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. 2021 Feb 17;17(3):861-868.
doi: 10.7150/ijbs.56091. eCollection 2021.

PTEN-mediated AKT/β-catenin signaling enhances the proliferation and expansion of Lgr5+ hepatocytes

Jimin Han  1   2   3 , Kaijun Lin  3 , Xuezheng Zhang  3 , Lingchen Yan  3 , Yu Chen  1   2   3 , Haiyan Chen  1   2   3 , Jianjun Liu  4 , Jia Liu  3 , Yaojiong Wu  1   2
Affiliations

PTEN-mediated AKT/β-catenin signaling enhances the proliferation and expansion of Lgr5+ hepatocytes

Jimin Han et al. Int J Biol Sci. .

Abstract

Rationale: Compelling evidence suggests that Lgr5+ hepatocytes repair liver damage by promoting the regeneration of hepatocytes and ductal cells in the case of liver injury. The PTEN-mediated AKT/β-catenin signaling plays a key role in the regulation of innate immune regulation in the liver. However, the signaling pathways that control Lgr5+ hepatocyte proliferation in the liver remain unclear. Methods: In order to assess the involvement of PTEN-mediated AKT/β-catenin signaling in the expansion of Lgr5+ hepatocytes upon liver injuries, the Lgr5-CreER; Rosa-mTmG lineage tracing system was used to target Lgr5+ hepatocytes. Results: The tracing of Lgr5+ hepatocytes showed that PTEN deletion and β-catenin activation significantly promoted the proliferation of Lgr5+ hepatocytes. In converse, the simultaneous inhibition of PTEN and β-catenin limited Lgr5+ hepatocyte proliferation in the liver. Our findings provide an insight into understanding how PTEN-mediated AKT/β-catenin signaling regulates the proliferation of Lgr5+ hepatocytes. Conclusion: The outcomes can improve the application potential of Lgr5+ hepatocytes in the treatment of liver injury diseases and provide a new treatment option for liver cancer.

Keywords: AKT/β-catenin; Lgr5; hepatocyte; liver regeneration.; proliferation.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interest exists.

Figures

Figure 1
Figure 1
Both PTEN ablation and increased β-catenin signaling in Lgr5+ hepatocytes promoted the expansion of Lgr5+ hepatocytes. (A) Experimental scheme for tamoxifen injection and analysis of liver samples in Lgr5CreER; mTmG mice at 21 days post-injection. (B) Targeting of Lgr5+ hepatocytes and their progeny (green) in the liver with Lgr5CreER; mTmG reporter mice. Note the sporadic distribution of mGFP-labeled cells in the liver. (C) Representative images showing positive Lgr5+ hepatocytes and their progeny in Lgr5CreER; mTmG; PTENflox/flox mice. (D) A low-magnification image of a liver sample from Lgr5CreER; Ctnnb1Δ(ex3); YFP mice (left panel). A magnified image of the left panel showed labeled Lgr5+ hepatocytes (right panel). Arrowheads indicate mGFP+ hepatocytes on day 21 after injection. (E) Phosphorylated β-catenin (white) immunostaining of liver tissue showing the specific colocalization of p-β-catenin and YFP (upper panel). Immunostaining of liver section from Lgr5CreER; Ctnnb1Δ(ex3); YFP mice showed that mGFP-labeled cells colocalized with Albumin (white) (lower panel). (F) Immunostaining for Albumin (white) revealed colocalization of mGFP with Albumin-positive hepatocytes. (G-H) Immunostaining of liver tissues from Lgr5CreER; mTmG and Lgr5CreER; mTmG; PTENflox/flox mice for p-AKT antibody (white). Arrowheads indicate Lgr5+ hepatocytes with an activated AKT pathway. n = 3 mice for each genotype; CV: central vein; Scale bar: 50 µm.
Figure 2
Figure 2
PTEN-mediated AKT/β-catenin signaling controls Lgr5+ hepatocyte proliferation. (A) The number of labeled cells in the liver from Lgr5CreER; mTmG; Ctnnb1flox/flox mice was significantly reduced upon β-catenin deletion. (B) Quantitation of mGFP+/YFP+ cells in liver samples from mice with different genotypes. (C) p-β-catenin immunostaining to show activation or inhibition of β-catenin signaling in the liver from different genotypes. (D) Sections of liver in Lgr5CreER; mTmG; PTENflox/flox; Ctnnb1flox/flox mice showing that β-catenin deletion in PTEN-deficient Lgr5+ hepatocytes strongly blocked the proliferation of Lgr5+ hepatocytes (left panel). The high-magnification view of the left panel depicts sporadic mGFP+ cells (top right). Another higher magnification image showing Lgr5-labeled cells (bottom right). Arrowheads denote mGFP+ cells. (E) Quantitation of mGFP+ cells in liver samples from mice with different genotypes. ****, P<0.0001. Data shown are means ± SEM (n = 3 biological replicated experiments). Scale bar: 100 µm.
Figure 3
Figure 3
Schematic summary of the distribution of Lgr5+ hepatocytes and their progeny under alteration of PI3K/AKT and Wnt/β-catenin signaling. (A-E) Schematic depiction of the proliferative ability of Lgr5+ hepatocytes (green) during normal liver homeostasis (A) and under changes of PI3K/AKT and Wnt/β-catenin signaling (B-E). (F) Forced activation of β-catenin induced the expansion of Lgr5+ hepatocytes. (G) Schematic illustration of the crosstalk between Wnt/β-catenin and PI3K/AKT signaling pathways and their role in the proliferation of Lgr5+ hepatocytes.
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