Supported by a Department of Veterans Affairs Senior Research Career Scientist Award (to J. K. Y.), Cancer Center Support Grant P30CA047904, and National Institutes of Health Grant R21DK073177 (to N.B.).
Steatohepatitis/Metabolic Liver Disease
Lack of de novo phosphatidylinositol synthesis leads to endoplasmic reticulum stress and hepatic steatosis in cdipt-deficient zebrafish
Article first published online: 2 MAY 2011
Copyright © 2011 American Association for the Study of Liver Diseases
Volume 54, Issue 2, pages 452–462, August 2011
How to Cite
Thakur, P. C., Stuckenholz, C., Rivera, M. R., Davison, J. M., Yao, J. K., Amsterdam, A., Sadler, K. C. and Bahary, N. (2011), Lack of de novo phosphatidylinositol synthesis leads to endoplasmic reticulum stress and hepatic steatosis in cdipt-deficient zebrafish. Hepatology, 54: 452–462. doi: 10.1002/hep.24349
- Issue published online: 25 JUL 2011
- Article first published online: 2 MAY 2011
- Accepted manuscript online: 12 APR 2011 08:18AM EST
- Manuscript Accepted: 28 MAR 2011
- Manuscript Received: 19 DEC 2010
Additional Supporting Information may be found in the online version of this article.
|HEP_24349_sm_suppinfoFig1.tif||1156K||Supporting Information Figure S1: The hi559 pancreatic tissue does not exhibit any overt abnormal architecture. (A) Expression of try (exocrine pancreas, arrow, left panel) and ins (endocrine pancreas, arrow, right panel) are similar in wild-type (left) and hi559 larvae (right). (B) H&E staining of sagittal sections (4 μ) through pancreatic region of 5-dpf wild-type and hi559 larvae. Endocrine pancreatic islet is indicated by arrow. The hi559 pancreas does not exhibit any overt defects in its architecture. ep, endocrine pancreas|
|HEP_24349_sm_suppinfoFig2.tif||1585K||Supporting Information Figure S2: cdipt morphants phenocopy hi559 phenotype. Embryos injected with p53 morpholino alone appears normal at 6 dpf (panel A), whereas embryos injected with p53 morpholino and two different cdipt splice-blocking morpholinos, cdipt-Mo1 (panel B) and cdipt-Mo2 (panel C) exhibit hepatomegaly, globular-appearing liver and smaller intestine at 6-dpf, resembling hi559 hepatic phenotype. The cdipt morphants survive through 7-dpf, but the liver and intestine exhibit severe defects (panel D). Injection of cdipt mRNA results in significant reduction of larvae showing hi559 phenotype (panel E)|
|HEP_24349_sm_suppinfoFig3.tif||1786K||Supporting Information Figure S3: Comparison of cdipt expression in wild-type and hi559 embryos during embryonic development. Developmental expression of cdipt at 1-dpf (A), 2-dpf (B), 3-dpf (C) and 4-dpf (D). Cdipt is expressed in the developing liver (arrow), intestine, brain, eye and branchial arches in wild-type embryos, whereas its expression is negligible in hi559 embryos. Wild-type on top, mutant below in each panel|
|HEP_24349_sm_suppinfoFig4.tif||1354K||Supporting Information Figure S4: TLC showing phospholipid profile of the wild-type and hi559 embryos. The PtdIns level (PtdIns band indicated by arrow) of the deyolked 5-dpf wild-type and hi559 larvae is unaltered. Sufficient amount of PtdIns is also present in the yolk (1-cell stage) suggesting maternal deposition of PtdIns in the yolk. The overall phospholipid profile does not differ qualitatively in wild-type and hi559 larvae. CH: Cholesterol, TG: Triglycerides, CB: Cerebrosides, PE: Phosphatidylethenolamine, PA; Phosphatidic Acid, PC: Phosphatidylcholine, PI: Phosphatidylinositol, PS: Phosphatidylserine|
|HEP_24349_sm_suppinfoFig5.tif||846K||Supporting Information Figure S5: Chemical inhibition of PIS replicates hi559 phenotype. More than 80% of the wild-type larvae treated with δ-HCH at 10 μM during 3 to 5-dpf exhibit hepatomegaly and a darkish liver (yellow outline) at 5-dpf similar to hi559 (A), and a fatty liver as shown by ORO staining (B). Data are representative of 3 biological clutches. Wild-type on top, mutant below in each panel|
|HEP_24349_sm_suppinfoFig6.tif||2145K||Supporting Information Figure S6: Distortion of sinusoid architecture in the hi559 liver. (A) Whole-mount alkaline phosphatase staining reveals abnormality of intrahepatic vasculature in severely steatotic hi559 liver, whereas rest of the larval vasculature appears normal. (B) Sagittal sections of larvae stained with alkaline phosphatase and eosin showing compressed sinusoids in the mutant liver (arrow). Liver is outlined by dotted line. ihv, intrahepatic vessels; isv, intersomitic vessels; siv, subintestinal vein. Scale bars, 20 μM|
|HEP_24349_sm_suppinfoFig7.tif||2107K||Supporting Information Figure S7: Pathways showing transcriptional upregulation of ERSR components. ERSR pathway from Ingenuity overlaid with microarray expression values. Color intensity is proportional to fold-change (red, upregulated; blue downregulated; white absent on array)|
|HEP_24349_sm_suppinfoFig8.tif||2576K||Supporting Information Figure S8: Diagrammatic representation of the proposed model of cellular mechanisms contributing to ER stress due to disruption of PtdIns synthesis. PtdIns and PI signaling are critical for proper functions of ER, secretory pathways, and ER-associated degradation (ERAD). Thus, we hypothesize that limited de novo PtdIns may affect one or combinations of these cellular processes causing an unresolved chronic ER stress|
|HEP_24349_sm_suppinfoFig9.tif||2019K||Supporting Information Figure S9: Zebrafish Cdipt is an orthologue of mammalian CDIPT. Sequence alignment shows striking similarity of the zebrafish Cdipt with its mammalian counterpart|
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