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The extracellular matrix of unfixed, unstained rat corneal stroma, visualized with high-resolution scanning electron microscopy and atomic force microscopy after minimal preliminary treatment, appears composed of straight, parallel, uniform collagen fibrils regularly spaced by a three-dimensional, irregular network of thin, delicate proteoglycan filaments. Rat tail tendon, observed under identical conditions, appears instead made of heterogeneous, closely packed fibrils interwoven with orthogonal proteoglycan filaments. Pre-treatment with cupromeronic blue just thickens the filaments without affecting their spatial layout. Digestion with chondroitinase ABC rids the tendon matrix of all its interconnecting filaments while the corneal stroma architecture remains virtually unaffected, its fibrils always being separated by an evident interfibrillar spacing which is never observed in tendon. Our observations indicate that matrix proteoglycans are responsible for both the highly regular interfibrillar spacing which is distinctive of corneal stroma, and the strong interfibrillar binding observed in tendon. These opposite interaction patterns appear to be distinctive of different proteo- glycan species. The molecular details of proteoglycan interactions are still incompletely understood and are the subject of ongoing research.

Specimens of human salivary glands have been studied by our modification of the AODO maceration method which, carried out on sections of controlled thickness, allows the analytical study of human bioptical material. Lately, our technique has been further improved and simplified by omitting the treatment with dimethylsulfoxide and by using osmium-ferrocyanide as secondary fixative. Following maceration with diluted OsO4, some of the sections also were shaken for 10-15 min with a rotating agitator. Already at low magnification, all parenchymal cells were clearly distinguishable for their complement of cytoplasmic organelles. Serous cells and mucous cells at the beginning of their secretory cycle were characterized by well developed RER and Golgi apparatus, while mature mucous cells exhibited only scanty organelles compressed among the secretory droplets. Mitochondria were tubular, and often branched and convoluted. When sectioned, these organelles, besides the usual plate-like cristae, showed tubular cristae as well. The SER of striated and excretory duct cells was well developed and consisted of a network of smooth anastomosing tubules in the apical cytoplasm where it probably represented the transcellular pathway for ion transport. In specimens subjected to shaking, cytoplasmic organelles were occasionally removed allowing a nonobstructed view of the inner side of the plasmalemma and its specializations. With this technique the intercellular canaliculi of serous cells also became appreciable from their cytoplasmic side. They appeared as ribbon-like irregular protrusions with walls fenestrated by holes, corresponding to the interior of microvilli deprived of the cytoskeleton, and, sometimes, with lateral expansions possibly related to the mechanism of exocytosis. Results reported here clearly showed the usefulness of the maceration method in providing additional data on the cytoarchitecture of epithelial cells of salivary glands. Furthermore, by allowing the visualization of internal surfaces previously hidden to direct inspection, our technique may open new horizons in morpho/functional studies of human salivary glands.

We performed an experimental study on 46 male rats to evaluate the vitality of superficial inferior epigastric neurovenous flaps performed following different procedures.Rats were divided into 5 groups: A (n=6), B (n=10), C (n=10), D (n=10) and E (n=10). On each rat a hemi-abdominal flap based on the superficial inferior epigastric pedicle was raised. In group A the flap was sutured into its original position, isolating it from underlying tissues by a latex glove patch. In group B an arteriectomy of the superficial inferior epigastric artery was performed. In group C the superficial inferior epigastric artery was sectioned at its origin, without dissecting it out of the pedicle. In group D and E the same procedures performed respectively in group B and C were followed, but the epigastric pedicles were immediately explanted for optic microscope observation. Flap vitality in groups A, B and C was checked after 7 days.Flaps in group A appeared completely vital: the glove did not interfere with flap survival. All flaps in group B looked necrotic. In group C, 9 flaps survived completely and 1 flap underwent complete necrosis. Pedicle specimens of the viable flaps (group C) were explanted for observation under the optic microscope. Each specimen from groups C, D, E revealed a rich vascular plexus in the connective tissue around the pedicle. Seven days after surgery, neo-microangiogenesis was evident.The results witness the importance of perivenous and perineural arterioles for neurovenous flaps survival, also considering the adaptive increase in capillary development in the days following surgery.

