Microchip for Immunomagnetic Sorting of Circulating Tumor Cells (CTCs).

Circulating tumor cells (CTCs) isolated directly from whole blood opens new perspectives for cancer monitoring and the development of personalized treatments. However, due to their rarity among the multitude of blood cells, it remains a challenge to recover them alive with high level of purity, i.e., with few remaining white blood cells, and in a time frame compatible with the clinical context. Microfluidic chips have emerged as promising tools to address these challenges. We propose a two-step workflow including a pre-enrichment step, performed by a size-based pre-enrichment system, and a purification step, performed by an immunomagnetic chip. Here, we describe the protocol for the fabrication of the immunomagnetic microchip, the preparation of the sample, and the procedure for injection into the microchip allowing the sorting of the CTCs.

In vivo hepatic lipid quantification using MRS at 7 Tesla in a mouse model of glycogen storage disease type 1a.

The assessment of liver lipid content and composition is needed in preclinical research to investigate steatosis and steatosis-related disorders. The purpose of this study was to quantify in vivo hepatic fatty acid content and composition using a method based on short echo time proton magnetic resonance spectroscopy (MRS) at 7 Tesla. A mouse model of glycogen storage disease type 1a with inducible liver-specific deletion of the glucose-6-phosphatase gene (L-G6pc(-/-)) mice and control mice were fed a standard diet or a high-fat/high-sucrose (HF/HS) diet for 9 months. In control mice, hepatic lipid content was found significantly higher with the HF/HS diet than with the standard diet. As expected, hepatic lipid content was already elevated in L-G6pc(-/-) mice fed a standard diet compared with control mice. L-G6pc(-/-) mice rapidly developed steatosis which was not modified by the HF/HS diet. On the standard diet, estimated amplitudes from olefinic protons were found significantly higher in L-G6pc(-/-) mice compared with that in control mice. L-G6pc(-/-) mice showed no noticeable polyunsaturation from diallylic protons. Total unsaturated fatty acid indexes measured by gas chromatography were in agreement with MRS measurements. These results showed the great potential of high magnetic field MRS to follow the diet impact and lipid alterations in mouse liver.

Targeted deletion of liver glucose-6 phosphatase mimics glycogen storage disease type 1a including development of multiple adenomas.

BACKGROUND AND AIMS: Glycogen storage disease type 1a (GSD1a) is an inherited disease caused by a deficiency in the catalytic subunit of the glucose-6 phosphatase enzyme (G6Pase). GSD1a is characterized by hypoglycaemia, hyperlipidemia, and lactic acidosis with associated hepatic (including hepatocellular adenomas), renal, and intestinal disorders. A total G6pc (catalytic subunit of G6Pase) knock-out mouse model has been generated that mimics the human pathology. However, these mice rarely live longer than 3 months and long-term liver pathogenesis cannot be evaluated. Herein, we report the long-term characterization of a liver-specific G6pc knock-out mouse model (L-G6pc(-/-)). METHODS: We generated L-G6pc(-/-) mice using an inducible CRE-lox strategy and followed up the development of hepatic tumours using magnetic resonance imaging. RESULTS: L-G6pc(-/-) mice are viable and exhibit normoglycemia in the fed state. They develop hyperlipidemia, lactic acidosis, and uricemia during the first month after gene deletion. However, these plasmatic parameters improved after 6 months. L-G6pc(-/-) mice develop hepatomegaly with glycogen accumulation and hepatic steatosis. Using an MRI approach, we could detect hepatic nodules with diameters of less than 1 mm, 9 months after induction of deficiency. Hepatic nodules (1 mm) were detected in 30-40% of L-G6pc(-/-) mice at 12 months. After 18 months, all L-G6pc(-/-) mice developed multiple hepatocellular adenomas of 1-10 mm diameter. CONCLUSIONS: This is the first report of a viable animal model of the hepatic pathology of GSD1a, including the late development of hepatocellular adenomas.

Quantification method using the Morlet wavelet for magnetic resonance spectroscopic signals with macromolecular contamination.

We study the Morlet wavelet transform on characterizing Magnetic Resonance Spectroscopic (MRS) signals acquired at short echo-time. These signals contain contributions from metabolites, water and a baseline which mainly originates from large molecules, known as macromolecules, and lipids. The baseline signal decays faster than the metabolite ones. Therefore, by making use of the time-scale representation of the wavelet, the two signals can be distinguished without any additional pre-processing. This is confirmed by the experimental results which show that the Morlet wavelet can correctly quantify the metabolite contributions even when a baseline is embedded in the MRS signals.

Influence of measured and simulated basis sets on metabolite concentration estimates.

By quantification of brain metabolites, localized brain proton MRS can non-invasively provide biochemical information from distinct regions of the brain. Quantification of short-TE signals is usually based on a metabolite basis set. The basis set can be obtained by two approaches: (1) by measuring the signals of metabolites in aqueous solution; (2) by quantum-mechanically simulating the theoretical metabolite signals. The purpose of this study was to compare the effect of these two approaches on metabolite concentration estimates. Metabolite concentrations were quantified with the QUEST method, using both approaches. A comparison was performed with the aid of Monte Carlo studies, by using signals simulated from both basis sets. The best results were obtained when the basis set used for the fit was the same as that used to simulate the Monte Carlo signals. This comparison was also performed using in vivo short-TE signals acquired at 7 T from the central region of rat brains. The concentration estimates, with confidence intervals, obtained using both basis sets were in good agreement with values from the literature. The in vivo study showed that, in general, the differences between the estimates obtained with the two basis sets were not statistically significant or scientifically important. Consequently, a simulated basis set can be used in place of a measured basis set.