Immune System Overview Mechanisms of Immunosuppression

Immune System Overview Mechanisms of Immunosuppression
By:Bob Luebke
Immunotoxicology Branch
Experimental Toxicology Division
NHEERL, ORD

Role of the Immune System in Homeostasis

* Bidirectional interaction with other systems
o Reproduction
o Endocrine
o CNS

Basics of Immunology
The Immune Response
Innate Immunity
Adaptive (Acquired) Immunity
-Phylogenetically ancient
-Limited recognition
-Rapid (minutes – hours)
- No cell proliferation required
-Limited memory (? mammals)
-First appeared in jawed fishes
- Infinite array of specificities
- Slow (days)
-Requires proliferation and differentiation
-Long-lasting memory

Basics of Immunology
* The adaptive immune response to antigen
Organs of the Immune System
Immune System Anatomy
Organs of the Immune System
Thymus: source of naive T cells
Fate of T Cells in the Thymus
Positive selection: optimal binding to self Ag prevents apoptosis
Negative selection: superoptimal binding to self Ag induces apoptosis
B cells: Tolerance to “Self”
Anergy: low expression of
IgM on surface; can’t bind Ag
Clonal ignorance: too few
copies of Ag in the periphery

Thymus size and architecture:
* May be very sensitive to xenobiotics
* Also sensitive to acute toxicity

Methods for Assessing Direct Immunotoxicity Associated with Exposure to Chemicals
Organs of the Immune System
Spleen: Antigen trapping and presentation, clonal expansion, cellular export
Organs of the Immune System
Lymph nodes: Antigen trapping and presentation, clonal expansion, cellular export
Cells of the Immune System
Innate Immune System: Granulocytes
Neutrophil (“PMN”)
* First responders
* Phagocytosis and killing of bacteria
* Inflammation

Eosinophil
* Allergy
* Killing parasite larvae
Basophil
* Circulating mast cells
* Allergy/anaphylaxis

Innate Immune System: Granulocytes
Neutrophil (“PMN”)
* First responders
* Phagocytosis and killing of bacteria
* Inflammation
Cells of the Immune System
Innate Immune System: Monocytes
Monocyte/macrophage
Macrophage with ingested
asbestos fiber (encarta.msn.com)
* Phagocytosis and killing of bacteria
* Antigen processing
* Inflammation
Adaptive Immune System: Lymphocytes
Activated B cell
Peripheral blood
Activated T cell (SEM)
* B cells: Mature into plasma cells, secrete antibody (IgM, IgG, IgA, IgE, IgD)
* T cells: T helper - produce stimulatory and regulatory cytokines
* T cells: T cytotoxic/suppressor – contact-dependent cytotoxicity,

regulation of immune response
* NK cells: direct killing of cells (innate arm of IS)

Plasma Cells Produce Antibodies
* IgM: Primary response, efficient agglutination
* IgG: Recall response, highest concentration
* IgA: Mucosal surfaces, trapping of microbes
* IgE: Allery/anaphylaxis

Factors Affecting Immunocompetence
* Age
* Gender
* Genotype
* Nutritional status
* Life style choices
* Acute toxicity
CONCEPT: Individual immunocompetence, in the absence of xenobiotic exposure, is complex, dynamic and affected by fixed and variable factors. At the population level, the “normal” range is broad.

Immunocompetence in the Young: Innate immunity
* Neutrophils
* NK cells
Immunocompetence in the Young: Adaptive immunity
* Humoral immunity
* Cellular immunity
* Resistance to infection
Advanced Age and Immunocompetence

* Innate Immunity
* Adaptive immunity
* Resistance to infection

Gender and Immunocompetence
Genotype and Immunocompetence
Lifestyle and Immunocompetence
* Recreational drug use
* Excessive use of alcohol
* Smoking
* Stress

Xenobiotic Exposure and Immunocompetence
Immune
System
Exposure
Suppression
Infection
Neoplasia
Modulation
Allergy
Autoimmunity

