Waqar R. R. Farid

Similar liver transplantation survival with selected cardiac death donors and brain death donors

The outcome of orthotopic liver transplantation (OLT) with controlled graft donation after cardiac death (DCD) is usually inferior to that with graft donation after brain death (DBD). This study compared outcomes from OLT with DBD versus controlled DCD donors with predefined restrictive acceptance criteria.

Hepatocyte‐derived microRNAs as serum biomarkers of hepatic injury and rejection after liver transplantation

Recent animal and human studies have highlighted the potential of hepatocyte‐derived microRNAs (HDmiRs) in serum as early, stable, sensitive, and specific biomarkers of liver injury. Their usefulness in human liver transplantation, however, has not been addressed. The aim of this study was to investigate serum HDmiRs as markers of hepatic injury and rejection in liver transplantation. Serum samples from healthy controls and liver transplant recipients (n = 107) and peritransplant liver allograft biopsy samples (n = 45) were analyzed via the real‐time polymerase chain reaction quantification of HDmiRs (miR‐122, miR‐148a, and miR‐194). The expression of miR‐122 and miR‐148a in liver tissue was significantly reduced with prolonged graft warm ischemia times. Conversely, the serum levels of these HDmiRs were elevated in patients with liver injury and positively correlated with aminotransferase levels. HDmiRs appear to be very sensitive because patients with normal aminotransferase values (<50 IU/L) had 6‐ to 17‐fold higher HDmiR levels in comparison with healthy controls (P < 0.005). During an episode of acute rejection, serum HDmiRs were elevated up to 20‐fold, and their levels appeared to rise earlier than aminotransferase levels. HDmiRs in serum were stable during repeated freezing and thawing. In conclusion, this study shows that liver injury is associated with the release of HDmiRs into the circulation. HDmiRs are promising candidates as early, stable, and sensitive biomarkers of rejection and hepatic injury after liver transplantation. Liver Transpl 18:290–297, 2012.

Sensitive detection of hepatocellular injury in chronic hepatitis C patients with circulating hepatocyte-derived microRNA-122

As chronic hepatitis C patients with progressive disease can present themselves with normal ALT levels, more sensitive biomarkers are needed. MicroRNAs are newly discovered small noncoding RNAs that are stable and detectable in the circulation. We aimed to investigate the association between hepatocyte‐derived microRNAs in serum and liver injury in patients with chronic hepatitis C. The hepatocyte‐derived miR‐122 and miR‐192 were analysed in sera of 102 chronic HCV‐infected patients and 24 healthy controls. Serum levels of miR‐122 and miR‐192 correlated strongly with ALT (R = 0.67 and R = 0.65, respectively, P < 0.001 for both). Median levels of miR‐122 and miR‐192 in HCV‐infected patients were 23 times and 8 times higher as in healthy controls (P < 0.001 for both). Even within the HCV‐infected patients with a normal ALT (n = 38), the levels of miR‐122 and miR‐192 were 12 times and 4 times higher compared with healthy controls (P < 0.001 for both). Multivariate logistic regression analyses showed that only miR‐122 was a significant predictor of the presence of chronic HCV infection (P = 0.026). Importantly, miR‐122 was also superior in discriminating chronic HCV‐infected patients with a normal ALT from healthy controls compared with the ALT level (AUC = 0.97 vs AUC = 0.78, P = 0.007). In conclusion, our study confirmed that liver injury is associated with high levels of hepatocyte‐derived microRNAs in circulation and demonstrated that in particular miR‐122 is a sensitive marker to distinguish chronic hepatitis C patients from healthy controls. More sensitive blood markers would benefit especially those patients with minor levels of hepatocellular injury, who are not identified by current screening with ALT testing.

Secreted Factors of Human Liver-Derived Mesenchymal Stem Cells Promote Liver Regeneration Early After Partial Hepatectomy

Rapid liver regeneration is required after living-donor liver transplantation and oncologic liver resections to warrant sufficient liver function and prevent small-for-size syndrome. Recent evidence highlights the therapeutic potential of mesenchymal stem cells (MSC) for treatment of toxic liver injury, but whether MSC and their secreted factors stimulate liver regeneration after surgical injury remains unknown. Therefore, the aim of this study is to investigate the effect of human liver-derived MSC-secreted factors in an experimental liver resection model. C57BL/6 mice were subjected to a 70% partial hepatectomy and treated with either concentrated MSC-conditioned culture medium (MSC-CM) or vehicle control. Animals were analyzed for liver and body weight, hepatocyte proliferation, and hepatic gene expression. Effects of MSC-CM on gene expression in a human hepatocyte-like cell line (Huh7 cells) were analyzed using genome-wide gene expression arrays. Liver regeneration was significantly stimulated by MSC-CM as shown by an increase in liver to body weight ratio and hepatocyte proliferation. MSC-CM upregulated hepatic gene expression of cytokines and growth factors relevant for cell proliferation, angiogenesis, and anti-inflammatory responses. In vitro, treatment of Huh7 cells with MSC-CM significantly altered expression levels of ~3,000 genes. Functional analysis revealed strong effects on networks associated with protein synthesis, cell survival, and cell proliferation. This study shows that treatment with MSC-derived factors can promote hepatocyte proliferation and regenerative responses in the early phase after surgical resection. MSC-CM may represent a feasible new strategy to promote liver regeneration in patients undergoing extensive liver resection or after transplantation of small liver grafts.