Long-term effects of glucocorticoid treatment in humans induce bone loss and increase the risk of fracture in the skeleton. The pathogenic mechanisms of glucocorticoid-induced osteoporosis (GIOP) are still unclear. The GIOP and its effects have been reproduced in several animal models including Danio rerio (zebrafish) embryo. The treatment of adult fish with prednisolone (PN) has shown a dose-dependent decrease of mineralized matrix in the scales. Large resorption lacunae are characterized by single TRAP-positive cells which migrate to the margin of the scale merging into a multinucleated structures. The treatment with PN of cultured scales did not increase TRAP activity suggesting that the massive presence of osteoclasts in the resorption sites could be likely the result of a systemic recruitment of monocyte–macrophage precursors. We observed that treatment with PN induced a significant decrease of the alkaline phosphatase (ALP) activity in scale scleroblasts if compared with untreated controls. Then, we investigated the total mineral balance under prednisolone treatment using a time-dependent double live staining. The untreated fish fully repaired the resorption lacuna induced by prednisolone, whereas treated fish failed. The presence of osteoclast resorption fingerprints on new matrix suggested that the osteoclast activity counterbalances the osteodepositive activity exerted by scleroblasts. The treatment with PN in association with alendronate (AL) has surprisingly resulted in a significant decrease of TRAP activity and increase of ALP compared to PN-treated fish in biochemical and histological assays confirming the action of alendronate against GIOP in fish as well in humans.

Carbon nanotubes (CNTs) have been extensively investigated and employed for industrial use because of their peculiar physical properties, which make them ideal for many industrial applications. However, rapid growth of CNT employment raises concerns about the potential risks and toxicities for public health, environment, and workers associated with the manufacture and use of these new materials. Here we investigate the main routes of entry following environmental exposure to multi-wall CNTs (MWCNTs; currently the most widely used in industry). We developed a novel murine model that could represent a surrogate of a workplace exposure to MWCNTs. We traced the localization of MWCNTs and their possible role in inducing an innate immune response, inflammation, macrophage recruitment, and inflammatory conditions. Following environmental exposure of CD1 mice, we observed that MWCNTs rapidly enter and disseminate in the organism, initially accumulating in lungs and brain and later reaching the liver and kidney via the bloodstream. Since recent experimental studies show that CNTs are associated with the aggregation process of proteins associated with neurodegenerative diseases, we investigated whether MWCNTs are able to induce amyloid fibril production and accumulation. Amyloid deposits in spatial association with macrophages and MWCNT aggregates were found in the brain, liver, lungs, and kidneys of exposed animals. Our data suggest that accumulation of MWCNTs in different organs is associated with inflammation and amyloid accumulation. In the brain, where we observed rapid accumulation and amyloid fibril deposition, exposure to MWCNTs might enhance progression of neurodegenerative and other amyloid-related diseases. Our data highlight the conclusion that, in a novel rodent model of exposure, MWCNTs may induce macrophage recruitment, activation, and amyloid deposition, causing potential damage to several organs., Video abstract

To evaluate normoglycemic overweight/obese (Ow/Ob) children whose post-load plasma glucose (2hPG) cut-point may be significantly associated with cardiometabolic risk factors (CMRFs) and whether this cut-point predicts preclinical signs of organ damage.One thousand seven hundred and thrity four normoglycemic Ow/Ob children were stratified into quintiles of 2hPG, the sixth group was constituted by 101 children with impaired glucose tolerance (IGT).Moving from the lower quintiles of 2hPG to IGT, the groups differed for Prepubertal stage, BMI, fasting PG, insulin levels, blood pressure, and lipids. To evaluate the best cut-off of 2hPG related to CMRFs, the area under the receiver operating characteristic curve and the Youden's index was calculated. Insulin resistance, high blood pressure, and high triglyceride/HDL-C ratio were associated with a 2hPG cut-off of 110 mg/dl. Children with 2hPG ≥110 mg/dl showed 1.3-3.2 fold higher risk to have high levels of ALT (as surrogate of nonalcoholic fatty liver disease) or increased carotid intima-media thickness.This study, performed in a large cohort of Ow/Ob children, shows that an atherogenic risk profile and preclinical signs of organ damage are associated with post-challenge elevations in plasma glucose still considered in the high normal range.