Loss of transforming growth factor β adaptor protein β-2 spectrin leads to delayed liver regeneration in mice

Abstract

Liver regeneration, following partial hepatectomy (PHx), occurs through precisely controlled and synchronized cell proliferation, in which quiescent hepatocytes undergo one to two rounds of replication, with restoration of liver mass and function. We previously demonstrated that loss of the Smad3/4 adaptor protein b-2 spectrin (b2SP) is associated with faster entry into S phase, and hepatocellular cancer formation. These observations led us to further pursue the role of b2SP in cell cycle progression in vivo. Liver regeneration studies with PHx in b2SP 1/2 mice reveal a surprising and significant decrease in liver/ body weight ratio at 48 hours after PHx in b2SP 1/2 mice in comparison to wildtype mice. At 48 hours after PHx we also observe decreased levels of cyclin E (2.4-fold, P < 0.05), Cdk1 (7.2-fold, P < 0.05), cyclin A, pRb (Ser249/Thr252), proliferative cell nuclear antigen (PCNA), cyclin D1 with elevated levels of pCdk1 (Thr14) (3.6-fold, P < 0.05). Strikingly, at 24 hours elevated levels of p53 (4-fold, P < 0.05), phospho-p53 (ser15 and ser20), and p21 (200-fold, P < 0.05) persisting to 48 hours after PHx further correlated with raised expression of the DNA damage markers pChk2 (Thr68) and cH2AX (S139). However, compromised cell cycle progression with loss of b2SP is not rescued by inhibiting p53 function, and that G 2 /M phase arrest observed is independent and upstream of p53. Conclusion: b2SP deficiency results in dysfunctional hepatocyte cell cycle progression and delayed liver regeneration at 48 hours after PHx, which is p53-independent. b2SP loss may increase susceptibility to DNA damage, impair cell cycle progression, and ultimately lead to hepatocellular cancer. (HEPATOLOGY 2011;53:1641-1650 L iver regeneration represents an example of precisely controlled and synchronized cell proliferation in vivo. Following two-thirds partial hepatectomy (PHx), 95% of normally quiescent hepatocytes exit G 0 , rapidly reenter the cell cycle, and undergo one or two rounds of replication, with restoration of liver mass and function.1 Cell cycle progression proceeds in a synchronized pattern following PHx. In mid to late G 1 , phosphorylation of the retinoblastoma protein (Rb) by Cdk4/6-cyclin D complexes initiates the cell cycle and mediates the G 1 /S-phase transition.2 Cdk2 then successively associates withAbbreviations: b2SP, b-2 spectrin; CKIs, cyclin-dependent-kinase-inhibitory proteins; ELF, embryonic liver fodrin; MAPK, mitogen-activated protein kinase; MEFs, mouse embryonic fibroblasts; MT, b2SP mutant; PHx, partial hepatectomy; TGFb, transforming growth factor beta.From the

TGF-β Signaling in Onset and Progression of Hepatocellular Carcinoma

“…One molecular mechanism for how liver cells can escape from the cytostatic TGF-β response was described by the work of Mishra and coworkers [14,15,16,17,18,19,20,21,22,23,24]. Initially, they found that disruption of the adaptor protein ELF, a β-spectrin, leads to disruption of TGF-β/Smad signaling.…”

Transforming growth factor‐β in liver cancer stem cells and regeneration

“…Currently, at least three stem cell populations are known to exist in both mouse27 and human adult liver28: 1) pericentral Axin2 + hepatocytes that can regenerate liver in normal homeostasis29; 2) periportal cells positive for Lgr530; 3) Prom1 + liver stem cells that are located within or adjacent to the Krt19 + bile duct epithelium 31. Human periportal cells also label for octamer 3/4 (Oct3/4), β2 spectrin (β2SP), and TBRII in both human and mouse liver regeneration 27, 28, 32.…”

“…Human periportal cells also label for octamer 3/4 (Oct3/4), β2 spectrin (β2SP), and TBRII in both human and mouse liver regeneration 27, 28, 32. Through previous studies in human liver donor and liver transplant specimens that represent human liver regeneration, as well as fulminant hepatic failure, putative liver progenitor/stem cell expansion has been observed during massive hepatic necrosis with fibrosis as well as submassive hepatic necrosis with hepatocytic lineage.…”

“…β2SP deficiency results in dysfunctional hepatocyte cell cycle progression and delayed liver regeneration at 48 hours after PHx. This defect is mediated by dysfunctional expression of cell cycle proteins and by increased DNA damage 27. Spatial and temporal expansion of TBRI and β2SP expression occurs as regeneration proceeds.…”

“…5). Mouse knockouts, human liver regeneration, and functional studies thus reveal a pivotal role for TGF‐β in suppressing cancer stem cells, as well as modulating processes of liver regeneration and fibrosis 27, 28. However, exactly how it is defined temporally with existing stem cell markers such as Axin1, Lgr5, Fancd2, and CD133 remains to be explored.…”