Phenotype characterization of human melanoma cells resistant to dabrafenib.

In the present study, the phenotype of melanoma cells resistant to dabrafenib (a B-RAF inhibitor) was investigated, to shed more light on melanoma resistance to B-RAF inhibition. Melanoma cells resistant to dabrafenib were generated using 3 different cell lines, A375, 397 and 624.38, all carrying B-RAFV600E, and they were characterized by cytofluorometric analysis, Ion Torrent technology, immunofluorescence and biochemistry. All dabrafenib-resistant cells showed, in addition to a re-activation of MAPK signaling, morphological changes compared to their sensitive counterparts, accompanied by an increase in CD90 (mesenchymal marker) expression and a decrease in E-cadherin (epithelial marker) expression, suggesting an epithelial-to-mesenchymal-like phenotypic transition. However, melanoma cells with TGF-ß1-induced epithelial-to-mesenchymal transition (EMT) were more sensitive to dabrafenib treatment compared to the sensitivity noted in the non-TGF­ß1­induced EMT melanoma cells, suggesting that TGF-ß1-induced EMT was not associated with dabrafenib resistance. Although dabrafenib-resistant cells exhibited increased cell motility and E-cadherin/vimentin reorganization, as expected in EMT, all of them showed unvaried E-cadherin mRNA and unchanged Snail protein levels, while Twist1 protein expression was decreased with the exception of A375 dabrafenib-resistant melanoma cells, where it was unaffected. These findings suggest a distinct active EMT-like process adopted by melanoma cells under drug exposure. Furthermore, dabrafenib-resistant cells exhibited stem cell-like features, with Oct4 translocation from the cytoplasm to peri-nuclear sites and nuclei, and increased CD20 expression. In conclusion, our data, in addition to confirming that resistance to dabrafenib is dependent on re-activation of MAPK signaling, suggest that this resistance is linked to a distinct active EMT-like process as well as stem-cell features adopted by melanoma cells.

Lead exposure and plasma mRNA expression in ERBB2 gene.

Epidemiologic data for carcinogenicity in those exposed to lead (Pb) suggests relations with cancers although the totality of the evidence is inconsistent. Alterations in the expression of ERBB receptors have been studied during the development and malignant transformation of different kinds of human tumors where they induce proliferation, angiogenesis and metastasis generation. Relevant clinical data demonstrate the role of ERBB2 receptors in the development and malignancy of human cancers. Therefore, the objective of the present investigation is to give more information on the link between plasma mRNA expression in ERBB2 gene and lead blood levels in a healthy population. Blood samples, socio­demographic, exposure and health data were obtained from 48 healthy men. Real­time polymerase chain reaction assays were performed to detect ERBB2 gene transcripts, ΔΔCt method was used to quantify gene expression. Pb blood level was assayed using high­resolution sector field inductively coupled mass spectrometry and is expressed in µg/dl. Plasma mRNA expression in ERBB2 gene was 6.44±3.07 ΔΔCt; Pb blood levels was 16.07±6.74 µg/dl. Regression analysis revealed a significant association (r2=0.5345; p<0.0001) between Pb levels and mRNA expression in ERBB2. So far, it has still not been established if the expression of ERBB2 receptors is influenced by Pb exposure. On the base of the above reported data, we believe an in vitro study might be useful, to understand the molecular mechanisms implicated.

MMP-9 overexpression is associated with intragenic hypermethylation of MMP9 gene in melanoma.

Tumor spreading is associated with the degradation of extracellular matrix proteins, mediated by the overexpression of matrix metalloproteinase 9 (MMP-9). Although, such overexpression was linked to epigenetic promoter methylation, the role of intragenic methylation was not clarified yet. Melanoma was used as tumor model to investigate the relationship between the DNA intragenic methylation ofMMP9 gene and MMP-9 overexpression at transcriptional and protein levels. Computational analysis revealed DNA hypermethylation within the intragenic CpG-2 region of MMP9 gene in melanoma samples with high MMP-9 transcript levels. In vitro validation showed that CpG-2 hotspot region was hypermethylated in the A375 melanoma cell line with highest mRNA and protein levels of MMP-9, while low methylation levels were observed in the MEWO cell line where MMP-9 was undetectable. Concordant results were demonstrated in both A2058 and M14 cell lines. This correlation may give further insights on the role of MMP-9 upregulation in melanoma.

c-Myc modulation: a key role in melanoma drug response.

