Page 106 - Vibrational Spectroscopic Imaging for Biomedical Applications

82 Cha pte r T h ree

same media (LNCap-FGC and PNT2-C2), whereas two were grown
in different media (BPH and PC-3). Importantly, the two cell lines that
were grown in identical media (LNCap-FGC and PNT2-C2) showed
significant separation, realized by anticorrelation on PC-2.
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In a follow-up study, Harvey et al. acquired conventional FTIR
spectra from PC-3 cells and LNCaP cells, each grown separately in
their optimum culture medium (as advised by ECACC protocols)
and a “foreign medium.” Unsupervised PCA of these spectra demon-
strated clustering of the two cell lines that was independent of the
culture medium in which they were grown, but was principally
dependent on the cell type (Fig. 3.10b). Thus, it may be concluded
that for at least prostate cell lines PC-3 and LNCaP, the influence of
the basic media under investigation in this study (RPMI 1640 and
Ham’s F-12) on the cell metabolites, which may be more significant
using other analytical modalities, is not the primary influence on
spectroscopic measurements. However, it must be acknowledged
that cells are a function of their environment (discussed in further
detail below). Thus, the same cell line grown in two different media
with relatively larger compositional differences (or containing potent
stimuli) will effect spectroscopic classification.
If the cell is exposed to an environment that does not sustain its
optimum growth and down-regulates the expression of biomolecular
features (such as cell surface antigens, hormone receptors, protein
expression), which characterize that cell type in vivo, then this may
ultimately render the cell to a new class. In vivo, it is well known that
stromal-cell interactions are particularly important in cancers such as
of the breast, where the stromal compartment plays a critical role in
directing proliferation and functional changes in the epithelium. 52
Moreover, environmental stimuli directing cell phenotype has been
53
recently studied with imaging FTIR by Krafft et al. In this study,
human mesenchymal stem cells were treated with osteogenic stimu-
latory factors that induced their differentiation. Differentiation was
detected by FTIR through changes in the amide I band shape (indica-
tive of protein composition/structural changes) and phosphate levels
(indicative of the expression of calcium phosphate salts).
Substrate Influences
In a similar manner to compositional differences in the growth media
that may or may not elicit changes in cell biochemistry and thus its
spectra, substrates can also induce morphological as well as func-
54
tional changes in the cell. Meade et al. studied the influence of a
range of substrates on the normal human epithelial keratinocyte cell
line (HaCaT) using a multimodal approach that included fluores-
cence, FTIR and Raman spectroscopies. The substrate extracellular
matrix (ECM) coatings under evaluation were two glycoproteins,
fibronectin and laminin and one protein, gelatin (derived from ther-
mal denaturing of collagen). Gelatin was coated onto MirrIR slides