Cell substrate interaction studies have also received

Cell-substrate interaction studies have also received extensive attention and are studied with acoustic sensors; cell adhesion response can vary with the cell type and surface coating as well as surface characteristics like topography, composition, energy and mechanical properties [19–23]. Up to now, the QCM-D (or TSM-R) has been used to study the attachment and spreading of various mammalian cell lines such as NIH 3T3 [24,25], MCF-7 [9], MDCK [26] and LG2 [11,27]. Cell surface-adhesion studies were performed on uncoated metal surfaces like gold [14], tantalum [28], titanium [29], hydroxyapatite [30] and silica [31]. Functionalization of metal surfaces includes adsorption of ECM proteins [32,33], modification by RGD peptides [34,35], formation of supported lipid bilayers [36,37] and multilayer buildup [38]. Among all the metal substrates, gold is used as a standard reference material while titanium finds applications in the orthopedic and dental implant industry. Fibrinogen, an extracellular protein found in blood in concentrations of 3mgmL, is also used as a surface coating [39,40]. Finally, the QCM-D has also been utilized in tumor disease studies. Frequency fluctuations have been monitored as an indicator of cell micromotility to assess the invasiveness of colon and pharynx derived malignant ceramide [41]. In other studies, the dynamic viscoelastic properties of normal (HMEC) and malignant (MCF-7) cells were evaluated upon adhesion and spreading on gold [9] while lectin-induced cell agglutination was employed to discriminate between normal (L-02) and cancer (Bel7402) hepatic cells [42].
Anaplastic Thyroid Cancer (ATC) is a fatal tumor malignancy of the human endocrine system; average survival rate is estimated in months and one-year survival is 15% [43]. ATC accounts for a minority (<2%) of all the thyroid malignancies and its rarity is counterbalanced by its aggressive phenotype. If the primary tumor is treated with surgery long term survival is possible [44]; nevertheless at later stages, the tumor is resistant to all methods of treatment like surgery, radiotherapy and chemotherapy [43,45]. Until now, there is little advance in disease-specific biomarker research, hence the disease is characterized by low prognosis [46,47]. In this study, we examined the potential of QCM-D as a diagnostic tool of ATC, by trying to elucidate differences between the adhesion pattern of normal thyroid and anaplastic thyroid cancer cells. Starting from the hypothesis that cancer cells may exhibit a different adhesion ability we decided to exploit possible differences in cell adhesion patterns on three surfaces, namely, titanium, gold and fibrinogen-coated gold. In parallel, scanning electron microscopy images were used to possibly correlate the morphology of cells on these surfaces with acoustic measurements. Results indicate that, indeed, screening of the two types of cells can be achieved by combining adhesion patterns of each cell line on the above three surfaces. This result points to the potential use of the QCM sensor as a diagnostic platform, complementary to other existing ones, for cell malignancy detection.

Materials and methods

Results

Discussion
Thyroid cancer has received little attention because of its rarity but demographic results suggest there is a need for better prognosis and treatment of the tumor. In fact, anaplastic tumor thyroid cells possess different genetic and phenotypic characteristics when compared to normal ones. Their different size, stage of differentiation and proteomic profile [49–51] could imply a different response upon adhesion to surfaces. In exploring this possibility, acoustic sensors and electron microscopy were employed to study adhesion of thyroid cells to various surfaces. Two cell lines were used here as a model in order to create an acoustic diagnostic tool for human thyroid cancer.
Real time acoustic signals were used to monitor cell adhesion events as a function of the cell concentration and surface-type. Four types of acoustic data were extracted from the real time graphs; the frequency change, which is related to the number of adhered cells; the dissipation change which is related to the viscoelastic properties of the bound cells; the acoustic ratio which refers to the mass-calibrated change in the viscoelastic properties of each cell line; and, finally, the rate of cell binding to the surface. Frequency and energy dissipation measurements were used to derive cell adhesion binding curves (Fig. 2). The binding curves were different for each surface. However, the trend in the acoustic responses was similar: both cell lines gave higher signals on the Ti surface followed by their adhesion to Au and last to Fg-Au (Ti>Au>Fg-Au). Au and Ti have different mechanical and surface properties regarding their hydrophilicity, topography etc. and here Ti proved to be a higher-adhesion substrate than Au. The Au surface is widely used as a reference surface and here is proved to have high-adhesion properties. On the other hand, fibrinogen seemed to be a low-adhesion substrate.