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PubMed1992

Histochemical and immunocytochemical characterizations of laminated bodies in the pancreas acinar lumen.

Combined adrenalectomy and castration cause numerous morphological alterations in the exocrine pancreas of the rat. Accumulation of laminated bodies (LB) in the acinar lumen is one of these alterations. A series of classical histological stains was applied to identify the components of these structures. A positive reaction was observed with periodic acid-Schiff, Congo red, and Alcian blue (pH 2.5). The first stain reacts with neutral and some acid mucosubstances and the second with amyloid, whereas the last one reacts with sulfated and nonsulfated acid mucosubstances. The LB also responded to Luxol fast blue, indicating the presence of lipids, an observation that is in agreement with the osmiophilic properties of these structures. A more specific identification of LB components was carried out with the immunocytochemical protein A-gold technique. Presence of gamma-glutamyltranspeptidase (gamma-GT) and GP2, two glycoproteins known to be secreted by the pancreas, was tested. The gamma-GT was associated with LB whereas GP2 was found in the lumen but not associated with these structures. Amylase was undetectable when LB occupied the lumen, suggesting that the process leading to production of LB also blocks secretory activity. To determine if diet influences LB accumulation in the pancreas acinar lumen, their frequency was compared in rats fed Purina Lab Chow or a lipid-free synthetic diet. A significant increase was observed in castrated-adrenalectomized rats fed the latter diet. This increase corresponded to a pronounced reduction in the number of zymogen granules (ZG) in the acinar cell. Our results show that LB are made of lipids, neutral mucosubstances, and nonsulfated acid mucosubstances, and that hormonal (steroids) and dietary factors (lipids) influence their accumulation.

Adrenalectomy↗
PubMed2004

Potential virulence-associated properties of Plesiomonas shigelloides strains.

Serotyping and some potential virulence-associated markers were investigated in Plesiomonas shigelloides strains isolated from humans, animals and aquatic environments. Surface properties of these strains were evaluated using Congo red binding, salt-aggregation test, bacterial adherence to xylene and motility. Production of pancreatic elastase, proteinase (consistent with subtilisin Carlsberg), triacylglycerol lipase, histidine decarboxylase and beta-hemolysin was also determined. In addition, detection of signal molecules such as C4-C8 unsubstituted N-acylhomoserine lactones (AHLs) was performed. The serological typing of the P. shigelloides strains showed that the isolates belonged to 13 different serovars. The majority of the strains were hydrophobic and motile. The strains produced low levels of elastase, proteinase and histidine decarboxylase whereas triacylglycerol lipase activity was relatively high. Only 23.3 % of the strains produced hemolysin. The AHLs signal molecules were not detected. P. shigelloides strains were able to produce a variety of potential virulence markers which may be involved in the pathogenesis of Plesiomonas-associated infections.

4-Butyrolactone↗
PubMed1999

[An unusual cause of hepatorenal symptoms].

CASE REPORT: A 65-year-old patient with normal blood pressure had an exclusive elevation of the cholestasis enzymes (alkaline phosphatase 297 U/l, gamma-GT 315 U/l) and elevated bilirubin levels (1.4 mg/dl) since August 1994. A biopsy of the liver in March 1995 showed features of a "subacute viral hepatitis"; DD drug-induced or toxic lesions. Serological tests gave no support for an acute hepatitis. Intra- or extrahepatic cholestasis could not be proved neither by ultrasound nor by an endoscopic retrograde cholangiopancreatography. Since November 1995 serum creatinine increased up to 1.7 mg/dl (March 1995 1.1 mg/dl) and proteinuria (2.1 g/d) developed. Due to worsening of renal function (serum creatinine 2.8 mg/dl) and increasing proteinuria (3.5 g/d) without nephrotic syndrome, a kidney biopsy was performed. Histologically an amyloidosis (type A lambda) was proven, involving glomerula, kidney vessels and tubules. Further biopsies from the stomach and the duodenum showed profound infiltration of the mucosa and submucosa with amyloid. Therefore, staining of the liver biopsy of March 1995 with congo red proved the diagnosis of liver amyloidosis. By a punch biopsy of the iliac crest a low-grade non-Hodgkin's lymphoma could be identified as the cause for this generalized amyloidosis. DISCUSSION: In the present case, the reason for these unusual hepatorenal symptoms with unclear cholestasis over years as the first clinical symptom and a succeeding progressive renal insufficiency with proteinuria could be found by the use of kidney biopsy and extending the analysis of a liver sample taken by biopsy 1 year ago. Immunoglobulin light chains produced by a low-grade non-Hodgkin's lymphoma caused a generalized amyloidosis type A lambda. CONCLUSION: As a consequence, by an occurrence of unusual hepatorenal symptoms with cholestasis and progressive renal failure, amyloidosis should be considered as a pathogenetic factor.

