Luciano Fonzi

Nestin Expression in Adult and Developing Human Kidney

Nestin is considered a marker of neurogenic and myogenic precursor cells. Its arrangement is regulated by cyclin-dependent kinase 5 (CDK5), which is expressed in murine podocytes. We investigated nestin expression in human adult and fetal kidney as well as CDK5 presence in adult human podocytes. Confocal microscopy demonstrated that adult glomeruli display nestin immunoreactivity in vimentin-expressing cells with the podocyte morphology and not in cells bearing the endothelial marker CD31. Glomerular nestin-positive cells were CDK5 immunoreactive as well. Western blotting of the intermediate filament-enriched cytoskeletal fraction and coimmunoprecipitation of nestin with anti-CDK5 antibodies confirmed these results. Nestin was also detected in developing glomeruli within immature podocytes and a few other cells. Confocal microscopy of experiments conducted with antibodies against nestin and endothelial markers demonstrated that endothelial cells belonging to capillaries invading the lower cleft of S-shaped bodies and the immature glomeruli were nestin immunoreactive. Similar experiments carried out with antibodies raised against nestin and α-smooth muscle actin showed that the first mesangial cells that populate the developing glomeruli expressed nestin. In conclusion, nestin is expressed in the human kidney from the first steps of glomerulogenesis within podocytes, mesangial, and endothelial cells. This expression, restricted to podocytes in mature glomeruli, appears associated with CDK5.

Influence of amalgam fillings on Hg levels and total antioxidant activity in plasma of healthy donors.

In order to evaluate the influence of specific factors on mercury (P-Hg) levels and antioxidant power (P-FRAP) in human plasma, 26 healthy donors were examined by a dentist, their plasma analyzed for Hg by atomic absorption spectrometry and for total antioxidant activity by FRAP method. Hg plasma concentration was found to be correlated with the number of amalgam fillings, suggesting that Hg released from fillings is a source of Hg in non-occupational exposed subjects. P-FRAP correlated negatively with P-Hg suggesting a pro-oxidant role of the Hg released from amalgam fillings. Though age by itself was not significantly correlated with P-FRAP, when considered together with P-Hg in multivariate analysis, it was found to be a major related cofactor. Multivariate analysis showed no influence of fish consumption or cigarette smoking on P-FRAP.

Ultrastructural Abnormalities in Respiratory Cilia and Sperm Tails in a Patient with Kartagener's Syndrome

Ultrastructural abnormalities of spermatozoa and respiratory cilia have been reported in a male patient with Kartageners syndrome and infertility. In this patient both respiratory cilia and sperm tails showed defects in radial spokes and dynein arms. Such defects are heretofore undescribed in the same subject with immotile cilia syndrome. Absence of both inner and outer dynein arms and absence of the inner dynein arms only were detected in spermatozoa and in respiratory tract cilia, respectively. Moreover, total absence of axoneme was seen in several sperm tails from this patient. The possibility that the features described are of genetic origin is discussed.

Nestin expression in rat adrenal gland

Abstract. The constituents of the intermediate filament network of adrenal gland cells have not been deeply investigated in vivo. Adrenocortical cells have been reported to express cytokeratins and vimentin, but the intermediate filament components of the adrenomedullary cells are still unknown. Nestin is an intermediate filament protein that is mainly expressed in the developing nervous and muscle systems. It has been reported to be unable to form filaments by itself and it co-assembles with vimentin. Using immunocytochemical and biochemical approaches, the present study demonstrates that nestin is expressed in situ either in the cortex or in the medulla of adult rat adrenal glands. Nestin-negative cells prevalently form the zona glomerulosa whereas the zona fasciculata and the zona reticularis are mainly nestin-immunoreactive. Nestin-positive cells always express vimentin-like immunoreactivity but several cells apparently expressing only vimentin are detectable too. Nestin is also expressed by adrenomedullary cells that also display a faint vimentin-like immunoreactivity. We hypothesise that the inconstant detection of nestin in adrenocortical cells depends on their different functional moments. Moreover, even though our data do not allow to confirm vimentin in adrenomedullary cells, in situ detection of nestin in the adrenal medulla indirectly supports in vivo expression of vimentin in chromaffin cells.

Rabbit Tonsil-associated M-cells Express Cytokeratin 20 and Take Up Particulate Antigen

M-cells are believed to play a pivotal role in initiation of the immune response. These cells, located in the epithelia that overlie mucosal lymphoid follicles, are responsible for the active uptake of particulate antigens and for their translocation to the underlying lymphoid tissue. The identification of reliable markers for M-cells is therefore extremely important for the study of the initial steps that lead to the immune response. For this purpose, we studied cytokeratin 20 (CK20) expression in the epithelium of rabbit palatine tonsils by immunofluorescence, confocal microscopy, and Western blotting. CK20+ cells were observed in all rabbit palatine tonsils examined. By Western blotting, one CK20-immunoreactive band was identified at 46 kD on samples of proteins from the intermediate filament-enriched cytoskeletal fraction of tonsil epithelium. Double labeling of CK20+ cells with cell-specific markers confirmed that such cells were actually M-cells. Moreover, CK20+ M-cells displayed a mature phenotype (they formed pockets harboring lymphoid cells) and were functionally competent because they could take up particulate antigens from the pharyngeal lumen. We conclude that CK20 is an M-cell marker for rabbit palatine tonsils. Moreover, we can hypothesize the use of M-cells as a possible site for antigen delivery of particle-based vaccines.

