## 3 Electrospinning
### Biomimetic Applications of Electrospun Nano Structures for Tissue Engineering Purposes
Dual extrusion electrospinning is one such example, which permits two different polymeric solutions to be electrospun simultaneously using different extrusion conditions [85]. This was performed by [PERSON] et al. whose hybrid scaffold made of silk fibroin nanofibers and PCL microfibers showed an increase in osteoblast cell differentiation [86]. Coaxial electrospinningning is another technique which uses two different polymer solutions. However, they are fed into the spinneret as inner and outer fluid to produce fibers with a different core and shell composition [87]. This feature has increased its significance not only in the field of tissue engineering but also in drug delivery, as these fibers have the capacity to encapsulate drugs, growth factors and antibiotics [87]. Temperature-assisted electrospinningning methods such as cryogenic electrospinningning and melt electrospinning are also used and can program the desired porosity into the scaffold [88; 89]. Cryogenic electrospinningning method uses ice crystals, spun along with the fibrous scaffold, which when removed by freeze drying leave behind voids through which cells can infiltrate [88]. The porosity can be controlled based on the size and amount of the ice crystals used [88]. Temperature-assisted electrospinningning, on the other hand, uses a melt blowing technology, where the polymer is melted with high temperature and is extruded out using controlled air flow [89]. This method can produce aligned fibers with desired porosity and hence was used to replicate the ECM environment of tendons [89; 90]. However, none of the above methods provide complete control over the fiber alignment and its 3D deposition. This has led researchers to use 3D printing technology in electrospinning [91; 92]. Near field-electrospinning is a relatively new method that prevents the bending and splitting instabilities of the electrospun fibers by reducing the distance between the spinneret and collector to a bare minimum (<3 mm). [PERSON] et al. included a 3D printing technology into this method by controlling the fibers deposition in a pre-programmed X-Y path [91]. [PERSON] et al. went a step further and used X-Y-Z moving stage (which replicates the technology used in hobbyist grade 3D printers) to produce a highly organized scaffold with 3D architecture [92]. More recently, a technique referred to as 3D jet writing a modified electrospinningning process, which yields 3D structures and scaffolds that have been shown to support the 3D expansion and differentiation of human mesenchymal stem cells in vitro (Figure 1) [93]. All the above examples clearly indicate the rapid progression of research in the field of electrospinning and scaffold technology to produce a structurally programmable and low-cost ECM or BM mimics for tissue engineering applications. Figure 1: In vitro hMSC culture on 3D jet writing scaffolds produces 3D microtissues. (**A**,**B**) 3D microfiber scaffolds were incubated with fibronectin and seeded with hMSCs to yield confluent cell structures. (**C**–**E**) hMSCs cultured on scaffolds (**E**) reveal significantly different morphologies and spatial distribution (3D) than cultures on glass (**C**), or nonwoven PLGA fiber mats (**D**) (both 2D). (**F**–**H**) hMSCs on scaffolds were incubated in osteogenic differentiation media and monitored using qPRC. The markers SP7 (**F**), RUNX2 (**G**), and BSP (**H**) were used as indicators of osteogenic differentiation. Values reported are fold increases over hMSCs cultured in growth medium. Large increases in these markers after three weeks are consistent with the onset of osteogenesis. Error bars represent \(\pm\) Std. Dev from three independent experiments. (**I**) Fluorescent staining of scaffolds for hydroxyapatite reveal substantial increases in matrix mineralization after two weeks of differentiation.