Mechanisms of Toxicity Flashcards
The mechanisms of toxicity are the fundamental ________ and ________ interactions responsible for the ______ and __________ of toxic responses.
* Large number of toxicants with ________ biological processes, so ultimately there are numerous potential mechanisms.
The fundamental chemical and biological interactions responsible for the genesis and longevity of toxic responses.
* Large number of toxicants with numerous biological processes, so ultimately there are numerous potential mechanisms
Why should we study the mechanisms of toxicity?
- To interpret descriptive toxicity data: We need to understand why one toxicant is more toxic than another.
- To estimate the probability that a chemical will cause harmful effects: This is because mechanistic studies generate data on doses that impair specific processes
- To develop procedures to prevent toxicity: E.g. The development of antidotes is based on the understanding of the mechanisms of toxicity. It is only possible to reverse toxic responses if you know how those responses occurred and develop procedures that counteract the underlying mechanisms
- To develop less hazardous drugs and chemicals, especially Selectively toxic chemicals. In order to treat cancer it is important that the drug you are using targets only the cancerous cells without affecting the adjacent normal cells.
Describe the steps in development of toxicosis
- Delivery of toxicant to target site. Once here, interacts with target molecule causing dysfunction or injury leading to toxicosis.
- Alter bio env –> injury –> toxicosis
- Cells impacted by toxicants no longer can perform processes –> toxicosis
- How is a toxicant delivered to its target site?
- The intensity of the toxic effect depends on?
- From site of exposure to the target organ
- Intensity of toxic effect depends on the concentration and persistence of the ultimate toxicant at the site of action
What is an ultimate toxicant? Provide examples.
A chemical species that reacts with endogenous target molecules or alters the biological microenvironment.
A. Acetaminophen is typically metabolized into N-acetyl-p-benzoquinone-imine which is
responsible for toxicity
B. Paraquat- During its process of metabolism results in the production of ROS, which then
discriminately affects cells
C. Lastly, it could be an endogenous molecule e.g. During sulfonamide toxicosis where sulfonamide
displays bilirubin from its binding sites in plasma albumin, and bilirubin then accumulates and is
responsible for the toxic response
What factors affect the concentration of the ultimate toxicant at the target site?
Work for toxicant: absorption, etc.
Work against: Distribution (from site of action)
The concentration of the ultimate toxicant at the target molecule depends on the relative effectiveness of the processes that increase or decrease its concentration at the target site.
The accumulation of the ultimate toxicant at its target site is facilitated by its absorption,
distribution to the site of action, reabsorption, and toxication (metabolic activation). Conversely, presystemic elimination, distribution away from the site of action, excretion, and detoxication oppose these processes and work against the accumulation of the ultimate toxicant at the target molecule.
Define the term absorption. What does it depend on?
Absorption
* Transfer of a xenobiotic from site of uptake
to systemic circulation. It depends on:
– Toxicant concentration at site of absorption (GI, resp, skin)
– Surface area of exposure/absorptive site (how much of substance is absorbed).
– Structure of the absorptive surface (thickness)
– Perfusion of the sub-epithelial region
– Physicochemical properties of the toxicant (lipid solubility)
Absorption is counterbalanced by?
Pre-systemic Elimination
* Also known as first-pass elimination
- Define pre-systemic elimination.
- How does this affect certain parts of the body?
- How does this affect the toxicant that was absorbed?
- It is the loss of a toxicant during transfer from
the site of exposure to the systemic circulation
– Most important for toxicants absorbed from the gastrointestinal tract - May contribute to increased injury of the digestive tract mucosa, liver, and lungs b/c these processes promote delivery to such sites (e.g. ethanol, iron salts, a-amanitin, and
paraquat) - Reduces toxicity of toxicants delivered to target sites via systemic circulation.
