The objective of this study was to investigate how molecular level changes in the collagen network affect its mechanical integrity. Our hypothesis is that the cleavage and unwinding of triple helices of collagen molecules significantly reduce the mechanical integrity of the collagen network in bone, whereas collagen crosslinks play a major role in sustaining the structural integrity of the collagen network. To test this hypothesis, the collagen molecular structure was altered in demineralized human cadaveric bone samples in the following two ways: heat induced unwinding and pancreas elastase induced cleavage of collagen molecules. Along with control specimens, the treated specimens were mechanically tested in tension to determine their strength, elastic modulus, toughness, and strain to failure. Also, the percentage of denatured collagen molecules and amounts of two major collagen crosslinks (hydroxylysylpyridinoline and lysylpyridinoline) were determined using high-performance liquid chromatography techniques. It was found that unwinding of collagen molecules may cause more reduction in stiffness (E) but less strain to failure (ef) than cleavage. Both collagen denaturation types cause similar changes in the strength (ss) and work to fracture (Wf) of the collagen network with no significant changes in hydroxylysylpyridinoline and lysylpyridinoline crosslinks. The results of this study indicate that the integrity of collagen molecules significantly affect the mechanical properties of the collagen network in bone, and that collagen crosslinks may play an important role in maintaining the mechanical integrity of the collagen network after collagen denaturation occurs.

Oral infections caused by Candida species, the most commonly isolated human fungal pathogen, are frequently associated with biofilms. Although Candida albicans is the predominant organism found in patients with oral thrush, a biofilm infection, there is an increasing incidence of oral colonization and infections caused by non- albicans Candida species, including C. glabrata, C. dubliniensis, and C. tropicalis, which are frequently more resistant to antifungal treatment. While single-species Candida biofilms have been well studied, considerably less is known about the dynamics of mixed- Candida species biofilms and how these dynamics are altered by antifungal treatment. To address these questions, we developed a quantitative polymerase chain reaction-based approach to determine the precise species composition of mixed- Candida species biofilms formed by clinical isolates and laboratory strains in the presence and absence of clinically relevant concentrations of 3 commonly used antifungals: fluconazole, caspofungin, and amphotericin B. In monospecies biofilms, fluconazole exposure favored growth of C. glabrata and C. tropicalis, while caspofungin generally favored significant growth of all species to a varying degree. Fluconazole was not effective against preformed mixed- Candida species biofilms while amphotericin B was potent. As a general trend, in mixed- Candida species biofilms, C. albicans lost dominance in the presence of antifungals. Interestingly, presence in mixed versus monospecies biofilms reduced susceptibility to amphotericin B for C. tropicalis and C. glabrata. Overall, our data suggest that antifungal treatment favors the growth of specific non- albicans Candida species in mixed- Candida species biofilms.

The hypothesis of this study is that the in situ thermostability of collagen in bone changes with age and such changes relate to the structural properties of collagen and/or the interaction between the collagen and mineral phases. To test the hypothesis, the effect of age on the in situ thermostability of collagen in human bone and its correlation with the mineral and collagen phases were investigated. In this study, 30 human cadaveric femurs were collected and divided into three age groups: young adults (20-45 years), middle aged (46-69 years), and elderly (over 70 years of age). The in situ thermostability of collagen was assessed using high performance liquid chromatography (HPLC) in terms of the amount of heat-induced collagen denaturation at eight temperatures between 25 degrees C and 140 degrees C. In addition to the density and weight fraction of the mineral and organic phases, the concentration of collagen crosslinks was measured to assess the structural integrity of collagen. The results of this study indicate that the in situ thermostability of collagen increases with increasing age, and such age-related changes correlate with the following: collagen molecular structure, amount of noncalcified collagen, and the fraction and density of the mineral phase in bone. These results suggest that the age-related changes in the in situ thermostability of collagen most likely relate to the collagen structure and its interaction with the mineral phase. In addition, the bone remodeling process may play a role in the age-related changes in collagen thermostability because noncalcified collagen is commonly associated with this process.