The relation of the properties of Congo red-stained amyloid fibrils to the -conformation.
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Amyloid↗
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Eyelid bleeding and atypical amyloidosis.
PURPOSE: To describe a case of atypical systemic amyloidosis in a patient who presented with recurrent subcutaneous bleeding of the eyelids and auricles. DESIGN: Observational case report. METHODS: A 52-year-old man with chronic cardiac failure, hypertension, weight loss, and impotence presented with recurrent bleeding of the eyelids and auricles, and was investigated for suspected amyloidosis. RESULTS: Biopsy specimens taken from the bone marrow, salivary gland, abdominal subcutaneous fat, and gastric mucosa, and eyelids were all negative for Congo red. Results of a conjunctival biopsy, which were returned after the patient's death, and cardiac muscle autopsy were positive for Congo red staining, indicating amyloidosis. CONCLUSION: The findings in this case show that the presentation of amyloidosis may be atypical. Ophthalmologists should consider a diagnosis of amyloidosis in patients with periorbital bleeding, even if biopsies are negative for Congo red. A conjunctival biopsy may be useful in establishing a diagnosis of primary systemic amyloidosis.
Amyloidosis↗
Purification and properties of an acidic protein from rat skin.
An acidic protein, extractable in neutral salt solutions from rat skin, was markedly enriched when precipitated by dialysis against 0.5 M acetic acid. After dissolving the precipitate in 0.5 M Tris-HCl buffer, pH 8.0, the protein was disaggregated by the addition of the nonionic detergent Triton X-100 and purified by chromatography on Sephadex G-100 and DEAE-Sephadex A-50 columns. The protein isolated under nondenaturing conditions appeared to be essentially homogeneous by its migration as a single band on (a) cellulose acetate membrane electrophoresis at pH 8.6; (B) 4% and 7.5% polyacrylamide gel electrophoresis at ph 8.9; (C) sodium dodecyl sulfate (10%) polyacrylamide gel electrophoresis at pH 7.0; and by (d) its complete freedom from collagen, the major contaminating protein. The molecular weight of the protein was determined as 76,000 +/- 2,000 from its electrophoretic mobility in sodium dodecyl sulfate polyacrylamide gels and 75,000 from its elution volume in Sephadex G-100 columns. Reduction and alkylation of the protein failed to generate smaller subunits. The amino acid composition of the protein showed that it was relatively rich in glutamic and aspartic acids, which together comprised 25% of its total residues. Hydrophobic amino acids like phenylalanine, leucine, isoleucine, valine, methionine, alanine, proline, and cystine accounted for about 34% of the total residues in the protein. No free NH2-terminal amino acid could be detected in the purified protein by the dansylation method. Each mole of protein contained 11 mol of phosphate. Triton X-100 was necessary for achieving nondestructive disaggregation of the acidic protein. Each mole of protein bound about 3200 mol of Triton X-100 or 10 mol of Congo red. While the detergent binding could be reversed by dialysis, Congo red formed a stable complex with the protein.
Amino Acids↗
The global transcriptional response to transient cell wall damage in Saccharomyces cerevisiae and its regulation by the cell integrity signaling pathway.
In the yeast Saccharomyces cerevisiae, environmental stress conditions that damage the cell wall lead to activation of the so-called "compensatory mechanism," aimed at preserving cell integrity through a remodeling of this extracellular matrix. Here we used DNA microarrays to investigate the molecular basis of this response to two agents that induce transient cell wall damage; namely Congo Red and Zymolyase. Treatment of the cells with these two agents elicited the up-regulation of 132 and 101 genes respectively, the main functional groups among them being involved in cell wall construction and metabolism. The main response does not occur until hours after exposure to the cell wall-perturbing agent. In some cases, this response was transient, but more sustained in others, especially in the case of the genes involved in cell wall remodeling. Clustering of these data together with those from the response to constitutive cell wall damage, revealed the existence of a cluster of co-regulated genes that was strongly induced under all conditions assayed. Those genes induced by cell wall damage showed an enrichment in DNA binding motifs for Rlm1p, Crz1p, SBF (Swi4p/Swi6p), Msn2p/Msn4p, Ste12p, and Tec1p transcription factors, suggesting a complex regulation of this response together with the possible involvement of several signaling pathways. With the exception of PHO89 and FKS2, none of the genes induced by Congo Red was up-regulated in a slt2 strain. Moreover, characterization of the transcriptional response to Congo Red in a rlm1 mutant strain revealed that only a few genes (i.e. PHO89, FKS2, YLR042C, and CHA1) were induced at least partially independently of the transcription factor Rlm1p, the rest being totally dependent on this transcription factor for their activation. Our findings consistently demonstrate that the cell integrity signaling pathway regulates the cell wall damage compensatory response, mainly through transcriptional activation mediated by Rlm1p.