An Appraisal of Intermediate Filament Expression in Adult and Developing Pancreas: Vimentin Is Expressed in α Cells of Rat and Mouse Embryos

Intermediate filaments are frequently used in studies of developmental biology as markers of cell differentiation. To assess whether they can be useful to identify differentiating pancreatic endocrine cells, we examined the pattern of expression of nestin, cyto-keratin 20, and vimentin on acetone-fixed cryosections of rat adult and developing pancreas. We also studied vimentin expression in mouse embryonic pancreas at E19. Cytokeratin 20 was found in all pancreatic epithelial cell lineages during the entire development of the rat gland and in the adult animals. Under our experimental conditions, therefore, cytokeratin 20 is not an exclusive marker of rat duct cells. Nestin was detected exclusively in stromal cells either in the adult or developing rat pancreas. Vimentin was observed within cells located in the primitive ducts of rat pancreas starting from E12.5. Their number rapidly increased, reaching its highest level in newborn animals. Vimentin was also spotted in α cells starting from E12.5 but disappeared soon after birth, likely identifying immature or recently differentiated α cells. In addition, vimentin was observed in duct and α cells of mouse developing pancreas showing that its expression in such cells is not an event restricted to the rat. This manuscript contains online supplemental material at http://www.jhc.org. Please visit this article online to view these materials.

Elastolytic activity in rabbit leukocyte extracts. Effects of the whole leukocyte homogenate on the rabbit lung

Abstract Studies performed on dogs treated with whole PMN homogenates from man and different animal species have shown that those obtained from rabbits hardly ever produce pulmonary emphysema. The presence in the cytoplasmic fraction of rabbit polymorphonuclear leukocytes of a powerful inhibitor towards neutral lysosomal proteinases may explain this different action. In this study we have investigated the degree of inhibition developed in vitro by the cytoplasmic fraction of rabbit PMNs towards elastolytic activity of granule (lysosomal) extracts. The whole homogenate of rabbit PMNs shows in comparison with granular fraction, a reduced elastolytic activity in vitro , together with a good emphysema-producing ability. These data and the characteristic aspects of the rabbit enzyme-mediated pulmonary damage are discussed in this study.

Pancreatic lymphatic system in rodents

The lymphatic network of the pancreas has been little investigated and recent studies have provided contrasting data. This research is aimed to supply the morphologic basis to outline the involvement of the lymphatic system in pancreatic pathology. Guinea pigs, rats, and mice were anesthetized with ether and sacrificed with the same anesthetic. Pieces of pancreas were processed for transmission electron microscopy. Semithin sections were observed by light microscopy and, after positive identification by transmission electron microscopy, lymphatics were followed with long series of consecutive sections to define their distribution. Lymphatics were detected in the pancreas of all the animals both in the inter and the intralobular sites. Closer relations with the exocrine parenchyma (ducts and acini) were observed in guinea pig pancreas. Remarkably, interesting relationships between lymphatics and endocrine tissue were observed in all the animals. Overall, however, the lymphatic network of rat pancreas was less develop and preferentially associated with blood vessels. The distribution of the pancreatic lymphatic network appears consistent with an active role in pancreatic pathology. Anat Rec 263:155–160, 2001.

Pulmonary changes induced in rabbits by long-term exposure to n-hexane

New Zealand male rabbits were exposed in an inhalation chamber to 3,000 ppm of n-hexane 8 h/day, 5 days/week for 24 weeks, resulting in a total of 120 exposures. After a further 120 days in clean air, morphologic examination of lungs by light and transmission electron microscopy (TEM) revealed three important exposure-related lesions. These consisted of air space enlargement centered on respiratory bronchioles and alveolar ducts (centriacinar emphysema), scattered foci of pulmonary fibrosis, and papillary tumors of non-ciliated bronchiolar epithelial cells. At TEM examination these tumors were rather homogeneous and were found to consist of cells showing both small and large electron dense ovoid granules in their cytoplasmic matrix, a feature that has been described for Clara cells.The implications of these findings are discussed.

Response of lung enzyme activities in rabbits following short-term exposure ton-hexane: Correlation between morphological and biochemical changes

The activity of lactatedehydrogenase, β-glucuronidase, glucose-6-phosphate dehydrogenase, acid and alkaline phosphatase was studied in lung homogenate from New Zealand rabbits exposed to 3000 p.p.m. ofn-hexane 8 h per day for 8 days or filtered air.In hydrocarbon-treated animals all enzymes examined, except alkaline phosphatase, were markedly increased.The biochemical changes correlated well with the morphological changes and the results of cytological evaluation of bronchopulmonary lavage. It is suggested that high values in lung lysosomal enzymes from treated rabbits reflect the acute inflammation whilst the increase in lung glueose-6-phosphate dehydrogenase may depend upon reparative process subsequent ton-hexane-induced lung damage.