BACKGROUND: Cholangiocarcinoma is a very aggressive tumor with poor survival. Therefore, early diagnosis and surgical resection are of paramount importance. Its diagnosis is difficult because access to the tumor is not easy. Biopsy is possible only for intrahepatic cholangiocarcinoma, which accounts for 10% of cases. Routine brush cytology from endoscopic retrograde cholangiopancreatography (ERCP) has a high specificity of 100% but unfortunately a low sensitivity of 30%. In this review we briefly describe new diagnostic techniques applicable to ERCP brush cytology specimens and targeting the genetic background of the disease, in particular fluorescence in situ hybridization (FISH) and digital image analysis (DIA).
DATE SOURCES: The PubMed database up to 2011 was used for the retrieval of relevant articles. The search terms FISH, fluorescence in situ hybridization, DIA, digital image analysis and cholangiocarcinoma were used. Both original and review articles were used.
RESULTS: FISH identifies cells with chromosomal abnormalities, mainly numerical aberrations, using a mixture of fluorescence-labeled probes. FISH offers a higher sensitivity than routine cytology, retaining a high level of specificity. The DIA criterion for malignancy is demonstration of aneuploidy. This technique increases the sensitivity to 40%, but the specificity remains low. Preliminary data from application to other tumors suggest that combination of FISH and DIA may be of further benefit.
CONCLUSIONS: The new techniques offer a significantly enhanced diagnostic efficacy in the evaluation of ERCP brush specimens. Apart from contributing to a more timely diagnosis, their wider application to cholangiocarcinoma may also facilitate the genetic study of the disease and add to our understanding of oncogenesis at the molecular level, with the prospect of identifying targets for novel therapeutic interventions.

BACKGROUND: Liver cell transplantation and bioartificial liver may provide metabolic support of liver function temporary and are prospective treatments for patients with liver failure. Mesenchymal stem cells (MSCs) are expected to be an ideal cell source for transplantation or liver tissue engineering, however the hepatic differentiation of MSCs is still insufficient for clinical application.
DATA SOURCES: A PubMed search on "mesenchymal stem cells", "liver cell" and "hepatocyte differentiation" was performed on the topic, and the relevant articles published in the past ten years were reviewed.
RESULTS: Hepatocyte-like cells differentiated from MSCs are a promising cell source for liver regeneration or tissue engineering. Although it is still a matter of debate as to whether MSC-derived hepatocytes may efficiently repopulate a host liver to provide adequate functional substitution, the majority of animal studies support that MSCs can become key players in liver-directed regenerative medicine. However the clinical application of human stem cells in the treatment of liver diseases is still in its infancy.
CONCLUSIONS: Future studies are required to improve the efficacy and consistency of hepatic differentiation from MSCs. It is necessary to better understand the mechanism to achieve transdifferentiation with high efficiency. More clinical trials are warranted to prove their efficacy in the management of patients with liver failure.

BACKGROUND: An updated definition of early allograft dysfunction (EAD) was recently validated in a multicenter study of 300 deceased donor liver transplant recipients. This analysis did not differentiate between donation after brain death (DBD) and donation after cardiac death (DCD) allograft recipients.
METHODS: We reviewed our prospectively entered database for all DBD (n=377) and DCD (n=38) liver transplantations between January 1, 2006 and October 30, 2011. The incidence of EAD as well as its ability to predict graft failure and survival was compared between DBD and DCD groups.
RESULTS: EAD was a valid predictor of both graft and patient survival at six months in DBD allograft recipients, but in DCD allograft recipients there was no significant difference in the rate of graft failure in those with EAD (11.5%) compared with those without EAD (16.7%) (P=0.664) or in the rate of death in recipients with EAD (3.8%) compared with those without EAD (8.3%) (P=0.565). The graft failure rate in the first 6 months in those with international normalized ratio ≥1.6 on day 7 who received a DCD allograft was 37.5% compared with 6.7% for those with international normalized ratio <1.6 on day 7 (P=0.022).
CONCLUSIONS: The recently validated definition of EAD is a valid predictor of patient and graft survival in recipients of DBD allografts. On initial assessment, it does not appear to be a useful predictor of patient and graft survival in recipients of DCD allografts, however a study with a larger sample size of DCD allografts is needed to confirm these findings. The high ALT/AST levels in most recipients of DCD livers as well as the predisposition to biliary complications and early cholestasis make these parameters as poor predictors of graft failure. An alternative definition of EAD that gives greater weight to the INR on day 7 may be more relevant in this population.

