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Biomedical subjects

A Zanini

Publications and source records attributed to A Zanini.

At least 19 recordsLinked to original sources
PubMed1995

Immunolocalization of secretogranin II and insulin in a nerve growth factor-differentiated insulinoma cell line.

RINm5F, a rat insulin-secreting pancreatic cell line, responds to nerve growth factor (NGF) by extending neurite-like processes. Secretogranin II (SgII), a marker of neuroendocrine secretory organelles, has recently been found to be a good marker of neuronal differentiation in both human neuroblastoma and rat pheochromocytoma cells. The present paper reports the results obtained from immunocytochemical studies, which show that NGF increases the expression of SgII-immunolabeled organelles in RINm5F cells. We also demonstrate that NGF increases the expression of insulin and that SgII and insulin are predominantly, but not always, colocalized. These results suggest that this insulinoma cell line may be a good model for studying the role of SgII in neuroendocrine secretion mechanisms.

Animals↗
PubMed1994

Pre-operative cervical preparation before first trimester missed abortion: a randomized controlled comparison between single or double intracervical administration of PGE2 gel.

The aim of this study was to evaluate whether a single intracervical application of prostaglandin E2 (PGE2) gel is as effective as a repeated administration with respect to the % of curettage for a missed abortion and to incidence of side effects. 32 consecutive patients with ultrasonographic diagnosis of missed abortion from 6th-13th week of gestation were randomly allocated to either single (group A) or repeated, 2 h apart (group B), intracervical application of PGE2 gel. No differences were observed in cervical dilatation before the administration of the gel between the two groups. In group B, evaluation of cervical dilatation 2 h after the first administration of PGE2 gel and before the second one did not show significant changes as compared to baseline values. The degree of cervical dilatation before surgery was significantly improved as compared to the initial dilatation in both groups; no significant difference was observed between the two study groups. A single administration of PGE2 gel followed by surgery 5 h later has the same effectiveness on cervical dilatation but fewer side effects than repeated administration of the drug 2 h apart.

Abortion, Missed↗
PubMed1993

Induction of labor with intravaginal prostaglandin E2 gel.

OBJECTIVE: The aim of our study was to evaluate the optimum dose of intravaginal prostaglandin E2 gel for induction for labor in nulliparous women with a relatively ripe cervix (modified Bishop score 4 or 5). METHOD: One hundred and sixty-seven nulliparous women at term with indications for the induction of labor were treated randomly with two doses of intravaginal 2.0 (group A) or 3.0 mg (group B) of prostaglandin PGE2 gel every 12 h. Data were analyzed by chi 2-test and Student's t-test. RESULTS: Of 87 patients 64 went into labor after gel application in group A, compared with 68/80 in group B (73.5% vs. 85.0%) (P = NS). A second gel administration was needed for 9 women in group A and 6 women in group B. More side effects (both local and systemic) were noted in group B than in group A (28.7% vs. 14.9%) (P = 0.03). In particular, more local (hyperstimulation or hypertonus) side effects were noted in group B (13.7% vs. 2.3%) (P = 0.01). CONCLUSION: The vaginal administration of 2.0 mg of PGE2 gel seems to be equally effective as 3.0 mg in terms of labor success rate with a significant lower incidence of side effects.

Administration, Intravaginal↗
PubMed1992

Immunodetection of secretogranin II in animal and human tissues by new monoclonal antibodies.

