MRI Findings in Patients with Clinical Onset Consistent with Infantile Neuroaxonal Dystrophy (INAD), Literature Review, Clinical and MRI Follow-up.

Infantile neuroaxonal dystrophy (INAD) is a rare autosomal recessive neurodegenerative disorder characterized by infantile onset and rapid progression of psychomotor regression and hypotonia evolving into spasticity. The neuroradiologic hallmark of the disease is represented by progressive cerebellar atrophy. Prior to the discovery of mutations in the PLA2G6 gene in family with INAD, the clinical diagnosis of the disease had been confirmed by the presence of spheroid bodies (SB) in a peripheral nerve biopsy. Various studies have found that some patients with mutations lacked SB and some without mutations had SB, indicating incomplete detection using either pathologic or molecular methods (7). This, together with the observation that the spectrum of clinical features associated with mutations in PLA2G6 is broader than previously described, has increased the usefulness of Magnetic Resonance (MR) in INAD diagnosis, particularly in the frequent occurrence of atypical cases, especially in the early stages of the disease. We retrospectively reviewed the MR studies of eight patients in whom clinical and imaging onset met the typical criteria for INAD. Their clinical and MR imaging (MRI) onset and follow-up were evaluated together with the neuroradiological findings reported in the literature in order to identify MRI features useful in differentiating INAD from other diseases with similar clinical onset and to discuss which of them are the most important, thus suggesting INAD diagnosis. Our contribution included the use of Proton Spectroscopy ((1)H-MR), diffusion weighted MR imaging (DWI) and diffusion tensor imaging (DTI) in the follow-up of seven of the eight patients. The literature reviewed included attempts to correlate clinical and MR data with the genotype in the group of patients carrying PLA2G6 mutations. From the limited and inhomogeneous cohort of patients included in our study, a correlation between the MR features, phenotype and genotype was not exhaustive.

'Normolipidemic' tendinous and tuberous xanthomatosis.

BACKGROUND: Multiple tendinous and tuberous xanthomas are characteristically associated with hyperlipidemic states. However, normolipidemic tendinous and tuberous xanthomas have been reported in the literature, with normal levels of cholesterol, cholestanol and plant sterols. OBJECTIVE AND METHOD: To delineate the disorder and to suggest its likely origin, a case of apparently normolipidemic severe tuberous and tendinous xanthomatosis was studied. Several lipoprotein and lipid analyses, clinical tests and histological studies were performed over a period of 5 years in the propositus and his family. RESULTS: At the first lipid analysis, no quantitative or qualitative alterations of the lipoprotein fractions or of the apoproteins AI, B, CII, CIII, E were detected in the propositus and xanthomatosis was classified as normolipidemic. During the follow-up, the patient showed a nonconstant hypertriglyceridemia and/or hypercholesterolemia associated with the presence of small and dense VLDL and LDL. An increase in apo-B was observed. There was an unusual quantity of conjugated dienes of arachidonic acid in the plasma and in the LDLs of the patient, present only in small traces in the control population. The family study and the long follow-up of the lipid analysis of the propositus were compatible with the diagnosis of familial combined hyperlipidemia. CONCLUSION: Our data highlight the importance of a critical review of studies regarding normolipidemic xanthomatosis, since only after an extensive follow-up and sequential analyses of lipoprotein fractions is it possible to exclude the presence of time variables and complex lipoprotein abnormalities.

Filamentous inclusions in nonneoplastic and neoplastic pancreas: an ultrastructural and immunogold labeling study.

