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Embedding methods for poly(L-lactic acid) microfiber mesh/human mesenchymal stem cell constructs
D D'Alessandro1, B Battolla, L Trombi
1CUCCS-RRMR, Center for the Clinical Use of Stem Cells-Regional Network of Regenerative Medicine, University of Pisa, Italy.
Micron (Oxford, England : 1993)
|April 3, 2009
Summary
This study presents a new histological method for analyzing poly(L-lactic acid) (PLLA) microfiber scaffolds seeded with human mesenchymal stromal cells (hMSCs). The method confirms cell viability and provides insights into cell distribution and extracellular matrix production in tissue engineering constructs.
Area of Science:
- Tissue Engineering
- Biomaterials Science
- Cell Biology
Background:
- Poly(L-lactic acid) (PLLA) fiber mesh scaffolds are explored for tissue repair, supporting stem cell growth and differentiation.
- Existing studies on PLLA microfiber/cell constructs lack detailed histological analysis, limiting understanding of cell-scaffold interactions and extracellular matrix (ECM) production.
- Histological evaluation is crucial for assessing cellular distribution, cell-scaffold integration, and ECM synthesis within engineered tissues.
Purpose of the Study:
- To develop and validate a robust histological and histochemical method for analyzing poly(L-lactic acid) (PLLA) microfiber/cell constructs.
- To investigate the compatibility and behavior of bone marrow-derived human mesenchymal stromal cells (hMSCs) within PLLA microfiber scaffolds over time.
Main Methods:
- Biodegradable non-woven fiber meshes were fabricated using hollow PLLA microfibers.
- Various embedding techniques (paraffin, Killik, acrylic resin) were evaluated; acrylic resin proved most suitable for histological analysis.
- PLLA microfiber/hMSC constructs were embedded in acrylic resin and analyzed using Toluidine Blue, Periodic Acid-Schiff (PAS), and Alcian Blue staining, alongside MTT and CCK-8 assays for cell viability.
Main Results:
- Acrylic resin embedding successfully preserved the structural integrity of PLLA microfiber/hMSC constructs for histological analysis.
- Histochemical staining confirmed cell viability and proliferation up to 6 weeks, consistent with MTT and CCK-8 data.
- Staining revealed detailed information on cell distribution, organization within the scaffold, and the production of ECM molecules, enriching previous SEM and quantitative findings.
Conclusions:
- A validated histological and histochemical method using acrylic resin embedding is proposed for analyzing PLLA microfiber/cell constructs.
- The study confirms the compatibility of PLLA microfibers with hMSCs, demonstrating sustained cell viability and ECM production.
- This methodological advancement provides deeper insights into cell-scaffold interactions, crucial for advancing PLLA-based tissue engineering applications.
