Immunological Disorders and Tests

Important words and concepts from Chapter 18, Black, 1999 (3/28/2003):

by Stephen T. Abedon (abedon.1@osu.edu) for Micro 509 at the Ohio State University

Course-external links are in brackets

Click [index] to access site index

Click here to access text’s website

Vocabulary words are found below

(1) Chapter title: Immunological Disorders and Tests

(a) [immunological disorders and tests (Google Search)] [index]

HYPERSENSITIVITY

(2) Hypersensitivity

(a) Hypersensitivities are inappropriate immune responses to foreign material that is either within or in contact with the body

(b) Essentially, the body mounts a sometimes dramatic immune response against an otherwise harmless, or at least less-harmful substance, thereby doing more harm to the body in the course of the immune response than might have the original allergen

(i) Type I: Immediate hypersensitivity

(ii) Type II: Cytotoxic hypersensitivity

(iii) Type III: Immune complex hypersensitivity

(iv) Type IV: Cell-mediated Hypersensitivity (Delayed Hypersensitivity)

(d) [hypersensitivity reactions (Google Search)]

(3) Anaphylaxis (anaphylactic shock)

(a) Anaphylaxis is a general term used to describe the detrimental effect(s) associated with hypersensitivities

(b) Anaphylaxis may be localized (annoying but not life threatening) or generalized (systemic and life threatening)

(c) Anaphylactic shock is a generalized anaphylaxis characterized by a significant, life-threatening drop in blood pressure

(d) [hypersensitivity reactions (Google Search)] [index]

(4) Prophylaxis

(a) Prophylaxis refers to the protective effects associated with an immune response

(b) [prophylaxis (Google Search)] [index] [check this]

(5) Immediate hypersensitivity (type I hypersensitivity; allergy)

(a) Immediate hypersensitivity occurs following the production of IgE antibodies against typically otherwise-harmless foreign antigens (which are known as allergens)

(b) Type I sensitivities are allergies

(6) Allergen

(a) An allergen is an antigen, the exposure to which results in a hypersensitivity reaction

(b) Note that allergens are non-self (i.e., foreign) antigens

(c) Since hypersensitivity (e.g., immediate hypersensitivity) is the result of a kind of specific immunity, an individual must be exposed to the allergen at least once (to sensitize the individual by inducing B cells that produce specific IgE antibodies) before exposures (subsequently) result in an allergic response

(d) [allergen (Google Search)] [index]

(7) Histamine (degranulation)

(a) The signs and symptoms of immediate hypersensitivity are a consequence of the release of histamine and other chemical mediators from body cells

(b) In the case of histamine, release occurs when IgE antibodies bound to basophils or mast cells bind to allergens

(c) Histamine is found intracellularly within vesicles (the granules within these cells) and degranulation is the term used to describe the release of histamine via the fusion of these vesicles with the basophil or mast-cell plasma membranes

(d) (in addition to histamine, prostoglandins and leukotrienes are reaction mediators that play important roles in mediating airway constriction)

(e) See Figure 18.1, The mechanism of immediate (Type I) hypersensitivity, or anaphylactic hypersensitivity

(f) [histamine, degranulation, degranulation and histamine (Google Search)] [index]

(8) Cytotoxic hypersensitivity (type II hypersensitivity)

(a) The term cytotoxic in cytotoxic hypersensitivity refers to host-cell damage caused by an over-zealous immune response

(b) Recall that a normal aspect of both specific and non-specific immune responses is extracellular killing, particularly the killing of host cells that are thought to be pathogen-infected

(c) Cytotoxic hypersensitivities are mediated by the binding of antibody's to body tissues which leads to the lysis of cells (either via ADCC or via the activation of complement)

(d) The negative consequences of not correctly matching blood types for transfusions are examples of the damaging effects of cytotoxic hypersensitivities (erythroblastosis fetalis is a related, additional example of a cytotoxic hypersensitivity)

(e) [cytotoxic hypersensitivity, type II hypersensitivity (Google Search)] [index]

