Elevated soluble TNFα levels and upregulated TNFα mRNA expression in purified peripheral blood monocyte subsets associated with high-grade hepatocellular carcinoma.

BACKGROUND: Chronic inflammation is involved in the initiation and progression of various cancers, including liver cancer. The current study focuses on the characterization of the peripheral immune response in hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA) patients, before and after surgical procedure, in order to assess the effect of tumor resection in the immune system homeostasis and to determine possible prognostic factors associated with high-grade tumors. We developed a whole-blood assay to monitor immune alterations and functional competence of peripheral monocytes in a group of 10 healthy individuals (HG), in 20 HCC patients and 8 CCA patients, by multi-color flow cytometry, qRT-PCR, and ELISA techniques. RESULTS: The qRT-PCR analysis showed an upregulation of TNFα expression by classical and intermediate monocytes purified from HCC patients presenting tumors in grade G3-G4 as compared to G1-G2 HCC patients. Moreover, ELISA assay confirmed elevated serum levels of TNFα in G3-G4 compared to G1-G2 HCC patients. A significant decrease of circulating non-classical monocytes was detected in both CCA and HCC patients before and after surgical procedure. In addition, a functional defect in circulating classical and intermediate monocytes was observed in both groups of cancer patients when compared to the HG, with partial recovery after the surgical intervention. CONCLUSIONS: This integrated analysis permitted the identification of altered functional competence of monocyte subsets in CCA and HCC patients. In addition, our results point to a potential role of TNFα as a prognostic peripheral biomarker in HCC patients, indicating the presence of high-grade tumors that should be further validated.

Functional and Phenotypic Characterization of Tumor-Infiltrating Leukocyte Subsets and Their Contribution to the Pathogenesis of Hepatocellular Carcinoma and Cholangiocarcinoma.

Hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA) represent the most common primary liver malignancies whose outcome is influenced by the immune response. In the present study, we evaluated the tumor-infiltrating leukocyte (TIL) populations in 21 HCC patients and 8 CCA patients by flow cytometry immediately after the surgical procedure. Moreover, CD4+ T cells, CD8+ T cells, monocytes, and macrophages were purified by cell sorting for further analysis of gene expression by quantitative reverse-transcription polymerase chain reaction. Regarding tumor-infiltrating macrophages, we observed a significantly higher expression of markers associated with M2 phenotype and a higher expression of PD-L1 in patients with HCC in comparison to CCA. In addition, for HCC patients, we found a significant increase in the expression of CD200R in macrophages from tumors that were in grade G3-G4 as compared to tumors in grade G1-G2. Besides, a significantly higher frequency of tumor-infiltrating lymphocytes, CD8+CD56+ T cells, and natural killer cells was detected in HCC biopsies in comparison to CCA. In summary, this study has revealed functional and phenotypic differences in TIL cell subpopulations between CCA and HCC, as well as among different histopathological grades and tumor aggressiveness degrees, and it has provided evidence to better understand the tumor immune microenvironment of CCA and HCC.

Influence of Ischemia Time in Injury of Deep Peribiliary Glands of the Bile Ducts Graft: A Prospective Study.

The deep peribiliary glands (DPBG) are a niche of progenitor cells in the wall of the biliary duct (BD) and are the second line of multiplication when severe lesion of the epithelium occurs. Previous studies have identified DPBG injury as a cause of post-liver transplant (LT) biliary stenosis; this complication is a major cause of post-LT morbidity. The incidence of biliary stenosis in our center is high (38.1%). This study evaluates the lesion of DPBG in response to ischemia. Graft BD was collected in adult LT between August 2016-July 2017, from donation after brain death. Samples of 45 grafts were collected at 2 moments: BD1-during graft preparation and BD2-before biliary anastomosis. Histological analysis of the samples was performed and then classified according to degree of lesion (0, ≤50%, and >50%). A comparison was made between the degree of lesion and graft ischemia, graft histology, donor, and procurement variables. The DPBG lesion was more frequent in BD2 (20.9% vs 7%, P = .079). BD2 lesions with DPBG lesions had higher medians and means at all times of ischemia. The difference was greater in the warm ischemia time (0: 43.3 ± 12.53 minutes vs ≤50%: 52.4 ± 14.38 minutes, P = .068). The group of BD1 with DPBG lesion presented superior median cold ischemia time (CIT). In the analysis of the remaining variables there were also no statistically significant differences. We concluded that during the period of CIT there is already lesion of the DPBG, which increases after reperfusion of the graft, in greater association with longer warm ischemia time.

