Liver is subject to various chronic pathologies, progressively leading to cirrhosis, which is associated with an increased risk of hepatocellular carcinoma. There is an urgent need for diagnostic and prognostic markers of chronic liver diseases and liver cancer. Spectroscopy-based approaches can provide an overview of the chemical composition of a tissue sample offering the possibility of investigating in depth the subtle chemical changes associated with pathological states. In this study, we have addressed the potential of spectroscopy-based approaches for diagnosis of Wilson disease as well as for investigating the composition and quantification of steatosis.
Wilson's disease (WD) is a rare autosomal recessive disease due to mutations of the gene encoding the copper-transporter ATP7B. The diagnosis is hampered by the variability of symptoms induced by copper accumulation, the inconstancy of the pathognomonic signs and the absence of a reliable diagnostic test. We investigated the diagnostic potential of X-ray fluorescence (XRF) that allows quantitative analysis of multiple elements. XRF experiments were first performed using synchrotron radiation to address the elemental composition at the cellular level. High-resolution mapping of tissue sections allowed measurement of the intensity and the distribution of copper, iron and zinc while preserving the morphology. Investigations were further conducted using a laboratory X-ray source for irradiating whole pieces of tissue. XRF on whole formalin-fixed paraffin embedded needle biopsies allowed profiling of the elements in a few minutes. The intensity of copper related to iron and zinc significantly discriminated WD from other genetic or chronic liver diseases with high specificity and sensitivity. This study established a definite diagnosis of Wilson's disease based on XRF. This rapid and versatile method can be easily implemented in a clinical setting.
Steatosis is one of the most important factors affecting liver allograft function. The gold standard to assess hepatic steatosis in liver grafts during the transplantation procedure is the histologic examination of frozen sections by a pathologist. The major issue is that assessment of hepatic steatosis on histologic sections is an imperfect and not reproducible method. We addressed the potential of Fourier transform-infrared (FTIR) microspectroscopy for grading steatosis on frozen tissue sections. The use of the bright infrared source emitted by synchrotron radiation allowed the investigation of the biochemical composition at the cellular level. We demonstrated that the progression of steatosis corresponds not only to the accumulation of lipids but also to dramatic changes in the qualitative composition of the tissue. We observed that dramatic biochemical changes occur in the non-steatotic part of the tissue despite its normal histological aspect, suggesting that the whole tissue reflects the grade of steatosis. We further developed a method for the quantification of lipid content on tissue section. This rapid method that takes only 1 minute has been implemented on laboratory instrument. Thus, the method can be easily used at the hospital for reliable assessment of graft quality control in liver transplantation.
François LE NAOUR
UMR-S1193, Hôpital Paul Brousse, Villejuif