Many physiological and pathological processes emerge from interactions between cellular populations rather than from the behavior of a single cell type. In neurodegenerative and retinal diseases, interactions between neurons, glial cells and immune populations play important roles in tissue homeostasis, inflammatory signaling and disease progression.
Human induced pluripotent stem cell (iPSC) technology enables defined human cell populations to be combined within controlled in vitro systems. These co-culture models provide valuable insights into cell-cell interactions and disease-associated cellular crosstalk that may not be apparent in monoculture systems.
In our new whitepaper, Modeling disease with co-culture of iPSC-derived cells: building complexity into neuroscience and ocular research, we explore the role of microglia in central nervous system (CNS) and retinal disease, the opportunities and practical challenges associated with developing complex co-culture models, and how advances in human iPSC technology are supporting the adoption of New Approach Methodologies (NAMs) in preclinical research.
Considerations for developing complex co-culture systems
As co-culture systems become increasingly sophisticated, biological complexity must be balanced with experimental robustness. Different cell types have distinct maturation timelines, metabolic requirements and environmental preferences, making careful optimization and characterization important when establishing complex co-culture systems.
Key considerations include the quality, viability and density of individual cell types, media composition, maturation processes, timing of cell addition and appropriate analytical readouts. Additional considerations arise when using patient-derived iPSC models, including donor-to-donor variation, reproducibility across experimental batches and the integration of functional readouts from multiple cell types.
Investigating cellular interactions in neurodegenerative disease
Although motor neuron degeneration remains the defining pathological feature of amyotrophic lateral sclerosis (ALS), disease progression is influenced by neighboring glial and immune cell populations. Astrocytes and microglia contribute to inflammatory signaling, neuronal stress responses and changes in the local microenvironment that affect motor neuron function and survival.
We generated iPSC-derived motor neurons from donors carrying ALS-associated mutations, alongside unaffected controls, and characterized their morphology, neuronal activity and disease-associated phenotypes. Incorporating astrocytes and microglia into a human tri-culture model altered motor neuron burst activity and synchronization, highlighting the influence of neuroimmune interactions on neuronal function.
These findings demonstrate how co-culture systems can reveal phenotypes not observed in neuronal monocultures, supporting more physiologically relevant disease modeling and the evaluation of novel therapeutics.
Extending co-culture models to retinal disease
Although neurological and retinal disorders have traditionally been studied separately, the retina is a developmental extension of the CNS and contains specialized neuronal and glial populations organized into highly interconnected cellular networks. These populations share many of the biological processes observed in the brain, including immune surveillance, inflammatory signaling, oxidative stress and complement activation.
Increasing evidence has highlighted the role of microglia and inflammation in disease progression, including in age-related macular degeneration (AMD). To study the role of inflammation in RPE atrophy during AMD progression in vitro, we developed a human iPSC-derived RPE-microglia co-culture model.
After identifying culture conditions that enabled prolonged co-culture of these two cell types, AMD-relevant stressors were introduced into the system. A2E induced little microglial activation in monoculture but triggered microglial proliferation and elevated secretion of inflammatory cytokines, including IL-1β, TNF-α, IL-6 and IL-10, when microglia were co-cultured with RPE cells.
In the presence of microglia, RPE cells also showed greater sensitivity to A2E- and blue light-induced stress, highlighting the importance of cell-cell interactions in shaping inflammatory responses.
We are extending this work through the development of a tri-culture system comprising iPSC-derived RPE cells, photoreceptors and microglia. This will enable investigation of how chronic stress affects communication between these three cell types and how these interactions contribute to inflammation and retinal degeneration.
Developing physiologically relevant human co-culture systems
Translational research continues to move toward increasingly sophisticated human-relevant systems that incorporate greater cellular diversity and more closely reflect tissue architecture. However, this drive toward complexity needs to be balanced with reproducibility, scale and accurate representation of diversity in patient populations.
The integration of microglia into three-dimensional brain and retinal organoids provides opportunities to investigate neuroimmune interactions within tissue-like environments. In parallel, advances in microphysiological systems and organ-on-chip technologies enable multiple human cell types to be combined within precisely controlled environments.
Combined with advances in single-cell transcriptomics, metabolomics, spatial biology, high-content imaging and computational analysis, complex human iPSC-derived cell models are expected to further improve the ability to study cellular interactions at high resolution and accelerate the development of more predictive disease models.
As broader industry-wide adoption of NAMs continues, human iPSC-derived co-culture systems provide scalable and reproducible insights into disease, improving translational relevance across neuroscience and ophthalmology.
Download the whitepaper
Explore how building complexity into cell-based in vitro models can reveal disease-relevant phenotypes while maintaining reproducibility, with examples from ALS and dry AMD.
