Polyglucosan body density in the aged mouse hippocampus is controlled by a novel modifier locus on chromosome 1.

Aging can be associated with the accumulation of hypobranched glycogen molecules (polyglucosan bodies, PGBs), particularly in astrocytes of the hippocampus. While PGBs have a detrimental effect on cognition in diseases such as adult polyglucosan body disease and Lafora disease, the underlying mechanism and clinical relevance of age-related PGB accumulation remains unknown. Here, we have investigated the genetic basis and functional impact of age-related PGB accumulation in 32 fully sequenced BXD-type strains of mice which exhibit a 400-fold variation in PGB burden in 16-18 month old females. We mapped a major locus controlling PGB density in the hippocampus to chromosome 1 at 72-75 Mb (linkage of 4.9 -logP), which we defined as the Pgb1 locus. To identify potentially causal gene variants within Pgb1, we generated extensive hippocampal transcriptome datasets and identified two strong candidate genes for which mRNA correlates with PGB density-Smarcal1 and Usp37. In addition, both Smarcal1 and Usp37 contain non-synonymous allele variations likely to impact protein function. A phenome-wide association analysis highlighted a trans-regulatory effect of the Pgb1 locus on expression of Hp1bp3, a gene known to play a role in age-related changes in learning and memory. To investigate the potential impact of PGBs on cognition, we performed conditioned fear memory testing on strains displaying varying degrees of PGB burden, and a phenome-wide association scan of ~12,000 traits. Importantly, we did not find any evidence suggesting a negative impact of PGB burden on cognitive capacity. Taken together, we have identified a major modifier locus controlling PGB burden in the hippocampus and shed light on the genetic architecture and clinical relevance of this strikingly heterogeneous hippocampal phenotype.

Comparative study of the primary cilia in thyrocytes of adult mammals.

Since their discovery in different human tissues by Zimmermann in 1898, primary cilia have been found in the vast majority of cell types in vertebrates. Primary cilia are considered to be cellular antennae that occupy an ideal cellular location for the interpretation of information both from the environment and from other cells. To date, in mammalian thyroid gland, primary cilia have been found in the thyrocytes of humans and dogs (fetuses and adults) and in rat embryos. The present study investigated whether the existence of this organelle in follicular cells is a general event in the postnatal thyroid gland of different mammals, using both immunolabeling by immunofluorescence and electron microscopy. Furthermore, we aimed to analyse the presence of primary cilia in various thyroid cell lines. According to our results, primary cilia are present in the adult thyroid gland of most mammal species we studied (human, pig, guinea pig and rabbit), usually as a single copy per follicular cell. Strikingly, they were not found in rat or mouse thyroid tissues. Similarly, cilia were also observed in all human thyroid cell lines tested, both normal and neoplastic follicular cells, but not in cultured thyrocytes of rat origin. We hypothesize that primary cilia could be involved in the regulation of normal thyroid function through specific signaling pathways. Nevertheless, further studies are needed to shed light on the permanence of these organelles in the thyroid gland of most species during postnatal life.

Apoptosis in breast carcinoma.

Apoptosis may play a major role in determining tumor growth and aggressiveness. The aim of this study was to examine the relationship between apoptosis, expression of bcl-2 and p53 proteins, proliferation index, and other clinicopathological features of breast carcinoma. Sixty-five formalin-fixed paraffin-embedded tissue sections from invasive ductal breast carcinomas were studied for the presence of apoptosis by the terminaldeoxynucleotidyl-transferase-mediated dUTP-FITC nick end-labeling (TUNEL) method. Immunohistochemical methods were also used to determine the expression of estrogen receptor, Ki67, bcl-2 and p53 proteins. The number of apoptotic cells ranged from 2.0 to 236.0/10HPF (mean 36.26, median 28.0). The observation of 30 apoptotic cells/10HPF was more common in tumors > 3 cm, of histological grade III, with a high mitotic index, Ki67 index > or = 300, and p53 positivity; however, statistical significance was found only for the histological grade. Grade I and III tumors displayed an inverse association between the apoptotic index and bcl-2 and p53 protein expressions; grade I tumors frequently expressed bcl-2 (19/28), lacked p53 (20/28), and presented a low number of apoptotic cells (18/28), whereas grade III tumors tended to express p53 (12/17), lacked bcl-2 (13/17), and displayed a high number of apoptotic cells/10HPF (12/17). Multivariate analysis for survival revealed that estrogen receptors and apoptosis were independent variables. These data suggest that apoptosis, rather than proliferation index or expression of bcl-2 or p53 proteins, is an independent factor for the prognosis of survival.

Glomerular basement membrane selective permeability in short-term streptozotocin-induced diabetic rats.

