FIG. 1.
Donor-dependent response of hMSCs in 1% MeHA hydrogels and pellets during chondrogenic differentiation. (A) hMSC donors laden in 1% MeHA hydrogels (20 million cells/mL) were cultured in CM+. Of the seven donors, three donors (18M, 22F, and 62M) maintained their cylindrical morphology with little contraction (left). Two other donors (42F and 57M) aggregated and separated from the hydrogels (middle; inset = a cell aggregate in 1% MeHA hydrogels after 1 week in CM+; scale bar = 1 mm). The remaining two donors (46F and 61F) failed to maintain their structural integrity and degraded (arrows) the MeHA hydrogels within 1 week (right; inset = disintegrated cells and/or small aggregates; scale bar = 100 μm). (B) Two young/healthy donors and one aged/arthritic donor (18M, 22F, and 62M) that demonstrated the least contraction had the greatest GAG and collagen content at 8 weeks (left panels) compared to two aged/arthritic donors (42F and 57M) that markedly self-aggregated. The remaining two aged/arthritic donors (46F and 61F) that failed to maintain a cylindrical geometry disintegrated within a few days and were unable to be assessed for biochemical properties. In pellet cultures, GAG production from those same donors was low overall compared to those in hydrogels, and GAG content in pellets was similar at 2 and 4 weeks (right panel). (C) Collagen production was slightly greater than GAG production, but the levels of collagen production were also similar between the donors, except for one young/healthy (18M) donor at 4 weeks (right panel). GAG and collagen contents were normalized to their respective starting cell number (construct = 106; pellet = 0.25 × 106) due to the variations in response (n = 3–4/group). (D and E) Due to the self-aggregation or disintegration, constructs from only three donors (yellow bars; 18M, 22F, and 62M) were assessed for mechanical properties at 8 weeks. (D) Equilibrium modulus (EY; kPa) of constructs from the 22F donor was 44% (30 kPa; #p < 0.05) lower than the 18M (52 kPa) and 62M (53 kPa) donors. (E) Constructs seeded with cells from donor 22F were less contractile (−22%; #p < 0.05) based on changes in volume (% change) compared to those from donors 18M (−52%) and 62M (−55%; light bars = 2w, medium bars = 4w, dark bars = 8w; n = 3–4/group; #p < 0.05). CM, chemically defined media; GAG, glycoaminoglaycan; hMSC, human mesenchymal stem cell; MeHA, methacrylated hyaluronic acid. Color images available online at www.liebertpub.com/tea