Mechanisms of Teratogenesis

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Mechanisms of Teratogenesis
Lynda B. Fawcett, Ph.D. Assistant Professor of
Pediatrics Thomas Jefferson University Alfred I
duPont Hospital For Children Wilmington, DE
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Structural Anomalies/Dysmorphology

• Malformation the normal developmental process is
  altered such that a given structure cannot form
  or forms improperly, the error is intrinsic to
  the morphogenetic process itself.
• Deformation alterations to already existing
  structures. Alterations in body shape, form, or
  position. Often due to uterine constraint
  (twinning, oligohydramnios etc). ex dislocation
  of the hip, clubfoot, and some facial anomalies
• Disruption alterations to already existing
  structures, often involving extensive destruction
  of tissue, and/or prevention of later tissue or
  organ formation. Can arise from mechanical or
  physiological factors. ex amniotic band
  syndrome, fetal vascular occlusion, placental
  emboli, localized or general hypoxia, vascular
  insults.

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Disruption Syndactyly/adactyly
Hypoxia, hemorrhage, necrosis ROS Usually
asymmetrical
EX Cocaine CVS
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Mechanism of action
Mode of Action
Outcomes
cell death altered cell-cell interactions cell
signaling reduced biosynthesis impaired cell
migration reduced/impaired proliferation alter
ed differentiation
ROS/Lipid peroxidation DNA damage Altered gene
expression Metabolic disturbance Enzyme
inhibition Growth factor imbalance Receptor
agonist/antagonist
Malformation Deformation Death Functional
deficit Growth retardation
Pathogenesis
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Cytotoxic Teratogens

• Irradiation
• 2Deoxyadenosine
• 4-CP (metabolites)
• Hyperthermia
• Ethanol

Whether a toxicant induces cell death depends on
dose and timing of exposure as well as the nature
of the toxicant
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Apoptosis
Necrosis
The blebs fuse and become larger, no organelles
in blebs
The nucleus begins to break apart, DNA breaks
into small pieces organelles in to blebs
Blebs
The cell membrane ruptures and releases the
cells contents, the organelles are not
functional
The cell breaks into several apoptotoc
bodies the organelles are still functional
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Apoptosis
SIGMA-ALDRICH
Receptor mediated - Extrinsic (TNF Signaling)
Mitochondrial Intrinsic CytC, caspase 9
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Characteristics of Cell Death Mediated
Teratogenesis Replication associated cytotoxicity

• Tissue susceptibility changes over time, however
  usually rapidly dividing undifferentiated tissues
  are most affected with agents that induce direct
  cytotoxicity
• Tissue sensitivity greatest during organogenesis
  in the neuroepithelium, limb buds and least in
  heart and yolk sac
• Characteristic malformations include NTDs,
  anopthalmia, limb defects but, generally, not
  heart defects (time dependent).
• Occurs because
• DNA more accessible in S phase
• Defects manifest sooner, critical mass, windows
  of oportunity
• Molecular mechanisms

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Tissue sensitivity Irradiation/mouse
studies/organogenesis

• Molecular basis for tissue sensitivity may be due
  to differential expression of P53 in tissues
• Temporal/spacial expression of P53 mirrors that
  of tissues most effected by cytotoxic teratogens
• Irradiation caused activation of P53 in rapidly
  proliferating, undifferentiated tissues (used a
  lac Z reporter model). Neuroepithelium, limb
  buds, arches (Gottli et al, 1997)

