PubMed Health⌕ Search

Biomedical subjects

M P Viola-Magni

Publications and source records attributed to M P Viola-Magni.

At least 19 recordsLinked to original sources
PubMed2003

Chromatin-associated sphingomyelin: metabolism in relation to cell function.

After the first histochemical demonstration by Chayen and Gahan of the presence of phospholipids and especially of sphingomyelin in chromatin, this became the object of long debate and of contradictory results. The general conclusion was that the presence of phospholipids may due to contamination during the isolation of chromatin. More recently the existence of a phospholipid chromatin fraction was confirmed by demonstrating that isolated hepatocyte nuclei, labelled by saturated and unsaturated radioiodination method, showed the presence of radioactivity only in the membrane and not in the isolated chromatin. The phospholipid composition showed an enrichment in sphingomyelin which increased during hepatocyte maturation or erythroleukemic cell differentiation induced by DMSO. A decrease in sphingomyelin was observed at the beginning of the S-phase in regenerating liver or in cultured proliferating cells. These changes were due to the presence of sphingomyelinase and sphingomyelin synthase in the chromatin, the activity of which paralleled the variation in sphingomyelin content. The sphingomyelin was co-localized with RNA as shown by biochemical and electron microscopy methods. Using bromo-uridine it was demonstrated that labelled RNA and sphingomyelin were present in actively transcribing nuclear regions. Isolated nuclear complexes after DNase and RNase digestion contained not only protein, but also RNA and sphingomyelin. After hydrolysis of sphingomyelin the RNAse-resistant RNA becomes RNAse sensitive. It can therefore be concluded that sphingomyelin and the related enzymes are present in the chromatin; sphingomyelin may have a role in RNA transcription protecting RNA by RNAse digestion before its transfer to the cytoplasm.

Animals↗
PubMed1997

Choline base exchange activity in rat hepatocyte nuclei and nuclear membranes.

Previous investigations have demonstrated the presence of phospholipids as a component of chromatin; however the mechanism of their synthesis, namely if they are synthesized in the nuclei or in the cytoplasm (microsomal fraction), from where they may eventually be transported to the nucleus, has not yet been clarified. The phosphatidylcholine, for example, can be formed, albeit in a limited amount, by an interconversion reaction between bases. The aim of the present research was to ascertain the presence of the enzyme complex responsible for this reaction in hepatocyte nuclei and in isolated nuclear membrane. The incorporation of [14C]-choline in phosphatidylcholine was assayed in microsomes, hepatocyte nuclei, liver nuclei and nuclear membranes of rat liver. The reaction was Ca(2+)-dependent and the specific activity was higher in microsomes but was present, albeit at a low level, also in nuclei and in nuclear membranes. Possible contaminations were excluded by specific microsomal markers and by the reaction time course. In fact, the nuclear reaction reached the maximum level slowly with respect to microsomes. Since the phosphatidylcholine extracted from the nuclei show an enrichment in unsaturated fatty acids of monoenoic fraction, such as oleic acid, the difference in reaction kinetics has been tentatively explained as due to the phosphatidylcholine fatty acid content. The presence of this base exchange enzyme complex may allow a fast change in chromatin phospholipid composition.

Animals↗
PubMed1996

[Hepatic resection in the fetal rabbit. Histologic comparison of tissue regeneration in the fetus versus the adult].