The most important site for pre-systemic loss is the _____ where upon delivery by the hepatic _____ vein, a significant fraction of toxicant may be metabolized and excreted via _____ and back into ___ before it gets into the systemic circulation. The other important site for pre-systemic elimination is the _____ where significant __________ and _________ for some toxicants can occur before those toxicants get into systemic circulation.
The process by which toxicants absorbed from GIT are taken to the _____, ________ and returned
to the GIT via _____ ____ is referred as enterohepatic circulation
liver, portal, bile, GIT, lungs, metabolism, elimination
liver, eliminated, bile duct
Within the GIT, the toxicant may be metabolized by enzymes of the gut ________ as well as enzymes in the cells of GI _______ and excreted before ________. This process is called?
What do the red arrows in the image below indicate?
microbiota, epithelium, absorption
Pre-systemic loss
In this diagram note the decrease in thickness of the red arrows from GIT to the systemic circulation which depicts presystemic elimination.
Enterohepatic circulation can go on for a _____ time and may involve the _______ of the metabolic reactions that took place in the liver within GIT. Provide an example of this?
Plasma conc. of toxicants that have significant enterohepatic circulation is characterized by ______ and even ______ peaks over time.
long, reversal
Eg Gut microbial glucuronidase can remove the glucuronide acid from the glucuronidated toxicant metabolite, thereby releasing the parent toxicant which is then reabsorbed.
secondary, tertiary
- What is the process of distribution facilitated by?
- What do each of these factors enable in terms of the development of toxicosis?
- See image below
- A) Porosity of capillary endothelium allows the passage of large molecules including protein-bound toxicants.
(B) Some toxicants can preferentially accumulate. For Lysosomes the accumulation occurs via pH trapping where the toxicant gets protonated in the acidic interior of the lysosomes preventing their efflux. For Mitochondria, accumulation occurs electrophoretically where toxicants are protonated in the intermembrane space which has got a high conc. of protons and thereafter they are sucked into
the alkaline mitochondrial matrix due to strong negative potential
Which mechanisms oppose distribution?
- Binding to Plasma Proteins- Dissociation from proteins is required for most xenobiotics to leave the blood and enter cells. Therefore, strong binding to plasma proteins delays and prolongs the
effects and elimination of toxicants. - Specialized Barriers - Brain capillaries have very low aqueous porosity because their endothelial
cells lack fenestrae and are joined by extremely tight junctions. This blood–brain barrier prevents the access of hydrophilic chemicals to the brain except for those that can be actively transported. - Distribution to Storage Sites- Some chemicals accumulate in tissues (i.e., storage sites) where
they do not exert significant effects. Such storage decreases the availability of these toxicants for
their target sites and acts as a temporary protective mechanism (e.g. lead in bones, chlorinated hydrocarbons in fat). - Association with Intracellular Binding Proteins- Binding to nontarget intracellular sites also
reduces the concentration of toxicants at the target site.
The best example is the binding of metals to low mol wt proteins known as MT. When metals are
bound by this protein they are not available to interact with sensitive target sites in the cell. - Lastly toxicants can be pumped out of cells. During the Export from Cells- Intracellular toxicants may be transported back into the EC space. This occurs in brain capillary endothelial cells. In their luminal membrane, these cells contain ATP-dependent membrane transporters (ATP-binding cassette or ABC
transporters) such as the multidrug-resistance protein (MDR1), or P-glycoprotein, which
extrudes chemicals and contributes to the blood-brain barrier
Explain how excretion and reabsorption oppose each other.
1. What are the main organs involved in the process of excretion?
2. What types of substances are eliminated via excretion? Which ones are not readily eliminated?
3. How does excretion differ from reabsorption?
Excretion
– Removal of xenobiotics from the blood and their
return to the external environment
* Main organs: kidney, liver → GI tract, lungs
- Primarily excrete hydrophilic or volatile substances.
* Lipophilic substances are not readily eliminated.
Differs from reabsorption in that the Amount of parent compound is reduced by biotransformation.