Filamentous inclusions (FI) are unusual, irregularly shaped cytoplasmic inclusions, which are mostly found in acinar cell carcinomas of the pancreas and are consequently thought to be an abnormal zymogen granule type. This study describes identical inclusions in acinar, centroacinar, and small duct epithelial cells from nonneoplastic pancreas, as well as those found in tumor cells from a mixed acinar-endocrine pancreatic carcinoma. An ultrastructural and immunogold labeling demonstration indicates that these inclusions are aggregates of intermediate filaments immunoreacting with the anti-cytokeratin AE1/AE3 mixture and with V9 clone anti-vimentin monoclonal antibodies. Their pleomorphic appearance, variable immunoreactivity, and frequent association with lipid droplets and secondary lysosomes, mostly of the angulate type, led to the hypothesis that the FI undergo a degenerative remodeling pathway similar to that proposed for hepatic Mallory bodies. A survey of the literature on FI and human tumors suggests that they are a variably expressed ultrastructural feature of tumor cells originating from exocrine cell-containing tissues, namely the pancreas and gastrointestinal tract.

Primary systemic amyloidosis presenting with polyneuropathy characterized by very long survival.

We report a case of primary systemic amyloidosis associated with IgA monoclonal gammopathy presenting with sensorimotor polyneuropathy. For 10 years the neurological symptoms were the only clinical manifestation. A great deal of therapy was given right from the onset of symptoms and the very long survival of the patient may have been due to these efforts.

[Wash and non-wash system in the perioperative recovery of blood in prosthetic surgery: ultrastructural assessment of corpuscular blood components]. / Sistema wash e non-wash nel recupero perioperatorio del sangue in chirurgia protesica: valutazione ultrastrutturale della parte corpuscolata del sangue.

AIM: The authors aimed to perform an ultrastructural morphological analysis of blood recovered using wash and non-wash systems in patients undergoing full cement-free hip replacement in order to evaluate the integrity of the various blood corpuscle components. EXPERIMENTAL PROTOCOL: An open prospective study in patients undergoing full cement-free hip replacement at the Orthopedics Division of S. Orsola-Malpighi Policlinico in Bologna. Materials of S. Orsola-Malpighi Policlinico in Bologna. MATERIALS AND METHODS: Blood recovered postoperatively using a non-wash system was studied in 6 patients. In a further 3 patients perioperatively recovered blood was studied after washing using Cell Saver Haemolite 2 before reinfusion. Red globules, white globules and plaelets were isolated from blood collected using these two different recovery systems and analysed by SEM. RESULTS: Study of the ultrastructural morphology of various corpusculated blood fractions. DISCUSSION AND CONCLUSIONS: From the data in our possession it appears that the ultrastructural morphology of the various corpuscle components of blood in subjects undergoing postoperative recovery is better preserved using a non-wash system. There was no sign of "polluting" material in terms of adipose cells or free bone fragments in either group.

Hereditary sensory and autonomic neuropathy with ataxia and late onset.

We report two brothers affected by a dominantly inherited form of hereditary sensory and autonomic neuropathy (HSAN), characterized by clinical features of sensory ataxia, and by late onset in the 6th decade. Sural nerve biopsy in the proband showed almost complete loss of myelinated fibers, and relative sparing of unmyelinated fibers. This family showed an atypical presentation of HSAN, which is usually characterized by acrodystrophic manifestations of infantile or juvenile onset. Although a few reports of HSAN presenting with late onset and/or ataxia appeared, this is the first report of a family with dominant HSAN characterized by late onset sensory ataxia.

Long-term cryopreservation of autologous veins in rabbits.

This study was undertaken to determine the effect of long-term cryopreservation on graft ultrastructure and endothelial cell viability in an animal model. The jugular veins from 12 New Zealand White rabbits were excised with a 'no-touch' technique and divided into four groups: control group (fresh veins); group 1, veins cryopreserved for 1 month; group 2, veins cryopreserved for 2 months; and group 3, veins cryopreserved for 3 months. Cryopreservation was accomplished by rapid freezing (-5 degrees C s-1 to -196 degrees C) in a solution of 17.5% dimethylsulphoxide and 20% fetal bovine serum and by storage in liquid nitrogen. Veins were then implanted as a carotid autograft (three grafts/group). At the time of graft implantation a segment of the paired matched vein was perfusion-fixed and evaluated by scanning and transmission electron microscopy, whereas the remainder were subjected to endothelial cell culture techniques to determine cell viability. Autografts were removed 1 month after implantation and subjected to similar evaluations. Histological changes seen in cryopreserved veins were dependent on preservation time and included focal endothelial cell blebbing, cytoplasmic vacuolization and disruption of cell-to-cell contacts. Smooth muscle cells showed mitochondrial swelling. Patency was identical in all groups (66.6%). Explants at 1 month were similar in histological appearance to fresh veins with a smooth endothelial cell lining arranged longitudinally and intact cell junctions. Endothelial cells could be cultured from fresh veins and 1-month-old explants but not from the cryopreserved graft surface before implantation. the present technique of cryopreservation leads to some damage of graft architecture and loss of endothelial cell viability.(ABSTRACT TRUNCATED AT 250 WORDS)

