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Biomedical subjects

R Bareggi

Publications and source records attributed to R Bareggi.

At least 55 records · Page 3Linked to original sources
PubMed1994

An immunohistochemical study of protein kinase C distribution in fetal mouse vertebral column.

Using polyclonal antibodies we have studied the distribution of protein kinase C in fetal mouse low thoracic vertebrae. By means of a pan protein kinase C antiserum recognizing the catalytic domain of the enzyme, we show that protein kinase C is markedly expressed in chondrocytes before birth. The enzyme seems to be very abundant in the more mature cells that are close to ossification centres as well as the periphery of the intervertebral disc, although it can also be detected in chondrocytes. In order to establish which protein kinase C isoenzyme(s) the chondrocytes produce, we employed polyclonal isoenzyme-specific antisera developed against three calcium-dependent isoforms (alpha, beta, gamma) and three calcium-independent isoforms (delta, epsilon, zeta). Secondary antibody conjugated to alkaline phosphatase revealed that chondrocytes markedly express the beta-isoform. Cells were also weakly stained by the anti-epsilon serum. The immunostaining was completely abolished by pre-incubating primary antibodies with the peptide antigens to which they were raised. These results suggest that protein kinase C (and particularly the beta isoform) could play an important role in mouse fetal chondrogenesis of the vertebral column.

Alkaline Phosphatase↗
PubMed1994

The growth of long bones in human embryological and fetal upper limbs and its relationship to other developmental patterns.

Measurements were made of the long bones of the upper limbs (humerus, ulna, radius) of 58 aborted embryos and fetuses, developmental age from 8 to 14 weeks, crown-rump length (CRL) between 38 and 116 mm. The specimens were cleared and double-stained, using alcian blue and alizarin red S for a differential detection of cartilage and bone. The values of both the total length (TL) and the ossified part (OL) of each long bone were related to the fetal developmental age previously estimated by freshly measured CRL. The relationship to another developmental pattern, i.e. the number of ossified centres in the vertebral column, suggested that the OL values could be much more significant than TL for the assessment of fetal growth.

Embryo, Mammalian↗
PubMed1994

On the presence of a secondary cartilage in the mental symphyseal region of human embryos and fetuses.

The presence of a secondary cartilage in the mental symphyseal region was examined in this study. A double-staining method with alcian blue and alizarin red S was performed on both whole human embryos and fetuses (developmental age between 8 and 17 weeks, crown-rump length, CRL, between 37 and 124 mm) and their disjointed mandibles. Histological and histochemical techniques were applied to transverse serial sections of whole disjointed fetal heads. The ossification process observed in the mental symphysis is quite different from that of the mandibular body, whose membranous ossification is induced by the contiguous Meckel's cartilage. No evidence of any fusion of Meckel's cartilage with the symphyseal cartilage, that lies within the symphyseal space, was detected. On the basis of these findings, we suggested that the mental secondary cartilage is able to change into bone according to an endochondral ossification process. Moreover, the role of mechanical causes in the development of the mental symphysis was hypothesized.

Cadaver↗
PubMed1994

A quantitative study on the spatial and temporal ossification patterns of vertebral centra and neural arches and their relationship to the fetal age.

A double-staining technique on 37 human embryos and fetuses (crown-rump length, CRL, between 38 and 116 mm) has been performed to study the ossification patterns of the vertebral column. Different growth sequences for centra and neural arches were observed. The survey of ossified centers suggested it was possible to relate significantly their appearance with the CRL. On the basis of already known data defining the developmental age in relationship to the latter parameter, we suggest their numerical evaluation as a further parameter for the assessment of the fetal age. Therefore, we have worked out a table that may be used either to determine the normal fetal growth, or when other parameters cannot be relied upon (i.e. in morphological diseases) for this aim.

Cervical Vertebrae↗
PubMed1993

The protein kinase inhibitor staurosporine induces morphological changes typical of apoptosis in MOLT-4 cells without concomitant DNA fragmentation.

The protein kinase inhibitor staurosporine induces ultrastructural changes typical of apoptotic cell death in MOLT-4 cells in a concentration range of 10-200 nM. The well known chromatin margination was indeed present, followed by characteristic nuclear protrusions. The formation of numerous homogeneously electron dense micronuclei was the final step of the process. Nevertheless we did not detect the distinctive internucleosomal DNA fragmentation which has been demonstrated to occur in a variety of cells exposed to agents causing apoptosis. Our results strengthen recent argument showing that DNA breakage cannot be considered the sole criterion for detection of apoptosis.

Alkaloids↗
PubMed1993

Subtraction of autofluorescent dead cells from the lymphocyte flow cytometric binding assay.

Flow cytometry allows the quantitative analysis of lymphocyte-target cell conjugates and the identification of the lymphocyte subset involved in the binding phenomenon. We recently described a methodology to identify the effector cells bound to K562 targets based on target cell autofluorescence coupled with lymphocyte staining by means of fluorescent monoclonal antibodies. Here we describe an implementation of the methodology that allows the subtraction of spontaneously dead targets to which lymphocytes may or may not adhere, thereby preventing the overestimation of the binding phenomenon and limiting its evaluation to living effector-target conjugates, thus preserving the specificity of the phenomenon.

