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Retroviruses. Groups of Retroviruses. Oncovirinae Tumor viruses and similar Lentiviruses Long latent period Progressive chronic disease Visna HIV Spumavirinae. important. important. Retroviruses known to cause human cancer. Human T cell lymphotropic virus -1 (HTLV-1)

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Groups of retroviruses
Groups of Retroviruses

  • Oncovirinae

  • Tumor viruses and similar

  • Lentiviruses

  • Long latent period

  • Progressive chronic disease

  • Visna HIV

  • Spumavirinae

important

important


Retroviruses known to cause human cancer

  • Human T cell lymphotropic virus -1 (HTLV-1)

  • Adult T cell leukemia, Sezary T-cell leukemia

  • Africa, Caribbean, Some Japanese Islands

  • Human T cell lymphotropic virus -2 (HTLV-2)

  • Hairy cell leukemia

  • HIV?


Hiv and aids
HIV and AIDS

  • Acquired Immunodeficiency Syndrome

  • Disease caused by an infectious agent:

  • a retrovirus


Hiv and aids an infectious agent

With giemsa stain at high magnification, the faint bluish dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

HIV and AIDSan infectious agent

In Los Angeles1967-1978 only two cases of Pneumocystis carinii pneumonia

All Homosexual

  • 1979 - 5 cases of Pneumocystis carinii


Hiv and aids1
HIV and AIDS dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

With dissemination to extrapulmonary sites, Pneumocystis carinii tends to produce foci with prominent calcification, as seen in the kidney here grossly.


Hiv and aids an infectious agent kaposi s sarcoma
HIV and AIDS dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavagean infectious agent – Kaposi’s Sarcoma

  • Early 1981 MMWR: 5 cases of Kaposi’s sarcoma

  • Hitherto: rare (immunocompromization)

  • 1981 - 26 cases of Kaposi’s sarcoma

    • Young

    • San Francisco and New York

    • All Homosexuals


Hiv and aids2
HIV and AIDS dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

Two rare diseases in the gay community linked to

IMMUNOSUPPRESSION

OPPORTUNISTIC INFECTIONS

Also

Lymphadenopathy

Hodgkin’s Lymphoma

  • Gay-Related Immune Deficiency

  • Acquired Immune Deficiency Syndrome (AIDS)


Hiv and aids3
HIV and AIDS dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

Distinguishing characteristics

  • Clusters of infected men

  • Apparent concentration within sexually interactive groups

  • High numbers of sex partners

Suggests an infectious agent


Hiv and aids4
HIV and AIDS dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

  • More evidence for an infectious agent:

  • Different ways of getting a similar syndrome

  • Blood transfusions

  • Intravenous drug use

  • Hemophilia (clotting factor)

Female sex partners of AIDS-positive IV drug users and hemophiliacs

Not just in the Gay community


Hiv and aids5
HIV and AIDS dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

Obvious agent: A virus……that is now in the blood supply

Primary route of transmission: Sex

AIDS is a sexually-transmitted disease


The cellular picture

HIV and AIDS dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

The Cellular Picture

Loss of one cell type throughout the course of the disease

CD4+ T4 helper cells

A fall in the CD4+ cells always precedes disease

In advanced disease the loss of another cell type

CD8+ cytotoxic killer cells

Suggests an infectious agent

A virus


AIDS Definition dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

  • AIDS is currently defined in persons older than 13 years as the presence of one of 25 conditions indicative of severe immunosuppression or

  • HIV infection in an individual with a CD4+ cell count of <200 cells per cubic mm of blood.

  • AIDS is therefore the end point of an infection that is continuous, progressive and pathogenic

  • With the prevalence of HIV in the developing world, HIV and its complications will be with us for generations


Hiv and aids the virus
HIV and AIDS dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavageThe Virus

  • The virus only grows on T4 cells that are proliferating in response to an immune stimulus

  • Therefore difficult to grow in culture

  • Robert Gallo : reverse transcriptase in activated T4 cells in blood of patients with AIDS : HTLV-3

  • Luc Montagnier: LAV

  • Human Immunodeficiency Virus (HIV)


Genosome
GENOSOME dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

  • Diploid Capped and polyadenylated

  • Positive sense (same as mRNA)

Viral RNA cannot be read as mRNA

New mRNA must be made

Virus must make negative sense DNA before proteins are made

Therefore virus must carry REVERSE TRANSCRIPTASE into the cell


The genome of hiv
The Genome of HIV dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

  • Three structural genes

  • LTRs

  • Extra open reading frames are clue to latency

  • These ORFs code for small proteins - antibodies in AIDS patients


A HIV has: dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

3 structure genes

GAG : internal proteins

ENV: Envelope glycoproteins

  • POL: Enzymes

  • Reverse transcriptase

  • Integrase

  • Protease (cuts polyproteins)


Hiv the virus

Membrane: host derived dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

Two glycoproteins: gp160 gp120 and gp41

gp41 is fusogen that spans the membrane

sugars: immunosilent

HIV - The Virus

vaccine problem


Hiv the virus1

p17: inner surface dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

p24: nucleocapsid

p9: nucleocapsid associated with RNA

HIV - The Virus

Group-Specific Antigens

GAG gene


The genome of hiv1
The Genome of HIV dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

Small non-structural proteins

mRNAs made by multiple splicing of genomic RNA

(c.f. mRNA for structural proteins)

  • EARLY

  • TAT: TransActivator of Transcription

  • REV: Regulator of Virion Protein Expression

  • NEF: Negative Regulatory Factor

  • LATE

  • VIF: Virion Infectivity Factor

  • VPU: Viral Protein U

  • VPR: Viral Protein R

TAT and REV are essential for HIV replication


Hiv life history
HIV - Life History dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

  • A retrovirus

  • Latency

  • Specific destruction of CD4+ cells

  • How does the virus enter the cell?


Hiv life history1
HIV - Life History dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

Entry into the cell

T4 (CD4+) cells are major target

Human HeLa Cell transfected with CD4 antigen

Human HeLa Cell

NOT INFECTED

INFECTED

But NOT the whole answer since this

does not happen if CD4 is transfected into a MOUSE cell


Hiv life history2
HIV - Life History dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

  • Fusion at ambient pH

  • No need for entry into lysosomes

  • Syncytia

Profound significance for AIDS progression

Profound significance for therapy


Hiv life history3

Co-Receptors dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

CD8+ Cells

MIP-1 alpha MIP-1 beta RANTES

Chemokines

Block HIV infection of macrophages

HIV - Life History

Why do CD4-transfected human cells become infected

but CD4-transfected mouse cells do not?

Human cells must possess a co-factor for infection that mouse cells do not


Hiv life history4
HIV - Life History dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

HIV

chemokine

Mutant CCR5

CD4

CCR5

CD4

CCR5

CD4

macrophage


Hiv and aids6
HIV and AIDS dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

Co-receptors

CCR5 is a chemokine receptor

25% of long term survivors are CCR5 or CCR2 mutants (deletions)

Many other chemokine receptors


Hiv life history5

Endocytosis dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

Fusion of membranes

Release of nucleocapsid to cytoplasm

Nucleus

HIV – life history


Hiv life history6
HIV - Life History dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

HIV carries with it:

  • Reverse transcriptase

  • Integrase

  • Protease

  • tRNA primer

HIV genes

GAG POL ENV

HIV has no oncogene but could still be oncogenic

vaccine problem


Hiv life history7

virus dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

virus

HIV – life history

RNA-dependent DNA Polymerase encoded by virus

REVERSE TRANSCRIPTASE

RNA genome

Reverse transcriptase

DNA genome

Integrase

Integrates

Host RNA polymerase II

RNA genome

host


Hiv life history8
HIV – life history dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

  • Parental RNA

  • RNA/DNA Hybrid

  • Linear DNA/DNA duplex

  • Circular Duplex DNA

  • Integration Replication (DNA genome in cell)

  • Transcription Viral RNA genome mRNA protein

Reverse transcriptase

Reverse transcriptase

Integrase

Host DNA polymerase

Host splicing enzymes

Host RNA pol II


Release of hiv
RELEASE OF HIV dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage


LIFE HISTORY OF dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

HIV


Hiv life history9
HIV - Life History dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

Latency - Cellular

Only activated T4 cells can replicate virus

Most infected T4 cells are rapidly lyzed but are replaced

Some T4 cells revert to resting state as memory cells which are long-lived

Memory T4 cells cannot replicate the virus unless the become activated

Macrophages do not show latency

Clinical Latency

HIV infection is not manifested as disease for years

During apparent clinical latency, virus is being replicated and cleared


Dynamics of cd4 t cells in an hiv infection

Chronically-infected memory T cells with provirus dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

Return to resting state

Infection

Long lived!

Reactivation

Long lived!

