Spatially Resolved, Functional Dissection of the Human Spermatogonial Stem Cell Niche

PROJECT SUMMARY The functions of human spermatogonial stem cells (SSCs), including self-renewal and differentiation, are required for the constant production of male gametes over a long reproductive lifespan; imbalances in this process directly contribute to infertility or germ cell-derived cancers. Despite this wide range of healthcare implications, the cellular and molecular regulation of human SSCs is poorly understood, in particular with respect to the microenvironment in which SSCs reside, termed the niche. Despite research efforts, a major gap in knowledge remains, i.e., how does the SSC niche influence the functions of human SSCs? Our long-term goal is to understand the molecular mechanisms that regulate human SSCs. Unlike rodents, for which genetic models are routinely used to evaluate the roles of the niche in regulating SSC functions in vivo, functional dissection of the human SSC niche has been challenging largely due to a lack of experimental tools. Single cell omics has enabled a molecular catalogue of human testicular cell types but falls short on providing insights into the spatial and functional interactions between SSCs and the niche due to tissue disassociation. We have recently established a spatial transcriptomics (ST) approach that successfully recapitulates many aspects of the mouse and human spermatogenesis. This approach quantifies genome-wide gene expression of individual testicular cells within intact tissue sections. Using this ST approach, coupled with a series of functional, cellular, and molecular analyses, we will test our central hypothesis that human SSCs are functionally regulated by the niche through a selective set of ligand-receptor (LR) interactions. First, we will perform a systematic characterization of LR interactions at the human SSC niche. Second, we will comparatively study the human and mouse SSC niches under the hypothesis that human specific LR pairs may be crucial to human SSC functions. Finally, we will examine the contribution of germline cells to the functions of human SSC niche. The successful completion of the proposed work will significantly enhance our mechanistic understanding of the functional role of the niche in regulating human SSC activities. It will also provide much needed insights into the etiology of male infertility, fertility preservation for cancer patients, and the successful establishment of in vitro spermatogenesis.