Main epidemiological and diagnostic aspects

HCV is an RNA virus that is characterized by structural regions (E1, E2 and NS1) located at the 5’ ends, which encode the core and envelope proteins, and non-structural regions (NS2, NS3, NS4 and NS5) located at the 3’ ends, involved in viral replication and in the lysis of infected cells.

The HCV virus is extremely heterogeneous and presents a high mutation rate in its variable E2 regions. Six HCV virus genotypes are known and some of them are divided into subtypes. Genotype 1 is the most widespread in industrialized countries, hence in Italy too (over 50% of cases). The series of mutations (including the mutation of only one amino acid) justifies the quasispecies concept that expresses the extreme gnomic variability of the virus, the important molecular base of the failed eradication and thus of ‘chronicization’. The HCV infection is widespread throughout the world and its prevalence varies between 1.5% in the US and 6-7% in Africa and South East Asia. The main risk factors of HCV infections are:

- active drug addiction;
- tattoos;
- organ transplant;
- transfusion of infected blood;
- dental care;
- accidental exposure to infected blood; and,
- sexual intercourse (rarely). (The risk allocated by US epidemiological studies to this method of transmission is below 3%).

HCV infections have a higher tendency to become chronic than HBV infections and their incubation period can vary from 6 to12 weeks. Almost always the acute forms are asymptomatic: in over 90% of cases the primary infection develops asymptomatically. It can be diagnosed by using the immunoenzymatic method (Elisa) to search for anti-HCV antibodies (AbHCV) at least 7-8 weeks after the contagion. Considering the relatively delayed appearance of these antibodies, in suspect cases an early diagnosis can be made by using genic amplification methods to search for the HCV RNA. In acute forms that tend to solve spontaneously the HCV RNA is inclined to disappear, while non-neutralizing anti-HCV antibodies still remain. The natural story of HCV infections foresees three possible patterns:

- in 20-25% of cases - complete healing with the normalization of transaminases, disappearance of the HCV RNA and later the disappearance of HCV antigens;

- ‘chronicization’ of the infection (60-70% of cases) with a more or less rapid development of chronic hepatitis (high ALT and more or less high levels of HCV RNA).

Degree of ‘chronicization’: correlations with ALT and histological activity

The transformation of chronic hepatitis in cirrhosis triggers a series of clinical signs that are common to other cirrhosis cases too. Besides, the causes of the transition from chronic HCV hepatitis to hepatocarcinoma (HCC), mediated or not by the degree of cirrhosis, are not entirely known. Special importance has been given to the virus’ possible carcinogenic role and, according to recent mouse models, to the imbalance between the function of the pro-apoptotic genes (fas) and the anti-apoptotic ones (bcl 2) - an over expression of the genes bcl 2 seems to exercise an important role in the induction of carcinogenesis.

Treatment notes

IFN-alpha is the choice treatment in chronic HCV hepatitis. Meta-analysis studies have proved that generally the best treatment specifies a dosage of 6 million units (MU) three times a week for 12 months and that the initial dosage must be taken daily. The IFN activity is related with its antiviral action (the removal of infected cells) and especially the immunomodulating cells (the induction of HLS class I molecule expression, which strengthens the innate immunity such as macrophages and natural killer lymphocytes). There are no certain factors to predict the response to interferon treatment but the following variables are favourably related with a better response:

- young age;
- female gender;
- low body weight;
- absence of cirrhosis; and,
- the disease’s short duration.

However we must not leave out the patient’s immune response characteristics. Along with these non-specific variables, the virus’ genotype too plays an important role: genotype 1 is the most resistant to treatment with IFN-alpha compared to genotypes 2 and 3. Another considered variable is the viral load before treatment. The study of HCV virus kinetics has not made significant progress and it is still not possible to draw replication models as in HIV infections. However, certain studies stress that a rapid decrease in the viral load, following the initial stages of treatment, is related to greater success.

