| Objective: To study the physicochemical properties of Alg-Sr hydrogel and toobserve the survival, proliferation and osteogenic differentiation of rabbit BMSCs init, then to provide theoretical foundation for the application of BMSCs/Alg-Srhydrogel in bone tissue engineering.Methods:(1)2.0wt%alginate sodium was prepared and then added to0.2mol/LSrCl2solution to produce the Alg-Sr hydrogel. Gross morphology, microstructure,modulus of compression, degradability and swelling capacity of Alg-Sr wereinvestigated accordingly, with the Alg-Ca obtained in the same method as a control.(2) BMSCs were gained by whole bone marrow culture, passaged by trypsindigestion and identified with immunocytochemistry.(3) BMSCs were then seededinto the Alg-Sr gels and the cell viability, proliferation were assessed by Live/Deadassay and DNA quantification, and simultaneously BMSCs were seeded in Alg-Caand same items were detected as a control.(4) Osteogenic differentiation of rabbitBMSCs embeded in Alg-Sr and Alg-Ca gels was evaluated by ALP activity,osteoblast specific gene expression level and calcium deposition.Results:(1) Alg-Sr and Alg-Ca gels were successfully fabricated intotransparent and spherical-shaped gels with the diameter at about3mm. Microstructureof Alg-Sr observed under SEM was three-dimensional and porous, as well as theAlg-Ca; the compression modulus of Alg-Sr was(186.53±8.37) kPa while Alg-Ca was(152.14±7.45) kPa, and the difference between them was significant (t=6.863，P<0.05); the degradability of Alg-Sr and Alg-Ca was about the same before day20(P>0.05), however, at day20,25and30the degradability of Alg-Sr was obviouslylower than that of Alg-Ca (P<0.05);no significant difference (t=0.927, P>0.05) wasobserved in the swelling rate of Alg-Sr (14.32%±1.53%) and Alg-Ca(15.25%±1.64%).(2) Cell morphology and identification withimmunocytochemistry(CD29(+)、 CD34()、 CD44(+)、 CD45()) showed thesuccessful obtain of rabbit BMSCs through the whole bone marrow culture.(3) Therewas the same morphologic change and growth trend of BMSCs3-D cultured in Alg-Sr and Alg-Ca gels; the morphology of BMSCs in alginate gels was spherical inthe first4days and spindle or stellate several days later; the DNA content of BMSCsin Alg-Sr (4.38±0.24) g was significantly higher than that in Alg-Ca (3.25±0.21) g,(t=7.923，P<0.05) after being3-D cultured in normal growth medium for21days.(4)12days after osteogenic induction, the ALP activity in Alg-Sr was(2.14±0.11)unit/100mL, obviously higher than (1.69±0.14)unit/100mL in Alg-Ca,(t=5.652, P<0.05); the mRNA expression level of OSX, Col1, Runx2and calciumdeposition of BMSCs embedded in Alg-Sr were notably higher than that in Alg-Ca(P<0.05).Conclusion: Alg-Sr gel shares the identical porous structure with Alg-Ca, whichbenefits the3-D culture of BMSCs. Nevertheless, Alg-Sr is characterized with thehigher mechanical strength and lower degradation rate compared to Alg-Ca, apartfrom that, the better promotion of Alg-Sr gel to proliferation and osteogenicdifferentiation of BMSCs indicates that Alg-Sr gel is a novel and excellent cellscaffold material for the regeneration and restoration of bone tissue. |
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