Electron microscopic and immunocytochemical profiles of human subcutaneous fat tissue microvascular endothelial cells.

The ultrastructural and immunocytochemical characteristics of microvascular cells from human subcutaneous fat tissue were studied after the addition of collagenase and Percoll density gradient, respectively. Monoclonal and polyclonal antibodies directed against antigens specific for endothelial cells (factor VIII, Ulex europaeus, CD31, and CD34), pericytes (muscle-specific actin and desmin), adipocytes (S-100 protein), and monocytes-macrophages (MAC 387 and 150.95 protein) were demonstrated by alkaline phosphatase monoclonal anti-alkaline phosphatase and protein A-gold techniques. In addition, to determine whether the harvesting method interfered with microvascular cell function, DOT immunoassays of factor VIII and CD34 were conducted on solutions recovered at collagenase incubation as well as after nylon filtration and Percoll administration, respectively. After the collagenase step, the vast majority of microvascular cells had the typical ultrastructural and immunophenotypical features of endothelial cells. In sharp contrast, following the Percoll step, only 1% to 18% of microvascular cells stained with factor VIII, Ulex europeaus, and CD31, whereas 90% of them expressed the CD34 antigen. Surprisingly, DOT immunoassay revealed the presence of factor VIII in the washing buffer recovered after the Percoll step only. Consequently the decreased expression of common endothelial cell markers (factor VIII, Ulex europaeus, and CD31) observed at the end of the cell isolation procedure was related to the adverse effects of Percoll on endothelial cell function. The CD34 surface molecule, being highly resistant, is particularly well suited for unequivocal characterization of microvascular cells as true endothelium.

A new polychrome stain and simultaneous methods of histological, histochemical and immunohistochemical stainings performed on semithin sections of Bioacryl-embedded human tissues.

We describe a new polychrome stain and simultaneous methods of histological, histochemical and immunocytochemical staining performed on sections from human tissues embedded in the new hydrophilic resin Bioacryl. The polychrome stain involves the sequential use of Harris' Haematoxylin, silver methenamine, Light Green and Eosin or Safranin dyes and provides a highly specific visualization of the overall cytological tissue architecture. When histochemical, immunocytochemical, and polychrome stains are performed together on the same section, crisp images are obtained, yielding simultaneous data of histochemical and immunological reactivities with clear tissue architecture.

An immunohistochemical study of inflammatory abdominal aortic aneurysms.

Seven cases of inflammatory abdominal aortic aneurysms (IAs) were studied by light microscopy, transmission electron microscopy (TEM) and immunohistochemistry. Microscopically, atherosclerosis coexisted with adventitial fibrosis and inflammation. The inflammatory component showed a follicular and a diffuse pattern. Fibrous entrapment of fatty tissue, adventitial vasculitis, neuritis were also common findings. By TEM, sparse smooth muscle cells having dilated cisternae of rough endoplasmic reticulum, large bundles of collagen fibres and oedematous, amorphous fibrillary elastin were observed. By immunohistochemistry, the follicles mostly contained CD22+ B-cells. T4- (CD2+/CD4+/CD8-), T8-(CD2+/CD4-/CD8+) cells as well as macrophages (CD4+/CD11c+) and follicular dendritic reticulum cells (DRC1+) were also detected. The monoclonal antibody Ki-67 reacted with 2-48% of germinal center cells. In the fibrous extrafollicular adventitia, actively synthesizing plasma cells prevailed over T4-cells, and macrophages. Some of the macrophages were also activated (CD4+/CD11c+/CD25+/CD30-). IgM, IgG and C3c deposits were detected in the fibrous zone, in the germinal centers, within adventitial vessels and nerves. HLA-DR antigen was diffusely expressed in cells populating both the fibrous and the follicular zones as well as in endothelial and Schwann cells. These findings suggest that IAs could develop in some individuals affected by advanced atherosclerosis of the abdominal aorta through a pathogenic B-cell response to locally presented antigens.