Aged↗
PubMed2002

Cellulase genes from the parabasalian symbiont Pseudotrichonympha grassii in the hindgut of the wood-feeding termite Coptotermes formosanus.

Cellulase genes of Pseudotrichonympha grassii (Hypermastigida: Eucomonymphidae), the symbiotic flagellate in the hindgut of the wood-feeding termite Coptotermes formosanus, were isolated and characterized. The nucleotide sequences of the major cellulase component in the hindgut of C. formosanus were determined based on its N-terminal amino acid sequence. The five isolated nucleotide sequences (PgCBH-homos) had an open reading frame of 1350 bp showing similarity to catalytic domains of glycoside hydrolase family (GHF) 7 members, and primary structure comparison with GHF7 members whose tertiary structures are well-characterized revealed the overall similarity between PgCBH-homo and the catalytic domain of a processive cellulase Cel7A (formerly CBHI) from the aerobic fungus Trichoderma reesei. Functional expression of PgCBH-homos in Escherichia coli, using the carboxymethylcellulose-Congo red assay, demonstrated the actual cellulolytic activity of PgCBH-homo. RT-PCR showed that PgCBH-homos were expressed, from the three flagellates in the hindgut, specifically in P. grassii.

Amino Acid Sequence↗
PubMed2005

Modelling of amyloid beta-peptide induced lesions using roller-drum incubation of hippocampal slice cultures from neonatal rats.

Pronounced neurodegeneration of hippocampal pyramidal neurons has been shown in Alzheimer's disease. The aim of this study was to establish an organotypic in vitro model for investigating effects of the amyloid beta (Abeta)-peptide on pyramidal neuron degeneration, glial cell activation and tau phosphorylation. Tissue cultures in a quasi-monolayer were obtained using roller-drum incubation of hippocampal slices from neonatal Sprague Dawley rats. Neuronal populations identified included N-methyl-D-aspartate (NMDA-R1) receptor immunoreactive pyramidal neurons, and neurons immunopositive for glutamic acid decarboxylase-65 (GAD65) or gamma amino butyric acid (GABA). Many neurons expressed phosphorylated tau as shown by pS(396), AD2 and PHF-tau immunostaining. Astrocytes, microglial cells and macrophages were also identified. The Abeta(25-35) peptide formed fibrillar networks within 2 days as demonstrated by electron microscopy. In the presence of the neurotoxic Abeta(25-35) peptide, but not Abeta(35-25), deposits developed in the tissue that were stainable with Thioflavine T and Congo red and showed the characteristic birefringence of Abeta plaques. Following Abeta(25-35) exposure, neurodegenerative cells were observed with Fluoro-Jade B staining. Further characterization of pyramidal neurons immunopositive for NMDA-R1 showed a decrease of cell number in the immediate surrounding of Abeta(25-35) deposits in a time- and concentration-dependent fashion. Similar effects on pyramidal neurons were obtained following exposure to the full-length, Abeta(1-40) peptide. Also, a loss of neuronal processes was seen with GAD65, but not GABA, immunohistochemistry after exposure to Abeta(25-35). Abeta(25-35)-exposed neurons immunopositive for phospho-tau showed degenerating, bent and often fragmented processes. Astrocytes showed increased GFAP-positive reactivity after Abeta(25-35) exposure and formation of large networks of processes. No obvious effect on microglial cells and macrophages could be seen after the Abeta(25-35) exposure. The developed in vitro system may constitute a useful tool for screening novel drugs against Abeta-induced alterations of tau and degeneration of hippocampal neurons.

Amyloid beta-Peptides↗
PubMed2000

Fibrobacter succinogenes S85 ferments ball-milled cellulose as fast as cellobiose until cellulose surface area is limiting.

