I would say even distribution of cells within the gel. Uniform distribution is the key and spacial dispersion dependant upon target tissue would also be important for cell-cell interactions. But also we are in the infancsy stage of bioprinting so we are still learning, furthermore we are still learning more about stem/progenitor cells.

As mentioned by Amin all of the factors that you mention are inter connected. Changing one might have a huge impact on the others. Currently the biggest challenge is producing functional 3D printed constructs which can be used right off the bat.

Assuming that you are referring to 'printing' mammalian cells; none of the above!

Regardless of the technique used to deposit living mammalian cells (of which there are many), the biggest challenge to bio-printing is the 'ink' – i.e. the physicochemical composition and properties of the extracellular material used as the vehicle to carry and deposit the cells. This material dictates both cell survival (including ability to cope with the stressors of printing - shear, compression, decompression, pH, T°C, osmolarity, light energy, etc.) and subsequent behaviour - proliferation, quiescence, differentiation, death (all forms), transdifferentiation, and ultimately ability to form multicellular tissue.

in reference to the above comment all of David's parameters are valid however, producing functional tissue and the requirements of that tissue will add further demands on the parameters. Therefore, under no circumstances are previous comments to be dismissed.

for example cellular dispersion within a printed construct if not dispersed evenly will possibly result in pockets of tissue "like" formation also the cells will add mechanical support to the scaffold. All the above are components of multifactorial system of tissue formation and should all be considered.

I agree with you entirely, William. Recommended reading!

My perspective is that while these parameters are important, however, they are secondary, being critical to the mechanics of the printing process. If the ink is inappropriate, secondary parameters are unimportant, because the process will not deliver the outcome you want!

To find the optimal bioink!

I think it is the blood vessels,especial the capillary connections.

Hydrogel for a specific tissue type and the number of cells required for printing

The rest follows ..

The printability of bioink, the control of multiple cell types for co-culture, and long-term cell culture performance study.

Right now, I would say the size of the bioprinted construct. Because with this scale-up, we need functional vascularization to keep every cells alive... In my opinion to overcome this problem, we need a dynamic maturation, so the use of a bioreactor. And we need a bioreactor dedicated to the transformation of a cellularized hydrogel into a functional tissue (control of pH, mechanical stress, nutiement and growth factor supply, waste evacution, ...)

I think it is 1. material characteristics, 2. hirachical blood supply, and 3. the maturation of the 3D printed object into a mature, tissue specific Matrix in a bioreactor. And 4. the team and its capacities, of course.

I agree with David and William. The bioink composition and the physicochemical properties should match both for printing and the cell survival and functionality.

If the bioink is printable at the abovesaid parameters by maintaining the cells viable, then you can take further for differentiation, maturation in bioreactor and the studies you wish to do. The porosity of the bioprinted construct is very important to keep it functional, which takes cares of the diffusion of nutrients, oxygen and metabolites. As a whole, various factors influence the bioprinting process and they are interlinked.

Hi daer Janaina,

Many thanks for this intersting qeustion.

In my opinion the most challanginh factor is the biocompatibilities to make the scaffold as much as possible close to in vivo nature, in order to ensure normal interactions between the cellular and ECM components.

Best wishes

Sarmad

Cell source

Cell number

Cell choice (stem cells, differentiated, primary)

One of the limits in achieving 3D bioprinted organs for transplant is size. Currently, researchers can create miniaturized tissue resembling natural tissue, but many of these constructs are not capable of achieving therapeutic impact due to their small size.

Read more at: https://www.idtechex.com/research/articles/challenges-on-the-road-to-3d-bioprinted-organs-00011400.asp

Yes Tarun... Clinically relevant size is a big challenge.. other aspects holds the way towards printing in large size.

Bioprinter as well as Bioink physicochemical properties like viscosity.

Dear Janaína Dernowsek,

3-dimensional (3D) printing also called additive manufacturing (AM) has found applications in a variety of industries including construction, food, aerospace and manufacturing. Using 3D bioprinting, researchers were able to construct several different tissues including bone, skin, cartilage, muscle and neural. 3D bioprinting has great advantages in building scaffolds over conventional approaches that it can position the cells precisely. Despite the successful studies and reported outstanding research efforts, the path to fully built a 3D bioprinted organ has yet to be accomplished and there are several challenges to be solved to further advance this exciting research theme.

Challenges

The bioprinter technology needs to increase resolution and speed and should be compatible with a wider spectrum of biocompatible materials. Higher resolution will enable better interaction and control in 3D microenvironment. Currently, printing process is slow so speeding up building the architectures is essential. To reach to a commercially acceptable level that is being able to mass produce requires faster printing and scaling up the process. Biomaterial’s viscosity and crosslinking mechanism determines its printability. The choice of cell source also determines the success of the printed construct.

In addition to technical challenges, there are ethical problems involved in 3D bioprinting as in many other aspects of bioengineering. These are equality, safety and human enhancement.

Ashish