Synthesis and antibacterial activity of four natural chalcones and their derivatives

Article abstract of DOI:10.1016/j.tetlet.2019.151165

Four natural chalcones bearing hydroxyisoprenyl or prenyl groups, named Paratocarpin E (2), Xanthoangelol D (3), Angusticornin A (4) and Kanzonol C (5), were prepared by employing the Claisen-Schmidt condensation as the key step. In an attempt to investigate the effect of the hydroxyisoprenyl group on biological activity, two of their derivatives were also prepared for antibacterial activity research. The synthesized compounds were investigated for their expected antibacterial activities against Gram positive bacteria (Bacillus subtilis, Staphylococcus aureus) as well as Gram negative bacteria (Escherichia coli, Pseudomonas aeruginosa). Paratocarpin E (2) was found to be the most potent against two Gram positive bacteria while the majority of the remaining compounds showed promising activity as well. However, all of the compounds were inactive against both Gram-negative bacteria.

Full text of DOI:10.1016/j.tetlet.2019.151165

Renal Failure
ISSN: 0886-022X (Print) 1525-6049 (Online) Journal homepage: https://www.tandfonline.com/loi/irnf20
The immunophenotyping of different stages of BK
virus allograft nephropathy
Ping Li, Dongrui Cheng, Jiqiu Wen, Xuefeng Ni, Xue Li, Kenan Xie & Jinsong
Chen
To cite this article: Ping Li, Dongrui Cheng, Jiqiu Wen, Xuefeng Ni, Xue Li, Kenan Xie & Jinsong
Chen (2019) The immunophenotyping of different stages of BK virus allograft nephropathy, Renal
Failure, 41:1, 855-861, DOI: 10.1080/0886022X.2019.1617168
To link to this article: https://doi.org/10.1080/0886022X.2019.1617168
© 2019 The Author(s). Published by Informa
UK Limited, trading as Taylor & Francis
Group.
Published online: 19 Sep 2019.
Submit your article to this journal
Article views: 86
View related articles
View Crossmark data
Full Terms & Conditions of access and use can be found at
https://www.tandfonline.com/action/journalInformation?journalCode=irnf20

RENAL FAILURE
2019, VOL. 41, NO. 1, 855–861
https://doi.org/10.1080/0886022X.2019.1617168
CLINICAL STUDY
The immunophenotyping of different stages of BK virus allograft
nephropathy
Ping Li, Dongrui Cheng, Jiqiu Wen, Xuefeng Ni, Xue Li, Kenan Xie and Jinsong Chen
National Clinical Research Center of Kidney Diseases, Jinling Hospital, Medical School of Nanjing University, Nanjing, China
ABSTRACT
ARTICLE HISTORY
Received 16 October 2018
Revised 29 April 2019
Accepted 30 April 2019
Objectives: To investigate the immunohistochemical features of different stages of BK virus
allograft nephropathy (BKVN) and further elucidate the underlying immunological mechanism
involved in the evolution of BKVN.
Methods: Fifty-two renal transplant recipients with biopsy proven BKVN were retrospectively
selected. According to the third edition of the American Society of Transplantation Infection
guidelines, 10 patients were categorized as having mild BKVN (stage A), 25 were moderate (stage
B) and 17 were severe (stage C). The differential infiltrations of CD3þ (T lymphocytes), CD4þ
(helper T lymphocytes), CD8þ (cytotoxic T lymphocytes), CD20þ (B lymphocytes), CD68þ (mac-
rophages) and CD138þ (plasma cells) cells and the expression of interleukin-2 receptor (IL-2R)
and human leukocyte antigen DR (HLA-DR) were compared among the three groups.
Results: CD3þ, CD4þ, CD8þ, CD20þ, CD138þ and CD68þ cells infiltrations, IL-2R and HLA-DR
expression were positive in the BKVN patients. Moreover, with increasing stages of BKVN, the
numbers of positively stained inflammatory cells and the expression of IL-2R were significantly
increased in the severe group compared to the mild group, whereas no statistically significant
differences were observed with regard to HLA-DR expression. Eosinophil and neutrophil infiltra-
tion could also be observed in moderate to advanced BKVN.