Consequences of Xenobiotic Exposure on Immunocompetence
“Pre-immune” Mechanisms of Defense
Immune Mediated Resistance to Infection
Organism Factors Influencing Host Resistance
Mechanisms of Chemically-induced Immunosuppression
Mechanisms of Suppression:
Effects on Supply of Cells
Mechanisms of Suppression: Effects on Supply of Cells
UVB (320-280 nm) exposure
Mechanisms of Suppression: Tolerance Induction (and then some)
Mechanisms of Suppression: Tolerance Induction (human studies)
Mechanisms of Suppression: Modulation of cytokine production
Mechanisms of Suppression: Th1/Th2 Polarization
Mechanisms of Suppression: Disruption of innate immunity
Human and Mouse Macrophage Responses to Ozone in vivo
Mechanisms of Chemically-induced Immunosuppression
Mechanisms of Suppression:
Summary
* Reduced supply of immune system cells
* Misdirection of the immune system
* Direct effects on cells
* Combination of effects
Decreased Host Resistance: Implications for Human Health

Immune System Overview Mechanisms of Immunosuppression

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Metabolic Disorders - Inborn Errors of Metabolism

Metabolic Disorders - Inborn Errors of Metabolism
By:Dr. Sara Mitchell

Overview
* Proteins - what are they and what do they do?
* Amino Acids - what are they and what do they do?

Eight Essential Amino Acids
* Tryptophan
* Lysine
* Methionine
* Phenylaline
* Theronine
* Valine
* Leucine
* Isolecucine

Inborn Errors of metabolism
* Affects amino acid & protein, carbohydrate, and lipid metabolism.
* Most disorders are autosomal recessive in transmission
* Most disorders are evident at or soon after birth.
* Early detection and treatment are essential to the prevention of irreversible cognitive impairment and early death

Newborn Screening: What is it?
* A test developed in 1961 by Dr. Robert Guthrie to evaluate infants for certain genetic anomalies, inborn errors of metabolism, and other disorders.

http://health.state.ga.us/programs/nsmscd/

Phenylketonuria (PKU):What is it?
* The most common amino acidemia. Classic PKU develops in the absence of the enzyme phenylalanine hydroxylase.
* Incidence

Phenylketonuria: How’s it happen?
* Cause
o absent Phenylalanine hydroxylase causes a build up phenylalanine
* Effect

Phenylketonuria
* Treatment
* Prognosis

Galactocemia: What is it?
* An inborn error of carbohydrate metabolism in which the hepatic enzyme galactose 1-phosphate uridine transferase is absent.
* Incidence

Galactocemia: How does it happen?
Galactocemia: What are the clinical manifestations?
Galactocemia: Diagnosis & Treatment
* Diagnosis
* Treatment

Metabolic Disorders - Inborn Errors of Metabolism.ppt

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Amino Acid Catabolism: Carbon Skeletons

Amino Acid Catabolism: Carbon Skeletons
Copyright © 1999-2007 by Joyce J. Diwan.
All rights reserved.

Molecular Biochemistry II

Amino Acid Carbon Skeletons
Amino acids, when deaminated, yield a-keto acids that, directly or via additional reactions, feed into major metabolic pathways (e.g., Krebs Cycle).
Amino acids are grouped into 2 classes, based on whether or not their carbon skeletons can be converted to glucose:

o glucogenic
o ketogenic.

Carbon skeletons of glucogenic amino acids are degraded to:
o pyruvate, or
o a 4-C or 5-C intermediate of Krebs Cycle. These are precursors for gluconeogenesis.
Glucogenic amino acids are the major carbon source for gluconeogenesis when glucose levels are low.
They can also be catabolized for energy, or converted to glycogen or fatty acids for energy storage.

Carbon skeletons of ketogenic amino acids are degraded to:
o acetyl-CoA, or
o acetoacetate.

Acetyl CoA, & its precursor acetoacetate, cannot yield net production of oxaloacetate, the gluconeogenesis precursor.
For every 2-C acetyl residue entering Krebs Cycle, 2 C leave as CO2.
Carbon skeletons of ketogenic amino acids can be catabolized for energy in Krebs Cycle, or converted to ketone bodies or fatty acids.
They cannot be converted to glucose.
The 3-C a-keto acid pyruvate is produced from alanine, cysteine, glycine, serine, & threonine.
Alanine deamination via Transaminase directly yields pyruvate.
Serine is deaminated to pyruvate via Serine Dehydratase.
Glycine, which is also product of threonine catabolism, is converted to serine by a reaction involving tetrahydrofolate (to be discussed later).

