Protective effect of S-adenosylmethionine on hepatic ischemia-reperfusion injury during hepatectomy in HCC patients with chronic HBV infection

Hepatocellular carcinoma (HCC), as a fatal disease, is the third leading cause of cancer death worldwide [1]. Its major cause is infection by hepatitis B virus (HBV), especially in the Asia-Pacific region. Currently, partial hepatectomy and orthotopic liver transplantation (OLT) have been considered as the most effectual treatment methods for HCC.

The Pringle maneuver is extensively applied as a standard procedure during liver resection to control blood loss, which can result in significant hepatic ischemia and reperfusion injury (HIRI). However, the amount of bleeding during surgery is closely related to higher postoperative complication rates [2]. Ischemia and reperfusion may induce oxidative stress and free radical formation [3], promote excessive serum liver enzyme release and further lead to severe organ dysfunction and organ failure. Due to the high rate of liver cell metabolism, it is an organ which is vulnerable to the influence of anoxia. However, the underlying mechanisms have not been fully clarified. To mitigate HIRI, several measures have been used in clinical application, such as ischemic preconditioning, ischemic postconditioning and pharmacological preconditioning.

S-adenosylmethionine (SAMe or AdoMet) is a natural substance and serves as a reduced glutathione, an endogenous methyl donor and ATP precursor in various tissues [4], and plays a crucial role in liver cells and especially for regulating the sensitivity of liver cells to various kinds of injuries. Chronic liver diseases lead to the depletion of SAMe [5, 6], which can exacerbate liver injury as reported by preclinical studies [7]. Recently, our previous study demonstrated that postoperative SAMe therapy can provide a protective effect on residual liver function of HCC patients [8]. However, there is a lack of clinical trials focusing on how to reduce the HIRI to human livers using pharmacological strategies.

For many years, SAMe has been used extensively as a useful drug in the treatment of certain diseases [9, 10]. The aim of the present study was to evaluate whether treatment with SAMe could alleviate HIRI in patients undergoing partial hepatectomy with the Pringle maneuver, especially for patients with HCC associated with chronic HBV infection and cirrhosis.

One hundred and thirty patients were assessed for potential participants. Study exclusion criteria were concomitant availability in another randomized controlled trial (n = 15), disagreement (n = 15) and inclusion criteria not being fulfilled (n = 10). Furthermore, after randomization, there were four patients in the PR group and five in the PO group who had to be excluded from analysis (the Pringle maneuver not having been used). Finally, 26 patients were included in the PR group, 25 in the PO group, and 30 in the control group.

HCC is one of the deadliest malignancies worldwide. Due to the obscure etiological agents and insufficient surveillance systems, HCC is frequently found at a late stage, often with severe liver cirrhosis and deteriorating hepatic functional reserve, especially in China. As is well known, the most important predisposing cause is infection with HBV [15]. Partial hepatectomy still remains the mainstay of therapy for HCC, not only for early stage, but also and especially for those who have large hepatic lesions. During liver surgery, clamping of the primary portahepatis is performed to control intraoperative blood loss, but this maneuver can cause serious liver function failure. Patients with cirrhosis are more impressionable to HIRI than patients with normal liver.

Under certain conditions, the protective effects of HIRI can be achieved at different time points to decrease the ischemic damage using techniques such as ischemic preconditioning [16], remote ischemic preconception [17], pharmacological preconditioning and ischemic postconditioning [18].

The present study demonstrated that the pharmacological preconditioning and postconditioning with SAMe exerted a protective effect on patients undergoing liver resection with clamping of the primary portahepatis. To the best of our knowledge, it was the first time that pharmacological preconditioning has been used in partial hepatectomy under continuous inflow occlusion, in which cirrhosis was the leading characteristic in up to 82.7% and all the patients were infected with HBV. Not only postoperative liver injury, but also the postoperative clinical outcomes were improved by pharmacological preconditioning and postconditioning. Also, the protective effect under preconditioning was much better than that under postconditioning, as represented by liver function, plasma cytokine and liver function assessment. Our preliminary results would suggest that increase of SAMe in hepatocytes through peripheral vein infusion would be advantageous in alleviating the HIRI.

