Stefanie Haegele

Evidence for serotonin as a relevant inducer of liver regeneration after liver resection in humans

Liver regeneration (LR) involves a complex interplay of growth factors and antagonists. In this context, platelet‐derived serotonin (5‐HT) has been identified as a critical inducer of LR in mice. Clinical evidence for a role of 5‐HT in LR in humans is lacking. Accordingly, serum and plasma 5‐HT was monitored perioperatively in 60 patients undergoing liver resection, of which 35 served as exploration and 25 as validation sets. Intraplatelet (IP) levels of 5‐HT were calculated by subtraction of plasma 5‐HT from serum values. Serum markers of liver function were used to evaluate LR and liver dysfunction (LD). In the exploration setting, IP 5‐HT levels significantly decreased after liver resection (P < 0.001) and gradually recovered during the first week. IP 5‐HT measured before surgery specifically predicted LD in the subsequent 7 days (area under the curve: 0.721; P = 0.029). Patients suffering from postoperative LD and morbidity were found to have reduced IP 5‐HT levels during the entire perioperative period. Furthermore, we validated that reduced preoperative IP 5‐HT (<73 ng/mL) was associated with an increased incidence of postoperative LD and morbidity (P =0.045 and P = 0.021) and were able to demonstrate that IP 5‐HT levels were an independent predictor of poor clinical outcome. Conclusions: These findings provide evidence that IP 5‐HT correlates with LR in humans: Patients with low IP 5‐HT before liver resection suffered from delayed hepatic regeneration. Therefore, IP 5‐HT levels may prove a helpful clinical marker to predict postoperative LD and clinical outcome before hepatic resection and initiate suitable interventions. (Hepatology 2014;60:257‐266)

The profile of platelet α‐granule released molecules affects postoperative liver regeneration

Platelets promote liver regeneration through site‐specific serotonin release from dense granules, triggering proliferative signaling in hepatocytes. However, the effects of factors derived from platelet α‐granules on liver regeneration are unclear, because α‐granules contain bioactive molecules with opposing functions. Because α‐granule molecules are stored in separate compartments, it has been suggested that platelets selectively release their α‐granule content dependent on the environmental stimulus. Therefore, we investigated the pattern of circulating α‐granule molecules during liver regeneration in 157 patients undergoing partial hepatectomy. We measured plasma levels of α‐granule‐derived factors in the liver vein at the end of liver resection, as well as on the first postoperative day. We observed a rapid accumulation of platelets within the liver after induction of liver regeneration. Platelet count and P‐selectin (a ubiquitous cargo of α‐granules) were not associated with postoperative liver dysfunction. However, low plasma levels of vascular endothelial growth factor (VEGF), but high levels of thrombospondin 1 (TSP‐1), predicted liver dysfunction after resection. Patients with an unfavorable postoperative α‐granule release profile (high TSP‐1/low VEGF) showed substantially worse postoperative clinical outcomes. The unfavorable postoperative α‐granule release profile was associated with increased postoperative portal venous pressure and von Willebrand factor antigen levels as a marker for intrahepatic endothelial dysfunction. Conclusion: The postoperative profile of circulating platelet‐derived factors correlates with the ability of the remnant liver to regenerate. Portal venous pressure and intrahepatic endothelial dysfunction might account for the selective granule release profile. Selective modulation of platelet α‐granule release in patients may represent an attractive target for therapeutic interventions to improve liver regeneration and clinical outcomes after partial hepatectomy. (Hepatology 2016;63:1675‐1688)

P2X1‐regulated IL‐22 secretion by innate lymphoid cells is required for efficient liver regeneration