Fetal tissues present peculiar features of repair after injury. Although the development of fetal hepatocytes have already been studied in vitro and in transplant models, an in vivo study of fetal liver regeneration is still missed in the literature, to the best of our knowledge. Eight time-dated pregnant California rabbits (23, 24, 25, 30 days of gestational age) and 2 adult male California rabbits were anesthetized following a standardized i.v. protocol (ketamine 50 mg/kg; xilazine 5 mg/kg; propiopromazine 0.75 mg/kg; spontaneous breathing; no anesthetic gas). All the pregnant does underwent a midline laparotomy and a minimal hysterotomy to approach a fetus per each animal. In 2 cases, 1 fetus was delivered and prior to sacrifice the fetal liver was sampled in toto (30 days of gestational age). These pregnancies were allowed to continue to term and were uneventful with a full-term spontaneous delivery of the remaining fetuses. In the other 6 pregnancies, after the hysterotomy, the fetal abdomen was entered through a right-sided longitudinal incision and the liver was partially resected by thermocauterization. Fetal abdomen was closed in 1 layer (non absorbable suture 7-0). The fetus was then returned in the uterus and, after amniotic fluid restoration with warmed saline, the hysterotomy was sutured in double layer (polyglycolic 5-0). Maternal abdomen was closed in 1 layer (polyglycolic 4-0) and the skin in a continuous overlying fashion (silk 3-0). The abdominal cavity of the 2 adult male rabbits was entered through a right subcostal incision. Partial liver resection was performed, and abdominal and skin closure followed the same techniques used for the pregnant does. The treated livers were then sampled in toto at 24, 48, 72 hrs and 4 days after surgery from the fetuses, and at 7 days from the adult rabbits. Histological stains were: H & E; Van Gieson; Masson; Alcian Bleu; PAS. Fetal histology showed a low inflammatory reaction poor in PMN cells with minimal deposition of collagen and a high amount of glycogen in the hepatocytes. The inflammatory response to resection was much more evident in the adult samples as much as the abundant intra and extra-cellular deposition of collagen associated to a minor amount of intracellular glycogen. The peculiar features of liver regeneration in the fetus, deserve further experimental studies.

Animals↗
PubMed1994

Different expression of beta-N-acetylhexosaminidase in mouse tissues.

The expression of genes encoding for the alpha and beta-subunits of the lysosomal enzyme beta-N-acetylhexosaminidase was investigated in different mouse tissues. It was found, using fluorogenic substrates, that the amounts of alpha and beta subunits were not the same in different tissues: alpha-subunit was more abundant in the brain, beta-subunit in epididymis and brain. The different isoenzyme patterns and specific activities in mouse tissues are due to the differences in the amount of hexosaminidase subunits. The mRNA, evaluated by Northern blotting analyses, revealed a greater expression of alpha-subunit in the testis and of beta-subunit in the brain and epididymis. The results indicate, therefore, that gene expression and the amount of subunits are in good relationship for beta-subunit, whereas there is no correlation for alpha-subunit.

Animals↗
PubMed1994

Rat liver chromatin phospholipids.

To shed light on the question whether the phospholipids present in chromatin are native or are due to contamination from nuclear membranes, we labeled the phospholipids of isolated nuclei and determined the amount of phospholipids (PL) and PL fatty acid composition in nuclei and chromatin. The hepatocyte nuclei were isolated and radioiodinated by the lactoperoxidase method under saturating and nonsaturating conditions, and the radioactivity associated with chromatin extracted from these nuclei was monitored. Whereas 97% the label was recovered in the nuclear membranes, only 0.08-0.6% was found in chromatin. The PL present in chromatin were relative to the amounts present in the entire nuclei and calculated as percentage of total, phosphatidylethanolamine (10%), phosphatidylserine (22%), phosphatidylinositol (19%) phosphatidylcholine (14%), and sphingomyelin (35%). In sphingomyelin of chromatin-associated PL an enrichment in polyunsaturated fatty acids was seen. The data indicated that the PL found in isolated chromatin do not seem to be due to contamination from the nuclear membrane.

Animals↗
PubMed1994

Changes in oncogene expression in ascite tumour cells during ageing.

The expression of two oncogenes, c-myc and c-fos, was studied in an ascitic tumour (ATPC+) at different times after implantation. The specific mRNA synthesis was analysed by Northern blot analysis. The presence of the oncogene proteins was shown by immunofluorescence using flow cytometry and referred to the distribution of the cells in the different cell phases. The results show that both oncogenes are expressed by ATPC+ tumour cells. c-myc is expressed 5, 8 and 12 days after implantation, although with a different intensity, and the protein is mainly present in S or S+G2 phase cells. The c-fos oncogene is expressed only 12 days after tumour implantation and the cells labelled with the specific antibody are mainly in G1 phase. We conclude that c-myc is principally correlated with proliferative activity, whereas c-fos is expressed by non-cycling cells.