The 4-C Krebs Cycle intermediate oxaloacetate is produced from aspartate & asparagine.
Aspartate transamination yields oxaloacetate.
Aspartate is also converted to fumarate in Urea Cycle. Fumarate is converted to oxaloacetate in Krebs cycle.
Asparagine loses the amino group from its R-group by hydrolysis catalyzed by Asparaginase.
This yields aspartate, which can be converted to oxaloacetate, e.g., by transamination.
The 4-C Krebs Cycle intermediate succinyl-CoA is produced from isoleucine, valine, & methionine.
Propionyl-CoA, an intermediate on these pathways, is also a product of b-oxidation of fatty acids with an odd number of C atoms.
The branched chain amino acids initially share in part a common pathway.
Branched Chain a-Keto Acid Dehydrogenase (BCKDH) is a multi-subunit complex homologous to Pyruvate Dehydrogenase complex.
Genetic deficiency of BCKDH is called Maple Syrup Urine Disease (MSUD).
High concentrations of branched chain keto acids in urine give it a characteristic odor.
Propionyl-CoA is carboxylated to methylmalonyl-CoA.
A racemase yields the L-isomer essential to the subsequent reaction.
Methylmalonyl-CoA Mutase catalyzes a molecular rearrangement: the branched C chain of methylmalonyl-CoA is converted to the linear C chain of succinyl-CoA.
The carboxyl that is in ester linkage to the thiol of coenzyme A is shifted to an adjacent carbon atom, with opposite shift of a hydrogen atom.

Recall that coenzyme A is a large molecule.
Coenzyme B12, a derivative of vitamin B12 (cobalamin), is the prosthetic group of Methylmalonyl-CoA Mutase.
A crystal structure of the enzyme-bound coenzyme B12.
Coenzyme B12 contains a heme-like corrin ring with a cobalt ion coordinated to 4 ring N atoms.
o methyl C atom of 5'-deoxyadenosine (not shown).
o an enzyme histidine N
When B12 is free in solution, a ring N of the dimethylbenzimidazole serves as axial ligand to the cobalt.
When B12 is enzyme-bound, a His side-chain N substitutes for the dimethylbenzimidazole.
Within the active site, the Co atom of coenzyme B12 has 2 axial ligands:
Homolytic cleavage of the deoxyadenosyl C-Co bond during catalysis yields a deoxyadenosyl carbon radical, as Co3+ becomes Co2+.
Reaction of this with methylmalonyl-CoA generates a radical substrate intermediate and 5'-deoxyadenosine.
Following rearrangement of the substrate, the product radical abstracts a H atom from the methyl group of 5'-deoxyadenosine.
This yields succinyl-CoA and the 5'-deoxyadenosyl radical, which reacts with coenzyme B12 to reestablish the deoxyadenosyl C-Co bond.

Methyl group transfers are also carried out by B12 (cobalamin).
Methyl-B12 (methylcobalamin), with a methyl axial ligand substituting for the deoxyadenosyl moiety of coenzyme B12, is an intermediate of such transfers.
E.g., B12 is a prosthetic group of the mammalian enzyme that catalyzes methylation of homocysteine to form methionine (to be discussed later).
o Vitamin B12 is synthesized only by bacteria.
Ruminants get B12 from bacteria in their digestive system.
Humans obtain B12 from meat or dairy products.
o Vitamin B12 bound to the protein gastric intrinsic factor is absorbed by cells in the upper part of the human small intestine via receptor-mediated endocytosis.
B12 synthesized by bacteria in the large intestine is unavailable.
Strict vegetarians eventually become deficient in B12 unless they consume it in pill form.
o Vitamin B12 is transported in the blood bound to the protein transcobalamin, which is recognized by a receptor that mediates uptake into body cells.

Explore via Chime
Methylmalonyl-CoA Mutase
with its prosthetic group,
Coenzyme B12.
Desulfo-CoA (without the
thiol) is at the active site.
The deoxyadenosyl moiety is lacking in the crystal.