Reabsorption
– Toxicants excreted in urine, bile & GI secretions may diffuse back/get reabsorbed into blood. REbaospriton depends on lipid solubility and is inversely proportional to the extent of ionization.
- Define toxication.
- How can toxication occur?
- Toxication is a biotransformation reaction in which metabolite is more toxic than the parent
xenobiotic. - It can occur through the acquisition of greater reactivity thru metabolism. E.g. the conversion of
parathion to paraoxon by the Cytochrome P450 enzyme system increases the toxicity of these
organophosphates. However, the most prevalent and important mechanism for increased toxicity is the acquisition of indiscriminate reactivity. This includes the formation of electrophiles which are chemical spp that are attracted towards the electrons, because electrons are –vly charged, electrophiles are +vly charged.
Toxicants can be converted to nucleophiles which are spp that are attracted towards the nucleus
because the nucleus is +vly charged nucleophiles carry –ve charge.
Toxicants can cause the formation of free radicals which are molecules that contain unpaired
electrons. As a result, this spp is highly unstable and reactive and basically always seeks to
donate or acquire an electron in order to stabilize.
Lastly toxicants can acquire indiscriminate reactivity by becoming redox active where they are able to accept or donate electrons
- What is detoxication?
- What does it depend on?
- Is the process of biotransformations that eliminate the ultimate toxicant or prevent its formation
- Detoxication mechanisms depend on the nature of the toxicant:
(A) Toxicants with no functional groups
* Detoxicated by addition of a functional group (-OH, -NH2, -COOH, etc.) followed by conjugation. Then they are …
* Catalyzed by phase I enzymes, e.g., CYP450
– Involves oxidation, reduction or hydrolysis - Toxicants with functional groups
– Conjugation (by phase II enzymes)
- An endogenous molecule, like glucuronic acid, sulphuric acid, or amino acid, is added by the process of conjugation that is catalyzed by phase 2 enzymes. - Free radicals
– e.g., O2*- is dismutated (simultaneous oxidation and reduction) and converted to water by catalase (CAT), glutathione peroxidase (GPx) or peroredoxin (Prx) - Protein toxins: E.g. venoms
can be enzymatically inactivated by proteases (e.g. venoms; cleaved into smaller units, such as dipeptides, that no longer exert toxicity).
When does detoxication fail?
- Toxicants can overwhelm detoxication
mechanisms
– Exhaustion of enzymes, antioxidants, etc - Inactivation of detoxifying enzymes
- Reversal of detoxification reactions
- Detoxication may produce harmful by-products (e.g., conversion of GSH to glutathione thiyl radical)
Describe step 2a: Interaction with Target Molecules
Toxic action is mediated by the reaction of the ultimate toxicant with _______ molecules. Such as ?
The outcome depends on:
I. Attributes of the ?
II. Types of reactions between the ________ _________ and the _______ molecules
III. Effects of toxicant on the _____ molecules
- Toxic action is mediated by the reaction of the ultimate toxicant with target molecules. Such as reactibility, accessibility, and function
The outcome depends on:
I. Attributes of the target molecules
II. Types of reactions between the ultimate toxicant and the target molecules
III. Effects of toxicant on the target molecules
I. Attributes of Target Molecules
* Practically all endogenous molecules are potential _______ for toxicants
* The most toxicologically relevant targets are ?
* The target molecule must be able to react with the ______ and have appropriate ______ and _____.
All toxicants do not contribute to toxicity?
targets, nucleic acids, proteins and membranes, configuration, accessibility
The type of reaction determines the type of _________ and ________ effects.
List these reaction types.
Longevity, toxic
“No Christine, His Electrons are Enzymatic”
- Binding is either be covalent or noncovalent: (A) Noncovalent binding= toxicants interacting with receptors, ion channels, and enzymes. Forms hydrogen and ionic bonds which have low binding energy why is why it’s reversible. (B) Covalent binding permanently alters the endogenous molecules and is irreversible = interaction of target molecules with electrophilic toxicants and free radicals.
