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 3

Bile acid transport protein expression is altered in liver from mtm zebrafish.

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Bile acid transport protein expression is altered in liver from mtm zebr...
(A) Immunofluorescence staining of whole-mount zebrafish at 7 dpf using anti-Bsep. In mtm larvae, Bsep staining was essentially undetectable. Scale bars: 20 μm. (B) Immunofluorescence staining of paraffin sections of whole zebrafish examined at 2 time points: 5 dpf and 7 dpf. Scale bars: 20 μm. (C) Coimmunofluorescence staining of sectioned 7 dpf zebrafish for GFP-CAAX, a membrane marker, as well as Mdr1. Costaining revealed reduced canaliculi numbers and altered morphology in mtm embryos. As seen in B, Mdr1 staining was also absent. Red arrows point to examples of canaliculi that were positive for both GFP-CAAX and Mdr1. Scale bars: 20 μm. (D) Quantification of Bsep+ canaliculi in WT versus mtm larvae in A at 7 dpf. Bsep+ puncta were reduced in mtm larvae (WT mean = 208.8 ± 110.3, mtm mean = 7.18 ± 4.71, **P = 0.0014, by unpaired, 2-tailed t test). (E and F) Quantification of Mdr1+ canaliculi in sectioned WT and mtm larvae from B. At 5 dpf (E), the WT and mtm images had similar numbers of puncta (WT mean = 10.33 ± 2.517, mtm mean = 8.667 ± 3.512, *P = 0.5406, by unpaired, 2-tailed t test). At 7 dpf, the number of Mdr1+ puncta in mtm larvae was greatly reduced (WT mean = 51 ± 15.62, mtm mean = 10 ± 2.65, **P = 0.011, by unpaired, 2-tailed t test). (G) Quantification of GFP+ canaliculi in C. There were significantly more intact canaliculi in WT livers than in mtm livers (WT mean = 12.0 ± 3.61, mtm mean = 1.0 ± 0, **P = 0.0062, by unpaired, 2-tailed t test).