Hepatitis B virus

Hepatitis B is an infectious disease of a person characterized by selective damage of the liver by a virus. This form of hepatitis is the most dangerous in its consequences among all known forms of viral hepatitis. Its causative agent is the hepatitis B virus (HBV).

For the first time the antigen of the hepatitis B virus was detected by B. Blumberg in the blood serum of the Australian aborigine in 1964, and the causative agent was discovered in 1970 by D. Dein (and others) and was called the Dane particles, since there was no complete certainty , that this is really a virus, not its components. Subsequently, all doubts disappeared, since in the Dane particles, genomic DNA and a viral DNA-dependent DNA polymerase were found. The virion contains three main antigens, for which the following designations were introduced in 1974:

• HBsAg is a superficial, or soluble, or Australian antigen.
• HBcAg is a core antigen (co-antigen).
• HBeAg is an antigen e, located in the core of the virion and, unlike HBcAg, is not only present in the virion, but also circulates in the blood in a free form or as a complex with the anti-HBeAg antibody. It is secreted into the blood from the hepatocytes with active replication of HBV.

The surface antigen - HBsAg - exists in the form of three morphologically different variants: 1) represents the supercapsid of the whole virion; 2) in large quantities occurs in the form of particles with a diameter of 20 nm, having a spherical shape; 3) in the form of filaments with a length of 230 nm. Chemically they are identical. HBsAg contains one common antigen a and two pairs of mutually exclusive type determinants: d / y and w / r, therefore there are four main subtypes of HBsAg (and, respectively, HBV): adw, adr, ayw and aur. Antigen a provides the formation of a common cross immunity to all subtypes of the virus.

Actually, the virion, the Dain particle, has a spherical shape and a diameter of 42 nm. The supercapsid of the virion consists of three proteins: the main (main), large and medium. The genome is enclosed in a capsid and is represented by a double-stranded annular DNA with a mass of 1.6 MD. The DNA consists of approximately 3200 nucleotides, but its "plus" -non is 20-50% shorter than the minus sign. With the 5 'end of the long filament, the virus-specific protein is covalently bound. The 5 'ends of both strands are complementary and form "sticky" sequences 300 nucleotides in length, so that the filaments are closed in the ring. The content of G + C in virion DNA is 48-49 mol%. In the core of the virion there is besides genomic DNA-viral DNA-dependent DNA polymerase. The "minus" -define DNA of HBV contains only four genes (S, C, P and X), but they are organized very compactly. The genes S, C, P, X strongly overlap and control the synthesis of the following products. Gene S codes for the synthesis of the main coat protein and contains all information about the surface antigen of HBsAg. In addition, it encodes the synthesis of medium and large envelope proteins. Proteins contain a common COOH-terminus, but their translation begins with three different initiator codons. Gene C codes for the synthesis of capsid proteins (HBcAg and HBeAg); although these proteins are coded by a single gene, the ways of their translation are different. Gene P is the largest. It includes part of all three other genes and encodes the enzymes necessary for replication of the virus. In particular, it encodes the reverse transcriptase, the domain of the enzyme RNA-ase H, the 5'-terminal protein of the "minus" -chain. Gene X codes for proteins that regulate the expression (expression) of all viral genes, in particular a protein with a mass of 17 kD, which is a gene transcription transactivator.

Proteins that form a surface antigen exist in glycosylated (gp) and non-glycosylated form. Glycosylated are gp27, gp33, gp36 and gp42 (the numbers are denoted by m in kD). Supercapsid HBV consists of the main, or basic, S-protein (92%); average M protein (4%) and large, or long, L-protein (1%).

• The main protein, p24 / gp27, or the main protein (protein S), is the main component of the HBV envelope. In the absence of other shell proteins, it polymerizes and forms spherical particles with a diameter of 20 nm, which consist of 100 polypeptide molecules.
• A large protein, p39 / gp42, or a long protein (L protein), is present in all three forms of HBsAg. It plays an important role in the morphogenesis of virions and in their exit from the cell. The L protein contains a sequence of protein M that is complemented at the N-terminus by sequences of 108 (ayw) or 119 (adw, adr, ayr) of amino acid residues encoded by the npe-Sl region of the S gene.
• The middle protein - gp33 / gp36, or the protein M, is also present in all three morphological forms of HBsAg. The protein M contains, at the N-terminus, a region of 55 amino acid residues encoded by the pre-52 region of the S gene. It is assumed that this site plays an important role in the recognition of hepatitis B virus by liver cells of a limited host range (human, chimpanzee monkey). The protein sequences encoded by the S-gene npe-S regions have high immunogenicity, and their determinants are located on the surface of the virion. Therefore, antibodies against these antigens play an important role in the formation of immunity against hepatitis B.

