2) The second line of defense is internal and nonspecific. This response does not require specificity, meaning that the cells involved in the response: Phagocytes (destroy microbes by phagocyting them, by digesting them) and Natural killer cells (destroy infected cells) act on a broad range of microbes without any kind of memory. This second line of defense is almost irrelevant in case of an HIV infection but involved in the immune response following an HIV infection (Acquisition of memory starts here too)
3) The third line of defense is internal and specific. It is the specific immune response. This response can be divided into 2 arms: The humoral response involving formation of antibodies by the lymphocytes B and the cell-mediated response involving the formation of a complex network of lymphocytes. This third line of defense is 100% involved in HIV infection and and immune response against HIV infection. I will come back later on the lecture to explain how the immune system responds to an HIV infection.
The HIV virus belongs to the retrovirus family where the genetic material consists of an RNA molecule. I should say HIV viruses because there is more than one strain of HIV. The more common being HIV1 and HIV2. You will see at the end that the diversity of this virus makes AIDS treatment very difficult. It has an outer lipidic bilayer envelope taken from the cells it infects. This envelope contains spikes which are glycoproteins involved in the recognition and infection of the cells, specially a glycoprotein called gp120. I will come back later on the mechanism of infection. The inner protein coat juxtaposes the outer envelope. Deeper into the virus are the core proteins which surround the single stranded RNA and the reverse transcriptase. This enzyme plays a key role in the infection process.
B. Target cells (Fig 39-15 Campbell book)
The target cells of the HIV virus are the lymphocytes T helpers. These cells express a membrane protein called CD4. (Cluster of differentiation #4). This protein is usually involved in the recognition of antigens in association with the molecules of the MHC (Major Histocompatibilty Complex), at the membrane of the APCs cells (Antigens presenting cells); macrophages. But in the case of HIV infection, it is also a receptor for the gp120 glycoprotein of the HIV virus. CD4 also is also expressed at the surface of macrophages (especially in lungs and brain). I am using this figure to comment the target cells to show you how the T Helpers cells are in the center of the immune system. They participate in the cell mediated response but are also involved in the regulation of the humoral response. Then before more explanations, you can see already see how their death can terribly affect the entire immune system.
C. Mechanism of HIV infection (Fig 17-7 Campbell book)
Let's take now a closer look on the life cycle of the HIV virus and on its mechanism of infection. Remember that the binding between HIV and T Helpers is mediated by 2 proteins. The gp120 at the surface of HIV and the CD4 at the surface of T Helpers. CD4 being the ligand for gp120.
1) The first step of the infection consists in the binding of gp120 to CD4. Once the HIV virus is attached to the T Helpers, it fuses with the plasma membrane of the T Helpers and releases its genetic material, single stranded RNA into its host.
2) Once the viral RNA is released into the cytoplasm of the T Helpers, the viral RNA Reverse Transcriptase transcripts the RNA into DNA. We now have an RNA-DNA hybrid molecule. The viral RNA is then degraded by a T Helpers RNAse. The remaining DNA serves as a template for the synthesis of a complementary DNA strand. We now have a double stranded DNA molecule.
3) The double stranded DNA then migrates into the nucleus of the host. It then integrates the genome of the host. This incorporated form of virus is called provirus. At this stage the virus is non infectious. It is in a latent form. This stage can last several months. It is why the apparition of antibodies against the virus I not immediate after infection. The immune system has to wait for macrophages to present the viral antigens as you will see later.
4) The proviral genes are then transcribed into RNA using the host machinery transcription. This RNA can be divided into 2 parts. One part is mRNA and serves as template for the translation of viral proteins and the other part is the viral RNA which will be the viral genetic material.
5) The final step consists in the RNA packaging, meaning assembly of viral RNA and viral protein to form a new virus. You can see that the host plasma membrane is used to form the outer envelope of the virus.
6) New viruses are then release into the extracellular fluid and will infect other cells. This budding of new viruses will also potentially kill the infected cells. At this point I would like to introduce a new concept (fig 14-51 molecular cell biology book). The concept of formation of syncytium. As you can see on this figure, before releasing new viruses, the membranes of the T helpers express the viral glycoproteins. One of them being the gp120. At this stage, the infected T helpers cells can fusion with uninfected T Helpers cells via the binding between the gp120 expresses on infected cells and the CD4 expressed on uninfected cells. This fusion forms a syncytium (multinucleate cytoplasm). A syncytium can involve several T Helpers cells and is lethal for the cells involved. There are then 3 ways for the virus to weaken the immune system.
D. Immune response to HIV infection (Scientific American figure)
Once the HIV virus replicated, the level of virus increases in the body. This increase induces then a response of the immune system. This response involves the humoral as well as the cell-mediated immunity. This response can be divided in several steps.
1) The free viral particles released into the body are ingested and digested by the macrophages. The small peptides resulting of this digestion are then expressed at the surface of the plasma membrane and act as antigens. Remember that beside its phagocyte function, the macrophage is also able to present antigens to the immune system. It belongs to the APCs class of cells (Antigens Presenting cells). This recognition involves the MHC molecule at the surface of the macrophage and the TcR (T cell Receptor) at the surface of the T Helpers cells.
2) Once the T Helper cell is activated by the binding of MHC/Antigen and TcR/CD4, it secretes cytokines or interleukines which are going to activate other immune cells.
