

Carbon↗

PubMed2001

Prognostic value of serum biological markers in patients with hepatocellular carcinoma.

PURPOSE: Prognosis of patients with hepatocellular carcinoma (HCC) is assessed by using indexes based on clinical and instrumental parameters. The Cancer of the Liver Italian Program (CLIP) staging system combines the Child-Pugh classification with tumor size, portal invasion, and alpha-fetoprotein and predicts the outcome of HCC patients more precisely than the Okuda staging system. Serum levels of a number of biological variables have been found to be increased in patients with HCC and are associated with different outcomes. Our aims in this study were to test the prognostic role of the serum levels of soluble intercellular adhesion molecule-1 (sICAM-1), soluble interleukin-2 receptor (sIL-2R), interleukin 6 (IL-6), and anti-p53 and to assess whether the addition of any of the above serum markers could further improve the predictive ability of the CLIP score. EXPERIMENTAL DESIGN: Serum levels of sICAM-1, sIL-2R, IL-6, and anti-p53 were assayed in 80 patients with HCC and correlated with their outcomes. Nonparametric procedures were applied to test correlations between serum sICAM-1, sIL-2R, IL-6, anti-p53, and other prognostic factors. For survival analyses, the product-limit method, log-rank test, and Cox proportional hazards model were applied. RESULTS: Only serum levels of sIL-2R correlated with survival, which was longer for patients with lower values (< or =950 units/ml). However, with multivariate analysis sIL-2R did not confirm its predictive role when tested with the CLIP score as a covariate, with a hazard of death of 1.51 (95% confidence interval, 0.76-3.01). CONCLUSIONS: Serum levels of sICAM-1, sIL-2R, IL-6, and anti-p53 are not useful as prognostic factors for HCC in clinical practice. They do not improve the predictive ability of the CLIP score.

Aged↗

PubMed2001

DNA fragmentation induced in K562 cells by nitrogen ions.

This study was aimed at investigating the radiation induced DNA fragmentation pattern as a function of cellular differentiation and radiation quality. DNA double strand breaks (DSB) induced by gamma-rays were analyzed in K562 human proerythroblasts before (AP cells) and after (D cells) differentiation induction while DNA DSB induced by 125 keV/micrometers N-ions have been studied in AP cells. Pulsed-Field Gel Electrophoresis (PFGE) of cellular DNA was used to determine the DSB yield by analysis of the Fraction of Activity Released (FAR) and of the fragmentation pattern in a specific size range (5.7-0.225 Mbp). The results so far obtained show that the DSB induction by gamma-rays is different if evaluated with the FAR or with the fragmentation analysis. The DSB yield obtained with the former method is about 1.4 times higher in AP respect to D cells while the latter method indicates that more fragments are produced in D cells. Comparison between gamma-rays and N-ions in AP cells shows that no significant differences are detected by the FAR analysis; otherwise fragmentation analysis demonstrates a higher effectiveness of nitrogen ions.

Cell Differentiation↗

PubMed2000

Disruption of the Rev3l-encoded catalytic subunit of polymerase zeta in mice results in early embryonic lethality.

Polymerase zeta (Pol zeta) is an error-prone DNA polymerase [1], which in yeast is involved in trans-lesion synthesis (TLS) and is responsible for most of the ultraviolet (UV) radiation-induced and spontaneous mutagenesis [2-4]. Pol zeta consists of three subunits: REV1, a deoxycytidyl-transferase [5]; REV7, of unclear function [6]; and REV3, the catalytic subunit. REV3 alone is sufficient to carry out TLS, but association with REV1 and REV7 enhances its activity [5, 7]. Experiments using human cells treated with UV radiation indicate also that mammalian Pol zeta is involved in TLS [7]. The peculiar mutagenic activity of Pol zeta [4,7,8] suggests a possible role in somatic hypermutation of immunoglobulin (Ig) genes [9]. Here, we report that, unlike in yeast where the REV3 gene is not essential for life [4], disruption of the mouse homologue (Rev3l) resulted in early embryonic lethality. In Rev3l(-/-) embryos, no haematopoietic cells other than erythrocytes could be identified in the yolk sac. Rev3l(-/-) haematopoietic precursors were unable to expand in vitro and no haematopoietic cells could be derived from the intraembryonic haematogenic compartment (splanchnopleura). Fibroblasts could not be derived from the Rev3l(-/-) embryos, and Rev3l(-/-) embryonic stem (ES) cells could not be obtained. This is the first evidence that an enzyme involved in TLS is critical for mammalian development.

Animals↗