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Toxicological Targets: Russell L. Carr Department of Basic Sciences College of Veterinary Medicine
Toxicological Targets: Russell L. Carr Department of Basic Sciences College of Veterinary Medicine
Russell L. Carr
Department of Basic Sciences
College of Veterinary Medicine
Definitions
Ø Toxicology = study of poisons
Ø Poison = any agent capable of producing a deleterious
response in a biological system
Types of Poisons
1. Toxin = toxic substances which are produced naturally
Ø Animal venoms (snakes, spiders, scorpions, insects, bacteria,
etc…)
Ø Plant poisons (poison ivy, mushrooms, etc…)
2. Toxicant = toxic substances which are produced by or are a by-
product of man’s activity
(pesticides, fertilizers, metals, oil spills, industrial solvents, etc…)
Ø For simplicity, we will refer to a poisonous chemical as a
toxicant but remember some toxins may have similar effects.
The Target
Ø Poisons are toxic because they interfere with some sort of normal
physiological process resulting in problems.
** As you can tell, the majority of things that a toxicant can act
on are Cellular Components and interaction of the toxicant
with these components interferes with Cell Function **
Why Is Interference With Cell Function
Important?
Ø It all has to do with the Organization of the Body
Ø First, let us introduce the Scheme of Organization
1. Cellular Components
Ø Within one cell, the components work together to keep the cell running.
Ø Failure of one component to operate effectively or over-stimulation of one
component may interfere with the ability of the other components to do their
job.
2. Cell
Ø Alteration of a cellular component has the potential to alter the overall
functioning of a cell.
Ø If a toxicant targets a certain type of cells, all of the cells of that type can be
affected and their contribution to maintaining normal functioning is eliminated
or diminished.
Why is Interference With Cell Function
Important?
Ø Scheme of Organization
3. Tissue
Ø Cells of different types often work together within a tissue to perform a function.
Ø The inability of one type of cell to function may negatively effect the
functioning of cells of other types in a tissue.
Ø A good example of cooperation of different cell types within a tissue occurs in
the brain where there are multiple cell types can be found in the same region.
v Some of these cells induce stimulation whereas others induce inhibition of
that stimulation.
v If the cells which induce stimulation are inappropriately stimulated, the
normal influence of the inhibitory cells will be overridden and the overall
output from that region will be hyperexcitatory.
v If the cells which inhibit stimulation are inappropriately stimulated,
inhibition of the stimulatory response will occur and no output will occur.
v Thus, the overall performance of the tissue is disrupted.
Why is Interference With Cell Function
Important?
Ø Scheme of Organization
4. Organ
Ø Organs (brain, heart, kidney) are composed of multiple type of tissue and each
tissue plays a role in the function of the organ.
Ø For example, let us consider the kidney:
v Certain tissues of the kidney filter the blood and excrete fluid and
macromolecules into the urine (glomerulus).
v Other tissues of the kidney selectively reabsorb water and other
macromolecules that the body needs to keep (tubules).
v There are certain markers (i.e., inulin) which are filtered out into the urine
are only reabsorbed when decreased filtration is required.
v If a toxicant affects a target in the tubules and causes non-selective
resorption, the tubules become more permeable to inulin, then inulin
causes the glomerulus to decrease urine filtration.
v This negatively affects the function of the entire kidney.
Why is Interference With Cell Function
Important?
Ø Scheme of Organization
4. Systems
Ø The different systems (nervous, circulatory, respiratory) are composed of
multiple organs and the systems interact with each other to keep the body
functioning.
Ø For example:
v The cardiovascular system feeds all of the other systems and blockage of
or reduction in blood flow will negatively affect each system.
v The renal system (kidneys) filter the blood to remove wastes from the
other systems.
v The respiratory system supplies oxygen and the digestive system supplies
nutrients for use by other systems and these are carried by the blood.
v The nervous system controls the functions of the cardiovascular,
respiratory, and digestive systems and is in turn fed by these systems.
v Thus, all the systems work together to help one another.