Understanding molecular mechanisms involved in melanoma resistance to drugs is a big challenge. Experimental evidences suggested a correlation between mutational status in B-RAF and melanoma cell susceptibility to drugs, such as paclitaxel, doxorubicin and temozolomide, which generate an accumulation of hydrogen peroxide (H2O2) in the cells. We investigated the survival phenotype and the protein level of c-myc, a B-RAF target molecule, in melanoma cells, carrying a different mutational status in B-RAF, upon paclitaxel, doxorubicin and H2O2 treatment. For the first time, we reported c-myc modulation is critical for melanoma drug response. It appeared drug-specific and post-transcriptionally driven through PP2A; in correlation, cell pre-treatment with okadaic acid (OA), a specific PP2A inhibitor, as well as PP2A silencing of melanoma cells, was able to increase melanoma cell drug-sensitivity and c-myc protein level. This is relevant for designing efficacious therapeutic strategies in melanoma.

Ran signaling in melanoma: implications for the development of alternative therapeutic strategies.

We performed a comparative study between two human metastatic melanoma cell lines (A375 and 526), and melanocytes (FOM78) by gene expression profiling and pathway analysis, using Gene Set Enrichment Analysis (GSEA) and Ingenuity Pathway Analysis (IPA) software. Genes involved in Ran signaling were significantly over-represented (p ≤ 0.001) and up-regulated in melanoma cells. A melanoma-associated molecular pathway was identified, where Ran, Aurora Kinase A (AurkA) and TERT were up-regulated, while c-myc and PTEN were down-regulated. A consistent high Ran and AurkA gene expression was detected in about 48% and 53%, respectively, of 113 tissue samples from metastatic melanoma patients. AurkA down-regulation was observed in melanoma cells, by Ran knockdown, suggesting AurkA protein is a Ran downstream target. Furthermore, AurkA inhibition, by exposure of melanoma cells to MLN8054, a specific AurKA inhibitor, induced apoptosis in both melanoma cell lines and molecular alterations in the IPA-identified molecular pathway. These alterations differed between cell lines, with an up-regulation of c-myc protein level observed in 526 cells and a slight reduction seen in A375 cells. Moreover, Ran silencing did not affect the A375 invasive capability, while it was enhanced in 526 cells, suggesting that Ran knockdown, by AurkA down-regulation, resulted in a Ran-independent enhanced melanoma cell invasion. Finally, AurK A inhibition induced a PTEN up-regulation and its action was independent of B-RAF mutational status. These findings provide insights relevant for the development of novel therapeutic strategies as well as for a better understanding of mechanisms underlying therapy resistance in melanoma.

Molecular screening in Sicilian families with hereditary non-poliposis colorectal cancer (H.N.P.C.C.) syndrome: identification of a novel mutation in MSH2 gene.

HNPCC is an autosomal inherited cancer syndrome characterized by germinal and somatic mutations of DNA mismatch repair (MMR) genes. The inherited mutation in one allele together with an acquired defect in the other allele of an MMR gene leads to accelerate tumor progression. In this study we analyzed a cohort of 11 subjects belonging to four Sicilian families with HNPCC suspected by molecular analysis of coding regions of hMSH2 (NC_000002) and hMLH1 (NC_000003) genes. Molecular analysis has detected the presence of two mutations in gene MSH2 and one mutation in MHL1 gene. In addition, we found a novel mutation consisting in a G deletion at 914 codon of the exon 16 in the MSH2 gene. This deletion leads to a stop codon due to a frame-shift, resulting in a truncated protein. We extended genetic analysis to the other family members and the same mutation was detected in three sisters and in one of the two healthy daughters. This mutation is correlated with clinical findings revealed in genealogic tree and it represents a novel mutation responsible of HNPCC.

Molecular analysis of the APC gene in Sicilian patients with familial adenomatous polyposis (F.A.P.).

Familial adenomatous polyposis (FAP) is an autosomal dominant inherited syndrome, caused by germline mutations in the adenomatous polyposis coli (APC) suppressor gene. Patients with colorectal polyps are more likely to develop a malignant condition with poor prognosis. Typical FAP is characterized by hundreds to thousands of colorectal adenomatous polyps and by several extra-colonic manifestations; an attenuated form of polyposis (AFAP), presenting less than 100 adenomas and later onset, has been reported. In this study we have examined five Sicilian families affected by FAP syndrome, in order to provide predictive genetic testing for the affected families, as well as to contribute to mutation catalog enrichment. We have detected different APC mutations in these five pedigrees, confirming the remarkable heterogeneity of the mutational spectrum in FAP.