(9) Immune complex hypersensitivity (type III hypersensitivity)

(a) One role of phagocytic cells (macrophages) is the removal of debris from body tissues (e.g., blood) and one kind of debris that results from specific immune reactions (specifically humoral immunity) are large complexes of antibody and antigen

(b) These complexes form as a consequence of the multivalent nature of both antibodies and antigens (i.e., an individual antibody molecule can bind to more than one epitope and thus, potentially, more than one antigen, while a large antigen or organism can display large numbers of individual epitopes)

(c) The phrase immune complex as in immune complex hypersensitivity refers to these antigen-antibody complexes, and type III hypersensitivity refers to an immune response that produces an excess of these immune complexes, particularly faster than macrophages (and the liver) can remove them

(d) The accumulation of these immune complexes can result in their depositing in otherwise healthy tissues followed by a damaging hypersensitivity immune response in those tissues to the not-engulfed immune complexes

(e) Certain autoimmune diseases (rheumatoid arthritis and lupus) are consequences of type III hypersensitivities as well as the serum sickness that results from a second exposure to an antitoxin

(f) [immune complex hypersensitivity (Google Search)] [index]

(10) Cell-mediated hypersensitivity (type IV hypersensitivity, delayed hypersensitivity)

(a) Cell-mediated hypersensitivity is mediated by T lymphocytes (rather than by antibodies)

(b) Cell-mediated hypersensitivity is also known as delayed hypersensitivity because the time between exposure to the eliciting antigen and the occurrence of symptoms can take many hours

(c) A common example of type IV hypersensitivity is poison ivy sensitivity (where, of course, the rash appears only after many hours—e.g., next day—following exposure to the poison ivy urushiol, the triggering oil)

(d) [cell-mediated hypersensitivity, delayed hypersensitivity (Google Search)] [index]

IMMUNODEFICIENCY

(11) Immunodeficiency

(a) Immunodeficiency is characterized by an inadequate immune response, either in general or against specific antigens or pathogens

(b) This inadequacy contrasts with the temporary inadequacy of specific immunity as immune responses normally develop following first-time exposure to antigens

(c) Instead, immunodeficiency is characterized by an abnormally under response to antigens over the long (as well as the short) term and is indicated by a weakness in the ability of the body to fight legitimate pathogens

(d) We may speak of immunodeficiencies as being either inborn (primary) or acquired (secondary)

(e) Things that can lead to acquired immunodeficiencies include:

(i) Drugs (e.g., anti-cancer chemotherapies)

(ii) Pathogens (e.g., HIV/AIDS)

(iii) Inadequate nutrition and injury

(iv) Some cancers

(f) [immunodeficiency -AIDS (Google Search)] [index]

(g) Extreme exposure to sunlight that comes from maintaining a deep tan can also lead to pathogen-fighting inadequacies [impacts of UV radiation on the globe today (UV Rays and Global Changes)] [the ultraviolet light in sunlight can also stimulate herpes infections and might stimulate HIV infection (AIDS Treatment News)… and other infections (UV Rays and Global Ghanges)] [safe sun? (MicroDude)] [index]

(12) Cyclosporin

(a) Cyclosporin is a transplant anti-rejection drug that intentionally serves to induce a highly specific immunodeficiency

(b) That is, cyclosporin interferes with cell-mediated immunity , which is one of the mechanisms by which organ-transplant rejection occurs

(c) Unfortunately, cell-mediated immunity is important in fighting viral infections, serving as the means by which virus-infected cells are destroyed by the immune system; consequently, individuals on a cyclosporin regimen are more susceptible to viral infections

(d) This immunosupression is not complete, however (i.e., the rest of the immune system still functions), thus allowing the benefits of the drug (significant boost in transplantation efficacy since it greatly reduces the need to type-match tissues) to outweigh the costs (increased susceptibility to viral infections)

