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Journal of Clinical Microbiology, April 1999, p. 1165-1167, Vol. 37, No. 4
Division of Pathology,
Received 2 July 1998/Returned for modification 9 November
1998/Accepted 21 December 1998
We performed immunohistochemical staining on the nontumorous renal
tissue of 45 patients with renal cancer but without progressive multifocal encephalopathy using JCV-specific antibody. For one patient
we found positive staining of the nuclei of the renal collecting ducts.
Immunoelectron microscopic examination of the positive cell nuclei
revealed electron-dense polyomavirus-like particles.
JC virus (JCV) is a member of the
polyomavirus group of viruses and is the causative agent of progressive
multifocal leukoencephalopathy (PML) (18). JCV infects
humans mostly at a young age without obvious clinical manifestations
and persists in the kidney tissue (6, 15, 19, 21).
Although the renal infection with JCV is not associated with clinically
apparent tissue damage, PML is a devastating central nervous system
disease that mostly occurs in immunocompromised patients (5, 18,
22). JCV has been detected in urine and renal tissue not only of
PML and immunocompromised patients but also those of the
immunocompetent general population (6, 8, 10, 12, 13, 14).
Although there have been many reports of studies of JCV in urine and
kidney tissue by DNA hybridization and/or PCR (6, 4, 7),
histological examinations of the JCV replication site in the kidney are
scarce. Dörries and ter Meulen (9) reported on the
detection of the JCV genome in the cells of renal collecting ducts
in a PML patient by in situ hybridization; however, renal JCV
localization by immunohistochemistry (IHC) and immunoelectron
microscopy (IEM) has not been reported. Recently, we developed a
JCV-specific antibody (JCAb1) that is effective with formalin-fixed
paraffin-embedded tissue (3). Here we report, for the first
time, on the localization of JCV in the kidney of an immunocompetent
patient without PML by IHC and IEM with a nontumorous part of the renal
tissue resected for renal cancer.
Kidney tissues resected for renal cancer from 45 patients were obtained
at the Department of Urology, Branch Hospital, Faculty of Medicine, the
University of Tokyo, and the Department of Urology, Mitui Memorial
Hospital. The kidney tissue collection included kidney tissues from 32 patients previously reported on for the detection of renal JCV
DNA (21). No patients were undergoing immunosuppressive or
anticancer drug therapy. The tissues were obtained from 33 males
and 12 females (average age, 65.1 years). Urine and/or frozen renal
tissues were collected, stored, and submitted for JCV DNA detection by
PCR as reported previously (21). Among the 45 patients, both
urine samples and kidney tissues were available for 38. For four
patients, only urine samples were available. For three patients,
only kidney tissues were available. JCV DNA was detected in the
kidney tissues of 20 of 41 patients and in the urine samples of 19 of
42 patients from whom urine was available. The tumor tissues were
negative for JCV DNA by PCR (data not shown).
Tissues for histological examination were fixed immediately after
nephrectomy in 10% formalin and were embedded in
paraffin. One block containing nontumorous kidney tissue was
selected from each patient, and 4-µm-thick sections were made.
JCAb1, a JCV-specific antibody used throughout this work, reacts
with a decapeptide of the VP1 protein of JCV and has been shown to not
cross-react with the closely related BKV or simian virus 40 (SV40)
polyomaviruses. The specificity of JCAb1 has been confirmed by
positive staining of JCV-infected IMR32 cells, negative staining of
BKV-infected 293 cells, and negative staining of SV40-infected IMR32
cells. The specificity has also been confirmed by positive staining of
the typical JCV-infected oligodendrocytes and astrocytes of
formalin-fixed paraffin sections of brain tissues from patients with
PML (3; unpublished data). Immunohistochemical
staining has been performed as reported previously with the peroxidase LSAB kit (DAKO), with visualization with diaminobenzidine (DAB) (3). Formalin-fixed paraffin sections of brain tissues from patients with PML were used as positive controls throughout this experiment. For the negative controls, JCAb1 was replaced by normal rabbit serum.
For examination by IEM, paraffin sections were processed in the same
way as they were for light microscopic IHC through the DAB coloring
step, but the microwave treatment was skipped to avoid tissue damage.
After visualization with DAB, the sections were processed as described
previously (2).
IHC-positive staining was obtained for tissue from one patient (patient
881073), whose kidney tissue was positive for JCV DNA but whose urine
was not available for examination. The patient had a nephrectomy on 25 April 1988, when he was 51 years old. His postoperative course has been
uneventful, with no signs or symptoms of either PML or a recurrence of
renal cell carcinoma until the time of submission of the manuscript of
this report. In the kidney tissue of patient 881073, strong staining
was observed in the lining epithelial cells of the collecting ducts in
a single area of the renal papilla of the section (Fig.