Uninfected unactivated memoryT cell pool

Dynamics of CD4 T cells in an HIV infection

Cell deathapoptosis etc

Uninfected activated T cell

Cell deathimmunedestruction

Adapted from Saag and Kilby Nat Med 5: 609, 1999


Latency
Latency dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

In the absence of any activating stimulus:

Homeostasis


Latency breaks

Immune response dot-like intracystic bodies of Pneumocystis carinii in lung are seen in this cytologic preparation from a bronchoalveolar lavage

T4 activated

HIV production

Latency Breaks

T4 resting




Hiv and aids9
HIV and AIDS disease

The cellular and immunological picture

The course of the disease

1. Acute Infection

  • High virus titer

  • Mild symptoms

  • Fall in CD4+ cells but recovers

  • Rise in CD8+ cells but recovers

  • A high virus titer (up to 10 million viruses per ml blood)

  • Macrophages infected

Macrophages bring HIV into the body if sexually transmitted


Hiv and aids10
HIV and AIDS disease

2. A strong immune response

  • Virus almost disappears from circulation

  • Good cytoxic T cell response

  • Soluble antibodies appear later against both surface and internal proteins

  • Most virus at this stage comes from recently activated (dividing) and infected CD4+ cells

  • CD4+ cell production compensates for loss due to lysis of cells by virus production and destruction of infected cells by CTLs


Hiv and aids11
HIV and AIDS disease

3. A latent state

  • Latency of virus and of symptoms

  • Virus persists in extra-vascular tissues

  • Lymph node dendritic cells

  • Resting CD4+ memory cells (last a very long time - a very stable population of cells) carry provirus


Hiv and aids12
HIV and AIDS disease

  • 10 billion HIV particles per day

  • Virus half life 5.7 hours

  • 100-10 million virions per ml blood (set point)

  • Small minority of T4 cells are infected

  • Virus found in lymph nodes


Hiv and aids13
HIV and AIDS disease

4. The beginning of disease

  • Massive loss of CD4+ cells

  • CD4+ cells are the targets of the virus

  • Cells that proliferate to respond to the virus are killed by it: Clonal deletion

  • Dendritic cells present antigen and virus to CD4 cells just as they are activated

  • Epitope variation allows more and more HIV to escape from immune response just as response wanes

  • Apoptosis of CD4+ cells

  • HIV patients with high T4 cell counts do not develop AIDS


Hiv and aids14
HIV and AIDS disease

5. Advanced disease - AIDS

  • CD8+ cells destroy more CD4+ cells

  • CD4 cell loss means virus and infected cells no longer controlled

  • As CD4+ cells fall below 200 per cu mm virus titer rises rapidly and remaining immune response collapses

  • CD8+ cell number collapses

  • Opportunistic infections

  • Death in ~2 years without intervention


Inexorable decline of cd4 t4 cells
Inexorable decline of CD4+ T4 cells disease

Why do all of the T4 cells disappear?

At early stages of infection only 1 in 10,000 cells is infectedLate 1 in 40

Of great importance to therapeutic strategy


Virus destroys the cell as a result of budding disease

But few cells are infected:

Early stage of infection 1:10,000

Late 1:40

HIV could kill sub population of precursor cells

People develop AIDS even when they have HIV that does not lyze cells

Why do all T4 cells disappear?

1. PUNCTURED MEMBRANE


Why do all t4 cells disappear 2
Why do all T4 cells disappear? - 2 disease

But syncytia not common

Most T4 cells are not HIV+

Could “sweep up” uninfected cells

Infected CD4 cell

Gp120 positive

Cells Fuse

Syncytia may be poor or ineffective at immune response

Uninfected CD4 cell

Gp120 negative

Killing of CD4 cells2. Syncytium Formation


Cytotoxic T cell disease

Why do all T4 cells disappear?

Killing of CD4 cells3. Cytotoxic T cell-mediated lysis

BUT: Most cells are not infected


Killing of CD4+ cells disease4. Binding of free Gp120 to CD4 antigen makes uninfected T4 cell look like an infected cellComplement-mediated lysis

Could account for the loss of uninfected T4 cells


Why do all t4 cells disappear
Why do all T4 cells disappear? disease

  • Apoptosis of T4 cells. Apoptosis of T4 cells is normal in clonal deletion to overcome autoimmunity

  • Also occurs with CD4 cells


CD8 cell disease(no CD4 antigen)

Macrophage

gp120

HIV

chemokine

G protein signal

?

?

Binding to CXCR4 results in expression of TNF-alpha on the cell surface

Binding to CXCR4 results in expression of TNF-alpha receptor II

CXCR4

Apoptosis of T cells


Death disease

CD8 T cell apoptotic bodies

CD8 cell

Macrophage

CXCR4


Macrophages may be infected by two routes

HIV disease

gp120

Anti-gp120

HIV

Macrophages may be infected by two routes

CD4

HIV gp120 binds to macrophage CD4 antigen

Virus is opsonized by anti gp120 antibodies which bind to macrophage Fc receptors - an enhancing antibody

Fc receptor

vaccine problem


Macrophages the trojan horse

Early HIV isolated during infection are diseasemacrophage tropic (have a macrophage chemokine co-receptor CCR5)

Virus probably infects patient via macrophages in semen

Infection by HIV leads to altered cytokine production “slim disease”

Slim disease very like Visna in sheep - also infects macrophages

Macrophages - The Trojan Horse

Macrophages form a reservoir outside the blood

Carry virus into different organs (brain)

Non-proliferating mature macrophages sustain HIV production for a long time without being killed by virus - - no latency


Population polymorphism
Population Polymorphism disease

HIV is a retrovirus

Retroviruses use host cell RNA polymerase II to replicate their genome

Pol II has a high error rate 1:2,000-10,000

HIV genome 9749 nucleotides

Therefore EVERY new virus has at least one mutation!

