Biomarkers

Feb 11, 2014

Biomarkers

Biochemical, physiological or histological changes

in organisms that are can be used to estimate either

exposure to chemicals or resultant effects

Behavioural changes??

Current emphasis is on Early Warning indicators of

stress at higher levels of organization

- individual

- population

- community

Two most relevant biomonitoring tools are relevant to:

1) Detoxification or adaptive responses

2) Reproductive effects

Biomarkers

A special type of endpoint

van der Oost R, Beyer J, Vermeulen NPE. 2003 Fish bioaccumulation and biomarkers in environmental risk assessment: a review. Environmental Toxicology and Pharmacology 13(2):57-149.

Advantages of Biomarkers

1) Rapid results

2) Subpopulation can be sampled

- vs. large scale population # monitoring

3) most yield info regarding actual toxic

mechanisms

Disadvantages of Biomarkers

1) Predicative capabilities are tenuous

2) Limited by data regarding “normal”

levels

3) lack of understanding by public and

funders

Criteria for Effective Biomarkers

1) General Specificity

Does the marker respond to one chemical

or is it a general indicator of stress?

Specific markers are more useful for

determining exposure to specific chemicals

but general markers are better for overall

health assessment.

2) Sensitivity

How sensitive is the marker in responding

to stress compared with traditional

measures of health? Eg. Condition Factor

Are there more sensitive biomarkers avail.

that are more predictive than the proposed

marker?

Criteria Cont’d

3) Biological Specificity

Is the marker responsive in just one type of

organisms or can it be used in other species?

Eg. MFO enzymes re not applicable to

molluscs since they metabolize organics

by a separate pathway (MFOs- Consider Later)

4) Interpretation

Can the results easily be interpreted? Are

they easily separated from natural

variation?

5) Timeline

The marker has to respond in a reasonably

short timeframe to be useful as a predictive

tool.

Criteria Cont’d

6) Permanence of Response

Is the response reversible? Is so, how

quickly?

Reversible = useful for establishing cause

and effect (eg. plasma ions)

Permanent = useful for establishing

exposure (eg. cancer)

7) Variability

Does the marker respond similarly in

individuals? What is the range of the

response? How does it compare to

natural variability of the parameter?

2 sources of variability

a) Physiological

Depends on the range of biological

responses exhibited by organisms.

b) Environmental

Variability due to the effects of

external factors. Eg. Ambient temp.

7) Variability cont’d

Criteria Cont’d

8) Linkage to higher levels of organization

Have to be able to answer “So what?”

question.

In early stages of development these

may be less clear…….

Criteria Cont’d

9) Applicability to the Field

Collection techniques, sample prep, and

even interpretation must be easily

transferred.

10) Validation in the Field

Ideally, the marker should have been

tested in wild populations.

11) Methodologies

Important to consider the analytical ease

and cost of using the biomarker and to

note the precision of the assay

(Precision = analytical reproducibility)

Consider 2 Biomarkers

a) MFO Enzymes

b) Metallothionein

1) MFO Enzymes

Mixed Function Oxidases or Mixed Funtion

Oxygenases also called CYP1A (Cytochrome P450

Family 1A)

Group of enzymes that metabolize several different

endogenous and exogenous compounds

10 Different Families in Mammals, >3

in fish and birds

Eg. Fatty acids, steroids, lipids, alcohol

and some contaminants

Activity expressed constitutively at a very low level

Found to respond to coplanar organochlorines

Coplanar Organochlorines

1) PCBs (Polychlorinated Biphenyls)

Cl

Cl

Cl

Cl

Ortho Position

- Cl here does not

favour co-planarity

Binding to Ah Binding to Ah Receptor Receptor

Induction of MFO Enzymes

Binding of Coplanar Organochlorines

To the Ah-Receptor

Translocation

to nucleus

Binding to gene

responsive element

MFO Enzymes As Biomarkers

Data available for variability due to:

sex, season, reproductive cycle,

temperature, handling stress,

dietary effects

BUT………

Links between MFO induction and population

level effects still weak.

Some evidence to suggest induction

is linked to inhibited reproduction, slower

growth and impaired reproduction.

2) Metallothionein

Low molecular weight protein (6-7 kDaltons) with

high cysteine content (30%)

Binds Group IB and IIB metals

Metallothionein cont’d

Each domain can bind different metals at one time

Affinity in the order:

Hg>Ag=Cu>Cd>Zn

But stoichiometry

7 for Hg, Cd, Zn

12 for Cu

12, 17 or 18 for Ag

Found in many aquatic organisms and all

vertebrates studied thus far…..also some inverts

Metallothionein cont’d

Also thought to function in the regulation of

toxicity of metals

2 lines of evidence:

1) Induced tolerance

- organisms pre-exposed to low

concentrations of metals have

greater tolerance for subsequent

exposures

2) Genetic Manipulations

- organisms with inhibited or

enhanced MTN expression have

either greater or lesser susceptibility

to metal toxicity

Metallothionein cont’d

Spillover Hypothesis:

Toxic effects of certain metals begins only

after the binding capacity of MTN is

exceeded.

At that point toxic metals “spill over” to

interact toxically with cellular constituents

Metallothionein cont’d

Other factors affecting MTN:

1) Physiological

developmental state, dietary Zn,

starvation, stress

Reproduction

[MTN] declines just prior to

spawning in fish and returns to

basal levels after spawning

Thought to be due to Zn

requirements to synthesize VTG

Metallothionein cont’d

Other factors affecting MTN:

1) Environmental

- metals

- arthritis, diabetes

- endotoxin

- epinephrine, glucocorticoids

- vitaminC, retinoic acid

- capture stress

- oxidative stress

So are MFO enzymes and Metallothionein useful biomarkers?

What about vitellogenin?

Next Class: Feb. 13

Midterm Exam

In Class

1 hour

Multiple choice with some definitions, short

answers