Paracrine signalling mediated by cytokine secretion is essential for liver regeneration after hepatic resection, yet the mechanisms of cellular crosstalk between immune and parenchymal cells are still elusive. Interleukin‐22 (IL‐22) is released by immune cells and mediates strong hepatoprotective functions. However, it remains unclear whether IL‐22 is critical for the crosstalk between liver lymphocytes and parenchymal cells during liver regeneration after partial hepatectomy (PH). Here, we found that plasma levels of IL‐22 and its upstream cytokine, IL‐23, are highly elevated in patients after major liver resection. In a mouse model of PH, deletion of IL‐22 was associated with significantly delayed hepatocellular proliferation and an increase of hepatocellular injury and endoplasmic reticulum stress. Using Rag1−/− and Rag2−/−γc−/− mice, we show that the main producers of IL‐22 post‐PH are conventional natural killer cells and innate lymphoid cells type 1. Extracellular adenosine triphosphate (ATP), a potent danger molecule, is elevated in patients immediately after major liver resection. Antagonism of the P2‐type nucleotide receptors, P2X1 and P2Y6, significantly decreased IL‐22 secretion ex vivo. In vivo, specific inhibition of P2X1 was associated with decreased IL‐22 secretion, elevated liver injury, and impaired liver regeneration. Conclusion: This study shows that innate immune cell‐derived IL‐22 is required for efficient liver regeneration and that secretion of IL‐22 in the regenerating liver is modulated by the ATP receptor, P2X1. (Hepatology 2016;63:2004‐2017)

Plasma thrombospondin 1 as a predictor of postoperative liver dysfunction.

Liver regeneration following liver resection involves a complex interplay of growth factors and their antagonists. Thrombospondin 1 has recently been identified as a critical inhibitor of liver regeneration by the activation of transforming growth factor β1 in mice, and preliminary data seem to confirm its relevance in humans. This study aimed to confirm these observations in an independent validation cohort.

Bivalent role of intra-platelet serotonin in liver regeneration and tumor recurrence in humans

BACKGROUND & AIMS Besides its critical role during liver regeneration, serotonin (5-HT) has been found to act as a mitogenic factor in several neoplastic entities. Accordingly, we aimed to evaluate whether intra-platelet 5-HT (IP5-HT) was associated with oncological outcome after liver resection and concomitantly evaluate its ability to serve as a therapeutic target to promote liver regeneration. METHODS A total of 96 patients undergoing liver resection for malignant liver tumors were prospectively included. Optimized plasma and serum preparation were performed and IP5-HT levels were determined. Patients were followed up for postoperative liver dysfunction (LD), morbidity, disease free and overall survival (OS). RESULTS We found increased preoperative IP5-HT levels in patients with disease recurrence at 6 and 12months (p=0.046, p=0.020, respectively). In clear contrast, patients suffering from postoperative morbidity, severe morbidity or LD had significantly reduced IP5-HT levels (p=0.011, p=0.035, p=0.003, respectively). Patients with high IP5-HT levels (>134ng/ml) suffered from an increased incidence of postoperative disease recurrence at 6 and 12months (p=0.045, p=0.006, respectively) but exhibited a reduction in morbidity, severe morbidity, and LD (p=0.006, p=0.008, p=0.005, respectively). We confirmed these results in our two largest subgroups, demonstrating that they were independent of tumor type. This bivalent effect of IP5-HT was also reflected in patients receiving selective serotonin reuptake inhibitor treatment, who displayed a reduction in disease recurrence accompanied by an increase in postoperative morbidity. Yet, both early disease recurrence and morbidity worsened OS. CONCLUSION Herein, we present first clinical evidence for IP5-HT being associated with early disease recurrence after liver resection in humans. Thus, pharmacological intervention at the level of platelets and platelet-derived 5-HT to promote liver regeneration should be considered with caution. A careful definition of indications and timing is needed to promote liver regeneration without inducing deleterious effects. LAY SUMMARY Preoperative intra-platelet serotonin (IP5-HT) levels seem to substantially affect patient outcomes after liver resection for liver tumors. While there is a narrow window of IP5-HT levels where liver regeneration and tumor progression is balanced, excessively high IP5-HT levels (>134ng/ml IP5-HT) lead to an increased incidence of early tumor recurrence and excessively low IP5-HT levels (<73ng/ml IP5-HT) lead to a higher rate of morbidity. Ultimately, overall survival is negatively affected by both postoperative early disease recurrence and morbidity. ClinicalTrials.gov-Identifier: NCT01700231.