BACKGROUND: Caudate lobectomy has long been considered technically difficult. This study aimed to elaborate the significance of early control of short hepatic portal veins (SHPVs) in isolated hepatic caudate lobectomy or in hepatic caudate lobectomy combined with major partial hepatectomy, and to describe the anatomical characteristics of SHPVs.
METHODS:?The data of 117 patients who underwent either isolated or combined caudate lobectomy by the same team of surgeons from 2005 to 2009 were retrospectively analyzed. From 2005 to 2007 (group A, n=55), we carried out early control of short hepatic veins (SHVs) only; from 2008 to 2009 (group B, n=62), we carried out early control of both SHVs and SHPVs. The two groups were compared to evaluate which surgical procedure was better. A detailed anatomical study was then carried out on the last 25 consecutive patients in group B to study the number and distribution of SHPVs during surgery.
RESULTS: Patients in group B had less intra-operative blood loss, less impairment of liver function, shorter postoperative hospital stay, fewer postoperative complications and required less blood transfusion (P<0.05). The number of SHPVs in the 25 patients was 183, with 7.3±2.7 per patient. The diameters of SHPVs were 1 to 4 mm. On average, 3.4 SHPVs/patient came from the left portal vein, 2.2 from the bifurcation, 1.4 from the right portal vein, and 0.3 from the main portal vein. On average, 3.3 SHPVs/patient supplied segment I of the liver, 0.4 for segment II, 2.1 for segment IV, 1.4 for segment V and 0.1 for segment VI.
CONCLUSION:?Early control of SHPVs in isolated or combined hepatic caudate lobectomy may be a useful method to decrease surgical risk and improve postoperative recovery.

BACKGROUND: Prior experimentation has shown that loss of the tyrosine kinase (TK) signaling domain of the Ron receptor leads to marked hepatocyte protection in a model of lipopolysaccharide-induced acute liver failure (ALF) in D-galactosamine (GalN)-sensitized mice. The aim of this study was to identify the role of Ron in the regulation of hepatic gene expression.
METHODS: Microarray analyses were performed on liver RNA isolated sequentially from wild-type (WT) and TK-/- mice during the progression of ALF. Gene array data were validated using Western and immunohistochemistry analyses as well as with ex vivo culture systems.
RESULTS: At baseline, 101 genes were differentially expressed between WT and TK-/- livers, which regulate processes involved in hypoxia, proliferation, apoptosis and metabolism. One hour after ALF induction, WT livers exhibited increased cytokine expression compared to TK-/- livers, and after 4 hours, an induction of suppressor of cytokine signaling (SOCS) genes as well as JAK-STAT pathway activation were prominent in TK-/- livers compared to controls.
CONCLUSION: Our studies suggest a novel hepato-protective mechanism in Ron TK-/- mice wherein increased and sustained SOCS production and JAK-STAT activation in the hepatocyte may inhibit the destructive proinflammatory milieu and promote survival factors which blunt hepatic death and the ensuing development of ALF.

BACKGROUND: Phospholipid scramblase 1 (PLSCR1) not only participates in the transbilayer movement of phospholipids, but also plays a role in the pathogenesis and progression of cancers. The present study aimed to evaluate the effect of silencing PLSCR1 expression by RNA interference in colorectal cancer (CRC) and metastatic liver cancer.
METHODS: The expression of PLSCR1 in CRC and metastatic liver cancer samples was assessed by immunohistochemistry. The cultured cells with the highest expression were selected for subsequent experiments. We designed three siRNA oligonucleotide segments targeted at PLSCR1. Successful transfection was confirmed. The biological behavior of the cells in proliferation, adhesion, migration and invasion was determined.
RESULTS: PLSCR1 protein expression increased significantly in the majority of CRC and metastatic liver cancer samples compared with normal samples. Lovo cells had the highest expression of PLSCR1. The siRNA-390 oligonucleotide segment had the best silencing effect. After transfection, Lovo cell proliferation was significantly inhibited compared with the controls in the MTT assay. Laminin and fibronectin adhesion assays showed Lovo cell adhesion was also significantly inhibited. In the migration assay, the number of migrating cells in the PLSCR1 siRNA-390 group was 50±12, significantly lower than the number in the siRNA-N group (115±28) and in the control group (118±31). In an invasion test, the number of invading cells in the PLSCR1 siRNA-390 group was 60±18, significantly lower than that in the siRNA-N group (97±26) and the control group (103±24).
CONCLUSIONS: PLSCR1 is overexpressed in CRC and metastatic liver cancer. Silencing of PLSCR1 by siRNA inhibits the proliferation, adhesion, migration and invasion of Lovo cells, which suggests that PLSCR1 contributes to the tumorigenesis and tumor progression of CRC. PLSCR1 may be a potential gene therapy target for CRC and associated metastatic liver cancer.

