SHP2 inhibition enhances Yes-associated protein–mediated liver regeneration in murine partial hepatectomy models
Ryan D. Watkins, EeeLN H. Buckarma, Jennifer L. Tomlinson, Chantal E. McCabe, Jennifer A. Yonkus, Nathan W. Werneburg, Rachel L. Bayer, Patrick P. Starlinger, Keith D. Robertson, Chen Wang, Gregory J. Gores, Rory L. Smoot
Ryan D. Watkins, EeeLN H. Buckarma, Jennifer L. Tomlinson, Chantal E. McCabe, Jennifer A. Yonkus, Nathan W. Werneburg, Rachel L. Bayer, Patrick P. Starlinger, Keith D. Robertson, Chen Wang, Gregory J. Gores, Rory L. Smoot
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Research Article Hepatology Therapeutics

SHP2 inhibition enhances Yes-associated protein–mediated liver regeneration in murine partial hepatectomy models

Abstract

Disrupted liver regeneration following hepatectomy represents an “undruggable” clinical challenge associated with poor patient outcomes. Yes-associated protein (YAP), a transcriptional coactivator that is repressed by the Hippo pathway, is instrumental in liver regeneration. We have previously described an alternative, Hippo-independent mechanism of YAP activation mediated by downregulation of protein tyrosine phosphatase nonreceptor type 11 (PTPN11, also known as SHP2) inhibition. Herein, we examined the effects of YAP activation with a selective SHP1/SHP2 inhibitor, NSC-87877, on liver regeneration in murine partial hepatectomy models. In our studies, NSC-87877 led to accelerated hepatocyte proliferation, improved liver regeneration, and decreased markers of injury following partial hepatectomy. The effects of NSC-87877 were lost in mice with hepatocyte-specific Yap/Taz deletion, and this demonstrated dependence on these molecules for the enhanced regenerative response. Furthermore, administration of NSC-87877 to murine models of nonalcoholic steatohepatitis was associated with improved survival and decreased markers of injury after hepatectomy. Evaluation of transcriptomic changes in the context of NSC-87877 administration revealed reduction in fibrotic signaling and augmentation of cell cycle signaling. Cytoprotective changes included downregulation of Nr4a1, an apoptosis inducer. Collectively, the data suggest that SHP2 inhibition induces a pro-proliferative and cytoprotective enhancement of liver regeneration dependent on YAP.

Authors

Ryan D. Watkins, EeeLN H. Buckarma, Jennifer L. Tomlinson, Chantal E. McCabe, Jennifer A. Yonkus, Nathan W. Werneburg, Rachel L. Bayer, Patrick P. Starlinger, Keith D. Robertson, Chen Wang, Gregory J. Gores, Rory L. Smoot

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Figure 4

NR4A1 in modulated by NSC in vitro and in vivo.

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NR4A1 in modulated by NSC in vitro and in vivo. (A) Nr4a1 mRNA relative ...
(A) Nr4a1 mRNA relative expression 40 hours after hepatectomy in vehicle- and NSC-treated mice (n = 3). Data are shown as mean ± SEM. (B) Liver lysates from resection specimen (0 hours) and 40 hours after hepatectomy probed for NR4A1, pNR4A1S351, and actin as a loading control in vehicle- and NSC-treated mice. (C) Primary mouse hepatocytes treated with NSC (10 μM) or vehicle for 24 hours and immunoblotted for pYAPY357, total YAP, pNR4A1S351, total NR4A1, and actin as a loading control. (D) Representative images of NR4A1 immunocytochemistry with mitochondrial (MitoTracker) and nuclei (DAPI) counterstains in isolated mouse hepatocytes treated with NSC (10 μM) or vehicle. Scale bars: 50 μm. (E) Mean fluorescence intensity (MFI) values for nuclear NR4A1 in NSC- (10 μM) or vehicle-treated hepatocytes. Median values represented by solid line, and IQR represented by dotted lines. In total, 35–50 nuclei were evaluated. (F) Mean Manders’ correlation coefficient of NR4A1 colocalized to mitochondria in vehicle- and NSC-treated hepatocytes. Correlation coefficient were calculated for 7 images (400×). Data are shown as mean ± SEM. (G) Relative caspase 3/7 activity in primary mouse hepatocytes 4 hours after glycochenodeoxycholate (GCDC) (50 μM) or vehicle treatment with or without NSC pretreatment (10 μM) for 16 hours, n = 3 (*P < 0.05, **P < 0.01). B and C are representative immunoblots from 2 independent experiments. Statistical analysis was performed with 2-tailed Student t test (E and F) and 1-way ANOVA (G).