Teratogens, Mutagens & Carcinogens

Embryogenesis #4

-3 germ layers form (*gastrulation*): cells in epiblast migrate into center of bilaminar disk -Now have a *trilaminar embryonic disk*: top=ectoderm (blue), mesoderm (red), endoderm (yellow) recall: primitive streak & primitive node

Mutagens include:

-Physical agents (X-rays and UV radiation) -Chemical agents (polycyclic aromatic hydrocarbons, ethidium bromide, mustard gas, vinyl chloride, benzene) -Biological agents (viruses, bacteria, transposons)

Teratogenesis

-from greek word for monster -origin or production of *malformed fetuses or offspring* resulting from *developmental exposure to toxins*

Mutagenesis

-production of a mutation (= a change in the genetic code)

Teratogenesis, Mutagenesis & Carcinogenesis

-typically non-lethal endpoints of toxicity -result from changes to nucleic acid function -if *germ cells* affected->can lead to heritable changes in offspring -if *somatic cells* affected-> can lead to changes in cellular growth & development

• List the criteria used to associate a rise in birth defects with a novel potential teratogen.

1) The agent must be present during the critical stage of development. 2) The agent produces a particular pattern of birth defects in animal studies. 3) The agent crosses the placenta and there is a dose-response relationship. 4) There is an abrupt increase in the frequency of a particular defect or group of defects (syndrome). 5) The increase of defects is associated with the use of a new drug or the widespread exposure to a chemical or environmental change. 6) There is an absence of other factors to explain the observations. 7) The mechanism of teratogenesis makes biological sense.

• How does the incidence and severity relate to the dosage of a teratogen?

1) With teratogens, there is a clear dose-response relationship. In general, the higher the dose, the more severe the resulting birth defect will be, as more and more cells in the embryo are damaged or destroyed. This is no dose-response relationship with mutagens. Once the critical genetic mutation(s) have occurred, the cancer cell begins to grow and follows a predictable path.

Damage at various stages:

1-2 weeks: Loss 3-7 weeks: major abnormalities 8-38 weeks: functional and minor abnormalities *week nine is beginning of fetal period

What is microcephaly?

Babies born with microcephaly have a smaller head than usual (less than <33 cm in circumference). Microcephaly can be either mild or severe, depending on the size of the head. The associated signs and problems of microcephaly include failure to thrive (slow weight gain and growth), high-pitched crying, little appetite or problems with feeding, muscle spasms, intellectual and developmental disabilities, impaired motor function, and seizures.

Embryogenesis #1

Blastula attaches to wall of uterus

• What is the function of E3 ubiquitin ligases?

E3 ubiquitin ligases are a family of proteins that participate in a reaction that ubiquitinates substrate proteins, which leads to their degradation by the proteosome.

Embryogenesis #7

Embryonic folding completes *SEE DIAGRAM p. 8!!!*

Embryogenesis #6

Embryonic folding: trilaminar structure-> a cylinder w/ 3 "layers" -Ectoderm folds over to form headfold, midgut and tailfold (from left to right) *looks sort of like a mushroom cap

Blastula

Hollow ball of cells w/ fluid in the middle

• How does Zika pass through the placenta to reach the fetus?

ZIKV passes through the placenta by infecting placental cells. Placental macrophages (known as Hofbauer cells) and trophoblasts are the main target cells of ZIKV infection. ZIKV efficiently targets human cortical NPCs, whereas immature neurons are less susceptible to ZIKV infection. Astrocytes, microglia, and oligodendrocyte precursor cells located throughout the developing cortex can be productively infected by ZIKV. The critical period for infection appears to be between 12 and 16 weeks gestation (note this overlaps the critical period for microcephaly caused by radiation).

Embryogenesis #5

Neurulation takes place (top becomes epidermis)-> neural tube (already had notochord) *See p. 6!!*

Phocomelia

congenital condition in which the proximal portions of the limbs are poorly developed or absent

• Why is thalidomide still prescribed today?

found to be highly effective at treating multiple myeloma (a cancer of white blood cells). Thalidomide also is a potent anti-inflammatory that blocks TNF-alpha by reducing the half-life of its mRNA. This eases the symptoms of the bacterial infection, leprosy

Embryoblast

inner cell mass in the blastula

Point Mutations

insertion, deletion, frame shift, transition, transversion

Chromosomal mutations

insertion, deletion, translocation, inversion, aneuploidy (monosomy, trisomy, polypoidy

