Spatial Analysis of High Parameter Imaging Data to Understand Early HIV-Cell Interactions in Colorectal Tissue

Abstract

Although Human Immunodeficiency Virus (HIV) was first discovered over 30 years ago,there remains no cure or vaccine for HIV. In Australia, HIV transmission almost exclusively occurs via sexual intercourse, with over 75% occurring as a result of anal intercourse. Due to technological limitations, there are still major gaps in understanding how HIV is transmitted across the human colorectum, with the role of key HIV target cells being unclear. Additionally, traditional microscopy methods have not been able to visualize the full repertoire of HIV target cells within tissue. To investigate cellular interactions during early HIV transmission, we topically infect fresh human colorectal tissue with HIV as a model of early HIV transmission in situ. We then make use of Cyclic Immunofluoresecence, a high parameter microscopy imaging approach, to visualise key HIV target cells, namely dendritic cells, macrophages and CD4+ T lymphocytes could be observed within the same tissue section. Furthermore, through the in situ hybridization technique RNAscope, we are able to visualize HIV with single-virion sensitivity. Cyclic Immunofluorescence and RNAscope provide a unique opportunity for visualizing the early events of HIV transmission, allowing the interactions of HIV with its target cells as well as the colorectal tissue structure to be captured. However, given the recent development of these techniques, the imaging data being produced provide many unique challenges in terms of computational analysis. In particular, the presence of autofluorescence makes it difficult to reliably assign marker expression to cells while spatial analytical techniques are limited when multiple cell types are measured simultaneously. This thesis presents three publications which address these analytical gaps, while also detailing the computational analysis techniques utilized in the analysis of the HIV imaging dataset.