Biomarkers to assess graft quality during conventional and machine preservation in liver transplantation

A global rising organ shortage necessitates the use of extended criteria donors (ECD) for liver transplantation (LT). However, poor preservation and extensive ischemic injury of ECD grafts have been recognized as important factors associated with primary non-function, early allograft dysfunction, and biliary complications after LT. In order to prevent for these ischemia-related complications, machine perfusion (MP) has gained interest as a technique to optimize preservation of grafts and to provide the opportunity to assess graft quality by screening for extensive ischemic injury. For this purpose, however, objective surrogate biomarkers are required which can be easily determined at time of graft preservation and the various techniques of MP. This review provides an overview and evaluation of biomarkers that have been investigated for the assessment of graft quality and viability testing during different types of MP. Moreover, studies regarding conventional graft preservation by static cold storage (SCS) were screened to identify biomarkers that correlated with either allograft dysfunction or biliary complications after LT and which could potentially be applied as predictive markers during MP. The pros and cons of the different biomaterials that are available for biomarker research during graft preservation are discussed, accompanied with suggestions for future research. Though many studies are currently still in the experimental setting or of low evidence level due to small cohort sizes, the biomarkers presented in this review provide a useful handle to monitor recovery of ECD grafts during clinical MP in the near future.

MicroRNA profiles in graft preservation solution are predictive of ischemic-type biliary lesions after liver transplantation

BACKGROUND & AIMS Ischemic-type biliary lesions (ITBL) are the second most common cause of graft loss after liver transplantation. Though the exact pathophysiology of ITBL is unknown, bile duct injury during graft preservation is considered to be a major cause. Here we investigated whether the release of cholangiocyte-derived microRNAs (CDmiRs) during graft preservation is predictive of the development of ITBL after liver transplantation. METHODS Graft preservation solutions (perfusates) and paired liver biopsies collected at the end of cold ischemia were analysed by RT-qPCR for CDmiR-30e, CDmiR-222, and CDmiR-296 and hepatocyte-derived miRNAs (HDmiRs) HDmiR-122 and HDmiR-148a. MicroRNAs in perfusates were evaluated on their stability by incubation and fractionation experiments. MicroRNA profiles in perfusates from grafts that developed ITBL (n=20) and grafts without biliary strictures (n=37) were compared. RESULTS MicroRNAs in perfusates were proven to be stable and protected against degradation by interacting proteins. Ratios between HDmiRs/CDmiRs were significantly higher in perfusates obtained from grafts that developed ITBL (p<0.01) and were identified as an independent risk factor by multivariate analysis (p<0.01, HR: 6.89). The discriminative power of HDmiRs/CDmiRs in perfusates was validated by analysis of separate brain death- (DBD) and cardiac death donors (DBD; p ≤ 0.016) and was superior to expression in liver biopsies (C=0.77 in perfusates vs. C<0.50 in biopsies). CONCLUSIONS This study demonstrates that differential release of CDmiRs during graft preservation is predictive of the development of ITBL after liver transplantation. This provides new evidence for the link between graft-related bile duct injury and the risk for later development of ITBL.

The Importance of Portal Venous Blood Flow in Ischemic‐Type Biliary Lesions after Liver Transplantation

Ischemic‐type biliary lesions (ITBL) are the most frequent cause of nonanastomotic biliary strictures after liver transplantation. This complication develops in up to 25% of patients, with a 50% retransplantation rate in affected patients. Traditionally, ischemia‐reperfusion injury to the biliary system is considered to be the major risk factor for ITBL. Several other risk factors for ITBL have been identified, including the use of liver grafts donated after cardiac death, prolonged cold and warm ischemic times and use of University of Wisconsin preservation solution. In recent years however, impaired microcirculation of the peribiliary plexus (PBP) has been implicated as a possible risk factor. It is widely accepted that the PBP is exclusively provided by blood from the hepatic artery, and therefore, the role of the portal venous blood supply has not been considered as a possible cause for the development of ITBL. In this short report, we present three patients with segmental portal vein thrombosis and subsequent development of ITBL in the affected segments in the presence of normal arterial blood flow. This suggests that portal blood flow may have an important contribution to the biliary microcirculation and that a compromised portal venous blood supply can predispose to the development of ITBL.