KEYWORDS
Renal transplantation; BK
virus allograft nephropathy;
renal pathology;
immunopathology; infection
Conclusion: Renal allograft damage caused by BKVN involved T lymphocyte-, B lymphocyte- and
mononuclear macrophage-mediated immune responses. Inflammatory cell infiltrations in the
renal allograft were probably the driving force for BKVN progression. Additionally, eosinophils
and neutrophils may be involved in the pathophysiological mechanism of BKVN.
Introduction
diagnosis and accurate staging of BKVN are essential to
the timely adjustment of immunosuppressive agents,
which help to delay graft function deterioration and
improve long-term allograft survival.
With the widespread use of potent and newer immuno-
suppressive drugs, the incidence of BK virus (BKV) infec-
tion after renal transplantation is rising [1–3]. BKV
allograft nephropathy (BKVN) has been one of the
major causes of allograft dysfunction and even graft
loss [1,4]. Unfortunately, the precise pathophysiological
mechanism of BKVN remains unclear, and there is still a
lack of specific and effective therapeutic interventions.
The differential diagnosis between BKVN and T cell-
mediated rejection is notoriously difficult, but the asso-
ciated treatments are diametrically opposed. Reducing
immunosuppression is successful in the former, but
potentially deleterious in the latter [5]. Furthermore,
during the treatment of BKVN, secondary acute rejec-
tion frequently ensues after the reduction in immuno-
suppressive agents [6,7]. Therefore, early effective
Kidney allograft biopsy remains the gold standard
for diagnosing BKVN based on molecular, cytological
and histological examinations [8]. Although the charac-
teristics of infiltrating cells in patients with BKVN have
been previously investigated [9–12], the influence
of inflammatory cell infiltration on BKVN progression
remains unknown. Therefore, we aimed to elucidate
the underlying immunological mechanism involved
in the evolution of BKVN by retrospectively reviewing
the immunohistochemical phenotypes of infiltrating
cells and the expression of interleukin-2 receptor (IL-
2R) and human leukocyte antigen (HLA-DR) in patients
with biopsy-proven BKVN.
CONTACT Jinsong Chen
chenjinsong_2017@sina.com
National Clinical Research Center of Kidney Diseases, Jinling Hospital, Medical School of
Nanjing University, 305 East Zhong Shan Road, Nanjing 210002, China
ß 2019 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group.
This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial License (http://creativecommons.org/licenses/by-nc/4.0/), which
permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

856
P. LI ET AL.
inclusions in renal tubular epithelial cells and/or
Bowman’s capsule epithelial cells, accompanied by
tubular epithelial cell necrosis and denudation of base-
ment membranes, as well as tubule interstitial infiltrates
and tubulitis. Positive SV40 staining is used to confirm
the diagnosis of BKVN (Figure 1) [13,14].
Materials and methods
Patients
Fifty-two patients diagnosed with BKVN by renal allo-
graft biopsy between June 2008 and July 2016 at
Jinling Hospital (Nanjing, China), Nanjing University
School of Medicine, were retrospectively identified.
Patients with other concomitant pathological changes,
such as rejection or calcineurin inhibitor toxicity, were
excluded from further analysis. Informed consent was
obtained from all patients, and the Human Subjects
Committee of Jinling Hospital (Nanjing, China),
Nanjing University School of Medicine, approved the
study protocols.
Classification of BKVN
According to the third edition of the American Society of
Transplantation Infection guidelines [13], BKVN patients
were divided into three groups: a mild group of 10
patients (stage A, viral cytopathic changes in almost nor-
mal renal parenchyma), moderate group of 25 patients
(stage B, more severe cellular damage with a combin-
ation of viral cytopathic changes and focal/multifocal
areas of tubular atrophy, and/or interstitial fibrosis),
and severe group of 17 patients (stage C, extensive tubu-
lar atrophy and interstitial fibrosis; Figure 2).
Diagnostic criteria for BKVN
The diagnosis of BKVN is based on detecting signs of
viral cytopathic changes, including intranuclear viral
Figure 1. (A) Light microscopy image of BK virus renal allograft nephropathy. The histological manifestations are characterized
by nuclear inclusion bodies in tubular epithelial cells (arrow indicates the hematoxylin-eosin-stained paraffin section, 400ꢀ). (B)
Immunostaining of BK virus-infected cells with anti-SV40 large T antigen antibodies showing the nuclei of renal tubular epithelial
cells have a transparent center and thorn-shaped periphery (arrow indicates the immunohistochemical staining, 400ꢀ).
