Revisiting the role of pathological analysis in transarterial chemoembolization-treated hepatocellular carcinoma after transplantation.

AIM: To define the histopathological features predictive of post-transplant hepatocellular carcinoma (HCC) recurrence after transarterial chemoembolization, applicable for recipient risk stratification. METHODS: We retrospectively reviewed the specimens of all suspicious nodules (total 275) from 101 consecutive liver transplant recipients which came to our Pathology Unit over a 6-year period. All nodules were sampled and analyzed, and follow-up data were collected. We finally considered 11 histological variables for each patient: total number of nodules, number of viable nodules, size of the major nodule, size of the major viable nodule, occurrence of microscopic vascular invasion, maximum Edmondson's grade, clear cell/sarcomatous changes, and the residual neoplastic volume. Survival data were computed by means of the Kaplan-Meier procedure and analyzed by means of the Cox proportional hazards model. The multivariate linear regression and a k-means cluster analysis were also used in order to compute the standardized histological score. RESULTS: The total number of nodules, the residual neoplastic volume (the total volume of all evaluated nodules minus the necrotic portion) and the microvascular invasion entered the Cox multivariate hazard model with HCC recurrence as dependent variable. The histological score was therefore computed and a cluster analysis sorted recipients into 3 risk groups, with 3.3%, 18.5% and 53.8% respectively of tumor recurrence rates and 1.6%, 11.1% and 38.5% of tumor-related mortality respectively at the end of follow-up. CONCLUSION: The histological score allows a reliable stratification of HCC recurrence risk, especially in those recipients found out to be beyond the Milan criteria after orthotopic liver transplantation (OLT).

The meaning of tissue and serum HCV RNA quantitation in hepatitis C recurrence after liver transplantation: a retrospective study.

BACKGROUND: While the role of serum HCV RNA quantitation in hepatitis C virus recurrence after liver transplantation is well established, the meaning of HCV RNA tissue quantitation is largely unclear, and no correlations with recipient outcome have been investigated yet. AIMS: To assess the predictive value, and a possible prognostic role, of tissue and serum HCV RNA in first post-transplant biopsies. METHODS: We retrospectively reviewed the first post-transplant biopsies of 83 recipients. Tissue and serum HCV RNA was quantitated by RT-PCR, and compared with serum, clinical and histological data. RESULTS: HCV RNA quantitation allowed us to categorise recipients into three different risk groups: (1) tissue HCV RNA ≤ 1.5 IU/ng with any serum HCV RNA; (2) tissue HCV RNA>1.5 IU/ng and serum HCV RNA < 40 × 10(6)copies/mL; (3) tissue HCV RNA>1.5 IU/ng and serum HCV RNA ≥ 40 × 10(6)copies/mL. Hepatitis C virus recurrence rates in the three groups were 68%, 91% and 100% (P=0.004); hepatitis C virus-related mortality was 0%, 14% and 45% respectively (P<0.001). CONCLUSIONS: This preliminary study on serum and tissue HCV RNA quantitation allows recipient "stratification" in prognostic groups, which could be applicable in the future for timely antiviral treatment and/or immunosuppression modulation.

Studies on immunoproteasome in human liver. Part I: absence in fetuses, presence in normal subjects, and increased levels in chronic active hepatitis and cirrhosis.

Despite the central role of proteasomes in relevant physiological pathways and pathological processes, this topic is unexpectedly largely unexplored in human liver. Here we present data on the presence of proteasome and immunoproteasome in human livers from normal adults, fetuses and patients affected by major hepatic diseases such as cirrhosis and chronic active hepatitis. Immunohistochemistry for constitutive (alpha4 and beta1) and inducible (LMP2 and LMP7) proteasome subunits, and for the PA28alphabeta regulator, was performed in liver samples from 38 normal subjects, 6 fetuses, 2 pediatric cases, and 19 pathological cases (10 chronic active hepatitis and 9 cirrhosis). The immunohistochemical data have been validated and quantified by Western blotting analysis. The most striking result we found was the concomitant presence in hepatocyte cytoplasm of all healthy subjects, including the pediatric cases, of constitutive proteasome and immunoproteasome subunits, as well as PA28alphabeta. At variance, immunoproteasome was not present in hepatocytes from fetuses, while a strong cytoplasmic and nuclear positivity for LMP2 and LMP7 was found in pathological samples, directly correlated to the histopathological grade of inflammation. At variance from other organs such as the brain, immunoproteasome is present in livers from normal adult and pediatric cases, in apparent absence of pathological processes, suggesting the presence of a peculiar regulation of the proteasome/immunoproteasome system, likely related to the physiological stimuli derived from the gut microbiota after birth. Other inflammatory stimuli contribute in inducing high levels of immunoproteasome in pathological conditions, where its role deserve further attention.

