A PILOT STUDY OF BIOMONITORING AMBIENT GENOTOXIC EXPOSURE

OF CHILDREN

A PILOT STUDY OF BIOMONITORING AMBIENT GENOTOXIC EXPOSURE

OF CHILDREN

L. Anna, B. Schoket, E. Győrffy, E. Erdei, P. Rudnai

National Institute of Environmental Health,

József Fodor National Centre for Public Health,

Budapest, Hungary

L. Anna, B. Schoket, E. Győrffy, E. Erdei, P. Rudnai

National Institute of Environmental Health,

József Fodor National Centre for Public Health,

Budapest, Hungary

AIM OF THE STUDYAIM OF THE STUDY

• To assess genotoxic exposure of children

to air pollution

• To explore the suitability of aromatic DNA adducts

as a biomarker of genotoxic exposure of children

• To assess genotoxic exposure of children

to air pollution

• To explore the suitability of aromatic DNA adducts

as a biomarker of genotoxic exposure of children

EPIDEMIOLOGICAL BACKGROUND OF THE STUDYEPIDEMIOLOGICAL BACKGROUND OF THE STUDY

• Investigation of the respiratory health of 1800 children of 8 to 10 years of age (Nov-Dec 1998)

• Six towns in Hungary, three of those with elevated air pollution

• Exposure and health questionnaires filled in by the parents

• Investigation of the respiratory health of 1800 children of 8 to 10 years of age (Nov-Dec 1998)

• Six towns in Hungary, three of those with elevated air pollution

• Exposure and health questionnaires filled in by the parents

STUDY POPULATION FOR THE BIOMONITORING STUDY

STUDY POPULATION FOR THE BIOMONITORING STUDY

Exposure categories

Polluted towns Control towns

ETS & Traffic ETS & TrafficETS only ETS onlyTraffic only Traffic onlyNo ETS & No traffic No ETS & No traffic

Exposure categories

Polluted towns Control towns

ETS & Traffic ETS & TrafficETS only ETS onlyTraffic only Traffic onlyNo ETS & No traffic No ETS & No traffic

N = 105N = 105

METHODOLOGYMETHODOLOGY

• Isolation of DNA from frozen whole blood (ca. 1.5 ml)

• Detection of aromatic DNA adducts by 32P-postlabelling

• Radioactivity measurement by electronic autoradiograph

• Statistical analyses by Fisher’s exact test and Mann-Whitney U test

• Isolation of DNA from frozen whole blood (ca. 1.5 ml)

• Detection of aromatic DNA adducts by 32P-postlabelling

• Radioactivity measurement by electronic autoradiograph

• Statistical analyses by Fisher’s exact test and Mann-Whitney U test

STRUCTURE OF BENZO[A]PYRENE DIOL-EPOXIDE-DNA ADDUCT

STRUCTURE OF BENZO[A]PYRENE DIOL-EPOXIDE-DNA ADDUCT

TYPICAL TWO-DIMENSIONAL DNA ADDUCT MAPS FROM CHILDREN’S BLOOD

a

ed

cb

f Blank

PERCENTAGE OF BLOOD SAMPLES WITH DNA ADDUCT SPOTS IN THE

POLLUTED AND CONTROL TOWNS

PERCENTAGE OF BLOOD SAMPLES WITH DNA ADDUCT SPOTS IN THE

POLLUTED AND CONTROL TOWNS

Pollute

d (49)

Control (

56)

Pollute

d (49)

Control (

56)

0

20

40

60

80

100

MAIN SPOT SECONDARY SPOT

Yes

No

Det

ecta

ble

sp

ot

(% o

f al

l sam

ple

s)

Pollute

d (49)

Control (

56)

Pollute

d (49)

Control (

56)

0

20

40

60

80

100

MAIN SPOT SECONDARY SPOT

Yes

No

Det

ecta

ble

sp

ot

(% o

f al

l sam

ple

s)

PERCENTAGE OF BLOOD SAMPLES WITH

DNA ADDUCT SPOTS

PERCENTAGE OF BLOOD SAMPLES WITH

DNA ADDUCT SPOTS

POLLUTED TOWNSPOLLUTED TOWNS CONTROL TOWNSCONTROL TOWNS

ETS&Traffi

c (18)

ETS only

(13)

Traffi

c only

(10)

No ETS&No T

raffi

c (8)

ETS&Traffi

c (18)

ETS only

(13)

Traffi

c only

(10)

No ETS&No T

raffi

c (8)

0

20

40

60

80

100

MAIN SPOT SECONDARY SPOT

Yes

No

Det

ecta

ble

sp

ot

(% o

f al

l sam

ple

s)

ETS&Traffi

c (7)

ETS only

(11)

Traffi

c only

(8)

No ETS&No T

raffi

c (30)

ETS&Traffi

c (7)

ETS only

(11)

Traffi

c only

(8)

No ETS&No T

raffi

c (30)

