Figure 5. Viability and metabolic activity of vitrified vs fresh and slow-frozen menisci.

(A) Transverse sections at half thickness show fresh (a, e, i, m), slow-frozen (b, f, j, n), 2-hour vitrified (c, g, k, o), and 3-hour vitrified (d, h, l, p) menisci. Images (a-d) are 2× objective magnification and stitched together. Images of regions (e-p) within dashed boxes (red = anterior (A), blue = central (C), and yellow = posterior (P)) presented at 10×. Scale bars: 2 mm (a-d), 200 μm (e-p). (B) Cell viability from fresh (N=4), slow-frozen (N=3), 2-hour vitrified (N=3), and 3-hour vitrified (N=3) menisci. p-value determined with two-way ANOVA with Bonferroni post-hoc test. (C) Cell viability quantified in outer (grey), middle (red), and inner (blue) layers within sagittal sections. No significant differences within layers of each group using two-way ANOVA with Bonferroni post-hoc test. (D) Metabolic activity between vitrified menisci for fresh (N=3; n=11), 2-hour (N=3; n≥10) and 3-hour (N=4; n≥11) durations, along with SF menisci (N=3; n≥11), over 0 to 4-day post warming. Metabolic activity normalized to corresponding average fresh meniscus values on day 0 within the same batch. N = number of independent menisci; n = number of sagittal sections. p-value determined via two-way ANOVA. Data are mean ± SD.