Secretogranin II (chromogranin C) is an acidic tyrosine-sulfated secretory protein, known to be a marker of neuroendocrine secretory products and of specific neuroendocrine tumours. In order to obtain anti-secretogranin II monoclonal antibodies for cell biology studies and, in particular, for clinical applications, we immunized mice with a secretogranin II-enriched fraction prepared from homogenates of bovine anterior pituitaries. Hybridoma supernatants obtained from the splenocytes of a hyperimmune mouse, screened with an enzyme-linked immunosorbent assay, were analyzed by both immunocytochemistry and two-dimensional immunoblotting. By using this experimental approach, we were able to identify two monoclonal antibodies (8G1 and 5A7) which recognize bovine secretogranin II. Both immunocytochemistry and immunoblotting revealed that one of them, the 5A7 antibody, cross-reacts with the human antigen. The distribution patterns of the immunoreactivity, obtained by immunocytochemistry with the 5A7 antibody in animal and human tissues, partially overlap those, obtained by using a polyclonal antiserum elicited against bovine secretogranin II, previously described. Moreover, the 5A7, but not the polyclonal antibody, reacts with some duodeno-jejunal cells. In conclusion, both the 5A7 and 8G1 antibodies can be useful for cell biology studies. The 5A7 antibody can be used for the detection of secretogranin II in human tissues and should be of help in clinical and pathological practices.

Animals↗
PubMed1992

Cervical priming and labor induction by multiple doses of intracervical prostaglandin E2 gel.

One hundred seventy-two term pregnant women with medical or obstetric conditions requiring induction of labor were treated with intracervical administration of 0.5 mg prostaglandin E2 in tylose gel. Multiple administrations were necessary in 42 cases (24.4%), two administrations in 31 cases (18.0%) and three administrations in 11 cases (6.4%). Intracervical administration of PGE2 tylose gel (0.5 mg dose) is useful to prime the cervix, induce labor, and significantly modify Bishop score.

Cervix Uteri↗
PubMed1992

Immunolocalization of secretogranin II, chromogranin A, and chromogranin B in differentiating human neuroblastoma cells.

In order to obtain further insights into the expression of the known markers of secretory neuroendocrine dense core organelles, secretogranin II (SgII), chromogranin A (CgA), and chromogranin B (CgB) during neuronal differentiation, the immunolocalization of these proteins was studied by means of double immunofluorescence in both undifferentiated and retinoic acid-differentiated SH-SY5Y human neuroblastoma cells. The majority of undifferentiated cells was not immunolabeled for all three proteins. In the majority of differentiated cells, a clearly punctate SgII immunolabeling indicative of the presence of secretory organelles was present in the Golgi region, at the cell periphery, along the neurites and in growth cones. Only relatively few of the SgII-immunolabeled cells were also immunolabeled for CgA and CgB, and in a single cell the three proteins were not always present in the same organelles. These results, obtained in a cultured cell line, confirm the not necessarily parallel distribution of SgII, CgA, and CgB observed in different neuroendocrine tissues and suggest that SgII may be the best marker of human neuroblastoma cell differentiation.

Biomarkers, Tumor↗
PubMed1991

Small GTP-binding proteins in human neuroblastoma cell lines.

The presence of small G proteins was investigated by [gamma-35S]GTP-binding in 3 human neuroblastoma cell lines. IMR-32, SK-N-BE and SH-SY5Y, before and after treatment with differentiating agents (dibutyryl-cAMP, 5-bromodeoxyuridine or retinoic acid) which induce the appearance of secretory organelles. One major component of about 24 kDa and 3 minor components of smaller Mr were found to bind specifically [gamma-35S]GTP in all 3 cell lines already before differentiation. Differentiation did not affect the expression of small G proteins in IMR-32 cells and only modestly affected it in the other two cell lines. The possibility that the expression of small G proteins in neuroblastoma cells is not coupled with the assembly of secretory organelles is discussed.

Cell Differentiation↗
PubMed1990

Co-localization of secretogranins/chromogranins with thyrotropin and luteinizing hormone in secretory granules of cow anterior pituitary.