BACKGROUND: While suppressor of cytokine signaling 3 (SOCS3) plays a crucial role in suppressing dysplasia and tumorigenesis, it also offers a typical instance of DNA methylation in the regulation of gene transcription, since the promoter region of the SOCS3 gene is rich in CpG islands (CGIs). During liver regeneration initiated by partial hepatectomy, SOCS3 acts as a suppressor to balance the acute-phase response and terminate the regeneration. This study aimed to determine whether the variation of SOCS3 expression throughout liver regeneration is also regulated by its DNA methylation.
METHODS: We established a 70% partial hepatectomy mouse model and the animals were sacrificed at indicated times to assess the SOCS3 expression. We performed bisulfite sequencing PCR and DNA sequencing to investigate the detailed cytosine methylation in the SOCS3 gene.
RESULTS: Within the promoter sequence, 58 CGIs were identified and 30 were found variously methylated before or after operation; however, methylation remained at a very low level. No evidence indicated that the total methylation level or the methylation of any CpG site regularly changed throughout liver regeneration.
CONCLUSION: DNA methylation or demethylation seems to be a relatively stable modification of cytosine, but not a dynamic and reversible process to regulate gene transcription in daily and acute pathophysiological events.

BACKGROUND: Early detection and treatment of hepatocellular carcinoma is crucial to improving the patients( survival. The hemodynamic changes caused by tumors can be serially measured using CT perfusion. In this study, we used a CT perfusion technique to demonstrate the changes of hepatic hemodynamics in early tumor growth, as a proof-of-concept study for human early hepatocellular carcinoma.
METHODS: VX2 tumors were implanted in the liver of ten New Zealand rabbits. CT perfusion scans were made 1 week (early) and 2 weeks (late) after tumor implantation. Ten normal rabbits served as controls. CT perfusion parameters were obtained at the tumor rim, normal tissue surrounding the tumor, and control liver; the parameters were hepatic blood flow, hepatic blood volume, mean transit time, permeability of capillary vessel surface, hepatic arterial index, hepatic arterial perfusion and hepatic portal perfusion. Microvessel density and vascular endothelial growth factor were correlated.
RESULTS: At the tumor rim, compared to the controls, hepatic blood flow, hepatic blood volume, permeability of capillary vessel surface, hepatic arterial index, and hepatic arterial perfusion increased, while mean transit time and hepatic portal perfusion decreased on both early and late scans (P<0.05). Hepatic arterial index increased (135%, P<0.05), combined with a sharp increase in hepatic arterial perfusion (182%, P<0.05) and a marked decrease in hepatic portal perfusion (-76%, P<0.05) at 2 weeks rather than at 1 week (P<0.05). Microvessel density and vascular endothelial growth factor showed significant linear correlations with hepatic blood flow, permeability of capillary vessel surface and hepatic arterial index, but not with hepatic blood volume or mean transit time.
CONCLUSION: The CT perfusion technique demonstrated early changes of hepatic hemodynamics in this tumor model as proof-of-concept for early hepatocellular carcinoma detection in humans.

