Impact of endothelial nitric oxide synthase activation on accelerated liver regeneration in a rat ALPPS model

Abstract

Background: Although the associating liver partition and portal vein ligation for staged hepatectomy (ALPPS) induces more rapid liver regeneration than portal vein embolization, the mechanism remains unclear.

Aim: To assess the influence of inflammatory cytokines and endothelial nitric oxide synthase (eNOS) activation on liver regeneration in ALPPS.

Methods: The future liver remnant/body weight (FLR/BW) ratio, hepatocyte proliferation, inflammatory cytokine expression, and activation of the Akt-eNOS pathway were evaluated in rat ALPPS and portal vein ligation (PVL) models. Hepatocyte proliferation was assessed based on Ki-67 expression, which was confirmed using immunohistochemistry. The serum concentrations of inflammatory cytokines were measured using enzyme linked immune-solvent assays. The Akt-eNOS pathway was assessed using western blotting. To explore the role of inflammatory cytokines and NO, Kupffer cell inhibitor gadolinium chloride (GdCl₃), NOS inhibitor N-nitro-arginine methyl ester (L-NAME), and NO enhancer molsidomine were administered intraperitoneally.

Results: The ALPPS group showed significant FLR regeneration (FLR/BW: 1.60% ± 0.08%, P < 0.05) compared with that observed in the PVL group (1.33% ± 0.11%) 48 h after surgery. In the ALPPS group, serum interleukin-6 expression was suppressed using GdCl₃ to the same extent as that in the PVL group. However, the FLR/BW ratio and Ki-67 labeling index were significantly higher in the ALPPS group administered GdCl₃ (1.72% ± 0.19%, P < 0.05; 22.25% ± 1.30%, P < 0.05) than in the PVL group (1.33% ± 0.11% and 12.78% ± 1.55%, respectively). Phospho-Akt Ser⁴⁷³ and phospho-eNOS Ser¹¹⁷⁷ levels were enhanced in the ALPPS group compared with those in the PVL group. There was no difference between the ALPPS group treated with L-NAME and the PVL group in the FLR/BW ratio and Ki-67 labeling index. In the PVL group treated with molsidomine, the FLR/BW ratio and Ki-67 labeling index increased to the same level as in the ALPPS group.

Conclusion: Early induction of inflammatory cytokines may not be pivotal for accelerated FLR regeneration after ALPPS, whereas Akt-eNOS pathway activation may contribute to accelerated regeneration of the FLR.

Keywords: Cytokines; Hepatectomy; Liver regeneration; Molsidomine; NG-Nitroarginine methyl ester; Nitric oxide.