Clinical And Biochemical Characteristics Biology Essay

This is a retrospective study including patients with HCC associated to chronic hepatitis C or cirrhosis related to HCV with a positive HCV RNA who visiting our hospital (Hepatology and Digestive Surgery department) between January 2000 and December 2010. We included in the study two groups of patients: cirrhosis-HCC group (CIR-HCC) and chronic hepatitis C with HCC group (CHC-HCC). CIR-HCC Group consisting of all patients who satisfied the following inclusion criteria: having a liver biopsy (minimum 18mm) with Metavir score of fibrosis F≥3 at base line and get a HCC. The CHC-HCC formed by patients with chronic hepatitis C diagnosis by liver biopsy and get HCC. The diagnosis of HCC was performed by liver biopsy (minimum 18mm) on hypervascular liver nodule or treated by liver resection or liver transplantation. All liver biopsy or surgical resections were reviewed at our pathology department by an experienced liver pathologist. The Criteria of exclusion was the HCC or cirrhosis or chronic hepatitis C diagnosed by other method than liver biopsy, patient treated by antiviral therapy for HCV infection, patient with coexistent causes of cirrhosis or liver disease than HCV or Coinfection HBV (Ab Hbs positive) or HIV. We excluded patients treated with IFN in order to avoid selection bias and factors that could interfered analysis. Several studies have shown the effect of IFN treatment on fibrosis. Moreover, the exclusion of patients who have not had resection even if they had a biopsy was justified in order to have enough of material for histological analysis.

Clinical and biochemical characteristics

We carried out a retrospective analysis of a prospectively maintained database containing all patients with HCC associated to HCV infection at our hospital.

Patients’ files were retrospectively reviewed and analyzed to obtain the following demographic, clinical, and biochemical data collected at the time of admission: age; sex; cause of cirrhosis; serologic markers of hepatitis B (hepatitis B surface antigen [HBsAg], antibody to hepatitis B core antigen, and antibody to hepatitis B surface antigen) and hepatitis C (antibody to hepatitis C virus [anti-HCV]); serum levels of aspartate aminotransferase, alanine aminotransferase, total bilirubin, albumin, and AFP; prothrombin activity, platelet count, viral load and mode of transmission HCV infection. Complications factors: Child±Pugh score, the presence or not of ascites, encephalopathy, esophageal varices, previous variceal haemorrhage, and a history of gastrointestinal bleeding at presentation. The absence of significant alcohol intake was estimated to less than 20 g/d of ethanol consumption. The relationship between metabolic syndrome and HCC in non-cirrhotic patients has been studied and was absent in our series. Metabolic syndrome has been defined by the association of diabetes type2, arterial hypertension, elevated triglyceride levels, low high-density lipoprotein cholesterol levels (hypercholesterolemia), and obesity (BMI > 30 kg/m²).

Histological data

Including METAVIR score (12), number and size of nodule, presence of microvascular invasion, presence of steatosis, iron overload, presence of NASH based on the presence of steatosis, clarification/ballooning of hepatocyte and lobular inflammation (13) (14). Each biopsy was individually coded and processed for histopathological interpretation by a single experienced liver pathologist (P. Bedossa) blinded to patient identity and clinical and biological data. A minimum of 6 portal tracts per biopsy specimen for noncirrhotic patients was deemed adequate for study analysis. All biopsies were processed routinely, formalin-fixed, serially sectioned, and stained with hematoxylineosin, Masson’s trichrome, and Sirius red for fibrosis evaluation and Perls staining for iron. A detailed descriptive scoring sheet was employed, including the grading and staging of chronic hepatitis or cirrhosis according to METAVIR classification, qualitative evaluation of steatosis (0, absent 1, present), predominant zonal distribution of fat (pericentral, periportal, azonal, diffuse), clarification/ballooning of liver cells (0, absent; 1, mild; 2, marked), perisinusoidal fibrosis (0, absent; 1, mild; 2, marked), Mallory hyaline (0, absent; 1, present) and presence of glycogenated nuclei (0, absent; 1, present) as well as other more less common histological features as described by Kleiner et al.(15).

