Investigations into the changes in biomechanics of liver cells upon HCV infection

Abstract

Biomechanics is an important regulator of cell function. Active (external forces) as well as passive (substrate stiffness) mechanical stimuli from the cell microenvironment can alter fundamental aspects of cell function such as metastasis and differentiation. Since diseases are a dysfunction of cellular function, there could be alterations of biomechanics due to diseases. These alterations could in turn contribute to some part of the disease through defective mechano-transduction. Alterations in the mechanical properties of cells were observed mostly in cancer and malaria and these changes are being exploited for diagnosis as a mechanical biomarker of the disease. In this bioengineering study, we investigated the changes in biomechanics of liver cells due to a viral infection. We consider Hepatitis C Virus infection and the changes in nuclear mechanics in the light of some of the previous work done in our lab. Biomechanical studies suffer from a dearth of standard equipment, experimental protocols, and analytical techniques. Hence, the development of devices, experimental and analytical techniques, wherever necessary, also form a part of this thesis. The new methods developed in this thesis not only serve the purpose of addressing the specific biological question addressed in this thesis, but also apply, in general, to other biomechanical studies. Therefore, this thesis is as much about the experimental and analytical techniques developed as it is about the specific biological phenomena discovered. We have designed and fabricated devices (perfusion culture system and microchannel flow setup), developed algorithms (nuclear segmentation using image processing, analyzing F-d curves from AFM), used computational modelling (mechanical model of nuclear morphology), and performed biological experiments (nuclear morphology, cell and nuclear stiffness, protein expression levels and response of cells to shear stress due to flow) to discover a hitherto unknown deregulation of the nuclear mechanics of liver cells due to HCV infection. The nucleus becomes more flexible and susceptible to shear stress due to flow. We also show that these alterations in the mechanics of the nuclei are due to the downregulation of Lamin A/C. It is known that nuclear shape affects gene expression, and disruption of the nuclear lamina can lead to gross changes in chromatin dynamics and regulation. Hence, our results suggest that some of the pathogenesis of HCV could be due to global changes in gene expression by the deregulation of nuclear morphology and mechanics.