Serum amyloid P component-DNA complexes are decreased in systemic lupus erythematosus. inverse association with anti-dsDNA antibodies.

OBJECTIVE: To study serum levels of serum amyloid P component (SAP) and SAP-DNA complexes in a population-based cohort of patients with systemic lupus erythematosus (SLE). METHODS: The study population comprised 82 unselected patients of predominantly Scandinavian ancestry with SLE according to current classification criteria. Serum samples were collected at baseline and serially for up to 2 years. SAP component and SAP-DNA complexes were measured by ELISA. Associations between SAP-DNA and clinical manifestations or serological findings were analyzed. Ninety healthy, age-matched blood donors served as controls. RESULTS: SLE patients had normal serum concentrations of SAP, whereas SAP-DNA complexes were decreased. Two-thirds of the SLE patients tested persistently SAP-DNA complex-negative. There was no relationship between the occurrence of SAP-DNA complexes and clinical manifestations. SAP-DNA-negative patients tended to have lower leukocyte counts and complement C3 levels, and higher erythrocyte sedimentation rates and C3d levels versus SAP-DNA-positive patients. There was an inverse association between the occurrence of anti-double-stranded DNA (anti-dsDNA) antibodies and SAP-DNA complexes. Co-occurrence of SAP-DNA complexes and anti-dsDNA antibodies was demonstrated in only one SLE patient, implying that 81/82 patients were discordant for the presence of anti-dsDNA antibodies and SAP-DNA complexes. CONCLUSION: The decreased level of SAP-DNA complexes in SLE patients and the inverse relationship between these complexes and anti-dsDNA antibody supports the concept that SAP component is implicated in the clearance of cell nuclear debris.

Recombinant human serum amyloid P component from Pichia pastoris: production and characterization.

Human serum amyloid P component (SAP) was expressed in the methylotrophic yeast Pichia pastoris. SAP cDNA was placed under control of regulatory sequences derived from the alcohol oxidase gene (AOX1), and its protein product was secreted using the Saccharomyces cerevisiae alpha-mating factor signal sequence. Recombinant SAP (r-SAP) was produced in a bioreactor with computer controlled fed-batch mode and purified by use of a C-terminal histidine tag. The yield of purified r-SAP was 3-4mg from 1L supernatant and 5-6mg from 1L cell paste, indicating that the majority of the produced SAP was not secreted. Treatment of the cell paste with EDTA increased the yield further by about 30%. The N-terminal of r-SAP purified from the supernatant showed non-complete cleavage of the alpha-mating factor signal sequence. Purified r-SAP, analyzed under native conditions, was shown to be a decamer, like purified human SAP (h-SAP), with monomers of 27kDa. Each monomer had one N-glycosylation site, positioned at the same site as for h-SAP. r-SAP bound to antibodies produced against h-SAP. Furthermore, r-SAP bound to ds DNA and influenza A virus subunits in a Ca(2+)-dependent manner and inhibited influenza A virus hemagglutination. These results indicate that r-SAP produced in P. pastoris has the same biological activity as purified h-SAP.

Only amyloidogenic intermediates of transthyretin induce apoptosis.

In diseases like Alzheimer's disease and familial amyloidotic polyneuropathy (FAP) amyloid deposits co-localize with areas of neurodegeneration. FAP is associated with mutations of the plasma protein transthyretin (TTR). We can here show an apoptotic effect of amyloidogenic mutants of TTR on a human neuroblastoma cell line. Toxicity could be blocked by catalase indicating a free oxygen radical dependent mechanism. The toxic effect was dependent on the state of aggregation and unexpectedly mature fibrils from FAP-patients who failed to exert an apoptotic response. Morphological studies revealed a correlation between toxicity and the presence of immature amyloid. Thus, we can show that toxicity is associated with early stages of fibril formation and propose that mature full-length fibrils represent an inert end stage, which might serve as a rescue mechanism.