Emerging targeted therapies for melanoma treatment (review).

Cutaneous melanoma is an aggressive cancer with a poor prognosis for patients with advanced disease. The identification of several key molecular pathways implicated in the pathogenesis of melanoma has led to the development of novel therapies for this devastating disease. In melanoma, both the Ras/Raf/MEK/ERK (MAPK) and the PI3K/AKT (AKT) signalling pathways are constitutively activated through multiple mechanisms. Targeting various effectors of these pathways with pharmacologic inhibitors may inhibit melanoma cell growth and angiogenesis. Ongoing clinical trials provide hope to improve progression-free survival of patients with advanced melanoma. This review summarizes the most relevant studies focused on the specific action of these new molecular targeted agents. Mechanisms of resistance to therapy are also discussed.

Immunolocalization of heparin-binding EGF-like growth factor (HB-EGF) as a possible immunotarget in diagnosis of some soft tissue sarcomas.

Heparin-binding EGF-like growth factor (HB-EGF), a member of the family of epidermal growth factors (EGFs), is involved in several biological processes and tumor formation. Several lines of evidence show that HB-EGF plays a key role in the acquisition of malignant phenotype. Studies show that HB-EGF expression is essential in oncogenesis of cancer-derived cell lines. HB-EGF is a promising target for cancer therapy. The aim of this study was to find new insights on the biological features of the soft tissue sarcomas, in order to consider the possibility to use HB-EGF as an immuno-target in histotype characterization and to facilitate therapeutic intervention. In our study we did HB-EGF-immunostaining on tissue samples collected from 43 human soft tissue sarcomas. We analyzed HB-EGF immunoexpression in some types of tumors such as clear cell sarcomas, leiomyosarcomas, phyllodes sarcomas, chondrosarcomas and liposarcomas. In relation to the different histotypes, we detected different immunostaining localization. From our results it was evident that pleomorphic cells, a signal of tumor progression, were HB-EGF immunostained, and this was accompanied by an extracellular matrix immunostaining. Moreover statistical analysis showed a correlation between HB-EGF immunostaining and the different types of analyzed soft tissue sarcomas. In conclusion, in some types of soft tissue sarcoma HB-EGF could be considered a useful diagnostic marker for their characterization.

Therapeutic potential of nitric oxide-modified drugs in colon cancer cells.

We have examined the influence of the nitric oxide (NO)-modified anti-inflammatory drug (S,R)-3-phenyl-4,5-dihydro-5-isoxasole acetic acid (VGX-1027) named GIT-27NO or the NO-modified antiviral drug saquinavir (Saq) named Saq-NO on two colon cancer cell lines, mouse CT26CL25 and human HCT116. The effects of the drugs on cell viability, apoptosis, proliferation, and metastatic potential were analyzed. The release of NO and oxygen and nitrogen species was also determined. The efficacy of the drugs was evaluated in vivo in BALB/c mice injected with CT26CL25 cells. Both agents suppressed the growth of colon cancer cells in vitro and reduced tumor volume in syngeneic BALB/c mice. However, their mechanisms of action were different because GIT-27NO released larger amounts of nitrite than Saq-NO in cell cultures and its antitumor action depended on the intracellular NO release inside the cells. On the contrary, Saq-NO released barely detectable amounts of NO and its antitumor action was NO-independent. In fact, cotreatment with an NO-peroxynitrite scavenger revealed that GIT-27NO but not Saq-NO acts through peroxynitrite-mediated cell destruction. At the cellular level, GIT-27NO prevalently induced proapoptotic signals followed by caspase-dependent apoptosis. In contrast, Saq-NO blocked cell proliferation, changed the adhesive, migratory, and invasive properties of the cells, and decreased metastatic potential in vivo. In conclusion, differences in NO release and oxidative stress generation between GIT-27NO and Saq-NO resulted in different mechanisms that caused cell death.

The tumor microenvironment in hepatocellular carcinoma (review).