Morula

Solid ball of cells

Lateral plate mesoderm->

Somatic mesoderm: bones, ligaments, dermis, and parietal (outer) layer of serous membranes Splanchnic mesoderm: visceral (inner) layer of serous membranes, blood vessels, CT of visceral organs

Paraxial mesoderm->

Somites: Skeletal muscles, CT, ribs and vertebrae

radiation

Take radiation for example. Radiation is fast moving energy emitted as particles or waves. It is commonly divided into two categories: non-ionizing and ionizing. Non-ionizing radiation is low frequency radiation that disperses energy through heat and increased molecular movement. It includes visible light, ultraviolet light, microwaves, ultrasounds, MRIs, etc. Ionizing radiation is dangerous high frequency radiation that includes alpha and beta particles, gamma rays, and X-rays. Ionizing radiation can directly damage proteins, DNA, and other molecules in cells by transferring so much energy that it frees electrons from atoms. Ionizing radiation can also generate free radical species that then go on to damage other molecules. Radiation is a teratogen, mutagen, and carcinogen.

Describe the policies (or lack thereof) in the 1950's that allowed the thalidomide tragedy to occur and the regulatory changes that were made in response.

the FDA had 60 days to review a drug application, otherwise it was given automatic approval. Regulatory changes abolished automatic drug approval, shifting the burden to the drug makers to scientifically prove that a new drug is safe and effective using the best contemporary methods, including using more than one animal model. Clinical trials of new drugs were required to undergo regulatory approval and oversight, and subjects had to give informed consent before participating. Drug makers had to share their internal records pertinent to drug safety and efficacy, throughout the testing and manufacturing process. The government also established authority over drug advertising and labeling and the power to immediately recall hazardous drugs. These monumental changes shifted the balance of power from the drug industry to governmental bodies.

Teratogen: Thalidomide

underdeveloped or no limbs

Teratogen: cigarette smoke

unfused pallet/lip (cleft lip/pallet)

Intermediate mesoderm->

urogenital system

• Describe the signaling pathways that pattern the limb bud. What is the hypothesis for how thalidomide might interfere with limb bud development? --last one needs answers

**simplify Researchers have studied how thalidomide blocks the development of the limb buds. Growth of the limb bud is regulated through a feedback system. At the distal part of the limb bud, there is a special ridge of ectoderm called the apical ectodermal ridge (AER). This highly proliferative area of cells lays down the limb tissue beginning with the proximal elements and continuing through the distal elements as the limb grows outward. The zone of polarizing activity (ZPA) at the base of the limb bud is an organizer that patterns the anterior-posterior axis of the limb. The ZPA secretes sonic hedgehog (Shh) which activates Ghremlin (Gre), which inhibits Bone morphogenic proteins (Bmp), which inhibit the production of Fibroblast growth factor (Fgf) in the AER. Thus, Shh secretion by the ZPA increases the activity of Fgf in the AER and the limb bud grows outward. However, when Fgf levels are high, Fgf actually starts to inhibit the activity of Gre (a self-regulating feedback loop), allowing Bmp's to become active and down regulate Fgf activity. This keeps Fgf levels at the correct level. Limb bud out-growth ceases when the limb has become long enough that the levels of Shh reaching the AER are too low to activate Gre. ---important---The model that has been proposed is that thalidomide binds to cereblon and inhibits its ability to ubiquitinate one or more of its substrates-

Mutagens

-Agents that induce mutations (often via DNA damage) -Mutations can lead to cancer (but remember mutations are also the fodder for evolution!) -occur in germline (entire organism has mutation) or somatic cells (part of organism has mutation, not in gametes) *must be in germline to be heritable A mutagen is an agent that causes changes to the sequence of an individual's DNA. If a mutagen can cause chrosomsomal breakages and rearrangements, such as translocations, deletions, and inversions, it is a clastogen. If it can cause aneuploidy, it is an aneuploidogen. Mutagens also cause less drastic changes: single base pair substitutions and small insertions and deletions. Finally, viral elements that insert themselves into a host genome are mutagens.

Preimplantation Development

-All happens in fallopian tubes -First 3 stages controlled almost entirely by maternal mRNA

Carcinogens

-Chemicals that induce uncontrolled cell division -Many carcinogens are mutagens -Some carcinogens are actually not carcinogenic until they are metabolized (e.g., by hepatocytes) via biotransformation Somatic mutations in these key oncogenes and tumor suppressors cause cancer to develop in that cell. However, not all carcinogens act by mutagenizing the DNA. Carcinogenic agents could also interfere with the immune system, signaling, or cell behavior in a way that accelerates cancer growth.