Composite gastric carcinoma and precursor lesions with amphicrine features in chronic atrophic gastritis.

A composite carcinoma of the gastric body consisting of endocrine and mucous epithelial cells with interspersed amphicrine cells is reported together with ultrastructural and immunocytochemical documentation of endocrine and nonendocrine differentiation. The tumor was associated with hypergastrinemia related to chronic atrophic gastritis (achlorhydria) and with multiple proliferative lesions, such as intramucosal microcarcinoid (IMC) and endocrine cell proliferations of the micronodular and linear type, which are currently regarded as carcinoid precursor changes. Ultrastructurally, a composite architecture with amphicrine features was demonstrated in the primary tumor, IMC, and liver metastases. On the other hand, the endocrine cell proliferations exclusively contained gastrin and enterochromaffinlike cells. Immunostaining with antibodies to calcitonin documented a number of positive cells both in the primary and in the metastatic sites. This is the first report of mixed exocrine-endocrine-amphicrine components both in a metastasizing carcinoma and in its precursor lesions in a chronic hypergastrinemic state. Unlike previously reported lesions, the endocrine component was unexpectedly composed of calcitonin cells, which are not usually present in the gastric mucosa.

A new acrylic resin formulation: a useful tool for histological, ultrastructural, and immunocytochemical investigations.

We describe a new formulation for a hydrophilic resin, mostly composed of glycol methacrylate and hydroxypropyl methacrylate and here referred to as bioacryl, that allows the performance of morphological and immunohistochemical investigations at both light and electron microscopic levels. Immunolocalizations performed on bioacryl-embedded tissues are characterized by high specificity with virtually absent background staining. Finally, the new resin yields satisfactory fine-structural preservation, resulting in ultrastructural images of better quality than those obtained with Lowicryl K4M.

Lysozyme localization in normal and diseased human gastric and colonic mucosa. A correlative histochemical, immunohistochemical and immunoelectron microscopic investigation.

The distribution of lysozyme in normal and pathological human gastric and colonic mucosa was studied by light and electron microscopic immunocytochemical techniques and compared with histological and histochemical features. Lysozyme was localized in pyloric glandular epithelial cells, mucous neck cells of fundic glands, Paneth cells and some crypt cells of the mature colonic mucosa. In addition, lysozyme was detected in a large spectrum of "immature" or "regenerative" epithelium: neck cells of the gastric regenerative zone, undifferentiated columnar cells of surface and hyperplastic interfoveolar crests of the stomach, regenerative cells in a healed gastric ulcer, some goblet cells in incomplete intestinal metaplasia, cells of the regenerative zone at the bottom of colonic crypts and, finally, fetal intestinal epithelium. Electron microscopically, we localized lysozyme in the central core of mucous granules in the pyloric gastric glandular epithelium and in the dense mucous granules in gastric mucous neck cells. Lysozyme was also detected in some immature mucin-producing cells of the gastric regenerative zone and in the rough endoplasmic reticulum of surface hyperplastic columnar gastric cells. At the electron microscopic level, a peculiar correlation between the immunopattern of lysozyme and the morphology of mucous granules has been postulated. All our data support and extend the view that the presence of lysozyme may be related to cell immaturity as well as to a regenerative state of the cell. Finally, the lysozyme distribution and its relation to mucosubstances in gastric and colonic carcinoma suggest that lysozyme should not be considered an exclusive marker of cells of gastric derivation.

