Amyloid P component and the diagnosis of amyloidosis.
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Amyloidosis↗
Biomedical subjects
M B Pepys
Publications and source records attributed to M B Pepys.
Induction in mice of human light-chain-associated amyloidosis.
Primary (idiopathic) or multiple myeloma-associated amyloidosis is characterized by the deposition in tissue of monoclonal light chains or light-chain fragments (AL amyloidosis). In contrast to other types of amyloidosis, information regarding the pathogenesis of light-chain-related amyloid has heretofore been limited due to the lack of a suitable in vivo model. The authors report the successful experimental induction of human AL amyloid deposits. The repeated injection into mice of Bence Jones proteins obtained from two patients with AL amyloidosis produced the histopathologic lesions characteristic of this disease. Partial dehydration of animals before protein injection resulted in the acceleration of amyloid formation. The human proteins were deposited as amyloid within the mouse renal blood vessel walls and parenchymal tissue, as well as in other organs. The deposits were Congo red-positive, exhibited green birefringence, and had a fibrillar ultrastructure. As evidenced immunohistochemically, the experimentally induced amyloid deposits consisted of the injected human light chains, and in addition, contained mouse amyloid P component (AP); mouse immunoglobulin (Ig) or inflammatory-associated amyloid A protein was not detected. Extraction and characterization of the amyloid deposits found within the mouse kidney revealed the presence of a predominantly intact human light polypeptide chain. Mice injected in identical manner with a non-amyloid-associated Bence Jones protein had no or only rare amyloid deposits. The experimental mouse model provides a means to ascertain the amyloidogenic potential of human monoclonal light chains and to study further the pathogenesis of AL amyloidosis.
Amyloid↗
Serum amyloid P component in chronic renal failure and dialysis.
A normal reference interval for serum amyloid P component (SAP) concentration in the serum was established in 500 healthy adult individuals (274 women, 226 men), by electroimmunoassay calibrated with standards of highly purified, isolated SAP. The mass of SAP in these was determined from the extinction coefficient of SAP at 280 nm measured here precisely for the first time by spectrophotometry and cryogenic drying. The mean (SD, range) SAP concentration was significantly lower in women: 24 mg/l (8, 8-55), compared to 32 mg/l (7, 12-50) in men (P less than 0.001). In renal insufficiency patients, 38 with chronic renal failure, 79 on hemodialysis and 66 on continuous ambulatory peritoneal dialysis, the mean values for SAP concentration were all significantly higher than normal (range of means, 39-59 mg/l in men and 35-42 mg/l in women), but did not correlate with serum creatinine, duration of dialysis or the presence of an acute phase response. The metabolism of SAP is thus altered in renal failure and is not normalized by dialysis, but it is not clear whether this is relevant to the pathogenesis of dialysis related arthropathy and amyloidosis.
Adult↗
Imaging of haemodialysis-associated amyloidosis with 123I-serum amyloid P component.
Long-term haemodialysis is frequently complicated by amyloid deposition in which the fibrils consist of beta 2-microglobulin. Dialysis-related amyloid disease causes extensive morbidity and has been associated with deaths in some cases. All amyloid deposits contain amyloid P component that is derived from the normal circulating protein, serum amyloid P component (SAP). We have used scintigraphic imaging after injection of 123I-labelled SAP to assess the distribution of amyloidosis in 38 patients receiving long-term haemodialysis for end-stage renal failure. There was focal localisation of tracer at all sites where histological examination confirmed amyloid deposition. Splenic uptake was seen in 12 patients, indicating splenic amyloidosis, but there was no evidence of other visceral involvement. 6 control subjects who had been dialysed for under 1.5 years showed no localisation of tracer, nor was there any uptake of 123I-labelled human serum albumin in 3 long-term dialysis patients with histologically confirmed amyloidosis and positive 123I-SAP images. Negative scans were also obtained in 5 patients who had been transplanted 0.8-2.4 years previously, despite past evidence of dialysis arthropathy (5) and histologically proven amyloidosis (4). 123I-SAP scintigraphy may be helpful as a non-invasive method for both the diagnosis and monitoring of dialysis-associated amyloidosis.
Amyloidosis↗
Isolation and characterization of the integral glycosaminoglycan constituents of human amyloid A and monoclonal light-chain amyloid fibrils.