Why is Interference With Cell Function
Important?
5. Organism
Toxicant Cell
Why no response?
v Signal not strong enough to elicit a response.
v Cell compensates such that inappropriate response does not occur.
v The damage in minimal and the cell repairs the damage.
v The damage is so severe and the cell undergoes spontaneous death
(apoptosis) and is replaced.
Whether or not a Toxicant will have an
Effect is Determined at the Cellular Level
Cellular No Response?
Component
Toxicant Cell
Response
•Excessive Excitation
Excitable Cells •Depression of Activity
Dysregulation of
Ongoing cell activity
Other Cells •Metabolic Alteration
•Increase Glandular Secretion
Role of
Target
Molecule
Energy Production
Membrane Function
Cell
Maintenance
S
P
S
E S E E
+ S +
P
S
E+S ES E+P
Enzymes
Ø Normally enzymes have two functions:
1. Activation
v The enzyme reaction can results in a chemical product which
induces some sort of physiological or biochemical response.
v The enzyme can also react with a cellular protein (change its
conformation) such that it is now free to react with chemicals
already present and thereby, induce a physiological response.
2. Inactivation
v The enzyme reaction destroys a substrate which was inducing
some sort of physiological or biochemical response.
v The enzyme can deactivate a cellular protein (change its
conformation) such that is insensitive to the effects of the
chemicals it would normally react with to induce a response.
Enzymes
Ø Many enzymes have two sites to which things can bind to:
1. Active Site
v This is the “business” portion of the enzyme which binds to the
substrate and produces the product.
2. Allosteric Site
v This is a site to which compounds can bind to that will regulate the
function of the enzyme (make it faster or slower).
Allosteric Site
Active Site
Toxicants and Enzymes
Ø The interaction of a toxicant with an enzyme can produce
several outcomes with respect to functioning of the enzyme.
1. Inhibition
v The toxicant can bind to the active site and block the production of
product. The toxicant which does this is called an inhibitor (I)
S
S
I E + I E
S
S
v The enzyme is inhibited and cannot function so either:
• The response being mediated by its substrate will continue.
• The response that needs to be mediated by the product will not occur.
v This is the most common type of toxicant-enzyme interaction.
Toxicants and Enzymes
2. Reduced Activity
v The toxicant can bind to the allosteric site and cause the enzyme to
slow down and not work as efficiently as before.
S S
S
E S E E
+ S + P
S S
v In this case, more substrate remains and less product is produced.
• If you need the product to mediate a response, the response
will not be as strong because less product is available
• If you need to stop a response being mediated by the
substrate, the time to stop the response will be delayed
because substrate is not broken down.
Toxicants and Enzymes
3. Enhanced Activity or Activation
v The toxicant can bind to the allosteric site and cause the enzyme to
speed up and work more efficiently than normal.
S P
S P
E S E E
+ S + P
P
S P
On the membrane
of organelles or
in the nucleus
Cell
+
+
Cell
Membrane +
Cell
Membrane
+ +
+
Cell
Membrane
Ca++
release
Initiates Cellular
or uptake Machinery
Protein Synthesis
Receptors on Organelles or in the Nucleus
3. Associated with organelles (may be
associated with an ion channel or a protein
on the organelle) or in the nucleus
(associated with DNA).
Ø Activate cellular machinery (short term
Cell G response).
Membrane EC Ø Activate gene expression and protein synthesis
(long term response).
Ca++
release
Initiates Cellular
or uptake Machinery
Protein Synthesis
Toxicant-Mediated Effects on
Receptor-Enzyme Cascade Complexes
Cell G
Membrane EC
Ca++
release
Initiates Cellular
or uptake Machinery
Protein Synthesis
Toxicant-Mediated Effects on
Receptor-Enzyme Cascade Complexes
Ca++
release
or uptake
Receptors in the Cytoplasm
Ø The receptors in the cytoplasm are designed to bind to endogenous ligands that can
cross the cell membrane.
Ø Once they bind to the ligand,
they move to the nucleus.