Statistical analysis

Data are expressed as median (range) or in percentages. The chi-square or fisher exact test was used to compare the categorical variables, and Wilcoxon test was used to compare the continuous variables. A P-value of <0.05 was considered significant. Analysis was performed using a SAS 9.2. Software (SAS Institute Inc., Cary, NC, USA).

Results

Study population

This study screened 437 patients who have been admitted for HCC associated to HCV during the period 2000-2010. We excluded 376 patients for the following reasons: 331 had transarterial chemoembolization (CEL), radiofrequency ablation (RF) or local-regional therapies (asymptomatic treatment or absence of treatment) or other exclusion criteria (as co-infection HIV) and 45 had previously IFN therapy (see Flow chart).

Finally the study population included 61 patients: 46 patients CIR-HCC vs 15 patients CHR-HCC (Table1). The overall performance status test (according the performance ECOG) was similar in the two groups. The median follow-up was 28 months (range: 1-143). The median age was 62 vs 69 years-old and p-value=0.154. Male were less in CIR-HCC than CHC-HCC (31(67%) vs 13(87%) p=0.037). Presence of markers past VHB infection were significant in non-cirrhotic patients than in cirrhotic group (5% vs 33% for Anti Hbs with p=0.048, 5% vs 40% for Anti-Hbc with p=0.001 and 5% vs 20% for both associated with p=0.001. There was a significant difference between two groups in term of albumin serum, AST, bilirubin total, platelets count and prothrombin. AFP serum level, globally considered, was not significantly different between the two groups (26 ng CIR-HCC in and 19 ng in CHC-HCC p=0.847). In addition no significant difference was observed in viral load between the two groups (5,6 log in CIR-HCC and 4,6 log in CHC-HCC, p=0.108). Genotype 1 were significantly less associated in CIR-HCC than CHC-HCC 27 (59%) vs (12 (80%) and p=0.001). The rate of patients with metabolic syndrome was not statistically different between CIR-HCC group 4(8%) vs CHC-HCC group 2(13%) and p-value=0.366. Despite 6(40%) of patient had arterial hypertension, 3(20%) had diabetes type2 and 2(13%) had hypercholesterolemia but their BMI were <30 kg/m² in CHC-HCC group.

Table1: Baseline clinical and blood parameters of 61 patients with HCV-HCC according presence of cirrhosis or not

Histology analysis

There was no difference in terms of tumor size, tumor type, extension of vascular invasion and BCLC classification between the two groups (Table 2). Tumor in CHC-HCC group were more often well-differentiated than CIR-HCC group (53% vs 33% and p=0.0004). The frequency of steatosis was significantly higher in CIR-HCC group than CHC-HCC group (48% vs 13%, p=<.0001). The presence of NASH was more frequent in CIR-HCC group than CHC-HCC (19% vs 0% and p=<.0001). (Table2.)

Table2: Pathological findings in 61 patients with HCV-HCC according presence of cirrhosis or not

Discussion

This study can make two observations: frequency HCC on HCV in absence of cirrhosis and identification of characteristics associated with the Nash in patients infected by chronic hepatitis C.

The current increase of the incidence of HCC in the prevalence of chronic HCV infection and NAFLD in the world seems be a consequence of the increasing prevalence of obesity and an ageing population. (16), (17), (18). However the own process that lead to malignant transformation of HCV-infected hepatocytes still remain unclear, but as most HCC related to HCV infection occurs on severe liver fibrosis or cirrhosis. The main assumption is that the mechanism of carcinogenesis seems more likely to be indirect, such that the process of regeneration and repair are important, rather than a direct oncogenic effect of HCV infection or the inflammatory response to the virus. Now it is well recognised that chronic HBV infection can lead to HCC in the absence of advanced fibrosis or cirrhosis (19), (20), (21), (10). Because in HBV virus, the integration of the host cell genome may be mutagenic directly by causing genomic instability, loss of tumour suppressor activity or over-expression of most of genes involved in regulation of cell cycle proliferation.

