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Biomedical subjects

F Orlandi

Publications and source records attributed to F Orlandi.

At least 19 recordsLinked to original sources

Quantitative analysis of proliferating sinusoidal cells in dimethylnitrosamine-induced cirrhosis. An immunohistochemical study.

Proliferating lipocytes (fat-storing cells or perisinusoidal stellate cells of the liver) were detected by in vivo incorporation of bromodeoxyuridine (BrdU) in an experimental model of cirrhosis in the rat by dimethylnitrosamine. Lipocytes were identified by sequential double immunohistochemical staining on frozen sections using anti-desmin antibodies as a marker of cytoplasmic intermediate filaments followed by anti-BrdU antibodies to identify S-phase nuclei in animals treated for 7, 14 or 21 days. The number of desmin-positive (lipocytes) and desmin-negative (Kupffer and endothelial cells) sinusoidal cells incorporating BrdU was recorded. The labelling index of lipocytes was calculated as the percentage of BrdU-labelled desmin-positive cells with respect to total number of lipocytes. In control animals, when the total number of lipocytes was 153.9 +/- 11/mm2 (mean +/- 1 S.E.) the number of desmin-positive S-phase sinusoidal cells never exceeded 6.8 +/- 1.2/mm2 with a maximum labelling index of 4.3 +/- 0.5%. At 7 days of treatment, the values were respectively 236 +/- 26.5/mm2, 53.2 +/- 5.9/mm2 and 22.6 +/- 0.5% (p less than 0.001 vs. controls), while, at 21 days they were 272.5 +/- 21.2/mm2, 23.3 +/- 4.0/mm2 and 8.5 +/- 1.1% respectively (p less than 0.01). These results show that hyperplasia of lipocytes represents an early reaction to dimethylnitrosamine-induced liver injury. The local accumulation of lipocytes appears to occur in areas where fibrous septa develop later on.

Animals

Steatosis associated with immotile cilia syndrome: an unrecognized relationship?

The present study deals with a case of hepatic parenchymal steatosis in a child with primary ciliary dyskinesia (immotile cilia syndrome) well documented by functional and ultrastructural evaluation of the ciliary epithelia. Hepatic steatosis was associated with ultrastructural evidence of retention of material either in the cisternae of the endoplasmic reticulum or in proximity of the Golgi apparatus of hepatocytes. It is suggested that the absence of dynein in the axoneme is probably part of a diffuse genetic defect which may extend to cytoplasmic, non axonemal, dynein and lead to a disturbance of various microtubule-dependent cell activities.

Biopsy, Needle

Categorization of cysts and steroid levels in breast cyst fluid.

Several reports indicate that patients with macrocysts have a two- to fourfold higher risk of developing cancer. The fluid filling the cysts (breast cyst fluid, BCF) contains unusual amounts of biologically active substances, including hormones and metabolites. The accumulation of steroid conjugates, such as androgen and estrogen sulfates, deserves interest. Measuring BCF cations (K+, Na+) permits classification of cysts into two major subsets (type I and type II), conceivably associated with a different degree and with a turnover of apocrine cells in the lining epithelium. Type I (high K+/Na+ ratio) and type II (low K+/Na+ ratio) cysts display different patterns of steroid analytes and steroid-binding proteins. There are many gaps in our understanding of the relationship between local steroids and hypersecretion of fluid in the terminal duct lobular units with eventual appearance of cysts. Accumulating biochemical and epidemiological data, however, point to recurrent, multiple type I cysts as a marker of endocrine risk, i.e., of a whole-organ promoting status toward proliferative premalignant lesions.

Breast Neoplasms

Prenatal diagnosis of haemoglobin disorders by cordocentesis at 12 weeks' gestation.

Prenatal diagnosis of haemoglobin disorders is accepted to be a useful procedure to avoid births of infants with homozygous diseases. Advances in sampling and molecular techniques, such as polymerase chain reaction (PCR) and chorionic villus sampling (CVS), have made earlier and safer first-trimester prenatal diagnosis possible. However, these procedures need previous studies of at-risk couples, which can be very time-consuming when a number of different beta-thalassaemia mutations occur in the region. We describe the possibility of making a first-trimester prenatal diagnosis by cordocentesis and fetal blood analysis at the 12th week of gestation. We found no statistically significant difference (p greater than 0.05) between beta/gamma values in fetuses at the 12th and 18th weeks of gestation. In seven affected fetuses aborted at the 12th week of gestation, the diagnosis was confirmed in all cases by PCR analysis. These findings suggest that early cordocentesis could be an alternative procedure to CVS and PCR analysis.

