HIV/AIDS Paul R Earl Facultad de Ciencias Biológicas Universidad Autónoma de Nuevo León San Nicolás, NL 66451, Mexico

1. HIV/AIDSPaul R EarlFacultad de Ciencias BiológicasUniversidad Autónoma de Nuevo LeónSan Nicolás, NL 66451, Mexico

2. AIDS is an infection of the immune system produced by HIV (human immunodeficiency virus). Every day, 15,000 humans are newly infected. The decrease of CD4 plus T lymphocytes well below 1000/ml (microliter) can indicate AIDS. In fact, less than 200 CD4+T /ml is defined as having AIDS. CD4+ cells are sometimes called T helper cells. This retroviral viremia may take 10 years to develop the illness called AIDS, and therefore the victim may not know he or she is infected and potentially able to spread AIDS. It also spreads from mother to baby.White blood cells = leukocytes = lymphocytes. Most are CD4+ and CD8+ T cells. T stands for matured in the thymus gland. B stands for bone marrow. Retrovirus is in reverse. Not DNA to RNA, but the opposite. RT means reverse transcription.

3. HIV testing is recommended for people who have had unprotected (did not use a condom) anal, oral or vaginal sex. HIV counseling and testing is also recom-mended for people who have tuberculosis, or a sexually transmitted disease like gonorrhea or attend a drug-treatment clinic, and others that have had multiple partners and had unprotected anal, oral or vaginal sex, or are partners of injection drug users. People often decide to get tested for HIV because they are worried about a recent contact that may have put them at risk. Latex condoms, when used consistently and correctly, are highly effective in preventing transmission of HIV.

4. Facing an HIV test may not be easy, but it is worth it. Today, people who are HIV positive stay healthy longer with treatment, especially if they act early. Antiretroviral drugs are relatively new, although known for over 20 years. Also, knowing your status allows you to take steps to protect others from infection. Having symptoms like swollen lymph glands, night sweats and weight loss might mean infection with HIV.

5. You may be interested in:Adherence to Antiretroviral Therapy in Adults: A Guide for Trainers.Presents guidelines for use in training health workers, including physicians, clinical officers and nurse counselors, in antiretroviral service delivery. Developed in Kenya.http://www.popcouncil.org/horizons/arvadhrnctrngmnl.html Adherence to Antiretroviral Therapy Acceptability of a Modified Directly Observed. Summarizes findings to understand client and health worker acceptability of different followup approaches to foster adherence to ARVs, the potential barriers to the approaches, and issues of disclosure and stigma.http://www.popcouncil.org/pdfs/horizons/mombasaform.pdfAIDSQuest: The HIV/AIDS Survey Library (updated June 2004). A compilation of surveys and questionnaires.http://www.popcouncil.org/horizons/AIDSquest/index.html

6. Where did HIV come from so suddenly ?HIV-1 was discovered in 1983-84 and fostered the Robert Gallo--Luc Montagnier debates. See Science 298 29 Nov 2002.The source of HIV-1/AIDS is chimpanzees, Pan troglodytes and other subSaharan monkeys. However strictly speaking, chimpanzees are apes. Species cross-infections occurs in subSaharan Africa. When this first involved man is unknown. Furthermore, the molecular clock needs repairs so 'when' cannot be found.

7. Who is infected with AIDS ?Although men who have sex with men--half the male cases--and injection drug users in the US are still the largest proportion of new HIV infections and AIDS cases each year, women, youth and black people are at higher risk now. Five million Americans are at high risk for HIV infection. Women make up nearly one-third of new HIV infections. In 1992, women accounted for 13.8% of people living with AIDS 5; yet by the end of 2001 that percentage had increased to 21%. One social issue is women's rights, especially in Africa & Asia, and their supposed greater susceptibility to AIDS.

8. About 27 nations, mostly African, have disasterously high levels of AIDS. The 5 most infected are 1/ Botswana, 2/Zimbabwe, 3/ Lesotho, 4/ Swaziland and 5/ Namibia. Asian countries are second to be infected: 1/ Thailand, 2/ India and 3/ Myanmar. In the New World, 1/ Haiti, 2/ Guyana and 3/ the Bahamas are prominently infected. For national demography, 2 indexes that seem applicible are Lx100/P = Living with HIVx100/Population in percent, and Dx100/L = Dead/Living in percent. Both are simple very high linear correlations that are lawlike.