0

20

40

60

80

100

MAIN SPOT SECONDARY SPOT

Yes

No

Det

ecta

ble

sp

ot

(% o

f al

l sam

ple

s)

ETS&Traffi

c (18)

ETS only

(13)

Traffi

c only

(10)

No ETS&No T

raffi

c (8)

ETS&Traffi

c (18)

ETS only

(13)

Traffi

c only

(10)

No ETS&No T

raffi

c (8)

0

20

40

60

80

100

MAIN SPOT SECONDARY SPOT

Yes

No

Det

ecta

ble

sp

ot

(% o

f al

l sam

ple

s)

ETS&Traffi

c (7)

ETS only

(11)

Traffi

c only

(8)

No ETS&No T

raffi

c (30)

ETS&Traffi

c (7)

ETS only

(11)

Traffi

c only

(8)

No ETS&No T

raffi

c (30)

0

20

40

60

80

100

MAIN SPOT SECONDARY SPOT

Yes

No

Det

ecta

ble

sp

ot

(% o

f al

l sam

ple

s)

ODDS RATIOS FOR DNA ADDUCT FORMATION

DUE TO ELEVATED AIR POLLUTION

(Main adduct spot)

ODDS RATIOS FOR DNA ADDUCT FORMATION

DUE TO ELEVATED AIR POLLUTION

(Main adduct spot)

Exposure OD 95% CI P

Polluted vs. Control 2.59 1.17 - 5.73 0.029

Polluted, ETS & Traffic vs.Control, No ETS & No Traffic 5.20 1.44 – 18.72 0.016

Exposure OD 95% CI P

Polluted vs. Control 2.59 1.17 - 5.73 0.029

Polluted, ETS & Traffic vs.Control, No ETS & No Traffic 5.20 1.44 – 18.72 0.016

INDIVIDUAL VARIATION IN AROMATIC DNA ADDUCT

LEVELS IN BLOOD SAMPLES FROM CHILDREN

INDIVIDUAL VARIATION IN AROMATIC DNA ADDUCT

LEVELS IN BLOOD SAMPLES FROM CHILDRENValues for the main adduct spotValues for the main adduct spot

Polluted townsPolluted towns Control townsControl towns

ETS&Traf

fic (1

8)

ETS (13)

Traf

fic (1

0)

NoETS&NoTraf

fic (8

)

ETS&Traf

fic (7

)

ETS (11)

Traf

fic (8

)

NoETS&NoTraf

fic(3

0)0.0

0.5

1.0

1.5

DN

A a

dd

uc

ts/1

08

nu

cle

oti

de

s

ETS&Traf

fic (1

8)

ETS (13)

Traf

fic (1

0)

NoETS&NoTraf

fic (8

)

ETS&Traf

fic (7

)

ETS (11)

Traf

fic (8

)

NoETS&NoTraf

fic(3

0)0.0

0.5

1.0

1.5

DN

A a

dd

uc

ts/1

08

nu

cle

oti

de

s

P=0.02for theMeans

P=0.02for theMeans

AROMATIC DNA ADDUCT LEVELS IN BLOOD SAMPLES FROM CHILDRENAROMATIC DNA ADDUCT LEVELS IN BLOOD SAMPLES FROM CHILDREN

Polluted townsPolluted towns Control townsControl towns

P=0.002P=0.002** **

Values for the main spot (Mean ± SD)Values for the main spot (Mean ± SD)

ETS&Traf

fic (1

8)

ETS (13)

Traf

fic (1

0)

NoETS&NoTraf

fic (8

)

ETS&Traf

fic (7

)

ETS (11)

Traf

fic (8

)

NoETS&NoTraf

fic(3

0)0.0

0.2

0.4

0.6

0.8

DN

A a

dd

uc

ts/1

08

nu

cle

oti

de

s

ETS&Traf

fic (1

8)

ETS (13)

Traf

fic (1

0)

NoETS&NoTraf

fic (8

)

ETS&Traf

fic (7

)

ETS (11)

Traf

fic (8

)

NoETS&NoTraf

fic(3

0)0.0

0.2

0.4

0.6

0.8

DN

A a

dd

uc

ts/1

08

nu

cle

oti

de

s

CONCLUSIONSCONCLUSIONS

• DNA adducts were detectable from frozen whole blood

• Detection limit was as low as 0.02-0.07 adducts in 108 nucleotides

• Two characteristic distinct adduct spots were related to the exposure

• Statistically significant difference was observed in DNA damage between the highest and lowest exposure categories

• Aromatic DNA adduct was a sensitive biomarker of genotoxic exposure of children to air pollution

• DNA adducts were detectable from frozen whole blood

• Detection limit was as low as 0.02-0.07 adducts in 108 nucleotides

• Two characteristic distinct adduct spots were related to the exposure

• Statistically significant difference was observed in DNA damage between the highest and lowest exposure categories

• Aromatic DNA adduct was a sensitive biomarker of genotoxic exposure of children to air pollution