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 Article has an altmetric score of 6

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 1

mtm zebrafish show evidence of hepatic steatosis and cholestasis.

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mtm zebrafish show evidence of hepatic steatosis and cholestasis. (A) O...
(A) Oil Red O staining shows hepatic steatosis in 5 dpf zebrafish larvae. Dashed lines outline the liver, where there is evidence of lipid accumulation in mtm zebrafish. Black arrow denotes the swim bladder, which is not properly inflated in mtm mutants. Scale bars: 100 μm. (B) Quantification of Oil Red O in liver (n = 10 fish for each condition). No WT larvae had severe steatosis compared with 66% of mtm larvae. (C) BODIPY feeding assay showed impaired bile flux in mtm larvae. WT and mtm zebrafish were fed AP-100 fish food mixed with BODIPY C12. Representative images are shown of fish with positive and negative gallbladder fluorescence. Blue arrow denotes the gallbladder. Original magnification, ×25. (D) Quantification of gallbladder fluorescence combining 2 independent biological replicates (n = 30 fish per replicate). Ninety-three percent of WT larvae exhibited gallbladder fluorescence after BODIPY exposure, whereas only 28% of mtm larvae showed gallbladder fluorescence (****P < 0.0001, by Fisher’s exact test). (E) Comparisons of individual bile acids at 5 dpf. TCA, TCDCA, and TDCA are 3 conjugated, hydrophobic bile acids that can cause toxicity when their levels are elevated. TCA and TDCA levels appeared unchanged, whereas TCDCA levels were elevated in mtm larvae (*P = 0.039, by unpaired, 2-tailed t test).