Nutrition Research Methodology 311has been estimated through RNA, DNA, or protein/
DNA values, respectively.
Furthermore, molecular biological approaches
have allowed numerous in vitro discoveries that have
aided our understanding of the genetic basis of
nutrient functions and metabolic states in vivo. The
polymerase chain reaction (PCR) can be used for
DNA and/or messenger RNA (mRNA) amplifi cation
to determine the genetic background and/or gene
expression in very small cellular samples. Transfec-
tion studies allow the insertion of DNA into cells to
examine nutrient function. Thus, cell lines that usually
lack the expression of a particular gene can be trans-
fected with DNA containing the gene promoter, as
well as all or part of the transfected gene of interest,
to study the interactions of various nutrients with the
expression of a particular gene. Conversely, knockout
cell lines allow us to investigate the consequences of
losing a specifi c gene. In either case, nutrient function
at the cell level and the cell–gene level may be studied
and provide defi nitive results. Gene regulation by
nutrients has been assessed in different isolated cells
and tissues using appropriate indicators and markers
of gene expression RNA levels.
The integration of biochemical and molecular
technologies into nutrition research allows the poten-
tial for an integrated systems biology perspective
examining the interactions among DNA, RNA protein,
and metabolites. Following the completion of the
human genome sequence, new fi ndings about
individual genes functions and their involvement in
body homeostasis is emerging. Thus, technologies to
achieve a simultaneous assessment of thousands of
gene polymorphisms, the quantitation of mRNA
levels of a large number of genes (transcriptomics) as
well as proteins (proteomics), or metabolites (metab-
olomics) is rapidly progressing. Advances in DNA and
RNA microarray-based tools as well in the application
of classic two-dimensional gel electrophoresis, various
Liquid chromatography-mass spectrometry (LC-MS)
techniques, image scanning, or antibody arrays is
contributing to unraveling the intimate mechanisms
involved in nutritional processes. Epigenetics studies
constitute a rising methodology to be applied in
nutritional research.
13.4 Animal models in nutrition research
Whole animal systems have been used in measuring
the utilization, function, and fate of nutrients. Thus,
a part of our knowledge regarding nutrition concepts
stems from animal experiments, which are often
extrapolated to humans and referred to as animal
models. There are many reasons for choosing an
animal study over a human study. We can and do
subject animals to experimental conditions that we
would ethically not be allowed to apply to humans.
For example, to study the manner in which a nutrient
infl uences the scale and histopathology of atheroscle-
rosis, animal studies are needed. Just as studies with
humans are governed by the rules of ethics commit-
tees, so too are studies with animals. These rules
involve the regulation of facilities, accommodation
and animal care, competence, alternatives to animal
experimentation, anesthesia and euthanasia proce-
dures, registration, supply of animals, and the involve-
ment of an ethical committee.
In general, the use of animals as models for human
nutrition research can be examined from three
aspects:
● the animal model
● the experimental diet and its delivery
● the experimental techniques available.The animal model
Many species have been used in the study of nutrition.
Many are pure-bred strains such as the Wistar rat,
the Charles River mouse, or the New Zealand white
rabbit. Some animal models have been specially
selected to exhibit particular traits, making them very
useful models for research. The Wattanable rabbit has
defective low-density lipoprotein (LDL) receptor
function, making this animal model very useful for
studying the role of diet in infl uencing LDL receptor-
mediated arterial disease. The ob/ob mouse develops
gross obesity because of an alteration in a genetic
profi le (leptin synthesis). In recent times there has
been a rise in the use of transgenic animal models that
have been produced through advanced molecular
genetic techniques. In such models, specifi c genes can
be inserted or deleted to fulfi ll specifi c functions. For
example, the peroxisome proliferator-activated recep-
tor-alpha (PPAR-α) is not expressed in one knockout
mouse model, giving rise to fat accumulation. Another
example of a transgenic mouse presents an overex-
pression of the Cu/Zn-superoxide dismutase enzyme.
The experimental diet and its delivery
The nature of the diet and its mode of delivery are
centrally important in understanding the role of