For tendon tissue engineering, tenocyte-seeded scaffolds are a promising approach. Under conventional 2D culture, however, tenocytes show rapid senescence and phenotype loss. Researches hypothesized that phenotype loss could be counteracted by simulated microgravity conditions.

In complex hand injuries, tendon grafting is often required. However, the number of suitable autologous donor tendons is limited, so that there is a demand for tissue engineered tendon grafts. These constructs consist of an acellular scaffold providing mechanical stability, and a seeded cell line providing regeneration and tissue function.

Several scaffold materials have been used with some success so far, such as chitosan, silk fibroin, PLGA or decellularized cadaver tendons. As both cultures in artificial matrices and high-density cultures are difficult to handle technically, they aimed to establish a 3D culture model by use of simulated microgravity. Simulated microgravity has been shown to induce three-dimensional cell formation (spheroids) in various cell types, such as thyroid cancer cells, corneal stromal cells.

They, therefore, hypothesized that spheroid formation would also take place in human tenocytes exposed to microgravity and that spheroid formation will counteract in vitro senescence in analogy to the 3D culture models mentioned above.

Tenocytes

As tenocytes are hard to obtain in a number sufficient for tendon tissue engineering, expansion in vitro is inevitable to the date. Under conventional culture conditions in a dish or flask, cells are plated in the 2-dimensional layer, which builds a fairly artificial environment, which has been shown to be detrimental to phenotype preservation in various cell lines.

Loss of differentiation potential into osteogenic and adipogenic lineage has been described for adipose-derived stem cells during monolayer culture. On the other hand, by 3D-cultivation in microfibers, human pluripotent stem cells have preserved their undifferentiated phenotype to a significantly higher passage than in culture on a matrigel surface. With simulated microgravity, they propose a 3D culture model that has several advantages: it is a cell-only environment that avoids problems known from scaffold systems, such as difficulties in harvesting cells. 

Furthermore, it is not limited in size and dimension, such as the hanging drop system, and it is not limited in its duration of application. No impaction of cell viability was observed in our experiments during the full observation period.

They have proposed a 3D culture model that is easy to use and may help to preserve tenocyte phenotype better than conventional 2D culture. This might facilitate tenocyte cultivation for tissue engineering purposes in the future.

With the further long-term expansion of tenocyte spheroids, this could also be a step towards scaffold-free tissue engineering of a tendon construct. The future experiment will evaluate these issues in detail, particularly concerning size, shape and mechanical stability.