Cellular behavior after mechanical stimulation on biofunctionalized polylactic acid nanofibers for tendon tissue engineering
Tutor / director / evaluatorMateos Timoneda, Miguel Ángel
Document typeMaster thesis (pre-Bologna period)
Rights accessOpen Access
Recently, tissue engineered solutions have been investigated to cure rotator cuff tears, whose curing is often long and painful. Scaffolds mimicking the natural structure and composition of natural tendons have been found to be a promising alternative. Tendons are typically composed of aligned collagen nanofibers on which tendon cells called tenocytes lay and produce their extracellular matrix. As a consequence, electrospinning was chosen as a technique to produce aligned nano-scaled fibers of poly(lactic acid), a biocompatible polymer. In the next steps, the surface of the fibers was functionalized with collagen and the resulting scaffolds were seeded with tenocytes isolated from tendon biopsies. The behavior of the cells was evaluated with and without mechanical stimulation of the scaffolds. First, the fibers were mechanically characterized and their diameter was indeed calculated to be in the nano-range, proving the efficiency of the electrospinning. The functionalization process was also characterized. Collagen immunofluorescence showed that covalent bonding of collagen on the fibers with carbodiimide chemistry (EDC/NHS) was more effective than the physical adsorption of collagen. Later, tenocytes isolated from a tendon biopsy were seeded on functionalized and non-functionalized scaffolds as well as on glass and a cells immunofluorescence was performed to verify the attachment of the cells on the different substrates. The area of the cells was higher on glass than on other substrates, however the cells were much more elongated on fibrous scaffolds and were all aligned in a single direction. It is very likely that the cells aligned along the fibers direction which would demonstrate that the scaffolds had a positive effect on the cells. Indeed, this cellular behavior is very close to what can be observed on natural tendons. The influence of collagen functionalized on the scaffolds was yet not clear. Although cells had a higher area and elongation on the first day of culture on functionalized scaffolds, the values of non-functionalized scaffolds became extremely similar after the third day of culture. A stimulation test on a scaffold seeded with tenocytes had to be ended prematurely due to experimental issues. The sample eventually was 75% stiffer but its yield strain was four times lower. Physical ageing was considered as the most consistent explanation over collagen production on a short stimulation period.