Alleles↗
Lactoferrin Glu561Asp facilitates secondary amyloidosis in the cornea.
AIM: To elucidate the pathogenic mechanism of amyloid formation in corneal amyloidosis with trichiasis. METHODS: Ophthalmological examination was performed in nine patients to determine secondary corneal amyloidosis with trichiasis. Congo red staining and immunohistochemistry using anti-human lactoferrin antibody were used for biopsied corneal samples. For genetic analyses, single strand conformation polymorphism (SSCP), direct DNA sequence analysis, and polymerase chain reaction (PCR) induced mutation restriction analysis (IMRA) were employed to detect lactoferrin gene polymorphism. RESULTS: All patients had had trichiasis at least for 1 year, and all amyloid-like deposits were found in one eye with trichiasis. Ophthalmological examination revealed that eight patients showed gelatinous type of amyloid deposition and one showed lattice type of amyloid deposition. Studies of biopsied corneal samples with Congo red stain revealed positive staining just under the corneal epithelial cells. Immunoreactivity of anti-human lactoferrin antibodies was recognised in all tissues with positive Congo red staining. Lactoferrin gene analysis revealed that seven patients were heterozygotic and two were homozygotic for lactoferrin Glu561Asp. The frequency of the polymorphism in the patients was significantly different from that in 56 healthy control subjects. CONCLUSION: Lactoferrin Glu561Asp is a key polymorphism related to facilitating amyloid formation in corneal amyloidosis with trichiasis.
Adolescent↗
Number of Abeta inoculations in APP+PS1 transgenic mice influences antibody titers, microglial activation, and congophilic plaque levels.
There have been several reports on the use of beta-amyloid (Abeta ) vaccination in different mouse models of Alzheimer's disease (AD) and its effects on pathology and cognitive function. In this report, the histopathologic findings in the APP+PS1 doubly transgenic mouse were compared after three, five, or nine Abeta inoculations. The number of inoculations influenced the effects of vaccination on Congo red levels, microglia activation, and anti-Abeta antibody titers. After three inoculations, the antibody titer of transgenic mice was substantially lower than that found in nontransgenic animals. However, after nine inoculations, the levels were considerably higher in both genotypes and no longer distinguishable statistically. The number of inoculations influenced CD45 expression, an indicator of microglial activation. There was an initial upregulation, which was significant after five inoculations, but by nine inoculations, the extent of microglial activation was equivalent to that in mice given control vaccinations. Along with this increased CD45 expression, there was a correlative reduction in staining by Congo red, which stains compact plaques. When data from the mice from all groups were combined, there was a significant correlation between activation of microglia and Congo red levels, suggesting that microglia play a role in the clearance of compact plaque.
Alzheimer Disease↗
A quantitative method for detecting deposits of amyloid A protein in aspirated fat tissue of patients with arthritis.