- H abstraction = removal of an H atom from a molecule by a radical.
This usually leads to the conversion of endogenous molecules to radicals/formation of carbonyls –> cross-linking of DNA. - Electron transfer = chemicals exchange electrons to oxidize/ reduce other molecules –> formation of harmful byproducts or inactivation of proteins. Eg Nitrite oxidizes Iron to Hb to form Iron 3 in methemoglobin and MetHb has low Oxygen carrying capacity
- Lastly, some toxins can act enzymatically on target proteins eg Ricin which is found in castor
beans can hydrolyze ribosomes whereas snake venom also contains hydrolytic enzymes that
destroy biomolecules
List the effects of toxicants on target molecules.
- Dysfunction of target molecules
– Activation (e.g. morphine/opioid receptor) or inhibition (e.g. strychnine/glycine receptor) - Destruction of target molecules
– Cross-linking, fragmentation, degradation - Formation of neoantigens
– Altered proteins evoke immune responses
(e.g., penicillin, halothane, nickel, urushiol- Poison ivy allergen)
Describe the Alteration of Biological
Microenvironment
- Alteration of H+ concentration (e.g., by acids or toxicants metabolized to acids) e.g. ethylene glycol (antifreeze)
- Physicochemical alteration of the lipid phase of cell membranes (e.g., by solvents, detergents) Occupy bilirubin binding site in albumin
- Occupation of a site or a space (e.g., CO2 - can occupy aklveolar space whihc is meant to be occupied by oxygen, sulfonamides - Occupy bilirubin binding site in albumin, ethylene glycol - Renals tubules)
How does toxicosis lead to cellular dysfunction?
(A) Impairment of cellular regulation
– Dysregulation of gene expression: transcription, translation, signal transduction and/or
extracellular signal production
* –> impaired cell division, protein synthesis and apoptosis –> neoplasia, teratogenesis, tissue involution
(B) Dysregulation of ongoing cellular activity
– Electrically excitable cells (neurons, muscle)
– Toxicants can also stimulate non-excitable cells such as e.g., exocrine cells
(C) Impairment of cell maintenance leading to cell death.
How does toxicosis lead to the dysregulation of electrically excitable cells?
- Alteration of neurotransmitter level: Toxicants may alter synaptic levels of NTs by interfering with their synthesis, storage, release, or removal from the vicinity of the receptor.
- Toxicant-neurotransmitter interactions: Some toxicants interact directly with NT receptors, including
(1) agonists that associate with the ligand-binding site on the receptor and mimic the natural ligand,
(2) antagonists that occupy the ligand-binding site but cannot activate the receptor,
(3) activators, and (4) inhibitors that bind to a site on the receptor that is not directly involved in ligand binding. In the absence of other actions, agonists and activators mimic, whereas antagonists and inhibitors block, the physiologic responses characteristic of endogenous ligands. - Alteration of signal transduction: E.g. Voltage-gated Na channels can be activated by DDT
- Impairment of signal termination: The cellular signal generated by cation influx is terminated by the removal of the cations through
channels or by transporters. Inhibition of cation efflux may prolong excitation
Eg Ba inhibits Ca-activated K channels
Thereby impairing the removal of K from cells and prolonging the excitation of those neuronal
cells
Under impaired internal maintenance, what effects do toxicants have?
– Toxicants can impair synthesis of ATP and
endogenous molecules, assembly of
macromolecules, membranes and organelles
and regulation of intracellular environment
* These impairments imperil cell survival
Under impaired external maintenance, what effects do toxicants have?
- Impaired external maintenance
– Impaired function of integrated systems, e.g.,
hemostasis –> excessive bleeding –> death
What are the Three primary biochemical disorders initiated by toxicants that can cause cell death?
