Assessment of Scaffolding Properties for Chondrogenic Differentiation of Adipose‐Derived Mesenchymal Stem Cells in Nasal Reconstruction

Abstract

IMPORTANCE Nasal reconstruction in patients who are missing a significant amount of structural nasal support remains a difficult challenge. One challenge is the deficiency of cartilage left within the nose as a consequence of rhinectomy or a midline destructive disease. Historically, the standard donor source for large quantities of native cartilage has been costal cartilage.OBJECTIVE To enable the development of protocols for new mesenchymal stem cell technologies as alternative procedures with reduced donor site morbidity, risk of infection and extrusion. DESIGN, SETTING, AND MATERIALSWe examined 6 popular scaffold materials in current practice in terms of their biodegradability in tissue culture, effect on adipose-derived mesenchymal stem cell growth, and chondrogenic fate commitment. Various biomaterials of matching size, porosity, and fiber alignment were synthesized by electrospinning and overlaid with rabbit adipose-derived mesenchymal cells in media supplemented or not with chondrogenic factors. Experiments were performed in vitro using as end points biomarkers for cell growth and chondrogenic differentiation. Polydioxanone (PDO), poly-3hydroxybutyrate-co-3-hydroxyvalerate (PHBV), PHBV-polycaprolactone, poly(L-lactide-cocaprolactone), poly(lactic-co-glycolic acid), and polystyrene scaffolds of 60% to 70% porosity and random fiber alignment were coated with poly(L)-lysine/laminin to promote cell adhesion and incubated for 28 days with 2.5 to 3.5 × 10 5 rabbit adipose mesenchymal cells. MAIN OUTCOMES AND MEASURESCell growth was measured by fluorometric DNA quantitation and chondrogenic differentiation of stem cells by spectrophotometric sulfated glycosaminoglycan (sGAG) assay. Microscopic visualization of cell growth and matrix deposition on formalin-fixed, paraffin-embedded tissue sections was performed, respectively, with nuclear fast red and Alcian blue. RESULTSOf 6 scaffold materials tested using rabbit apidose mesenchymal cells, uncoated scaffolds promoted limited cell adhesion but coating with poly(L)-lysine/laminin enabled efficient cell saturation of scaffold surfaces, albeit with limited involvement of scaffold interiors. Similar growth rates were observed under these conditions, based on DNA content analysis. However, PDO and PHBV/PCL scaffolds supported chondrogenic fate commitment better than other materials, based on soluble sGAG analysis and microscopic observation of chondrogenic matrix deposition. The mean (SD) sGAG scaffold values expressed as fold increase over control were PDO, 2.26 (0.88), PHBV/PCL, 2.09 (0.83), PLCL, 1.36 (0.39), PLGA, 1.34 (0.77), PHBV, 1.07 (0.31), and PS, 0.38 (0.14). CONCLUSIONS AND RELEVANCEThese results establish materials, reagents, and protocols for tissue engineering for nasal reconstruction using single-layer, chondrogenically differentiated, adipose-derived mesenchymal stem cells. Stackable, scaffold-supported, multisheet bioengineered tissue may be generated using these protocols.LEVEL OF EVIDENCE NA.

Applications of Electrospinning for Tissue Engineering in Otolaryngology

“…26 San Marina and colleagues constructed electrospun scaffolds of 5 different polymer blends, with preliminary findings suggesting polydioxanone (PDO) and poly-3-hydroxybutyrate-co-3-hydroxyvalerate (PHBV), with and without the addition of PCL, showed the greatest amount of matrix deposition when a nasal chondrogenic supplement was applied in an in vitro model (Table 1). 27…”

Polydioxanone-Based Membranes for Bone Regeneration

“…PDO is a safe synthetic graft material and is well known as a guide for tissue regeneration to stimulate bone and cartilage development in vivo [ 19 , 46 , 47 , 48 , 49 ]. PDS ® as an absorbable barrier material (polydioxanone, Johnson & Johnson, Ethicon GmbH & Co KG, Norderstedt, Germany) was used for periodontal regenerative surgery, which helped to eliminate a second surgical procedure for the removal of non-absorbable membranes [ 49 ].…”

Tissue-Engineered Grafts from Human Decellularized Extracellular Matrices: A Systematic Review and Future Perspectives

“…To date, nasal reconstruction in patients who are missing a significant amount of structural nasal support remains a difficult challenge. Historically, the standard donor source for large quantities of native cartilage has been costal cartilage [41] but some studies also proposed tissue engineering approaches from human cadaver nasal cartilages [42]. Schwarz et al [32] developed a cost-effective and simple decellularization method based on osmotic treatment in distilled water (dH 2 O) and standard chemicals (sodium hydroxyde, ethanol, guanidine hydrochloride, sodium acetate, and hydrogen peroxide), without any enzymatic digestion.…”