Glasgow Researchers Explore Stem Cell Engineering with Bacteria Filled Microgels

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Researchers from the University of Glasgow are expanding their research into bioprinting, with their findings outlined in the recently published ‘Bacteria laden microgels as autonomous 3D environments for stem cell engineering.’ The team has developed a microfluidic system comprised of one step, allowing both stem cells and genetically engineered non-pathogenic bacteria to be contained in an alginate microgel.

While most techniques rely on droplet extrusion, here the researchers are creating a more efficient system via a one-step droplet microfluidic method. Fabrication of the pearl-lace microgels occurs at physiological pH without any sheathing material, with channel dimensions and overall design meant to avoid shear stress on the cells and encourage viability.

“The fabricated gel-construct is unique in a way that it has both compartmentalized units as in individual microcapsules as well as the connectivity found in fibrous constructs,” state the researchers. “It is also noted that the compartmentalized microunits and the link connecting them are highly tunable resulting in highly mono-dispersed pearl-lace interlinking structures.”

Droplet-based microfluidic setup. (A) Schematic representation of the microfluidic device and encapsulation of prokaryotic and eukaryotic cells. (B) Image of the capillary based microfluidic device. (C) A snapshot of pearl formation in microfluidic device with indication of parameters used to quantify assembled pearls. (D) Thread thickness graph with corresponding flow rates (Y-axis: water flow; X-axis: oil flow). (E) Thread thickness graph. (F) Pearl area graph.

For this project, the researchers created an in vitro 3D model ‘for investigating the commensalism symbiosis between eukaryotic (bone marrow mesenchymal stem cells, hBM-MSC) and prokaryotic cells (engineered non-pathogenic bacteria Lactococcus lactisL. lactis).’

While bacteria are commonly used as an affordable ‘production organism’ for proteins in bioprinting, they can also act as a mechanism for directing both cell growth and differentiation. The researchers also used a bacteriostatic antibiotic, sulfamethoxazole, to prevent growth of harmful bacteria.

Four 3D printed shapes were fabricated, including a line, triangle, square, and circle, and arrangement as follows:

  • Line – two circular discs (180-degree angle)
  • Three for triangle (60-degree internal angle)
  • Four for square (90-degree internal angle)
  • Eight for a circle-like shape (135-degree internal angle, octagon)

The microfluidic systems allowed the researchers to create ‘mono-disperse’ constructs which are suitable for applications like pharmacological screenings, biological studies, and personalized medicine.

SEM images of cell-laden alginate constructs. Phase contrast images of alginate microgels with MSC in basal media (A); alginate microgel with MSC in osteogenic media (B); alginate microgel with MSC containing two colonies of L. lactis (C) either expressing FNIII 7-10 or BMP-2 in a constitutive manner. Samples were fixed after two weeks of culture. Scale bar: 100 μm. SEM images of alginate construct with MSC in basal media, the images show mark impressed by cell on the cross section of an alginate construct (D); alginate microgel with MSC and two colonies of L. lactis overpopulating the space (E); alginate microgel with MSC in osteogenic media, round entities covering the cells, cavities and fine membrane-like constructs (F). The hydrogels were slightly dehydrated/shrunken compared to their state in aqueous media

“The connectivity of pearl-lace hydrogels can provide a way of gradient studies in which the population of each cell type and so its relative density can be controlled. It can also be utilized for time-series indexing studies as well as providing a mean for a low-cost, easy to fabricate 3D bio-printing prototypes as demonstrated in this study,” concluded the researchers.

“Microgel in this study has been utilized as a proof of concept for modeling a tuneable platform in which both hydrogel acting as an ECM as well as production and release of growth factor can be both engineered at a low cost with high precision spatio-temporal control. It has been an attempt to further engineer more aspects of an in vitro system, paving the way for study of cells and their interaction with adjustable dynamic ECM-like environment with greater control. “

As the progress of bioprinting continues to take hold in global research, scientists create new bioprinting inks, 3D printed microsurfaces, progressive microfluidic techniques, and more. What do you think of this news? Let us know your thoughts! Join the discussion of this and other 3D printing topics at 3DPrintBoard.com.

Mono-dispersity, encapsulation and viability of micro-beads. (A) Pearl-size (long axis) distribution of produced hydrogel of two miscible fluid streams under laminar flow conditions using flow rates of 500 μl h-1 for the two inner phases and 5000 μl h-1 for the outer phase. N ≥ 5 – 10 microgels were analysed for each condition. The mean length of the formed pearl was 167 μm with a RSD of 3.2%. Scale bar: 100 μm. (B) Cell (MSC and L. lactis) encapsulation efficiency by the formed hydrogel. The cell counts at each time point are the result of 8 measurements sequentially acquired at 30-minutes intervals at room temperature for 2 hours. (C) Fluorescent images of two weeks old alginate hydrogels with L. lactis and MSC. The hydrogel was stained with BacLight Bacterial Viability Kit for L. lactis and Viability/Cytotoxicity Kit, for MSC. Both kits stain viable cells in green (SYTO® 9 and Calcein AM) and non-viable cells in red (Propidium iodide and Ethidium homodimer-1), a 50:50 mixture of the kits were used for co-culture. Scale bar: 100 μm. (D) The viability graph of L. lactis, MSCs and co-culture.

[Source / Images: Bacteria laden microgels as autonomous 3D environments for stem cell engineering]

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