We investigated the co-localization in secretory granules of secretogranins/chromogranins, thyrotropin, and luteinizing hormone in ultra-thin frozen sections of cow anterior pituitary by double immunoelectron microscopy, using specific antibodies and protein A-gold particles of different sizes. The distribution of secretogranin II, chromogranin A, and chromogranin B (secretogranin I) was largely similar. In cells containing secretory granules of relatively small size (100-300 nm) and low electron density (identified as thyrotrophs and gonadotrophs by immunolabeling for the respective hormone) and in cells containing both small (170-250 nm) and large (300-500 nm) secretory granules of low electron density (also identified as gonadotrophs), all three secretogranins/chromogranins were detected in most if not all granules, being co-localized with the hormone. In cells containing both relatively large (400-550 nm), electron-dense granules and small, less electron-dense secretory granules (150-300 nm), identified as somatomammotrophs by double immunolabeling for growth hormone and prolactin, all three secretogranins/chromogranins were predominantly detected in the subpopulation of small, less electron-dense granules containing neither growth hormone nor prolactin. Interestingly, this granule subpopulation of somatomammotrophs was also immunoreactive for thyrotropin and luteinizing hormone. These data show that somatomammotrophs of cow anterior pituitary are highly multihormonal, in that the same cell can produce and store in secretory granules up to four different hormones and, in addition, the three secretogranins/chromogranins. Moreover, selective localization of the secretogranins/chromogranins together with thyrotropin and luteinizing hormone in a subpopulation of secretory granules of somatomammotrophs indicates the preferential co-packaging of the secretogranins/chromogranins and these hormones during secretory granule formation.

Animals↗
PubMed1990

Deprivation of growth hormone-releasing hormone early in the rat's neonatal life permanently affects somatotropic function.

This work investigated in rats whether passive immunization against the endogenous GHRF in the early postnatal period led to permanent alterations of somatotropic function, similar to those observed in several human growth disorders, e.g. constitutional growth delay (CGD). On postnatal days 1, 2, 4, 6, 8, and 10, rats were given an anti-GHRF-serum (GHRH-Ab, 100 microliters/rat, sc) and were tested 1, 30, and 60 days after this treatment for basal and GHRH-stimulated GH secretion both in vivo and in vitro. GHRH-Ab reduced both basal and GHRF-stimulated GH secretion at all intervals and induced marked and chronic impairment of growth rate. The following differences were observed in the GHRH-Ab treated rats compared to normal rabbit serum-treated controls: 1) GH biosynthesis (incorporation of L-[3H]leucine into the electrophoretic band of GH): reduction of about 70%, 1 day but not 30 days after treatment; 2) Pituitary weight: significant reduction in absolute weight (30-40%) at all posttreatment intervals, and relative weight, 1 and 30 days after treatment. 3) Pituitary GH concentration: significant reduction in GH content (about 40%) but not concentration, at all posttreatment intervals; 4) Percentage of somatotrophs (immunocytochemistry): about 40% reduction 1 day, but not 30 and 60 days after treatment; 5) Hypothalamic somatostatin messenger RNA (mRNA) levels in situ hybridization): selective reduction (40%) in the periventricular nucleus 1 day but not 30 days after treatment; 6) Hypothalamic somatostatin cell number (immunocytochemistry): no significant changes in any hypothalamic area at any interval; 7) Pituitary somatostatin binding (in situ autoradiography): significant reduction, 1 day and 30 days after treatment; 8) Somatostatin inhibition of GH release "in vitro": somatostatin effect on GH release was reduced 30 days after treatment. These and previous data indicate that: 1) Transient deprivation of GHRF in the immediate postnatal period of the rat leads to permanent impairment of growth rate and somatotropic function; 2) GHRF deficiency itself or through reduction of GH secretion impairs somatostatin functions temporarily in the hypothalamus and permanently in the pituitary; 3) This rat model may mimic some forms of growth disorders in humans and holds promise as useful tools for investigating the underlying pathophysiological mechanisms.

Animals↗
PubMed1990

Pre-induction cervical ripening with prostaglandin E2 gel: intracervical versus intravaginal route.

The aim of our study was to evaluate the best method for cervical ripening before a classical induction with amniotomy and oxytocin. One hundred term pregnant patients who presented an unfavorable cervix and an indication for the induction of labor were assigned randomly to either 0.5 mg prostaglandin (PG) E2 gel intracervically (N = 52) or 3 mg PGE2 gel intravaginally (N = 48). The intravaginal gel had a greater effect on cervical ripening according to a modification of the Bishop score than did intracervical gel, but it had a higher incidence of side effects.