Preoperative prediction of hepatocellular carcinoma tumour grade and micro-vascular invasion by means of artificial neural network: a pilot study.

BACKGROUND & AIMS: Hepatocellular carcinoma (HCC) prognosis strongly depends upon nuclear grade and the presence of microscopic vascular invasion (MVI). The aim of this study was to develop an artificial neural network (ANN) that is able to predict tumour grade and MVI on the basis of non-invasive variables. METHODS: Clinical, radiological, and histological data from 250 cirrhotic patients resected (n=200) or transplanted (n=50) for HCC were analyzed. ANN and logistic regression models were built on a training group of 175 randomly chosen patients and tested on the remaining testing group of 75. Receiver operating characteristics curve (ROC) and k-statistics were used to analyze model accuracy in the prediction of the final histological assessment of tumour grade (G1-G2 vs. G3-G4) and MVI (absent vs. present). RESULTS: Pathologic examination showed G3-G4 in 69.6% of cases and MVI in 74.4%. Preoperative serum alpha-fetoprotein (AFP), tumour number, size, and volume were related to tumour grade and MVI (p<0.05) and were used for ANN building, whereas, tumour number did not enter into the logistic models. In the training group, ANN area under ROC curves (AUC) for tumour grade and MVI prediction were 0.94 and 0.92, both higher (p<0.001) than those of logistic models (0.85 for both). In the testing group, ANN correctly identified 93.3% of tumour grades (k=0.81) and 91% of MVI (k=0.73). Logistic models correctly identified 81% of tumour grades (k=0.55) and 85% of MVI (k=0.57). CONCLUSION: ANN identifies HCC tumour grades and MVI on the basis of preoperative variables more accurately than the conventional linear model and should be used for tailoring clinical management.

Predictive value of frozen-section analysis in the histological assessment of steatosis before liver transplantation.

Histological quality assessment of donated livers is a key factor for extending the cadaveric donor pool for liver transplantation. We retrospectively compared frozen-section analysis with routine histological permanent slides and the outcomes of grafts in liver biopsies from 294 candidate donors. The kappa concordance coefficient of agreement between frozen-section analysis and routine histological analysis was very good for macrosteatosis (kappa = 0.934), microsteatosis (kappa = 0.828), and total steatosis (kappa = 0.814). The correlation between the mean amounts of macrosteatosis, microsteatosis, and total steatosis in frozen and permanent sections was also significant (P < 0.001, Spearman's test). Macrosteatosis and microsteatosis were overestimated to >30% in 4 of 32 cases (12.5%) and in 23 of 62 cases (37.1%), respectively. The only 2 histological parameters of frozen sections able to predict graft dysfunction within 7 days of transplantation were macrosteatosis and total steatosis (P = 0.018 and P = 0.015, respectively, Mann-Whitney test). None of the other histopathological features evaluated in frozen sections, including portal inflammation, lobular necrosis, myointimal thickening, biliocyte regression, cholestasis, hepatocellular polymorphism, lipofuscin storage, and fibrous septa, were significantly correlated with the graft outcome. The frozen-section histological evaluation of biopsies from cadaveric liver donors is an accurate, time-effective, and predictive method for the assessment of graft suitability.

EML4-ALK rearrangement in non-small cell lung cancer and non-tumor lung tissues.

