Genetic analysis of DNA excreted in urine: a new approach for detecting specific genomic DNA sequences from cells dying in an organism.
BACKGROUND
Cell-free DNA from dying cells recently has been discovered in human blood plasma. In experiments performed on animals and humans, we examined whether this cell-free DNA can cross the kidney barrier and be used as a diagnostic tool.


METHODS
Mice received subcutaneous injections of either human Raji cells or purified (32)P-labeled DNA. DNA was isolated from urine and analyzed by measurement of radioactivity, agarose gel electrophoresis, and PCR. In humans, the permeability of the kidney barrier to polymeric DNA was assessed by detection in urine of sequences that were different from an organism bulk nuclear DNA.


RESULTS
In the experiments on laboratory animals, we found that approximately 0.06% of injected DNA was excreted into urine within 3 days in a polymeric form and that human-specific ALU: sequences that passed through the kidneys could be amplified by PCR. In humans, male-specific sequences could be detected in the urine of females who had been transfused with male blood as well as in DNA isolated from urine of women pregnant with male fetuses. K-ras mutations were detected in the urine of patients with colon adenocarcinomas and pancreatic carcinomas.


CONCLUSIONS
The data suggest that the kidney barrier in rodents and humans is permeable to DNA molecules large enough to be analyzed by standard genetic methodologies.
Institute of Carcinogenesis, Cancer Research Center, Moscow, Russia 115478.
I
I
Botezatu
O
O
Serdyuk
G
G
Potapova
V
V
Shelepov
R
R
Alechina
Y
Y
Molyaka
V
V
Ananév
I
I
Bazin
A
A
Garin
M
M
Narimanov
V
V
Knysh
H
H
Melkonyan
S
S
Umansky
A
A
Lichtenstein
D000818:Animals; D001803:Blood Transfusion; D016923:Cell Death; D015179:Colorectal Neoplasms / Q000737:chemistry / Q000473:pathology / Q000652:urine; D004247:DNA / Q000235:genetics / Q000652:urine*; D004587:Electrophoresis, Agar Gel; D005260:Female; D005333:Fetus / Q000737:chemistry; D011905:Genes, ras; D016678:Genome; D006801:Humans; D007668:Kidney / Q000378:metabolism; D008297:Male; D051379:Mice; D008809:Mice, Inbred C3H; D009154:Mutation; D010190:Pancreatic Neoplasms / Q000737:chemistry / Q000652:urine; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D014407:Tumor Cells, Cultured; D014998:Y Chromosome
D004247:DNA