Fibrobacter succinogenes S85 grew rapidly on cellobiose (0.31 h(-1) and the absolute rate of increase in fermentation acids was 0.68 h(-1). Cultures that were provided with ball-milled cellulose initially produced fermentation acids and microbial protein as fast as those provided with cellobiose, but the absolute cellulose digestion rate eventually declined. If the inoculum size was increased, the kinetics decayed from first to zero order (with respect to cells) even sooner, but in each case the absolute rate declined after only 20 to 30% of the cellulose had been fermented. Congo red binding indicated that the cellulose surface area of individual cellulose particles was not decreasing, and the transition of ball-milled cellulose digestion corresponded with the appearance of unbound cells in the culture supernatant. When bound cells from partially digested cellulose were removed and the cellulose was re-incubated with a fresh inoculum, the initial absolute fermentation rate was as high as the one observed for undigested cellulose and cellobiose. Based on these results, cellulose digestion by F. succinogenes S85 appears to be constrained by cellulose surface area rather than cellulase activity per se.

Animals↗
PubMed2000

Bilateral metachronous periosteal tibial amyloid tumors.

Localized primary periosteal amyloid tumors are extremely rare. A case of bilateral tibial amyloid tumor is presented. A 62-year-old woman initially presented with a painful mass in the anterior aspect of the right leg. There was no evidence of underlying systemic disease, including chronic infection or malignancy. Based on the results of resistance with Congo red staining to treatment with potassium permanganate and positivity for kappa light chain, we classified this particular case as AL-type amyloidosis. The patient noticed a swelling in the opposite leg 2 years later. The second tumor was also an AL-type amyloidoma. Amyloid tumors are generally solitary. This is the first case of bilateral periosteal amyloid tumors of the AL-type occurring in the tibiae.

Amyloidosis↗
PubMed2006

Xanthan gum: an economical partial substitute for agar in microbial culture media.

Xanthan gum, microbial desiccation-resistant polysaccharide prepared commercially by aerobic submerged fermentation from Xanthomonas campestris, has been successfully used alone and in combination with agar for microbial culture media. As illustrative examples, eight bacteria and eight fungi were grown on media solidified with either agar (A, 1.5%), xanthan gum (X, 1%), or combinations of both (0.9% X + 0.1% A, 0.8% X + 0.2% A, 0.7% X + 0.3% A, 0.6% X + 0.4% A). All fungi and bacteria exhibited normal growth and differentiation in all these treatments. Rather, growth of most of the fungi was better on xanthan (alone) and xanthan + agar media than agar medium. As the media gelled with xanthan gum alone flow, it was not possible to incubate Petri plates in inverted position. Moreover, because of the softness, streaking of bacteria was difficult on such media. However, these problems could be overcome by partially replacing xanthan gum with 0.3% agar. Bacterial enumeration studies carried out for Serratia sp. and Pseudomonas sp. by serial dilution and pour-plate method on agar (1.5%), 0.7%/0.6% X + 0.3%/0.4% A yielded similar counts. Selective media, succinate medium for Pseudomonas sp., and MacConkey broth medium for Escherichia coli gelled with 0.7%/0.6% X + 0.3%/0.4% A did not support growth of other bacteria when inoculated along with the above-mentioned bacteria. Likewise, differential medium, CRMA (Congo red mannitol agar) gelled with xanthan-agar combination could differentiate between Agrobacterium tumefaciens and Rhizobium sp.

Agar↗
PubMed1998

[Amyloid: microscopic demonstration, classification and clinical correlation].

Amyloid is a biophysically defined, biochemically non-uniform protein which is deposited in the tissue, forming a cross-beta configuration. In paraffin sections it is demonstrated using Congo red staining according to Puchtler et al. and a polarizing microscope with a tension-free clean optic system that shows a typical apple-green birefringence. The identification of amyloid has to be followed by immunostaining for AA, ATTR, A lambda, A kappa and A beta 2 microglobulin and possibly further types of amyloids for classification purposes. Considering the localization, expansion and type localized and generalized types probably can be differentiated. The latter should be correlated to a basic disease. The treatment of the basic disease is the only chance for reducing or eliminating the amyloid deposits. In this review, the modern morphological methods for demonstration and both immunohistological and clinical classification of amyloid and amyloidoses are presented. Rare types are mentioned in the tables.