Human immunodeficiency virus type-1 (HIV-1) infection of endothelial cells in vitro: a virological, ultrastructural and immuno-cytochemical approach.

In an attempt to better understand the role of endothelial cells during HIV-1 infection, we report a virological and ultrastructural study on isolated endothelial cells from human adipose tissue, infected by HIV-1 in vitro. Supernatants from cultures showed the presence of p24 antigen and reverse transcriptase activity starting two days after HIV inoculation. A significant decrease of viral rescue was observed in cycloheximide treated cells confirming a de novo synthesis of viral products. SEM analysis individualized several surface slender projections and interdispersed virus-like particles in the infected cells. Furthermore, transmission electron microscopy (TEM) results showed cellular aspects of HIV phagocytosis and virus budding, suggesting that endothelial cells may represent a CD4 negative cell target of HIV-1 infection.

Peroneal muscular atrophy with hereditary spastic paraparesis (HMSN V) is pathologically heterogeneous. Report of nerve biopsy in four cases and review of the literature.

Peroneal muscular atrophy (PMA) associated with hereditary spastic paraparesis (HSP) is a nosologically ill-defined disease, which has been classified by Dyck as hereditary motor and sensory neuropathy type V (HMSN V). Nerve biopsy has been rarely reported in this condition. We examined sural nerve biopsies in four patients, demonstrating the following: severe myelinated fiber loss especially of large fibers, with moderate (one case) or prominent (one case) onion bulb formation; selective decrease of large fibers with moderate Schwann cell hyperplasia (one case); normal myelinated fiber population with minimal changes (one case). After reviewing previously reported cases we, conclude that in PMA with HSP sural nerve biopsy may show features either of hypertrophic type of PMA, of neuronal type, or of spinal type; thus, it seems inappropriate to allocate PMA with HSP in a unique subtype of HMSN. In addition, HSP may be not associated with peripheral neuropathy, and thus the classification in the HMSN group may be incongruous. A proper classification of PMA with HSP may be in the "complicated" forms of HSP according to Harding [Lancet I: 1151-1155 (1983)]; however, the nosology of this condition needs to be further elucidated, possibly on the basis of the underlying molecular genetic mechanisms of HSP and PMA.

Human immunodeficiency virus type 1 infection of endothelial cells in vitro.

In order to establish whether endothelial cells are involved in immunodeficiency virus type 1 (HIV-1) infection, we performed a virological study on endothelial cells isolated from human adipose tissue and infected with HIV-1 in vitro. Supernatants from cultures showed a reverse transcriptase activity starting one day after HIV inoculation. Viral rescue was significantly impaired in cycloheximide treated cells confirming a de novo synthesis of viral products.

Conventional and high resolution scanning electron microscopy of biological sectioned material.

Intracellular structures of embedded biological tissues (rat kidney, myocardium and small intestine) were observed by conventional-scanning electron microscopy (C-SEM) and high-resolution scanning electron microscopy (HR-SEM) after glass knife sectioning. C-SEM of semi-thin sections of material processed the same way as conventional transmission electron microscopy (TEM) provided strong backscattered electron (BSE)-dependent, two-dimensional secondary electron images (SEI(-)) which precisely integrated and further extended previous light microscopy (LM) observation of the same specimen. In addition, the three-dimensional (3-D) arrangement of intracellular organelles was appreciated using a mixture of acetone-soluble acrylic resin in place of epoxy resin embedding. Since the identification of such structures was hampered by the use of conventional fixations we introduced osmium maceration as a preliminary step to remove excess cytoplasmic matrix from the specimen. Consequently, semi-thin sections for LM and thin sections for TEM were obtained by sectioning of the tissue blocks. After resin removal, the sections were successfully observed in 3-D under a C-SEM. Finally, the deembedded, osmium treated sections proved to be smooth enough to facilitate deposition of continuous, ultra-thin (1 nm) chromium films and, therefore, HR-SEM studies of macromolecular cell membrane structures.