Female

Enumeration of S-phase cells in normal rat liver by immunohistochemistry using bromodeoxyuridine-antibromodeoxyuridine system.

The visualization of incorporation sites of the thymidine analog bromodeoxyuridine (BrdU) into DNA, detected by immunocytochemistry, has been proposed as an index of the percentage of S-phase cells in a variety of tissues and as an easy, less expensive alternative to autoradiography. This technique has not yet been applied to the study of physiological cell renewal in the normal liver. In the present study, results obtained with this method in the liver of normal young adult rats is reported. BrdU was administered in vivo and subsequent incorporation was detected by the PAP technique using monoclonal anti-BrdU antibodies. The nuclei exhibiting a positive reaction within the liver were few and accounted for about 0.45% of all hepatocytes. Positive cells were located preferentially in zone 1, which contained 82.7% of the labeled cells. Zone 2 contained 15.4%, while only 1.9% of the labeled cells were found in zone 3. Positive-staining Kupffer cell nuclei were rare (about 0.5% of all Kupffer cells) and were distributed randomly in the hepatic lobule. These findings provide quantitative data about hepatocyte renewal in the normal liver in the absence of a growth stimulus. The simplicity and the reproducibility of this technique suggests that further application of this method in situations assessing hepatic regeneration are indicated.

Animals

Hyperthyroidism due to a pituitary adenoma composed of two different cell types, one secreting alpha-subunit alone and another cosecreting alpha-subunit and thyrotropin.

A 37-yr-old female presented with clinical signs and symptoms of mild hyperthyroidism, high serum levels of free T4 (24.2 pmol/L), free T3 (11.7 pmol/L), and sex hormone-binding globulin (157 nmol/L) as well as measurable (by immunofluorometric assay) serum TSH concentrations (1.9 mU/L) in the absence of any known methodological interference. The above finding indicated the presence of hyperthyroidism due to inappropriate secretion of TSH, whose neoplastic origin was documented by computed tomographic scan showing a 1-cm pituitary adenoma. The diagnosis was confirmed by elevated alpha-subunit levels (9.2 micrograms/L) and alpha-subunit/TSH molar ratio (25.2) as well as absent TSH suppression after T3 administration. TRH injection (200 microgram, iv) caused impaired TSH (from 3.0 to 4.8 mU/L) and unexpectedly exaggerated alpha-subunit (from 8.8 to 18.2 micrograms/L) responses. Such a discrepancy was also observed after other dynamic tests. Double gold particle immunostaining of the adenomatous tissue removed at surgery showed that all of the cells contained secretory granules positive for alpha-subunit, while very few cells were positive for TSH beta and alpha-subunit. In conclusion, the present study demonstrates the existence of TSH-induced hyperthyroidism due to a pituitary adenoma composed of two different cell types: one secreting alpha-subunit alone and another cosecreting alpha-subunit and TSH.

Adenoma

Phenotypic analysis of inflammatory infiltrate in rats with dimethylnitrosamine-induced cirrhosis.

The present study is concerned with the immunohistochemical characterization in situ of the mononuclear infiltrate accompanying the formation of septa and the development of cirrhosis in the liver of rats treated with dimethylnitrosamine (DMN), i.p. (10 microliters/kg, 3 days a week for 3 weeks). Monoclonal antibodies against macrophages, pan T cells, T cell subsets and B cells have been applied on cryostat sections of animals given DMN for 7, 14 and 21 days. The maximum increase of macrophages and lymphocytes was observed at days 7 and 14 respectively. At all times T lymphocytes appeared as the major component of the inflammatory infiltrate with a largely predominant population of cytotoxic/suppressor T cells. At day 21, with evidence of nodulation of parenchyma, macrophages levelled off while T cells remained numerous without changes in the inducer-helper T cells/cytotoxic-suppressor T cells ratio which remained always less than 1. B cells were always few. These findings illustrate the early influx of lymphocytes in DMN-induced liver injury and help to define the lymphocyte subsets associated with inflammation and fibrosis in a reproducible animal model.