Synthesis of viral proteins is tightly controlled at the level of transcription and translation. In the transcription of the viral genome, two types of mRNA are synthesized:

• a smaller - 2100 nucleotides - encodes the main and middle proteins of the membrane;
• large - 3,500 nucleotides, i.e., longer than genomic DNA itself; it contains terminal repeats of 100 nucleotides in length.

This kind of mRNA encodes the capsid protein and the products of the P gene. It is also a template for viral DNA replication. As part of the genome, there are enhancers (transcription enhancers) - regulatory elements that activate the expression of all viral genes and act primarily in the liver cells. In particular, gene S is expressed at a very high level only in the liver cells and under the influence of steroid hormones. This circumstance explains why chronic hepatitis B and liver cancer (hepatoma) are observed in men more often than in women who have a lower level of steroid hormones.

Other regulatory elements of the hepatitis B virus modulate (control) the levels of synthesis of individual proteins. For example, a large protein is synthesized only in small amounts. Most of it on the surface of infectious virions. And the main protein and, to a lesser extent, the middle protein are synthesized in huge quantities and leave the cells in the surface antigen particles, which in the serum contain many times more than the mature virions. The number of surface antigen particles can be 1011 -1013 per 1 ml of blood (several hundred μg).

Hepatitis B virus is isolated in a new family of viruses - Hepadnaviridae, genus Orthohepadnavirus. Similar hepadnaviruses were found in various animals (earth proteins, marmots, chipmunks, Peking ducks).

Reproduction of hepadnaviruses takes place in an unusual way. In particular, the replication of genomic DNA occurs through the intermediate link - RNA, ie, with the mechanism of reverse transcription.

The life cycle of the hepatitis B virus.

• Adsorption on the cell.
• Penetration into the cell by the mechanism of receptor-mediated endocytosis (bordered fossa -> bordered vesicle -> lysosome -> nucleocapsid yield and penetration of the viral genome into the hepatocyte nucleus).
• Intracellular reproduction.

During the penetration into the cell, the short ("plus") DNA chain elongates (completes). In the nucleus, the cellular DNA-dependent RNA polymerase synthesizes an RNA of 3,500 nucleotides (pregenes) and smaller mRNAs for the synthesis of viral proteins. Then, the pregene and the viral DNA polymerase are packed into a newly synthesized capsid, which is transferred to the cytoplasm. Here, the reverse transcription of the pregenoma occurs. It synthesizes a new "minus" -define DNA. After the synthesis of the minus DNA is completed, the pregenomic RNA is destroyed. The virion DNA polymerase on the "minus" chain synthesizes a "plus" -chain. Viral DNA, now double-stranded, can exist in the cell for quite a long time and return to the nucleus for the next replication cycle. If the new virus particle does not undergo further replication, then the nucleocapsid that is formed, passing through the cell membrane, is covered with super-capsid, budding from the cell, and the short "plus" chain of DNA immediately stops there. That's why the length of this thread varies. In the typical acute form of hepatitis B, serologically the following serological markers appear successively in the blood: HBsAg, HBeAg and antibodies (IgM, IgG): anti-HBcAg. Anti-HBeAg and anti-HBsAg.

The hepatitis B virus does not contain an oncogene, but it is established that by introducing into the cell chromosome (at different sites), the viral DNA can induce in them various genetic alterations - deletions, translocations, amplifications, which can cause liver cancer - one of the most serious consequences of viral hepatitis B.

[1], [2], [3], [4], [5], [6], [7], [8],

Resistance of the hepatitis B virus

The hepatitis B virus is highly resistant. At room temperature it remains viable for 3 months, in the frozen state - for several years. The virus is completely inactivated by autoclaving (120 ° C), boiling for 30 minutes, dry heat at 180 ° C for 60 minutes, at 60 ° C for 10 hours. It is stable in an acidic medium, but is destroyed in alkaline. The virus dies when treated with H202, chloramine, formalin, phenol and with UV irradiation.

Pathogenesis and symptoms of hepatitis B

The virus is hematogenously injected directly into the liver. In the pathogenesis of hepatitis, autoimmune humoral and cellular responses play an important role. It is assumed that the defeat of hepatocytes is associated not so much with the direct action of the virus itself as with the host's immunological reactions that arise in connection with the modification of the cell membrane by viral proteins that induce the formation of autoantibodies to liver cells. Therefore, developing chronic hepatitis and cirrhosis of the liver can be considered as an autoimmune disease.

Cellular autoimmune reactions to viral proteins contained in the hepatocyte membrane are mediated by T-cytotoxic lymphocytes and other hepatic killer cells. Therefore, acute dystrophy of the liver can be considered as a reaction of rejection of a peculiar heterograft.