The frequency of HCV-HCC in absence of cirrhosis remains unknown (11). Our findings show that 4% (18 patients) of HCC may arise in HCV without cirrhosis. According resection treatment, 14% of patient had HCC-HCV in absence of cirrhosis. This figure seems the same than 16% that found Yeh et al. (22) on combining prevalence of HCV-positive cases among all resections for, which was significantly higher than the general prevalence of HCV-infected individuals in the US population.

In the present study, we characterized risks factors as NASH or metabolic syndrome in the occurrence of HCC in HCV infection in absence of cirrhosis. NASH has been proposed as a possible cause of cirrhosis even though most of the histologic characteristics of NASH are not present in cryptic cirrhosis (23), (24), (25). In the absence of reliable clinical and new biochemical markers, diagnosis of NASH relies purely on histopathological bases (14), (15).

Within the context of NAFLD (Non-Alcoholic Fatty Liver Disease), only patients with biopsy-proven NASH have convincingly been shown to progress to cirrhosis; therefore, it is of major importance to clearly delineate the histopathological criteria of these entities NALFD and NASH in CHC. However, the histopathological criteria required for diagnosis of NASH are ambiguous, because it result of a progressive disease associating several different characteristics and different definitions and specific grading have been proposed (15), (26), (27). Within the context of CHC, lesions related to the background chronic inflammatory disease increase the complexity of NASH identification, because some of these elementary features might overlap (steatosis, lobular inflammation). Therefore, diagnosis of NASH in patients with CHC requires careful histopathological evaluation of selected criteria, the relevance of which should be validated by clinico-biological correlates. More specifically, it remains to be determined whether histopathological criteria for the definition of NASH in patients with CHC-HCC are relevant to characterize the association of NASH in patients with HCC associated to CHC in absence of steatopathy or metabolic syndrome. For those raison we selected patients underwent resection or had a good biopsy (18mm minimum). In our study only 9(19%) patients had NASH and all had cirrhosis vs 0% in noncirrhotic group (p=<.0001), 48% had steatosis in CIR-HCC vs 13% in CHC-HCC (p=<.0001). While no patient had NASH and only 13% had steatosis in HCC without cirrhosis group. In this condition the hypothesis of steatopathy or NAFLD seems unrealistic in those patients. The mechanisms of pathogenesis of NASH in occurrence of HCC in setting of CHC infection have not been elucidated. Diabetes and obesity have been previously related to HCC risk (28) (29) (30). In this study, we examined the role of metabolic syndrome and the combined effect of its components, and their interaction, as well as the modifying effect of HCV on the relation between metabolic syndrome and liver cancer risk. But only 2 (22%) patients had diabetes associated to syndrome metabolic in absence of obesity in CHC-HCC group. The mean of BMI was 25 ranged [19-28] even if 20% had diabetes and 13% had metabolic syndrome in CHC-HCC group. If the association of NASH and obesity increase the risk of HCC in chronic HCV, in our serie this association is absent (31), (32). Some data suggesting that hepatic steatosis, obesity and diabetes type II increase the risk of occurrence of HCC in HCV infection suggest the idea that they are risk factors for the development of HCC in NASH. In this study those combinations were not found and may not explain the occurrence of HCC in noncirrhotic group.