Amyloid fibrils were isolated by extraction in water from the livers and spleens of four patients who had died of monoclonal, light-chain (AL)-type, systemic amyloidosis and one with reactive systemic, amyloid A protein (AA)-type amyloidosis. Each fibril preparation contained 1-2% by weight of glycosaminoglycan (GAG) which was tightly associated with the fibrils and not just co-isolated from the tissues with them. After exhaustive digestion of the fibrils with papain and Pronase, the GAGs were specifically precipitated with cetylpyridinium chloride and were identified by cellulose acetate electrophoresis and selective susceptibility to specific glycosidases. All the preparations contained approximately equal amounts of heparan sulphate and dermatan sulphate. There was no evidence for the presence of chondroitin sulphate or other GAGs. Fine structural analysis by oligosaccharide mapping in gradient polyacrylamide gels, following partial digestion with specific glycosidases, showed very similar structures among the heparan sulphates and the dermatan sulphates, respectively. GAGs were also extracted by solubilizing amyloid fibrils in 4 M-guanidinium chloride followed by CsCl density-gradient ultracentrifugation. Although a minor proportion of the GAG material obtained in this way was apparently in the form of proteoglycan molecules, most of it was free GAG chains. The presence in amyloid fibrils of different types, in different organs and from different patients of particular GAG classes with similar structures supports the view that these molecules may be of pathogenic significance.
Amyloidosis↗
Tissue vitronectin in normal adult human dermis is non-covalently bound to elastic tissue.
Vitronectin is a multifunctional human plasma glycoprotein that is also found in constant association with elastic tissue fibers in normal adults. We have investigated the nature of the association of vitronectin with elastic tissue, and compared it to that of other elastic fiber-associated proteins, namely fibrillin and amyloid P component. Samples of normal human dermis were incubated with a variety of extraction agents, including high molar salt solution, non-ionic detergent (Nonidet P-40), the reducing agents dithiothreitol or 2-mercaptoethanol, and the chaotropic agents sodium dodecyl sulfate or guanidine hydrochloride. Vitronectin purified from serum typically migrates as two bands of 75 and 65 kD. By contrast, immunoblotting studies of residual dermal material after extraction with the various agents revealed only lower molecular weight (58, 50, 42, 35, and 27 kD) anti-vitronectin reactive bands. Although these bands may represent degradation products of vitronectin generated as a result of the extraction procedure, we cannot exclude the possibility that tissue vitronectin is distinct from plasma vitronectin. Anti-vitronectin reactive polypeptides co-migrating with the 58-, 50-, and 42-kD bands were solubilized following extraction with sodium dodecyl sulfate or guanidine hydrochloride, but not with the other extraction agents. Immunofluorescence studies using residual dermal material after extraction with guanidine hydrochloride demonstrated a marked reduction in elastic fiber staining intensity with anti-vitronectin and anti-amyloid P component, but not with anti-fibrillin. Thus the majority, if not all of dermal vitronectin, is, like amyloid P component, non-covalently associated with, and not an integral constituent of, elastic fibers.
Blood Proteins↗
Studies in vivo and in vitro of serum amyloid P component in normals and in a patient with AA amyloidosis.
Pure serum amyloid P component (SAP) was isolated from a normal donor pool, from individuals with the different genotypes of an MspI restriction fragment length polymorphism (RFLP) linked to the SAP gene, and from a patient with AA amyloidosis. The SAP preparations were all identical and all behaved as a single homogeneous species in polyacrylamide gel electrophoresis, isoelectric focussing, reverse-phase chromatography, binding in vitro to phosphoethanolamine-Sepharose (binding constant 2.4 x 10(7) l/mol) and AL amyloid fibrils (1.6 x 10(8) l/mol), and binding to amyloid deposits in vivo in mice with casein-induced amyloidosis. The in vivo metabolism of 125I-SAP from a single donor was normal and identical in three healthy individuals representing the three different MspI RFLP genotypes. There is thus no frequent polymorphism of SAP in normal subjects, and SAP altered with respect to the characteristics studied here is not a necessary condition for pathogenesis of systemic AA amyloidosis.
Amyloidosis↗
Plasma leucocyte elastase concentrations in smokers.
The associations between cigarette smoking, plasma leucocyte elastase concentration, peripheral leucocyte count and FEV1 were examined in 148 men, 72 of whom were current cigarette smokers, 40 of whom were ex-smokers, and 36 who had never smoked. All men were part of a long-term survey. Smokers had significantly higher plasma leucocyte elastase concentrations than ex-smokers or those who had never smoked. Mean current FEV1 was lower, and the annual decline in FEV1 in the preceding 10 years was faster in smokers than the other two groups. A few smokers had slight increases in serum C-reactive protein concentrations. Although peripheral blood leucocyte counts were higher in smokers than in non-smokers or ex-smokers, no association was found in any of the three groups of men between plasma elastase concentration and peripheral leucocyte count, nor between either of these two variables and annual decline in FEV1 or current level of FEV1. There was also no relation between plasma elastase concentration and reported daily cigarette consumption or mixed expired carbon monoxide in smokers. The results indicate that some male smokers have increased in vivo release of elastase from peripheral blood neutrophils at a time when there is no evidence of acute infection. Because leucocyte elastase is a strong candidate for pulmonary tissue damage, further studies of the mechanisms that increase plasma concentrations are indicated.