Embryogenesis #3

-Connecting stalk forms -amniotic cavity and yolk sac get larger -extraembryonic mesoderm forms

Factors that affect teratology

-Dose and frequency of exposure -Thresholds -Genetics -Timing and critical developmental windows

Embryogenesis #2

-Embryoblast-> bilaminar disk: epiblast & hypoblast (what becomes the embryo) *epiblast closest to uterine wall -amniotic sac starts to form (b/t epiblast and uterine wall side)

Carcinogenesis

-a progression of changes at cellular, genetic, and epigenetic level that reprogram a cell to undergo *uncontrolled cell division*, thus forming a malignant mass

Teratogens

-chemicals that *cause birth defects* (death, malformation, growth retardation, functional defect) -Medications, recreational drugs, alcohol, infectious agents, or environmental contaminants -Usually interfere with *cell differentiation* in a developing *embryo/fetus*

• What is the threshold dose for a teratogen?

2) Most teratogens have a dose below which all embryos are unaffected. This is called the threshold dose. Below it, the embryos are safe. There is no similar completely safe dose of mutagens. A degree of risk that a critical gene could be randomly mutated exists at any dose of mutagen.

Periods of fetal development:

3 weeks: CNS 3.5 weeks: Heart 4.5 weeks: ears, eyes, limbs 6.75 weeks: teeth, palate 7 weeks: external genitalia

• Discuss how the timing of exposure to a teratogen affects the severity of a birth defect.

3) There is a critical period during development during which a teratogen causes its associated birth defects. The teratogen has to be present when the target organ is developing to be able to disrupt its formation. In contrast, mutagens can act at any time during a person's life to cause cancer causing mutations. 4) For both teratogens and mutagens, the fraction of people who develop disease increases with increasing dose. The higher dose of mutagen, the more likely a critical cancer gene will be randomly mutated and cancer will begin. The higher the dose of teratogen, the more likely an embryo will be above its individual maximum tolerance level.

Teratogen: Rubella Virus

Microcephaly, PDA, cataracts

• How was it concluded that the Zika virus causes microcephaly?

The results revealed the presence of the ZIKV in samples of their amniotic fluid. Shortly thereafter, the virus was also found in the brains of miscarried fetuses with microcephaly. ZIKV was then found to infect and kill neuronal progenitor cells in model systems. C

• What are the modes of transmission and process of replication of ZIKV?

Transmission of ZIKV is mostly through zoonosis. ZIKV is spread between people by the Aedes aegypti mosquito, which lives throughout the world in tropical areas. When a mosquito bites someone, it injects some fluids into the skin to numb the area. The virus is passed into the person within these fluids. If a person is experiencing an active infection, the virus can be picked up by mosquitoes when they drink that person's blood. The virus cannot survive outside of the body. Most human infections of ZIKV are transmitted by the Aedes mosquito, but the virus can also spread directly through sexual contact and blood transfusions.

• What birth defects are caused by exposure to thalidomide? What is the time sensitive window and dose threshold for thalidomide syndrome?

primarily seen to the limbs (upper limbs are more commonly affected than lower limbs), ears, heart, kidney, and genitals. Thumb abnormalities are the most frequent deformity of thalidomide syndrome. Phocomelia (69%), Genital defects (3%) include absence of the testis and malformation of the uterus. the most time sensitive window for thalidomide syndrome is thought to occur between days 20 and 36 after fertilization. This corresponds to Carnegie stages 12 and 13, Exposure on days 20-24 leads to anotia, while exposure 25-29 leads to microtia. Exposure from days 24-29 causes upper limb defects, while exposure from days 27-31 causes lower limb defects. Triphalangism was determine to arise from exposure on days 32-36. Reports indicate that a single 50 mg tablet of thalidomide during the time-sensitive window is sufficient to cause birth defects in 50% of pregnancies.

• What is the hypothesis for why ZIKV spread from Southeast Asia to the Americas after 2013?

single substitution (Ser139 to Asn) in the viral polyprotein substantially increased ZIKV infection in model systems, and an infected person travelled over, and since everyone was not exposed before, and it was stronger, it spread easier

Teratogen: Alcohol

small head, small eyes, short nose, thin lip, behavioral issues