Fabrication of Nanofibrous Scaffold Grafted with Gelatin Functionalized Polystyrene Microspheres for Manifesting Nanomechanical Cues of Stretch Stimulated Fibroblast.

Cells sense mechanical cues in their microenvironments and effectively transduce them into intracellular signaling events, which subsequently orchestrate adaptive changes in cells cytoskeletal components and other adhesion proteins. Thus far, although diverse nanostructured scaffold systems have been utilized for evaluating such mechanical cues, most of them do not recapitulate the in vivo conditions (ECM matrix, 3D culture), which by themselves obscure natural cellular responses in the first place. To address this drawback, in the present work, a hybrid aligned nanofibrous scaffold grafted with gelatin functionalized polystyrene microspheres has been adapted for assessing fibroblast cells mechanobiology in response to mechanical stimulation by stretch cycles. In this work, with the help of a rationally designed hybrid scaffold, we present clear evidence of that fact that fibroblast cells when subjected to mechanical stress tend to manifest higher magnitude traction force on the underlying hybrid scaffold, which consequently introduces proportional distortion in orientation of aligned nanofibers scaffold as well as the polystyrene microspheres. A clear insight into sequestration of focal adhesion proteins and cellular cytoskeletal dynamics that actuate such changes was perceived and quantified by confocal microscopy, field emission-scanning electron microscopy, flow cytometer, and atomic force microscopy (peak force QNM). The mechanical stress induced alteration in regulation of genes associated with focal adhesion proteins and extracellular matrix proteins was also quantified by real-time polymerase chain reaction. In summary, the work presents a biologically relevant 4D hybrid scaffold and validates its implications for evaluation of cell mechanobiology.