abstract 159141 Use of a human breast cancer cell proliferation
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Use of a human breast cancer Use of a human breast cancer cell proliferation assay as cell proliferation assay as an exposure assessment tool an exposure assessment tool for total bioaccumulated for total bioaccumulated xenoestrogens in channel xenoestrogens in channel catfish caught in catfish caught in Pittsburgh’s three rivers. Pittsburgh’s three rivers. Frank Houghton PhD, Conrad Volz DrPH, MPH, Yan Liu Frank Houghton PhD, Conrad Volz DrPH, MPH, Yan Liu MPH, Christopher Price, Mary Elm BS, Devra Lee Davis MPH, Christopher Price, Mary Elm BS, Devra Lee Davis PhD, MPH, Maryann Donovan MPH, PhD, and Patricia K PhD, MPH, Maryann Donovan MPH, PhD, and Patricia K Eagon PhD. Eagon PhD. Department of Gastroenterology, Hepatology and Nutrition, School Department of Gastroenterology, Hepatology and Nutrition, School of Medicine, University of Pittsburgh, Department of Environmental of Medicine, University of Pittsburgh, Department of Environmental and Occupational Health, Graduate School of Public Health, and Occupational Health, Graduate School of Public Health, University of Pittsburgh, Center for Environmental Oncology, University of Pittsburgh, Center for Environmental Oncology, University of Pittsburgh Cancer Institute. University of Pittsburgh Cancer Institute.
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Transcript of abstract 159141 Use of a human breast cancer cell proliferation
Abstract #159141Abstract #159141Use of a human breast cancer Use of a human breast cancer cell proliferation assay as an cell proliferation assay as an exposure assessment tool for exposure assessment tool for total bioaccumulated total bioaccumulated xenoestrogens in channel xenoestrogens in channel catfish caught in Pittsburgh’s catfish caught in Pittsburgh’s three rivers.three rivers.Frank Houghton PhD, Conrad Volz DrPH, MPH, Yan Liu Frank Houghton PhD, Conrad Volz DrPH, MPH, Yan Liu MPH, Christopher Price, Mary Elm BS, Devra Lee Davis MPH, Christopher Price, Mary Elm BS, Devra Lee Davis PhD, MPH, Maryann Donovan MPH, PhD, and Patricia K PhD, MPH, Maryann Donovan MPH, PhD, and Patricia K Eagon PhD.Eagon PhD. Department of Gastroenterology, Hepatology and Nutrition, School of Department of Gastroenterology, Hepatology and Nutrition, School of Medicine, University of Pittsburgh, Department of Environmental and Medicine, University of Pittsburgh, Department of Environmental and Occupational Health, Graduate School of Public Health, University of Occupational Health, Graduate School of Public Health, University of Pittsburgh, Center for Environmental Oncology, University of Pittsburgh, Center for Environmental Oncology, University of Pittsburgh Cancer Institute.Pittsburgh Cancer Institute.
Accumulation of xenoestrogenic compounds in waterways is a world-wide public health problem.
Fish are sensitive sentinels of waterway xenoestrogen contamination.
Introduction
Effects of xenoestrogens on male fish
• Disruption of testicular development
• Elevation of serum vitellogenin levels
• Conversion to intersex phenotype
Potential sources of xenoestrogens in Pittsburgh’s
three rivers• Legacy waste from steel mill sites
• Untreated sewage
• Residential waste
• Agricultural runoff
Routes of exposure to xenoestrogens in water
• Recreational activities
• Subsistence fishing
• Municipal drinking water
AimTo assess the efficacy of a cell proliferation assay (CPA) to determine the presence of estrogenic substances in the flesh of channel catfish caught in Pittsburgh’s three rivers
Methods
• Fish caught• Location of fish catch recorded• Biometric parameters recorded• Fish fillets extracted• Extracts tested for estrogenicity
via cell proliferation & estrogen receptor binding assays
Extraction Method• 1g (±5%) flesh, skin, and fat
• Extracted with CHCl3/CH3OH (9:1 v/v)
• Reduced to residue using N2 gas
• Stored at -20OC under N2
• Solubilized in ethanol/glycerol (70:30 v/v)
Cell proliferation assay (CPA)Cell lines used:
MCF-7: Estrogen receptor-positive BT20: Estrogen receptor-negative
Cell treatments:Physiological estrogen (estradiol, 10-9M)Fish extracts (varying doses)Medium only (negative control)
Cell number assessed: CellTiter96Proliferation index (PI)
Absorbance of treated well
Mean of negative control wells (n=10)PI=
Estrogen response profile(idealized)
Contro
l
Estrad
iol
1/400
0
1/300
0
1/200
0
1/150
0
1/100
01/5
001/2
001/1
000.000.250.500.751.001.251.501.752.002.25
No Response
Strong ResponseModerate Response
Treatment
Pro
lifer
atio
n In
dex
Estrogen response profile(channel catfish 138)
0.00
0.25
0.50
0.75
1.00
1.25
1.50
1.75
2.00
Treatment
Pro
lifer
atio
n In
dex
no response
Estrogen response profile(channel catfish 100)
0.00
0.25
0.50
0.75
1.00
1.25
1.50
1.75
2.00
Treatment
Pro
lifer
atio
n In
dex
moderate response
Estrogen response profile(channel catfish 47)
0.000.250.500.751.001.251.501.752.002.25
Treatment
Pro
lifer
atio
n In
dex
strong response
CPA Results• N=21
• 1 extract exhibited a strong response
• 13 extracts displayed a moderate response
• 7 extracts produced no response
Highland Park
Point State Park
Kittanning,Armstrong County
Monongahela River
Results
In general, extracts that are stimulators of cell proliferation also compete for ER binding (data not presented)
Conclusions
These results support the hypothesis that bioaccumulated xenoestrogens are highest in channel catfish caught nearest to legacy-contaminated locations and areas of untreated sewage overflows.
Community participants:
Venture Outdoors
Clean Water Action
Bassmasters
Individual local anglers
Support:
The DSF Charitable Trust & The Heinz Foundation through the Center for Environmental Oncology,
University of Pittsburgh Cancer Institute, and
The Veterans Research Foundation of Pittsburgh
Acknowledgements