A fusion gene, echinoderm microtubule associated protein like 4-anaplastic lymphoma kinase (EML4-ALK), with transforming activity has recently been identified in a subset of non-small cell lung cancer (NSCLC), but its pathogenetic, diagnostic, and therapeutic roles remain unclear. Both frequency and type of EML4-ALK transcripts were investigated by reverse transcription PCR in 120 frozen NSCLC specimens from Italy and Spain; non-neoplastic lung tissues taken far from the tumor were used as controls. In cases carrying the fusion transcript, we determined EML4-ALK gene and protein levels using fluorescence in situ hybridization, Western blotting, and immunoprecipitation. We also analyzed ALK protein levels in paraffin samples from 662 NSCLC specimens, including the 120 cases investigated in the molecular studies. EML4-ALK transcripts (variants 1 and 3) were detected in 9 of 120 NSCLC samples but were not specific for NSCLC since they were also found in non-cancerous lung tissues taken far from the tumor. Notably, no transcripts were detected in matching tumor samples from these patients. Fluorescence in situ hybridization analysis of cases expressing EML4-ALK transcripts showed that only a minority of cells harbored the EML4-ALK gene. None of these cases was found to express the EML4-ALK protein as examined by immunohistochemistry, Western blotting, and immunoprecipitation. The EML4-ALK transcript cannot be regarded as a specific diagnostic tool for NSCLC. Our results show therefore that the causal role and value of EML4-ALK as a therapeutic target remain to be defined.

Blood monitoring of granzyme B and perforin expression after intestinal transplantation: considerations on clinical relevance.

BACKGROUND: The use of biomarkers for rejection monitoring represents a major goal in intestinal transplantation. We analyzed the blood expression of Granzyme B (GB) and Perforin (PF) in the following pathological conditions after intestinal transplantation: acute rejection (AR), Epstein-Barr virus (EBV) and cytomegalovirus (CMV) infection, and posttransplant lymphoproliferative disease (PTLD). The diagnostic accuracy and the clinical utility of these tests are finally discussed. METHODS: GB and PF levels were measured by real time polymerase chain reaction on peripheral blood samples from 32 intestinal recipients. Blood samples (n=494) after comparison of clinical, histological, and microbiological data were assigned to the following groups: normal (n=307), AR (n=30), EBV infection (n=107), CMV infection (n=25), and PTLD (n=25). RESULTS: Mean levels of GB and PF in the AR (GB=279.7; PF=256.7), PTLD (GB=199; PF=185.9), EBV (GB=133.2; PF=143.7), and CMV (GB=151.3; PF=144) groups were significantly higher than in the normal group (GB=100.1; PF=101.1) (all P<0.05, except for PF in CMV infection). The best accuracy was obtained for the diagnosis of AR with sensitivity and specificity of 80% and 79% for GB and 70% and 79% for PF, respectively. The area under the receiver-operator characteristics curve was 0.87 for GB and 0.82 for PF. CONCLUSIONS: GB and PF are diagnostic molecular markers of AR. GB and PF blood levels are also increased in case of viral infections or PTLD. Serial blood testing for GB and PF might be predictive of early intestinal graft dysfunction and should be interpreted in the context of the histological and virological analyses.

11C-choline positron emission tomography/computerized tomography for preoperative lymph-node staging in intermediate-risk and high-risk prostate cancer: comparison with clinical staging nomograms.

