Collagen cross-linking with Au nanoparticles.

Tiopronin (N-(2-mercaptopropionyl)glycine)-protected gold nanoparticles (TPAu) were cross-linked to collagen via EDC (1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide) coupling. On average, each TPAu forms eight amide bonds with collagen lysine moieties. The resulting gels were studied with environmental SEM, TEM, micro-DSC, and TNBS assay. The porous structure of collagen was significantly altered by cross-linking, resulting in the reduction of the pore size from ca. 140 to <1 microm depending on the concentration of nanoparticles. The collagenase biodegradation assay showed improved stability of cross-linked material. The cell viability assay, CellTiter96, indicates that the gold nanoparticles are not toxic at the concentrations used in gel synthesis. This new material has potential for the delivery of small molecule drugs as well as Au nanoparticles for photothermal therapies, imaging, and cell targeting.

Altered expression of transcription factors and genes regulating lipogenesis in liver and adipose tissue of mice with high fat diet-induced obesity and nonalcoholic fatty liver disease.

OBJECTIVE: To determine whether expression of transcription factors and lipogenic enzymes is altered in liver and adipose tissue of mice with obesity, insulin resistance, and nonalcoholic fatty liver disease. METHODS: Mice were fed chow containing 9% of calories from standard fat (SF) or 20% of calories from high fat (HF) and killed after 9 months in the fasted or fed state. MEASUREMENTS: Liver injury was evaluated by histology and serum aminotransferase levels. Transcription factor expression was measured by real-time PCR. Lipogenic enzymes were measured by real-time PCR and Western blots. RESULTS: HF mice weighed more, had insulin resistance, hepatic steatosis, and focal pericellular hepatic fibrosis. Hepatic expression of sterol regulatory element-binding protein-1c, carbohydrate response element-binding protein, liver X receptor-alpha, acetyl-CoA carboxylase (ACC), and fatty acid synthase (FAS) decreased during fasting in SF and HF mice; however, FAS expression and protein content were higher in the liver of fasted HF mice than of fasted SF mice. In adipose tissue, expression of sterol response element-binding protein-1c, carbohydrate response element-binding protein, liver X receptor-alpha, peroxisome proliferator-activated receptor-gamma, ACC, and FAS decreased with fasting in mice fed SF, but not in HF mice. ACC and FAS expression and protein content remained higher during fasting in HF than in SF mice. CONCLUSION: Feeding a nutritionally complete diet containing a moderate increase in fat produces obesity and steatohepatitis. During fasting, hepatic FAS expression and protein content are increased in HF mice. Transcription factor expression, and lipogenic enzyme expression and protein concentration do not decline during fasting in adipose tissue from HF mice. De-novo lipogenesis may persist in liver and adipose tissue during fasting in obesity/nonalcoholic fatty liver disease.

Electrochemical Time-of-Flight in crosslinked collagen matrix solution: implications of structural changes for drug delivery systems.

Electrochemical Time-of-Flight (ETOF) method was used for the first time to measure the diffusion coefficient of 4-hydroxy-TEMPO, a molecular probe in collagen I matrix solution as a function of its concentration and the extent of crosslinking with glutaraldehyde (GA). The values of the diffusion coefficient were correlated with Circular Dichroism (CD) and viscosity data to assess the changes of the structure of a collagen matrix. The low value of probe diffusion coefficient indicates that the molecular collagen contributes to large diffusion hindrance of the medium. The combined Brinkman or effective medium model and the Carman-Kozeny model were used to estimate the average diameter of a matrix pore as a function of collagen solution composition. We show that 0.5% and 1% (w/w) collagen matrix crosslinked with the addition of GA above 0.1% (v/w) forms matrix with pores larger than in native collagen. This result suggests the existence of a micro-phase separation in the crosslinked collagen matrix. The implications of these findings for the design of small molecule drug delivery systems are discussed.

The microstructure of collagen type I gel cross-linked with gold nanoparticles.

Scanning electron microscopy, transmission electron microscopy, rheometry, and electrochemistry were used to provide insight into the microstructure of collagen type I gel (1%, w/v) modified with the tiopronin-protected (N-(2-mercaptopropionyl)glycine) gold nanoparticles (TPAu), a multivalent crosslinker. The cross-linking reaction, performed via EDC (1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide) coupling, results in compliant, mechanically stable and continuous gels. The gels contain unusual interconnected collagen-TPAu particles. Electrochemical measurements of 4-hydroxy-(2,2,6,6-tetramethylpiperidine-1-oxyl) (4HT) diffusion within the gel reveal that the gel hindrance is nearly independent of the TPAu concentration. The properties of the collagen-TPAu gel make it suitable for potential biomedical applications, such as delivery of small molecule drugs.