In this article, we explore the potential for SIMS imaging of unknown biological samples by employing traditional TOF-SIMS and accurate mass determination via FTICR-SIMS for direct molecular ion identification of biological components in D. discoideum during aggregation. However, the covalently crosslinked MeHA hydrogels undergo minimal degradation by hydrolysis after extended incubation in aqueous environment. As a result, hydrogels fabricated with high MeHA concentrations hinder the spatial distribution of cartilaginous matrix synthesized by the encapsulated hMSCs. An ideal scaffold material for tissue engineering should be able to degrade in concert with the matrix elaboration by the seeded cells. Numerous studies have attempted to develop scaffolds that can degrade either by hydrolytic or enzymatic actions. Cells are known to remodel their surrounding extracellular matrixby secreting catabolic enzymes, such as matrix metalloproteinases. Hydrogels that are designed to be sensitive to MMP actions have been demonstrated by several previous studies. These studies also revealed that cell-mediated scaffold degradation substantially impacts the differentiation of mesenchymal stem cells. For example, a recent study demonstrated that the MMP-sensitive PEGhydrogels promoted the chondrogenesis of encapsulated human MSCs. However, few studies have examined the effect of cell-mediated matrix degradation on hMSC chondrogenesis and subsequent hypertrophy in hyaluronic acid hydrogels. Furthermore, many of the previous investigations used Neosperidin-dihydrochalcone MMP-cleavable peptides containing dual cysteine residues to crosslink acrylated macromers. The resulting hydrolytically degradable thiol-ether-ester bonds confound the effect of enzymatic degradation of the hydrogels by cell-secreted MMPs. Chondrogenically induced MSCs display the propensity to continue differentiating towards a hypertrophic phenotype, resulting in extensive calcification of the ECM after ectopic transplantation. This process is similar to that observed in the terminal differentiation of hypertrophic chondrocytes in the growth plate and articular cartilage, where the components and structures of the ECM regulate chondrocyte hypertrophy and matrix calcification. Reciprocally, hypertrophic chondrocytes produce increased MMPs such as MMP13 to remodel their surrounding cartilage matrix in a way that facilitates the ensuing mineralization. Therefore, the dynamic Salvianolic-acid-B interplay between the degradational properties of the scaffold materials and MMPs synthesis by chondrogenically induced hMSCs may influence hypertrophic differentiation of these MSCs, and consequently the calcification of the neocartilage matrix. Previously we showed that the crosslinking densities of chondrocytes or hMSC-laden HA hydrogels influence neocartilage formation, subsequent hypertrophy and neocartilage calcification. In addition, a recent study demonstrated that MMP-sensitive but hydrolysis-insensitive HA hydrogels can be fabricated by crosslinking maleimide modified HA macromers with MMP-cleavable peptides. However, further understanding on the regulation of hMSCs hypertrophy and tissue calcification is needed to ensure successful clinical application of hMSCs for cartilage repair. We hypothesize that MMP-sensitive HA hydrogels will regulate the chondrogenic and hypertrophic differentiation of the encapsulated hMSCs. Thus, the objective of this study is to evaluate the effect of cell-mediated degradation of HA hydrogels.