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Abstract

The human prostate is a significant source of disease burden for adult males. In the malignant context, prostate cancer is the most common non-cutaneous cancer in males. Benign prostatic hyperplasia is also common, affecting a majority of males over the age of 60. This high disease burden has been a focus of study for many years, but an answer to the ultimate question of what predisposes the prostate to such a high level of disease has eluded researchers. One possible explanation for the disease burden of the prostate may lie in the capacity of the prostate for hormone-dependent regeneration. This is a process by which the prostate shrinks in the absence of androgen signaling, and regenerates to its original size when androgen is reintroduced to the system. The study of hormone-dependent regeneration has allowed researchers to analyze the progenitor cell populations of the prostate. In this thesis, single cell RNA-Seq approaches were used to investigate the progenitor cell populations of the prostate. First, I investigated cells harvested from monolayer and organoid culture conditions to better understand the progenitor populations present in these models and differences between the models overall. This yielded evidence that prostate progenitor cells expressing Keratin 13 were preserved in both monolayer and organoid conditions. This presence of prostate progenitor cells was validated using immunofluorescence microscopy targeting Keratin 13 protein. In comparing the single cell RNA-Seq data from the two culture conditions, we were able to observe an enrichment of proliferating populations in the monolayer sample and an enrichment of intermediate cells in the organoid sample. Further comparison of these in vitro samples with in vivo prostate data gathered by another lab provided evidence that the in vitro samples were enriched for proliferating cells while the in vivo sample contained terminally differentiated cell populations that were not observed in vitro. These data provide an in-depth validation of the preservation of prostate progenitor cells in commonly used in vitro models, as well as providing insights into the different cell populations selected for in monolayer and organoid culture conditions respectively. Application of single cell RNA-Seq approaches to in vivo mouse prostate led to the identification of a luminal progenitor cell population in both the intact and castrate mouse prostate. These cells expressed luminal keratins as well as multiple putative progenitor cell markers. The presence of luminal progenitor cells in the mouse prostate was also validated using both immunofluorescence microscopy and flow cytometry. Pathway analysis of the expression data from luminal progenitor cells allowed for the selection of candidate factors likely to contribute to the prostate progenitor cell phenotype. Small molecule inhibitor treatment targeting two of these factors, Yap1 and Bcl-2, caused a significant decrease in in vitro regeneration of organoids derived from both mouse and human cells. These results provide in-depth expression data for luminal progenitor cells and also identify factors necessary for the prostate progenitor cell phenotype. These factors can be leveraged to better understand luminal progenitor cells and possibly be used to treat prostate disease in the future.

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