TY - JOUR
T1 - Genes and Pathways Promoting Long-Term Liver Repopulation by Ex Vivo hYAP-ERT2 Transduced Hepatocytes and Treatment of Jaundice in Gunn Rats
AU - Peterson, Esther A.
AU - Polgar, Zsuzsanna
AU - Devakanmalai, Gnanapackiam S.
AU - Li, Yanfeng
AU - Jaber, Fadi L.
AU - Zhang, Wei
AU - Wang, Xia
AU - Iqbal, Niloy J.
AU - Murray, John W.
AU - Roy-Chowdhury, Namita
AU - Quispe-Tintaya, Wilber
AU - Maslov, Alexander Y.
AU - Tchaikovskaya, Tatyana L.
AU - Sharma, Yogeshwar
AU - Rogler, Leslie E.
AU - Gupta, Sanjeev
AU - Zhu, Liang
AU - Roy-Chowdhury, Jayanta
AU - Shafritz, David A.
N1 - Funding Information:
P30 DK41296 Core Facilities: Administrative Core, Anna Caponigro, for typing manuscript; Animal Model, Stem Cell and Cell Therapy Core for providing DPPIV−/− rats, Wistar RHA rats, and Gunn rats for hepatocyte repopulation studies and David Neufeld for liver perfusions and preparations of hepatocytes; Molecular Biology and Next Generation Technology Core for preparation of hepatocyte RNA fractions and cDNA libraries for RNAseq studies; Imaging and Cell Structure Core, Hillary Guzick, for preparing sequential images stitched together for computerized analysis of liver repopulation by transplanted hepatic cells; Genetic Engineering and Gene Therapy Core for preparation of recombinant lentiviruses; Daqian Sun for conducting FACS analysis and purification of repopulating and host hepatocytes; and Kith Pradhan for analysis of RNAseq data, to identify specific locations of single nucleotide differences between hYAP1 versus rat Yap1, and hERT2 versus ESR1 in FACS-purified hepatocyte preparations.
Publisher Copyright:
© 2018 The Authors. Hepatology Communications published by Wiley Periodicals, Inc., on behalf of the American Association for the Study of Liver Diseases.
PY - 2019/1
Y1 - 2019/1
N2 - Hepatocyte transplantation is an attractive alternative to liver transplantation. Thus far, however, extensive liver repopulation by adult hepatocytes has required ongoing genetic, physical, or chemical injury to host liver. We hypothesized that providing a regulated proliferative and/or survival advantage to transplanted hepatocytes should enable repopulation in a normal liver microenvironment. Here, we repopulated livers of DPPIV− (dipeptidyl peptidase-4) rats and Ugt1a1 (uridinediphosphoglucuronate glucuronosyltransferase 1a1)-deficient Gunn rats (model of Crigler-Najjar syndrome type 1), both models without underlying liver injury, for up to 1 year by transplanting lenti-hYAP-ERT2 (mutated estrogen receptor ligand-binding domain 2)-transduced hepatocytes (YAP-Hc). Yap (yes-associated protein) nuclear translocation/function in YAP-Hc was regulated by tamoxifen. Repopulating YAP-Hc and host hepatocytes were fluorescence-activated cell sorting–purified and their transcriptomic profiles compared by RNAseq. After 1 year of liver repopulation, YAP-Hc clusters exhibited normal morphology, integration into hepatic plates and hepatocyte-specific gene expression, without dysplasia, dedifferentiation, or tumorigenesis. RNAseq analysis showed up-regulation of 145 genes promoting cell proliferation and 305 genes suppressing apoptosis, including hepatocyte growth factor and connective tissue growth factor among the top 30 in each category and provided insight into the mechanism of cell competition that enabled replacement of host hepatocytes by YAP-Hc. In Gunn rats transplanted with YAP-Hc+tamoxifen, there was a 65%-81% decline in serum bilirubin over 6 months versus 8%-20% with control-Hc, representing a 3-4-fold increase in therapeutic response. This correlated with liver repopulation as demonstrated by the presence of Ugt1a1-positive hepatocyte clusters in livers and western blot analysis of tissue homogenates. Conclusion: Tamoxifen-regulated nuclear translocation/function of hYAP-ERT2 enabled long-term repopulation of DPPIV−/− and Gunn rat livers by hYAP-ERT2-transduced hepatocytes without tumorigenesis. This cell transplantation strategy may offer a potential therapy for most of the inherited monogenic liver diseases that do not exhibit liver injury.
AB - Hepatocyte transplantation is an attractive alternative to liver transplantation. Thus far, however, extensive liver repopulation by adult hepatocytes has required ongoing genetic, physical, or chemical injury to host liver. We hypothesized that providing a regulated proliferative and/or survival advantage to transplanted hepatocytes should enable repopulation in a normal liver microenvironment. Here, we repopulated livers of DPPIV− (dipeptidyl peptidase-4) rats and Ugt1a1 (uridinediphosphoglucuronate glucuronosyltransferase 1a1)-deficient Gunn rats (model of Crigler-Najjar syndrome type 1), both models without underlying liver injury, for up to 1 year by transplanting lenti-hYAP-ERT2 (mutated estrogen receptor ligand-binding domain 2)-transduced hepatocytes (YAP-Hc). Yap (yes-associated protein) nuclear translocation/function in YAP-Hc was regulated by tamoxifen. Repopulating YAP-Hc and host hepatocytes were fluorescence-activated cell sorting–purified and their transcriptomic profiles compared by RNAseq. After 1 year of liver repopulation, YAP-Hc clusters exhibited normal morphology, integration into hepatic plates and hepatocyte-specific gene expression, without dysplasia, dedifferentiation, or tumorigenesis. RNAseq analysis showed up-regulation of 145 genes promoting cell proliferation and 305 genes suppressing apoptosis, including hepatocyte growth factor and connective tissue growth factor among the top 30 in each category and provided insight into the mechanism of cell competition that enabled replacement of host hepatocytes by YAP-Hc. In Gunn rats transplanted with YAP-Hc+tamoxifen, there was a 65%-81% decline in serum bilirubin over 6 months versus 8%-20% with control-Hc, representing a 3-4-fold increase in therapeutic response. This correlated with liver repopulation as demonstrated by the presence of Ugt1a1-positive hepatocyte clusters in livers and western blot analysis of tissue homogenates. Conclusion: Tamoxifen-regulated nuclear translocation/function of hYAP-ERT2 enabled long-term repopulation of DPPIV−/− and Gunn rat livers by hYAP-ERT2-transduced hepatocytes without tumorigenesis. This cell transplantation strategy may offer a potential therapy for most of the inherited monogenic liver diseases that do not exhibit liver injury.
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U2 - 10.1002/hep4.1278
DO - 10.1002/hep4.1278
M3 - Article
AN - SCOPUS:85113976949
SN - 2471-254X
VL - 3
SP - 129
EP - 146
JO - Hepatology Communications
JF - Hepatology Communications
IS - 1
ER -