SAMe is a metabolically pleiotropic molecule, which occurs in multiple cellulate reactions and serves as a key precursor of glutathione [19]. It can eliminate free radicals to deter the cellular damage [20] and act as a methyl donor in myriad biological and biochemical events [21]. Also, it participates in the reaction of transmethylation, transsulfuration and aminopropylation to preserve hepatocyte viability and proliferation. Patients with cirrhosis often have severe hepatocyte dysfunction, which can be significantly improved by increasing SAMe levels in liver cells [10, 11], especially in hepatocytes after partial hepatectomy [10]. Moreover, SAMe and its metabolite can modulate proinflammatory cytokines, such as TNF-α and interleukins. Additionally, it can relieve cholestasis, which is caused by cirrhosis, viral hepatitis and surgical injury [9], the underlying mechanisms of which are frequently associated with depletion of SAMe and glutathione [20]. Recently, Liu et al. [22] reported that the X protein of HBV can reduce SAMe and enhance the Mmethionine adenosyltransferase 2A (MAT2A) gene expression to inhibit apoptosis in hepatocytes. However, dual effects of SAMe have also been dramatically observed. Varela-Rey et al. [23] found that SAMe could impair liver regeneration, and another study [24] showed it could inhibit mitogenic activity through acting on the hepatocyte growth factor (HGF). These findings were not consistent with our present study. However, due to different emphasis of study design and limitations of current reported data, the underlying mechanisms and detailed effects of SAMe need our further exploration.

The change of liver function is an extremely complex process, the details of which are difficult to elucidate. The principal biochemical parameters used as indicators of hepatocyte damage after liver resection are ALT and AST. In this study, ALT and AST in the PR group were significantly lower than those in the control group after resection, and the mean levels in the PO group after resection were lower than in the control group, but there was no statistical significance. In subgroup analyses, non-cirrhosis patients had statistically lower mean levels of ALT and AST. However, the changes in measured value were more acute in cirrhosis than in non-cirrhosis patients. These findings suggest that the cirrhotic liver was sensitive to the protective effect of SAMe.

Surgical damage during resection can lead to the elevation of serum bilirubin. Our results showed that supplying of exogenous SAMe could significantly reduce serum bilirubin in the PR group compared to that in the control group on Day1, Day3 and Day5; in the PO group, it was significantly lower than that in the control group after administration of SAMe on Day1. The underlying mechanism may be related to the fact that the supply of SAMe can restore the membrane fluidity of hepatocytes and mitochondria, strengthen the metabolism and increase the production of ATP.

ICGR15 is one of the most popular methods for examination of liver function and can reflect the operative risk before partial hepatectomy and assess the prognosis of patients with cirrhosis. Real-time ICG elimination tests [25] are often used as a new method to monitor the functional reserve of human liver. In this study, treatment with SAMe before surgery made the liver functional reserve much better than in the group without administration of SAMe.

IL-6 and TNF-α are both well-known superactive proinflammatory mediators that mediate the inflammatory response to tissue damage. Through analyzing the plasma cytokine levels, we found that preconditioning with SAMe prompted IL-6 and TNF-α production in the PR group at levels that were significantly lower than those in the PO and control groups. Considering that SAMe can modulate proinflammatory cytokines in liver injury [10], the mechanism is likely related to the downregulation of NF-κB [26]. Taking into account the change of transaminases among three groups, our data indicated that hepatocyte dysfunction after liver resection with blood inflow occlusion was mediated by the excess increase of IL-6 and TNF-α, but validation of this deduction needs further studies.

Moreover, the beneficial effects of SAMe in clinical outcomes can also be seen in this study. The patients who received SAMe had lower surgical complications, and a significantly shortened hospital stay was also observed in the PR group, mainly due to damage limitation and the rapid recovery of liver function. However, the underlying mechanisms are still unclear. Whether they work through the regulation of NF-κB, or influence the expression of some microRNAs, should be elucidated in further studies.

In conclusion, our preliminary results demonstrated that preoperative administration of SAMe was useful for protecting the HIRI in HCC patients with chronic HBV infection and cirrhosis. Pharmacological preconditioning with SAMe can not only decrease liver injury after hepatic resection but also promote the life quality of patients after surgery. This strategy can be easily applied to relieve HIRI, especially in patients who have chronic HBV infection and cirrhosis.

Written informed consent was obtained from each patient for publication of this report and any accompanying images.