(e) In addition to viruses, cyclosporin increases tumor risks, an observation that is consistent with the tumor-fighting role of cell-mediated immunity, but, apparently, may also be a consequence of cyclosporin actually promoting the growth of certain tumors [Nature review on cyclosporin and TGF Beta (Biocognizance.com)]

(f) To prevent the rejection of transplanted organs, organ-transplant recipients must remain on a cyclosporin regimen for life

(g) [cyclosporin (Google Search)] [index]

(13) Acquired Immune Deficiency Syndrome (AIDS)

(a) The most-popularly understood cause of immunodeficiencies is, of course, AIDS, which is an immunodeficiency brought on by the infection with the Human Immunodeficiency Virus (HIV)

(b) (note that AIDS typically stands for acquired immunodeficiency syndrome as well as the immune deficiency phrase used in your text; a Google search for "acquired immunodeficiency syndrome" gives 79,800 hits on 3/14/02 while a Google search on the same day for "acquired immune deficiency syndrome" gives 54,600 hits)

(c) Immunodeficiency caused by HIV occurs because this virus preferentially infects host immune system cells, specifically those that carry the antigen that designates T lymphocytes as helper T lymphocytes (but the same antigen also is carried by macrophages and other cell types)

(d) HIV ultimately kills the cells it infects (e.g., via cell-mediated immunity by the body against HIV-infected cells); this creates a constant drain on the number of helper T cells present in the body, which in turn interferes with the functioning of both the cell-mediated and the humoral arms of specific immunity

(e) The virus is always replicating and the body is always fighting off the virus, with the virus mutating to evade specific immunity (more scientifically stated, with mutationally generated evavion-capable HIV variants are selected by specific immunity), and the specific immunity of the body must periodically produce new primary immune responses against the new variants of the virus

(f) Thus, HIV infection is characterized by

(i) an initial (~6 week) period of flu-like disease before specific immunity brings the infection under control

(ii) a steady-state period during which viral replication is kept more-or-less under control, with some break outs of viral replication as immune-system evading virus variants arise (this steady state can occur over many years, usually <10)

(iii) a gradual decline in immune system resilience and functioning until the growth of newly arising virus variants is no longer successfully brought back under immune-system control (AIDS)

(g) See Figure 18.22, CDC classification of HIV disease and AIDS

(h) The immunodeficiency characterized by AIDS is actually only the end-product of a long decline in immune system functioning and represents only the end stage of a typically decade-long disease process; that is, not all individuals who are HIV infected have AIDS (though all people with AIDS are HIV infected), but most people who are HIV infected (95%+), who are not successfully treated using modern antiviral chemotherapeutics, will eventually succumb to AIDS

(i) As a further complication, note that most HIV-infected people do not die with AIDS as a direct cause, but instead from secondary infections that are brought on the increases in susceptibility to infection that results from immunodeficiency

(j) Various external links: [index]

(i) [AIDS (Google Search)]

(ii) [The AIDS Knowledge Project]

(iii) [AIDS lectures: (1) definitions, origins, and prevalence, (2) the virus, (3) HIV disease and therapy, (4) the human immune response, (5) the biology the stages of HIV disease, (6) how is HIV transmitted? (7) preventing HIV transmission, (8) HIV testing, (9) AIDS and social issues (University of Michigan Bio 118)]

(iv) [does HIV prevention work? (JAMA HIV/AIDS Information Center)]

(v) [early impact on HIV infection, effects of treatment (JAMA HIV/AIDS Information Center)]

(vi) [the origin of AIDS (HIV InSite)]

(14) Human Immunodeficiency Virus (HIV)

(a) HIV is a plus-stranded, diploid, single-stranded RNA virus

(b) HIV is an enveloped virus that derives its envelope from the host-cell plasma membrane

(c) Also as part of the maturation of an HIV virion the virus envelope proteins are formed via the proteolytic cleavage of a precursor (larger) protein (without this cleavage the resulting virus particle is not functional and it is this cleavage that is blocked by anti-HIV protease inhibitors)

