Transplantation of human intestinal organoids into the mouse mesentery: A more physiologic and anatomic engraftment site

Cortez, Alexander R.; Poling, Holly M.; Brown, Nicole; Singh, Akaljot; Mahé, Maxime M.; Helmrath, Michael A.

doi:10.1016/j.surg.2018.04.048

Cited by 47 publications

(44 citation statements)

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“…Similarly, microinjection of E. coli into the HIO lumen has permitted the study of epithelial response to early gut colonization (Hill et al, 2017a, 2017b). However, vascularization of HIOs has been restricted to in vivo models, whereby HIOs are transplanted into highly vascularized regions of immunocompromised mice (Cortez et al, 2018; Watson et al, 2014). In these environments, HIOs undergo extensive vascularization by the murine host tissue, and increase in complexity to resemble mature intestinal tissue (Finkbeiner et al, 2015b; Watson et al, 2014).…”

Section: Introductionmentioning

confidence: 99%

Differentiation of human intestinal organoids with endogenous vascular endothelial cells

Holloway

Czerwinkski

et al. 2020

Preprint

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22Human pluripotent stem cell (hPSC)-derived intestinal organoids (HIOs) generated using directed 23 differentiation lack some cellular populations found in the native organ, including vasculature. Using 24 single cell RNA sequencing (scRNAseq), we have identified a transient population of endothelial cells 25 (ECs) present early in HIO differentiation that are lost over time in culture. Here, we have developed a 26 method to enhance co-differentiation and maintenance of ECs within HIOs (vHIOs). Given that ECs are 27 known to possess organ specific gene expression, morphology and function, we used bulk RNAseq 28 and scRNAseq to interrogate the developing human intestine, lung, and kidney in order to identify 29 organ-enriched EC-gene signatures in these organ systems. By comparing organ-specific gene 30 signatures along with markers validated by fluorescent in situ hybridization to HIO ECs, we find that 31 HIO ECs grown in vitro share the highest similarity with native intestinal ECs relative to kidney and 32 lung. Together, these data show that HIOs can co-differentiate a native EC population that are properly 33 patterned with an intestine-specific EC transcriptional signature in vitro. 34 35 36 37 38 39 40 41 42 43 44 65highly vascularized regions of immunocompromised mice (Cortez et al., 2018; Watson et al., 2014). In 66 these environments, HIOs undergo extensive vascularization by the murine host tissue, and increase in 67 complexity to resemble mature intestinal tissue (Finkbeiner et al., 2015b; Watson et al., 2014). 68However, co-culture approaches or co-differentiating HIOs with a native vasculature prior to in vivo 69 engraftment has not yet been achieved. 71Here, we performed single cell RNA sequencing (scRNAseq) at various timepoints across HIO 72 differentiation in vitro and observed a transient population of endothelial-like cells (ECs) present within 73 HIOs early during differentiation; however, these cells are not maintained during prolonged culture 74 under standard growth conditions. This suggested that early during HIO differentiation, cells within the 75 culture are capable of giving rise to EC-like cells. Based on these observations, we hypothesized that a 76 modified directed differentiation approach would allow the induction and maintenance of a more robust 77 EC population within HIOs (termed vHIO). Our findings demonstrate that modified culture conditions 78 allow a ~13-fold increase in the induction of EC-like cells within HIOs without impacting the other HIO 79 cell populations present (i.e. epithelium, mesenchyme), and support the survival of this population of 80 ECs within HIOs in culture for months. 82Since organ-specific morphology in vascular beds has long been appreciated (Aird, 2007), and 83 organ-specific transcriptional signatures and functions have been described in mouse (Ding et al., 84 2011; Kalucka et al., 2020; Lee et al., 2014; Nolan et al., 2013) and human tissues (Chi et al., 2003; 85 Marcu et al., 2018), we further sought to determine if HIO ECs were properly...

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Section: Introductionmentioning

confidence: 99%

Differentiation of human intestinal organoids with endogenous vascular endothelial cells

Holloway

Czerwinkski

et al. 2020

Preprint

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“…Importantly, the mesentery shares a blood supply with the intestine, and is thought to contribute to intestinal function, including peristalsis, immune function, and tissue repair [7]. While our previous studies have demonstrated that the mesentery is a viable alternative site for HIO transplantation, a direct comparison between the two transplantation sites has not been performed [8]. Here, we aim to characterize potential similarities and differences between tHIOs engrafted into both sites.…”

Section: Introductionmentioning

confidence: 99%

Evaluation of transplantation sites for human intestinal organoids

et al. 2020

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Our group has developed two transplantation models for the engraftment of Human Intestinal Organoids (HIOs): the renal subcapsular space (RSS) and the mesentery each with specific benefits for study. While engraftment at both sites generates laminated intestinal structures, a direct comparison between models has not yet been performed. Embryonic stem cells were differentiated into HIOs, as previously described. HIOs from the same batch were transplanted on the same day into either the RSS or mesentery. 10 weeks were allowed for engraftment and differentiation, at which time they were harvested and assessed. Metrics for comparison included: mortality, engraftment rate, gross size, number and grade of lumens, and expression of markers specific to epithelial differentiation, mesenchymal differentiation, and carbohydrate metabolism. Mortality was significantly increased when undergoing mesentery transplantation, however engraftment was significantly higher. Graft sizes were similar between groups. Morphometric parameters were similar between groups, however m-tHIOs presented with significantly fewer lumens than k-tHIO. Transcript and protein level expression of markers specific to epithelial differentiation, mesenchymal differentiation, and carbohydrate metabolism were similar between groups. Transplantation into both sites yields viable tissue of similar quality based on our assessments with enhanced engraftment and a dominant lumen for uniform study benefiting the mesenteric site and survival benefiting RSS.

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“…For obvious reasons, they are also limited in their capacity to reach endpoint differentiation or full scale by virtue of their in vitro unperfused state. In large part, these models are also lacking a neural and immune system, although some approaches are including such cellular components either as additional elements during differentiation (Schlieve et al 2017) and/or transplantation for subsequent vascularization (Cortez et al 2018;Daviaud et al 2018).…”

Section: Building a Kidney Organoid From Pluripotent Stem Cellsmentioning

confidence: 99%

Kidney organoids: accurate models or fortunate accidents

2019

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There are now many reports of human kidney organoids generated via the directed differentiation of human pluripotent stem cells (PSCs) based on an existing understanding of mammalian kidney organogenesis. Such kidney organoids potentially represent tractable tools for the study of normal human development and disease with improvements in scale, structure, and functional maturation potentially providing future options for renal regeneration. The utility of such organotypic models, however, will ultimately be determined by their developmental accuracy. While initially inferred from mouse models, recent transcriptional analyses of human fetal kidney have provided greater insight into nephrogenesis. In this review, we discuss how well human kidney organoids model the human fetal kidney and how the remaining differences challenge their utility.

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Transplantation of human intestinal organoids into the mouse mesentery: A more physiologic and anatomic engraftment site

Abstract: The mouse mesentery is a viable location for the transplantation of human intestinal organoids, yielding grafts of reproducible size and quality. This improved model serves to advance functional and translational studies of human intestinal organoids.

Cited by 47 publications

References 12 publications

Differentiation of human intestinal organoids with endogenous vascular endothelial cells

Differentiation of human intestinal organoids with endogenous vascular endothelial cells

Evaluation of transplantation sites for human intestinal organoids

Kidney organoids: accurate models or fortunate accidents

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