Characterization of the EBV-B cell line–derived iPSCs. (A) Reprogramming genes were not detected in PCR analysis of genomic DNA, indicating generation of integration-free iPSCs. Day 2 transfected cells were included as a positive control, and parental cells were used as a negative control. (B) AAT sequence of human iPSC lines generated from patient fibroblasts and EBV-B cell lines. The inherited Z mutation was preserved after reprogramming of both cell types. Human iPSC lines derived from either healthy volunteers (iH16) or other disease (iLC2, liver cirrhosis), which do not carry the Z mutation, were used as controls. (C) EBV-related genes (EBNA-1, EBNA-2, OriP, LMP-1, and BZLF-1) were analyzed by PCR analysis of genomic DNA obtained from human fibroblasts, parental B cells, and the B-cell line–derived iPSCs. PCR results reveal the absence of EBV DNA in B-cell line–derived iPSCs after passage 20. (D) Three sets of PCR primers were used to detect IGH rearrangements occurring in committed B cells. A clonal control was included as a positive control, and human ESC (H9) was used as a negative control. Both the parental EBV-B cell lines and the cell line–derived iPSC lines showed positive signals of IGH rearrangement. (E-F) Disease modeling potentials. On the basis of our hepatic differentiation protocol, all these patient iPSCs were able to directly differentiate into mature hepatocyte-like cells expressing albumin (green, left) and AAT (red, right); representative images are shown with iAF3 and iAB5 (E). Numerous PASD-positive inclusion bodies (one of the most important AAT deficiency–related liver pathologic features, pink) were detected within mature hepatocytes derived from the AAT patient iPSCs (iAF3, iAB5) but not in the control iPSC (iLC2)-derived hepatocytes (F). The level of PASD-positive inclusion body was decreased with carbamazepine treatment (iAF3, 10μM for 5 days, bottom right).