Another important role seems to be played by the new type of pegilated IFN – administered weekly it assures a better compliance and constant and continuous antiviral activity. In fact, one of the greatest problems related with the use of traditional IFN is the loss of a constant antiviral activity; this could enable a special replication process in a virus that tends to invade non-hepatic cells too. The IFN and ribavarin combination has been remarkably successful and effective. This association results in a response rate of about 80% in formerly non-responsive individuals, while in naive patients (those never treated), a much higher response percentage (about 60%) can be obtained after 48 weeks, by only administering IFN. According to the results of certain studies, this association seems to strengthen the IFN’s immunomodulating action; in particular, it seems targeted at the activity of CTL lymphocytes (O. Perrella, data not published). Other studies also suggest the association of IFN + ribavarin + amantadin, but controlled trials are required to document this combination’s real effectiveness. The main knowledge we have of the immune response during chronic HCV hepatitis and the progress of basic immunology let us consider the future use of certain cytokines as therapeutic possibilities. This strategy’s molecular base would be related to the modulation of the inflammatory process promoted by a Th1 type intrahepatic immune condition. A recent study published on Gastroenterology, Nelson et al., 2000, experimented with IL-10 in HCV patients with a high index of fibrosis. According to the trial’s preliminary data, this cytokine reduced the degree of fibrosis and improved other necro-inflammatory indexes. However the widespread use of immunomodulating treatment still seems premature because the immunopathogenic process is still not completely known and because the manipulation of the immune system must be guided by a greater knowledge of basic immunology.

Immunopathogenesis

The HCV viral infection develops significantly towards chronic hepatitis, cirrhosis of the liver and lastly, even without the cirrhosis stage, at times into hepatocarcinoma. This infection’s natural history and the increasing information on immune response mechanisms suggest that cell immunity plays an important role in the mechanisms that influence the infection’s progress. This participation seems to have a double valence that is related both to the guest’s inability to generate an effective viral clearance and to the virus’ interference with the immune response’s function. Lymphocytes B are another target of the C virus, besides hepatocytes, and this explains the C virus’ involvement in the genesis of lymphoproliferative and autoimmune diseases. However, how the virus enters the cell still remains unexplained - a second receptor has been theorized for the signal’s penetration. The immune system’s role in limiting the clearance in viral infections lies in a sort of balance between the phenomena that induce the activation and those that cause its conclusion.

Various components of the immune system, based on the coordinated intervention of lymphocytes and on the elaboration of the various cytokines, participate in this regulation. These, according to their Th1 or Th2 response, determined both by the antigen quantity and by the background present in the surrounding “microenvironment”, can often cause the disease to develop differently. This molecular basis characterizes the tissue infected by the virus and in whose cells a more or less intense viral replication takes place. One of the more intriguing and fascinating problems of the immunopathogenesis of chronic HCV hepatitis is the cytokine pattern that can influence the hepatic disease’s development. A Th1 type energetic initial pattern would be more associated with healing from a C virus infection, while an initial expansion of the immune response polarized towards Th2 would make the disease chronic. In the course of the infection, the virus releases viral antigens that activate both B-lymphocytes and T-lymphocytes; these produce substances that trigger viral clearance and can also damage the tissue involved when the inflammatory process is prolonged. This creates an immunopathological picture in which the disease can be backed by the immune response itself. For example the C virus’ infection draws a Th1 type response, which, if it does not succeed in eradicating the virus, enables its possible mutation, thus triggering a new Th1 response, prolonging the inflammatory process and hence the tissue damage (immunological escape phenomenon) Figure 1.

Immunological studies on hepatic tissue have confirmed that a strong and persistent Th1 response influences the chronic disease’s progress. The constantly formed new viral alterations act as new antigens on the immune system, influencing its chronic immune mechanism. Another possible mechanism that explains the virus’ persistence could be the induction of the T-lymphocyte energy and apoptosis mechanism: by producing an immune microenvironment that is characterized by Th2 cytokines, the C virus could increase the lymphocytes’ energy by failing to express co-signals. The C virus can act at various levels in the immune response:

- humoral response;
- HLS class I lymphocyte response; and,
- HLA class II lymphocyte response.

Experimental studies on animal models (chimpanzees) have proved how neutralizing antibodies are really produced in the course of the infection but they are unable to keep up with the virus’ constant mutations. The neutralizing antibodies’ ineffectiveness was also confirmed by recent studies on E2 epitomes. Lastly, there still remain obscure regions in the immunopathogenesis of chronic HCV hepatitis in which a balance between the Th1 and Th2 immune responses and especially between the networks that regulate the immune response influence the infection’s natural history. Unlike chronic HBC hepatitis, where the role of immunity is clearly related to the disease’s history, many questions are still unexplained in hepatitis.

Oreste Perrella
VII Division of Intective Disease and Immunology
"D, Cotugno" - Napoli

Translated by interpres sas