In diabetes, the glomerular basement membrane undergoes thickening and structural alterations with loss of glomerular permselectivity properties. However, the onset of the alterations at early phases of diabetes is unclear. Aiming to determine the functional and structural alterations of the glomerular wall in the early stages of diabetes, we have studied the distribution of endogenous circulating albumin and type IV collagen in the glomerular basement membrane, using the immunocytochemical approach. The streptozotocin-injected hyperglycemic rat was our animal model. Renal tissues were examined after 10 days, 2, 4 and 6 months of hyperglycemia. Upon immunogold labelings, changes in the glomerular permeability to endogenous albumin were found altered as early as upon ten days of hyperglycemia. In contrast, no structural modifications were detected at this time point. Indeed, glomerular basement membrane thickening and an altered type IV collagen labeling distribution were only observed after four months of hyperglycemia, suggesting that functional alterations take place early in diabetes prior to any structural modification. In order to evaluate the reversibility of the glomerular alterations, two-month-old diabetic animals were treated with insulin. These animals showed a significant restoring of their glomerular permselectivity. Our results suggest a link between glycemic levels and alteration of glomerular permeability in early stages of diabetes, probably through high levels of glycated serum proteins.

Sarcomatoid carcinoma of the skin: report of a case.

We report the histological, immunohistochemical and ultrastructural findings of an exophytic cutaneous tumor composed of a mixture of typical basal cell carcinoma (BCC) and malignant fibrous histiocytoma. Nine previously reported carcinosarcomas of the skin are reviewed. We prefer the term "sarcomatoid carcinoma" for this rare neoplasm. Only the BCC showed a positive immunoreaction to cytokeratin; the sarcomatous component was negative, but it did express vimentin, and, focally, smooth-muscle-specific actin and KP1 (CD68). Both components showed p53 immunostaining.

Mesenchymal chondrosarcoma of the retroperitoneum. Report of a case diagnosed by fine needle aspiration biopsy with immunohistochemical, electron microscopic demonstration of S-100 protein in undifferentiated cells.

BACKGROUND: Mesenchymal chondrosarcoma (MC), first described in 1959 by Lichtenstein and Bernstein, is an uncommon tumor of the soft tissues and bone defined by the coexistence of nests of well-defined cartilaginous tissue within a proliferation of primitive mesenchymal cells. CASE: A 27-year-old, white female consulted a physician for a left lumbar mass associated with sciatica-type pain, asthenia, anorexia and weight loss. Abdominal computed tomography showed a 20-cm, retroperitoneal neoplasm that occupied the lower half of the left abdominal flank. Fine needle aspiration biopsy showed clusters of small round cells mixed with islets of mature cartilage. Immunomarking for electron microscopy, carried out with 15 nm of A protein/colloidal gold, showed specific immunoreactivity to S-100 protein in the nuclear chromatin, nucleoli, Golgi apparatus, plasma membrane and cytosol of cells with more developed organelles. CONCLUSION: The diagnosis of MC rests on recognition of the biphasic component of small, undifferentiated cells and cartilage islets. In aspirates with only undifferentiated cells, demonstration of ultrastructure-specific immunoreactivity to S-100 protein may contribute to the final diagnosis.

Immunocytochemical investigation of the in vivo endocytosis by renal tubular epithelial cells.

The internalization and degradation of glomerular filtered serum proteins by the proximal tubular epithelium has been extensively studied by microperfusion methods. By using a cationic probe that easily traverses the glomerular wall into the urinary space, we have performed a morpho-cytochemical and quantitative study of the in vivo endocytotic activity of the proximal tubular epithelial cell. Bovine serum albumin (BSA) was tagged with dinitrophenol (DNP) and cationized to pI over 8. It was introduced into the circulation of normal mice for 5, 10, and 30 minutes and the distribution of the labeling was determined by protein A-gold immunocytochemistry, using specific antiDNP antibodies on tissue sections of routinely aldehyde-fixed, osmiumpostfixed, and Epon-embedded kidneys. Cationic BSA-DNP was detected at the endothelial and epithelial sides of the glomerular basement membrane, and over capillary and tubular basement membranes. In the proximal tubular epithelial cell, labeling was present over microvilli as well as over endosomal and lysosomal compartments, with labeling intensities varying from one compartment to the other. Morphometric evaluations of the labeling demonstrated a progressive incorporation of the probe from microvilli and endocytic compartments at 5 minutes to endocytic and lysosomal compartments at 10 and then 30 minutes. When considering labeling densities, no significant differences were found on microvilli and basolateral membranes between times of circulation; however, the labeling density over endosomal and lysosomal compartments was very intense at 10 minutes compared with 5 minutes, decreasing at 30 minutes. Results from this study validate the cationic albumin tagged with DNP as a tool in the study of the quantitative aspects of protein endocytosis at the ultrastructural level, in the kidney tubular epithelium.