BACKGROUND: Conventional imaging (CI) techniques are inadequate for lymph node (LN) staging in prostate cancer (PCa). OBJECTIVES: To assess the accuracy of (11)C-Choline positron emission tomography/computerized tomography (PET/CT) for LN staging in intermediate-risk and high-risk PCa and to compare it with two currently used nomograms. DESIGN, SETTING, AND PARTICIPANTS: From January 2007 to September 2007, 57 PCa patients at intermediate risk (n=27) or high risk (n=30) were enrolled at two academic centres. All patients underwent preoperative PET/CT and radical prostatectomy with extended pelvic LN dissection (PLND). Risk of LN metastasis (LNM) was assessed using available nomograms. MEASUREMENTS: Sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and number of correctly recognized cases for LNM detection at PET/CT were assessed. The accuracy of PET/CT for LNM detection was compared with the accuracy of nomograms for LNM prediction by using receiver operating characteristic (ROC) analysis. RESULTS AND LIMITATIONS: Fifteen patients (26%) had LNMs, and a total of 41 LNMs were identified. On a patient analysis, sensitivity, specificity, PPV, NPV, and number of correctly recognized cases at PET/CT were 60.0%, 97.6%, 90.0%, 87.2%, and 87.7% while, on node analysis, these numbers were 41.4%, 99.8%, 94.4%, 97.2%, and 97.1%. The mean diameter (in mm) of the metastatic deposit of true-positive LNs was significantly higher than that of false-negative LNs (9.2 vs 4.2; p=0.001). PET/CT showed higher specificity and accuracy than the nomograms; however, in pairwise comparison, the areas under the curve (AUCs) were not statistically different (all p values >0.05). CONCLUSIONS: In patients with intermediate-risk and high-risk PCa, (11)C-Choline PET/CT has quite a low sensitivity for LNM detection but performed better than clinical nomograms, with equal sensitivity and better specificity.

Real time RT-PCR approach for the evaluation of ERBB2 overexpression in breast cancer archival samples: a comparative study with FISH, SISH, and immunohistochemistry.

We tested the reliability of real time reverse transcription polymerase chain reactions as an alternative method for the assessment of ERBB2 status in paraffin-embedded tissues of 83 patients with breast cancer and 20 non-neoplastic controls. PCR was also compared with the immunohistochemical (IHC) HercepTest score and with fluorescence (FISH) and silver (SISH) in-situ hybridization, in 42 selected cases. ERBB2 mRNA was overexpressed in 26/83 (31%) breast cancer samples, using a cutoff calculated as the mean value of the controls plus 3 SD or with the receiver operating curve. The PCR test showed a 96% sensitivity and a 100% specificity when compared with FISH, with an area under the receiver operating curve of 98.4%. Overexpression of ERBB2 at PCR was also significantly correlated with amplification in FISH (P<0.001, Mann-Whitney test) and in SISH (P<0.001, Mann-Whitney test), and with the IHC HercepTest scores 2 or 3 (P<0.001, Spearman rank correlation). FISH, SISH, and IHC were also compared with each other. ERBB2 amplification in FISH significantly correlated with that in SISH (P=0.002, chi test with a concordance of the 87%), but not with IHC HercepTest scores (P=0.214, chi test). Real time PCR is a reliable and cost-effective method for the assessment of ERBB2 status in archival breast cancer samples, compared with FISH. Its introduction in routine diagnostic pathology practice is feasible even if it requires amendments to the current clinical oncology protocols.

Diagnostic role of circulating free plasma DNA detection in patients with localized prostate cancer.

To analyze the potential diagnostic relevance of free plasma DNA (FPDNA), we enrolled 64 patients with localized prostate cancer (CaP). FPDNA was quantified by real-time polymerase chain reaction assessment of the HTERT gene in blood samples from 64 patients with CaP and 45 healthy males. Methylation of the GSTP1 gene was used to confirm the neoplastic origin of FPDNA in selected cases. The mean +/- SD levels of FPDNA were higher in patients with CaP (15.4 +/- 10.9 ng/mL) than in control subjects (5.5 +/- 3.5 ng/mL; P <.001). By using the best cutoff value, the sensitivity of the test was 80%, the specificity was 82%, the area under the receiver operating characteristic curve, 0.881. High FPDNA values were significantly associated with pathologic T3 stage (P = . 035). Methylation of the GSTP1 gene was found in 4 (25%) of 16 FPDNA samples and 15 (94%) of 16 tissue samples. Quantification of FPDNA discriminates between patients with CaP and healthy subjects and correlates with pathologic tumor stage. FPDNA is a candidate biomarker for early diagnosis and monitoring of CaP.

Tissue hepatitis C virus RNA quantification and protein expression help identify early hepatitis C virus recurrence after liver transplantation.