BACKGROUND: Micro-RNAs (miRNAs) are small, non-coding RNA species considered to fine-tune basic cellular functions by modulating target gene translation and/or mRNA stability. A common G/C polymorphism (rs2910164) in the precursor (pre-) miR-146a gene engaged in NF-κB signaling and apoptosis pathways has been reported to modulate the genetic risk of hepatocellular carcinoma by increased G-allelic production of mature miR-146a. We investigated rs2910164 in a large European-based cholangiocarcinoma (CCA) cohort.
METHODS: We recruited 182 CCA patients and 350 controls in three academic medical centers. Genotyping for rs2910164 was performed by PCR-based assays with 5-nuclease and fluorescence detection. Genotype frequencies were tested for consistency with the Hardy-Weinberg equilibrium using an exact test; allelic and genotypic differences between the patients and controls were assessed by the Chi-square test and Armitage s trend test. Exploratory subgroup analyses included gender, tumor localization (extra- versus intrahepatic CCA) and early-onset CCA.
RESULTS: Genotype distributions were consistent with the Hardy-Weinberg equilibrium. No significant differences in either allele or genotype distributions were detected between the CCA and control groups or the respective subgroups investigated. However, there was a trend for a protective effect of the heterozygous single-nucleotide polymorphism state GC, as indicated by an underrepresentation in the CCA group in general (29% vs 35%; P=0.18) and, in particular, for extrahepatic tumor sites (26% vs 35%; OR=0.67; 95% CI, 0.43-1.02; P=0.065).
CONCLUSIONS: Our data do not support a prominent contribution of the pre-miR-146a sequence variant in the genetic predisposition to CCA. However, current studies functionally characterizing rs2910164 have proposed that distinct repertoires of target genes are addressed by genotype-specific mature miR-146a species. Given the detected trend towards a potentially protective role of GC heterozygosity, a subtle modulation of genetic CCA risk by the pre-miR-146a GC genotype may exist and should be evaluated further.

BACKGROUND: Cholangiocarcinoma, a type of malignant tumor, originates from epithelial cells of the bile duct. Perineural invasion is common path for cholangiocarcinoma metastasis, and it is highly correlated with postoperative recurrence and poor prognosis. It has been reported that muscarinic acetylcholine receptor M3 (mAChR M3) is widely expressed in digestive tract cancer, and may play an important role in the proliferation, differentiation, transformation and carcinogenesis of tumors. This study was to explore the effect of mAChR M3 on the growth of cholangiocarcinoma cells in vitro and provide a new approach to the pathogenesis and treatment of cholangiocarcinoma.
METHODS: Streptavidin-biotin complex immunohistochemistry was carried out to assess the expression of mAChR M3 in surgical specimens of cholangiocarcinomas (40 cases) and normal bile duct tissues (9), as well as to investigate nerve infiltration. The cholangiocarcinoma cells were treated with different concentrations of selective M-receptor agonist pilocarpine and M-receptor blocker atropine sulfate to induce changes in cell proliferation. The experimental data were analyzed by the Chi-square test.
RESULTS: The strongly-positive expression rate of mAChR M3 was much higher in poorly-differentiated (69%, 9/13) than in well- and moderately-differentiated cholangiocarcinomas (30%, 8/27) (χ2=5.631, P<0.05). The strongly-positive mAChR M3 expression rate in hilar cholangiocarcinoma (50%, 14/28) was higher than that in cholangiocarcinomas from the middle and lower common bile duct (25%, 3/12) (χ2=2.148, P<0.05). Cholangiocarcinomas with distant metastasis had a strongly-positive expression rate (75%, 9/12), which was much higher than those without distant metastasis (29%, 8/28) (χ2=7.410, P<0.01). The absorbance value in the pilocarpine+atropine group was significantly higher than the corresponding value in the pilocarpine group.
CONCLUSIONS: The expression of mAChR M3 is influenced by the extent of differentiation, distant metastasis and the site of cholangiocarcinoma. It also plays a key role in the proliferation and metastasis of cholangiocarcinoma.

BACKGROUND: Although invasive ductal adenocarcinoma of the pancreas (PDAC) manifests as a relatively uniform histomorphological feature of the pancreatobiliary type, it may be complicated by metaplastic changes and heterogeneous gastric and intestinal elements. This study aimed to investigate the complication rate and clinicopathological significance of such heterogeneous elements.
METHODS: Fifty-nine patients who underwent resection of PDAC were examined in this study. Immunohistochemically, tumors showing high expression (>25%) of the intestinal-type (INT) marker CDX2 were classified as PDAC with INT. Those with high expression (>25%) of the gastric-type (GAS) marker MUC5AC were classified as PDAC with GAS, while those with high expression of both markers were classified as PDAC with INT/GAS. These patients were compared with those with PDAC of the negative group in which neither markers was highly expressed to examine their clinicopathological significance.
RESULTS: In the 59 patients, 31 (52.5%) showed high CDX2 or MUC5AC expression. Twenty-eight patients (47.5%) belonged to a negative group, 11 (18.6%) to a PDAC with INT group, 15 (25.4%) to a PDAC with GAS group, and 5 (8.5%) to a PDAC with INT/GAS group. No significant differences were observed for age, gender, size, localization, T classification, or prognosis among the four groups. Although the PDAC with GAS group had well differentiated types significantly more than the other groups, the rate of lymph node metastasis in this group was significantly higher (PDAC with GAS: 73%; other groups: 36%).
CONCLUSION: Complications with heterogeneous elements are not uncommon in PDAC, and this should be considered during the diagnosis and treatment of PDAC along with histogenesis of the disease.