The incubation period lasts from 45 to 180 days, an average of 60 to 90 days. The clinical course of hepatitis B is characterized by great diversity; the disease can occur: in a latent form, detected only by laboratory methods, in a typical icteric form and in a malignant form that ends lethal. The duration of the pre-jaundiced stage ranges from one day to several weeks. The icteric period, as a rule, is long and characterized by well-marked symptoms (jaundice, hyperbilirubinemia, darkening of the urine, icteric sclera). Prolonged form is observed in 15-20% of patients, and in 90% of them chronic hepatitis B develops. In patients with lingering form, autoimmune processes are often observed, accompanied by an increased content of anti-hepatitis antibodies, which are detected by immunosorbent assay (IFM). In children, hepatitis B is mild and often without jaundice, in young children - mostly asymptomatic.

Postinfectious immunity (humoral and cellular) is long, lifelong, due to virus neutralizing antibodies (anti-HBsAg) in the absence of a surface antigen in the blood. Often observed latent immunization due to repeated contact with HBV, which is the cause of widespread immunity to the virus among the population. Usually patients with acute form of hepatitis B recover completely as the antibodies accumulate to it. However, in some cases, despite the high level of viral antigen in the blood (a circumstance explaining why parenteral infection occurs most often), antibodies to it are not produced. The virus remains in the liver, and a person for a long time, sometimes for life, becomes a chronic carrier. This circumstance is obviously connected with a weak immune response. One of the most common outcomes of chronic hepatitis B is cirrhosis and liver cancer, which develops after a latent period of up to 30-50 years.

Epidemiology of hepatitis B

The source of infection with the hepatitis B virus is only a person. Contrary to past beliefs that infection with the hepatitis B virus occurs exclusively parenterally, it is now proven that it is found in various secrets and excreta: in saliva, nasopharyngeal discharge, feces, tear fluid, in sperm, menstrual blood, etc. Thus, infection occurs not only parenterally, but also sexually and vertically (from the mother to the fetus), that is, practically infecting the hepatitis B virus in possibly different ways.

Hepatitis B in the world killed as many people as during all the years of World War II. The number of carriers of HBV, according to WHO, is from 0.1 to 20% of the population of different countries or regions.

[9], [10], [11], [12], [13], [14], [15], [16],

Diagnosis of hepatitis B

Currently, the main method for diagnosis of hepatitis B is the use of a reverse passive hemagglutination reaction (ROSGA) to detect a virus or its surface antigen, HBsAg. As already noted, the blood of the surface antigen contains many times more than the virus itself (100-1000 times). For the reaction of ROPGA, erythrocytes are sensitized with antibodies against the hepatitis B virus. If there is an antigen in the blood, a hemagglutination reaction occurs. ROPGA is simple, convenient, very specific. To detect antibodies to the HBsAg virus antigen, various immunological methods (DSC, RPGA, IFM, RIM, etc.) are used. In addition, PCR variants are used to detect HBV and its antigens.

Various immunological methods can be used to detect antibodies to the viral antigen (HBsAg) in the serum of the patient (DSC, RPGA, precipitation reaction, IFM, RIM, etc.).

[17], [18], [19], [20], [21], [22],

Specific prevention of hepatitis B

Taking into account the high incidence of hepatitis B, and also that there are a lot of HBV carriers in the world, WHO vaccinations against hepatitis B are mandatory and must be carried out in the first year of life. Two types of vaccines are suggested for vaccination. To prepare one of them, the plasma of virus carriers is used as raw material, since the virus antigen in it is contained in quantities sufficient to prepare the vaccine. The main condition for the preparation of this type of vaccine is their complete safety, i.e. Complete inactivation of the virus, which is provided by vaccine preparation technology. To manufacture a vaccine of a different type, genetic engineering methods are used, in particular, a recombinant clone of yeast that produces the surface antigen of the hepatitis B virus is used to prepare the antigenic material.

Both vaccines are highly effective (protect 95% of vaccinated). The duration of postvaccinal immunity is not less than 5-6 years. Vaccines have been created for both adults and infants and young children - the most important component in the global fight against hepatitis B. The full course of vaccination consists of three injections:

I dose - immediately after birth; II dose - after 1-2 months; III dose - until the end of the first year of life.

These vaccines are included in the WHO expanded program of immunization and are combined with the calendar of its implementation (on the recommendation of WHO, at the 1st year of life vaccinations against tuberculosis, poliomyelitis, hepatitis B, measles, tetanus, diphtheria, whooping cough).

Gammaglobulin, containing antibodies against HBV, is used for emergency passive immunoprophylaxis to persons who have had contact with a hepatitis B patient.

To treat hepatitis B (acute and chronic forms) use interferon and amixin (for the induction of its endogenous synthesis). In the treatment of chronic hepatitis B, a new drug lamivudine (synthetic nucleoside) is effective.