OBJECTIVE: To describe a new, quantitative, and reproducible method for detecting deposits of amyloid A protein in aspirated fat tissue and to compare it with smears stained with Congo red. METHODS: After extraction of at least 30 mg of abdominal fat tissue in guanidine, the amyloid A protein concentration was measured by a monoclonal antibody-based sandwich ELISA. RESULTS: The concentrations in 24 patients with arthritis and AA amyloidosis (median 236, range 1.1-8530 ng/mg tissue) were higher (p < 0.001) than in non-arthritic controls, uncomplicated rheumatoid arthritis, and other types of systemic amyloidosis (median 1.1, range 1.1-11.6 ng/mg tissue). Patients with extensive deposits, according to Congo red staining, had higher concentrations than patients with minute deposits. CONCLUSION: This is a new, quantitative, and reproducible method for detecting deposits of amyloid A protein in aspirated fat tissue of patients with arthritis, even when minute deposits are present as detected in smears stained with Congo red.
Abdomen↗
Amphotericin B binds to amyloid fibrils and delays their formation: a therapeutic mechanism?
The membrane-active antifungal agent amphotericin B (AmB) is one of the few agents shown to slow the course of prion diseases in animals. Congo Red and other small molecules have been reported to directly inhibit amyloidogenesis in both prion and Alzheimer peptide model systems via specific binding. We propose that it is possible that AmB may act similarly to physically prevent conversion of the largely alpha-helical prion protein (PrP) to the pathological beta-sheet aggregate protease-resistant isoform (PrP(res)) in prion disease and by analogy prevent fibrillization in amyloid diseases. To assess whether AmB is capable of binding specifically to amyloid fibrils as does Congo Red, we have used the insulin fibril and Abeta 25-35 amyloid model fibril system. We find that AmB does bind strongly to both insulin (K(d) = 1.1 microM) and Abeta 25-35 amyloid (K(d) = 6.4 microM) fibrils but not to native insulin. Binding is characterized by a red-shifted AmB spectrum indicative of a more hydrophobic environment. Thus AmB seems to have a complementary face for amyloid fibrils but not the native protein. In addition, AmB interacts specifically with Congo Red, a known fibril-binding agent. In kinetic fibril formation studies, AmB was able to significantly kinetically delay the formation of Abeta 25-35 fibrils at pH 7.4 but not insulin fibrils at pH 2.
Alzheimer Disease↗
Decolourization of azo dyes and a dye industry effluent by a white rot fungus Thelephora sp.
A white rot fungus Thelephora sp. was used for decolourization of azo dyes such as orange G (50 microM), congo red (50 microM), and amido black 10B (25 microM). Decolourization using the fungus was 33.3%, 97.1% and 98.8% for orange G, congo red and amido black 10B, respectively. An enzymatic dye decolourization study showed that a maximum of 19% orange G was removed by laccase at 15 U/ml whereas lignin peroxidase (LiP) and manganese dependent peroxidase (MnP) at the same concentration decolourized 13.5% and 10.8%, orange G, respectively. A maximum decolourization of 12.0% and 15.0% for congo red and amido black 10B, respectively, was recorded by laccase. A dye industry effluent was treated by the fungus in batch and continuous modes. A maximum decolourization of 61% was achieved on the third day in the batch mode and a maximum decolourization of 50% was obtained by the seventh day in the continuous mode. These results suggest that the batch mode of treatment using Thelephora sp. may be more effective than the continuous mode for colour removal from dye industry effluents.
Azo Compounds↗
A microenvironmental probe of elastin. Properties of a solubilized Congo red-elastin complex.
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Amino Acids↗
Postantibiotic effects and postantibiotic sub-MIC effects of ciprofloxacin, pefloxacin and amikacin on the biological properties of Salmonella strains.
The postantibiotic effect (PAE) and the postantibiotic sub-MIC effect (PASME) of ciprofloxacin, pefloxacin and amikacin were studied for Salmonella typhimurium and S. enteritidis strains. PAE was induced by 2 x and 4 x MIC of antibiotics studied for 0.5 h. After PAE and PASME their effect on prophage induction of a lysogenic S. typhimurium strain and on Congo red binding for both strains as a marker of their surface hydrophobicity was examined. The longest PAE was found after treatment with ciprofloxacin, higher values being observed with S. typhimurium. PAEs of pefloxacin and amikacin were much lower, except for the suprainhibitory concentration 4 x MIC of amikacin with S. enteritidis (6.9h). PASMEs of ciprofloxacin did not allow any regrowth of either strain. For other antibiotics the PASMEs were different while concentrations of 2 x MIC + 0.2 x MIC and 0.3 x MIC, and of 4 x MIC + 0.1 x MIC, 0.2 x MIC and 0.3 x MIC of amikacin did not allow any regrowth of S. enteritidis. PAEs of the antibiotics tested did not affect the Congo red binding by both Salmonella strains, but the PAEs of ciprofloxacin and pefloxacin expressively induced a prophage of lysogenic S. typhimurium strain. We noted the influence of Congo red binding after applying 4 x MIC + 0.1 x MIC, 0.2 x MIC and 0.3 x MIC of amikacin for S. typhimurium and 2 x MIC + 0.1 x MIC for S. enteritidis.