(d) HIV is a retrovirus that employs the enzyme reverse transcriptase to process its single-stranded RNA genome into a double-stranded DNA genome

(e) This double-stranded DNA genome is then inserted into a host chromosome

(f) See Figure 10.13, Replication of RNA viruses

(g) Not all inserted genomes are immediately active, thus allowing some virus-infected cells to evade immune system recognition (as well as drug treatment) over long periods (years, perhaps decades) thus making it nearly impossible to cure an HIV infection

(h) There are two major groups of HIV viruses in circulation among humans, HIV-1 which is probably derived from a chimpanzee virus (the revenge of the chimpanzees, who probably passed on the virus to humans as “bush meat”) and HIV-2 which is probably derived from a monkey virus (one kind of SIV or simian immunodeficiency virus) (ditto re: the revenge of…) [Nature on HIV origin (Biocognizance.com)] [the AIDS pandemic is new, but is HIV new? (Systematic Biology)]

(i) HIV-1 is by far the more prevalent (in the U.S.) and the more virulent of the two

(j) [HIV (Google Search)] [anti-HIV strategies (and additional HIV information) (Biocognizance.com)] [index]

(15) HIV epidemiology

(a) HIV/AIDS is a pandemic disease with estimates of world-wide cumulative prevalence (i.e., including those that have died—so far a minority) as high as 50 million people or more

(b) HIV is transmitted via body fluids such as semen and blood

(i) During unprotected vaginal intercourse (the prominent route of transmission in sub-Sahara Africa) or during anal intercourse (in both cases the recipient is the more susceptible to infection)

(ii) From needle sharing during intravenous drug use

(iii) From the transfusion of blood or blood products (rare since the implementation of immunological testing of the blood supply)

(iv) From mother to child either in utero, during passage down the birth canal, or from breast milk

(d) “It is not possible to acquire the HIV virus by donating blood because new, sterile needles are used.”

(e) Health-care workers should observe universal precautions to avoid exposure to blood-borne pathogens including HIV

(f) [HIV epidemiology (Google Search)] [index]

(16) HIV vaccination

(a) Difficulties in developing vaccines: (not responsible for material under this subheading, i.e., subheading (a))

(i) While from a public health point of view vaccines are wonderful things, in practice it is not necessarily easy to engineer effective vaccines against a given disease

(ii) Reasons that vaccine development is not always a fruitful endeavor can include:

· limited range:

a given vaccine tends to be effective only against individual serovars of pathogen species (some species have hundreds of serovars—a serovar is a strain that is differentiated from other strains of a given organism by serological means)

· disease isn't immunizing:

for some pathogens even exposure to disease (the ultimate form of immunization) does not confer active immunity

· rapid evolution:

development of vaccines against particularly rapidly evolving pathogens (such as HIV) is also difficult because the pathogen, essentially, is an immunologically moving targets—at best such vaccines are rapidly made obsolete by pathogen evolution (e.g., anti-influenza vaccines)

· exacerbation of disease:

vaccines of certain types, against certain pathogens can actually exacerbate disease when it occurs

· cause of disease:

live vaccines retain at least some potential for causing the disease they are charged with preventing; this is especially true with regard to immunodepressed individuals (e.g., live polio vaccine)

· cost-benefit problems:

successful vaccine delivery is not always economically or politically justifiable

(b) In the mid-to-late 1980s optimism was high that an anti-HIV vaccine could be rapidly developed. This optimism was based on the premise that molecular techniques in biology had advanced so far that the development of a molecular (recombinant, subunit) vaccine against any pathogen was possible given the application of sufficient resources.

(c) Stemming from this optimism the more-easily developed whole live or killed vaccine strategies were rejected as too dangerous:

(i) a live HIV could infect indefinitely, possibly reactivated as a pathogen given future host immunodepression

(ii) a dead HIV might not be completely dead, or completely harmless given subsequent exposure to living HIV

(d) However, it turns out that HIV possesses many of the qualities that would lead one to predict difficulty in vaccine development:

(i) There exists numerous and extensive serological variation among wild isolates (contrast polio for which only three serovars are known).