Animals↗
PubMed1992

TEM cytochemical study of the localization of phospholipids in interphase chromatin in rat hepatocytes.

The electron microscopy cytochemical detection of phospholipids in well-defined areas in the interphase nuclei of hepatocytes has been obtained by the acid haematein test, modified for electron microscopy and by the phospholipase A2-colloidal gold method. The specificity of both methods were controlled by enzymatic digestion with phospholipase. The main intra-nuclear localization of phospholipids is at the border between the condensed and dispersed chromatin, where non-ribosomal RNA is also revealed by RNase-gold labelling. Phospholipids are detected, too, over the clusters of interchromatin granules and in the fibrillar component of the nucleolus.

Animals↗
PubMed1991

Age-related changes in the cell proliferation of ATPC+ mouse ascites tumour.

The growth of ATPC+, an ascites tumour derived from a spontaneous mammary carcinoma in BALB/c+ mice, was studied at different ages. It was observed that the number of cells increases rapidly during the first 5 days after implantation. Thereafter, the cell number increases more slowly, reaching a plateau after 8 days. This slowing-down is not due to a reduction in the growth fraction but to a lengthening of the cell cycle. Between 5 and 8 days the duration of all phases increases, including the S phase, which increases from 5.2 h in 5-day tumours to 8.2 h in 8-day tumours. In 12-day tumours both the cell cycle and S phase are only slightly longer than in 8-day tumours whereas the growth fraction is reduced. The slowing-down of cell growth can be attributed to growth fraction reduction rather than cell loss, which is maximal in the 5-day tumour. At this age the time course of the percent labelled cells and of the number of grains/nucleus suggests reutilization of [3H]-thymidine. Incorporation of [3H]-thymidine/cell decreases sharply in 12-day tumours due to a reduced availability of thymidine, which is degraded to thymine in the in vivo ascitic fluid faster than in 8-day tumours. This indicates an age-related change in the ascitic fluid composition.

Aging↗
PubMed1987

Synthesis of chromatin phospholipids.

The presence of a phospholipid fraction associated with chromatin has been demonstrated by biochemical technique in rat hepatocytes. The composition of this fraction determined by chromatography with respect to that of the nuclei is characterized by low content of phosphatidylserine and high content in phosphatidylethanolamine. Also the synthesis and turnover studied after injection of [32P]O4(2-) show a different behaviour: the peak of activity is after 6 hrs in nuclei and microsomes, whereas in chromatin it occurs after 9 hrs. A second peak is evident after 24 hrs in chromatin and microsome phospholipids. Differences have been also shown by analyzing the single phospholipid radioactivity in time. The behaviour of chromatin phospholipids has also been studied during DNA premitotic synthesis in regenerating liver. It has been shown that there is no difference in synthesis in relation to that of DNA in nuclear phospholipids, whereas the specific activity of chromatin phospholipids begins to increase twelve hours after hepatectomy and continues throughout the period of the first mitotic wave, thus bringing to a summation with the beginning of the second wave. The role of this phospholipid fraction in relation to DNA synthesis and gene expression is discussed.

Animals↗
PubMed1987

Cytochemical and autoradiographic studies of the localization and turnover of chromatin and nucleolar associated phospholipids in plant and animal tissues.

The localization of chromatin-associated phospholipids has been demonstrated on chromosomes and on chromatin of interphase nuclei by cytochemical methods either in plant and in animal tissues. Three methods of fixation are suggested which can be combined which two cytochemical methods for phospholipids detection. An additional method is represented by autoradiographic technique after incorporation of a radioactive precursor such as [3H] ethanolamine. This method has been used with good results in nuclei of lateral root apices of Vicia faba on 1 micron thick section, thus avoiding any possible contamination by nuclear membrane. In these nuclei the phospholipids appear associated with the chromatin and more intensely with the nucleolar regions. The positivity of the cytochemical reaction disappears after extraction of fixed tissue with acidified methanol chloroform and after treatment of unfixed sections with phospholipase D. The use of phospholipid precursor has allowed the study of chromatin-phospholipids synthesis in root apices of Vicia faba with respect to timings of the cell cycle. The results show that there is a strong case for a pattern of chromatin phospholipid synthesis which operates during S phase. Concerning the role of phospholipids it is suggested that they may be linked to acidic protein and may have a structural function, particularly on the nucleoli.