Amikacin↗
[Evaluation of the usefulness of selected virulence markers for the identification of virulent Yersinia enterocolitica strains. I. Phenotypic markders associated with Plasmid pYV].
The species Yersinia enterocolitica includes either pathogenic or non-pathogenic strains. Therefore it is necessary to differentiate virulent bacilli from other. It is well known that pathogenic strains of Y. enterocolitica bearing virulence associated plasmid called pYV, which could be demonstrated by its isolation or detected by the presence of specific, phenotypic properties directly related with this plasmid. The aim of the presented paper was to check the ability of some phenotypic virulence markers associated with pYV, to detection of pathogenic Y. enterocolitica strains. In the presented work 152 (130 carrying pYV) clinical strains of Y. enterocolitica O3 isolated mainly from stool were examined for the presence of phenotypic virulence markers such as: calcium dependency, Congo-red binding, autoagglutination and agglutination with Mangifera indica extract. Both first features were detected parallel, on the same plate, using CRMOX (Congo-red, Magnesium Oxalate) agar. The detection of the tested markers in the examined strains was compared with the presence of virulence plasmid. The obtained results confirmed the observations done by other authors that Y. enterocolitica strains, in which bacilli bearing the virulence plasmid predominate, exhibit all tested phenotypic properties whereas the plasmid-cured isogenic strains show no one of these features. Therefore all the tested markers could be useful for detection of virulent Y. enterocolitica strains directly isolated from patients. The most useful virulence markers in bacteriological study seems to be calcium dependency and Congo-red binding, examined together by the use of CRMOX agar, because they confirm the presence of the virulence plasmid by parallel detection of two physiologically different features associated with this plasmid. In addition CRMOX agar allows for the examination rough strains while agglutination tests do not.
Bacterial Proteins↗
Mutagenicity of selected sulfonated azo dyes in the Salmonella/microsome assay: use of aerobic and anaerobic activation procedures.
A selection of 16 sulfonated azo dyes of both the monoazo type and diazo dyes based on benzidine, o-tolidine and o-dianisidine were assayed for mutagenicity in Salmonella typhimurium strains TA98 and TA100 employing both aerobic and anaerobic preincubation procedures. 3 food dyes, FD & C Red No. 40 and Yellows No. 5 and No. 6 were non-mutagenic in all tests. 5 dyes were mutagenic with aerobic treatment (trypan blue, Pontacyl Sky Blue 4BX, Congo Red, Eriochrome Blue Black B, dimethylaminoazobenzene) and 6 were mutagenic aerobically with riboflavin and cofactors (Deltapurpurin, trypan blue, Pontacyl Sky Blue 4BX, Congo Red, methyl orange, Ponceau 3R). Anaerobic preincubation involving enzymatic reduction of the dyes led to a different pattern of mutagenicity, with trypan blue giving much enhanced mutagenicity; Eriochrome Blue Black B, Pontacyl Sky Blue 4BX, Deltapurpurin and Congo Red exhibiting similar activity to aerobic preincubation; and methyl orange and Ponceau 3R yielding no mutagenicity. The results are interpreted with respect to an hypothesis involving partial reduction of the azo bond under differing degrees of aerobiosis via azo-anion radicals and hydrazo intermediates.
Aerobiosis↗
An immunoperoxidase study of senile cerebral amyloidosis with pathogenetic considerations.