Amyloid↗
PubMed1999

Nuclear inclusions in spinocerebellar ataxia type 1.

Spinocerebellar ataxia type 1 is due to a CAG repeat expansion in the gene encoding ataxin-1. In a case with an expansion of 56 repeats, intranuclear inclusions were found only in neurons, both in severely affected regions (such as the pons) and in areas where the lesions were inconspicuous (such as the cortex or the striatum). The inclusions were labelled by a monoclonal antibody directed against long polyglutamine stretches (1C2); they were also detected by the anti-ubiquitin antibody. They were faintly eosinophilic, Congo red negative and were not stained by thioflavin S or by ethidium bromide.

Adult↗
PubMed2003

Endoscopic management of laryngo-tracheobronchial amyloidosis: a series of 32 patients.

Between June 1983 and December 2002, 32 patients were treated for primary localized laryngo-tracheobronchial amyloidosis (LTBA) at our institution. For enrollment in this retrospective study, at least one positive biopsy with Congo red stain and a diagnostic test battery excluding systemic or secondary amyloidosis were mandatory. The most common presenting symptom was dyspnea, which was observed in 75% of cases. Endoscopic appearance was described as submucosal plaques and nodules with a cobblestone appearance in 44% of patients, tumor-like in 28% and circumferential wall thickening in 28% of cases. Tracheobronchopatia osteochondroplastica was associated with LTBA in 22% of patients, both at the first treatment (four cases) or during follow-up examinations (three). Synchronous or metachronous disease in the larynx and tracheobronchial tree was observed in 47% of cases, while only one patient showed lung involvement with extensive amyloidosis of the trachea and bronchi. All but two patients were endoscopically treated. One of these was affected by a tracheal amyloidoma cured by endoscopic debulking and further tracheal resection-anastomosis. The other patient presented massive laryngo-pharyngeal involvement and was therefore treated by total laryngectomy. A total of 58 endoscopic procedures (range: 1-7; mean: two per patient) were carried out, 11 with CO2 laser (for supraglottic, glottic and selected subglottic lesions) and 47 with Nd:YAG laser (for the most part of subglottic lesions and amyloid deposits located in trachea and bronchi). Four patients required a tracheotomy and three necessitated a T-tube stent during management, which was subsequently removed in all cases. One patient was lost to follow-up, nine were endoscopically controlled elsewhere (Group A) and 22 were followed at our institution (Group B). In Group A, the patient with tracheal amyloidoma is asymptomatic and free of additional LTBA localizations 8 years after surgery. The remaining eight are asymptomatic, but with persistent endoscopic signs of amyloidosis. In Group B, five patients died: one from heart failure and another one from pneumonia 3 and 4 days after surgery, respectively. In one case death was from unrelated causes, and in the two remaining patients it was from respiratory failure due to uncontrolled bilateral bronchial amyloidosis in the subsequent 5 to 6 years. One patient is completely cured 5 years after surgery, and 16 are asymptomatic with persistence of LTBA.

Adult↗
PubMed2005

Keratoepithelin in secondary corneal amyloidosis.

BACKGROUND: Amyloid is found in several corneal dystrophies, including distinct lattice corneal dystrophies (LCD) and Avellino corneal dystrophy. Recently, point mutations in the transforming growth factor-beta-induced gene (TGFBI) encoding for keratoepithelin (KE) have been demonstrated in these corneal disease entities. We intended to investigate if KE was also a component of the rarely seen secondary corneal amyloid deposits. METHODS: Immunohistochemical staining with a polyclonal antibody against KE was performed on formalin-fixed paraffin-embedded tissue of five corneal buttons with secondary amyloid obtained after keratoplasty. Secondary amyloidosis was due to Fuchs endothelial dystrophy (FED) with bullous keratopathy and/or recurrent erosions in all cases. The diagnosis had been established by light microscopy using Congo red staining. Two cases of LCD type I served as positive controls and three corneas with FED and one with keratoconus without amyloid served as negative controls. RESULTS: All corneas with secondary amyloidosis as well as LCD type I revealed positive staining in the respective amyloid deposits. KE was localized in the subepithelial pannus and in the anterior stroma in the corneas with secondary amyloidosis. In the specimens with LCD type I it was distributed in the amyloid deposits located in the anterior and mid-stroma. Staining for KE showed a granular appearance in all cases. The intensity of staining was variable among the specimens. CONCLUSIONS: KE is found not only in primary amyloid deposits of hereditary corneal dystrophies, but also in secondary amyloidosis of the cornea of diverse ethiologies.

