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Biomedical subjects

M B Pepys

Publications and source records attributed to M B Pepys.

At least 145 records · Page 8Linked to original sources
PubMed1988

Hormone binding globulins undergo serpin conformational change in inflammation.

A surprising recent finding is that thyroxine binding globulin (TBG) and cortisol binding globulin (CBG), are members of the serine protease inhibitor (serpin) superfamily. Apparently evolution has completely adapted the serpin structure for its new role in these proteins as a transport agent, as there is no evidence of any retained protease inhibitory activity. This drastic change in function raises the question as to why such a complex molecular framework has been selected for the relatively simple task of hormone transport? To function as inhibitors the serpins have a native stressed (S) conformation that makes them vulnerable to proteolytic cleavage, the cleavage being accompanied by an irreversible transition to a stable relaxed (R) form. We demonstrate here that TBG and CBG have retained the stressed native structure typical of the inhibitor members of the family and we provide evidence that the S-R transition has been adapted to allow altered hormone delivery at inflammatory sites.

Amino Acid Sequence↗
PubMed1988

A pentameric form of human serum amyloid P component. Crystallization, X-ray diffraction and neutron scattering studies.

Human serum amyloid P component crystallizes from sodium acetate buffer (pH 5.5) in the presence of calcium and polyethylene glycol 6000, at 4 degrees C. The space group is P2(1) and the cell parameters are a = 69.0 A (1 A = 0.1 nm), b = 99.3 A, c = 96.8 A, beta = 96.1. Density considerations supported by neutron scattering and gel filtration experiments indicate that the species crystallized is pentameric. The orientation of the pentamer 5-fold axis is determined and a crystal packing for the discs is proposed.

Crystallization↗
PubMed1988

Diagnostic radionuclide imaging of amyloid: biological targeting by circulating human serum amyloid P component.

The specific molecular affinity of the normal plasma protein, serum amyloid P component (SAP), for all known types of amyloid fibrils was used to develop a new general diagnostic method for in-vivo radionuclide imaging of amyloid deposits. After intravenous injection of 123I-labelled purified human SAP there was specific uptake into amyloid deposits in all affected patients, 7 with systemic AL amyloid, 5 with AA amyloid, and 2 with beta 2M amyloid, in contrast to the complete absence of any tissue localisation in 5 control subjects. Distinctive high-resolution scintigraphic images, even of minor deposits in the carpal regions, bone marrow, or adrenals, were obtained. This procedure should yield much information on the natural history and the management of amyloidosis, the presence of which has hitherto been confirmed only by biopsy. Clearance and metabolic studies indicated that, in the presence of extensive amyloidosis, the rate of synthesis of SAP was greatly increased despite maintenance of normal plasma levels. Furthermore, once localised to amyloid deposits the 123I-SAP persisted for long periods and was apparently protected from its normal rapid degradation. These findings shed new light on the pathophysiology of amyloid and may have implications for therapeutic strategies based upon specific molecular targeting with SAP.

Adult↗
PubMed1988

Specific localization and imaging of amyloid deposits in vivo using 123I-labeled serum amyloid P component.

Highly specific, high-resolution scintigraphic images of amyloid-laden organs in mice with experimentally induced amyloid A protein (AA) amyloidosis were obtained after intravenous injection of 123I-labeled serum amyloid P component (SAP). Interestingly, a much higher proportion (up to 40%) of the injected dose of heterologous human SAP localized to amyloid and was retained there than was the case with isologous mouse SAP, indicating that human SAP binds more avidly to mouse AA fibrils than does mouse SAP. Specificity of SAP localization was established by the failure of the related proteins, human C-reactive protein and Limulus C-reactive protein, to deposit significantly in amyloid and by the absence of human SAP deposition in nonamyloidotic organs. However, only partial correlations were observed between the quantity of SAP localized and two independent estimates, histology and RIA for AA of the amount of amyloid in particular organs. It is not clear which of the three methods used reflects better the extent or clinical significance of the amyloid deposits but in vivo localization of radiolabeled SAP, detectable and quantifiable by gamma camera imaging, is apparently extremely sensitive. These findings establish the use of labeled SAP as a noninvasive in vivo diagnostic probe in experimental amyloidosis, potentially capable of revealing the natural history of the condition, and suggest that it may also be applicable generally as a specific targeting agent for diagnostic and even therapeutic purposes in clinical amyloidosis.

Amyloidosis↗
PubMed1988

Insulin as an amyloid-fibril protein at sites of repeated insulin injections in a diabetic patient.

A patient with Type 1 (insulin-dependent) diabetes mellitus developed localised amyloidosis at the sites of his injections of porcine insulin. A major amyloid fibril protein was extracted and, by means of its amino acid composition and amino acid sequence, it was shown to contain intact insulin molecules. Porcine insulin is the tenth protein and the first foreign protein to be chemically identified in human amyloid fibrils.

Adult↗
PubMed1988

Amyloid P component binds to keratin bodies in human skin and to isolated keratin filament aggregates in vitro.