3) One type of actived cells are the lymphocytes B. You have already seen that activation of lymphocytes B results in the production of antibodies specific to the antigen, the HIV viral peptides. At this point we can detect free circulating antibodies in blood. This is the common way to detect HIV infection. For example, before doing any blood transfusion to a patient, doctors use only HIV negative blood. Mention PcR, in this case this technique allows to detect the genetic material of HIV virus when the virus circulates freely into the blood. Even if PcR detection is a more precise way of detection of any virus because we can detect it earlier, it is not the routine test. Then the free antibodies released into the body are going to bind the free virus. The binding occurs because of the specificity. The last step of the humoral response consists in the binding of the complex antibodies/antigens to macrophages via receptor for the constant regions of the immunoglobulins. This binding results in the phagocyte and digestion of the virus and finally in the death of the virus. This is how the humoral immunity depletes our body from HIV viruses.
4) The other response to HIV infection is the cell-mediated response. The secretion by the T Helpers of cytokines and interleukines is also able to activate the lymphocytes cytototix. The activated cytotoxic cells are then able to recognize the infected T Helpers cells. The binding between the NK cells and the infected T Helpers cells results in the death of the infected cells via secretion of lethal molecules.
I would like at this point make a break and summerize the major points. I know it is complicated because of the complexity of the immune system and because the target cells for HIV are the cells which play a key role in the immune response. Comment of the course of HIV infection figure.
1) So far we have seen the first step of an HIV infection is the increase of virus, released by the lymphocytes T Helpers. The level of virus into the blood is called viremia. It is why after an HIV infection the viremia increase rapidly because a lot of cells are infected and because they produce a lot of new viral particles. This is the acute phase of infection.
2) Then, in response to this increase of viremia, the immune system is activated. The number of antibodies against viral peptides increases. This is the humoral response.
3) The number of activated T Helpers and T cytotoxic increases simultaneously. This is the cell-mediated response.
During this asymptomatic phase, activated cells in response to HIV virus are at their highest level. The consequence of this extremely powerful response is a decrease in viremia. But somehow, and nobody knows so far. The immune system is overwhelmed and does not respond anymore. There is nomore humoral and cell-mediated response resulting in the increase of the viremia and the death of the infected patient.
I am now going to tell you how to control the life cycle of the HIV virus. Comment fig p1257, Science Review. I am using this complex figure to make my point but I do not expect you to memorize everything. I want to show you that there are indeed several ways to control the life cycle of the HIV virus, that scientists are making a lot of progress and that an ideal treatment would be the combination of several drugs that act at different levels in the life cycle to increase the chance of controlling efficiently the life cycle.
This figure is divided in 3 parts. The central part is a schematic of the life cycle. On the left are mentioned the different steps in the cycle with numbers. On the right are mentioned the different drugs that interfere with the cycle with corresponding numbers. You can see that for each step, there is one or several corresponding drugs. This is a very optimistic statement. I am now going to focus on the drugs currently used in therapeutics today.
1) The most common drug used is AZT (ziduvidine). This molecule acts at the reverse transcription step when the single stranded viral RNA molecule is copied into a DNA molecule by the viral reverse transcriptase. The AZT is a nucleotide mimic, it incorporates into the DNA during the process of reverse transcription. Then, the reverse transcriptase is not able anymore to go further in the process of the DNA molecule elongation. The result is a stop of the reverse transcription. No DNA synthesized, no new viruses released into the blood. There are also 2 others similar drugs used in therapeutics: ddI and ddC. I do not want to sound pessimistic but the sad reality is that these drugs are poorly effective and not represent a major hope. The paradox is these drugs is that there are more efficient during the terminal phase than during the asymptomatic phase. It is why scientists had developed an other type of drug.
2) Inhibitors of viral protease. You may have heard about them because the CNN channel made them public a month ago. At the end of the life cycle of the virus, viral proteins are synthesized from the mRNA. The first proteins to be synthesized are longer than required in their final forms. The protease cut them in smaller, functional proteins able to form a new virus. If this cutting process is inhibited, the new viral particle is malformed and then noninfectious. This approach seems to be very promisive.
But once again, the ideal scenario would be to find the perfect combination of drugs in order to control the life cycle at different stages and then increase the chances of interfering with the formation of new viruses. Mention soluble CD4
B. Vaccines Fig p1261 Science Review
An other way to approach HIV treatment is to develop vaccines against the HIV virus. Scientists made a lot of progress but are still trying to come up with a strategy in the elaboration of an efficient vaccine. They are confronted to a very challenging problem. How to make an efficient vaccine using existent methodology when they have to face the HIV virus. Almost all effective antiviral vaccines are based on two approaches:
1) The "live attenuated" strategy uses the entire virus but in a weaken form. Such viruses are made by gene deletion. In a case of the HIV virus, this approach has been rejected because of the retroviral nature of the virus. During its proviral form, such deletion into the viral genetic material can induce cancer.
2) The "whole killed" approach consists in killing the virus by eating it, by chemical treatments or by irradiating it. The resulting virus has still its glycoproteins at the surface but not able to integrate the cells. This second approach has also been rejected because there is always a possibility than not every single viral particle was killed. So, scientists found this strategy too dangerous.
The problem is to find a vaccine that is effecient and completely inoffensive for the immune system. This usually simple approach becomes very complicated when we have to face an HIV vaccine.
3) Scientists are now working on an alternative strategy where they selectively choose peptides from the surface glycoproteins that are recognized as antigens by the immune system. They however will have to face an other problem: What region to choose? One particularity of the HIV virus is its extremely rapid ability to mutate. For example someone who has been infected by a given strain of HIV could later on produce an other type of HIV. The diversity is not only from person to person but also within a given individual. The efforts are concentrated in finding constant and similar region for all the HIV virus forms.