ABSTRACT
The goal of this chapter is to provide the biomechanical engineer with a description of the fundamental tools and procedures that are currently used to access the quality and quantity of genes made by specific tissues and cells. Genes can be controlled at many levels, as shown schematically in Fig. 3.1. The control of gene expression begins in the nucleus where a combination of DNA sequences (
cis
acting) and proteins (transcription factors, or
trans
acting) interact in the nucleus to make (transcribe) RNA, which then transits to the cytoplasm. By using the ribosome machinery (made of 18S and 28S RNA embedded in proteins) (see Fig. 3.2), the mRNA is translated into proteins that are either used within the cell or secreted and function outside the cell. This chapter will focus exclusively on the analysis of mRNA expression. The methods described are designed to compare the quantity (amount) and quality (specific type of gene) of mRNA activation in control and experimental situations. Thus, the techniques described are used to evaluate the response of a cell or tissue to environmental influences such as biomechanical stress specifically at the ‘‘mRNA level.’’ By examining the nature of the RNA that is altered one can extrapolate the corresponding changes in protein expression and develop a framework for understanding the molecular
events that accompany changes in skeletal structure and function. Altered protein production results in altered bone matrix composition that can, in turn, affect bone elasticity.
