Del Mar Photonics - Newsletter April 2011
Description of the application of multiphoton imaging from one of our potential customers
Diagnosing liver fibrosis: a new scoring method based on nonlinear microscopy
Liver fibrosis refers to the scarring response of the liver to external aggressions, like hepatitis B or C viruses (HBV, HCV) infecting more than 500 million people worldwide or excessive alcohol intake. The over-accumulation of fibrous tissue (mainly collagen) modifies liver function, and ultimately leads to cirrhosis and life threatening complications such as primary liver cancer. Cirrhosis is the leading cause for liver transplantation in France. In Rennes, more than 3800 patients with HCV infection are treated in CHU Pontchaillou.
Thus, liver chronic diseases constitute an important public health issue, and it is important to have a reliable diagnosis and early detection of fibrosis. Liver biopsy is the gold standard for assessing liver fibrosis and cirrhosis but the semi-quantitative scores used until now are inappropriate for accurate follow-up (especially for the assessment of the effect of antifibrotic treatments) and are subjected to possible intra and inter observer lecture variations. Therefore, this study was motivated by the need to develop a robust scoring system for diagnosing liver fibrosis/cirrhosis that avoids the risk of inter-observer variations and allows for precise quantitative measurement of the amount of collagen, as well as providing new informations on the structure of collagen deposits and 3D reconstruction of the ECM network arrangement.
Nonlinear microscopy is a unique tool which provides intrinsic optical sectioning and high in-depth imaging due to the inherent localization of the nonlinear excitation at the objective focal volume, while drastically reducing out-of-focus photobleaching and phototoxicity. Moreover, type-I or III collagen, the main component of the fibrosis deposit in liver, can be selectively imaged by the second harmonic generation (SHG) signals without staining. From this specific contrast, we developed a new scoring method in assessing human liver fibrosis . This multidisciplinary work is an example of fruitful collaboration between physicists (IPR/UMR CNRS 6251), biologists (EA Seraic/IRSET, IFR140) and hepatologists and pathologists (CHU Pontchaillou).
Although SHG scoring may lead to a more accurate diagnosis of liver fibrosis, our study also opens many perspectives . In particular, we are working in using the potentialities of 3D reconstruction of collagen networks, as well as orientational patterns [3,4], to get a better insight into the collagen deposit at various scales and fibrosis levels. Feasibility of endoscopic SHG exploration for rodent models is also planned.
Legend: Nonlinear microscopy of human liver biopsies. Two-photon excitation fluorescence (TPEF) and Second-harmonic generation (SHG) were acquired simultaneously in 2D or 3D image stacks. SHG images reveal selectively fibrillar collagen of type I or III without staining. A score proportional to the amount of collagen deposit was derived from these images (Image courtesy of Universite de Rennes 1).
 Gailhouste L., Le Grand Y., Odin C., Guyader D., Turlin B., Ezan F., Desille Y., Guilbert T., Bessard A., Fremin C., Theret N., Baffet G., "Fibrillar collagen scoring by second harmonic microscopy: A new tool in the assessement of liver fibrosis." Journal of Hepatology, 52, (3), 398-406 (2010).
 Bedossa P., Editorial: "Harmony in liver fibrosis..." Journal of Hepatology, 52, (3), 313-314 (2010).
 Odin C., Guilbert T., Al Kilani A., Boryskina O.P., Fleury V., Le Grand Y., "Collagen and myosin characterization by orientation field second harmonic microscopy.", Optics express, 16, (20), 16151-16165 (2008).
 Odin C., Le Grand Y., Renault A., Gailhouste L., Baffet G.,"Orientation Fields of Non-Linear Biological Fibrils by Second Harmonic Generation Microscopy." Journal of Microscopy, 229, 32-38 (2008).
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