9. Twenty countries with LP and DL not having reasonable correlations.

10. The pathogen: virus HIV-1/2Virions enveloped; slightly pleomorphic; spherical; 80-100 nannometers (nm) in diameter. Surface projections of envelope small (surface appears rough), or distinct (8 nm long glycoprotein); spikes; dispersed evenly over all the surface. Capsids isometric to spherical, or rod-shaped or a truncated cone. HIV is purified by ultracentrifugation and found in the band for retroviruses.The genes of HIV-1 encode for 9 protiens in 3 classes: 1/The major structural proteins, Gag, Pol and Env, 2/The regulatory proteins, Tat & Rev, and 3/ The accessory proteins, Vpu, Vpr, Vif and Nef.

11. HIV structure after Milan Nermut

12. Histocompatibility molecules are glycoproteins (gp) present in the cellular membranes. Class I molecules serve to display antigens on the surface of the cell so that they can be recognized by T cells. The 3 different types of class I molecules designated: 1/ HLA-A, 2/HLA-B 3/ HLA-C. HLA means human leukocyte antigen. These differ only in their heavy chains. All share the same type of beta-2 microglobulin. The genes encoding the different heavy chains are clustered on chromosome 6 in the major histocompatibility complex (MHC). Human class II molecules are all HLA-D. The genes encoding them are also located in the MHC. CD8+ T cells are only able to respond to antigens presented by class I molecules.

13. The specificity of binding resides in a receptor for antigen: the B cell receptor (BCR) for antigen, and the T cell receptor (TCR) respectively. Both BCRs and TCRs share the properties of these membrane proteins. They are present in thousands of identical copies exposed at the cell surface. They made before the cell ever encounters an antigen. TCRs are encoded by genes assembled by the recombination of segments of DNA and have a unique binding site.

14. CD4+ T cells bind an epitope consisting of an antigen fragment lying in the groove of a class II histocompatibility molecule like a hotdog. CD4+ T cells are essential for both the cell-mediated and antibody-mediated parts of the immune system: The role of the CD8+ T cells is to monitor all the cells of the body, ready to destroy any that express foreign antigen fragments in their class I molecules.

15. The attack on the immune systemWhen the number of CD4+ T lymphocytes drops below 400-500/ml due to destruction by HIV, the ability of the patient to mount an immune response declines. The patient become hypersusceptible to miscellaneous pathogens, especially viruses and fungi, that normally inhabit our tissues. Eventually the patient likely dies of these opportunistic infections.Some other infected lymphocytes selfdestruct, i. e., commit suicide. The halflife of some of these cells can be less than a day.

16. DiagnosisEstablishing HIV serology or the diagnosis of HIV infection in the blood, is recommended for several categories of people, including: 1/ people with other sexually transmitted diseases, 2/ people in "high risk" categories, including men who have sex with men, 3/ injection drug users, 4/ men and women who have had unprotected sex outside of a mutually monogamous relationship, as well as the sexual partners, 5/ people who have or have had sex with multiple partners, 6/ persons who think they are at risk, 7/ pregnant women, 8/ people with active tuberculosis,

17. Diagnosis depends on ELISA that then can be corroborated by Western blot.HIV-1 ELISA (enzyme-linked immunosorbent assay) is usually the first test performed on potentially HIV-infected blood. Results may be positive, negative or undetermined. ELISA is a sensitive, yet unspecific test therefore a positive ELISA must be confirmed by a positive Western blot test, which is more specific but less sensitive. ELISA usually measures the presence of antibodies against HIV, not the virus itself. Western Blot measures viral protein bands like p24.

18. After exposure to HIV, a period of time known as the window period elapses before a measureable amount of antibody is produced (seroconversion). This period rarely lasts longer than 6 months. False-negative results may occur during the window period. HIV-1 tests will detect antibodies to most variants of HIV-1 except for rare subtype O. ELISA will detect 80 % of people with HIV-2 infection, but Western blot test results can be indeterminate. HIV-2 tests and a combined HIV-1/HIV-2 test are available.

19. Another diagnostic system is enzyme immuno assay (EIA). See Lisse et al, Scand J Immunol 1997, 45: 637-644. EIA is sometimes abbreviated (IA) as the immunoalkaline phosphatase method. EIA gives higher counts of CD4+cells and lower of CD8+ T cells than FC. For EIA, monoclonal antibodies against the CD4+ and CD84+ cell receptors were used. Postive control blood smears with known percentages of T cells were included with each sample. Lymphocytes are counted to 200 in triplicate for each case. Persue also Erber WN, Pinching AJ & Mason DY. Lancet 1984, May 12: 1042-1046, using monoclonal antialkaline phosphatase and Fast Red counterstaining with hematoxylin and mounting in Apathy's aqueous medium.

