Biomedical subjects

M C de Beer

Publications and source records attributed to M C de Beer.

28 records · Page 2Linked to original sources

PubMed1994

Mouse serum amyloid A protein (SAA5) structure and expression.

A novel member of the mouse serum amyloid A protein family, SAA5, has been identified as a normal apolipoprotein component of non-acute-phase high density lipoprotein (HDL). The structure of SAA5 was derived from a clone isolated from a normal Balb/c liver cDNA library. The clone predicts a pre-SAA5 molecule of 130 residues from which an 18-residue leader peptide is cleaved. The mature molecule has an octapeptide insert spanning from position 70 to 77. Similar inserts are found in human C-SAA and, paradoxically, in acute-phase SAA molecules of a number of other species. There is 48% amino acid identity between apo-SAA5 and the other mouse SAA proteins and 57% identity between the human C-SAA and apo-SAA5. The SAA5 mRNA is three times larger than previously identified SAA mRNAs. Although SAA5 is constitutively expressed in the liver, it has a rapid albeit muted response to inflammatory stimuli. The increase of SAA5 mRNA is due to increased transcription rather than mRNA stabilization. Plasma SAA5 levels during the acute phase are biphasic, either because of translational control or displacement from HDL and rapid clearance. We propose that constitutive SAAs (SAA5) on normal HDL contribute to its normal physiological role, whereas the dramatically inducible family members (SAA1, SAA2, SAA3) equip this particle for an altered functional role during inflammation.

Amino Acid Sequence↗

PubMed1994

Serum amyloid A protein family. Differential induction by oxidized lipids in mouse strains.

During inflammation, serum amyloid A (SAA) protein increases up to 1000-fold and can become a major component of high-density lipoprotein (HDL). We have identified a new apolipoprotein molecule (SAA5) as a distinct member of the SAA family. It differs from the inflammatory isotypes (SAA1, SAA2, and SAA3) not only in structure but in the fact that it is constitutive on normal HDL where it accounts for more than 90% of total SAA. Whereas all members of the SAA family, including SAA5, responded to endotoxin administration, SAA5 contrasted with other SAAs in its resistance to induction either by a high-fat, high-cholesterol atherogenic diet or the injection of mildly oxidized LDL (MM-LDL). These data provide further evidence that the induction of inflammatory molecules by oxidized lipids is selective. In atherosclerosis-susceptible C57BL/6 mice and atherosclerosis-resistant C3H/HeJ mice, the inflammatory SAA isotypes (SAA1, SAA2, and SAA3) responded in a strain-specific manner to oxidized lipids either generated with feeding of the atherogenic diet or introduced by MM-LDL injection. We hypothesize that the constitutive SAA5 molecules on normal HDL may contribute to its normal physiological role and that the dramatic induction of the inflammatory SAA subfamily equips the particle for an altered yet related functional role appropriate to the inflammatory state.

Animals↗

PubMed1993

Structural prerequisites for serum amyloid A fibril formation.

Most studies of experimental amyloid A protein (AA) amyloidosis in mice have been performed in type A mice with BALB/c as the prototype. In these mice the products of two genes, SAA1 and SAA2, are the major apo-SAA isoforms on high density lipoprotein (HDL). Of these two isoforms, that differ at nine amino acids, only apo-SAA2 is rapidly cleared and deposited as amyloid fibrils. No mouse strain has ever been shown to be completely resistant to amyloid induction. We have found the CE/J mouse strain to be exceedingly resistant to amyloidogenesis. Data indicate that this resistance is not due to a lack of apo-SAA synthesis but rather resides in the unique apo-SAA isoform in this strain. CE/J mice have a single major apo-SAA isoform (pI 6.15) the product of a single gene. This is a hybrid molecule with features of both apo-SAA1 and apo-SAA2, differing from the latter at only six amino acids. When CD studies were performed to explore the structural relationship of this isoform to apo-SAA1 and apo-SAA2, we found that when bound to heparan sulfate proteoglycan the CE/J pI 6.15 isoform fails to undergo the beta-sheet folding typical for apo-SAA2. This evidence suggests that the folding effect of heparan sulfate proteoglycan on apo-SAA2 is important in amyloid formation.

Amino Acid Sequence↗

PubMed1993

Syrian and Armenian hamsters differ in serum amyloid A gene expression. Identification of novel Syrian hamster serum amyloid A subtypes.

