Loss of Mtm1 causes cholestatic liver disease in a model of X-linked myotubular myopathy
Sophie Karolczak, … , Chunyue Yin, James J. Dowling
Sophie Karolczak, … , Chunyue Yin, James J. Dowling
Published July 25, 2023
Citation Information: J Clin Invest. 2023;133(18):e166275. https://doi.org/10.1172/JCI166275.
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Research Article Hepatology Muscle biology

Loss of Mtm1 causes cholestatic liver disease in a model of X-linked myotubular myopathy

Abstract

X-linked myotubular myopathy (XLMTM) is a fatal congenital disorder caused by mutations in the MTM1 gene. Currently, there are no approved treatments, although AAV8-mediated gene transfer therapy has shown promise in animal models and preliminarily in patients. However, 4 patients with XLMTM treated with gene therapy have died from progressive liver failure, and hepatobiliary disease has now been recognized more broadly in association with XLMTM. In an attempt to understand whether loss of MTM1 itself is associated with liver pathology, we have characterized what we believe to be a novel liver phenotype in a zebrafish model of this disease. Specifically, we found that loss-of-function mutations in mtm1 led to severe liver abnormalities including impaired bile flux, structural abnormalities of the bile canaliculus, and improper endosome-mediated trafficking of canalicular transporters. Using a reporter-tagged Mtm1 zebrafish line, we established localization of Mtm1 in the liver in association with Rab11, a marker of recycling endosomes, and canalicular transport proteins and demonstrated that hepatocyte-specific reexpression of Mtm1 could rescue the cholestatic phenotype. Last, we completed a targeted chemical screen and found that Dynasore, a dynamin-2 inhibitor, was able to partially restore bile flow and transporter localization to the canalicular membrane. In summary, we demonstrate, for the first time to our knowledge, liver abnormalities that were directly caused by MTM1 mutation in a preclinical model, thus establishing the critical framework for better understanding and comprehensive treatment of the human disease.

Authors

Sophie Karolczak, Ashish R. Deshwar, Evangelina Aristegui, Binita M. Kamath, Michael W. Lawlor, Gaia Andreoletti, Jonathan Volpatti, Jillian L. Ellis, Chunyue Yin, James J. Dowling

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Figure 7

Liver-specific Mtm1 expression rescues the cholestatic phenotype of mtm zebrafish.

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Liver-specific Mtm1 expression rescues the cholestatic phenotype of mtm ...
The fabp:mtm1-GFP transgene was introduced into the mtm-mutant zebrafish line. The resulting fish were analyzed for morphological and functional changes associated with cholestasis. (A) Crossing scheme for introducing the fabp:mtm1-gfp transgene into the mtm zebrafish line. Transgenic fish were outcrossed twice to mtm1+/Δ8 fish, resulting in clutches of larvae for experiments that contained WT and mtm fish with and without the transgene. (B) A BODIPY assay was used to measure bile flux (WT – GFP = 88%, WT + GFP = 96%, mtm – GFP = 30%, mtm + GFP = 65%). In the mtm transgene–positive group, there were more mtm mutants with normal bile flux when compared with the mtm transgene–negative group, as measured by positive gall bladder fluorescence (P = 0.0532, 1-sided Fisher’s exact test). (C) Visualization of liver-specific Mtm1 expression from the fabp:mtm1-GFP transgene. In WT fish, Mtm1 localized to the plasma membrane and to subapical structures that were Mdr1+ and Rab11+ by immunostaining (top two rows). In mtm zebrafish, hepatocyte-expressed Mtm1 restored bile canalicular architecture and bile transporter expression to the bile canaliculi. Coimmunostaining of whole-mount embryos revealed the reexpression of Mdr1 puncta in 7 dpf mtm larvae. Scale bars: 10 μm.