The use of decellularized adipose tissue to provide an inductive microenvironment for the adipogenic differentiation of human adipose-derived stem cells.

The development of an engineered adipose tissue substitute, capable of supporting reliable, predictable, and complete fat tissue formation, would be of significant value in the fields of plastic and reconstructive surgery. Towards the goal of engineering an optimized microenvironment for adipogenesis, a decellularization strategy was developed for adipose tissue, which yielded 3-D scaffolds with preserved extracellular matrix architecture. A significant volume of scaffolding material could be obtained from a human tissue source that is commonly discarded. Histology, immunohistochemistry, and scanning electron microscopy confirmed the efficacy and reproducibility of the approach, and also indicated that the basement membrane was conserved in the processed matrix, including laminin and collagen type IV. Seeding experiments with human adipose-derived stem cells indicated that the decellularized adipose tissue (DAT) provided an inductive microenvironment for adipogenesis, supporting the expression of the master regulators PPARgamma and CEBPalpha, without the need for exogenous differentiation factors. High levels of adipogenic gene expression and glycerol-3-phosphate dehydrogenase activity were observed in the induced DAT scaffolds, as compared to cells grown in monolayer or cell aggregate culture. The protein data emphasized the importance of the cell donor source in the development of tissue-engineering strategies for large-volume soft tissue regeneration.

Harnessing the purinergic receptor pathway to develop functional engineered cartilage constructs.

OBJECTIVE: Mechanical stimulation is a widely used method to enhance the formation and properties of tissue-engineered cartilage. While this approach can be highly successful, it may be more efficient and effective to harness the known underlying mechanotransduction pathways responsible. With this aim, the purpose of this study was to assess the effect of directly stimulating the purinergic receptor pathway through exogenous adenosine 5'-triphosphate (ATP) in absence of externally applied forces. METHODS: Isolated bovine articular chondrocytes were seeded in high density, 3D culture and supplemented with varying doses of ATP for up to 4 weeks. The effects on biosynthesis, extracellular matrix accumulation and mechanical properties were then evaluated. Experiments were also conducted to assess whether exogenous ATP elicited any undesirable effects, such as: inflammatory mediator release, matrix turn-over and mineralization. RESULTS: Supplementation with ATP had a profound effect on the growth and maturation of the developed tissue. Exogenous ATP (62.5-250 microM) increased biosynthesis by 80-120%, and when stimulated for a period of 4 weeks resulted in increased matrix accumulation (80% increase in collagen and 60% increase in proteoglycans) and improved mechanical properties (6.5-fold increase in indentation modulus). While exogenous ATP did not stimulate the release of inflammatory mediators or induce mineralization, high doses of ATP (250 microM) elicited a 2-fold increase in matrix metalloproteinase-13 expression suggesting the emergence of a catabolic response. CONCLUSIONS: Harnessing the ATP-purinergic receptor pathway is a highly effective approach to improve tissue formation and impart functional mechanical properties. However, the dose of ATP needs to be controlled as not to elicit a catabolic response.

Estimation of ozone with total ozone portable spectroradiometer instruments. I. Theoretical model and error analysis.

Inexpensive devices to measure solar UV irradiance are available to monitor atmospheric ozone, for example, total ozone portable spectroradiometers (TOPS instruments). A procedure to convert these measurements into ozone estimates is examined. For well-characterized filters with 7-nm FWHM bandpasses, the method provides ozone values (from 304- and 310-nm channels) with less than 0.4% error attributable to inversion of the theoretical model. Analysis of sensitivity to model assumptions and parameters yields estimates of ±3% bias in total ozone results with dependence on total ozone and path length. Unmodeled effects of atmospheric constituents and instrument components can result in additional ±2% errors.

Estimation of ozone with total ozone portable spectroradiometer instruments. II. Practical operation and comparisons.

We used a microcomputer-controlled total ozone portable spectroradiometer instrument #21 (MTOPS21) to measure solar radiation at 298, 304 and 310 nm in Greenbelt, Md., during 1995. One day's ozone measurements from a Brewer instrument (B105) were used to calibrate the 304- and 310-nm channel ratios to a theoretical model. Total ozone estimates were then determined for the entire MTOPS21 data set. Differences between individual B105 and MTOPS21 ozone estimates show a 1% drop as solar zenith angles increase and depend on atmospheric attenuation and SO(2) variation at the ±2% level. Daily average values agree well (<0.5% average offset, 2% standard deviation).