The biology of aging is focused on the identification of novel pathways that regulate the underlying processes of aging to develop interventions aimed at delaying the onset and progression of chronic diseases to extend lifespan. However, the research on the aging field has been conducted mainly in animal models, yeast, Caenorhabditis elegans, and cell cultures. Thus, it is unclear to what extent this knowledge is transferable to humans since they might not reflect the complexity of aging in people. An organoid culture is an in vitro 3D cell-culture technology that reproduces the physiological and cellular composition of the tissues and/or organs. This technology is being used in the cancer field to predict the response of a patient-derived tumor to a certain drug or treatment serving as patient stratification and drug-guidance approaches. Modeling aging with patient-derived organoids has a tremendous potential as a preclinical model tool to discover new biomarkers of aging, to predict adverse outcomes during aging, and to design personalized approaches for the prevention and treatment of aging-related diseases and geriatric syndromes. This could represent a novel approach to study chronological and/or biological aging, paving the way to personalized interventions targeting the biology of aging.
1. Introduction: Organoids, Spheroids, and Matrix-Embedded 3D Cultures
Patient-derived organoids (PDOs) are self-organized 3D tissue cultures that are derived from stem cells. Isolated patients’ stem cells differentiate to form an organ-like tissue comprising multiple cell types. Organoids have self-renewal and self-organization capabilities and retain the characteristics of the physiological structure and function of their source
[1][2][1,2]. Recent culturing advances aim to create the right environment for the stem cells so they can follow their own genetic instructions to self-organize, forming organoid structures that resemble miniature organs composed of many cell types. This approach provides tractable in vitro models of human physiology and pathology, thereby enabling interventional studies that are difficult or impossible to conduct in human subjects
[1][3][1,3]. Attempts to model the biology of human organs—including the differentiation of human stem cells in 2D, in either the presence or absence of a 3D matrix; bio-printing of human cells; and the culture of cells in a microfluidic device (“organ-on-a-chip”)—were made prior to the emergence of organoids and have shown some potential for drug screening or human disease research.
During the late twentieth and early twenty-first centuries, the use of classical cell lines and animal model systems in biomedical research has helped to improve our understanding of cellular signaling pathways, to identify potential drug targets and to guide the design of candidate drugs for pathologies including cancer and infectious diseases, among others
[3][4][3,4]. Recent studies have identified biological processes that are specific to the human body, such as brain development, metabolism, and the test of drug efficacy that cannot be modeled in animal or cell models. Nevertheless, extrapolating results from these model systems to humans has become a major bottleneck in the drug discovery process. Therefore, the emergence of human in vitro 3D cell cultures, such as organoids, spheroids, and matrix-embedded 3D cultures has received widespread attention due to the potential to overcome these limitations
[4][5][6][4,5,6]. These 3D structures of cultured cells recapitulate important aspects of in vivo organ development and biological function. Such cultures can be crafted to replicate much of the complexity of an organ or to express selected aspects of its physiology like producing only certain types of cells
[1][2][3][1,2,3].
Spheroids form by spontaneous aggregation of cells followed by the binding of cell surface integrins to the extracellular matrix (ECM). After initial cell–cell contact, cells upregulate E-Cadherin, which accumulates on the cell surface and then the spheroid becomes a compact structure through strong intercellular E-cadherin interactions
[6]. Different spheroid models have been described based on their cellular sources. Multicellular tumor spheroids (MCTS) are often made from cancer cell lines but rarely from tumor tissues. MCTS show little histological resemblance to the original tumor, but they mimic metabolic and proliferation gradients of the in vivo tumor and model clinically relevant resistance to chemotherapy. The advantages of MCTS are that they are clonal, simple to expand into large cultures, and suitable for high-throughput systems
[5][6][7][5,6,7].
In contrast, organoids grow from stem cells, which can divide indefinitely and produce different types of cells as part of their progeny. Organoids allow genetic and pharmacological manipulation in a complex cellular context that reflects human biology and enable investigations of the early stages of organ development and disease onset. They complement (and may eventually replace) animal models in many areas of preclinical drug development. Moreover, they provide patient-specific “avatars” for drug development and precision therapies, including treatments for cancer, rare genetic diseases (such as cystic fibrosis), and complex multifactorial disorders (such as epilepsy). Finally, they promise to contribute to regenerative medicine, with the goal of producing functional biological structures that can be transplanted into patients
[3][4][5][6][7][8][9][3,4,5,6,7,8,9].
