Patient-derived enteroids provide a platform for the development of therapeutic approaches in microvillus inclusion disease
Meri Kalashyan, Krishnan Raghunathan, Haley Oller, Marie-Theres Bayer, Lissette Jimenez, Joseph T. Roland, Elena Kolobova, Susan J. Hagen, Jeffrey D. Goldsmith, Mitchell D. Shub, James R. Goldenring, Izumi Kaji, Jay R. Thiagarajah
Meri Kalashyan, Krishnan Raghunathan, Haley Oller, Marie-Theres Bayer, Lissette Jimenez, Joseph T. Roland, Elena Kolobova, Susan J. Hagen, Jeffrey D. Goldsmith, Mitchell D. Shub, James R. Goldenring, Izumi Kaji, Jay R. Thiagarajah
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Research Article Gastroenterology

Patient-derived enteroids provide a platform for the development of therapeutic approaches in microvillus inclusion disease

Abstract

Microvillus inclusion disease (MVID), caused by loss-of-function mutations in the motor protein myosin Vb (MYO5B), is a severe infantile disease characterized by diarrhea, malabsorption, and acid/base instability, requiring intensive parenteral support for nutritional and fluid management. Human patient–derived enteroids represent a model for investigation of monogenic epithelial disorders but are a rare resource from MVID patients. We developed human enteroids with different loss-of function MYO5B variants and showed that they recapitulated the structural changes found in native MVID enterocytes. Multiplex immunofluorescence imaging of patient duodenal tissues revealed patient-specific changes in localization of brush border transporters. Functional analysis of electrolyte transport revealed profound loss of Na+/H+ exchange (NHE) activity in MVID patient enteroids with near-normal chloride secretion. The chloride channel–blocking antidiarrheal drug crofelemer dose-dependently inhibited agonist-mediated fluid secretion. MVID enteroids exhibited altered differentiation and maturation versus healthy enteroids. γ-Secretase inhibition with DAPT recovered apical brush border structure and functional Na+/H+ exchange activity in MVID enteroids. Transcriptomic analysis revealed potential pathways involved in the rescue of MVID cells including serum/glucocorticoid-regulated kinase 2 (SGK2) and NHE regulatory factor 3 (NHERF3). These results demonstrate the utility of patient-derived enteroids for developing therapeutic approaches to MVID.

Authors

Meri Kalashyan, Krishnan Raghunathan, Haley Oller, Marie-Theres Bayer, Lissette Jimenez, Joseph T. Roland, Elena Kolobova, Susan J. Hagen, Jeffrey D. Goldsmith, Mitchell D. Shub, James R. Goldenring, Izumi Kaji, Jay R. Thiagarajah

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Figure 7

Genome-wide transcriptomic analysis reveals potential targets for rescue of MVID enteroids.

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Genome-wide transcriptomic analysis reveals potential targets for rescue...
(A) Volcano plot showing log2 fold change (FC) and false discovery rate (FDR) showing genes with significantly up- and downregulated expression (red) in healthy enteroids (n = 3) following DAPT treatment (10 μM). (B) Volcano plot showing genes with significantly up- and downregulated expression (red) between healthy enteroids and MVID enteroids (MYO5B KO) (n = 3). (C) Volcano plot showing genes with significantly up- and downregulated expression (red) in MVID enteroids (MYO5B KO) following DAPT treatment (10 μM). (D) Plot of genes with significantly altered expression (green dots) between MVID and healthy against MVID + DAPT. Red dots indicate genes changing in opposite directions following DAPT treatment (filtered genes). (E) Dot plot of filtered genes by change in expression and base mean expression, with dot size indicating fold change and color indicating FDR. Highlighted genes are based on previous functional data indicating a plausible biological role. (F) Pathway analysis showing most significant Gene Ontology (GO) terms, Human Protein Atlas (HPA) terms, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways.