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

HIV/AIDS
Paul R Earl
Facultad de Ciencias Biológicas
Universidad Autónoma de Nuevo León
San Nicolás, NL
66451, Mexico
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.
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 drugtreatment 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.
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.
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.pdf
AIDSQuest: The HIV/AIDS Survey Library (updated June
2004). A compilation of surveys and questionnaires.
http://www.popcouncil.org/horizons/AIDSquest/index.html
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.
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.
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.
Twenty countries with LP and DL not having
reasonable correlations.
The pathogen: virus HIV-1/2
Virions 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.
HIV structure
after Milan
Nermut
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/ HLAC. 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.
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.
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.
The attack on the immune system
When 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.
Diagnosis
Establishing 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,
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.
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. Falsenegative 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.
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.
More HIV pathology
The progressive depletion of CD4+ T
lymphocytes is the cardinal event
in the pathogenesis of
infection 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.
CD4+ and CD8+ counts
The 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.
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.
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’’ (FACSCalibur; 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.
Viral load tests
The 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.
The Amplicor HIV-1 Monitor Test (RT-PCR test)
made by Roche Molecular Systems is FDAapproved for establishing HIV disease
prognosis
Review of the pathogenisis
Once 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.
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.
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.
What's new ? Treatment
Effective treatment--antiretroviral treatment
(ART)--is still new. Antiretroviral drugs have
changes AIDS from a purely fatal
to 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.
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.
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.
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.
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.
Protesters against AZT in Vancouver,
British Columbia, Canada
What's next
A 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).
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.
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.