Figure 2. Histological patterns of BK virus renal allograft nephropathy (hematoxylin-eosin-stained paraffin section, 400ꢀ). (A)
Stage A: viral cytopathic changes in nearly normal renal parenchyma, (B) Stage B: more severe cellular damage with a combin-
ation of viral cytopathic changes and focal/multifocal areas of tubular atrophy and/or interstitial fibrosis, and (C) Stage C: exten-
sive tubular atrophy and interstitial fibrosis.

RENAL FAILURE
857
Renal biopsies
Biopsies were performed upon protocol or clinical
indication under the guidance of ultrasonography.
Haematoxylin-eosin (HE), periodic acid Schiff (PAS),
periodic acid-silver methenamine (PASM) and Masson
trichrome staining were routinely performed. Fresh-
frozen tissue was analyzed by immunofluorescence
microscopy using a conventional panel of antibodies
against Immunoglobulin A (IgA), IgG, IgM, C3, C1q
and C4d. C4d staining was detected by an indirect
immunofluorescence technique with a primary affin-
ity-purified mouse anti-human monoclonal antibody
(Quidel, San Diego, CA) and a fluorescein isothio-
cyanate-labelled affinity-purified secondary rabbit
anti-mouse IgG antibody (Dako, Glostrup, Denmark).
Positive C4d staining was defined as bright linear
staining along the capillary basement membranes
that involved more than ten percent of the sampled
capillaries according to the Banff Meeting guide-
lines [15].
Immunohistochemical staining
Figure 3. Immunohistochemical characterization of the inflam-
matory infiltrates in BK virus renal allograft nephropathy.
Formalin-fixed, paraffin-embedded renal biopsy sec-
tions were deparaffinized in xylene for 20 min and
rehydrated in a graded ethanol series (100–70%) for
8 min. Endogenous peroxidase were deactivated with
3% hydrogen peroxide for 5–510 min at room tem-
perature after deparaffinization. Antigen retrieval was
performed with a sodium citrate buffer (10 mM, pH ¼
6) for 10 min. For the detection of infiltrating cell sub-
populations, CD3 (T lymphocytes), CD4 (cytotoxic T
lymphocytes), CD8 (suppressor T lymphocytes), CD20
(B lymphocytes), CD68 (macrophages) and CD138
(plasma cells) were regularly detected with mouse
monoclonal antibodies as follows: anti-CD3 (CD3-PS1,
Gene Tech, Shanghai, China), anti-CD4 (CD4-1F6, Gene
Tech, Shanghai, China), anti-CD8 (C8/144B, Dako,
Glostrup, Denmark), anti-CD20 (L26, Dako), anti-CD68
(KPI, Dako) and anti-CD138 (MI 15, Dako; Figure 3). In
addition, IL-2R expression was detected by an anti-IL-
2R mouse monoclonal antibody (ACT-1, Dako) and
HLA-DR staining was evaluated using an anti-HLA-
DR monoclonal antibody (TAL.1B5, Dako) by visually
assessing the approximate proportion of positively
stained tubules. Immunohistochemical staining for
BKV antigens was performed using an anti-BKV large
T antigen mouse monoclonal antibody (BK-T (0).1,
Chemicon International, Billerica, MA).
immunohistochemically positive cells, including IL-
2R-expressing cells in the unscarred cortical paren-
chyma was performed by the Aperio scanner and
expressed as the total number of positive cells per
square millimeter, while the expression of tubular HLA-
DR was assessed as the overall percentage of positive
tubular cross-sections in the entire biopsy sample.
Statistical analysis
Statistical analyses were conducted using SPSS (v25.0,
SPSS, Chicago, IL) software. Categorical variables were
expressed as percentages and compared using Fisher’s
exact test with the Bonferroni correction for p values
among the groups. Continuous variables with normal
distribution are presented as the mean standard
deviation and compared using one-way analysis of
variance (ANOVA) followed by post-hoc pairwise com-
parisons using least significant difference (LSD) tests
or a nonparametric method, as appropriate. p < 0.05
was accepted as statistically significant.