We compared tissue hepatitis C virus (HCV) RNA polymerase chain reaction quantification and HCV immunohistochemistry (IHC) to histology in biopsy tissues in order to differentiate between acute rejection and HCV hepatitis recurrence early after orthotopic liver transplantation (OLT). We analyzed the first biopsy performed because of alteration of serum aminotransferases in 65 consecutive OLT patients with HCV genotype 1b. In the histological analysis, we quantified the portal tracts, Councilman bodies, Councilman body/portal tract (CP) ratio, steatosis, and Knodell and Ishak scores. The 52 patients (80%) with histological HCV recurrence [recurrence-positive (Rec+)] were separated from the 6 (9%) with acute rejection and the 7 (11%) with undetermined pathological features [recurrence-negative (Rec-)]. HCV RNA strongly correlated with HCV IHC, regardless of the histological diagnosis (P < 0.001). Both HCV RNA and HCV IHC were significantly associated with CP ratio (P = 0.041 and P = 0.008). No statistical correlation was found between HCV RNA, HCV IHC, and the other histopathologic features or the hepatitis scores. HCV RNA, HCV IHC, and CP ratio were the only variables able to discriminate between Rec+ and Rec- patients (Mann-Whitney test P < 0.001, P < 0.001, P = 0.014). In conclusion, a combined evaluation of histology, tissue HCV RNA, and HCV IHC significantly discriminated between OLT patients with or without HCV recurrence.

Ex vivo HR-MAS magnetic resonance spectroscopy of normal and malignant human renal tissues.

UNLABELLED: The aim of the present study was to examine the metabolic profile of normal and tumoral renal tissues by ex vivo high resolution magic angle spinning magnetic resonance spectroscopy (HR-MAS MRS). PATIENTS AND METHODS: Five patients, three affected by clear cell renal cell carcinoma (RCC) and two by papillary RCC, were examined. A radical nephrectomy was performed in each. In all patients, fresh tissue samples taken from normal cortex, normal medulla and tumor were collected and analyzed by mono-dimensional HR-MAS MRS. RESULTS: The spectra of human normal cortex and medulla showed the presence of differently distributed organic osmolytes as markers of a physiological renal condition. The marked decrease or disappearance of these metabolites and the high lipid content (triglycerides and cholesteryl esters) is typical of clear cell RCC, while papillary RCC are characterized by the absence of lipids and very high amounts of taurine. CONCLUSION: This paper demonstrates that ex vivo HR-MAS MRS is a viable and powerful means of probing for molecular information in human normal and tumoral renal tissues. This research will constitute the basis for a biochemical classification of renal neoplastic pathologies, especially for RCCs, which can be thus evaluated by in vivo MRS for clinical purposes. Moreover, these data may contribute to a better knowledge of the molecular processes for the basis of the onset of renal carcinogenesis.

Application of a fluorescent PCR method for molecular diagnosis of posttransplant lymphoproliferative disorders on routine tissue sections.

Molecular detection of monoclonality can play an important role in the diagnosis of posttransplantation lymphoproliferative disorders (PTLD). To permit accurate molecular diagnosis of PTLD even on very small amounts of DNA extracted from routinely embedded histologic material, we adapted a commercially available PCR protocol (for FR-1, -2 and -3 regions), originally designed for use on fresh/frozen samples. We applied this approach on routine biopsy/surgical material of 10 PTLD (from nine patients). All three FR regions were always amplified, indicating that the extracted DNA was of medium quality. All five PTLD morphologically classified as lymphomas were monoclonal in at least one FR region. Thus, using the WHO histologic, immunohistochemical, and clinical criteria as the reference standard, the approach provided 100% sensitivity for detection of monoclonal malignancies, supporting the validity of the method. Of five specimens classified morphologically as polymorphic PTLD, three displayed a solitary IgH gene rearrangement peak, consistent with the presence of a monoclonal B-cell population (ie, monoclonal polymorphic PTLD). This rapid and straightforward procedure, which allows identification of a wide range of IgH rearrangements, could facilitate molecular analysis of PTLD in routine practice, while limiting consumption of valuable diagnostic material.

Genomic allelotyping for distinction of recurrent and de novo hepatocellular carcinoma after orthotopic liver transplantation.