Samples of human cerebral cortex were obtained from twelve autopsied patients with Alzheimer's disease or "normal" aging. Rabbit or goat anti-human antisera to the following plasma proteins: IgG, F(ab')2, Fc, kappa and lambda light chains, IgM, IgA, fibrinogen, albumin, C3, lysozyme, haptoglobin, macroglobulin, and microglobulin; antibodies to the following intracellular proteins: glial fibrillary acidic (GFA) protein, filamin, actin, non-muscle myosin, tubulin, cholinergic vesicle proteins, and neurofilament (NF) proteins were utilized in the immunoglobulin peroxidase bridge. Amyloid cores of classical or perivascular plaques and dyshoric angiopathy exhibited a strong reaction for intact IgG and for both of its light chains, moderate reactions for lysozyme, fibrinogen, albumin and IgA, and weak reactions for IgM, C3, Fc, F(ab')2, haptoglobin, macroglobulin and microglobulin. Antibodies to all three NF proteins, individually and pooled, stained dyshoric and plaque amyloid, while antibodies to other intracellular proteins did not. The coronae of classical plaques and many primitive plaques stained for GFA, but inconsistently for IgG, both light chains, lysozyme, actin, tubulin, and NF proteins. Affected vessels of three patients with Congophilic angiopathy were reactive for all plasma proteins (especially IgG, fibrinogen, and albumin) and for NF proteins. NF staining in Congophilic blood vessels, although variable, revealed a peripheral or adventitial distribution, whereas plasma proteins tended to be localized in the media of the vessel wall. The distributions of Congo red and NF positivity were often identical. Both NF and Congo red staining was sensitive to oxidation. Isolated NF proteins were Congophilic and capable of displaying apple-green birefringence. A hypothesis concerning the role of NF proteins in senile cerebral amyloid is presented.
Amyloid↗
The effects of prenatal administration of azo dyes on testicular development in the mouse: a structure activity profile of dyes derived from benzidine, dimethylbenzidine, or dimethoxybenzidine.
In mice and rats, prenatal exposure to the dye Congo red permanently reduces the number of germ cells in male and female offspring. In the current investigation, nine other dyes structurally related to Congo red were examined for developmental testicular toxicity. In this study, the structural component of the dyes responsible for the prenatal induction of germ cell aplasia was identified. We found that only benzidine-based dyes altered testicular development and caused hypospermatogenesis in mice during adulthood. Dimethyl- and dimethoxybenzidine-based dyes were without effect. Pregnant mice were dosed orally on Days 8-12 of gestation with a benzidine-, dimethylbenzidine-, or a dimethoxybenzidine-based dye and the testes of 45- to 50-day-old male offspring were examined. The testes of postpubertal male offspring exposed to the benzidine-based dyes, Congo red, diamine blue, and Chlorazol Black E, were small and contained some tubules completely devoid of germ cells, but the dimethylbenzidine-based dyes, trypan blue, Evans blue, and benzopurpurin 4B, and the dimethoxybenzidine-based dye, Chicago sky blue, did not alter testicular development in this manner. Azoic diazo component 48, a dimethoxybenzidine congener, and two other diazo dyes, naphthol blue black and Sudan III, were also without effect on the germ cells. Experiments with Chlorazol Black E (CBE) indicate that the period of susceptibility in the male fetus is limited to the period of primordial germ cell migration and division. When CBE was administered on Days 8-10 of gestation it reduced testis weight after puberty by 30%, while treatment after Day 13 did not affect testicular function.(ABSTRACT TRUNCATED AT 250 WORDS)
Abnormalities, Drug-Induced↗
Light scattering analysis of fibril growth from the amino-terminal fragment beta(1-28) of beta-amyloid peptide.