BACKGROUND: The piggy-back caval anastomosis technique is widely used in orthotopic liver transplantation although it carries an increased risk of complications, including outflow obstruction and Budd-Chiari syndrome. The aim of this study is to clarify the anatomy and variations of hepatic veins (HVs) draining into the inferior vena cava (IVC), and to classify the surgical techniques of piggy-back liver transplantation (PBLT) based on the anatomy of HVs which can reduce the occurrence of complications.

METHODS: PBLT was performed in 248 consecutive cases at our hospital from January 2004 to August 2011. The anatomy of recipients HVs was determined when removing the native diseased livers. Both anatomy of HVs and short HVs draining into the IVC were recorded. These data were collected and analyzed.

RESULTS: We classified anatomic variations of HVs in the 248 livers into five types according to the way of drainage into the IVC: type I (trunk type of left and middle HVs), 142 (57.3%) patients; type II (trunk type of right and middle HVs), 54 (21.8%); type III (trunk type of left, middle and right HVs), 14 (5.6%); type IV (non-trunk type of left, middle and right HVs), of which, type IVa, 16 (6.5%), in the same horizontal plane; type IVb, 18 (7.3%), in different horizontal planes; and type V (segment type), 4 (1.6%). The patients whose HVs anatomy belonged to types I, II and III underwent classical piggy-back liver transplantation. Type IVa patients had classical PBLT via HV venoplasty prior to piggy-back anastomosis, while type IVb patients and type V patients could only have modified PBLT.

CONCLUSION: This study demonstrates that HVs can be classified according to the anatomy of their drainage into the IVC and we can use this classification to choose the best operative approach to PBLT.

ABSTRACT: Terlipressin has been shown to improve both pulmonary and systemic hemodynamics in stable cirrhotic patients with pulmonary hypertension, whereas other vasoconstrictors may cause pulmonary pressures to deteriorate. We investigated the pulmonary and systemic hemodynamic effects of the first terlipressin dose (2 mg) in 7 cirrhotic patients with PH presenting with variceal bleeding (n=4) or hepatorenal syndrome (n=3). Terlipressin decreased pulmonary vascular resistance (158.8±8.9 vs 186.5±13.9 dynes • sec • cm-5; P=0.003) together with an increase in systemic vascular resistance (2143±126 vs 1643±126 dynes • sec • cm-5; P<0.001). Terlipressin should be the vasoconstrictor treatment of choice when patients present with variceal bleeding or HRS.

Living donor liver right lobe transplantation using donors with variation of the right sectorial portal vein is considered a challenging procedure in terms of the donor(s safety and the complexity of reconstruction in the recipient. We describe an innovative technique to reconstruct double portal vein orifices via a deceased donor iliac vein graft.?The postoperative course of the recipient was uneventful. Doppler ultrasound on the fourth postoperative month revealed equivalent flow in both portal vein branches. Reconstruction of double right portal vein branches using a cryopreserved iliac vein is a valuable technique for utilizing right lobe grafts with challenging portal vein anatomy.

Significant intraoperative bleeding and injuries to vital structures in the liver remnant can occur during liver parenchymal transection using the conventional clamp-crushing technique. We performed liver resection on 242 patients using a mosquito clamp-crushing technique combined with a self-assembled saline-linked diathermy for liver parenchymal transection. The mean blood loss was 215 mL (range 20-1100). There was no damage to the vital structures in the liver remnant. The mean liver transection time was 53 minutes (range 15-125). Our technique has the advantages of reducing blood loss, exposing vessles clearly and being simple, cheap and efficacious.