Adult↗
C-reactive protein as an index of disease activity. Comparison of tenidap, cyclophosphamide and dexamethasone in rat adjuvant arthritis.
C-reactive protein (CRP) concentrations are a useful plasma protein measure that correlate with disease severity and radiographic progression in rheumatoid arthritis (RA). We compared 3 drugs with different mechanisms, i.e., tenidap, dexamethasone and cyclophosphamide, on both CRP levels and soft tissue swelling in the rat adjuvant arthritis model. CRP rose from a normal concentration of approximately 400 micrograms/ml during the first phase of adjuvant arthritis to approximately 1200 micrograms/ml (primary response), then fell to approximately 900 micrograms/ml and rose again as the disease became systemic during the secondary response to approximately 1400 micrograms/ml. When treatment was administered prophylactically, tenidap and dexamethasone suppressed both the primary and secondary CRP and swelling responses. Cyclophosphamide was without effect in the primary response, but inhibited both swelling and CRP in the secondary response. When therapeutic treatment was begun after secondary disease was established, only tenidap and dexamethasone inhibited CRP and swelling. Both dexamethasone and cyclophosphamide decreased lymphocyte numbers during treatment whereas lymphocyte numbers were elevated during tenidap treatment, suggesting a different mechanism of action for tenidap. CRP levels were more closely linked to the rate of change of paw swelling (disease progression) than to paw volume.
Animals↗
Evaluation of systemic amyloidosis by scintigraphy with 123I-labeled serum amyloid P component.
BACKGROUND: In systemic amyloidosis the distribution and progression of disease have been difficult to monitor, because they can be demonstrated only by biopsy. Serum amyloid P component (SAP) is a normal circulating plasma protein that is deposited on amyloid fibrils because of its specific binding affinity for them. We investigated whether labeled SAP could be used to locate amyloid deposits. METHODS: Purified human SAP labeled with iodine-123 was given intravenously to 50 patients with biopsy-proved systemic amyloidosis--25 with the AL (primary) type and 25 with the AA (secondary) type--and to 26 control patients with disease and 10 healthy subjects. Whole-body images and regional views were obtained after 24 hours and read in a blinded fashion. RESULTS: In the patients with amyloidosis the 123I-SAP was localized rapidly and specifically in amyloid deposits. The scintigraphic images obtained were characteristic and appeared to identify the extent of amyloid deposition in all 50 patients. There was no uptake of the 123I-SAP by the control patients and the healthy subjects. In all patients with AA amyloidosis the spleen was affected, whereas the scans showed uptake in the heart, skin, carpal region, and bone marrow only in patients with the AL type. Positive images were seen in six patients in whom biopsies had been negative or unsuccessful; in all six, amyloid was subsequently found on biopsy or at autopsy. Progressive amyloid deposition was observed in 9 of 11 patients studied serially. CONCLUSIONS: Scintigraphy after the injection of 123I-SAP can be used for diagnosing, locating, and monitoring the extent of systemic amyloidosis.
Adult↗
Pentraxin-chromatin interactions: serum amyloid P component specifically displaces H1-type histones and solubilizes native long chromatin.
Pure serum amyloid P component (SAP) and native long chromatin, mixed together at wt/wt ratios between 1:1 and 1:2 in the presence of physiological concentrations of NaCl and calcium, both remained in solution, whereas each alone precipitates rapidly under these conditions. This solubilization accompanies the binding of SAP to chromatin and the displacement of H1-type histones, which are essential for condensation and higher order folding of chromatin. Such binding of SAP to chromatin is remarkable since displacement of H1 and H5 by salt alone requires approximately 0.5 M NaCl. SAP also bound to nucleosome core particles forming soluble complexes with an apparent stoichiometry of 1:2, a result that is compatible with attachment of SAP at the nucleosome dyad, the site of H1 in intact chromatin. SAP thus undergoes a specific, avid interaction with chromatin that promotes its solubilization and may thereby contribute to the physiological handling of chromatin released from cells in vivo. In contrast, C-reactive protein (CRP) did not bind significantly to either chromatin or to core particles at physiological ionic strength. Incubation of chromatin with either normal serum, or acute phase human serum containing raised levels of CRP, did not induce complement activation regardless of the presence of added SAP or CRP, nor was any cleavage of DNA observed.
Animals↗
Preparative isolation of human beta 2-microglobulin from urine.