Significant Contribution of the Portal Vein to Blood Flow Through the Common Bile Duct

Objective:The aim of this study was to determine the contribution of the hepatic artery, gastroduodenal artery, and portal vein to the microvascular blood flow in the common bile duct (CBD). Background:Biliary complications are a common cause of graft loss after liver transplantation. The occurrence is, partly, attributed to hepatic artery thrombosis, which is considered to be the sole provider of blood flow to the bile ducts. However, the contribution of the portal vein and the gastroduodenal artery to the bile ducts is unknown. Methods:Microvascular blood flow in the CBD was determined in 15 patients who underwent a pancreaticoduodenectomy with a combination of laser Doppler flowmetry and reflectance spectrophotometry. Microvascular blood flow was measured at baseline, during clamping the portal vein, during clamping the hepatic artery, and during clamping both. After transection of the CBD, these 4 measurements were repeated. Results:Compared with baseline measurements, the microvascular blood flow through the CBD decreased to 62% after clamping the portal vein, 51% after clamping the hepatic artery, and 31% after clamping both. After the CBD was transected, these 3 measurements were 60%, 31%, and 20%, respectively. Conclusions:Historically, the hepatic artery has been considered mainly responsible for biliary blood flow. We show that after transection of the CBD, mimicking the situation after liver transplantation, the contribution of the portal vein to the microvascular blood flow through the CBD is 40%. This study emphasizes the importance of the portal vein, and disturbances in portal venous blood flow could contribute to the formation of biliary complications after liver transplantation.

The ins and outs of microRNAs as biomarkers in liver disease and transplantation

Ongoing research is being conducted in the field of transplantation to discover novel, noninvasive biomarkers for assessment of graft quality before transplantation and monitoring of graft injury after transplantation. MicroRNAs (miRNAs) are among the most promising in this field. MiRNAs are small noncoding RNAs that function as important regulators of gene expression in response to cellular stress and disease. An advantage that makes miRNAs attractive candidates for biomarker research is their fast release from cells in response to stress and injury, which can occur via different routes. In the context of liver transplantation (LT), noninvasive measurement and stability of extracellular miRNAs in blood, bile, and graft perfusates has been linked to cell‐type specific injury and early graft outcome following LT. Furthermore, specific intrahepatic miRNA expression patterns have been associated with graft survival and recurrent disease, like hepatitis C virus‐related fibrosis and hepatocellular carcinoma. Therefore, miRNAs with strong predictive value and high sensitivity and specificity might be successfully applied to assess hepatic injury and to diagnose (recurrent) liver disease before, during and after LT. In this review, the current features and future prospects of miRNAs as biomarkers in and out of the liver are discussed.

Relationship between the histological appearance of the portal vein and development of ischemic‐type biliary lesions after liver transplantation

Ischemic‐type biliary lesions (ITBLs) are a major cause of morbidity after liver transplantation (LT). Their assumed underlying pathophysiological mechanism is ischemia/reperfusion injury of the biliary tree, in which the portal circulation has been proposed recently to have a role. The aim of this study was to investigate whether early histological changes, particularly in the portal vein, predispose patients to ITBLs. A case‐control study of 22 LT recipients was performed through a retrospective assessment of more than 30 histological parameters in 44 intraoperative liver biopsy samples taken after cold ischemia (time 0) and portal reperfusion (time 1). Eleven grafts developed ITBLs requiring retransplantation (the ITBL group), and 11 matched controls had normally functioning grafts 11 years after LT on average (the non‐ITBL group). Additionally, 11 liver biopsy samples from hemihepatectomies performed for metastases of colorectal cancer (CRC) were assessed similarly. Analyses showed no significant histological differences at time 0 between the ITBL and non‐ITBL groups. However, the time 1 biopsy samples from the ITBL group showed smaller portal vein branches (PVBs) significantly more often than the samples from the non‐ITBL group, which also showed persisting paraportal collateral vessels. Larger PVBs and paraportal collateral vessels were also found in the CRC group. A morphometric analysis confirmed these findings and showed that PVB measurements were significantly lower for the ITBL group at time 1 versus the ITBL group at time 0 and the non‐ITBL and CRC groups (they were largest in the CRC group). Thus, the PVB dimensions decreased in the ITBL group in comparison with the time 0 biopsy samples, and they were significantly smaller at time 1 in comparison with the dimensions for the non‐ITBL and CRC groups. In conclusion, a smaller PVB lumen size in postreperfusion biopsy samples from liver grafts, suggesting a relatively decreased portal blood flow, is associated with a higher incidence of ITBLs. These findings support recent clinical studies suggesting a possible pathophysiological role of portal blood flow in the oxygenation of the biliary tree after LT. Liver Transpl 19:1088‐1098, 2013.