Modeling inherited metabolic disorders of the liver using human induced pluripotent stem cells
S. Tamir Rashid, Sebastien Corbineau, Nick Hannan, Stefan J. Marciniak, Elena Miranda, Graeme Alexander, Isabel Huang-Doran, Julian Griffin, Lars Ahrlund-Richter, Jeremy Skepper, Robert Semple, Anne Weber, David A. Lomas, Ludovic Vallier
S. Tamir Rashid, Sebastien Corbineau, Nick Hannan, Stefan J. Marciniak, Elena Miranda, Graeme Alexander, Isabel Huang-Doran, Julian Griffin, Lars Ahrlund-Richter, Jeremy Skepper, Robert Semple, Anne Weber, David A. Lomas, Ludovic Vallier
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Modeling inherited metabolic disorders of the liver using human induced pluripotent stem cells

Abstract

Human induced pluripotent stem (iPS) cells hold great promise for advancements in developmental biology, cell-based therapy, and modeling of human disease. Here, we examined the use of human iPS cells for modeling inherited metabolic disorders of the liver. Dermal fibroblasts from patients with various inherited metabolic diseases of the liver were used to generate a library of patient-specific human iPS cell lines. Each line was differentiated into hepatocytes using what we believe to be a novel 3-step differentiation protocol in chemically defined conditions. The resulting cells exhibited properties of mature hepatocytes, such as albumin secretion and cytochrome P450 metabolism. Moreover, cells generated from patients with 3 of the inherited metabolic conditions studied in further detail (α1-antitrypsin deficiency, familial hypercholesterolemia, and glycogen storage disease type 1a) were found to recapitulate key pathological features of the diseases affecting the patients from which they were derived, such as aggregation of misfolded α1-antitrypsin in the endoplasmic reticulum, deficient LDL receptor–mediated cholesterol uptake, and elevated lipid and glycogen accumulation. Therefore, we report a simple and effective platform for hepatocyte generation from patient-specific human iPS cells. These patient-derived hepatocytes demonstrate that it is possible to model diseases whose phenotypes are caused by pathological dysregulation of key processes within adult cells.

Authors

S. Tamir Rashid, Sebastien Corbineau, Nick Hannan, Stefan J. Marciniak, Elena Miranda, Graeme Alexander, Isabel Huang-Doran, Julian Griffin, Lars Ahrlund-Richter, Jeremy Skepper, Robert Semple, Anne Weber, David A. Lomas, Ludovic Vallier

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Figure 4

In vitro modeling of GSD1a using disease-specific human iPS cells.

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In vitro modeling of GSD1a using disease-specific human iPS cells. (A) G...
(A) GSD1a disease-specific human iPS cells differentiated to hepatocytes displayed functional activity characteristic of primary human hepatocytes, including intracellular presence of albumin, albumin secretion, and active CytP450 metabolism. Error bars denote SEM. (B) PAS staining showing excessive accumulation of intracellular glycogen in GSD1a disease-specific human iPS cell–derived hepatocytes compared with human iPS cell–derived hepatocytes from control subjects. n = 3. (C) BODIPY staining showed excessive accumulation of intracellular lipid in GSD1a disease-specific human iPS cell–derived hepatocytes compared with human iPS cell–derived hepatocytes from control subjects. n = 3. (D) Disease-specific human iPS cell–derived hepatocytes appropriately upregulated transcriptional targets of glucagon, as shown by quantitative RT-PCR analysis of PEPCK, glucose-6-phosphatase (G6P), and IGFBP1 expression analyzed 0, 1, 2, and 3 hours after stimulation with 100 nM glucagon hydrochloride. Error bars denote SEM. n = 3. (E) GSD1a disease-specific human iPS cell–derived hepatocytes secreted more lactate than did human iPS cell–derived hepatocytes from control subjects, as assessed by ELISA analysis of a 24-hour collection of cell culture medium. Error bars denote SEM. n = 3. Original magnification, ×40 (A; B, bottom; and C).