The tumor microenvironment has been largely studied as a dynamic system orchestrated by inflammatory cells, including cancer cells, stroma as well as the extracellular matrix. It is useful to describe and predict the phenotypic characteristics of cancer. Furthermore, a better understanding of its interplay with the various aspects of the tumor cells may be utilized for the discovery of novel molecular targets. Liver cancer is considered a model of the relation occurring between the tumor micro-environment and tumor development. The chronic inflammatory status of the liver, sustained by the infection of hepatitis viruses, as well as the production of cytokines and growth factors within the parenchyma, lead to an intricate microenvironment. The identification of novel molecular therapeutic targets may improve the outcome of patients with liver cancer as it remains the third leading cause of cancer death worldwide. In the present study, the tumor microenvironment in hepatocellular carcinoma (HCC) was explored by a review of the literature. Studies on hepatitis virus infections and the consequent chronic inflammatory status were examined. In this context, immune-mediated and/or virus-related molecular mechanisms have been hypothesized as being responsible for liver cancer development. The interlink among HCC microenvironment components, comprising cellular elements, cytokines, growth factors and several proteins is also described together with the role of matrix metalloproteinases in HCC development. Finally, the rationale for targeting tumor-stromal interface is summarized in the context of new therapeutic opportunities.

Gene alterations in the PI3K/PTEN/AKT pathway as a mechanism of drug-resistance (review).

The most common therapeutic approach for many cancers is chemotherapy. However, many patients relapse after treatment due to the development of chemoresistance. Recently, targeted therapies represent novel approaches to destroy cancer cells. The PI3K/PTEN/AKT pathway is a key signaling pathway involved in the regulation of cell growth. Dysregulated signaling of this pathway may be associated with activating mutations of PI3K-related genes. Analyses of these mutations reveal that they increase the PI3K signal, stimulate downstream Akt signaling, promote growth factor-independent growth and increase cell invasion and metastasis. In this review, we summarize the PI3K/PTEN/AKT pathway genetic alterations in cancer and their potential clinical applications.

Characterization of human melanoma cell lines and melanocytes by proteome analysis.

We have analyzed the proteomes of two human melanoma cell lines (A375 and 526), and of the human melanocytes, (FOM 78), by two-dimensional electrophoresis (2D-PAGE) and liquid chromatography - tandem mass spectrometry (LC-MS/MS). Our comparative proteomic analysis revealed that six proteins were over-expressed in both melanoma cell lines as compared to melanocytes: galectin-1, inosine-5'-monophosphate dehydrogenase 2, serine/threonine-protein phosphatase 2A 65 kDa regulatory subunit A alpha isoform, protein DJ-1, cyclophilin A and cofilin-1. We show, for the first time, that only specific isoforms of these molecules are over-expressed in melanoma. Different protein profiles were also found between each individual melanoma cell line and the melanocytes. s-Methyl-5-thioadenosine phosphorylase, ubiquitin and ribosomal protein S27 a precursor, the basic form of protein DJ-1, annexin a1, proliferation associated protein 2g4, isoform alfa-enolase of alfa-enolase, protein disulfide-isomerase precursor, and elongation factor 2 were more strongly expressed in A375 cells compared to melanocytes. In 526 cells, 60s acidic ribosomal protein p1 and calreticulin precursor were more highly expressed than in melanocytes. These molecular differences may help in better understanding melanoma development and its different responsiveness to therapies. The identified proteins could be exploited as biomarkers or therapeutic targets for melanoma.

Decreased expression of GRAF1/OPHN-1-L in the X-linked alpha thalassemia mental retardation syndrome.

BACKGROUND: ATRX is a severe X-linked disorder characterized by mental retardation, facial dysmorphism, urogenital abnormalities and alpha-thalassemia. The disease is caused by mutations in ATRX gene, which encodes a protein belonging to the SWI/SNF DNA helicase family, a group of proteins involved in the regulation of gene transcription at the chromatin level. In order to identify specific genes involved in the pathogenesis of the disease, we compared, by cDNA microarray, the expression levels of approximately 8500 transcripts between ATRX and normal males of comparable age. METHODS: cDNA microarray was performed using total RNA from peripheral blood mononuclear cells of ATRX and normal males. Microarray results were validated by quantitative real-time polymerase chain reaction. RESULTS: cDNA microarray analysis showed that 35 genes had a lower expression (30-35% of controls) while 25 transcripts had a two-fold higher expression in comparison to controls. In the microarray results the probe for oligophrenin-1, a gene known for its involvement in mental retardation, showed a decreased hybridization signal. However, such gene was poorly expressed in blood mononuclear cells and its decrease was not confirmed in the quantitative real-time RT-PCR assay. On the other hand, the expression of an homologous gene, the GTPase regulator associated with the focal adhesion kinase 1/Oligophrenin-1-like (GRAF1/OPHN-1-L), was relatively high in blood mononuclear cells and significantly decreased in ATRX patients. The analysis of the expression pattern of the GRAF1/OPHN-1-L gene in human tissues and organs revealed the predominant brain expression of a novel splicing isoform, called variant-3. CONCLUSIONS: Our data support the hypothesis of a primary role for altered gene expression in ATRX syndrome and suggest that the GRAF1/OPHN-1-L gene might be involved in the pathogenesis of the mental retardation. Moreover a novel alternative splicing transcript of such gene, predominantly expressed in brain tissues, was identified.

