Fatty-acid uptake in prostate cancer cells using dynamic microfluidic raman technology

It is known that intake of dietary fatty acid (FA) is strongly correlated with prostate cancer progression but is highly dependent on the type of FAs. High levels of palmitic acid (PA) or arachidonic acid (AA) can stimulate the progression of cancer. In this study, a unique experimental set-up consisting of a Raman microscope, coupled with a commercial shear-flow microfluidic system is used to monitor fatty acid uptake by prostate cancer (PC-3) cells in real-time at the single cell level. Uptake of deuterated PA, deuterated AA, and the omega-3 fatty acids docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) were monitored using this new system, while complementary flow cytometry experiments using Nile red staining, were also conducted for the validation of the cellular lipid uptake. Using this novel experimental system, we show that DHA and EPA have inhibitory effects on the uptake of PA and AA by PC-3 cells

Increased optical pathlength through aqueous media for the infrared microanalysis of live cells

The study of live cells using Fourier transform infrared spectroscopy (FTIR) and FTIR microspectroscopy (FT-IRMS) intrinsically yields more information about cell metabolism than comparable experiments using dried or chemically fixed samples. There are, however, a number of barriers to obtaining high-quality vibrational spectra of live cells, including correction for the significant contributions of water bands to the spectra, and the physical stresses placed upon cells by compression in short pathlength sample holders. In this study, we present a water correction method that is able to result in good-quality cell spectra from water layers of 10 and 12 μm and demonstrate that sufficient biological detail is retained to separate spectra of live cells based upon their exposure to different novel anti-cancer agents. The IR brilliance of a synchrotron radiation (SR) source overcomes the problem of the strong water absorption and provides cell spectra with good signal-to-noise ratio for further analysis. Supervised multivariate analysis (MVA) and investigation of average spectra have shown significant separation between control cells and cells treated with the DNA cross-linker PL63 on the basis of phosphate and DNA-related signatures. Meanwhile, the same control cells can be significantly distinguished from cells treated with the protein kinase inhibitor YA1 based on changes in the amide II region. Each of these separations can be linked directly to the known biochemical mode of action of each agent. Keywords: Synchrotron radiation (SR), Fourier transform infrared spectroscopy (FTIR), Infrared microspectroscopy (IRMS), Cancer, Single cell, Drug-cell interaction

Live single cell analysis using synchrotron FTIR microspectroscopy: development of a simple dynamic flow system for prolonged sample viability

Synchrotron radiation Fourier transform infrared microspectroscopy (SR-microFTIR) of live biological cells has the potential to provide far greater biochemical and morphological detail than equivalent studies using dehydrated, chemically-fixed single cells. Attempts to measure live cells using microFTIR are complicated by the aqueous environment required and corresponding strong infrared absorbance by water. There is also the additional problem of the limited lifetime of the cells outside of their preferred culture environment. In this work, we outline simple, cost-effective modifications to a commercially available liquid sample holder to perform single live cell analysis under an IR microscope and demonstrate cell viability up to at least 24 hours. A study using this system in which live cells have been measured at increasing temperature has shown spectral changes in protein bands attributed to α-β transition, consistent with other published work, and proves the ability to simultaneously induce and measure biochemical changes. An additional study of deuterated palmitic acid (D31-PA) uptake at different timepoints has made use of over 200 individual IR spectra collected over ∼4 hours, taking advantage of the ability to maintain viable cell samples for longer periods of time in the measurement environment, and therefore acquire greatly increased numbers of spectra without compromising on spectral quality. Further developments of this system are planned to widen the range of possible experiments, and incorporate more complex studies, including drug-cell interaction

Analysis of fixed and live single cells using optical photothermal infrared with concomitant raman spectroscopy

This paper reports the first use of a novel completely optically based photothermal method (O-PTIR) for obtaining infrared spectra of both fixed and living cells using a quantum cascade laser (QCL) and optical parametric oscillator (OPO) laser as excitation sources, thus enabling all biologically relevant vibrations to be analyzed at submicron spatial resolution. In addition, infrared data acquisition is combined with concomitant Raman spectra from exactly the same excitation location, meaning the full vibrational profile of the cell can be obtained. The pancreatic cancer cell line MIA PaCa-2 and the breast cancer cell line MDA-MB-231 are used as model cells to demonstrate the capabilities of the new instrumentation. These combined modalities can be used to analyze subcellular structures in both fixed and, more importantly, live cells under aqueous conditions. We show that the protein secondary structure and lipid-rich bodies can be identified on the submicron scale

3D DESI-MS lipid imaging in a xenograft model of glioblastoma : a proof of principle

Desorption electrospray ionisation mass spectrometry (DESI-MS) can image hundreds of molecules in a 2D tissue section, making it an ideal tool for mapping tumour heterogeneity. Tumour lipid metabolism has gained increasing attention over the past decade; and here, lipid heterogeneity has been visualised in a glioblastoma xenograft tumour using 3D DESI-MS imaging. The use of an automatic slide loader automates 3D imaging for high sample-throughput. Glioblastomas are highly aggressive primary brain tumours, which display heterogeneous characteristics and are resistant to chemotherapy and radiotherapy. It is therefore important to understand biochemical contributions to their heterogeneity, which may be contributing to treatment resistance. Adjacent sections to those used for DESI-MS imaging were used for H&E staining and immunofluorescence to identify different histological regions, and areas of hypoxia. Comparing DESI-MS imaging with biological staining allowed association of different lipid species with hypoxic and viable tissue within the tumour, and hence mapping of molecularly different tumour regions in 3D space. This work highlights that lipids are playing an important role in the heterogeneity of this xenograft tumour model, and DESI-MS imaging can be used for lipid 3D imaging in an automated fashion to reveal heterogeneity, which is not apparent in H&E stains alone

Mind the Gap: Transitions Between Concepts of Information in Varied Domains

The concept of 'information' in five different realms – technological, physical, biological, social and philosophical – is briefly examined. The 'gaps' between these conceptions are dis‐ cussed, and unifying frameworks of diverse nature, including those of Shannon/Wiener, Landauer, Stonier, Bates and Floridi, are examined. The value of attempting to bridge the gaps, while avoiding shallow analogies, is explained. With information physics gaining general acceptance, and biology gaining the status of an information science, it seems rational to look for links, relationships, analogies and even helpful metaphors between them and the library/information sciences. Prospects for doing so, involving concepts of complexity and emergence, are suggested