Correlations between second harmonic signal, microstructure, and mechanics of contracting collagen gels

摘要

Second harmonic generation (SHG) from collagen provides an optical signal that can yield detailed information about collagen microstructure when imaged with laser scanning microscopy, from both collagen-based engineered tissue and connective tissues from animals. Therefore SHG images may provide information that correlates with bulk tissue mechanical properties, or at least a component of those properties resulting from collagen. In order to probe these correlations, we used multiphoton microscopy to gather SHG signal intensity and depth decay information from fibroblast-seeded contracting collagen hydrogels. These gels were polymerized at pH 6 to engineer a tissue with large diameter collagen fibers and large pores between fibers, and pH 9 to produce smaller diameter collagen fibers with smaller pores. Both gels initially contained 4 mg/ml collagen; after 16 days of floating culture, the pH 6-polymerized gels had contracted to 4.4 ± 0.6% of their original volume, and the pH 9-polymerized gels to 10.7 ± 2.7%. During this time period, the bulk compressive moduli (CM) of the gels increased ～9.2-fold and ～ 1.4-fold for the pH 6 and pH 9 polymerization conditions, respectively. Correspondingly, the SHG signal at the tissue surface increased ～25-fold and ～19-fold for the pH 6 and pH 9 gels, respectively; whereas the effective SHG attenuation coefficient increased ～4.5 and ～5.8-fold, respectively. Meaningful linear correlations only existed between the CM and surface SHG signal and the CM and SHG attenuation coefficient for pH 6-polymerized gels, indicating a possible influence of fibroblast activity on the CM of the pH-9 polymerized gels.