Administration, Intravaginal↗
PubMed1989

Immunohistochemical localization of secretogranin II in the rat cerebellum.

Secretogranin II (chromogranin C) is a peptide related to chromogranin A and secretogranin I (chromogranin B) which is secreted by a regulated pathway from both neurons and endocrine cells. In the present study we have determined by light microscopic immunocytochemistry its distribution in the cerebellum and in adjacent brain stem regions. Secretogranin II was found to be widely distributed throughout the gray matter of these regions. Highly immunoreactive structures in the cerebellar cortex included the majority of climbing fibers, a large number of mossy fibers, sparse varicose fibers in the molecular layer and a subpopulation of neuronal perikarya in the granule cell layer. The location and shape of these neurons are very similar to those of a novel type of cerebellar neurons which has been recently described. A moderate level of immunoreactivity was observed on fibers travelling among Purkinje cells and parallel to the pial surface in the Purkinje cell layer. A variable, but in general low, degree of immunoreactivity was also detectable in the perikarya of Purkinje cells. In the deep cerebellar nuclei a loose network of secretogranin II-positive fibers was visible. Neurons of the nuclei, however, were non-immunoreactive. A dense network of highly immunoreactive fibers was found throughout the brain stem regions adjacent to the cerebellum. Our results indicate that secretogranin II has in the cerebellum and adjacent regions a distribution more widespread than that of known regulatory peptides and suggest that the peptide-mediated signaling in the cerebellum plays a role more important that has been acknowledged so far.

Animals↗
PubMed1989

Preparation and characterization of anti-human chromogranin A and chromogranin B (secretogranin I) monoclonal antibodies.

Chromogranin A, chromogranin B/secretogranin I and chromogranin C/secretogranin II are acidic sulphated and phosphorylated secretory proteins present in a large number of endocrine and neuronal tissues. It has been suggested that these proteins may be useful immunohistochemical markers for human tumours of endocrine origin and their measurement in plasma has been proposed as a diagnostic tool in patients with these tumours. In order to obtain anti-human chromogranins/secretogranins antibodies for clinical applications, we immunized mice with whole chromaffin granules isolated from human pheochromocytoma. The immune sera analysed by two-dimensional immunoblotting were found to recognize chromogranins/secretogranins and other unidentified proteins and to react in immunocytochemistry with pheochromocytoma as well as with a number of endocrine cells of different types. Hybridoma supernatants obtained from the splenocytes of a hyperimmune mouse, screened with an enzyme-linked immunosorbent assay, were analysed by both immunocytochemistry and two-dimensional immunoblotting. By using this experimental approach we were able to identify several monoclonal antibodies against human chromaffin granule components. In particular, we have characterized one anti-human chromogranin A and one anti-human chromogranin B/secretogranin I monoclonal antibody which showed a very specific pattern both in immunocytochemistry and in two-dimensional immunoblotting.

Antibodies, Monoclonal↗
PubMed1989

Human neuroblastoma cells acquire regulated secretory properties and different sensitivity to Ca2+ and alpha-latrotoxin after exposure to differentiating agents.

IMR-32 human neuroblastoma cells are unable to release [3H]dopamine in response to secretagogues. However, they express a normal complement of membrane receptors and ion channels which are efficiently coupled to second messenger production. In the present study we took advantage of the ability of this cell line to differentiate in vitro in the presence of either dibutyrryl-cAMP or 5-bromodeoxyuridine, to analyze any developmentally regulated changes in its secretory properties. Uptake, storage, and release of [3H]dopamine were studied biochemically and by autoradiography. The calcium ionophore ionomycin, phorbol 12-myristate 13-acetate and the presynaptic acting neurotoxin alpha-latrotoxin were used in both control and differentiated cells as secretagogue agents. The presence of secretory organelles was investigated by electron microscopy; the expression of secretory organelle markers, such as chromogranin/secretogranin proteins (secretory proteins) and synaptophysin (membrane protein), was detected by Western blotting and immunofluorescence. The results obtained indicate that IMR-32 cells acquire regulated secretory properties after in vitro drug-induced differentiation: (a) they assemble "de novo" secretory organelles, as revealed by electron microscopy and detection of secretory organelle markers, and (b) they are able to store [3H]dopamine and to release the neurotransmitter in response to secretagogue stimuli. Furthermore, secretagogue sensitivity was found to be different, depending on the differentiating agent. In fact, dibutyrryl-cAMP treated cells release [3H]dopamine in response to alpha-latrotoxin, but not in response to ionomycin, whereas 5-bromodeoxyuridine treated cells release the neurotransmitter in response to both secretagogues. All together these results suggest that IMR-32 cells represent an adequate model for studying the development of the secretory apparatus in cultured human neurons.

