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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 anti-diarrheal drug, Crofelemer, dose-dependently inhibited agonist-mediated fluid secretion. MVID enteroids exhibited altered differentiation and maturation versus healthy enteroids. Gamma-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- and glucocorticoid-induced protein kinase 2 (SGK2), and NHE regulatory factor 3 (NHERF3). These results demonstrate the utility of patient-derived enteroids for developing therapeutic approaches to MVID.

ORGANISM(S): Homo sapiens

PROVIDER: GSE240928 | GEO | 2023/08/16

REPOSITORIES: GEO

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Lysophosphatidic acid treatment improves brush border maturation and apical SGLT1 activity in Myo5b deficient mice, a model of MVID

Project description:Myosin Vb (Myo5b) is an essential trafficking protein for membrane recycling in gastrointestinal (GI) epithelial cells and its inactivating mutation causes the congenital diarrheal disease, microvillus inclusion disease (MVID). We previously reported that Myo5b deficiency in mice causes mislocalization of SGLT1 and NHE3, but retained apical function of CFTR, resulting in malabsorption and secretory diarrhea. Activation of lysophosphatidic acid (LPA) receptors can improve diarrhea, but the effects of LPA on MVID symptoms is unclear. We therefore tested whether LPA treatment can ameliorate the epithelial deficits in Myo5b knockout (KO) mice. Methods: Adult tamoxifen-induced, intestine-specific, Myo5b KO (VillinCreERT2;Myo5bflox/flox) and littermate control mice were treated with LPA, an LPAR2 agonist (GRI977143), or vehicle for 4 days after a single tamoxifen injection. Apical SGLT1 and CFTR activities were measured in Üssing chambers. The localization of membrane transporters was evaluated by immunostaining in mouse tissues and enteroids. RNA sequencing and enrichment analysis were performed with isolated jejunal epithelial cells. Results: Daily treatment with LPA decreased the frequency of multivesicular bodies and the expression of cathepsins, but did not affect the formation of microvillus inclusions in Myo5b KO mice. LPA partially restored the brush border height and the localization of SGLT1 and NHE3 in Myo5b KO small intestine and enteroids. SGLT1-dependent short-circuit current (Isc) was increased and abnormal CFTR activities were suppressed in LPA-treated jejunum compared to that of vehicle-treated Myo5b KO mice. Conclusions: Cell autonomous LPA signaling may modulate a Myo5b-independent trafficking mechanism and the brush border maturation, demonstrating a therapeutic potential for LPA in the treatment of MVID.

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