Skeletal muscle progenitors are sensitive to collagen architectural features of fibril size and cross linking.

Muscle stem cells (MuSCs) are essential for the robust regenerative capacity of skeletal muscle. However, in fibrotic environments marked by abundant collagen and altered collagen organization, the regenerative capability of MuSCs is diminished. MuSCs are sensitive to their extracellular matrix environment but their response to collagen architecture is largely unknown. The present study aimed to systematically test the effect of underlying collagen structures on MuSC functions. Collagen hydrogels were engineered with varied architectures: collagen concentration, cross linking, fibril size, and fibril alignment, and the changes were validated with second harmonic generation imaging and rheology. Proliferation and differentiation responses of primary mouse MuSCs and immortal myoblasts (C2C12s) were assessed using EdU assays and immunolabeling skeletal muscle myosin expression, respectively. Changing collagen concentration and the corresponding hydrogel stiffness did not have a significant influence on MuSC proliferation or differentiation. However, MuSC differentiation on atelocollagen gels, which do not form mature pyridinoline cross links, was increased compared with the cross-linked control. In addition, MuSCs and C2C12 myoblasts showed greater differentiation on gels with smaller collagen fibrils. Proliferation rates of C2C12 myoblasts were also higher on gels with smaller collagen fibrils, whereas MuSCs did not show a significant difference. Surprisingly, collagen alignment did not have significant effects on muscle progenitor function. This study demonstrates that MuSCs are capable of sensing their underlying extracellular matrix (ECM) structures and enhancing differentiation on substrates with less collagen cross linking or smaller collagen fibrils. Thus, in fibrotic muscle, targeting cross linking and fibril size rather than collagen expression may more effectively support MuSC-based regeneration.

Measures of Bone Mineral Carbonate Content and Mineral Maturity/Crystallinity for FT-IR and Raman Spectroscopic Imaging Differentially Relate to Physical-Chemical Properties of Carbonate-Substituted Hydroxyapatite.

Bone mineral carbonate content assessed by vibrational spectroscopy relates to fracture incidence, and mineral maturity/ crystallinity (MMC) relates to tissue age. As FT-IR and Raman spectroscopy become more widely used to characterize the chemical composition of bone in pre-clinical and translational studies, their bone mineral outcomes require improved validation to inform interpretation of spectroscopic data. In this study, our objectives were (1) to relate Raman and FT-IR carbonate:phosphate ratios calculated through direct integration of peaks to gold-standard analytical measures of carbonate content and underlying subband ratios; (2) to relate Raman and FT-IR MMC measures to gold-standard analytical measures of crystal size in chemical standards and native bone powders. Raman and FT-IR direct integration carbonate:phosphate ratios increased with carbonate content (Raman: p < 0.01, R2 = 0.87; FT-IR: p < 0.01, R2 = 0.96) and Raman was more sensitive to carbonate content than the FT-IR (Raman slope + 95% vs FT-IR slope, p < 0.01). MMC increased with crystal size for both Raman and FT-IR (Raman: p < 0.01, R2 = 0.76; FT-IR p < 0.01, R2 = 0.73) and FT-IR was more sensitive to crystal size than Raman (c-axis length: slope FT-IR MMC + 111% vs Raman MMC, p < 0.01). Additionally, FT-IR but not Raman spectroscopy detected differences in the relationship between MMC and crystal size of carbonated hydroxyapatite (CHA) vs poorly crystalline hydroxyapatites (HA) (slope CHA + 87% vs HA, p < 0.01). Combined, these results contribute to the ability of future studies to elucidate the relationships between carbonate content and fracture and provide insight to the strengths and limitations of FT-IR and Raman spectroscopy of native bone mineral.

Mucin O-glycans suppress quorum-sensing pathways and genetic transformation in Streptococcus mutans.

Mucus barriers accommodate trillions of microorganisms throughout the human body while preventing pathogenic colonization1. In the oral cavity, saliva containing the mucins MUC5B and MUC7 forms a pellicle that coats the soft tissue and teeth to prevent infection by oral pathogens, such as Streptococcus mutans2. Salivary mucin can interact directly with microorganisms through selective agglutinin activity and bacterial binding2-4, but the extent and basis of the protective functions of saliva are not well understood. Here, using an ex vivo saliva model, we identify that MUC5B is an inhibitor of microbial virulence. Specifically, we find that natively purified MUC5B downregulates the expression of quorum-sensing pathways activated by the competence stimulating peptide and the sigX-inducing peptide5. Furthermore, MUC5B prevents the acquisition of antimicrobial resistance through natural genetic transformation, a process that is activated through quorum sensing. Our data reveal that the effect of MUC5B is mediated by its associated O-linked glycans, which are potent suppressors of quorum sensing and genetic transformation, even when removed from the mucin backbone. Together, these results present mucin O-glycans as a host strategy for domesticating potentially pathogenic microorganisms without killing them.