Tissue engineering offers excellent opportunities, such as creating mechanically viable structures in vitro that can functionally replace damaged or diseased tissues in vivo.
From this point of view, bone tissue engineering is very promising. Indeed, it is possible to promote the growth of bone-forming cells through seeding systems on three-dimensional scaffolds within bioreactors.
There are several bioreactor technologies for culturing engineered tissues for in vivo implantation. These include spinner flask bioreactors.
Spinner Flask Bioreactors, what they are and how they work
Spinner flask bioreactors are often used to seed cells on scaffolds and obtain constructs for bone tissue regeneration.
This is a type of bioreactor with a very simple design.
Inside the culture chamber, the scaffolds are attached to needles secured to the lid, immersed in a liquid-state culture medium. At the bottom of the chamber is a magnetic stirring rod that, as it rotates, imparts a convective force that allows continuous mixing of the liquid surrounding the scaffolds.
In this way, the cells remain in suspension and the movement of the fluid promotes nutrient transport.
Advantages of Spinner Flask bioreactors for bone tissue engineering
In the study “Formation of three-dimensional cell/polymer constructs for bone tissue engineering in a spinner flask and a rotating wall vessel bioreactor” (Sikavitsas, Bancroft et Mikos, 2001), promising data were recorded on the use of the spinner flask bioreactor for the development of cartilage and bone tissue equivalents.
This study investigates the cell culture conditions of three-dimensional polymeric scaffolds seeded with rat marrow stromal cells (MSCs) grown in three different bioreactors: spinner flask, rotating wall vessel and static culture.
In particular, compared with static culture, spinner flask culture demonstrated:
- Day 7: 60% increase in cell proliferation;
- Day 14: all cell/polymer constructs showed the highest alkaline phosphatase (AP) activity, with a 2.4 times higher value than the constructs cultured under static conditions;
- Day 18: peak secretion of osteocalcin (OC), which reached a total value 3.5 times higher than that of static culture;
- Day 21: calcium deposition was 6.6 times higher than that of constructs in static culture.
During the experiment, the spinner flask bioreactor also recorded better data than the rotating-wall vessel bioreactors.
On day 21, calcium deposition in the spinner flask culture was more than 30 times higher than that in the rotating-wall vessel culture. Moreover, in the rotating-wall vessel culture, no considerable AP activity and OC secretion were detected throughout the culture period.
Critical issues of Spinner Flask Bioreactors for bone tissue engineering
Does this mean that spinner flask bioreactors are the ultimate answer to bone tissue engineering?
Not exactly. In fact, this culture system has some critical issues.
First, the convective forces generated by the rotation of the magnetic stirrer at the bottom of the chamber induce uneven transport of nutrients and gases to the cellularized scaffolds.
In fact, the generation of convective forces does not extend into the interior of the scaffolds. All this limits the transport of nutrients, causing uneven distribution of cells and mineral deposits in the scaffolds.
A direct consequence is that mineralization is limited only to cells exposed outside the scaffold.
The aforementioned study by Sikavitsas, Bancroft et Mikos also highlights that “the histology sections revealed a dense cell layer on the surface of the scaffolds and a considerably lower cell distribution in the scaffold’s interior.”
In addition, the existence of convective forces in the bioreactors exposes the resident cells on the outer surface of the scaffolds to stressful shear forces.
The bioreactor that solves these critical issues
Despite the promising results obtained from culturing in spinner flask bioreactors, improved culture conditions are needed to enable cell growth through cellular polymer constructs.
The main problem with spinner flasks lies in the difficulty of nutrient and gas transport within the scaffolds, which is crucial to achieve homogeneous distribution of cells “in and out” of the scaffold.
The innovative BioAxFlow bioreactor, developed by the Cellex team specifically for tissue engineering, transports nutrients and oxygen more efficiently, ensuring improved cell culture.
Gentle fluid dynamics ensures the transport of nutrients and gases, allowing permeation into the cellular media. Indeed, in BioAxFlow, cell culture medium is continuously pumped from below gently with the help of an external peristaltic pump.
The flow generated ensures continuous movement of the medium, promoting constant exposure of cells to nutrients and growth factors. This ensures optimal growth conditions both outside and inside the scaffolds, resulting in more homogeneous cell adhesion and proliferation.
This revolutionary fluid dynamics also reduces stress generated by shear forces, adversely affecting cells adhered to the scaffolds.
In support of this, the Cellex team tested BioAxFlow with SAOS-2 (osteosarcoma) cells, and the data revealed excellent results regarding cell adhesion and growth and homogeneous distribution of cells on the scaffolds.
Sources
“Formation of three-dimensional cell/polymer constructs for bone tissue engineering in a spinner flask and a rotating wall vessel bioreactor”, Vassilios I. Sikavitsas, Gregory N. Bancroft, Antonios G. Mikos, 2001
“A fluid dynamics-model system for advancing Tissue Engineering and Cancer Research studies: Dynamic Culture with the innovative BioAxFlow Bioreactor”, Giulia Gramigna, Federica Liguori, Ludovica Filippini, Maurizio Mastantuono, Michele Pistillo, Margherita Scamarcio, Antonella Lisi, Giuseppe Falvo D’Urso Labate, Mario Ledda, 2025