1A and B), which shows the focal nature
of the intrarenal distribution of JCV infection. Granular positive
staining was recognized in the nuclei of these cells. Some cells with
strongly positive nuclei showed faint cytoplasmic staining. A single to
several positive cells were found in each positive duct, but these were
not associated with degenerative or necrotic changes. Some of the
positive cells appeared to be protruding into the tubular lumen, with
the positive nuclei of the cells protruding into the luminal side. This
might reflect a process of exfoliation into the tubular lumen (Fig.
1B). No specific staining was observed in the other segments of the
renal tubules, in the glomerulus, in the interstitial kidney tissues, or in the tumorous tissues adjacent to the normal kidney tissue. The
discrepancy between the number of PCR-positive patients (n = 20) and the number of IHC-positive patients (n = 1) may be due to (i) the relative low sensitivity of the IHC
method compared with that of PCR, (ii) viral latency (the presence of
viral DNA without VP1 antigen expression), and (iii) a focal
distribution of cells with viral replication.
IEM revealed that the collecting ducts had positive epithelial cells,
which concurred with our light microscopic findings. The electron-dense
staining was localized in the nuclei of ductal cells. Some of these
positive cells were located in the luminal side and were lying on the
negative cells (Fig. 2A). JCV replication may affect the cellular adhesion to the basement membrane and/or to the
adjacent cells, or JCV replication may be facilitated in the
exfoliating cells. The cells loaded with high viral contents may
exfoliate into the lumen and may serve as a source of the urinary JCV
(7, 12, 24). Although most of the IEM-positive electron-dense material was of a particular or a granular nature, the
virus-like nature of the particles was clear only in some limited
areas. These virus-like particles measured 35 to 45 nm in diameter
(16, 17, 20) and aligned in beads or clusters (Fig. 2B),
which were compatible with polyomavirus particles. Filamentous forms
were not observed. Cytoplasmic virus-like particles were not readily
discernible, although the cytoplasmic structures were poorly preserved.
0095-1137/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
Immunohistochemical Detection of JC Virus in Nontumorous Renal
Tissue of a Patient with Renal Cancer but without Progressive
Multifocal Leukoencephalopathy
ABSTRACT
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FIG. 1.
IHC of the renal medulla. The tissue was counterstained
with hematoxylin. (A) The arrowheads and the arrow indicate positive
collecting ducts. The ducts indicated by arrowheads each contain one
positive cell. Magnification, ×120. (B) High-power view of the
positive duct indicated by an arrow in panel A. Note the two strongly
positive (black in this photo) nuclei and the three weakly positive
nuclei that can be seen to be protruding into the lumen. Magnification,
×480.
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FIG. 2.
IEM of the positive duct. The tissue was counterstained
with uranyl acetate. (A) Low-power view of the positive collecting
duct. Arrow, a positive cell; L, the lumen; *, a negative cell. Note
that the positive cell protrudes into the lumen and overlies the
negative cells. Bar, 4 µm. Magnification, ×2,600. (B) High-power
view of the positive nuclei in panel A. Arrowheads indicate
representative areas containing electron-dense polyomavirus-like
particles. Bar, 200 nm. Magnification, ×50,000.
No inflammatory cell infiltration was observed in the area containing JCV-positive collecting ducts either by IHC or by IEM. Although Dörries and ter Meulen (9) also found no inflammatory cell infiltration in the area with ISH-positive cells in a patient with PML, the implication suggested by our findings is different. In typical patients with PML, the lack of inflammatory reactions in central nervous system lesions has been ascribed to the presence of generalized immunodeficiency, including specific cellular immunity against JCV (1, 11, 23). By the same token, the renal reaction may lack a cellular immune response in patients with PML. On the other hand, our findings suggest a lack of an apparent local cellular immune response against the JCV-infected tubular epithelial cells even in the immunocompetent patient. This kind of virus-host interaction may closely be related to mechanisms of persistent infection of JCV in renal tissue.
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ACKNOWLEDGMENTS |
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We are grateful to Y. Yogo, Institute of Medical Science, the University of Tokyo, for helpful suggestions.
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FOOTNOTES |
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* Corresponding author. Mailing address: Division of Pathology, Department of Toxicology, Tokyo Metropolitan Research Laboratory of Public Health, 3-24-1 Hyakunincho, Shinjukuku, Tokyo 169, Japan. Phone: 81-3-3363-3231, ext. 5700. Fax: 81-3-3368-4060. E-mail: naoto{at}tokyo-eiken.go.jp.
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Journal of Clinical Microbiology, April 1999, p. 1165-1167, Vol. 37, No. 4
0095-1137/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
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