Distribution of p53, GST, and MTHFR polymorphisms and risk of cervical intraepithelial lesions in sicily.

OBJECTIVES: Host factors, including genetic polymorphisms, may explain some of the individual differences in cervical cancer occurrence, and susceptibility information may be useful to address effective and specific preventive strategies for different countries. The purpose of the present study was to investigate the role of p53 codon 72, glutathione S-transferase class mu (GSTM1), glutathione S-transferase class theta (GSTT1), and methylenetetrahydrofolate reductase (MTHFR) C677T polymorphisms on the risk for infection and/or of cervical intraepithelial lesions in women attending a colposcopy service in Catania, Sicily, with an already reported high prevalence of human papillomavirus. METHODS: To identify the association among individual genetic polymorphisms, human papillomavirus infection, and histological findings, a case-control study was designed. Furthermore, to assess the combined effects of these polymorphisms on cervical cancer risk, combined genotype frequencies were compared among case patients and controls. RESULTS: Women homozygous for the p53 codon 72 Arg genotype were at a 5.6-fold higher risk for developing cervical intraepithelial neoplasia (CIN) 2 or 3 compared with those showing homozygosity for the Pro genotype or heterozygosity for the Pro/Arg genotype. The GSTM1 and GSTT1 null genotypes were overrepresented in infected patients and in women with CIN 2 or 3, although without any significant associations. A decreased risk for CIN of individuals homozygous for the MTHFR T allele was shown. CONCLUSIONS: After multiple logistic analyses, the presence of the allele 677T of the MTHFR gene was the best explaining protective factor against cervical carcinogenesis, and the allelic distribution in the control group followed the Hardy-Weinberg equilibrium expectations. However, the findings of our study still remain to be confirmed by additional and larger population-based surveys.

Human papillomavirus infection: low-risk and high-risk genotypes in women in Catania, Sicily.

INTRODUCTION: Human papillomavirus (HPV) infection has been strongly and consistently associated with cervical carcinoma and its cytologic precursors, such as squamous intraepithelial lesions. A cross-sectional survey was conducted with the aim of estimating the prevalence of cervical HPV infection in women attending a service of colposcopy in Catania, Eastern Sicily, Italy. METHODS: The prevalence of type-specific HPV was examined in women with negative colposcopic results and cervical intraepithelial neoplasia grades 1, 2, or 3, with the aim of providing some cross-sectional figures on the local epidemiology of HPV infection. RESULTS: Human papillomavirus DNA was found in 62.1% of women with negative colposcopic results and in 73.2% with positive colposcopic results. Among high-risk types, a predominance of HPV-16 (51.5% of infected women) was shown followed by HPV-56 (29.7%). An age-related pattern was described with a peak in HPV prevalence among women younger than 25 years, followed by the expected decline in prevalence and a second characteristic peak in the perimenopausal or postmenopausal years, useful to design future control strategies. CONCLUSIONS: The age-related pattern of HPV prevalence and the presence of uncommon high-risk genotypes and their role in the pathogenesis of cervical cancer need to be addressed by specific epidemiologic studies to design large-scale screening programs and multivalent vaccine strategies.

Ultrasonographic study of gallbladder wall thickness and emptying in cirrhotic patients without gallstones.