Amyloid A (AA) amyloidosis is widespread throughout the animal kingdom. Several factors including: 1) precursor production; 2) precursor structure; 3) precursor degradation; and 4) precursor/product interaction with the pentraxin serum amyloid P have been implicated in amyloidogenesis, but the exact sequence of events leading to AA fibril formation and deposition remains unclear. Most models of experimental amyloidosis, including golden Syrian hamsters (Mesocricetus auratus), involve massive and repeated inflammatory stimulation; however, the model of spontaneous amyloidosis with aging in female, but not male, Syrian hamsters permits analysis of amyloidogenic factors in the absence of inflammation. Another genus, the Armenian hamster (Cricetulus migratorius), differs from Syrian hamsters both in gender-specific serum amyloid P expression and susceptibility to AA amyloidosis. In this study, we describe novel SAA molecules in the Syrian hamster in the presence and absence of inflammation. We demonstrate that, based on isoelectric separation, the Syrian hamster SAA proteins can be separated into two broad subfamilies. Plasma SAA concentration in female Syrian hamsters increases spontaneously with age, and fragments of a basic SAA isotype expressed both hepatically and extrahepatically are selectively deposited as AA fibrils. After inflammatory stimulation, the patterns of SAA gene expression in Syrian and Armenian hamsters differ. In Syrian hamsters, both hepatic SAA mRNA and the high density lipoprotein apoSAA content increase approximately 1000-fold; in Armenian hamsters, hepatic SAA mRNA is limited in quantity and different in structure; and although plasma SAA proteins increase three- to fivefold, apoSAA is not detectable in high density lipoprotein. The results suggest that regulation and site of precursor production as well as precursor structure influence AA amyloidogenesis in these two hamster genera.

Amino Acid Sequence↗

PubMed1993

Characterization of the inbred CE/J mouse strain as amyloid resistant.

Inbred CE/J mice have been identified as extremely resistant to azocasein-induced amyloidosis relative to five commonly used inbred strains, A/J, CBA/J, C57BL/6J, C3H/HeN, and SJL/J. The enhanced amyloid resistance in CE/J mice seems to derive from the novel structure of the SAA gene family in CE/J mice, as determined by Southern blot hybridization analysis of SAA gene structure and isoelectric focusing analysis of acute phase SAA proteins in the six inbred strains. In CE/J mice, a single, novel SAA isoform of pI 6.15 is present, whereas in the other strains the amyloidogenic SAA2 isoform (pI 6.3) is codominantly expressed with SAA1 (pI 6.45). Two other inbred strains, PERU and IS/CAM, share common SAA specific HindIII DNA fragments with CE/J mice. Wild-derived Mus musculus mice differ from all of the inbred strains studied, both in SAA gene structure and in the pattern of SAA isoform production; two isoforms, one pI 6.15 and the other pI 6.3 (corresponding to SAA2), were codominantly expressed. Only the pI 6.15 isoform, not SAA1 and 2, was produced by CE/J mice in response to lipopolysaccharide, casein, silver nitrate, interleukin-1, or tumor necrosis factor; tumor necrosis factor was a weaker stimulus than interleukin-1 for the pI 6.15 isoform as it is for SAA1 and 2 production in the other inbred strains. This study provides a new line of evidence supporting the role of precursor structure as a determining factor in murine amyloid A amyloidosis and provides a valuable model for studies of amyloidogenesis.

Acute-Phase Reaction↗

PubMed1992

Mouse serum amyloid A protein. Complete amino acid sequence and mRNA analysis of a new isoform.

Four serum amyloid A protein (SAA) genes and two gene products, apo-SAA1 and apo-SAA2 were identified in BALB/c mice (type A). SJL/J mice (type B) are thought to be defective in apo-SAA2 expression. A unique variant of mouse apo-SAA was identified in SJL/J mice by isoelectric-focusing analysis of high-density lipoprotein from endotoxin-treated mice. Complete amino-acid-sequence analysis of this quantitatively major form of SJL/J apo-SAA (pI 5.9) showed it to be identical with the apo-SAA2 isoform from BALB/c mice, except for the substitution of aspartic acid for alanine at position 101. Isoform-specific analysis of mRNA from liver of BALB/c and SJL/J mice and their F1 hybrid progeny (CSJLF1/J) mice revealed further differences in the 3' untranslated regions of the genes, not only encoding apo-SAA2 and apo-SAA pI 5.9, but also apo-SAA1. The SAA genes of SJL/J mice thus differ from BALB/c in exon 4. Additional minor isoforms corresponding to apo-SAA2 (pI 6.3) in SJL/J mice and apo-SAA (pI 5.9) in BALB/c mice were identified. We propose that, when analysing a multigene family such as SAA, thorough analysis at the protein level should complement molecular-biological approaches where the use of a too-limited repertoire of probes can obscure complexities.

Amino Acid Sequence↗

PubMed1992

Identification of novel members of the serum amyloid A protein superfamily as constitutive apolipoproteins of high density lipoprotein.

A novel serum amyloid A protein (SAA) has been identified as a normal apolipoprotein component of non-acute phase high density lipoprotein. This novel SAA has been designated "constitutive" SAA (C-SAA) to distinguish it from "acute phase" SAA (A-SAA). C-SAA was partially sequenced, and immunochemical analyses indicated that it constitutes a distinct subclass of apolipoproteins within the SAA superfamily. A C-SAA cDNA clone was isolated from a human liver library and sequenced. The clone predicts a pre-C-SAA molecule of 130 residues from which an 18-residue leader peptide is cleaved. The 112-residue mature molecule is 8 residues longer than human A-SAA; the size difference is due to the presence of an octapeptide between positions 70 and 77 that is not found in the corresponding region of human A-SAA. Paradoxically, octapeptides of similar composition are found at similar positions in the A-SAAs of a number of other species. The C-SAA octapeptide specifies the first two residues of a NSS tripeptide, the only potential N-linked glycosylation site in the molecule. Studies indicate that approximately 50% of these sites are glycosylated, thereby giving rise to two size classes, 14 and 19 kDa, of C-SAA in vivo. Human acute phase liver contains little C-SAA mRNA relative to the levels of A-SAA mRNA, and the treatment of PLC/PRF/5 hepatoma cells with monocyte-conditioned medium does not induce C-SAA mRNA concentrations to detectable levels, in contrast to the massive induction of A-SAA mRNA observed. C-SAA is therefore not a major acute phase reactant.