Sinusoidal dilation increases the risk of complications in hepatectomy for CRCLM - Protective effect of bevacizumab and diabetes mellitus, serum gamma-glutamyltranspeptidase as predictive factor.

INTRODUCTION: Advances in neoadjuvant chemotherapy (NCT) have allowed surgical treatment in otherwise unresectable patients with colorectal liver metastases (CRLM). It is well known that NCT induces liver lesions such as sinusoidal obstruction syndrome (SOS) and steatohepatitis (SH). However, whether it affects postoperative morbidity remains controversial. The aim of this study was both to evaluate the impact of NCT on liver parenchyma and postoperative morbidity, and to identify preoperative predictive markers for liver injury. PATIENTS AND METHODS: Among 140 patients undergoing liver resection for CRLM between 2010 and 2013, 70 underwent systemic NCT. Liver function tests, pathology, postoperative morbidity and mortality were compared between the two groups. RESULTS: Univariate analysis revealed NCT as a cause of sinusoidal dilation (p = 0.09), peliosis (p = 0.028) and moderate and severe SOS (p = 0.004) and bevacizumab as a protective agent against moderate and severe SOS (p = 0.045). Diabetic patients were identified as having a lower incidence of sinusoidal dilation (p = 0.034) and a higher incidence of steatosis (p = 0.003). Multivariate analysis confirmed sinusoidal dilation as an independent cause for morbidity (p = 0.02) and liver-specific complications (p = 0.016). Preoperative level of GGT was identified as predictive factor for moderate and severe SOS and peliosis (p < 0.001 and p = 0.004, respectively). CONCLUSION: The administration of NCT induces SOS-lesions, but can be partially prevented by bevacizumab and diabetes. Sinusoidal dilation is associated with increased postoperative morbidity. Preoperative GGT levels can be useful to predict the presence of SOS.

Hepatocellular carcinoma and chemotherapy: the role of p53.

BACKGROUND: Hepatocellular carcinoma (HCC) is the most common primary neoplasm of the liver. A major proportion of HCCs also present mutation of the gene that encodes p53, which confers chemoresistance. The main goal of this work is to investigate the effect of cisplatin, doxorubicin and 5-fluoruracil (5-FU) in three human HCC cell lines which differ in p53 expression. METHODS: HepG2 (expressing normal p53), HuH7 (expressing mutated p53) and Hep3B2.1-7 (not expressing p53) cell lines were cultivated in the presence of cisplatin, doxorubicin and 5-FU. Cell proliferation was evaluated by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay (MTT assay). The type of cell death and Bax and Bcl2 activation were assessed by flow cytometry. RESULTS: It was found that for all of the cell lines studied, the agent that gave the most satisfactory results was doxorubicin. 5-FU demonstrated no activity in these cell lines. CONCLUSIONS: For all the cell lines studied, doxorubicin was the most satisfactory agent. In HepG2 and HuH7 cell lines, it can activate Bax with statistical significance.

Perioperative tumor cell dissemination in patients with primary or metastatic colorectal cancer.

