Lysophosphatidic Acid Increases Maturation of Brush Borders and SGLT1 Activity in MYO5B-deficient Mice, a Model of Microvillus Inclusion Disease

Abstract

BACKGROUND & AIM: Myosin VB (MYO5B) is an essential trafficking protein for membrane recycling in gastrointestinal epithelial cells. The inactivating mutations of MYO5B cause the congenital diarrheal disease, microvillus inclusion disease (MVID). 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 effect of LPA on MVID symptoms is unclear. We investigated whether LPA administration can reduce the epithelial deficits in MYO5B-knockout mice. METHODS: Studies were conducted with tamoxifen-induced, intestine-specific knockout of MYO5B (VilCre ERT2 ;Myo5b flox/flox) and littermate controls. Mice were given LPA, an LPAR2 agonist (GRI977143), or vehicle for 4 days after a single injection of tamoxifen. Apical SGLT1 and CFTR activities were measured in Üssing chambers. Intestinal tissues were collected, and localization of membrane transporters was evaluated by immunofluorescence analysis in tissue sections and enteroids. RNA sequencing and enrichment analysis were performed with isolated jejunal epithelial cells. RESULTS: Daily administration of LPA reduced villus blunting, frequency of multivesicular bodies, and levels of cathepsins in intestinal tissues of MYO5B-knockout mice compared with vehicle administration. LPA partially restored the brush border height and the localization of SGLT1 and NHE3 in small intestine of MYO5B-knockout mice and enteroids. The SGLT1-dependent short-circuit current was increased and abnormal CFTR activities were decreased in jejunum from MYO5B-knockout mice given LPA compared with vehicle. CONCLUSIONS: LPA may regulate a MYO5B-independent trafficking mechanism and brush border maturation, and therefore be developed for treatment of MVID.

Recruitment of Polarity Complexes and Tight Junction Proteins to the Site of Apical Bulk Endocytosis

“…In support of this concept, adult inducible VillinCre ERT2 ;Myo5b floxed/floxed mice show many intestinal alterations after loss of Myo5b including diarrhea, however, they do not have an intestinal barrier defect based on tissue resistance and dextran permeability measurements. 11,12 We observed that in Myo5b KO mice, polarity complex proteins Crumbs3, Patj, Pals1, and aPKC all were present in intracellular inclusions. These findings appear to mirror observations with enteric pathogens.…”

“…[3][4][5][6][7][8][9] MVID is characterized by severe congenital diarrhea, resulting from an inability to properly absorb water coupled with active chloride secretion, which drives further dehydration. [10][11][12][13][14] The presence of intracellular inclusions containing microvilli is a hallmark of MVID. 6,15 We previously showed that inclusions in Myo5b-deficient enterocytes form through apical bulk endocytosis, which requires Pacsin2 and Dynamin2.…”

Advanced Microscopy for Liver and Gut Ultrastructural Pathology in Patients with MVID and PFIC Caused by MYO5B Mutations

“…MVID enterocytes regularly display conspicuous lysosomes and other large compartments that partially contain still identifiable subcellular compounds and apical marker proteins, including apical sodium transporters sodium/glucose cotransporter 1 (SGLT1) and NHE3, for example [ 68 ]. According to morphological criteria those latter compartments have been tentatively classified as autophagosomes and autophagolysosomes [ 28 , 31 ], but without formal proof by autophagy markers.…”

Exploiting Alternative Brush Border Trafficking Routes to Treat Microvillous Inclusion Disease

“…9,12,13 This process can also be activated by physiologic stimuli, including Na þ -glucose cotransport. 12,14 In this issue of Gastroenterology, Kaji et al 15 asked if LPA-induced NHE3 exocytosis required MYO5B or, alternatively, be EDITORIALS exploited to stimulate delivery of apical transporters to the brush border in MVID.…”

“…Defective MYO5B function in microvillous inclusion disease (MVID) leads to loss of brush border transporters as well as essential microvillous structural components (center). Kaji et al15 show lysophosphatidic acid (LPA) can activate a MYO5B-independent exocytic pathway that partially restores brush border transporter expression and microvillous structure (right). engender optimism.…”