Figure 7B. MTT assay of C2C12 myoblast cells incubated onto well plate (control) and cellulose acetate (CA) and cellulose acetate nanofibers with annatto extract (CA@A) over 2 days and 7 days. ***p<0.05 (Student’s t-test)
Cells grown in a monolayer (control) were maintained under the same conditions and used for comparison. The MTT results (Figure 7B) obtained after 2 days of cell culture found lower cell viability in the scaffolds, indicating that the monolayer control was more suitable than nanofiber scaffolds for cell attachment and proliferation during the initial days of culture.
After 7 days of culture, however, the cells in the monolayer reached growth arrest, while the viability rate for the cells cultivated onto the scaffolds increased. Growth arrest in the control may be regulated by cell-cell contact or contact inhibition [43]. This was not observed in cells cultivated onto nanofibers due to their large surface area, indicating that scaffolds are beneficial for longer-duration cell culture.
The viability of cells cultivated onto the CA@A nanofiber was also found to be higher than for those cultivated onto the CA nanofiber. Previous authors have shown annatto extract to be rich in antioxidant components that can improve cellular proliferation [44]. Furthermore, considering the more hydrophilic nature of cellulose acetate nanofiber with annatto (as demonstrated by the contact angle), this surface can also facilitate interactions between cells and matrices, leading to improved attachment and proliferation of C2C12 myoblasts [37]. These results suggest that CA@A nanofibers induce myoblast proliferation, which is essential for scaffolds utilized in cultivated meat expansion.