2. Approaches to Generate a Patient-Derived Human Organoid: Surgical Resections, Liquid Biopsy, and iPSC-Derivation
The first organoids were developed from tissues of animal models. However, some biological aspects are unique to humans, and as a consequence, these models show limitations recapitulating human pathology. In that sense, PDOs emerged as a model to study cancer, infectious diseases, and inheritable genetic disorders
[10][11][12][13][10,11,12,13]. The generation of organoids requires the use of stem cells, which can be either (a) pluripotent stem cells (PSCs)—embryonic stem cells (ESC) and induced pluripotent stem cells (iPSCs)—or (b) adult stem cells (ASCs)
[7][8][9][13][7,8,9,13]. For instance, the source of PSC is restricted to iPSCs that are generated through the reprogramming of somatic cells, avoiding the ethical concerns of the use of ESC. iPSCs have the potential to generate all three germ layers. Differentiation into distinct cell and tissue types can be controlled in vitro by the sequential use of different factors that mimic in vivo organ development
[7][11][7,11]. In contrast, ASCs can be obtained from tissues with regenerative ability, and they have a limited differentiation potential. In that case, the starting material for the generation of the organoids is normal or malignant human tissue that can be obtained from surgical resection or biopsy
[10]. In fact, the generation of organoids from these sources allow the expansion and maintenance of this valuable material. The development of the organoids requires the use of specific growth factors depending on the tissue of origin, and they are mainly restricted to the growth of epithelial cells
[14]. For that reason, the ASC-derived organoids are less complex than the iPSCs-derived, which might include mesenchymal and epithelial constituents
[12]. On the other hand, tissue-derived organoids may recapitulate the genetic and epigenetic signature of the original organ
[14], but iPSCs can lose this kind of information due to the dedifferentiation process required for the establishment of the cell line
[15], thus hampering the use of iPSCs for preclinical models. In addition, liquid biopsies contain circulating tumor cells (CTCs), which although scarce in material could be good candidates to generate 3D structures, expanding the approaches that could be used to generate organoids
[16]. In any case, the procedures to establish organoids rely on the self-renewal and differentiation of tissue-resident stem cells that expand in culture and self-organize into complex three-dimensional structures. Once established, organoids can be initiated from cryopreserved material, cultured using largely traditional cell culture techniques and equipment, and then expanded and cryopreserved for future use
[17].
3. Organoids as a Model to Study Aging Signature across Tissues
Aging is the major risk factor for most chronic diseases. As a consequence of an increase in lifespan over the years, the elderly population is growing
[18]. Frequently, this extension in longevity is not being accompanied with an increase in health-span
[19]. Therefore, studying the underlying mechanisms of aging and developing interventions that target the aging process has become a priority field of research for most of the governments and private research agencies worldwide
[19].
Organoids might provide a new valuable tool to model the changes that occur during aging across tissues and to study the development of age-associated diseases. Impaired processes and/or damage at the molecular and cellular levels accumulate as we age, leading to a decrease in the reserve capacity or resilience, ultimately developing the aging phenotypes, which have been divided into four domains: body composition, energetic imbalance between availability and demand, homeostatic dysregulation, and neurodegeneration
[18][19][20][18,19,20]. Genomic instability, telomere attrition, epigenetic alterations, loss of proteostasis, deregulated nutrient sensing, mitochondrial dysfunction, cellular senescence, stem cell exhaustion, and altered intercellular communication have been identified as important hallmarks of aging in mammals
[20]. Modeling these hallmarks using organoids seems to be possible (
Table 1).
Table 1. Summary of studies using organoids to model aging.
Type of Organoid |
Addressed Hallmark of Aging |
Main Findings |
Reference |
Gut |
Stem cell exhaustion; deregulated nutrient sensing |
Lower O.F.E.; altered crypt formation |
[21][22][23][24][25][21,22,23,24,25] |
Epigenetic changes; cellular senescence |
Increased senescence markers; altered DNA methylation |
[21][26][27][21,26,27] |
Stem cell exhaustion; deregulated nutrient sensing |
CR increased O.F.E.; reduced mTOR signaling |
[28][29][28,29] |
Stem cell exhaustion; deregulated nutrient sensing |
NR supplementation increased O.F.E. |
[30] |
Altered intercellular communication |
Chronic inflammation led to NF-κB activation and cellular transformation |
[31] |
Genomic instability |
Tissue-specific mutational profile; tumor development |
[32][33][32,33] |
Liver |
Genomic instability |
Tissue-specific mutational profile; tumor development |
[32][34][32,34] |
Skin |
Cellular senescence; altered intercellular communication |
Increased senescence markers; decreased ECM synthesis |
[35][36][37][35,36,37] |
Cellular senescence; altered intercellular communication |
Adipose stem cells prevent skin senescence |
[38] |
Altered intercellular communication |
Altered TGF-β/Smad signaling |
[39] |
Tendon |
Stem cell exhaustion; cellular senescence; altered intercellular communication |
Lower O.F.E; decreased ECM synthesis; increased senescent markers |
[40] |
Lung |
Stem cell exhaustion; cellular senescence; telomer attrition |
Lower O.F.E; shortened telomeres; increased senescent markers |
[41] |
Breast |
Genomic instability |
Tumor development |
[42] |
Gastric |
Genomic instability; epigenetic changes; altered intercellular communication |
PDO characterization; altered Wnt signaling |
[43] |
Pancreatic |
Genomic instability; altered intercellular communication |
PDO characterization; altered Wnt signaling |
[44] |
Brain |
Loss of proteostasis |
Amyloid plaques and tau aggregates |
[45][45[46],46] |