Distinction between recurrent and de novo hepatocellular carcinoma (HCC) after orthotopic liver transplantation (OLT) bears important clinical and therapeutic implications. Techniques for molecular profiling of clinically suspected de novo and recurrent HCC are required since the histological/clinical discrimination of donor vs. recipient tumor origin is difficult. Multiple PCR amplification of 16 highly polymorphic short tandem repeat (STR) DNA sequences (routinely used for paternity and forensic assays) was applied in two patients who developed a second HCC after OLT. In both patients the technique provided reliable evidence that the two second HCC were recurrences of the primary tumor. Multiple STR genetic allelotyping is an effective tool for clear-cut discrimination of donor/recipient origin of a second HCC after OLT. Its application could be of great therapeutic relevance for such OLT patients.

Partial necrosis on hepatocellular carcinoma nodules facilitates tumor recurrence after liver transplantation.

BACKGROUND: The presence of partial necrosis in hepatocellular carcinoma (HCC) nodules is a common histologic finding after liver transplantation, but its correlation with tumor recurrence has never been investigated. METHODS: we retrospectively reviewed the outcome of 54 patients with a single histologically proven HCC after liver transplantation. All cases had a survival of more than 6 months, and patients treated preoperatively had a transarterial chemoembolization (TACE) procedure. Since 1996, our center has applied the Milan criteria. Correlations between tumor recurrences and clinicopathologic variables, including the presence of partial necrosis, were performed. Etiologic factors for HCC partial necrosis were also investigated. RESULTS: Sixteen of 54 (29.6%) HCC nodules presented partial necrosis, and 4 (25%) of them developed HCC recurrence compared with 1 of 38 (2.6%) cases without this histologic finding (P<0.05). Partial necrosis was related to TACE procedure (P<0.05), patient age less than 50 years (P<0.05), and tumor diameter greater than 2 cm (P<0.05). Multivariate analysis showed only TACE as an independent variable. The other variables related to the five (9.3%) tumor recurrences were HCC diameter greater than 2 cm (P<0.05), year of liver transplantation before 1996 (P<0.05), and the presence of satellite nodules (P<0.05). The Cox regression analysis showed the presence of partial necrosis as an independent variable related to tumor recurrence. The analysis of the recurrence-free survival confirmed the results of the recurrence rate. CONCLUSION: Partial necrosis was a risk factor for tumor recurrence after liver transplantation. Patients and procedures should be selected while also bearing in mind the side-effect of incomplete necrosis of the nodules.

A histopathologic screening method for rational use of organs from prostate-specific antigen-positive multiorgan donors: the Italian Emilia-Romagna Region experience.

Reduction of waiting-list length requires extension of the organ-donor pool to elderly males bearing an higher risk of prostate cancer incidence. Prostate-cancer screening in organ donors is currently based on prostate-specific antigen (PSA) assays (total PSA and free/total PSA). However, the specificity of these assays is restricted, limiting risk-benefit analysis. Since 2001, 33 multiorgan donor candidates presenting within Emilia-Romagna (Italy) with suspect ultrasonography or abnormal PSA values were submitted to a histopathologic screening method of the entire prostate based on extemporary frozen-section analysis (maximum 1 hour) of over 50% of the organ at 0.1 mm cutting levels. Extemporary diagnosis of adenocarcinoma was made in 12 (36%) cases, corresponding to 4.5% of the male candidates aged more than 50 years in the donor pool. In all cases, the final diagnosis confirmed the extemporary analysis. As well as maximizing safety, this novel approach should permit more refined risk-benefit analysis.

Liver transplantation for hepatocellular carcinoma: further considerations on selection criteria.