The 5-C Krebs Cycle intermediate a-ketoglutarate is produced from arginine, glutamate, glutamine, histidine, & proline.
Glutamate deamination via Transaminase directly yields a-ketoglutarate.
Glutamate deamination by Glutamate Dehydrogenase also directly yields a-ketoglutarate.
Histidine is first converted to glutamate. The last step in this pathway involves the cofactor tetrahydrofolate.
Tetrahydrofolate (THF), which has a pteridine ring, is a reduced form of the B vitamin folate.
Within a cell, THF has an attached chain of several glutamate residues, linked to one another by isopeptide bonds involving the R-group carboxyl.
THF exists in various forms, with single-C units, of varying oxidation state, bonded at N5 or N10, or bridging between them.
In these diagrams N10 with R is r-aminobenzoic acid, linked to a chain of glutamate residues.
The cellular pool of THF includes various forms, produced and utilized in different reactions.
N5-formimino-THF is involved in the pathway for degradation of histidine.
Reactions using THF as donor of a single-C unit include synthesis of thymidylate, methionine, f-methionine-tRNA, etc.
In the pathway of histidine degradation, N-formiminoglutamate is converted to glutamate by transfer of the formimino group to THF, yielding N5-formimino-THF.
Because of the essential roles of THF as acceptor and donor of single carbon units, dietary deficiency of folate, genetic deficiencies in folate metabolism or transport, and the increased catabolism of folate seen in some disease states, result in various metabolic effects leading to increased risk of developmental defects, cardiovascular disease, and cancer.

Aromatic Amino Acids
Aromatic amino acids phenylalanine & tyrosine are catabolized to fumarate and acetoacetate.
Hydroxylation of phenylalanine to form tyrosine involves the reductant tetrahydrobiopterin. Biopterin, like folate, has a pteridine ring.
Dihydrobiopterin is reduced to tetrahydrobiopterin by electron transfer from NADH.
Thus NADH is secondarily the e- donor for conversion of phenylalanine to tyrosine.
Overall the reaction is considered a mixed function oxidation, because one O atom of O2 is reduced to water while the other is incorporated into the amino acid product.
O2, tetrahydrobiopterin, and the iron atom in the ferrous (Fe++) oxidation state participate in the hydroxylation.
O2 is thought to react initially with the tetrahydrobiopterin to form a peroxy intermediate.
Phenylalanine Hydroxylase includes a non-heme iron atom at its active site.
X-ray crystallography has shown the following are ligands to the iron atom:
His N, Glu O & water O.
(Fe shown in spacefill & ligands in ball & stick).
deamination via transaminase) accumulate in blood & urine.
Mental retardation results unless treatment begins immediately after birth. Treatment consists of limiting phenylalanine intake to levels barely adequate to support growth. Tyrosine, an essential nutrient for individuals with phenylketonuria, must be supplied in the diet.
Genetic deficiency of Phenylalanine Hydroxylase leads to the disease phenylketonuria.
Phenylalanine & phenylpyruvate (the product of phenylalanine
Tyrosine is a precursor for synthesis of melanins and of epinephrine and norepinephrine.
High [phenylalanine] inhibits Tyrosine Hydroxylase, on the pathway for synthesis of the pigment melanin from tyrosine. Individuals with phenylketonuria have light skin & hair color.
Methionine S-Adenosylmethionine by ATP-dependent reaction.
SAM is a methyl group donor in synthetic reactions.
The resulting S-adenosylhomocysteine is hydrolyzed to homocysteine.
Homocysteine may be catabolized via a complex pathway to cysteine & succinyl-CoA.
Or methionine may be regenerated from homocysteine by methyl transfer from N5-methyl-tetrahydrofolate, via a methyltransferase enzyme that uses B12 as prosthetic group.
The methyl group is transferred from THF to B12 to homocysteine.
Another pathway converts homocysteine to glutathione.
In various reactions, S-adenosylmethionine (SAM) is a donor of diverse chemical groups including methylene, amino, ribosyl and aminoalkyl groups, and a source of 5'-deoxyadenosyl radicals.
But SAM is best known as a methyl group donor.

Examples:
S-adenosylmethionine as methyl group donor
o methylation of bases in tRNA
o methylation of cytosine residues in DNA
o methylation of norepinephrine epinephrine
o conversion of the glycerophospholipid
phosphatidyl ethanolamine phosphatidylcholine via methyl transfer from SAM.
Enzymes involved in formation and utilization of S-adenosylmethionine are particularly active in liver.
Liver has important roles in synthetic pathways involving methylation reactions, & in regulation of blood methionine.
Methyl Group Donors

Methyl group donors in synthetic reactions include:

* methyl-B12
* S-adenosylmethionine (SAM)
* N5-methyl-tetrahydrofolate (N5-methyl-THF)
Lysine & Tryptophan

Amino Acid Catabolism: Carbon Skeletons.ppt

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