Dermal keratin bodies, consisting mainly of keratin intermediate filament aggregates (KIFA) coated with IgM anti-KIF autoantibodies, are present in normal human skin and occur in increased quantities in certain skin diseases. Keratin bodies are normally rapidly removed, but in primary localized cutaneous amyloidosis (PLCA) they are converted by an unknown mechanism to amyloid. Amyloid P component (AP), a glycoprotein identical to, and derived from, the normal plasma protein serum amyloid P component (SAP), is present in all forms of amyloid including PLCA. We investigated the interaction between SAP, keratin bodies, and KIFA. Immunofluorescence staining of normal skin using fluoresceinated anti-SAP and rhodamine-conjugated anti-IgM, or AE-1/AE-3 anti-keratin antibodies followed by Texas Red-conjugated anti-mouse immunoglobulin, showed that 52% +/- 4 (mean +/- sem, n = 6) of keratin bodies bound anti-SAP. Similar findings were present in a biopsy from a patient with lichen planus. Isolated KIFA, prepared by 8M urea extraction of normal human epidermis or cultured keratinocytes, were preincubated with normal human serum as a source of SAP and then stained with fluoresceinated anti-SAP. Bright fluorescence seen when the incubation medium contained Ca++ was absent in the presence of ethylenediamine tetraacetic acid. Specific Ca++-dependent binding of SAP to KIFA was confirmed using immunoblotting. Binding of SAP to KIFA did not prevent their degradation following exposure to trypsin or alpha-chymotrypsin. Similarly, partial enzymatic digestion of KIFA did not abrogate their ability to bind SAP. Our findings, that SAP is associated with keratin bodies in skin and exhibits Ca++-dependent binding to KIFA in vitro, identify keratin filaments as a newly recognized ligand for SAP.

Benzothiazoles↗
PubMed1988

Comparison of the role of complement in immunity to Schistosoma mansoni in rats and mice.

In vivo depletion of C3 with cobra venom factor (CoF) was used to demonstrate the participation of complement in the innate immunity to S. mansoni and in the acquired immunity of both actively and passively immunized rats. Complement was shown to play an important role in innate immunity, being more involved later in larval migration (Days 8-13 post-infection) than at earlier times (Days--1-3 and Days 3-8 post-infection). Furthermore, the specific component of immunity conferred by immune serum transferred at the lung-migration stage also required complement for optimal expression. This supports the notion that both innate and acquired immunity act not against the much studied early post-penetration stages, but primarily against the lung stages. Although decomplementation at earlier stages of parasite migration (up to 3 days post-infection) did cause some reduction of innate immunity, there was no evidence of any effect on the levels of resistance actively induced by exposure to irradiated cercariae. This suggests that, while complement may play a role in innate immunity during the skin-migration phase, specific complement-mediated attrition does not play a crucial role at this time. The situation was very different in the mouse model, since no involvement of complement in either innate or irradiated vaccine-induced immunity could be demonstrated within the first 15 days of infection. Thus, there appear to be phases in the parasite migration in rats, but not in mice, during which complement becomes a critical factor in both innate and acquired immunity to S. mansoni.

Animals↗
PubMed1987

Serum amyloid P component is the major calcium-dependent specific DNA binding protein of the serum.

Serum amyloid P component (SAP), a member of the highly conserved pentraxin family of plasma proteins, was found to be the only protein in whole normal or acute phase serum which underwent specific calcium-dependent binding to either single or double-stranded DNA immobilised on gel. Isolated purified SAP also bound to long chromatin, to H1-stripped chromatin and to native DNA in solution at physiological ionic strength. Pure SAP which had been immobilised on gel, specifically bound nucleosome core particles from solution. These observations strongly suggest that SAP may bind to extracellular chromatin and DNA in vivo and that this may be its physiological role.

Animals↗
PubMed1987

Isolation and characterisation of goat C-reactive protein.

A pentraxin was isolated from acute phase goat serum by its calcium-dependent affinity for agarose, and although it did not bind to phosphorylcholine immobilised on Sepharose, its binding to agarose was reversed by exposure to fluid phase phosphorylcholine. It was identified as goat C-reactive protein on the basis of its immunochemical cross-reactivity with human and bovine C-reactive protein. The molecule was composed of five identical, glycosylated, non-covalently associated subunits, each of molecular weight approx. 24,000. Acute phase serum levels in a small number of samples were not significantly different from normal levels (means 72 and 55 micrograms/ml, respectively), suggesting that goat C-reactive protein is not a major acute phase reactant. No other pentraxin was detected in goat serum.

Animals↗
PubMed1987

Acute-phase high-density lipoprotein in the rat does not contain serum amyloid A protein.

