Gastrointestinal
Loss of p27Kip1 accelerates DNA replication after partial hepatectomy in mice

https://doi.org/10.1016/S0022-4804(03)00052-0Get rights and content

Abstract

Background. Liver regeneration after partial hepatectomy (PH) is accomplished by a synchronous replication of hepatocytes. Both positive and negative regulators of cyclin-dependent protein kinase (Cdk) have been implicated in hepatocyte proliferation, but their specific roles in vivo remain to be clarified. To investigate the specific role of p27Kip1, a member of the Cip/Kip family of Cdk inhibitors, in cell-cycle regulation during liver regeneration, p27-knockout mice were studied after PH.

Materials and methods. Under ether anesthesia, mice were subjected to 70% PH. Animals were sacrificed at intervals after the surgery, and the remnant liver was harvested and analyzed.

Results. In p27-deficient mice, the timing of DNA synthesis was significantly accelerated with a perturbation in the ordered distribution of proliferating cells in the hepatic lobule. p27 deficiency, however, did not affect the whole population of cycling cells, the number of apoptotic cells, or liver injury and mortality after PH.

Conclusion. These data provide in vivo evidence that p27 functions as a brake in the “start” of the hepatocyte cell cycle, thereby coordinating temporally and spatially the onset of DNA synthesis of hepatocytes within the hepatic lobules.

Introduction

An extended hepatectomy is adopted to remove advanced biliary tract carcinomas 1, 2, 3. The risk of liver surgery increases in proportion to the amount of liver parenchyma resected, mainly as a result of liver failure after the surgery. Recovery of the remnant liver after such surgery, therefore, is critical for the outcome of the hepatectomized patient4, 5. Liver regeneration after partial (70%) hepatectomy (PH) has been extensively studied in rodents[6]. In this process, most hepatocytes reenter the cell cycle and arrest in a relatively synchronous manner [7], which is supposedly controlled by a precise balance between proliferative and antiproliferative signals 8, 9.

Eukaryotic cell-cycle progression is regulated by a family of cyclin-dependent kinases (Cdks)[10]. Cdk activity is positively regulated by association with regulatory subunits, cyclins, and is negatively regulated by binding to a group of Cdk inhibitors (CKIs)11, 12, 13. One CKI, p27Kip1 binds to a wide range of cyclin/Cdk complexes including cyclin D/Cdk4 and Cdk6 and cyclin E/Cdk2, and inhibits their activities14, 15. p27Kip1 deficiency in mice causes an increase in body size, multiple organ hyperplasia, retinal dysplasia, and pituitary tumors by increasing the number of postmitotic, properly differentiated cells16, 17, 18. p27Kip1-deficient primary hepatocytes have been shown to exhibit an increased activity of Cdk2, and could proliferate more easily than wild-type hepatocytes17, 19. These observations indicate that p27Kip1 plays a crucial role in regulating cell-cycle progression both in vitro and in vivo.

p27Kip1 was shown to modulate Cdk activities prior to DNA synthesis after PH, and p27Kip1 has been proposed to be involved in the initiation of cell proliferation during the liver regeneration process20, 21, 22, 23. In this study, we addressed the role of p27Kip1 in cell-cycle regulation during liver regeneration after PH by using p27Kip1 knockout mice. Our results demonstrated that p27Kip1 functions as a brake in the “start” of the hepatocyte cell cycle, but loss of p27Kip1 does not significantly affect the later stage of liver regeneration after PH.

Section snippets

Animals

p27Kip1 heterozygous mice [16] were kindly provided by Nippon Roche Research Center (Kamakura, Japan), and were bred and maintained under specific pathogen free conditions. Mice used in this study were 9- to 12-week-old male mice lacking p27Kip1 and isogenic wild-type mice, whose genetic background is a mixture of C57BL/6 and 129SV. The genotypes of the mice were confirmed by polymerase chain reaction (PCR) amplification of genomic DNAs using primers corresponding to the endogenous p27Kip1 gene

Results

Based on a rodent model of PH, we performed surgical treatment with p27Kip1 knockout and isogenic wild-type mice. The mortality rate during the initial 5 days after 70% PH was 6.3% in p27-/- mice and 6.8% in p27+/+ mice, indicating that there was no significant difference between the two groups (P = 0.9496). Furthermore, no significant difference was observed between the two groups in the serum levels of a marker protein, alanine transaminase, which reflects the degree of liver injuries (data

Discussion

p27Kip1 originally was isolated as an inhibitor of the cyclin D-Cdk4 and cyclin E-Cdk2 complexes, and has been shown to be involved in the G1-S progression of the mammalian cell cycle14, 15. p27-deficient primary hepatocytes exhibit an increased activity of Cdk2, and proliferate more easily than wild-type hepatocytes17, 19. Thus, p27 acts as a negative regulator of cell cycle progression of hepatocytes in vitro. In the regenerating liver in rats, p27 bound to both cyclin D-Cdk4 and cyclin

Acknowledgements

The authors gratefully acknowledge Nippon Roche K.K. (Tokyo, Japan), Keiko Nakayama and Kei-ichi Nakayama (Kyushu University, Fukuoka, Japan) for providing p27-heterozygous mice, Ken Watanabe (Department of Geriatric Research, National Institute for Longevity Sciences, Obu, Japan) for his critical reading of the manuscript, Osamu Miyaishi (Aichi Medical University, Aichi, Japan), Shin Tanaka (National Institute for Longevity Sciences), and Kaori Takai-Kajihara (Department of Geriatric Research,

References (35)

  • G.M. Ledda-Columbano et al.

    Early increase in cyclin-D1 expression and accelerated entry of mouse hepatocytes into S phase after administration of the mitogen 1, 4-Bis[2–(3, 5-Dichloropyridyloxy)] benzene

    Am. J. Pathol.

    (2000)
  • R.L. Jirtle et al.

    Modulation of insulin-like growth factor-II/mannose 6-phosphate receptors and transforming growth factor-beta 1 during liver regeneration

    J. Biol. Chem.

    (1991)
  • T.C. Chen et al.

    Mutational analysis of the p27(kip1) gene in hepatocellular carcinoma

    Cancer Lett.

    (2000)
  • Y. Nimura et al.

    Hepatic segmentectomy with caudate lobe resection for bile duct carcinoma of the hepatic bilus

    World J. Surg.

    (1990)
  • Y. Nimura et al.

    Combined portal vein and liver resection for carcinoma of the biliary tract

    Br. J. Surg.

    (1991)
  • Y. Nimura et al.

    Hepatopancreatoduodenectomy for advanced carcinoma of the biliary tract

    Hepatogastroenterology

    (1991)
  • M. Nagino et al.

    Logistic regression and discriminant analyses of hepatic failure after liver resection for carcinoma of the biliary tract

    World J. Surg.

    (1993)
  • Cited by (0)

    View full text