Perioperative Von Willebrand Factor Dynamics are Associated with Liver Regeneration and Predict Outcome after Liver Resection

von Willebrand Factor (vWF) was found to mediate platelet influx during the early phase of liver regeneration in mice. Furthermore, increased vWF‐antigen (vWF‐Ag) levels were shown to be predictive for outcome of patients with chronic liver disease. Accordingly, we aimed to assess the relevance of perioperative vWF‐Ag dynamics in terms of liver regeneration and clinical outcome in patients undergoing liver resection (LR). Accordingly, we observed that vWF‐Ag and its activity—estimated by ristocetin cofactor measurement—increased immediately after induction of liver regeneration and was associated with platelet accumulation within the liver. However, a significant vWF‐Ag burst was only observed in patients with unaffected postoperative liver regeneration. E‐selectin, as an established marker for endothelial cell activation, was found to correlate with vWF‐Ag in the liver vein after induction of liver regeneration (R = 0.535, P = 0.022). Preoperative vWF‐Ag levels significantly predicted postoperative liver dysfunction (LD; N = 95; area under the curve, 0.725; P = 0.009). Furthermore, a cutoff of vWF‐Ag ≥182% was defined to identify patients with a higher risk for postoperative LD or morbidity. This was confirmed within an independent mulitcenter validation cohort (N = 133). Ultimately, multivariable analysis revealed that vWF‐Ag was an independent predictor of postoperative LD and morbidity. Conclusion: Within this study, we were able to provide evidence that an initial vWF burst is required to allow for adequate platelet accumulation and concomitant liver regeneration post‐LR and might be abolished as a consequence of intrahepatic endothelial cell dysfunction. We were further able to reveal and validate the potential of preoperative vWF‐antigen levels to predict poor postoperative outcome in patients undergoing LR. Despite the pathophysiological relevance of our findings, vWF‐Ag seems to be a valuable tool for preoperative risk assessment in patients undergoing LR. (Hepatology 2018;67:1516‐1530)

Deficiency in Thrombopoietin Induction after Liver Surgery Is Associated with Postoperative Liver Dysfunction

Background and Aims Thrombopoietin (TPO) has been implicated in the process of liver regeneration and was found to correlate with hepatic function in patients with liver disease. With this investigation we aimed to determine if perioperative TPO levels were associated with postoperative outcome in patients undergoing liver resection. Methods Perioperative TPO was analyzed prior to liver resection as well as on the first and fifth postoperative day in 46 colorectal cancer patients with liver metastasis (mCRC) as well as 23 hepatocellular carcinoma patients (HCC). Serum markers of liver function within the first postoperative week were used to define liver dysfunction. Results While circulating TPO levels significantly increased one day after liver resection in patients without liver cirrhosis (mCRC) (P < 0.001), patients with underlying liver disease (HCC) failed to significantly induce TPO postoperatively. Accordingly, HCC patients had significantly lower TPO levels on POD1 and 5. Similarly, patients with major resections failed to increase circulating TPO levels. Perioperative dynamics of TPO were found to specifically predict liver dysfunction (AUC: 0.893, P < 0.001) after hepatectomy and remained an independent predictor upon multivariate analysis. Conclusions We here demonstrate that perioperative TPO dynamics are associated with postoperative LD. Postoperative TPO levels were found to be lowest in high-risk patients (HCC patients undergoing major resection) but showed an independent predictive value. Thus, a dampened TPO increase after liver resection reflects a poor capacity for hepatic recovery and may help to identify patients who require close monitoring or intervention for potential complications.

Perioperative Non-Invasive Indocyanine Green-Clearance Testing to Predict Postoperative Outcome after Liver Resection

Background Postoperative liver dysfunction may lead to morbidity and mortality after liver resection. Preoperative liver function assessment is critical to identify preexisting liver dysfunction in patients prior to resection. The aim of this study was to evaluate the predictive potential of perioperative indocyanine green (ICG)-clearance testing to prevent postoperative liver dysfunction and morbidity using standardized outcome parameters in a routine Western-clinical-setting. Study Design 137 patients undergoing partial hepatectomy between 2011 and 2013, at the general hospital of Vienna, were included. ICG-clearance was recorded one day prior to surgery as well as on the first and fifth postoperative day. Postoperative liver dysfunction was defined according to the International Study Group of Liver Surgery and evaluation of morbidity was based on the Dindo-Clavien classification. Statistical analyses were based on non-parametric tests. Results Preoperative reduced ICG—plasma disappearance rate (PDR) as well as increased ICG—retention rate at 15 min (R15) were able to significantly predict postoperative liver dysfunction (Area under the curve = PDR: 0.716, P = 0.018; R15: 0.719, P = 0.016). Furthermore, PDR <17%/min. or R15 >8%, were able to accurately predict postoperative complications prior to surgery. In addition to this, ICG-clearance on postoperative day 1 comparably predicted postoperative liver dysfunction (Area under the curve = PDR: 0.895; R15: 0.893; both P <0.001), specifically, PDR <10%/min or R15 >20% on postoperative day 1 predicted poor postoperative outcome. Conclusion PDR and R15 may represent useful parameters to distinguish preoperative high and low risk patients in a Western collective as well as on postoperative day 1, to identify patients who require closer monitoring for potential complications.