Antibodies, Monoclonal↗
PubMed1993

Absence of high levels of DNA polymerase alpha activity in the nuclear matrix prepared from mouse erythroleukemia cells.

We have examined the association of DNA polymerase alpha activity with the nuclear matrix prepared by different techniques from mouse erythroleukemia cells. At variance with the data obtained using other cell types we have found that only a small amount (less than 2%) of nuclear DNA polymerase alpha activity resisted extraction with high-ionic strength buffers, even if nuclei were heat-stabilized by incubation at 37 degrees C for 45 min prior to subfractionation. The recovery of DNA polymerase alpha activity bound to the matrix was unaffected by the type of extracting agent used (NaCl or (NH4)2 SO4), by the extraction sequence or by the method employed for obtaining nuclei. These results could indicate that in some types of cells the nuclear matrix is not involved in DNA replication.

Animals↗
PubMed1993

Interferon rapidly induces modifications of chromatin sensitivity to DNase I in Daudi lymphoma cells.

The DNase I sensitivity of total chromatin has been analyzed in nuclei isolated from control and interferon-treated Daudi cells. The electrophoretic analysis of DNA has evidenced a different pattern of DNA fragment size produced by DNase I in nuclei isolated from control cells compared to interferon-treated samples. This feature is supported by a different recovery of acid soluble chromatin and is accompanied by modifications of in vitro RNA synthesis along with initiation and elongation of RNA chains. No changes have been evidenced in nuclei isolated from Daudi-resistant cells under the same experimental conditions. These data might be interpreted as a transient modulation, induced by interferon, of chromatin structure in terms of chromatin condensation which, in turn, activates the RNA synthesis after the transduction into the nucleus of the interferon-generated signals.

Burkitt Lymphoma↗
PubMed1993

Developmental pathways of vertebral centra and neural arches in human embryos and fetuses.

The ossification pathways of both vertebral centra (i.e., vertebral bodies) and neural arches were studied in human embryos and fetuses (CR-length between 38 and 116 mm). A clearing and double-staining method for whole embryo or fetus, using alcian blue and alizarin red S, allowed an easy and precise detection of the morphology of the whole vertebral column and every single vertebra. Both cartilaginous and bony components were clearly visible. Different temporal and topographical patterns of ossification were shown for the centra and arches; the latter were respectively proximal-distal (i.e., bidirectional from a defined starting tract in T10-L1) and cranial-caudal (i.e., monodirectional). The patterns could be related to the morphogenetic processes of other structures (i.e., muscles and nerves). Moreover, the numerical survey of ossification centers provided a possible parameter for the determination of the fetal developmental age. This could be useful in the study of pathological conditions.

Alcian Blue↗
PubMed1993

Evidence for nuclear phosphoinositidase C activity in the antiproliferative signals produced by interferon in Burkitt lymphoma cells.

The influence of interferon alpha on nuclear phosphoinositidase C (PIC) in Daudi cells has been analysed. Results showed an early increase of PIC activity detectable within 90 min of interferon treatment concomitant with an increase of diacylglcerol (DAG) levels. Since the interferon-induced DAG production is not modified by the addition of propranolol, a compound known to inhibit production of DAG from phosphatidylcholine hydrolysis, it is suggested that the interferon antiproliferative signal is transduced into the nucleus via the inositol lipid pathway. A parallel analysis performed on intact cells showed a rapid inhibition of PIC activity accompanied by an increase of DAG level thus suggesting that interferon-generated signals at plasma-membrane level use pathways different from that of inositol lipids. A selected clone of Daudi cells resistant to interferon action provided a control for specificity of results.

Burkitt Lymphoma↗
PubMed1993

Decrease in nuclear phospholipids associated with DNA replication.

Lipid metabolism in nuclei is very active and appears involved in the transduction of signals to the genome in response to agonists acting at the plasma membrane level. However, the precise topology of nuclear lipid metabolism and the relationship between nuclear lipids and crucial events of the cell function, such as DNA replication, have not been fully elucidated. By using a recently developed cytochemical method for detecting phospholipids inside the nucleus of intact cells at the electron microscope level, we have analyzed the changes in intranuclear phospholipids in DNA-replicating versus resting cells, which are both present in the same sample of regenerating liver after partial hepatectomy. The pattern of DNA synthesis in replicating cells has been monitored by electron microscope immunocytochemistry after bromodeoxyuridine (BrdU) labeling. The data obtained, which allow a fine localization and a quantitative analysis of both DNA synthesis and phospholipid distribution, indicate a significant reduction in the phospholipids detectable inside the nucleus in all steps of the S phase. This could depend on an increased nuclear phospholipid hydrolysis, whose products should in turn activate some of the enzymes involved in the control of DNA replication.