Results
Baseline patient characteristics
All immunohistochemical sections were scanned
by using a full-slice digital scanner (Aperio ScanScope
XT Turbo Scanner, Leica, Germany). The counting of
The baseline patient characteristics at biopsy are listed
in Table 1. Among the 52 patients, 37 were male and

858
P. LI ET AL.
Table 1. The clinical characteristic of patients.
p Value
Characteristic
Total
Mild group
Moderate group
Severe group
p12
p13
p23
Number
Male, n (%)
Age (years)
52
10
8 (80%)
33.9 8.9
2.11 0.80
25
17
–
–
NS
NS
–
NS
NS
37 (71.2%)
37.1 9.9
2.77 1.20
17 (68%)
37.3 11.6
2.63 1.08
12 (70.6%)
38.6 7.5
3.37 1.34
NS
NS
NS
Scr at time of biopsy(mg/dL)
BKV viral load before graft biopsy, Log10 (copies/mL)
Urine
0.001
0.042
9.84 1.21
4.28 1.07
21.6 19.7
9.40 1.04
4.20 1.02
19.1 16.8
10.08 1.35
4.48 1.25
19.0 14.2
9.77 1.11
4.07 1.04
26.8 27.2
NS
NS
NS
NS
NS
NS
NS
NS
NS
Plasma
The time of biopsy from renal transplantation (months)
Immunosuppression at time of biopsy
Tac þ MMF þ Pred, n (%)
48 (92.3%)
2(3.8%)
2(3.8%)
9 (90.0%)
0
1 (10.0%)
23 (92.0%)
1 (4.0%)
1 (4.0%)
16 (94.1%)
1 (5.9%)
0
NS
NS
NS
NS
NS
NS
NS
NS
NS
CyA þ MMF þ Pred, n (%)
Others, n (%)
BKV: BK virus; CyA: cyclosporine; MMF: mycophenolate mofetil; NS: not significant statistically; Pred: prednisone; p12: p values for mild and moderate
group; p13: p values for mild and severe group; p23: p values for moderate and severe group; Scr: serum creatinine; Tac: tacrolimus.
Table 2. Inflammatory cellular constituents in different groups of BKVN.
p Value
Total
Mild group
Moderate group
Severe group
(n ¼ 52)
(n ¼ 10)
(n ¼ 25)
(n ¼ 17)
p12
NS
NS
NS
NS
NS
<0.001
NS
NS
p13
0.002
p23
0.023
0.031
0.040
NS
<0.001
0.005
NS
CD3 (cells/mm2)
645.08 320.88
375.00 192.63
289.47 137.00
299.96 189.20
115.88 103.26
760.02 292.25
24 (46.2%)
455.75 265.78
254.17 152.63
226.00 134.83
175.33 110.11
25.92 24.39
449.92 163.10
4 (40%)
600.55 245.37
353.50 146.38
268.45 103.94
301.59 141.94
70.00 45.35
754.36 170.45
11(44%)
816.21 356.11
476.21 216.44
353.89 151.35
376.79 236.45
190.63 150.43
962.42 298.66
9 (52.9%)
CD4 (cells/mm2)
0.001
0.010
0.003
<0.001
<0.001
NS
CD8 (cells/mm2)
CD20 (cells/mm2)
CD138 (cells/mm2)
CD68 (cells/mm2)
T cell-rich aggregates, n (%)
B cell-rich aggregates, n (%)
Macrophage aggregates (CD68), n (%)
21 (40.4%)
15 (28.8%)
3 (30%)
9 (36%)
7 (28%)
9 (52.9%)
8 (47.1%)
NS
0.012
NS
NS
0
NS
BKVN: BK virus allograft nephropathy; NS: not significant statistically; p12: p values for mild and moderate group; p13: p values for mild and severe group;
p23: p values for moderate and severe group.
15 were female. The average age and serum creatinine
(Scr) level of the patients at biopsy was 37.1 9.9 years
and 2.77 1.20 mg/dL, respectively and the time of
BKVN diagnosis was 21.6 19.7 months after transplant-
ation. Noticeably, the differences in the Scr level among
the three groups were statistically significant (p < 0.05).
Additionally, the BKV viral load in the plasma and urine
before graft biopsy of all the patients was positive;
however, there were no significant differences among
the groups. The post-transplant maintenance immuno-
suppressive protocols of the patients mainly consisted
of tacrolimus (Tac) or cyclosporine A (CyA), mycopheno-
late mofetil (MMF) and prednisone (Pred) at the time of
biopsy. All patients were negative for donor-specific
antibodies (DSA) in the blood.
the numbers of CD3þ, CD4þ, CD8þ, CD20þ, CD68þ
and CD138þ cells in the severe group were significantly
higher than those in the mild group (p < 0.05).
