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Model IgG monoclonal autoantibody-anti-idiotype pair for dissecting the humoral immune response to oxidized low density lipoprotein.

ABSTRACT: Increasing evidence implicates IgG autoantibodies against oxidized forms of low density lipoprotein (oxLDL) in the pathophysiology of atherosclerotic arterial disease. However, insufficient knowledge of their structure and function is a key gap. Using an elderly LDL receptor-deficient atherosclerotic mouse, we isolated a novel IgG3k against oxLDL (designated MAb LO1). LO1 reacts with copper-oxidized LDL, but minimally with native LDL. Further analysis showed that MAb LO1 also reacts in vitro with malondialdehyde-conjugated LDL (MDA-LDL), a known key epitope in copper-oxidized LDL preparations. By screening a phage library expressing single chain variable region antibodies (scFv), we selected an anti-idiotype scFv (designated H3) that neutralizes MAb LO1 binding to MDA-LDL. Amino acid substitutions between H3 and an irrelevant control scFv C12 showed that residues in the H3 CDRH2, CDRH3, and CDRL2 are all critical for MAb LO1 binding, consistent with a conformational epitope on H3 involving both heavy and light chains. Comparison of amino acids in H3 CDRH2 and CDRL2 with apoB, the major LDL protein, showed homologous sequences, suggesting H3 has structural similarities to the MAb LO1 binding site on MDA-LDL. Immunocytochemical staining showed that MAb LO1 binds epitopes in mouse and human atherosclerotic lesions. The MAb LO1-H3 combination therefore provides a very promising model for analyzing the structure and function of an individual IgG autoantibody in relation to atherosclerosis.

SUBMITTER: Chang SH

PROVIDER: S-EPMC3326269 | biostudies-literature | 2012 Apr

REPOSITORIES: biostudies-literature

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Model IgG monoclonal autoantibody-anti-idiotype pair for dissecting the humoral immune response to oxidized low density lipoprotein.

Chang Shang-Hung SH Johns Michael M Boyle Joseph J JJ McConnell Ellen E Kirkham Paul A PA Bicknell Colin C Zahoor-ul-Hassan Dogar M M Edwards Robert J RJ Gale-Grant Oliver O Khamis Ramzi R Ramkhelawon Kurrun V V KV Haskard Dorian O DO

Hybridoma (2005) 20120401 2

Increasing evidence implicates IgG autoantibodies against oxidized forms of low density lipoprotein (oxLDL) in the pathophysiology of atherosclerotic arterial disease. However, insufficient knowledge of their structure and function is a key gap. Using an elderly LDL receptor-deficient atherosclerotic mouse, we isolated a novel IgG3k against oxLDL (designated MAb LO1). LO1 reacts with copper-oxidized LDL, but minimally with native LDL. Further analysis showed that MAb LO1 also reacts in vitro wit ...[more]

PMID: 22509912

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Project description:Background and aimsAs the incidence of nonalcoholic steatohepatitis (NASH) continues to rise, understanding how normal liver functions are affected during disease is required before developing novel therapeutics which could reduce morbidity and mortality. However, very little is understood about how the transport of proteins and cells from the liver by the lymphatic vasculature is affected by inflammatory mediators or during disease.MethodsTo answer these questions, we utilized a well-validated mouse model of NASH and exposure to highly oxidized low density lipoprotein (oxLDL). In addition to single cell sequencing, multiplexed immunofluorescence and metabolomic analysis of liver lymphatic endothelial cells (LEC)s we evaluated lymphatic permeability and transport both in vitro and in vivo.ResultsConfirming similarities between human and mouse liver lymphatic vasculature in NASH, we found that the lymphatic vasculature expands as disease progresses and results in the downregulation of genes important to lymphatic identity and function. We also demonstrate, in mice with NASH, that fluorescein isothiocyanate (FITC) dextran does not accumulate in the liver draining lymph node upon intrahepatic injection, a defect that was rescued with therapeutic administration of the lymphatic growth factor, recombinant vascular endothelial growth factor C (rVEGFC). Similarly, exposure to oxLDL reduced the amount of FITC-dextran in the portal draining lymph node and through an LEC monolayer. We provide evidence that the mechanism by which oxLDL impacts lymphatic permeability is via a reduction in Prox1 expression which decreases lymphatic specific gene expression, impedes LEC metabolism and reorganizes the highly permeable lymphatic cell-cell junctions which are a defining feature of lymphatic capillaries.ConclusionsWe identify oxLDL as a major contributor to decreased lymphatic permeability in the liver, a change which is consistent with decreased protein homeostasis and increased inflammation during chronic liver disease.

Dataset Information

Model IgG monoclonal autoantibody-anti-idiotype pair for dissecting the humoral immune response to oxidized low density lipoprotein.

Publications

Model IgG monoclonal autoantibody-anti-idiotype pair for dissecting the humoral immune response to oxidized low density lipoprotein.

Similar Datasets

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