Amino Acid Sequence↗

PubMed1991

Identification of a novel serum amyloid A protein in BALB/c mice.

Four serum amyloid A protein (SAA) genes and two SAA gene products, SAA1 and SAA2, were identified in BALB/c mice. Using analytical isoelectric focusing we have identified a quantitatively significant new member of the SAA family and designated it 'SAA5'. This protein has characteristics never before described for any SAA molecule. In the highly conserved region between amino acids 33 and 44, identical in all SAAs from all species examined, SAA5 had four amino acid substitutions. In addition, the induction of SAA5 by lipopolysaccharide had different kinetics from that of the other mouse SAAs. Our data suggest that the mouse SAA gene family is more complex in composition and regulation than previously surmised.

Amino Acid Sequence↗

PubMed1989

Human serum amyloid A protein. The assignment of the six major isoforms to three published gene sequences and evidence for two genetic loci.

Serum amyloid A protein (apo-SAA) is an acute-phase reactant and an apolipoprotein of high density lipoproteins (HDL). Six major isoforms of apo-SAA occur in humans (pI 6.0, 6.4, 7.0, 7.4, 7.5, 8.0). In this report we have rationalized the phenotypic expression of apo-SAA isoforms with published apo-SAA structures predicted from apo-SAA cDNA's pA1 and pSAA82 and the genomic DNA SAAg9. The six apo-SAA isoforms fall into three pairs, pI 6.0/6.4, 7.0/7.5, and 7.4/8.0, which are products of cDNA pA1, cDNA pSAA82, and genomic DNA SAAg9, respectively. The second of each isoform pair (i.e. pI 6.4, 7.5, and 8.0) is the "primary" product: a 104-residue peptide with the NH2-terminal sequence Arg-Ser-Phe-Phe. Each primary product is processed either to a major 103-residue peptide with the NH2-terminal sequence Ser-Phe-Phe or processed to a minor 102-residue product which results from the loss of both an Arg and a Ser residue from the NH2 termini. These "secondary" products have the lower pI values of 6.0, 7.0, and 7.4, respectively. The isoelectric points of the SAAg9 products were confirmed by expression of SAAg9 in transfected mouse L-cells. Both the pI 8.0 and 7.4 isoforms were present in cellular extracts, suggesting that post-translational modification of apo-SAA may occur intracellularly. However, the greater relative abundance of the pI 7.4 isoform extracellularly suggests that the major conversion may occur after secretion. Whereas the gene corresponding to the pA1 cDNA sequence does not show allelic variation, the segregation characteristics of the pI 7.0/7.5 and 7.4/8.0 isoform pairs amongst individuals suggests that these isoforms are the products of genes (with sequences corresponding to pSAA82 and SAAg9, respectively) which are allelic variants at a single locus distinct from that for the pI 6.0/6.4 isoform pair.

Amino Acid Sequence↗

PubMed1987

Neutrophil association and degradation of normal and acute-phase high-density lipoprotein 3.

The interaction of normal and acute-phase high-density lipoproteins of the subclass 3 (N-HDL3 and AP-HDL3) with human neutrophils and the accompanying degradation of HDL3 apolipoproteins have been studied in vitro. The chemical composition of normal and acute-phase HDL3 was similar except that serum amyloid A protein (apo-SAA) was a major apolipoprotein in AP-HDL3 (approx. 30% of total apolipoproteins). 125I-labelled AP-HDL3 was degraded 5-10 times faster than 125I-labelled N-HDL3 during incubation with neutrophils or neutrophil-conditioned medium. Apo-SAA, like apolipoprotein A-II (apo-A-II), was more susceptible than apolipoprotein A-I (apo-A-I) to the action of proteases released from the cells. The amounts of cell-associated AP-HDL3 apolipoproteins at saturation were up to 2.8 times greater than N-HDL3 apolipoproteins; while apo-A-I was the major cell-associated apolipoprotein when N-HDL3 was bound, apo-SAA constituted 80% of the apolipoproteins bound in the case of AP-HDL3. The associated intact apo-SAA was mostly surface-bound as it was accessible to the action of exogenous trypsin. alpha 1-Antitrypsin-resistant (alpha 1-AT-resistant) cellular degradation of AP-HDL3 apolipoproteins also occurred; experiments in which pulse-chase labelling was performed or lysosomotropic agents were used indicated that insignificant intracellular degradation occurred which points to the involvement of cell-surface proteases in this degradation.

Acute-Phase Proteins↗