– ATP depletion
– Sustained elevation of intracellular Ca2+
– Overproduction of ROS (Reactive Oxygen Species)
–> These disorders result in Mitochondrial Permeability Transition (MPT): an abrupt increase in mitochondrial inner membrane permeability.
The permeability increase causes dissipation of the proton gradient of mitochondrial inner mb
resulting in the cessation of ATP synthesis. In addition, water flows into the mitochondria causing swelling and this eventually can result in the rupture of mitochondria
What are the consequences of MPT in the case of:
1. Few
2. Many
3. All/most mitos
The outcome of mitochondrial permeability transition depends on the number of mitochondria
affected.
If only a few mitochondria are affected = mitophagy and the cell
survives.
If many mitochondria in the cell undergo MPT = caspase is activated, and the cell undergoes apoptosis.
If most or all the mitochondria are affected = cessation of ATP synthesis and ATP depletion.
Without energy, the cell undergoes necrosis.
- Necrosis results when MPT affects?
- Necrosis is characterized by?
Necrosis
* Results when MPT affects most or all mitochondria
* Necrosis is characterized by cell/organellar swelling and lysis of plasma membrane
* There is disconnection of cytoskeleton from plasma membrane and membrane blebbing
* It is a random, uncontrolled and disorderly disintegration of cells associated with inflammation
In regards to MPT, when does apoptosis occur?
Apoptosis, or programmed Cell Death, results when MPT affects many but NOT all mitochondria
* Characteristics of cell death by apoptosis:
– Cells shrink but retain intact membranes
– Condensation of nuclear and cytoplasmic materials
– Apoptotic bodies (membrane-bound cell fragments)
* Damaged cells (apoptotic bodies) are eliminated without inflammation
This diagram summarizes cell death by necrosis and apoptosis.
At the bottom is cell death by necrosis where the cell ______, becomes _____ and ultimately _______ releasing its contents to the _______. This is then followed by severe ________.
At the top is cell death by apoptosis in which the cell _____ and the chromatin ______. The cell then breaks down into small ______-bound fragments known as _______ ________ which are ______.
There is no _______ in this case.
swells, leaky, ruptures, surrounding, inflammation
shrinks, condenses, membrane, apoptotic bodies, phagocytosed, inflammation
- Solvents, detergents, and venoms damage what part of the cell?
- Aminoglycosides damages what part of the cell?
- Phalloidin damages what part of the cell?
- Ricin damages what part of the cell?
- Damage to plasma membranes
- Damage to lysosomal membranes
- Destruction of cytoskeleton
– e.g., (a mushroom toxin) - Disruption of protein synthesis
– e.g., (castor bean toxin)
Describe the step of repair or dysrepair.
- Toxicants may alter macromolecules that if not repaired cause damage at higher levels of biological hierarchy
- Repair mechanisms arrest progression of injury
Failure of the repair mechanisms results in the progression of the damage from the molecular
level to the cell, tissue and organ levels.
At what step does molecular repair fail?
- How are oxidized proteins repaired?
- How are denatured or altered proteins repaired?
- How are permanently damaged proteins repaired?
- Oxidized proteins are repaired by reduction or electron transfer
– Requires enzyme action and reducing equivalents
* e.g., repair of oxidized thiol groups involves the thioredoxin/glutaredoxin system and NADPH (Nicotinamide adenine dinucleotide phosphate, reduced) - Denatured or altered proteins undergo refolding by heat shock proteins
- Permanently damaged proteins are eliminated by proteolytic degradation
- How are perioxidized lipids repaired?
- How are perioxidized fatty acids repaired?
- Peroxidized lipids are repaired by a complex process involving reductants and enzymes e.g., GSH, glutathione peroxidase, and glutathione reductase
– NADPH recycles the reductants (GSH) oxidized in the process - Peroxidized fatty acids in phospholipids can be replaced with normal fatty acids after preferential hydrolysis by phospholipase A2
Why is mitochondrial DNA prone to damage? Is nuclear DNA prone to damage?