Human beta 2-microglobulin was isolated from the urine of patients with tubular proteinuria by sequential ion exchange and gel filtration chromatography on Sepharose Q fast flow and Sephacryl S-200 respectively. Large volumes of starting material could be processed and yielded a homogeneous preparation of purity 99% or greater with a recovery of about 50% of the immunochemically demonstrable beta 2-microglobulin. This method provides protein suitable for structural studies and investigation of beta 2-microglobulin amyloidogenesis.
Chromatography, Agarose↗
C-reactive protein and bacterial infection in preterm infants.
Serum C-reactive protein (CRP) concentration was measured by a new solid phase ligand-binding radiometric monoclonal antibody immunoassay in a prospective study of 193 consecutively born preterm infants. In 104 with no clinical or laboratory evidence of infection the median CRP in cord serum was 0.125 mg/l (range 0.011-6.0 mg/l), at 24 h it was 1 mg/l (0.016-7.0) and at 48 h 2 mg/l (0.400-8.0). The present highly sensitive assay has enabled these normal ranges to be defined for the first time, at levels below the threshold of non-labelled immunoassays and of all commercially available CRP assays. The values in cord serum were significantly lower than in normal healthy adults (median 0.8 mg/l, range 0.07-29 mg/l, n = 468). Arterial catheterisation and endotracheal intubation, in the absence of infection, did not appear to elevate CRP, nor did cerebral germinal layer or intraventricular haemorrhage. Among nine infants with confirmed septicaemia eight had a serum CRP level raised at least once during the first 48 h and serum CRP in the other one increased 250-fold in 24 h before treatment was started. Using this assay, serum CRP is a useful and rapidly available adjunct to clinical assessment in diagnosis and exclusion of bacterial infection in the early neonatal period, has encouraged us to withhold or discontinue antibiotics and also has a role in monitoring response to treatment.
Antibodies, Monoclonal↗
A primed state exists in vivo following histological regression of amyloidosis.
Using a sensitive, quantitative, and non-invasive in vivo method, based on the specific binding of serum amyloid P component to amyloid fibrils, we have directly documented the spontaneous resolution of AA amyloid deposits in mice, and the prolonged existence thereafter of a primed state of enhanced susceptibility to further amyloid deposition. These results may have important implications for understanding and management of amyloidosis in humans.
Amyloid↗
Metabolic studies of radioiodinated serum amyloid P component in normal subjects and patients with systemic amyloidosis.
125I-Serum amyloid P component (SAP), injected intravenously into 10 normal subjects, remained predominantly intravascular with mean (SD) T1/2 (half time) in plasma of 24.5 (5.9) h. The fractional catabolic rate of 68 (19)% of the plasma pool per day was more rapid than other reported human plasma proteins. All radioactivity was excreted in the urine by 14 d. In 16 patients with monoclonal gammopathy or chronic inflammatory diseases, but without amyloidosis, 125I-SAP metabolism was normal. However, among 45 patients with biopsy-proven systemic amyloidosis (25, amyloid A type; 20, amyloid L type), 125I-SAP was cleared from the plasma more rapidly, accumulated in the amyloid deposits, and persisted there. The T1/2 in amyloid, measured directly with 131I-SAP, was 24 d. Repeat studies after 6-18 mo were notably consistent in normals but changed significantly in amyloid patients, generally correlating with clinical signs of disease progression. Measurements of 125I-SAP turnover may thus be of value for diagnosis and monitoring of amyloidosis. Analysis of SAP metabolism in amyloidosis suggests that plasma SAP is in dynamic equilibrium with a very large amyloid pool, and in two autopsies the total mass of SAP in the amyloid deposits was 2,100 and 21,000 mg, respectively.
Adult↗
Campylobacter pylori, duodenal ulcer disease, and gastrin.
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Adult↗
Serum amyloid P component binds to cell nuclei in vitro and to in vivo deposits of extracellular chromatin in systemic lupus erythematosus.
Serum amyloid P component (SAP) is the single plasma protein that, from the milieu of whole normal human serum, undergoes specific calcium-dependent binding to isolated DNA and chromatin in vitro. We now report for the first time that SAP in whole serum also undergoes calcium-dependent binding to nuclei of epidermal cells in sections of normal human skin and to nuclei of fixed Hep-2 cells, a human epithelial cell line. Furthermore, and most importantly, SAP was detected in association with unusual globular dermal deposits of nuclear material in skin biopsies from two patients with systemic lupus erythematosus. This is the first evidence for binding of SAP to extracellular chromatin in vivo and supports the idea that SAP may have an important physiological role in the disposal of this material.
Cell Line↗
Imaging amyloidosis in Still's disease.
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Adrenal Glands↗