Lymphoid and macrophage aggregates as well as gran-
ulomas were observed in some BKVN patients (Figure
4). Additionally, eosinophil and neutrophil infiltrations
could also be found in moderate to severe BKVN
patients (Table 3).
The expression patterns of IL-2R and HLA-DR in
different groups of BKVN patients
As shown in Table 3, the expression of IL-2R in renal
allografts and HLA-DR in renal tubular epithelial cells
also increased with the progression of BKVN. The
expression of IL-2R in the severe group was significantly
higher than that in the mild group (p ¼ 0.015), whereas
there were no significant differences in the expression
of HLA-DR among the three groups (p > 0.05).
Comparison of the inflammatory infiltrates among
the different groups of BKVN patients
The numbers of CD3þ, CD4þ, CD8þ, CD20þ, CD68þ
and CD138þ cells at different stages of BKVN are shown
in Table 2. The expression of these infiltrating cells
increased with the progression of BKVN (Figure 3).
Infiltrating cells were mainly concentrated in atrophic
renal tubules, especially those with viral inclusion
bodies in both the cortex and medulla. Furthermore,
C4d deposition in different groups of
BKVN patients
C4d focal deposition was noted in 2/10 patients (20%)
in the mild group, 1/25 (4%) in the moderate group
and 0/17 (0%) in the severe group.

RENAL FAILURE
859
Figure 4. Inflammatory changes in BK virus renal allograft nephropathy. (A,B) T and B lymphocyte aggregates were highlighted
with staining for CD3 and CD20, respectively (immunohistochemical staining, 400ꢀ) and (C) Syncytial granulomatous inflamma-
tion with clusters of macrophages was noted (hematoxylin-eosin-stained paraffin section, 400ꢀ). Inset (C): clusters of macro-
phages were highlighted with staining for CD68 stain (immunohistochemical staining, 400ꢀ).
Table 3. The other inflammatory reaction and the expression of IL-2R and HLA-DR in different groups of BKVN.
p Value
Total
Mild group
Moderate group
Severe group
(n ¼ 52)
(n ¼ 10)
(n ¼ 25)
(n ¼ 17)
p12
p13
p23
Granulomatous inflammation, n (%)
Eosinophil infiltration, n (%)
Neutrophil clusters, n (%)
IL-2R (cells/mm2)
1 (1.9%)
4 (7.7%)
0
0
1 (5.9%)
1 (5.9%)
–
NS
0.034
NS
NS
NS
NS
NS
0.015
NS
NS
NS
NS
NS
NS
0
0
3 (12%)
10 (40%)
141.18 48.79
5.40 10.60
16 (30.8%)
140.87 88.09
6.85 12.72
6 (35.3%)
171.16 108.69
10.35 16.98
93.01 92.33
4.50 8.32
HLA-DR (%)
BKVN: BK virus allograft nephropathy; IL-2R: interleukin-2 receptors; HLA: human leukocyte antigen; NS: not significant statistically; p12: p values for mild
and moderate group; p13: p values for mild and severe group; p23: p values for moderate and severe group.
CD8þ, CD20þ, CD138þ and CD68þ cells as well as IL-
Discussion
2R and HLA-DR expression increased with the progres-
Since the development and widespread use of reliable
sion of BKVN. Noticeably, the increases in the numbers
and newer immunosuppressive drugs, BKV infection
of CD3þ, CD4þ, CD8þ, CD20þ, CD138þ and CD68þ
has become common after renal transplantation and
cells were significant, indicating that these cells may
BKVN has become an important cause of graft loss
play important roles in BKVN progression.
[2,3,16]. The reported incidence of biopsy proven BKVN
varies from 1 to 10% [1,13] with subsequent graft loss
in 10 to 80% of cases [17]. BKVN is characterized by the
presence of viral cytopathic changes in the epithelium
of the renal tubules and urothelial lining, as well as
tubulointerstitial inflammation and tubular atrophy/
interstitial fibrosis [18]. Infected cells have an enlarged
nucleus with a gelatinous basophilic inclusion resulting
CD3þ T cells are mainly composed of CD4þ and
CD8þ T cells. These cells are the main source of IL-2R
secretion. Studies have shown that CD3þ T cells play
crucial roles in the initiation and progression of BKVN
and that both CD8þ and CD4þ T cells are involved in
the recognition and clearance of BKV [19,23]. Moreover,
it has been confirmed that CD4þ T cells have a specific
polyfunctional antiviral effect on BKV infection [4,14,24].
from the accumulation of newly formed virions.
