LeeAnn O. Bailey, Newell R.
Washburn, Carl G. Simon Jr., Edward S. Chan,
Francis W. Wang
Biomaterials Group, Polymers Division, National Institute of Standards and Technology, Gaithersburg, MD. 20899-8545
The
introduction of tissue engineering approaches for the repair and replacement
of human body components extends the application and importance of synthetic
materials (biomaterials). Implanted biomaterials frequently evoke inflammatory
responses, which are quite complex and not well defined at present. The
goals of this work are to develop improved measurement methods for the
quantification of cellular inflammatory responses to biomaterials and to
obtain data that culminates in an enhanced understanding of the ways in
which the body responds to the introduction of biomaterials. To evaluate
the biocompatibility of materials, we have established a system that allows
for the analysis and quantitation of cellular inflammatory responses in
vitro. Elevated cytokine production serves as an indicator of inflammatory
responses, thus we monitored the levels of tumor necrosis factor alpha
and interleukin-1b
produced by cells. In this study, the inflammatory responses of non-transformed
murine macrophages (Raw 264.7) were analyzed. The cells were incubated
with polymethylmethacrylate (PMMA) microspheres in the presence and absence
of lipopolysaccharide (LPS) at 8 h and 18 h. The analysis of the genetic
material obtained from the cells was quantitated using Real-Time Reverse
Transcriptase Polymerase Chain Reaction (RT-PCR). The cell populations
treated with LPS alone or PMMA microspheres alone resulted in an elevation
of cytokine levels, while cells incubated with LPS and PMMA particles demonstrated
a synerginistic effect by producing a marked increase in the level of cytokine
expression, 336 fold greater than that of the untreated control. Florescence
microscopy studies that assessed cellular viability were also performed
and are consistent with the RT-PCR results. Thus, we have developed an
in vitro assay for quantifying cellular inflammatory responses.
Furthermore, we have demonstrated that the presence of a bacterial component
and wear debris particles can work in concert to elevate cytokine production.