Synthesis and characterization of polypropylene fumarate and poly[(R)-3 hydroxybutyrate]-based polyurethane scaffolds for tissue engineering / Bushra Naureen

Bushra , Naureen (2022) Synthesis and characterization of polypropylene fumarate and poly[(R)-3 hydroxybutyrate]-based polyurethane scaffolds for tissue engineering / Bushra Naureen. PhD thesis, Universiti Malaya.

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Abstract

Degeneration of living cells/tissues, and organs is a global concern. Biodegradable tissue engineering scaffolds (TES) synthesized from natural or synthetic polymeric biomaterials aid tissue repair. Polyurethanes (PU) are prominent synthetic polymers for TES synthesis. However, PU TES based on polycaprolactone (PCL) and polylactic acid (PLA) polyols have slow degradation, limited biocompatibility (cell adhesion), and hindrance towards growth factors (GF) immobilization due to single functional group (COOR) per monomer unit. Hence, there is a need to explore alternative polyols and chain extenders (CE) to incorporate multiple functional groups per monomer unit of PU TES. This study aimed to synthesize new 3D PU foam scaffolds (PUFS) with more hydrolysis susceptible and crosslinkable functional groups, designed for enhanced/variable in-vitro degradation rate, biocompatibility, and GF immobilization/release (IR) potential. Four 3D PUFS (PU1,PU2,PU3, and PU4) with tunable in-vitro degradation rates were synthesized through addition polymerization reaction between polypropylene fumarate (PPF) the polyol, 1,6-hexamethylene diisocyanate (HDI) and variable concentrations of CE poly[(R)-3-hydroxybutyrate] (P3HB). To enhance PUFS biocompatibility and GF IR potential, PU4 was modified with four concentrations of κ-carrageenan (CMPU: PU4C1,PU4C2,PU4C3, and PU4C4). Different characterization techniques were employed for the analysis of PPF (GPC,FTIR,1H and 13C NMR) and PUFS (FTIR,1H and 13C NMR,FESEM,TGA/DTA and 70 days in-vitro degradation analysis (n=8)). Moreover, PU4 and CMPU were examined via FTIR and EDX. The biocompatibility of PU4 and CMPU scaffolds (n=4) were investigated by in-vitro cell culture (C2C12 myoblast) through (%) resazurin reduction assay (till day-21), live/dead, and SEM images (both up to 14 days). The GF(IGF-1) IR potential of PU4 and CMPU (n=3) was determined by ELISA kits and microplate reader for 14 days. GPC provided the molecular weight (Mn = 455.67 g mol-1, Mw = 587.15 g mol-1) and PDI (1.29) of the PPF. The results of FTIR or 1H and 13C NMR spectra confirmed the synthesis of PPF (C=C,COOR), PUFS (C=C,COOR,NHCOO) and CMPU (C=C,COOR,NHCOO,O=S=O,C-O-C,O-SO3-) with multiple functional groups. The EDX spectra presented an increase of sulfur (%) composition from 0.24% to 4.0% with the increase of incorporated κ-carrageenan in PU4. The PUFS FESEM images revealed pores (242-340μm) and void spaces (735-860μm) beneficial for TE. The PUFS (PU1,PU2,PU3,PU4) TGA/DTA curves showed an onset thermal decomposition temperature of 267.17ºC, 267.28ºC, 256.0ºC, and 240.72ºC, respectively. The results of day-70 PUFS in-vitro degradation analysis revealed a significant difference between the weight loss (%) (PU1=4.09±0.18%, PU2=8.03±0.17%, PU3=13.14±0.12%, and PU4=20.21±0.19%). Thus, confirmed the formation of PUFS with variable degradation rates (PU4>PU3>PU2>PU1) based on incorporated P3HB concentration (hydrolyzable COOR). The results of biocompatibility assessment day-21 (%) resazurin reduction (PU4=67.54±7.68%, PU4C1=71.40±5.07%, PU4C2=75.79±6.10%, PU4C3=82.42±6.82% and PU4C4=83.37±7.68%), live/dead and SEM images (day 1,7,14) proposed that C2C12 myoblasts adhesion, proliferation, metabolic activity, and viability on PUFS followed a trend (PU4C4>PU4C3>PU4C2>PU4C1>PU4). A similar trend was depicted from the results of IGF-1(ng ml-1) release from PU4 and CMPU scaffolds at all timepoints (day 1,3,7,10,14), suggesting successful synthesis of PUFS with enhanced biocompatibility and IGF-1 release potential, i.e., directly related to the incorporated κ-carrageenan concentration.

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