Ø In the nucleus, they bind to
DNA and stimulate gene
transcription leading to protein
synthesis.
Ø The resulting proteins that
are synthesized will elicit a
Response response.
Ø Steroid hormones
(estrogen and testosterone)
Protein Synthesis
are good examples of
ligands for these receptors.
Receptors in the Cytoplasm
The toxicant enters the cell, binds to the receptor, and causes a
response.
Response
Protein Synthesis
Receptors in the Cytoplasm
The toxicant binds to the receptor but blocks the action of the
normal ligand so no response.
Receptors in the Cytoplasm
Some toxicants can interfere with the binding of the ligand-
receptor complex to specific response elements on the DNA.
Allosteric Sites on Receptors
ØAs with enzymes, some receptors have allosteric binding sites.
ØAs with enzymes, binding of a ligand to this site will alter the
response of the receptor.
Toxicants and Receptors
Ø How can you model the effects of toxicants on receptor?
Ø The same principle we introduced for enzymes applies here:
Start Simple and Work Your Way Up
Ø The reaction is similar to that of enzymes but slightly different.
E+R ER
Ø As with enzymes, receptors have values associated with them:
v A specific affinity for their endogenous ligand (Kd)
• When measuring the effects of toxicants which bind to the allosteric
site, the Kd will be affected.
• So you can also have measures of affinity for the allosteric site.
• The receptor will also have a certain affinity for the toxicant that
binds to its active site.
v A specific number of receptors (Bmax)
Toxicants and Receptors
v A specific number of receptors (Bmax)
• Prolonged exposure to toxicants which active receptors will cause the
body to decrease the Bmax to reduce the activation (down-regulation).
• Prolonged exposure to toxicants which block receptors will cause
the body to increase the Bmax to improve the chance for activation
(up-regulation).
v These values can be determined experimentally.
Ø After determining the basic interactions of the toxicant with the receptor,
you can do the same types of things as with enzymes and move up and
measure the effect of those interactions on the response that the receptor
mediates.
Ø You can also progress to measuring the toxicant effects on cell to cell
interactions, cell to tissue interactions, tissue to tissue interactions, tissue
to organ interactions, and so on….
Ø Remember the further you move up, the more complex the situation
becomes.
Reactive Oxygen Species
Ø Other common toxicants are Reactive Oxygen Species.
Ø These are produced naturally by the body but can be produced
following exposure to environmental chemicals.
Ø These are gotten rid of by Anti-oxidants.
Ø The body has anti-oxidants to scavenge these compounds
when you produce them naturally.
Ø Dietary intake can add anti-oxidants. This is why we have the
big push in the “natural supplements” industry to get you to
buy their anti-oxidant products
Ø When you are exposed to an environmental chemical, the
breakdown of that chemical causes these Reactive Oxygen
Species to be produced above the normal levels of production
by your body.
Reactive Oxygen Species
Ø The Reactive Oxygen Species are highly reactive and will bind
to almost anything including enzymes, receptors, and DNA.
Ø This binding can damage proteins (enzymes and receptors) and
cause a 3-D conformational change in protein structure such
that they no longer function correctly.
Ø Binding to DNA by certain species will cause the
fragmentation of DNA we mentioned earlier (this is one of the
few things that causes fragmentation of DNA).
Ø So, in essence, we can have alteration of cell function by
changing the structure of the cellular components through the
action of non-specific toxicants as well as specific toxicants.
Conclusion
Ø The toxic effects of every poison is directly related to cell
dysfunction.
Ø The cell dysfunction induced by a toxicant is the result of its
interaction with a target molecule.
Ø This interaction can occur at:
v the “business” region (ligand or substrate binding site)
v other regions (allosteric site or other sites which induce a
conformational change in structure)
Ø This interaction can cause effects not only on the target
molecule itself but on other molecules which require the target
molecule to be fully functional .
Ø This chain of effects can cause dysfunction at various levels
including the cell, tissue, organ, system, and finally whole
organism.