BACKGROUND AND AIM: Gallbladder wall thickening and impaired contractility are currently reported in cirrhotic patients and often related to portal hypertension and hepatic failure. The purpose of this work was to evaluate, by ultrasonographic method, gallbladder wall thickness and gallbladder emptying after a standard meal in normal subjects and in patients with compensated liver cirrhosis without gallstones. METHODS: Twenty-three patients with Child-Pugh class A liver cirrhosis and twenty healthy controls were studied. Gallbladder wall thickness (GWT), gallbladder fasting volume (FV), residual volume (RV), and maximum percentage of emptying (%E) were calculated. Measurements of mean portal velocity, portal vein flow, and serum albumin were performed too. Statistical analysis was assessed by Student's "t test" for unpaired data. RESULTS: GWT was 0.60 +/- 0.22 cm in cirrhotic patients and 0.21 +/- 0.06 cm in controls (P < .0001). FV and RV were, respectively, 37.8 +/- 3.7 cm(3) and 21.8 +/- 3 cm(3) in cirrhotic patients, 21.9 +/- 4.2 cm(3) and 4.6 +/- 2.2 cm(3) in healthy volunteers (P < .0001). %E was smaller in cirrhotics (42.6 +/- 7.8) as compared to controls (80.3 +/- 7.2; P < .0001). CONCLUSIONS: In patients with compensated liver cirrhosis without gallstones gallbladder wall thickness is increased whereas its contractility is reduced. These early structural and functional alterations could play a role in gallstone formation in more advanced stages of the disease.

Activation of the osteopontin/matrix metalloproteinase-9 pathway correlates with prostate cancer progression.

PURPOSE: Prostate cancer remains the second most frequent cause of tumor-related deaths in the Western world. Additional markers for the identification of prostate cancer development and progression are needed. Osteopontin (OPN), which activates matrix metalloproteinases (MMP), is considered a prognostic biomarker in several cancers. "In silico" and experimental approaches were used to determine whether OPN-mediated MMP activation may be a signal of prostate cancer progression. EXPERIMENTAL DESIGN: Pearson correlation coefficients were computed for each OPN/MMP pair across seven publicly available prostate cancer gene expression data sets. Using Gene Set Enrichment Analysis, 101 cancer-related gene sets were analyzed for association with OPN and MMP-9 expression. OPN, MMP-9, MMP-2 tissue inhibitor of metalloproteinase-1 plasma levels, and MMP gelatinase activity were measured by ELISA and zymography in 96 and 92 patients with prostate cancer and benign prostatic hyperplasia, respectively, and 125 age-matched healthy men. RESULTS: Computational analyses identified a significant correlation only between MMP-9 and OPN, and showed significant enrichment scores in "cell proliferation", "genes constituting the phosphoinositide-3-kinase predictor", "proliferation signature", and "tumor metastasis" gene sets in association with both OPN and MMP-9. Plasma analyses revealed a significant increase in OPN and MMP-9 levels and activity in patients with prostate cancer in association with clinical variables (prostate-specific antigen > 4 ng/mL and Gleason score > 7). Significant correlation between OPN and MMP-9 levels were also observed. Mean plasma levels of OPN and MMP-9 decreased in patients with prostate cancer within 6 months after prostatectomy. CONCLUSIONS: The concordant computational and experimental data indicate that the extent of OPN pathway activation correlates with prostate cancer progression.

[Prognostic index of acute hepato-cellular damage]. / Indice prognostico del danno epato-cellulare acuto in rapporto alla rigenerazione epatica.

INTRODUCTION: Fulminant hepatic failure is the end result of many different acute damage to the liver. In the present study we compared the clinical to the experimental experience and we postulated the usage of Vascular Endothelial Growth Factor in the clinical arena as a potential treatment in alternative to liver transplantation. MATERIALS AND METHODS: Twelve patient diagnosed with fulminat hepatic failure have been enclosed in the present study. Each patient underwent trans-jugular liver biopsy in order to assess the degree of liver necrosis as well as the following biochemical investigation: AST ALT, Total Bilirubin, _gt, alkaline phosphatase, prothrombin time. RESULTS: Twenty-five percent of those patients required support in the Intensive Care Unit without need for transplantation. Forty-one percent of those patients underwent liver transplantation, and 36% of them died before the liver become available. These results were compared with an experiment, previously performed by our group, where 260 rats were poisoned with CCl4 and subsequently treated with Vascula Endothelial Growth Factor (VEGF). CONCLUSION: The rate of the hepatic regeneration has been found to be critical in the prognosis of patients diagnosed with fulminant hepatic failure.