(ii) We lack data on having the disease actually being immunizing; after all, HIV-infected individuals successfully control their infections for years without actually eliminating the infection, nor staving-off disease

(iii) HIV is the poster child for rapidly evolving pathogens; almost nothing else mutates or evolves faster than HIV

(iv) See immediately above discussion of the dangers of whole vaccine use

(e) Even if a disease-preventing vaccine existed, how many of us would volunteer to receive a vaccine that

(i) By definition made us HIV seropositive (if not necessarily HIV infected)

(ii) That could prevent (in most cases) the progression of an HIV infection towards AIDS, but could not actually prevent the occurrence of an HIV infection

(iii) Possessed a less-than full (<100%) potential to prevent the progression of infection towards AIDS

(f) Furthermore, consider that those who are most at-risk for HIV infection (in the U.S., at least) are also the same individuals (with the likely exception of upper- and middle-class homosexuals) who are most-likely to fall through the cracks of health-care systems and therefore the least likely to be vaccinated

(g) An effective anti-HIV vaccine may never arrive, and when it does it may not be able to achieve its promise for bringing the HIV pandemic under control (especially if vaccination serves as a signal to everyone and their mother to go out and have unprotected sex with multiple partners)

(h) In the mean time the best we can do is to live an HIV-defensive life and, as a society, to vigorously protect our tissue (e.g., blood) supplies through vigorous serological screening for HIV infection

(i) [HIV vaccination (Google Search)] [HIV biology, vaccine-strategy emphasis (Bio 160: Vaccine Development)] [index]

IMMUNOLOGICAL TESTING (SEROLOGY)

(17) Immunological tests (serology)

(a) A variety of experimental methods exist that employ immunological reagents, particularly antibodies

(b) These tests are typically employed

(i) as a means of testing for the presence of certain antigens in experimental unknowns

(ii) for quantifying the presence of specific antigens

(iii) as a means of detecting the anigen’s owner (e.g., a pathogen), or

(iv) as a means of characterizing an immune response (including detecting and quantifying antibodies)

(c) Because the majority of these tests employ antibodies (as opposed to cell-mediated immune responses) and since the crudest and earliest-to-be-worked-with antibody-containing reagent is serum (i.e., the liquid portion of blood once clotting has taken place), the study and development of such tests is called serology

(d) Here, for the sake of brevity, we will consider in depth two of the more-modern serological methods, the ELISA and the Western blot, particularly to enhance our understanding of how laboratories assay for HIV seroconversion

(e) [immunological tests, serological tests, serology (Google Search)]

(18) Serum

(a) Whole blood may be allowed to clot upon exposure to air (oxygen); if one removes the solid portion of the clot (using centrifugation), the remaining liquid is called serum

(b) Serum contains large quantities of proteins including high concentrations of antibodies

(d) [serum (Google Search)] [index]

(19) Seroconversion

(a) Seroconversion is the production of antibodies following exposure to an antigen

(b) The production of specific antibodies can be used as a diagnostic for previous exposure to specific antigens (e.g., HIV)

(20) ELISA

(a) The ELISA technique (which stands for Enzyme-Linked Immunosorbent Assay) is a method by which tagged antibodies are used to visualize specific proteins

(b) This immunological technique is very powerful because, by varying proteins and antibodies, it allows a rapid detection of very specific proteins or antibodies

(i) Binding of a substance, such as a protein or a specific antibody, to the plastic well of a assay plate

(ii) Washing excess (unbound) substance from the well

(iii) Blocking unbound plastic with an otherwise inert substance (such as skim milk—and then washing)

(iv) Probing with a substance that binds to the first substance (e.g., an antibody to the bound protein or a protein to the bound antibody—and then washing)

(v) Probing with a substance that is linked to an enzyme (this second probe can be done simultaneously with the first probe, e.g., an enzyme-linked antibody as the only probe—and then washing)