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(No Transcript)
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P53 KO studies
Consistent findings / apoptosis, cell cycle
check G1/S /- apoptosis and necrosis -/-
necrosis, no cell cycle check (M,S)
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Replication associated cytotoxicity
Open NT, reduced tissues Anopthalmia Growth
retardation
Control 11.5 d
AA deficiency (Met)
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Demethylating agents. Whole Mount Immunostaining
165 kD NF
Control
-AA (Met)
CL (-SAM)
NTD only ?165 Kd NF Mechanism probably more
specific, not general cytotoxicity
NTD GR, Reduced Size Protein Somite Pairs
limb buds Anopthalmia No heart defects
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Effects of Homocysteine on Rat Embryos Cultured
in Vitro from 9.5-11.5 p.c Cardiac Defects
Control
Hcy
Hcy
Control
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Effects of Hcy on Rat Embryos Cultured in Vitro
from 9.5-11.5 p.c Somite/Vertebral/Arch
Abnormalities (450 ?g/ml)
control
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• Although defects such as NTDs may result from
  death, other mechanisms can also result in NTD
• Specific mechanisms of action
• Receptor mediated - directly or indirectly
• Display some type of ligand specificity (enzyme)
• downstream alterations in cell signaling (growth
  factors differentiation)
• altered expression of key genes involved in
  morphogenesis.
• Closure defects have discrete window of
  susceptibility, and less chance for recuperation.
  for NTD reduced proliferation in neuroepithelium,
  failure of neural fold elevation, failure of
  neural fold fusion due to specific mechanisms may
  resemble cytotoxicity at term

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Tissue specificity and TeratogenesisSpecific
Mechanism Usually receptor mediated

• Only affect tissues bearing appropriate
  receptors
• ex sex steroids - genitourinary

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Receptor-Mediated Teratogenesis
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Retinoic Acid

• Accutane 13-cis retinoic acid, used for
  dematological and oncology therapies

RAE Spectrum of defects and potential for all
organ systems to be affected Most common defects
are craniofacial, external ear (BA 1-2)
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Specificity Teratogenesis RA

• Vitamin A derivatives involved in signaling
  pathways during normal development - Morphogen
  directly alter/control developmental programs.
• 1) Induces differentiation or apoptosis of
  various cell types breast carcinoma, prostate
  cancer, AL
• 2) Produces homeotic mutations in experimental
  animals (limb duplications)
• Proposed Mode of action for defects
• Promotes excessive cell death in regions
  undergoing programmed cell death (Alles Sulik,
  89, 90 etc)
• Alters specification of tissues (NC) HOX genes
• Alters expression of other RARE genes

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RA receptors

• RAR ??? (all trans and 9-cis) RXR???(9 cis)
• Multiple isoforms exist for all RAR
• RAR/RXR form heterodimers RXR homodimers,
  heterodimers with thyroid hormone R, peroxisome
  proliferators and others
• RAR, RXR differentially expressed in embryo
  temporal and spatial pattern
• RA interacts with other receptors (CRABP)

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CRABP1
RA
RA
RA
RA
RA
RAR/RXR
CRABP-II
(???)
RA
Other involvment
CRABP1, II
Biosynthesis RA ADH - ETOH hypervitamitosis vitami
n deficiency
Transcription RARE
AP-1 transcription factor block
RAR/RXR
Nucleus
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• RAR? -/- Vitamin A deficiency, Resistant to RA
  induced limb defects. Agonists
  RAR?gtRAR?gtRAR? for inducing limb defects
  in wild type
• RAR? -/- Normal, Resistant to BA 1-2 fusion
  by RA
• / RAR? agonist caused BA fusions
• RAR? upregulated in limbs following RA
• RAR? -/- Vitamin A deficiency defects, normal
  craniofacial and posterior axial skeleton
  resistant to RA induced posterior axial
  skeleton truncation but not cranial, partial
  resistance to other except limb
• RXR? -/- Lethal, cardiac. Resistant to RA induced
  limb defects

Limb
Craniofacial
Posterior
Cardiomorphogenesis
(Matt et al., 2003 Collins and Mao, 1999)
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Chromatin condensation suppresses RAREs (RARß2)
VPA
Potential for increased RARß2 destabilization of
chromatin Inappropriate expression of other
genes (p21 p53 cell death)
RARE
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Summary

• Teratogenic mechanisms can be broadly separated
  into specific (receptor mediated) and
  non-specific (cell death)
• Teratogens can have both specific and
  non-specific mechanisms (alcohol)
• Even general cytotoxic/genotoxic teratogens may
  have a molecular component that alters tissue
  susceptibility
• Initial molecular mechanisms have the potential
  to result in myriad responses cell death,
  differentiation, altered signaling. Pathogenesis
  must be considered when evaluating the action of
  teratogens
• Understanding the molecular mechanisms of
  teratogens furthers our understanding of
  development, and allows for the development of
  therapeutics with reduced chance of
  teratogenicity