Sequential analysis of kidney stone formation in the Aprt knockout mouse.
BACKGROUND
We have previously shown that, as in human adenine phosphoribosyltransferase (APRT) deficiency, Aprt knockout mice form 2,8-dihydroxyadenine (DHA) renal stones. The disease develops earlier and is more severe in male than in female mice. To examine the biological bases for these differences, the area occupied by DHA crystals was quantified in kidney sections from male and female mice (strain 129) aged one day to eight months and this parameter was correlated with changes in renal histopathology. Aprt heterozygous and wild-type mice were used as controls.


METHODS
Following anesthesia, the left kidney was removed and immediately frozen in dry ice. Unstained cryosections were examined by polarized light to determine total area of birefringent particles. The right kidney was perfused and embedded in plastic, and stained sections were viewed by light microscopy to examine the histopathology and to determine the location of the birefringent particles. A pathological score was assigned to the histological findings. The scores from the right kidney were compared with crystal/particle area in the left kidney, and the data were analyzed using two-way analysis of variance. The chemical composition of the particles was determined by x-ray diffraction analysis. Several stone fragments from the bladder were also examined by scanning electron microscopy (SEM).


RESULTS
Crystals were detected in kidney sections from one- to two-day-old Aprt knockout mice. The crystal burden remained low in both sexes throughout the study except in males at the 120- to 240-day period. Furthermore, there was a substantial degree of renal pathology, primarily seen as interstitial fibrosis, in those males with a very high level of stone formation. The crystalline material was identified as 6-amino-2,8(3,9)-purine dione, a tautomeric form of DHA. SEM indicated that the crystals were spherical, with a diameter of 10 to 20 microm. Tissue staining and fixation procedures dramatically reduced the amount of birefringent material in kidney sections. Aprt heterozygotes of both sexes had low levels of crystalline material in the kidneys and no pathology. Birefringent material or pathological changes were not seen in kidneys from wild-type mice.


CONCLUSIONS
Both male and female Aprt knockout mice accumulate DHA. However, the area occupied by DHA crystals was significantly greater in 120- to 240-day-old males compared with the females of similar age. Also, substantial renal pathology was detected in kidneys of male mice that had very high levels of stone material.
Evan
A P
AP
Department of Anatomy and Cell Biology, Indiana University School of Medicine, Indianapolis, Indiana 46202-5120, USA. evan@anatomy.iupui.edu
Bledsoe
S B
SB
Connors
B A
BA
Deng
L
L
Liang
L
L
Shao
C
C
Fineberg
N S
NS
Grynpas
M D
MD
Stambrook
P J
PJ
Youzhi
S
S
Sahota
A
A
Tischfield
J A
JA
D000225:Adenine / Q000031:analogs & derivatives* / Q000378:metabolism; D000228:Adenine Phosphoribosyltransferase / Q000172:deficiency* / Q000235:genetics; D000367:Age Factors; D000818:Animals; D005260:Female; D005838:Genotype; D007669:Kidney Calculi / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D007686:Kidney Tubules, Distal / Q000737:chemistry / Q000473:pathology*; D007687:Kidney Tubules, Proximal / Q000737:chemistry / Q000473:pathology*; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D008855:Microscopy, Electron, Scanning; D012737:Sex Factors; D016707:Tissue Fixation; D014961:X-Ray Diffraction
C002993:2,8-dihydroxyadenine; D000228:Adenine Phosphoribosyltransferase; D000225:Adenine