(vi) Addition of substances that undergo a color reaction in the presence of the enzyme tag

(vii) Only if all of the steps work in this assay (e.g., protein bound to plastic followed by antibody bound to protein followed by enzyme-tagged antibody bound to the first antibody) will the color reaction occur since washing removes all unbound reagents from the reaction well

(viii) A positive color reaction thus is used as a test for the presence in an experimental unknown of the presence of one of the necessary components (e.g., a specific protein or a specific antibody); see a microtiter plate used for ELISA with color reaction increasingly intense going from bottom to top à

(d) See Figure 18.34, Enzyme-linked immunosorbent assay (ELISA) is a modification of RIA

(e) The use of the ELISA technique is extensive in microbiology and immunology, in both the clinic and research, but it is perhaps best known as the primary means by which people and blood are tested for HIV seroconversion (as illustrated below)

(f) [ELISA (Google Search)]

(21) HIV antibody test

(a) The ELISA commonly employed to test for HIV seroconversion specifically is at test for the presence of anti-HIV antibodies in blood

(b) The ELISA consists of (second Roman numerals are from general ELISA description above)

(i) (i) partially purified HIV antigen is bound to plastic (picture above right.. not albumin employed to block plastic)

(ii) (iv) patient serum is used as the antibody probe (middle, right)

(iii) (v) enzyme-linked anti-human antibody is then used to probe for the presence of bound serum (bottom, left)

(iv) the enzyme-linked antibody remains bound in a well only if the patient’s serum contains antibodies that bind to HIV proteins, and for the most part an individual will possess anti-HIV antibodies (of sufficient titer) only if they have been infected with the HIV virus

(c)

(d) “Generally, serum antibodies to HIV can be detected by indirect ELISA within 6 weeks of infection.” (p. 529)

(e) If this ELISA test indicates seroconversion (i.e., the presumed presence of anti-HIV antibodies; bottom, right in above illustration), then a second, more rigorous test is employed to rule out false positives (that is, tests that falsely indicate HIV seroconversion)

(f) The more rigorous method typically employed is a Western blot assay

(g) [HIV antibody test (Google Search)] [index]

(22) Western Blot

(a) History (not responsible for material under this subheading, i.e., subheading (a))

(i) There exists a series of gel-based blotting methods known as the Southern Blot, the Northern Blot, and the Western Blot (the originator was named Southern and scientists being scientists subsequent blotting methods were named within this tradition)

(ii) The Southern Blot separates DNA on a gel (different sizes migrate at different rates) and probes with DNA (e.g., radioactive DNA); the tagged DNA is the visualizer of the DNA in the gel so only those gel DNAs (bands) that are probed for are visualized

(iii) The Northern Blot separates RNA on the gel and probes with DNA

(b) The Western Blot separates proteins on the gel (producing a protein profile) and probes with antibodies

(c) Those antibodies are labeled for example with radioactive elements (or various enzymes) [Western blot chemiluminescence reagents (NEN Life Sciences Products]

(d) In the case of HIV testing, the proteins on the gel are HIV proteins and the antibodies come from the serum of individuals; similar to variations on the ELISA technique, these human antibodies, if they bind HIV proteins, are visualized by labeled anti-human antibodies

(e) See Figure 18.36, Western blotting test for HIV antigens in blood (note how blotting techniques get their name from the transfer—blotting—of, in this case, proteins in the gel to a non-gel material, which is the material that is probed with the labeled materials)

(f) The nice thing about Western blotting is that the results can be very specific where protein bands are only visualized if the antibodies bind the protein, and then only those protein-bound antibodies are then visualized (e.g., a mixture of all of the proteins in a cell could be probed with a single monoclonal antibody that visualized only a single protein type from that mixture)

(g) However, the Western blot is also more time-consuming and expensive than the ELISA so is used in HIV testing only to further characterize ELISA positives

(h) [western blot assay (Google Search)] [index]