Animals↗
PubMed1986

Phospholipids in chromatin: incorporation of [32P]O4(2-) in different subcellular fractions of hepatocytes.

Nuclear, chromatin and microsomal fractions were isolated from hepatocytes prepared from rats injected with [32P]O4(2-) and killed subsequently at times between 1 and 48 h. Specific activities of the total phospholipids (PL) were determined for each subcellular fraction. The major points noted were the initial specific activity of the chromatin PL was half that of both nuclear and microsomal PL at 1 h; the first peak of labelling occurred at 6 h in both nuclear and microsomal PL, but was 3 h later (9h) in the chromatin PL; and a second peak of labelling occurred in the chromatin and microsomal PL, but not in those of the nuclei. On fractionation of the PL, the major and most metabolically active components were phosphatidylcholine + phosphatidylethanolamine, whilst sphingomyelin accounted for only about 8 per cent of the total PL. The chromatin and microsomal fractions were somewhat similar in their labelling patterns though with a delayed peaking of activity in the chromatin. This is indicative of a synthesis and transport of PL from the microsomes to the chromatin.

Animals↗
PubMed1985

Phospholipids in plant and animal chromatin.

Isolated hepatic nuclei and hepatic chromatin have been analysed for their DNA, RNA, protein and phospholipid content. The protein/DNA ratio is 3 for nuclei and 1.95 for chromatin extracted from Triton X-100 treated nuclei. The phospholipids, (2.36 +/- 0.91 (S.D.) per cent of the total nuclear material), are lost during the chromatin preparation mainly during the Triton X-100 washings of the nuclei. Nevertheless, 10 per cent of the total nuclear phospholipids remain bound to the chromatin. The comparative analysis of both nuclei and chromatin shows a difference in phospholipids and fatty acid composition. Thus, the chromatin-associated phospholipid cannot be attributed simply to contaminating nuclear membrane. This is supported by the autoradiographic study of semi-thin sections of interphase nuclei from root apices of Vicia faba in which [3H] ethanolamine is clearly localized in the chromatin and nucleolar regions of the nuclei.

Animals↗
PubMed1984

The use of (3H) thymidine-adsorbed on activated charcoal for the study of non-premitotic DNA synthesis in newborn rat hepatocytes.

DNA synthesis in newborn rat hepatocytes was studied in the first three days of life by means of repeated injections of (3H) thymidine. One group of animals was treated with the label adsorbed on activated charcoal (experimental group) and another group (controls) was given the label diluted in saline. The specific activity of DNA was higher in control group, but its increase was not linear with time; in the experimental group, the radioactivity was lower, but its increase with time was linear. The percentage of labeled nuclei was higher in the experimental animals than in the controls and increased linearly with time. The average number of grains/nucleus was considerably smaller in the experimental group than in the controls, in which also the percentage of labeled cells showed considerable variations during the first three days of life. It is concluded that activated charcoal adsorption increases label availability with time and, by keeping a lower label concentration in the pool, reduces the risk of radiation damage.

Animals↗
PubMed1983

Behaviour of tyrosine amino transferase and convertase during the first hours after hepatectomy in rats.

The activity of rat liver tyrosine amino transferase (TAT) increases after hepatectomy with a first prominent peak at 8 h and a second peak at 18 h. This change in activity is probably due to de novo enzyme synthesis since it is prevented by actinomycin-D (AMD). In the same period an increase of the lysosomal converting enzyme (convertase) which catalyses the in vitro transition of TAT from form I to form III, has been observed; this is not accompanied by changes of other lysosomal enzymes, such as acid phosphatase and cathepsin L. The activity of convertase is equal to that of the controls (sham operated animals) 2 h after hepatectomy, increases three times at 5 h, maintains the same value at 8 h and then decreases slowly to control level after 24 h. The correlation between the activity changes of the two enzymes strongly suggests a physiological role of convertase in TAT turnover.

Acid Phosphatase↗