The selection criteria in liver transplantation for HCC are a matter of debate. We reviewed our series, comparing two periods: before and after 1996, when we started to apply the Milan criteria. The study population was composed of patients with a preoperative diagnosis of HCC, confirmed by the pathological report and with a survival of >1 year. Preoperative staging as revealed by radiological imagining was distinguished from postoperative data, including the variable of tumor volume. After 1996 tumor recurrences significantly decreased (6 out of 15 cases, 40% vs. 3 out of 48, 6.3%, P < .005) and 5-year patient survival improved (42% vs. 83%, P < .005). Not meeting the Milan criteria was significantly related to higher recurrence rate (37.5% vs. 12.7%, P < .05) and to lower 5-year patient survival (38% vs. 78%, P < .005%) in the preoperative analysis, but not in the postoperative one. The alfa-fetoprotein level of more than 30 ng/dL and the preoperative tumor volume of more than 28 cm3 predicted HCC recurrences in the univariate and mutivariate analysis (P < .005 and P < .05, respectively). The ROC curve showed a linear correlation between preoperative tumor volume and HCC recurrence. Milan criteria significantly reduced tumor recurrences after liver transplantation, improving long-term survival. In conclusion, the efficacy of tumor selection criteria must be analyzed with the use of preoperative data, to avoid bias of the postoperative evaluation. Tumor volume and alfa-fetoprotein level may improve the selection of patients.

Predictive value of biological markers for hepatocellular carcinoma patients treated with orthotopic liver transplantation.

PURPOSE: To help stratify candidates with hepatocellular carcinoma (HCC) for orthotopic liver transplantation (OLT), biomarkers are needed that are capable of predicting recurrence of disease (ROD). We investigated the prognostic role in this setting of immunohistochemical markers reported previously to predict poor prognosis in HCC patients treated with resection. EXPERIMENTAL DESIGN: Eighty-three patients with HCC who underwent OLT between 1987 and 2001 with a minimum clinical follow up of 12 months were included in this retrospective study. We analyzed immunohistochemical expression of the adhesion molecules E-cadherin and beta-catenin (membrane/nuclear localization), MIB-1 proliferative index and the cyclin-dependent kinase inhibitor p27, alongside the main clinical-pathological variables. RESULTS: At univariate analysis, vascular thrombosis, high MIB-1 index, lower membrane expression of E-cadherin and beta-catenin, and nuclear beta-catenin localization were associated with ROD. At multivariate analysis, only MIB-1 index, low equal E-cadherin (with respect to non-neoplastic surrounding tissue), and nuclear beta-catenin appeared as independent predictors of ROD. The logistic regression analysis model indicated that detection of any one parameter was associated with at least 88% estimated risk of ROD (up to 99% for all three). CONCLUSIONS: We propose these three molecular parameters as an additional tool for rational selection of OLT candidates among HCC patients (stratification according to the risk of ROD might help provide a similar life expectancy for cirrhotic candidates with and without HCC).

Helicobacter species sequences in liver samples from patients with and without hepatocellular carcinoma.

AIM: Only a minority of patients carrying a defined viral aetiologic agent develop cirrhosis and ultimately hepatocellular carcinoma (HCC), the mechanism underlying the worsening is still undefined. Experimental infection by Helicobacter hepaticus in mice causes chronic hepatitis and HCC and recently, more Helicobacter species (Helicobacter spp.) have been detected in the liver of patients suffering from cholestatic diseases and HCC arising from non-cirrhotic liver. We investigated whether Helicobacter spp. sequences could be detected in the liver of patients with cirrhosis and HCC compared to subjects with metastasis to liver from colon cancer. METHODS: Twenty-three liver samples from patients operated upon for HCC superimposed on hepatitis C virus (HCV)-related cirrhosis and 6 from patients with resected metastases from colorectal cancer, were tested by polymerase chain reaction for presence of genomic 16S rRNA of Helicobacter genus using specific primers. DNA sequencing and cag A gene analysis were also performed. RESULTS: Genomic sequences of Helicobacter spp. were found in 17 of 20 (85%) liver samples from patients with HCC and in 2 of 6 samples from patients with liver metastasis. In three samples of the first group the result was uncertain. H pylori was revealed in 16 out of 17 positive samples and Helicobacter pullorum in the other. CONCLUSION: Helicobacter spp., carcinogenic in mice, were found at a higher frequency in the liver of patients with HCV-related cirrhosis and HCC than those in patients without primary liver disease.