Functional, metabolic and transcriptional maturation of human pancreatic islets derived from stem cells

Source

PDF: raw/sources/balboa_2022_functional_metabolic_and_transcriptional.pdf
Article: Nature Biotechnology
Labs: Timo Otonkoski + Diego Balboa (University of Helsinki; also CRG Barcelona)
Status: deep ingested 2026-04-09

Starting material: hPSCs (3 iPSC lines for robustness testing)
Protocol innovations over baseline SC-islet methods:
Adherent differentiation until pancreatic progenitor stage (S4)
Optimized S4: nicotinamide + EGF + Activin A + ROCK inhibitor
Microwell aggregation step → ~80% PDX1+/NKX6-1+ pancreatic progenitor clusters
Improved final maturation stage (S7) in suspension culture
S7 omits ALK5 inhibitor and adds aurora kinase inhibitor ZM447439 (anti-proliferative) + T3 + N-acetyl cysteine (NAC)
Timepoints: S7w0 (start of final maturation) → S7w6 (6 weeks of maturation)
Comparison: primary adult human islets
Readouts: morphometry, GSIS (glucose-stimulated insulin secretion), metabolomics, electrophysiology, exocytosis, single-cell transcriptomics (in vitro + 6 months post-engraftment in mice)

Biphasic GSIS develops during in vitro S7 maturation — by S7w2 onwards, SC-islets show biphasic responses similar to primary islets.
4-fold increase in insulin content during first 3 weeks of S7.
SC-β cells acquire dense-core insulin granules resembling those of primary β cells (ultrastructural).
Cytoarchitecture reorganizes: from periphery-localized INS+ cells at S7w0 → polarized GCG+/INS+ clusters at S7w3 → intermingled mature cytoarchitecture at S7w6 (mirrors fetal islet development).
Ki-67+ proliferation reduces by 80% during S7 — maturation linked to cell cycle exit.
ZM / NAC / T3 all required for the maturation effect.
SC-EC (enterochromaffin-like) byproduct cells reduce from ~13% → ~6.5% during S7 — another ZM-dependent outcome.
Electrophysiology, signaling, and exocytosis approach primary islet levels.
Glycolytic and mitochondrial glucose metabolism still differ from primary islets — maturation is not complete.
scRNA-seq over 6 months of in vivo engraftment shows continued maturation trajectory converging toward primary islet transcriptome.

Most comprehensive maturation benchmark of SC-islets vs primary in the corpus.
Establishes the "how mature are the cells really" framework for stem-cell-derived β cells.
Identifies specific molecular maturation axes (cytoarchitecture, granule morphology, GSIS biphasicity, proliferation) and their dependencies.
Complements Hogrebe 2021 (which is primarily about production protocol).

Metabolic differences (glycolysis, mitochondrial metabolism) persist even after 6 weeks of S7.
Maturation is assessed in vitro; full equivalence with adult β cells may require in vivo maturation.
Primary islet comparison used donors at the lower end of functional responses — caveat for generalization.

Together with Hogrebe 2021, establishes the current best-practice pair for pancreatic β cell generation (production + maturation).
Represents the "functional fidelity" axis for pancreatic organoids — parallel to what Bhaduri 2020 did for brain organoids.

Why do glycolytic and mitochondrial metabolism remain different even after functional maturation?
Can the ZM / NAC / T3 cocktail be transferred to other endocrine differentiation systems?
What is the cytoarchitectural reorganization mechanism (mechanical? paracrine?)?
How do Hogrebe 2021's planar SC-β cells compare on these maturation benchmarks?

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