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Human SAA1-derived amyloid deposition in cell culture: a consistent model utilizing human peripheral blood
Wataru Ishii1, Juris J Liepnieks, Toshiyuki Yamada
1Department of Medicine (Neurology and Rheumatology), Shinshu Unviersity School of Medicine, Matsumoto, Japan.
Abstract:
Amyloid A (AA) amyloidosis is a fatal disease caused by extracellular deposition of fibrils derived from serum AA (SAA). AA amyloid fibril formation has previously been modeled in macrophage cultures using highly amyloidogenic mouse SAA1.1, but attempts to do the same with human SAA invariably failed. Our objective was to define conditions that support human SAA-derived amyloid formation in peripheral blood mononuclear cell (PBMC) cultures. Two conditions were found to be critical - omission of fetal calf serum and use of StemPro34, a lipid-enriched medium formulated for hematopoietic progenitor cells. Cultures maintained in serum-free StemPro34 and provided with recombinant human SAA1 in the complete absence of amyloid-enhancing factor exhibited amyloid deposition within 7 d. Amyloid co-localized with cell clusters that characteristically included cells of fibrocytic/dendritic morphology as well as macrophages. These cells formed networks that appeared to serve as scaffolding within and upon which amyloid accumulated. Cells in amyloid-forming cultures demonstrated increased adherence, survival and expression of extracellular matrix components. Of the three human SAA1 isoforms, SAA1.3 showed the most extensive amyloid deposition, consistent with it being the most prevalent isoform in Japanese patients with AA amyloidosis. Attesting to the reproducibility and general applicability of this model, amyloid formation has been documented in cultures established from eight PBMC donors.
Insights
Researchers developed a new method to model human amyloid A (AA) amyloidosis using peripheral blood mononuclear cells (PBMC). This breakthrough allows for studying AA amyloid fibril formation in vitro, crucial for understanding this fatal disease.
Area of Science:
- Biochemistry
- Cell Biology
- Pathology
Background:
- Amyloid A (AA) amyloidosis is a fatal condition characterized by the extracellular deposition of serum amyloid A (SAA) protein fibrils.
- Previous attempts to model human AA amyloidosis in vitro using macrophage cultures with human SAA were unsuccessful.
Purpose of the Study:
- To establish conditions supporting the formation of amyloid fibrils from human SAA in peripheral blood mononuclear cell (PBMC) cultures.
- To develop a reliable in vitro model for studying human AA amyloidosis.
Main Methods:
- Utilized peripheral blood mononuclear cells (PBMC) cultured in serum-free StemPro34 medium.
- Introduced recombinant human SAA1 to the PBMC cultures without amyloid-enhancing factors.
- Incubated cultures for 7 days to observe amyloid deposition.
Main Results:
- Successful induction of amyloid deposition in human SAA-treated PBMC cultures within 7 days.
- Amyloid co-localized with cell clusters, including fibrocytic/dendritic cells and macrophages, which formed a scaffolding network.
- SAA1.3 isoform demonstrated the most significant amyloid deposition.
- The model proved reproducible across cultures from multiple PBMC donors.
Conclusions:
- Defined critical conditions (serum-free StemPro34 medium) for in vitro human SAA-derived amyloid formation in PBMC cultures.
- Identified specific cell types and their interactions that facilitate amyloid deposition.
- This novel model provides a platform for investigating the mechanisms of human AA amyloidosis and testing therapeutic interventions.