Mitochondrial DNA is prone to damage: lacks
histones and efficient repair mechanisms. Nuclear DNA is very stable.
Histones are rich in _________ and _______ and repair DNA into ___ chromosomes.
arginine, lysinine, new
Decribe the process fo direct repair.
- Direct repair
– Involves enzymatic reversal of covalent DNA
modification e.g. by DNA photolyase following UV
modification or by alkyl transferases (remove alkyl
groups from DNA) after chemical-induced alkylation. UV-induced thymine dimers
repaired by photoreactivation,
in which energy from visible
light is used to split the bonds
What can be seen below?
DNA Repair: UV-induced thymine dimers repaired by photoreactivation, in which energy from visible light is used to split the bonds
The first one is direct repair where DNA photolyase cleaves adjacent pyrimidine bases dimerized
by UV light. This enzyme utilizes energy from visible light.
Another mechanism of direct repair involves the use of alkyl transferases which removes alkyl
groups from alkylated DNA.
Describe the process of excision repair.
What is base excision used for? Nucleotide excision?
Base or nucleotide is excised and replaced
– Base excision: used for lesions that do not cause major helix distortion
– Nucleotide excision: used for removal of bulky adducts that distort the double helix structure
What can be seen in the image below?
The diagram here shows base excision repair. Basically, the altered base or dimer is recognized
by a DNA glycosylase and removed leaving the sugar-phosphate bond intact.
This creates a site without a purine or pyrimidine base known as an AP site which is recognized
by AP endonuclease.
The AP endonuclease cuts the phosphodiester bond,
and the missing part (gap) is resynthesized by a DNA polymerase and sealed in place by a DNA
ligase.
Describe recombinational/postreplication repair.
- Occurs when excision of a bulky adduct or pyrimidine dimer fails to occur before replication begins
- This results in a gap opposite the dimer in the newly synthesized strand
- Recombination with the undamaged parental strand fills the gap
- Parental strand gap is filled using daughter strand as template
Thereafter, the pyrimidine dimer is removed by excision repair.
Because recombinational repair occurs after DNA replication, in contrast with excision repair, it
is also called postreplication repair.
Cellular Repair
* Not a widely applied strategy to overcome toxic cell injuries. In most tissues damaged cells are removed
and replaced
* Important in repair of peripheral neurons (axons) and is mediated by macrophages and Schwann cells
– Macrophages remove debris & produce cytokines and growth factors which activate Schwann cells
– Schwann cells support growth and synthesize adhesion molecules
Note: CNS contains inhibitory glycoproteins & chondroitin sulfate
proteoglycans which prevent axonal regrowth
Tissue Repair
* Deletion (by apoptosis) of the injured cells
and regeneration (by mitosis and cell
migration)
* Regeneration of the extracellular matrix by
stellate cells, mesangial cells and fibroblasts
* Reintegration of new cells and matrix into
tissues and organs. Requires the adhesion
molecules cadherins (Cell-cell adhesion), connexins (Gap junction) & integrins (link Cells within ECM)
When does repair failure occur?
- Damage can overwhelm repair mechanisms
Oxidation > reduction, enzymes/cofactors consumed - Toxicant-induced injury adversely affects the
repair process, e.g., stoppage of mitosis - Some toxic injuries cannot be repaired
- Repair can contribute to toxicity
through Depletion of ATP and reductants such as NADPS or glutothione. - Repair may go astray and lead to uncontrolled
proliferation –> neoplasia and fibrosis
What are the adverse consequences of dysrepair?
- Necrosis (injury overwhelms repair)
- Fibrosis (excess abnormal extracellular
matrix)
– Causes loss of elasticity, compresses normal cells
and blood vessels, increases diffusion barriers - Carcinogenesis
– Results from failure of DNA repair, apoptosis or
termination of cell proliferation