Animals↗
PubMed1992

Interferon-mediated intracellular signalling. Modulation of different phospholipase activities in Burkitt lymphoma cells.

The effect of interferon-alpha on Daudi lymphoma cells either sensitive or resistant to the action of this cytokine has been analysed in terms of phospholipase C (PLC) and D (PLD) activities. Results have shown a combined modulation of PIP2-specific phospholipase C and phospholipase D. In particular, a decreased activity of PIP2-specific PLC has been found, concomitant to a PLD-mediated phosphatidylcholine hydrolysis, suggesting that the intracellular signalling activated by interferon in Daudi cells involves a phospholipase D/phosphohydrolase pathway.

Burkitt Lymphoma↗
PubMed1992

The effect of in vitro heat exposure on the recovery of nuclear matrix-bound DNA polymerase alpha activity during the different phases of the cell cycle in synchronized HeLa S3 cells.

HeLa S3 cells were synchronized by a double thymidine block or aphidicolin treatment and the levels of nuclear matrix-bound DNA polymerase alpha activity were then measured using activated calf thymus DNA as template. The nuclear matrix was obtained by 2 M NaCl extraction and DNase I digestion of isolated nuclei incubated at 37 degrees C for 45 min prior to subfractionation. In all phases of the cell cycle 25-30% of nuclear DNA polymerase alpha activity remained matrix-bound, even when cells were in the G1 phase. No dynamic association of DNA polymerase alpha activity with the matrix was seen, at variance with previous results obtained in regenerating rat liver. The variations measured in matrix-bound activity closely followed those detected in isolated nuclei throughout the cell cycle. If nuclei were not heat-stabilized very low levels of DNA polymerase alpha activity were measured in the matrix (1-2% of total nuclear activity). Heat incubation of nuclei failed to produce any enrichment in matrix-associated newly replicated DNA, whereas the sulfhydryl cross-linking chemical sodium tetrathionate did. Therefore the results obtained after the heat stabilization procedure do not completely fit with the model that envisions the nuclear matrix as the active site where eucaryotic DNA replication takes place.

Aphidicolin↗
PubMed1992

Changes in polyphosphoinositide levels in rat liver nuclei in response to prolactin, a known hepatic mitogen.

The effect of prolactin action on nuclear polyphosphoinositide synthesis was investigated in isolated rat liver nuclei. An increased uptake of phosphate from [gamma 32P] adenosinetriphosphate was observed in both phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate with a maximum response at 10(-12) M concentration of hormone. Pulse-chase experiments in isolated nuclei following prolactin treatment indicate that the observed increase in accumulation of phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate is mainly due to a decrease in their rate of turnover possibly induced by a change in activity of polyphosphoinositide-specific monoesterases. In vitro prolactin also reduces the activity of nuclear phospholipase C specific for phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate. Moreover, this feature is strongly supported by the concomitant decrease in nuclear diacylglycerol mass. Thus these data suggest that once prolactin reaches the nucleus an intranuclear signalling is evoked through inositol lipid metabolism.

Animals↗
PubMed1987

Histochemical study on catecholamines in human coronary endothelium.

Specific catecholaminergic granules had been previously described in the endothelial cells of blood and lymphatic vessels. 2 histochemical techniques were used in this work for detecting catecholamines in human coronary vessels: both the postfixation with OsO4-KI mixture and the formaldehyde induced fluorescence (FIF) reaction. Ultrastructural examinations of bioptic specimens processed with the OsO4-KI staining showed a marked positivity in the coronary endothelial cells, as well as in the smooth muscle fiber cells of the coronary arteriolae and the adventitial nerve endings. These findings were confirmed by a high level of fluorescence in the same structures, obtained using the FIF reaction. Myocardial fiber cells never reacted. Therefore, an important role of the endothelium of human coronary vessels in the turn-over of catecholamines could have been supposed.

Biopsy↗
PubMed1986

Localization of an aminoglycoside (streptomycin) in the inner ear after its systemic administration. A histochemical study using fluorescence microscopy.

We used the simple method of direct cytofluorescence to detect the presence of the aminoglycoside, streptomycin, in the inner ear after its systemic administration. In the cochlea, fluorescence was observed in the organ of Corti, the spiral ganglion, the nerve fibres, the vascular stria and Reissner's membrane; in the vestibulum, fluorescence was seen in the crista ampullaris and the planum semilunatum. The localization of the drug was related to the distribution of its specific receptor, triphosphoinositide (TPI); therefore, it is reasonable to assume that aminoglycosides exert their toxic effects by binding to TPI.

Animals↗
PubMed1979

Lipids in the Harder's gland of certain rodents. I: Neutral lipids.

The major components of the lipid fraction from Harder's glands of mice and rats were studied, including examination of the influence of age and sex on absolute and relative amounts of the most relevant components. The data, based on thin layer chromatographic analyses, is discussed in detail and possible explanations suggested for the biological roles which the different components may play.

Aging↗