*3.2.2. Ligands and ligand densities*

**Figure 4. Schema of renoprotective effects of ADSCs and synthetic ECM (CS-IGF-1C hydrogel).** When co-trans‐ planted into AKI model, CS-IGF-1C hydrogel could protect delivered ADSCs, facilitated their paracrine and anti-in‐ flammatory effects, and inhibit ECM synthesis in kidney, which result in enhanced angiogenesis, regeneration and alleviated fibrosis after kidney injuries. Consequently, CS-IGF-1C hydrogel therapy leads to improved functional and

Although it has commonly acknowledged that signals transduced by ECM could direct stem cell fate, there is increasing evidence that physical properties of ECM could also make a great impact on cell behaviours [48–50]. Some of these factors are proven to be of great influence, but we still have a long way to go and a lot of work to do to establish a complete theory. For example, in response to injury, the accumulation of ECM is excess and abnormal, which would cause significant changes to the stiffness of ECM and ultimately lead to tissue fibrosis [51,52].

To test the effect of different stiffness (EY) on cell behaviors, substrates with EY ranging from <1 kPa to 30 kPa were synthesized [53]. The results showed that ECM stiffness has influence on cell proliferation as well as cell differentiation. For instance, neural stem/progenitor cells (NSPCs) could proliferated on substrates with EY <10 kPa. On soft substrates (<1 kPa), neuronal differentiation was promoted; whereas, on relatively stiff substrates (>7 kPa), oligodendrocyte

structural recovery of kidney [47]. Reprinted by permission of the publisher.

328 Composition and Function of the Extracellular Matrix in the Human Body

**3.2. Physical interaction**

*3.2.1. Stiffness and elasticity*

When cells were seeded on ECM with different ligand densities, changes in stem cell viability, size, and shape provided the direct evidence that ligand immobilized to ECM could not be easily separated from the biophysical effects of matrix [58,59]. The spatial arrangement of ligands had a significant influence on MSC behavior [44]. Through manipulating of the ratio of polystyrene-block-poly (ethylene oxide) copolymers (PS-PEO-Ma) in mixtures of block copolymer and polystyrene homopolymer, the lateral spacing of RGD (arginine-glycineaspartic acid) peptides was under control. With increased lateral spacing, osteogenesis of MSC was reduced while adipogenic differentiation was increased, which was consistent with the results of gene expression levels and alkaline phosphatase activities.

Moreover, the type of ligands covalently linked to ECM could also influence stem cell fate determination. The differentiation of MSCs on different composition of adhesion ligands with the same concentration was various. MSCs cultured on fibronectin or laminin matrices tended to undergo adipogenic differentiation; whereas, MSCs cultured on ECM containing collagen preferred to adopt a neurogenic outcome [60].

#### *3.2.3. Macro/nano-scale topography*

Recent development also demonstrated that macro/nano-scale of ECM was another important physical parameter that could not only change the shape of stem cells but also influence the behaviour of stem cells. A preliminary study demonstrated Zyxin played an important role in nanotopographical feature-facilitated changes in stem cells [48]. On 350 nm grating, expression of Zyxin was down-regulated, which was associated with the accelerated speed of migration and the decreased intracellular tension. Likewise, McMurray et al. revealed that modification in surface nanotopography of thermoplastic polycaprolactone (PCL) would influence intra‐ cellular tension, which could maintain the multipotency of stem cells and diminish spontane‐ ous differentiation of MSC [61]. Moreover, his current study further illustrated that nanoscale spatial organization of cell-adhesive ligands bound to ECM could affect lineage commitment of MSCs [62]. By using nanopatterning techniques, arginine-glycine-aspartate (RGD) was covalently linked to the surface of poly (ethyleneglycol) (PEG) hydrogels with different nanospacing. It was interesting to identify that large RGD nanospacing was beneficial for osteogenesis; small RGD nanospacing was conducive to adipogenesis.