Animals

The risks of early cordocentesis (12-21 weeks): analysis of 500 procedures.

Five hundred cordocenteses were performed between 12 and 21 weeks. The indications were thalassaemia (386), rapid karyotyping (97), feto-maternal allo-immunization (10), rubella (6), and toxoplasmosis (1). One hundred and ten pregnancies underwent termination on the basis of the result, while 20 of the 370 pregnancies intended to continue were lost to follow-up. Amongst these were 16 fetal losses (4.3 per cent) and 22 premature deliveries (5.9 per cent); no other complications were reported. Four adverse prognostic factors were identified: (a) cord bleeding; (b) fetal bradycardia; (c) prolonged procedure time; and (d) anterior insertion of the placenta. There was no 'obvious' difference in fetal loss rate with advancing gestation until 19-21 weeks, when the risk of fetal loss decreased to 2.5 per cent.

Blood Specimen Collection

Evidence that plasma membrane fluidity of isolated hepatocytes is modified by exposure to microtubule-depolymerizing drugs.

The role of microtubules on membrane fluidity has been investigated on freshly isolated whole rat hepatocytes prepared by the perfusion method and exposed either to the microtubule-depolymerizing drugs colchicine and vincristine or to beta-lumicolchicine, a colchicine analog deprived of biological activity. Exposure of hepatocytes to 6.3 microM colchicine or to 3.0 microM vincristine led to a significant decrease of membrane fluidity as measured by fluorescence polarization of trimethylammoniodiphenylhexatriene (TMA-DPH). No changes were observed in cells exposed to 10.0 microM beta-lumicolchicine. These observations support the hypothesis that the microtubular system plays a role in the modulations of physico-chemical properties of the plasma membrane.

Animals

Modulation of extracellular matrix components during dimethylnitrosamine-induced cirrhosis.

Liver fibrosis was induced in rats after administration of dimethylnitrosamine (DMN) intraperitoneally three times a week for 3 weeks. Incomplete septa appeared after 7 days and evidence of nodulation of the parenchyma was observed after 21 days. Both distribution of extracellular matrix components (collagen type I, type III and type IV, laminin, fibronectin, heparan sulphate proteoglycan) and the distribution of desmin as a marker of lipocytes (Ito cells) and of iso-alpha-smooth muscle actin were studied with immunoperoxidase. Changes in the distribution of extracellular matrix components outlined both the formation of septa and the development of nodules with changes in the sinusoidal pattern evoking aspects of capillarization. The number of desmin-positive cells increased in DMN-treated animals, showing a prominent reaction in the fibrous septa. In the normal liver, lipocytes were positive for laminin and negative for actin, but septal and juxta-septal lipocytes were positive for both antigens, suggesting the presence of transitional cells with mixed immunoreactivity. This was confirmed by ultrastructural studies which showed typical intraseptal myofibroblasts and other elements exhibiting the structural features of both myofibroblasts and lipocytes.

Actins

Immunohistochemical identification of proliferating cells following dimethylnitrosamine-induced liver injury.

The present study is concerned with changes in the number and localization of S-phase cells in the liver of rats exposed to dimethylnitrosamine (DMN). S-phase cells were detected by immunohistochemistry after injection of bromodeoxyuridine (BrdU) and exposure of paraffin sections of liver tissue to the antibody anti-BrdU. With respect to controls, the number of S-phase cells increased four to fivefold in DMN-treated animals in the first week of treatment and remained significantly higher thereafter, in association with the formation of septa. At all times, the labelling index was higher in littoral cells than in hepatocytes. No labelling was observed in biliary cells. This behaviour is different from that reported in other situations, for instance in regeneration after partial hepatectomy, which suggests that besides hepatocytes and littoral cells replacement, an involvement of the latter cell line in the inflammatory reaction, synthesis of extracellular matrix components and formation of septa may account for this particular pattern.

Animals