Adult↗
PubMed2001

Analysis of aqueous humour proteins of eyes with and without pseudoexfoliation syndrome.

UNLABELLED: Pseudoexfoliation syndrome (PEX) has been suggested to represent a blood-aqueous barrier impairment leading to a higher protein content in aqueous humour of eyes with PEX. However, the nature of a prospective PEX protein has not yet been described. We set out to reevaluate protein content and examine protein composition for prospective PEX protein candidates in aqueous humour of eyes with PEX syndrome. Aqueous humour of 52 patients with PEX and 38 without PEX signs was sampled during cataract or glaucoma surgery. Total aqueous protein concentration in the samples was analysed in 43 PEX specimens and 32 non-PEX specimens according to Bradford. Aqueous protein composition of all samples was determined by sodium dodecylsulphate polyacrylamide gel electrophoresis (SDS PAGE) and silver staining. Screening for amyloids was performed in nine PEX samples and six non-PEX samples by Congo Red staining and polarised light microscopy. Aqueous protein concentration was not significantly increased in PEX eyes in comparison with non-PEX eyes. Furthermore, we could not detect any characteristic difference in protein band sizes of the two groups after SDS PAGE. However, we were able to show the presence of amyloid exclusively in aqueous humour of PEX patients. CONCLUSION: our results do not confirm a generally higher protein concentration in pseudoexfoliation syndrome eyes. This does not necessarily contradict a blood-aqueous barrier impairment but illustrates the variance in protein concentration between and within the two groups. No characteristic protein band allocatable to pseudoexfoliation syndrome proteins could be detected in any of the samples. However, our findings support the theory that the pseudoexfoliation syndrome is associated with an amyloid of a serum protein.

Aged↗
PubMed2004

Novel sporophyte-like plants are regenerated from protoplasts fused between sporophytic and gametophytic protoplasts of Bryopsis plumosa.

Protoplasts of the marine coenocytic macrophyte Bryopsis plumosa (Hudson) C. Agardh. [Caulerpales] can easily be obtained by cutting gametophytes or sporophytes with sharp scissors. When a protoplast isolated from a gametophyte was fused with a protoplast isolated from a sporophyte of this alga, it germinated and developed into either one of two completely different forms. One plant form, named Type G, appeared quite similar to a gametophyte, and the other, named Type S, looked similar to a sporophyte. While the Type G plant contained many small nuclei of gametophyte origin together with a single giant nucleus of sporophyte origin, the Type S plant contained many large nuclei of uniform size. These large nuclei in the Type S plant had metamorphosed from the gametophytic nuclei, and were not formed through division of the giant nucleus of sporophyte origin. Fragments of the Type S plant, each having such a large nucleus, developed into creeping filaments that look very similar to sporophytes. While cell walls of gametophytes and Type G plants were stained by Congo-red, those of the thalli of regenerated Type S plants and sporophytes were not stained by the dye. This indicated that the large nuclei of the Type S plant did not express genes for xylan synthesis, which are characteristic of gametophytes. Two-dimensional gel electrophoretic analysis revealed that most of the proteins synthesized in the Type S plant were identical to those of sporophytes. These results strongly suggest that in the Type S plant, the gametophytic nuclei are transformed into sporophyte-like nuclei by an unknown factor(s) produced by the giant nucleus of sporophyte origin and that the transformed nuclei express the set of genes characteristic of sporophytes. Despite morphological similarity, however, the regenerated Type S plant could not produce zoospores, because its large nuclei did not divide normally. The transformed large nuclei of gametophyte origin still seemed to be in the haploid state.

Cell Fusion↗
PubMed2002

Pathology, diagnosis and pathogenesis of AA amyloidosis.

