| Increase developmental maturity of pluripotent stem cell–derived organoids | Long-term culture (Nicholas et al., 2013; Takasato et al., 2016; Tzatzalos et al., 2016) | Stem cells are highly sensitive to culture conditions, impeding robust protocols |
| | In vivo maturation (Huch et al., 2013; Watson et al., 2014; Takebe et al., 2015) | |
| | Mechanical or electrical conditioning of muscle and cartilage (Ruan et al., 2016) | |
| | Improved culture conditions and differentiation protocols | |
| | Acceleration by small molecules (Chambers et al., 2012) | |
| Source culturable, age-varied human cells | Surgical discards from elective surgery, transplant trimmings, and tissue peripheral to tumors | Hard to source certain tissues, especially healthy, culturable adult cells |
| | | Cells adapt to culture and are not infinitely renewable |
| | | Nontumor peripheral tissue may differ from healthy tissue |
| In vitro aging to model the effects of specific age-related lesions and provide a source of artificially aged cells | Induced senescence by DNA damage or environmental stress (Busuttil et al., 2003; Parrinello et al., 2005; Nasto et al., 2013) | Controversial which treatments best phenocopy aging |
| | Long-term culture (Dos Santos et al., 2015) | |
| | Progeria mutations (Liu et al., 2011; Zhang et al., 2011; Miller et al., 2013) | |
| | Direct reprogramming of aged cells (Mertens et al., 2015) | |
| Mimic the effects of the aged ECM in vitro | ECM from aged donors (Gullapalli et al., 2005; Williams et al., 2014; Stearns-Reider et al., 2017) | Controversial which treatments best phenocopy aging |
| | Glycation crosslinking (Rodriguez-Teja et al., 2015) | ECM extraction from tissues can alter its mechanical properties, microstructure, and composition |
| | Enzymatic crosslinking (Levental et al., 2009) | |
| Develop aging-relevant experimental readouts | Epigenetic clock (Hannum et al., 2013; Horvath, 2013) | Unclear which signs of aging are most important |
| | Mutational analysis (Blokzijl et al., 2016) | Require signs of aging that change appreciably across the span of an experiment |
| | Protein/DNA oxidation | |
| | Protein aggregation | |
| | Tissue-specific functional assays | |
| Long-term maintenance of organoids in a stable, growth-arrested state | Improved culture media | Relatively few published maintenance conditions |
| | Improved ECM and bioreactors | Vetting culture conditions is lengthy |
| Complete modeling of whole organs and physiological systems | Co-culture with immune cells, stromal cells, and microbiome bacteria (Parrinello et al., 2005; Engevik et al., 2015; Plaks et al., 2015) | Few good techniques for sophisticated organoid construction |
| Increase throughput and reproducibility | Vascularization (Auger et al., 2013) | Powerful techniques are often cumbersome and low-throughput |
| | Innervation (Workman et al., 2017) | |
| | In vivo implantation (Watson et al., 2014; Takebe et al., 2015) | |
| | Microfluidic access to apical/basal fluid reservoirs and fluid transport between organ systems (Vernetti et al., 2017) | |
| | Organ-specific ECM (Voytik-Harbin et al., 2007; Zhang et al., 2009; O’Brien et al., 2010) | |
| | Defined artificial ECMs may decrease lot-to-lot variability (Gjorevski et al., 2016) | |
| | Growth factor distribution within ECM gels for spatial control over growth and differentiation (Wylie et al., 2011) | |
| | Cell patterning for control over initial organoid shape and composition (Nelson et al., 2008; Todhunter et al., 2015) | |
| | 3D-printed gels amenable to perfusion with control over large-scale tissue structure (Kolesky et al., 2016) | |
| | Morphological screening and sorting to enrich for correctly formed organoids (Arora et al., 2017) | |