Engineering musculoskeletal tissue by sound

Micro-Pellet patterning within specific hydrogels to control cell sprouting


Highlights

  • Micro-Pellets (cell spheroids) can be patterned in a variety of hydrogels

  • Hydrogel properties can be adjusted to control cell sprouting

  • Custom-tailored mimiX labware allows for simultaneous micro-pellet patterning in 12-well plates

Recent efforts in the field of Tissue Engineering have increasingly considered bottom-up approaches and the assembly of living and non-living building blocks into functional tissues by sound (1).

In this regard, spheroids, or micro-pellets are particulary interesting, since they allow for easy patterning with sound and provide enhanced cell-cell contacts compared to individual cells. Additionally, micro-pellets of mesenchymal stem cells (MSC), combining several hundred cells within a dense cluster, are promising candidates to induce cell lineage commitment to engineer cartilage of clinically relevant size, or vascularised bone tissue constructs (2, 3).

Here, we are exploiting the use of sound waves to pattern micro-pellets into tight assemblies. By using ZAVA, a hydrogel from the mimix biomaterials portfolio with MMP cleavable sites and RGD adhesion peptides, cells can sprout and migrate from the micro-pellet, while cell sprouting and migration is limited in the hydrogel BAKU, which does not contain MMP-cleavable sites or RGD. Overall, cymatiX and the mimiX hydrogel portfolio can be used to address specific needs in tissue engineering.


Figure1

A) Spheroids were formed in AggreWell Plates(StemCellTechnologies) and patterned in B)Fibrin, C) BAKU, D) ZAVA or E)Agarose


 

Figure 2

The hydrogel composition can be tuned in order to control cell sprouting from the spheroids A) with RGD and MMP-cleavable sites, B) without. mimiX offers hydrogels that were specifically designed to meet these criteria. C) High magnification image of spheroids patterned in ZAVA, a hydrogel with RGD and MMP cleavable sites. Cell sprouting from the spheroids is apparent. Spheroids were stained for actin with phalloidin 633.


 

Figure3

To ultimately engineer a vascularised osteochondral construct, green fluorescent protein expressing human umbilical vein endothelial cells(GFP-HUVEC) were patterned into concentric rings (i).Subsequently, micro-pellets formed from MSC were patterned on top (ii) to form a bilayered construct.We envisage to develop truly 3-dimensional, multi-layered biografts by use of sound.


 

Experimental Conditions

Biomaterial: Fibrin,ZAVA, BAKU, Agarose

Cell Type: Endothelial Cells, Human Mesenchymal Stem Cells

Labware: mimiX Labware for 12-well plate or petri dish


References

(1) Guex et al., The waves that make the pattern, Materials Today Bio, 2021.

(2) Futrega et al., A single day of TGF-β1 exposure activates chondrogenic and hypertrophic differentiation

pathways in bone marrow-derived stromal cells Communications Biology, 2021.

(3) Nilsson Hall et al., Patterned, organoid-based cartilaginous implants exhibit zone specific functionality forming osteochondral-like tissues in vivo Biomaterials, 2021.