Every possible single mutation arises daily

1% of all possible double mutations arise daily

The HIV that infects a patient is very different from that seen by the time AIDS appears

vaccine problem


Population polymorphism1
Population Polymorphism disease

  • Initial infecting virus is macrophage-tropic (has CCR5 as co-receptor)

  • These are non-syncytium-inducing strains (INFECTIOUS)

  • (Note: most vaccines have been made against syncytium-inducing T4 cell tropic strains)

  • As virus mutates, it changes subtypes of cells that it infects as the ability to bind different co-receptors changes


Population polymorphism2
Population Polymorphism disease

  • Early in infection:

  • Macrophage-tropic

  • Non-syncytium-inducing

  • Slowly replicating

  • Late in infection

  • T4 cell tropic

  • Syncytium-inducing

  • High titer virus

vaccine problem


Population polymorphism3
Population Polymorphism disease

  • The most variable protein is gp120

  • Amino acid sequence within a single patient varies by 1-6%

  • Up to 30% in population

vaccine problem

  • Glycosylation masks conserved sites

vaccine problem

Co-infection may result in recombination

vaccine problem


Population polymorphism4
Population Polymorphism disease

  • Variation in reverse transcriptase leads to resistance to nucleoside analogs

drug problem

  • Variation in protease leads to resistance to protease inhibitors

drug problem

Polymorphism due to high mutation rate as a result of lack of proof-reading in reverse transcriptase and RNA pol II

Sub-populations arise with altered cell tropism


Other cells infected by hiv cd4
Other cells infected by HIV diseaseCD4-

  • Epithelial cells of bowel and vagina

  • Endothelial cells of brain

  • Brains cells : Astroglia, oligodendroglia

  • Galactocerebroside receptor


AIDS Statistics disease

  • Approximately 40,000,000 people in the world are HIV-infected

  • Approximately 8500 new HIV infections occur daily around the world

  • Over 90% of these are in developing countries.

  • 1000 are in children less than 15 years of age.

  • Of adult infections, 48% are in women and 15% in individuals 15-25 years.

  • As of June 2002, 793,026 Americans reported with AIDS. At least 457,667 of them have died. .

  • Prior to the introduction of combination therapies for HIV, AIDS incidence was increasing at a rate of just under 5% each year.


AIDS Statistics disease

  • Sub-Saharan Africa

  • About 1 million new cases of AIDS per year

  • 24 million people with HIV infection

  • AIDS is responsible for a decrease in life expectancy and increase in child mortality. Child mortality rates in East Africa will double by 2010 and adult life expectancy has already declined by 2 years in that region.

  • Several countries in sub-Saharan Africa report infection rates of 20-25%, especially urban areas.

  • Botswana: 35.8% of adult population infected

  • In Zambia, 1 in 5 urban girls is HIV-positive by the age of 20


Anti hiv strategies

  • Education disease

  • Sexually transmitted

  • Not highly infectious

  • Chemotherapy

  • Mutation selectionResistance

  • but

  • Suppress replication No capacity for mutation

Anti-HIV Strategies


Anti hiv strategies1
Anti-HIV Strategies disease

Highly

Active

Anti-

Retroviral

Therapy

HAART: Two nucleoside analog RT inhibitors and 1 protease inhibitor

Now also: Two nucleoside analog RT inhibitors and 1 non nucleoside


Does hiv cause aids
Does HIV Cause AIDS? disease

Single common factor between:

  • Gay San Franciscans

  • New York I.V. drug users

  • African heterosexuals

  • Hemophiliacs

  • Spouses of hemophiliacs and drug users

  • Children of hemophiliacs and drug users


Does hiv cause aids1

  • HIV diseaseprecedes AIDS in every population in which AIDS occurs

  • Infection by cloned virus

  • SIV HIV

  • Simian AIDS Human AIDS

Does HIV Cause AIDS?


Remember
Remember! disease

  • Education led to leveling off of rate of increase in AIDS

  • HAART has greatly slowed death rate

  • The fact that fewer people are dying per year from the infection means that the number of HIV-infected people in the population is rising!

  • Unless education continues to be successful and unless we can cure infected people of virus, the problem of virus spread is and will continue to be with us