Biological Transport↗
PubMed1989

Immunoreactivity of hormonally-characterized human endocrine cells against three novel anti-human chromogranin B(B11 and B13) and chromogranin A (A11) monoclonal antibodies.

Two novel monoclonal antibodies, called B11 and B13, directed exclusively against human chromogranin B (CgB) and another antibody, A11, specific for human chromogranin A (CgA), were obtained by immunization of mice with chromaffin granules, the fusion of their splenocytes, the screening of hybridomas supernatants by ELISA and immunohistochemistry, and characterization of the antibodies by two-dimensional immunoblotting. The antibodies were used in immunohistochemical tests to investigate the distribution of CgA and CgB in hormonally-identified cells of the human endocrine system. The A11 antibody confirmed the occurrence of CgA in gut EC, ECL, gastrin, secretin and neurotensin cells, pancreatic A and PP cells, parathyroid chief cells, pituitary TSH and gonadotrope cells and adrenal medullary cells. Only a fraction of CgA-immunoreactive cells in the human gut and pancreas showed C-terminus arginine-glycinamide immunoreactivity, suggesting pancreastatin storage. Both CgB antibodies showed immunoreactivity in gastrin cells, intestinal (but not gastric) EC cells, pancreatic A and PP cells, pituitary TSH and gonadotrope cells and adrenal medullary cells. In addition, the B11 antibody stained thyroid C cells and the B13 antibody stained the Golgi area of pituitary GH cells. It is concluded that most CgB is stored in the same cells showing CgA, although some CgA-rich cells, like gastric EC and ECL cells. lacked B11 and B13 immunoreactivities and some CgA-poor cells, like human thyroid C cells, showed intense B11 immunostaining.

Antibodies, Monoclonal↗
PubMed1988

The gonadotrope polypeptide (GP 87) released from pituitary cells under luteinizing hormone-releasing hormone stimulation is a secretogranin II form.

Cultured gonadotrope cells from 14 day old female rat pituitaries have been shown to release a highly acidic protein when incubated with LHRH: the gonadotrope polypeptide (GP 87). Moreover, a new tyrosine-sulfated acidic protein, secretogranin II (Sg II), clearly distinct from the chromogranin species, was described in the secretory granule matrix of endocrine cells secreting peptide hormones by the regulated pathway. Recently, the release of Sg II from female rat pituitary stimulated by LHRH was demonstrated in vitro. Several physicochemical (Mr; pI) and biological (cellular localization in the pituitary; LHRH-stimulated release) properties are common to Sg II and GP 87. To verify if these 2 polypeptides are similar or distinct components, other physicochemical characteristics (heat-stability, sulfation, phosphorylation) were compared using isotope incorporation followed by either 1- or 2-dimensional polyacrylamide gel electrophoresis and autoradiography. Furthermore, the similarity of GP 87 to Sg II was studied by immunoblotting on nitrocellulose sheets following electrophoresis of intracellular and secreted proteins. Antisera raised against bovine Sg II (extracted from whole pituitaries) and against rat GP 87 (released into the medium of cultured pituitary cells stimulated by LHRH) were used. The overall data presented here suggest that GP 87 is the Sg II form contained in, and released by, gonadotrope cells.

Animals↗