Serum amyloid A protein (SAA) is an acute-phase apolipoprotein of high-density lipoprotein (HDL). Its N-terminal sequence is identical with that of amyloid A protein (AA), the subunit of AA amyloid fibrils. However, rats do not develop AA amyloidosis, and we report here that neither normal nor acute-phase rat HDL contains a protein corresponding to SAA of other species. mRNA coding for a sequence homologous with the C-terminal but not with the N-terminal part of human SAA is synthesized in greatly increased amounts in acute-phase rat liver. These observations indicate that the failure of rats to develop AA amyloid results from the absence of most of the AA-like part of their SAA-like protein, and that the N-terminal portion of SAA probably contains the lipid-binding sequences.

Acute-Phase Proteins↗
PubMed1987

Imaging of experimental amyloidosis with 131I-labeled serum amyloid P component.

131I-labeled human serum amyloid P component, which was injected into mice with experimentally induced systemic AA amyloidosis and into controls, became specifically localized and was retained in amyloidotic organs. In comparison, it was rapidly and completely eliminated from unaffected tissues and from control animals. Distinctive images of this amyloid-specific deposition of labeled serum amyloid P component were derived from whole body scanning, in vivo, of amyloidotic mice. These findings suggest that such imaging may have applications for the diagnosis and quantitation of amyloid deposits in humans.

Amyloidosis↗
PubMed1987

Measurement of serum C-reactive protein concentration after renal transplantation.

A prospective study of serial serum C-reactive protein (CRP) concentrations was made on 21 patients who had received renal allografts. CRP was raised during 27 of 32 rejection episodes and in all of five episodes of rejection associated with infection. CRP values were persistently elevated in three irreversible rejection episodes. Significantly raised CRP concentrations were documented in 14 of 20 episodes of infection. In some cases CRP was a predictive indicator of rejection or infection. In all cases of infection or rejection associated with a rise in CRP, CRP values fell following successful treatment with appropriate agents. Serial CRP measurement used in conjunction with other clinical and biochemical parameters appears to be valuable clinically following renal transplantation.

Adult↗
PubMed1987

Immunochemical cross-reactions between pentraxins of different species.

Monospecific antisera were raised by immunization with pentraxins, C-reactive protein (CRP) or serum amyloid P component (SAP), that had been isolated from man and a number of different vertebrate species. These antisera were used to test sera or serous fluids from a range of other vertebrate species and also some invertebrates. Cross-reactions between species were comparatively rare but were seen among primates and among ruminants. There was also cross-reactivity between these two groups. Antisera to plaice pentraxins did not react with sera from higher vertebrates but did recognize antigens in sera from other flat fish. No cross-reactivity was observed between CRP and SAP either within or between species despite the known homologies of amino acid sequence and similarities of structure within the family. Several reactions occurred with sera in which either CRP or SAP had previously not been sought or in which only preliminary investigations had been performed. The proteins detected were: CRP in goat, cow, sheep, cat and lemon sole; SAP in monkey and dab. These findings significantly extend the range of species in which pentraxins are known to be present.

Animals↗
PubMed1987

C-reactive protein in dogs.

Serum concentrations of C-reactive protein (CRP) in dogs with various diseases or undergoing various procedures were measured by specific immunoassay. In 20 healthy dogs from various sources, values were all less than 5 mg/L, but in 22 healthy dogs from a single source, values ranged from less than 5 mg/L in 14 dogs and from 8 to 67 mg/L in 8 dogs. Increased concentrations of serum CRP were attained 24 hours after injection of casein (n = 9; median 188 mg/L), ovariohysterectomy (n = 11; median, 144 mg/L), or elective, nonacute orthopedic surgery (n = 10; median, 83 mg/L). After inoculation of Leptospira interrogans serovar canicola (n = 5), the behavior of serum CRP as an acute-phase reactant provided a sensitive and precise objective reflection of in vivo response. The CRP concentration in random single-serum samples from 73 dogs with other inflammatory and noninflammatory disorders ranged from normal (less than 5 mg/L) to 246 mg/L and generally correlated with the extent and activity of disease.

Animals↗
PubMed1987

Identification and isolation of two pentraxins from bovine serum.

Two distinct pentraxin proteins were isolated from bovine serum by calcium-dependent affinity chromatography on high pyruvate agarose gel. One of these proteins cross-reacted specifically with certain rabbit anti-human CRP antisera and was therefore designated as bovine CRP. The other cross-reacted specifically with a sheep anti-human SAP antiserum and was therefore designated as bovine SAP. Although the mixture of these two pentraxins was not resolved by gel filtration chromatography, they were separated by solid phase absorption of the CRP with immobilized rabbit anti-human CRP antibodies. Their identity as pentraxins was confirmed by their electron microscopic appearance. Bovine CRP was composed of a single type of non-glycosylated subunit whilst bovine SAP contained two major types of glycosylated subunits and a minor polypeptide, the glycosylation of which was not determined. Serum concentrations were in the range of 5-40 mg/l and neither protein behaved as an acute phase reactant. No bovine serum protein undergoing calcium-dependent binding to phosphoryl choline or pneumococcal C-polysaccharide was obtained.

Animals↗