20. More HIV pathologyThe progressive depletion of CD4+ T lymphocytes is the cardinal event in the pathogenesis ofinfection by HIV.HIV infects lymphocytes and produces fresh crops of virus particles that leave the cell, often killing it in the process. The new virus particles spread to new target cells. Killer cells: NK, CD8+, try to stop this viral replication.

21. CD4+ and CD8+ countsThe techniques for lymphocyte counts and how many copies of HIV are in the plasma or blood are indeed sophisticated and thereby EXPENSIVE. They are flow cytometry (FC) and the DNA polymerase chain reaction (PCR). When used for RNA it has reverse transcriptase abbreviated RT-PCR. FC can analyze 12 fluorencent substances at one time. Over 30,000 of these FC instruments are in circulation. Still, the instrument costs over $50,000.

22. The blood-cytosphere mixture is added to a red cell lysing solution (2% acetic acid and 0.025% crystal violet stain), shaken, and pipetted onto a 0.1-mm-deep hemacytometer. With a light microscope at 340 magnification, lymphocytes with 4 or more CD4 1 or CD8 1 cytospheres attached to their surface are counted, while all others are excluded.

23. The single approach is to CD4+ T only. Some are ’volumetric’ (Cytoron, Ortho 3 ; Galaxy, DAKO) and can determine volumes directly. Others are ‘bead-based’’ (FACS-Calibur; BD; Elite, Coulter) and need known numbers of microbeads added to the tubes to calculate absolute counts. Percents from FC can be compared using a common automated blood counter. For much more on fluorescence activated cell sorters (FACS) and flow cytometry (FC), read Herzenberg et al., Clin Chem 48: 1819-1827, 2002.

24. Viral load testsThe amout of HIV or number of copies in blood is called the plasma viral load (PVL). Three different PVL tests can be used: 1/ The polymerase chain reaction (PCR) test, 2/ The branched-chain DNA (bDNA) test and 3/ The nucleic acid sequence-based amplification (NASBA) test. All of these tests work well.Changes in PVL are given in log to the base 10. A log change means 10 times more or 10 times less than a previous PVL test result. The number of copies can be in many thousands. Thus, logs are used. Therapy might be started when PVL is over 10,000. If therapy is in progress, several PVL tests a year monitor the status. If PVL goes up, medication needs to be changed, quite obviously. The goal is to clear detectable virus from the blood in 16-24 weeks.

25. The Amplicor HIV-1 Monitor Test (RT-PCR test) made by Roche Molecular Systems is FDA-approved for establishing HIV disease prognosis

26. Review of the pathogenisisOnce HIV has entered the body, the immune system initiates antiHIV antibody and cytotoxic T cell production. However, it can take 1-6 months for an individual exposed to HIV to produce measurable quantities of antibody.HIV enters the body and binds to dendritic cells which carry the virus to CD4+ T cells into lymphoid tissue establishing the infection.CD4+ T lymphocytes produce antibodies and CD8+ T cells police the population, killing defective lymphocytes. These 2 types make up most of the white cells.

27. Lymphocytes include T-cells and B-cells. T-cells are divided into CD4 and CD8 cells, both of which have learned to recognize antigens and distinguish them from tissues that belong to the body. They recognize an antigen only when it is shown to them by an APC. This is why the immunity they provide is called cell-mediated immunity. B-cells recognize antigens directly or with the help of T-cells. They produce and release antibodies in the plasma. This antibody production is called humoral immunity.

28. The process of activation may cause T cells that are not specific for HIV to become activated and undergo apoptosis (cell death). Many uninfected T cells die in HIV infected individuals. Peripheral lymphoid depletion is met by increased production of T cells from stem cells in the bone marrow. This process produces more actively dividing T cells which can be infected by HIV. Over time, the ability of the bone marrow to maintain increased T cell production is eroded, while mutations in the virus result in the evolution cytopathic variants that escape immunologic destruction. This impaired production results in the eventual collapse of the immune system.

