Development of a pH-responsive nanofibrous scaffold for therapeutic applications in bone tissue engineering: A comparative evaluation of three-layered vs single-layered fibers

Bone tissue engineering requires multifunctional scaffolds that mimic the extracellular matrix while providing controlled therapeutic delivery. In this study, a three-layered electrospun fibrous scaffold based on a tri-polymer/silica matrix loaded with tetracycline hydrochloride (TC) was fabricated using sequential electrospinning for pH-responsive drug delivery. Silica was prepared through the sol-gel route. Characterisation, including XRD, FTIR, and FESEM, confirmed the successful formation of layered and interconnected fibers with adequate porosity, wettability, mechanical strength and thermal stability. Scaffolds effectively inhibited the growth of E. coli and S. aureus and showed excellent compatibility with blood (hemolysis < 2%), cell viability, enhanced ALP activity (60.47 ± 2.7 U/g), and calcium mineralization through ARS staining. A prolonged release of TC was achieved over 292 h, in a controlled manner, showing a stimulated release at pH 5.5 compared with pH 7.4. This indicated a targeted and pH-responsive release at infected and inflamed bone defect regions. Kinetics followed a concentration-dependent release with a Fickian diffusion mechanism. The prepared three-layered fibers were compared with a single-layered counterpart to evaluate the performance. Overall, the developed three- layered scaffold serves as a biomimetic platform for targeted stimuli-responsive delivery applications in bone tissue engineering.