Research

Research in the Dobrinski lab is focused on mammalian germ line stem cell biology and the regulation of its niche. Germline stem cells (spermatogonial stem cells, SSCs) form the basis of male fertility by developing into sperm. SSCs are the only cells in an adult body that divide and can contribute genes to subsequent generations, making them valuable for regenerative medicine and targets for genetic manipulation. Our lab currently works on developing strategies to model and study the effects of the testicular microenvironment on germ cell differentiation and to enable full spermatogenesis in vitro.

Study of human and animal disease requires appropriate models. While there are many rodent models, larger animal models such as pigs more closely represent human physiology, size and longevity. Testicular maturation takes much longer in pigs and humans in comparison to rodents, and we’ve described dynamic changes in the metabolic phenotype of germ and somatic cells during prepubertal development in pig and human testis (Voigt et al., 2021, 2022, 2023). Considering these similarities, by doing research in porcine models, we expect our findings to be more translatable within reproductive health research. 

We pioneered SSC transplantation in non-rodent models (Honaramooz et al., 2002a, 2003a, 2003b), and first reported germline transgenesis in large animals through modification and transplantation of SSCs (Honaramooz et al., 2008a; Zeng et al., 2012, 2013; Tang et al., 2015; Lara et al., 2023), enabling the generation of genetically modified non-rodent animal models (Fig 1). We also developed testis xenografting for complete recapitulation of mammalian spermatogenesis while maintaining accessibility in a mouse host (Honaramooz et al., 2002b, 2004, 2007), translating this approach to cattle, horses, sheep, cats, humans, and endangered species (Snedaker et al., 2004; Rathi et al., 2005, 2006; Arregui et al., 2008, 2014). Besides being an accessible system to study testis morphogenesis and sperm production in vivo (Arregui & Dobrinski, 2014), sperm from xenografts can support embryo development after ICSI (Honaramooz et al., 2008b, Fayomi et al., 2019) (Fig 1). These approaches present a potential avenue for fertility preservation and restoration.

Fig. 1: Schematic representation of SSC transplantation (top) and xenografting (bottom) approaches. Adapted from Honaramooz et al., 2002; Tang et al., 2012. Scale bar: 5 mm

Fig. 1: Schematic representation of SSC transplantation (top) and xenografting (bottom) approaches. Adapted from Honaramooz et al., 2002; Tang et al., 2012. Scale bar: 5 mm

Fig. 2: Testicular organoid formation in microwells (top), organoid cytoarchitecture schematics (bottom left) and immunostaining (bottom right). Adapted from Sakib et al., 2019. Scale bars: 100 µm.

Fig. 2: Testicular organoid formation in microwells (top), organoid cytoarchitecture schematics (bottom left) and immunostaining (bottom right). Adapted from Sakib et al., 2019. Scale bars: 100 µm.

There have been many approaches to studying the testicular microenvironment and recapitulating spermatogenesis in vitro, including platforms like testicular organoids (Fig 2). These are 3D mini testicles that can perform in a culture dish many functions that usually take place in the testicles. Our laboratory has developed organotypic testis organoids from rodents, pigs, non-human primates and human immature testicular cells (Sakib et al., 2019, 2022), allowing us to investigate cell–cell interactions, SSC niche signalling, reproductive toxicology, and the structural organization required to support germ cell maintenance and differentiation, as well as testicular maturation.

Extracellular vesicles (EVs), including exosomes, microvesicles and apoptotic bodies, are micro- and nano-sized vesicles secreted by cells into the extracellular space that are increasingly recognized as a significant intercellular communication system. We study EVs as critical mediators of cell-cell communication within the testicular microenvironment and how they influence spermatogonial development (Thiageswaran et al., 2022) (Fig 3). By characterizing their molecular cargo and functional effects, we aim to define how EV-mediated signalling regulates germ cell fate decisions and contributes to the establishment and maintenance of a supportive SSC niche.

Fig. 3: Extracellular vesicles derived from Sertoli cells (A, electron microscopy; B, confocal microscopy, green) may contribute to the SSC niche (C). Adapted from Thiageswaran et al., 2022. Scale bars: A = 0.2 µm; B = 8 µm.

Fig. 3: Extracellular vesicles derived from Sertoli cells (A, electron microscopy; B, confocal microscopy, green) may contribute to the SSC niche (C). Adapted from Thiageswaran et al., 2022. Scale bars: A = 0.2 µm; B = 8 µm.

Fig. 4: Porcine iPSCs can be maintained in both static (A,C) and suspension culture (B,D), maintaining expression of pluripotency markers (C,D). Adapted from Lara et al., 2021. Scale bars: 25 µm.

Fig. 4: Porcine iPSCs can be maintained in both static (A,C) and suspension culture (B,D), maintaining expression of pluripotency markers (C,D). Adapted from Lara et al., 2021. Scale bars: 25 µm.

Induced pluripotent stem cells (iPSCs) are adult cells genetically reprogrammed to a pluripotent stem cell state, acquiring the ability to self-renew and differentiate into nearly any cell type of the body, holding enormous potential for disease modelling, drug discovery, and regenerative medicine. We use pig (Burrell et al., 2019) and human iPSCs as a scalable and versatile platform to generate and genetically modify germ cell–like and somatic cell-like cells (Fig 4). This approach enables mechanistic studies of germline development and supports the reconstruction of spermatogenesis in vitro.