Fine needle aspiration cytology of primary soft tissue tumors. Morphologic analysis of the most frequent types.

We reviewed 98 cases of fine needle aspiration of soft tissue tumors with histologic confirmation performed during an eight-year period and propose a working morphologic classification based on the most prominent cytologic features. Six main tumor groups are recognized: myxoid, round cells, spindle cells, pleomorphic cells, polygonal cells and well differentiated. We believe that this classification allows recognition of the most common soft tissue tumors while helping with the differential diagnosis of other neoplasia, primary or secondary, with similar morphology and a similar presentation.

Fine needle aspiration of peripheral neuroepithelioma of soft tissues.

Peripheral neuroepithelioma of soft tissue is a malignant primitive neuroectodermal tumor that appears both in children and adults and usually has a poor outcome. Fine needle aspiration on two patients with tumors in the lower limbs showed small round cells with unipolar processes and a tendency to form Homer-Wright rosettes. The cells had a round to oval nucleus with fine chromatin, up to four small, conspicuous nucleoli and vacuolated, periodic acid-Schiff-positive cytoplasm. The diagnosis was supported by electron microscopic study of the aspirate, which showed features of neuroblastic differentiation (i.e., neurosecretory granules), and by histologic, immunohistochemical and cytogenetic study of the resected tumors.

Nucleolar organizer regions in pigmented skin lesions. Value in the differential diagnosis of Spitz nevi.

Forty-one cases of typical melanocytic skin lesions (15 intradermal nevi, 14 Spitz nevi and 12 malignant melanomas) were used to investigate the value of staining of nucleolar organizer regions (NORs) in the differential diagnosis of such pigmented lesions. Histologic sections were stained by the silver colloid (Ag) method, with and without the prior use of a melanin blocking agent. There were statistically significant differences in the mean numbers of AgNORs per nucleus between the groups of lesions studied (1.658 for intradermal nevi, 3.0042 for Spitz nevi and 6.669 for malignant melanomas). Sections treated with potassium permanganate (melanin blocking agent) prior to staining showed an obvious increase in the AgNOR scores in all groups; this increase was highest for Spitz nevi. Although AgNOR staining allows a distinction to be made between intradermal nevi and malignant melanomas, the striking overlap between the counts for Spitz nevi and malignant melanomas precludes the use of this technique as the sole method for establishing the diagnosis of malignancy. Other clinical and morphologic data are especially required to make the diagnosis of Spitz nevi.

Neuroendocrine (NE) cells in rat neonatal lungs. A histochemical and immunocytochemical study.

A study has been carried out to determine the presence of NE cells in the newborn rat lung. The localization of these cell was achieved by an argyrophil method. Both single NE and NEB, were found. Immunoperoxidase techniques were performed to determine NSE, serotonin and calcitonin production in these NE formations.

Cytologic diagnosis of a case of recurrent adamantinoma.

The cytologic features of a case of recurrent adamantinoma of the tibia diagnosed by fine needle aspiration (FNA) in a 31-year-old patient are described. The FNA smears contained single-lying cells and cells arranged in small clusters and Indian files, with some nuclear molding. Three cell types were present: (1) large polygonal cells with chromatin clearing, (2) smaller cells with poorly defined cytoplasm and dense chromatin and (3) fusiform cells. The first two were interpreted as epithelial in nature and the third one as being derived from the stroma. These cytologic features appear to be sufficiently characteristic to suggest an FNA diagnosis of adamantinoma, especially in the case of a recurrent tumor.

Demonstration of sugar residues in the ultimobranchial tubule and thyroid C-cells of the rat using peroxidase labelled lectins.

The distribution of various sugars, that contribute to the composition of the carbohydrate components of the glycoprotein in the ultimobranchial tubule and the thyroid C-cells has been studied, using eight peroxidase-labelled lectins (Dolichos biblorus, Glycine max, Bauhinia purpurea, Helix pomatia, Arachis hipogaea, Ulex europaeus I, Limax flavus and Sophora japonica). The results obtained seem to show that there are receptors for Dolichos biflorus, Glycine max, Bauhinia purpurea, Helix pomatia, Arachis hipogaea, Ulex europaeus I and Limax flavus in the ultimobranchial tubule, suggesting the existence of N-acetylgalactosamine, galactose, L-fucose and sialic acid. However, the C-cells show only a positive reaction with Ulex europaeus I indicating the existence of fucosyl residues in those cells. All these findings support the contention that the ultimobranchial body might contribute decisively not only to the formation of C-cells but also, partially, to the constitution of certain follicular cells.