BIO 4127 8 - Endocrine Disrupting Chemicals

What are Endocrine Disrupting Chemicals?

Chemicals that mimic or upset hormone levels

- Not designed to do that

- Effects are very damaging

- Ex: DDT and bird eggs

EDC examples

Estrogens, Atrazine, Prozac

Estrogens in the aquatic environment (1980s)

Started outside of London, UK in the 1980s. Fisherman reported that fish had both testes and ovaries

What was measured in the fish in 1980s UK?

Vitellogenins were measured in male fish. Found that there were pollutants in the river that were estrogenic (E1, E2, EE2).

E1

estrone

E2

estradiol

EE2

Ethynylestradiol

- adding ethylene group to E2, increases receptor affinity and half-life

- used in contraceptive pill

Sexual Development in frogs (fish, turtle) and estrogens

The functional sex of a fish, frog, or turtle can be changed because hormones (Estrogens) can override genetics (even if they have sex chromosomes)

In frogs, you can produce all male/female populations by changing what?

the hormones they're exposed to as tadpoles

Estrogens and estrogenic molecules (6)

- Estradiol (E2)

- Ethinylestradiol (EE2)

- Methoxychlor

- Octylphenol

- B-Sitosterol

- Bisphenol A

What processing do E2 and EE2 undergo to become water soluble?

They are sulfonated and glucuronated to become water soluble (less biologically active as well) to be excreted

How are estrogens re-activated in the environment?

Anaerobic bacteria can de-sulfonate and re-activate it in the environment

Methoxychlor

Insecticide that doesn't seem estrogenic, but it does have low affinity for E2 receptor

Octylphenol

Breakdown product of manufacturing surfactants (both toxic and estrogenic)

- Nonylphenol is also toxic and estrogenic.

- Both are weak estrogens

B-Sitosterol

Phyto-estrogen from plants

Bisphenol A

(BPA) weak estrogen, good at polymerization (was used in plastics)

What was observed in male fish exposed to estrogenic compounds?

Male fish produced vitellogenin (egg-yolk proteins) this is usually the job of female fish liver when exposed to E2

- huge induction VTG mRNA

What is a biomarker to determine if a body of water is contaminated with an EDC?

Vitellogenin in male fish blood

Intersex frog and fish possess?

ovotestes

- can be caused due to exposure to EE2 at sensitive stage

After sex determination in amphibians, what happens?

sexual differentiation occurs

- This is a steroid sensitive phase of development. In frogs this occurs usually before there is an increase in TH and metamorphosis

How did inappropriate EE2 exposure change the sex ratio in frogs?

EE2 shifted the sex ratio towards females and increased the incidence of intersex

- from 50/50 to 70%f, 10%m, 20%i

Does North American water treatment treat for estrogens?

No, estrogens are entering our water

Atrazine

a widely used herbicide

- Pre-emergent, agricultural, spring application

Atrazine as an EDC

Known to affect sex hormone production in mammals, reptiles - likely acting on aromatase

- Same effects on human cells in vitro

- Dramatically alters sexual development in fish and frogs

Atrazine and testosterone

Atrazine decreases testosterone by stimulating aromatase to increase the conversion of testosterone to estrogen

- seen in fish

Atrazine 0.1 ppb (ng/L) causes what in Leopard frogs?

development of hermaphrodite male

- oocyte-like structures inside testes

Atrazine and gonadal development

Causes gonadal abnormalities

At certain atrazine concentrations it can induce gonadal dysgenesis and testicular oogenesis

- at higher concentrations there are less abnormalities

What proportion of North Americans are on antidepressants?

1 in 10 North Americans

- 250,000 million prescriptions annually

3 most commonly prescribed antidepressants

Sertraline (Zoloft), Fluoxetine (Prozac), Paroxetine (Paxil)

Fluoxetine (FLX, Prozac) mechanism of action

selective serotonin reuptake inhibitor

Fluoxetine in the environment is resistant to...?

photolysis and hydrolysis

- Occurs in norfluoxetine as well

- Adsorps to sludge

Concentrations of FLX in the water

up to 540 ng/L

Effects of waterborne FLX

Not a toxic effect at low levels, but it will disrupt normal endocrine functions

- Environmental FLX is relatively low

What is the research paper investigating?

Transgenerational effects of FLX on cortisol and behaviour in male zebrafish

Serotonin roles

*Programming of the stress axis

- Mood

- Social interaction

- Stress

- Appetite

- Reproduction

- Growth

What is the problem with FLX?

FLX is highly prescribed

- pre-partum and post-partum depression too

- Some women, while pregnant, can be prescribed antidepressants

Does FLX cross the placenta?

Yes, it was measured in amniotic fluid and cord blood

FLX paper experimental methods

Exposed F0 male zebrafish to either high or low dose SSRI for 6 days after fertilization and then tracked generations. Stress response measured at 6 months old

FLX effect on stress response across generations

F0 to F3 - fish exposed to SSRI had lower basal CORT and lower CORT upon stress compared to control

- Conclusion: FLX reduces cortisol levels in male adults

FLX effect on steroidogenesis

FLX modulates pathways associated with steroidogenesis in kidney. Affected many enzymes involved in steroidogenesis

- studied with RNA-Seq

Behaviour assay used in paper

Novel tank test

- when placed in a new tank, a zebrafish should explore the tank

Novel tank test on FLX results

In F0 both low and high dose FLX zebrafish exhibited lower exploratory behaviour compared to control, but by F1 it was restored in low dose.

High FLX decreases exploratory behaviour and only is returned in generation 3 (F3)

What is observed when cortisol synthesis is chemically inhibited in male zebrafish?

We see similar effects to the FLX exposed fish

- Showed that it is through the inhibition of cortisol that there is decreased locomotive behaviour

- MET treated fish did not have a stress response

Implications of results of FLX in zebrafish paper

Cortisol disruption by FLX is of high concern for humans and aquatic animals.

In humans, chronic hypocortisolism, has been associated with burnouts, chronic fatigue, fibromyalgia, immune disorders, and post-traumatic stress disorder

The children of moms who took antidepressants, were studied and they had low CORT growing and were disengaged in class