Early prediction of postoperative liver dysfunction and clinical outcome using antithrombin III-activity

Background and aims Antithrombin III (ATIII) has been reported to be associated with liver pathologies and was shown to predict outcome in patients undergoing liver resection for hepatocellular carcinoma. We now aimed to assess whether perioperative ATIII-activity could predict postoperative outcome in patients without underlying liver disease, as well as in a routine clinical setting of patients undergoing hepatic resection. Methods ATIII-activity was evaluated preoperatively and on the first (POD1) and fifth day after liver resection in a retrospective evaluation cohort of 228 colorectal cancer patients with liver metastasis (mCRC). We further aimed to prospectively validate our results in a set of 177 consecutive patients undergoing hepatic resection. Results Patients developing postoperative liver dysfunction (LD) had a more pronounced postoperative decrease in ATIII-activity (P<0.001). ATIII-activity on POD1 significantly predicted postoperative LD (P<0.001, AUC = 84.4%) and remained independent upon multivariable analysis. A cut-off value of 61.5% ATIII-activity was determined using ROC analysis. This cut-off was vital to identify high-risk patients for postoperative LD, morbidity, severe morbidity and mortality (P<0.001, respectively) with a highly accurate negative predictive value of 97%, which could be confirmed for LD (P<0.001) and mortality (P = 0.014) in our independent validation cohort. Further, mCRC patients below our cut-off suffered from a significantly decreased overall survival (OS) at 1 and 3 years after surgery (P = 0.011, P = 0.025). Conclusions The routine laboratory parameter ATIII-activity on POD1 independently predicted postoperative LD and was associated with clinical outcome. Patients with a postoperative ATIII-activity <61.5% might benefit from close monitoring and timely initiation of supportive therapy. Trial registration ClinicalTrials.gov NCT01700231

Preoperative von willebrand factor – antigen predicts clinical outcome after liver resection: a prospective, international, multicenter trial

Backgroundand Aims:vWF-antigenhasbeenshowntobeincreased in patients with portal hypertension and to predict mortality in patients with chronic liver disease. This study aimed to assess the clinical utility of preoperative vWF-antigen levels to predict poor postoperative outcome in patients undergoing liver resection in a routine clinical setting. Methods: 95patientsundergoingliverresectionservedasprospective exploration cohort and results were validated in an independent cohort of 133 patients for 4 different institutions. VWF-Ag was evaluated perioperativelyand postoperative outcomewas recorded. Results: Preoperative vWF-antigen levels significantly predicted postoperative liver dysfunction (LD, area under the curve [AUC]: 0.725, P=0.009). Furthermore, a cut-off of vWF-antigen ≥182% was defined to identify patients with a higher incidence of postoperative LDormorbidity(LD:≥182%:33.3%,<182%:5.9%,P<0.001;morbidity: ≥182%: 74.1%, <182%: 44.1%, P=0.008). We confirmed our results within a prospective validation cohort (LD: ≥182%: 20.0%, <182%: 5.2%, P=0.008; morbidity: ≥182%: 56.4%, <182%: 35.1%, P=0.015). Analyzing the entire cohort, we found that patients exceeding the cut-off suffered from a significantly increased incidence of postoperative LD, morbidity, prolonged hospitalization, intensive care unit stayand mortality. Conclusions: Within this study we were able to reveal and subsequently validate the potential of preoperative vWF-antigen levels to predict poor postoperative outcome in patients undergoing liver resection. As vWF-antigen is easily determined and available in most standard laboratories, it seems to be avaluable clinical marker to allow for preoperative risk-stratification of patients undergoing liver resection.