Amyloid is defined as a proteinaceous tissue deposit that shows a typical green birefringence in polarised light after staining with Congo red, the presence of non-branching linear fibrils of indefinite length with an approximate diameter of 10-12 nm and a distinct X-ray diffraction pattern consistent with Pauling's model of a cross-beta fibril. Approximately 45% of generalised amyloidoses are secondary or reactive (AA) amyloidosis. Among the causes of AA amyloidosis are rheumatic diseases, idiopathic diseases, inherited diseases, infectious diseases and malignant tumours. Recent decades have provided significant advances in our understanding of the pathology and pathogenesis of AA amyloidosis. Its pathogenesis is multifactorial involving many variables such as primary structure of the precursor protein, acute phase response, the presence of non-fibril proteins (e.g. amyloid P component, apolipoprotein E, glycosaminoglycans, proteoglycans and basement membrane proteins), receptors, lipid metabolism and proteases. Study of the pathogenesis of AA amyloidosis has provided many insights into the nature of conformational diseases, which may help in the understanding of other members of this particularly heterogeneous group of diseases, such as Alzheimer's disease and transmissible spongiform encephalopathies.

Amyloidosis↗
PubMed2005

Heterogeneity in distribution of amyloid-positive islets in type-2 diabetic patients.

Amyloid-containing (A+) islets are characteristic for type-2 diabetes (T2D), but their abundance seems variable among patients. It is unclear whether the distribution of A+ islets follows a certain pattern or occurs randomly throughout the pancreatic organ. We investigated the topography of A+ islets in eight pancreata of T2D patients and eight sex- and age-matched non-diabetic subjects. Transversal sections of head, body and tail segments were stained with synaptophysin combined with Congo red to map/quantify islet tissue and amyloid. In the eight T2D pancreata, the overall percentage of A+ islets varied from 4% to 85%. Further analysis in body and tail indicated that peripheral regions exhibited higher percentages of A+ islets than central regions (averages of, respectively, 30% and 17%, P<0.05). Non-diabetic control pancreata also exhibited A+ islets, albeit at a 25-fold lower frequency; a tendency towards higher percentage of A+ islets in peripheral versus central regions was also observed. The higher percentage A+ islets in peripheral regions was associated with a higher density and relative islet over exocrine surface area. These observations on heterogeneity in abundance and distribution of A+ islets need consideration when sampling tissue for studies on human islet amyloidosis. The present methodology allows us to further investigate the susceptibility to amyloidosis of islets in peripheral regions of the pancreas.

Aged↗
PubMed1998

Amyloid tumours in the soft tissues of the legs. Case report and review of the literature.

We present a case of multiple amyloidomas occurring in the calves of a 61-year-old woman, without systemic amyloidosis or plasma cell dyscrasia. The disappearance of Congo red positivity after potassium permanganate treatment and immunohistochemical results showed that this was a case of reactive AA amyloidosis. True soft tissue amyloidomas are extremely rare, and this is the first case of AA amyloidoma in the soft tissues of the legs.

Amyloid↗
PubMed1998

Basement membrane proteins, apolipoprotein E and glycosaminoglycans in pituitary adenomas and their correlation to amyloid.

The aim of the study was to confirm earlier reports of the association of amyloid deposits in growth hormone (GH)-producing adenomas of the pituitary and the presence of glycosaminoglycans, basement membrane proteins and apolipoprotein E (apo E). Serial sections from 17 amyloidotic and 11 nonamyloidotic, sparsely granulated, GH-producing adenomas obtained from patients presenting with acromegaly were stained with Congo red and Alcian blue, and also with antisera directed against fibronectin, collagen IV, laminin and apo E. Glycosaminoglycans were found in capillaries of every adenoma and were also related spatially to amyloid deposits. Immunostaining of both nonamyloidotic and amyloidotic adenomas demonstrated the presence of fibronectin, collagen IV and laminin in the basement membranes of surrounding nonadenomatous tissue and tumour vessels. In approximately half the amyloidotic adenomas, each basement membrane protein presented with a distinct spatial relationship to amyloid deposits. Apo E was found in 88% of the amyloidotic adenomas within the amyloid deposits, and in six cases intracellular immunostaining was also evident in folliculo-stellate cells. The results are consistent with the presence of glycosaminoglycans, basement membrane proteins and apo E in the amyloid deposits of pituitary adenomas.

Adenoma↗