“Miniguts” from plucked human hair meet Crohn’s disease

Abstract

Human pluripotent stem cells represent a powerful tool to study human embryonic development and disease but also open up novel strategies for cell replacement therapies. Their capacity to give rise to every cell type of the human body, meanwhile, enables researchers to generate high yields of mesodermal, ectodermal, but also endodermal-derived tissues such as hepatic, pancreatic, or intestinal cells. Another progress in the field came with the advent of 3-dimensional culture conditions, so-called organoids, which facilitate maturation of stem cells and in turn more faithfully recapitulate human tissue architecture. While several studies reported the derivation of organoid cultures from adult intestinal tissue, the derivation of intestinal organoids derived from plucked human hair of Crohn’s disease patients has not been reported. The current research project reports such successful generation and characterization of induced pluripotent stem cells (iPSCs) derived from hair sheet keratinocyte cultures of a patient with Crohn's disease. Stepwise differentiation along the intestinal lineage showed no differences in intermediate stages such as definitive endoderm formation. We also directed the patterned primitive gut tube toward intestinal organoids resembling the cellular architecture of human “miniguts”. As expected from current pathophysiological knowledge on Crohn's disease, there were no obvious morphological differences in the “miniguts” derived from healthy control and diseased patient-induced pluripotent stem cells. Taken together, our platform will enable for detailed and complementary phenotyping of the pathophysiology of Crohn’s disease in a novel disease-in-a-dish format.

Zusammenfassung

Humane pluripotente Stammzellen stellen ein wertvolles Instrument zur Erforschung menschlicher Embryonalentwicklung und zahlreicher Erkrankungen dar; darüber hinaus ergeben sich perspektivisch neuartige Strategien für Zellersatztherapien. Die Fähigkeit von Stammzellen, sich in jede Zellart des menschlichen Körpers zu entwickeln, ermöglicht es mittlerweile, aus allen drei Keimblättern (Mesoderm, Ektoderm und Endoderm) hochdifferenzierte Gewebestrukturen wie etwa Leber-, Pankreas- oder Darmzellen in größeren Mengen zu generieren. Ein weiterer Fortschritt trat mit der Einführung von 3-dimensionalen Kulturbedingungen ein, die eine komplexe Ausreifung von Stammzellen erheblich erleichtern. Dadurch können sog. Organoide entstehen, welche die menschliche Gewebsarchitektur sehr genau widerspiegeln. Während mehrere Studien die Generierung von Organoiden aus adultem Darmgewebe beschreiben, wurde bisher die Erzeugung von Organoiden aus Haarwurzel-Keratinozyten von Patienten mit Morbus Crohn noch nicht beschrieben. In der hier vorliegenden Arbeit berichten wir erstmalig über die erfolgreiche Herstellung und Charakterisierung von induzierten pluripotenten Stammzellen (iPSCs) abstammend von einem Patienten mit M. Crohn. Die schrittweise Differenzierung dieser iPS-Zellen hin zu Darmgewebe zeigte keine Unterschiede bei der Erzeugung von Endoderm. Eine weitere Differenzierung des komplexen Gewebes bis hin zur Entstehung von Strukturen, die Darmorganoiden („Miniguts“) in höchstem Maße ähnlich sind, war möglich. Wie aufgrund des derzeitigen Wissenstands über M. Crohn zu erwarten ergaben sich keine erkennbaren morphologischen Veränderungen im Vergleich zu Organoiden einer gesunden Kontrollperson. Mit der vorliegenden Arbeit legen wir die Grundlage für ein neues In-vitro-Modellsystem, das weitergehende Untersuchungen, insbesondere der Interaktion von Zellen des Immunsystems, Antigenen, Inflammationsmediatoren und den komplexen Darmorganoiden ermöglichen wird.

Schlüsselwörter

chronisch entzündliche Darmerkrankung - Morbus Crohn - induziert pluripotente Stammzellen - intestinale Organoide

Key words

Inflammatory Bowel Disease - Crohn's Disease - induced pluripotent stem cells - intestinal organoids

^# Equal contribution.

^$ These authors jointly supervised this work.

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