However presence of VHB markers (Ac anti Hbs (33%), Ac anti Hbc (40%) and both associated (20%)) were significant in patients without cirrhosis than in cirrhotic group (5% vs 33% for Anti Hbs with p=0.048, 5% vs 40% for Anti-Hbc with p=0.001 and 5% vs 20% for both associated with p=0.001. This suggests the possible occult HBV infection in these patients. Occult infection with hepatitis B virus (HBV), defined as persistent detectable viral genome in serum or liver while HBV surface antigen (HBsAg) is undetectable. But this definition was improved with European Association for the Study of the Liver (EASL) (33) , as well as The Taormina Consensus Conference in 2008, defined "Occult infection with hepatitis B" as the "presence of HBV DNA in the liver of individuals testing HBsAg-negative with currently available assays" (34) and introduced a cutoff value for serum HBV DNA (< 200 IU/mL). The relationship between occult HBV infection and serological markers of HBV infection has been studied before, and the prevalence of occult HBV infection was usually higher in subjects positive for either anti-HBs or anti-HBc or for both anti- HBs and anti-HBc than in those negative for all serological markers (46% to 80% versus 20 to 50%) (35) (36). There is much evidence suggesting that HBV is a risk factor for the development of HCC in the case of occult infection as well, and this occurs both in patients with HCV-related and cryptogenic chronic liver disease (37) (38). Moreover, it recently demonstrated that the potential mechanisms whereby overt HBV might induce tumor formation are maintained for the most part in the occult status (39). However, all the studies performed in the field were crosssectional examinations of cases in which carcinoma hepatocellular already occurred, whereas to our knowledge no prospective study to date has investigated the fate of chronic hepatitis patients in terms of the development of HCC in accordance with their occult HBV status. Growing evidence revealed a high frequency of occult HBV infection in patients with HCV-related HCC.

The prevalence of anti-HBc and or anti-HBs varies from 50% to 90% in patients with anti-HCV positive HCC. The prevalence of serum and intrahepatic HBV-DNA ranges from 0% to 18%, and 13% to 80%, respectively, in these patients (40) (41). More recently, this occult HBV infection has frequently been identified in patients with chronic HCV infection (42) , and in such patients this occult infection may be associated with severe liver damage and even the development of hepatocellular carcinoma (HCC) (43). Squadrito et al. (38) provided further evidence that occult HBV may be associated with the neoplastic transformation of a chronic liver disease, and definitively confirm that the presence of HBV genomic sequences distinguishes those individuals at higher risk of developing HCC among Ag HBs negative Individuals. In their study Fifty-three patients (39%) were occult HBV carriers and 81 (61%) were not. Nine patients developed HCC during the follow-up; eight were positive and one was negative for occult HBV (P = 0.002). Fukuda et al study showed that HCV genotype 1b was a more frequent coinfection in these patients; this is not supported by other studies (44). In our serie genotype 1b was more frequent (60%). Into HCV virus, some HCV proteins, such as non-structural proteins NS3, NS4B and NS5A have a regulatory effect on cellular promoters and seems interact with some of cellular proteins involved in the carcinogenesis mechanism under certain particular conditions (45) (46). More, hepatocytes from patients with chronic HCV infected by genotype 1 may undergo replicative senescence, which may predispose to malignancy (47). Therefore evidence for a carcinogenic role for HCV infection is lacking. These conditions suggests that age > 62, genotype1, Markers of occult hepatitis B could be risk factors for HCC development in the absence of cirrhosis in patients with chronic hepatitis C. Hence the monitoring of these patients by ultrasound scans may be useful. This will allow screening of HCC at curative stage, curative treatment and a better prognosis.

As all retrospective study, this study has some limitations according selection of patients….

This study allows understanding the association of NASH in the physiopathological of HCC in CHC in absence of NASH that will lead to change manners in clinical care of these patients. To understand the precise mechanism of occurrence of HCC in CHC in absence of cirrhosis, we have to better define the pathogenic role of each risk factor and epidemiology of occult HBV infection. The pathogenic role of occult HBV infection in HCV-related HCC remains to be determined. Consequently, testing patients with CHC for HBV genomes appears to be a powerful tool for the identification of subjects who need to be more carefully monitored to allow early diagnosis of HCC.