beta-Amyloid protein (beta-A/4) is the major protein component of Alzheimer disease-related senile plaques and has been postulated to be a significant contributing factor in the onset and/or progression of the disease. In the senile plaque, beta-A/4 appears as bundles of amyloid fibrils. The biological activity of beta-A/4 may be related to its state of aggregation. In this work, self-assembly, fibril formation, and interfibrillary aggregation of beta(1-28), a synthetic peptide homologous with the amino-terminal fragment of beta-A/4, were investigated. The predominant form of beta(1-28) detected by size-exclusion chromatography and polyacrylamide gel electrophoresis was apparently a tetramer which does not bind Congo red. Aggregates containing cross-beta sheet structures which bind Congo red and thioflavin T were observed at concentrations of approximately 0.3 mg/ml or greater. Concentrations of 0.5-1 mg/ml were necessary for aggregation into fibrils to be detectable by classical or quasielastic light scattering. Both fibril elongation and fibril-fibril aggregation occur over the time scale investigated. The kinetics of aggregation were much faster at physiological salt concentrations than at lower ionic strength. Ionic strength also appeared to influence the morphology of the fibril aggregates. The data indicate that sample preparation method and sample history influence fibril size and number density.
Amino Acid Sequence↗
Amyloid myopathy: clinicopathologic study of 16 cases.
Amyloid deposition in skeletal muscle is a well-recognized but rare occurrence. Sixteen such cases seen in a 17-year period (1979 to 1996) out of a total of 3,937 muscle biopsy specimens (0.004%) form this study group. Either Congo red or sulfated alcian blue stains were routinely performed in each biopsy to screen for amyloid. Patients in this study (eight men, eight women) ranged in age from 42 to 90 years (mean, 61 years) at initial presentation. The most common symptoms at presentation included weakness/fatigue (n = 10), autonomic symptoms (n = 8), and weight loss/decreased appetite (n = 7). Five patients had a concomitant malignancy (myeloma, n = 3; malignant carcinoid tumor, n = 1; melanoma, n = 1). Two patients had known hereditary forms of amyloidosis. Five patients had amyloid diagnosed on another organ biopsy (excluding peripheral nerve). Histologically, amyloid was deposited in the interstitium or perivascular region in 14 muscles and endomysial region in seven muscles. All cases were confirmed with Congo red staining (apple green birefringence) or by electron microscopic identification of fibrillary amyloid material. Scattered angular atrophic esterase-positive muscle myofibers indicative of acute denervation atrophy were seen in 14 muscles. Eight muscles showed small group atrophy, and seven showed myofiber type grouping. Scattered regenerating muscle fibers were seen in nine cases, degenerating myofibers in six, and foci of chronic endomysial and perivascular inflammation in two. Four muscles showed type II muscle fiber atrophy. A concomitant sural nerve biopsy specimen was evaluated in seven patients; all seven contained amyloid, confirmed either by Congo red staining or electron microscopic examination. In two nerves, there was a mild loss of myelinated axons; four had a moderate loss, and one, severe loss. Six of seven nerves showed predominantly axonopathic changes. In conclusion, (1) the prevalence rate of amyloid myopathy in muscle biopsy specimens was low (in this series, 0.004%); (2) only a minority of patients had multiple myeloma, and most presented with muscle weakness/fatigue or autonomic symptoms; (3) most of the muscles showed neurogenic features histologically; (4) all concomitant sural nerve biopsy specimens contained amyloid, and most showed a predominance of axonopathic changes.
Adult↗
Feline insular amyloid: histochemical distinction from secondary systemic amyloid.
Amyloid in islets of Langerhans from 48 domestic cats, one human, one non-human primate, and one raccoon was compared with secondary systemic amyloid from three domestic cats, one dog, one human, and one cow to determine affinity for Congo red dye after treatment of paraffin-embedded tissue sections with potassium permanganate and dilute sulfuric acid. Insular amyloid from all six species was resistant to pretreatment with potassium permanganate, i.e., affinity for Congo red was retained, whereas secondary systemic amyloid from all species was sensitive to the potassium permanganate pretreatment. Other stains did not distinguish between insular and secondary systemic amyloid. The potassium permanganate-Congo red staining procedure thus can be used to differentiate insular from secondary systemic amyloid in the cat and other species. The results also indicate that insular amyloid and secondary systemic amyloid are of different chemical composition and pathogenesis.
Adrenal Glands↗