INTRODUCTION: Although there is general correlation between the TNM stage of colorectal cancer (CRC) and its prognosis, there is often significant variability of tumor behaviour and individual patient outcome, which is unaccounted for by pathologic factors alone. Our aim was to estimate perioperative tumor cell dissemination in patients with primary or CRC liver metastases as a possible factor influencing the outcome. METHODS: Forty patients were prospectively enrolled in the study from the year 2007 to 2008. Eighteen patients had histologically proven CRC (50% rectal, 44% colonic, 6% colonic and rectal). Sixteen patients (47%) had CRC liver metastases only. The remaining six patients who underwent colon or liver resection for benign conditions, acted as the control group. All patients with malignant pathologies had R0 resections. Blood samples were taken before the surgical incision (T0), immediately after tumor resection (T1) and at the end of the surgical intervention (T2). Data acquisition was performed using a dual-laser FACSCalibur flow cytometer. Circulating malignant cells were identified as being CD45-/cytokeratin+. RESULTS: The analysis of patients overall (CRC resection subgroup and hepatectomy subgroup) revealed that there was no statistically significant difference of the tumoral cell count in the blood per million of hematopoietic cells at T0, T1 and T2. CONCLUSIONS: This study demonstrates no differences in the detected circulating numbers of tumor cells at different stages of surgical intervention.

Resection of hepatocellular carcinoma: the effect of surgical margin and blood transfusion on long-term survival. Analysis of 209 consecutive patients.

BACKGROUND/AIMS: Certain prognostic factors affect the postoperative mortality and long-term survival of patients following hepatic resection for hepatocellular carcinoma (HCC) and may change the surgical strategy. METHODOLOGY: 209 consecutive patients underwent hepatic resection for HCC in our hospital. Seventy-three patients underwent major resection and 136 underwent minor resections. We looked for correlations between clinical, biological, surgical and pathological factors and postoperative mortality, disease-free survival and overall survival. RESULTS: The postoperative mortality rate was 7.7% (it fell to 0% in the last two years). The cumulative overall five-year survival rate was 27% and the overall disease-free survival rate was 7.3%. Multivariate analysis identified: (1) two independent prognostic factors for postoperative mortality: age and tumor size; (2) one risk factor for tumor recurrence: intraoperative blood transfusion, and (3) three independent prognostic factors for overall survival: infiltrative tumor type, surgical margin <10 mm and intraoperative blood transfusion. CONCLUSIONS: In addition to routine staging of the tumor, the preoperative evaluation of HCC patients should include tests to determine whether the tumor is infiltrative or expansive and whether it will be possible to obtain a surgical margin (>10 mm). This procedure should make it possible to propose an appropriate neoadjuvant treatment only to these patients. The prevention of intraoperative bleeding or blood transfusion should improve the disease-free and overall survival rates in HCC patients.

Treatment of tumour recurrence after resection of hepatocellular carcinoma. Analysis of 97 consecutive patients.

OBJECTIVE: To evaluate the long-term results of aggressive treatment of HCC recurrence. METHODS: Two hundred and nine consecutive patients underwent hepatic resection for HCC in our hospital. Tumour recurrence was diagnosed in 97 (51%) of the 190 patients with curative resection. Sixteen underwent hepatic resection: two right hepatectomies, one three-segmentectomy, one left hepatectomy, five two-segmentectomies, six segmental resections and one subsegmentectomy. Two patients with metastasis in the spine were submitted to a vertebral body resection. Twenty-five patients were treated with percutaneous ethanol injection or intra-arterial chemoembolization. Fifty-four patients with a poor performance status and liver function or multiple extra hepatic recurrences did not receive any treatment. RESULTS: There were no operative deaths. The postoperative mortality rate was 5.5% (one patient). The cumulative overall survival after the second resection was respectively 89%, 46% and 31% at 1, 3 and 5 years. There was a significant difference in survival between patients treated with repeat resection and those submitted to a non-surgical or conservative treatment (p<0.0001). There were no differences in operative deaths, postoperative mortality and morbidity between the first and second hepatic resection. CONCLUSIONS: Aggressive management with combined resection or loco regional therapy for intrahepatic recurrence and resection of isolated extra-hepatic recurrence may offer long-term survival in selected patients. Second liver resection for recurrence of HCC can be safely performed.