Mapping the Cell-Surface N-Glycoproteome of Human Hepatocytes Reveals Markers for Selecting a Homogeneous Population of iPSC-Derived Hepatocytes

• Published in: Stem cell reports Vol. 7; no. 3; pp. 543 - 556
• Main Authors: Mallanna, Sunil K., Cayo, Max A., Twaroski, Kirk, Gundry, Rebekah L., Duncan, Stephen A.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 13.09.2016; Elsevier
• Subjects: Biomarkers; Cell Differentiation; Cluster Analysis; Hepatocytes - cytology; Hepatocytes - metabolism; Humans; Induced Pluripotent Stem Cells - cytology; Membrane Glycoproteins - metabolism; Organ Specificity - genetics; Phenotype; Proteome; Proteomics - methods; Resource
• ISSN: 2213-6711; 2213-6711
• DOI: 10.1016/j.stemcr.2016.07.016

When comparing hepatic phenotypes between iPSC-derived hepatocyte-like cells from different liver disease patients, cell heterogeneity can confound interpretation. We proposed that homogeneous cell populations could be generated by fluorescence-activated cell sorting (FACS). Using cell-surface capture proteomics, we identified a total of 300 glycoproteins on hepatocytes. Analyses of the expression profiles during the differentiation of iPSCs revealed that SLC10A1, CLRN3, and AADAC were highly enriched during the final stages of hepatocyte differentiation. FACS purification of hepatocyte-like cells expressing SLC10A1, CLRN3, or AADAC demonstrated enrichment of cells with hepatocyte characteristics. Moreover, transcriptome analyses revealed that cells expressing the liver gene regulatory network were enriched while cells expressing a pluripotent stem cell network were depleted. In conclusion, we report an extensive catalog of cell-surface N-linked glycoproteins expressed in primary hepatocytes and identify cell-surface proteins that facilitate the purification of homogeneous populations of iPSC-derived hepatocyte-like cells. •Identified N-linked glycoproteins occupying surface of primary human hepatocytes•SLC10A1, CLRN3, and AADAC are expressed on ∼25% of iPSC-derived hepatocytes•FACS with these markers enriches hepatic character in iPSC-derived hepatocytes•SLC10A1 sorted cells provide homogeneous populations of hepatocyte-like cells Most iPSC differentiation protocols generate hepatocytes with heterogeneous expression of mature hepatic markers, which confounds analyses of complex traits and subtle phenotypes. Duncan and colleagues utilize proteomics, transcriptional profiling, and FACS to identify hepatocyte-restricted cell-surface proteins that mark a subpopulation of iPSC-derived hepatocytes, allow purification of iPSC-derived hepatocytes with enriched hepatic character, and minimize variability between differentiations.