29. What's new ? TreatmentEffective treatment--antiretroviral treatment (ART)--is still new. Antiretroviral drugs have changes AIDS from a purely fatalto a chronic disease.Treatment that prolongs life encourages people to get tested for AIDS. They know they can be treated, even hopefully in Africa. It also means that a low-priced counting system for CD+ T cells is crucial for following the disease state, and the evaluation of antiretroviral and antiprotease drugs.The final problem might be the high cost of drugs, e. g., zidovudine can cost over $175 a month.

30. There are currently 5 major classes of antiretroviral drugs in general use: 1/ nucleoside analogues, 2/ reverse transcriptase inhibitors (NRTIs), 3/ nonnucleoside reverse transcriptase inhibitors (NNRTIs), 4/ protease inhibitors (PIs) and 5/ fusion inhibitors. NRTIs function by inhibiting the synthesis of DNA by reverse transcriptase (RT), the viral enzyme that copies viral RNA into DNA in the newly infected cell. Nucleoside analogues bear a structural resemblance the nucleosides: purine nucleosides adenosine (A) and guanosine (G), and the pyrimidine nucleosides thymidine (T) and cytidine (C). Nucleoside analogues are triphosphorylated within the cell.

31. Nucleotide analogues resemble monophosphorylated nucleosides, and therefore require only 2 additional phosphorylations to become active inhibitors of DNA synthesis. RT fails to distinguish the phosphorylated NRTIs from their natural counterparts and attempts to use the drugs in the synthesis of viral DNA. When an NRTI is incorporated into a strand of DNA being synthesized, the addition of further nucleotides is prevented, and a full-length copy of the viral DNA is not produced. Rather than acting as false nucleotides, NNRTIs bind to RT to inhibit the enzyme's activity.

32. FDA also has approved a second class of drugs for treating HIV infection. These drugs, called protease inhibitors, interrupt virus replication at a later step in its life cycle. They include 1/ Ritonavir (Norvir), 2/ Saquinivir (Invirase), 3/ Indinavir (Crixivan). 4/ Amprenivir (Agenerase), 5/ Nelfinavir (Viracept) and 6/ Lopinavir (Kaletra). As HIV can become resistant to any of these drugs, a combination treatment can be used to effectively suppress the virus. When RT inhibitors and protease inhibitors are used in combination, it is referred to as highly active antiretroviral therapy, or HAART, and can be used by people who are newly infected with HIV as well as people with AIDS.

33. A number of drugs are available to help treat opportunistic infections to which people with HIV are especially prone. These drugs include 1/ Foscarnet and ganciclovir to treat cytomegalovirus (CMV)eye infections, 2/ Fluconazole to treat yeast and other fungal infections, 3/ Trimethoprim/sulfamethoxazole (TMP/SMX) and also 4/ pentamidine to treat Pneumocystis carinii pneumonia.

34. Protesters against AZT in Vancouver, British Columbia, Canada

35. What's nextA vaccine? Is the risk of infection high enough to warrant vaccine protection? Likely not except for African countries where an AIDS vaccine might be the only hope. What about eradication of HIV? Will condom use eventually eliminate HIV? This revolves around the risk and of course human behavior. What about all the other sexually transmitted diseases? One in 5 persons may have a hidden sexual disease. Which populations need AIDS protection? Again, the realistic protecter is the condom.The WHO-UNAIDS HIV Vaccine Initiative (HVI) was established in January 2000. Also, programs are conducted by the Vaccine Research Center (VRC) of the National Institute of Allergy and Infectious Diseases (NIAID).

36. An ideal vaccine against HIV-1 would induce neutralizing antibody levels which would provide sterilizing immunity. Still, the levels of antibody required may not be achievable by vaccination. What about escaoe viral mutants? The necessity of stimulating B cells that produce such neutralizing antibodies seems obvious.Live attenuated viruses have consistently provided protection against infection with SIV and SIV/HIV-1 chimeras in nonhuman primates. The safety requirement for the vaccine excludes this method with humans. Regardless, a few HIV-infected people never show signs of illness.

37. How can we ERADICATE HIV/AIDS ?The epidemiological role of HIV/AIDS is clear enough in developed countries, but the data for heavily burdened countries cannot be trusted.The HIV-1 mature envelope glycoprotein (gp) complex plays a pivotal role in the early events of virus attachment and entry into the target cell. Neutralizing antibodies found in the sera of infected individuals are primarily directed against the Env complex. The complex is arranged in a trimeric configuration of heterodimers, each consisting of a gp120 surface subunit noncovalently associated with a gp41 transmembrane subunit.