Study of myoblast differentiation using multi-dimensional scaffolds consisting of nano and micropatterns

Abstract

BackgroundThe topographical cue is major influence on skeletal muscle cell culture because the structure is highly organized and consists of long parallel bundles of multinucleated myotubes that are formed by differentiation and fusion of myoblast satellite cells. In this technical report, we fabricated a multiscale scaffold using electrospinning and poly (ethylene glycol) (PEG) hydrogel micropatterns to monitor the cell behaviors on nano- and micro-alignment combined scaffolds with different combinations of angles.ResultsWe fabricated multiscale scaffolds that provide biocompatible and extracellular matrix (ECM)-mimetic environments via electrospun nanofiber and PEG hydrogel micro patterning. MTT assays demonstrated an almost four-fold increase in the proliferation rate during the 7 days of cell culture for all of the experimental groups. Cell orientation and elongation were measured to confirm the myogenic potential. On the aligned fibrous scaffolds, more than 90% of the cells were dispersed ± 20° of the fiber orientation. To determine cell elongation, we monitored nuclei aspect ratios. On a random nanofiber, the cells demonstrated an aspect ratio of 1.33, but on perpendicular and parallel nanofibers, the aspect ratio was greater than 2. Myosin heavy chain (MHC) expression was significantly higher i) on parallel compared to random fibers, ii) the 100 μm compared to the 200 μm line pattern. We confirmed the disparate trends of myotube formation that can be provoked through multi-dimensional scaffolds.ConclusionWe studied more favorable environments that induce cell alignment and elongation for myogenesis by combining nano- and micro-scale patterns. The fabricated system can serve as a novel multi-dimensional platform to study in vitro cell behaviors.

Microcarriers for Upscaling Cultured Meat Production

“…Therefore, shear stresses generated during a microcarrier based dynamic culture could be tailored to meet the stimulus required for differentiation, through the tuning of operational parameters, or the design of MCs that allow for the culture of cells onto regions of controlled shear stress, as has been recently reported by Wu et al (195). Micro-patterned MCs providing topographies favoring myogenic differentiation, such as aligned patterns (93) could also potentially support a one-step bioprocess.…”

“…C2C12 cells cultured on a micropatterned surface including pillars showed better cell attachment whereas proliferation and spreading were higher on non-patterned surfaces (85). Better proliferation of C2C12 cells on a randomly oriented nanofibers was observed than on aligned ones, nevertheless, a better fusion and alignment of myoblasts was observed on the latter (93). There is consensus that nanofibrous surfaces, by mimicking the extracellular matrix (ECM), are promoting cell attachment and proliferation (94).…”

“…The necessary nutrients can be provided through a switch from a "proliferation medium" to a "differentiation medium, " but providing the physical environment that the cells need in order to differentiate is more challenging. The substrate requirements for the proliferation and differentiation phases are typically different in terms of surface chemistry and topography (88,91,93). Stiffness requirements on the other hand shouldn't be difficult to combine for the proliferation and differentiation phase.…”

The fabrication of 3D surface scaffold of collagen/poly (L‐lactide‐co‐caprolactone) with dynamic liquid system and its application in urinary incontinence treatment as a tissue engineered sub‐urethral sling: In vitro and in vivo study

“…Electrospinning is reported to be an efficient and economical process to fabricate scaffolds for tissue engineering applications. One of the advantages of electrospun fibers is the high surface‐to‐volume ratio which is extremely suitable for cell growth . Promising as electrospinning technique is, the fabrication of scaffolds with controlled 3D property remains a problem, due to its nature of layer by layer deposition pattern of the nanofibers.…”

“…One of the advantages of electrospun fibers is the high surface-tovolume ratio which is extremely suitable for cell growth. 15 Promising as electrospinning technique is, the fabrication of scaffolds with controlled 3D property remains a problem, due to its nature of layer by layer deposition pattern of the nanofibers. To solve this problem, previous methods have included salt leaching, ice crystal formation, sacrificial fibers or increasing the fiber diameter.…”