Immunohistochemistry has been widely adopted for
the diagnosis of viral infections in immunocomprom-
Large T antigen preferentially stimulates CD8þ T cells,
while the virion protein 1 of BKV preferentially stimu-
lates CD4þ T cells [20]. CD4þ T cells can control BKV
ised patients. Previous studies have shown that the
infiltrating cells in BKVN predominantly comprise
mononuclear cells, including lymphocytes and macro-
phages [11,14,19–21]. Nevertheless, the precise patho-
physiological mechanisms of BKVN remain unclear. To
our knowledge, this is the first study to investigate the
immunohistochemical features of different stages of
BKVN based on the American Society of Transplantation
Infection guidelines [13].
infection by secreting interferon-c (IFN-c), tumor necro-
sis factor-alpha and IL-2 [24]. In this study, with BKVN
development, CD4þ and CD8þ T cells numbers in the
kidney allografts increased. Additionally, this study
highlighted that IL-2R expression was also correlated
with BKVN progression, supporting the conclusion that
the T cell-mediated immune response plays a pivotal
role in BKVN.
Recent studies have demonstrated that the per-
centage of B lymphocytes in BKVN increases signifi-
cantly [10,11,21], favoring the involvement of humoral
In agreement with prior studies [11,14,19,21,22], our
study showed that the infiltration of CD3þ, CD4þ,

860
P. LI ET AL.
immunity in the pathogenesis of BKVN, but the pre-
cise immunopathological mechanisms remain unclear.
Part of the virus could increase B lymphocyte counts
by secreting a B cell-activating factor that is mediated
by NF-kB signaling pathways [25–29]. Nevertheless, it
remains unknown whether BKV causes allograft dam-
age by a similar mechanism. We also found a marked
increase in CD20þ and plasma (CD138þ) cell numbers
with concordant BKVN progression, which was consist-
ent with previous investigations [11,21]. These obser-
vations suggested that humoral immunity may be
involved in the immunological reaction against BKV.
Nevertheless, in our study, there was no significant
difference regarding the expression of HLA-DR among
the groups. Furthermore, only three of the 52 patients
had focal positive C4d expression in renal allograft
peritubular capillaries and DSA levels in the blood
were also negative without histological evidence of
antibody-mediated rejection [15]. Therefore, we specu-
lated that there was a difference in activated B
lymphocyte function between BKVN and antibody-
mediated rejection.
There is currently no reported mechanism for the
significant increase in CD68þ cell infiltration in BKVN.
We assumed that BKV can activate macrophages in a
manner similar to other viruses via the CCL2/CCR2 path-
way that mediates monocyte migration to an inflamed
site [30,31]. Additionally, the encroachment of BKV dir-
ectly upon renal tubular epithelial cells may be another
possible explanation. Furthermore, Braga et al. [32]
found that the factors released after BKV infection could
activate macrophages through Toll-like receptors and
the MyD88 signaling pathway to promote renal allo-
graft fibrosis.
molecules, whereas CD4þ T cells scan viral epitopes
presented by MHC class II molecules on antigen-pre-
senting cells such as macrophages, B lymphocytes and
dendritic cells. On the other hand, activated T lympho-
cytes regulate the proliferation, differentiation and
immune function of B lymphocytes and macrophages
by secreting a variety of cytokines to control viral infec-
tion, such as IL-2 and IFN-c. Therefore, T lymphocyte-, B
lymphocyte- and mononuclear macrophage-mediated
immune responses are involved during BKV infection in
renal allografts and these immune components inter-
act intimately.
The main limitation of this study is that renal allo-
graft biopsies were performed for-cause instead of per-
protocol. Serial biopsies from patients with BKVN help
to define the histological evolution of BKVN and pro-
vide insight into the underlying mechanism.
In conclusion, we observed renal allograft damage
caused by BKVN involving T lymphocyte-, B lympho-
cyte- and mononuclear macrophage-mediated immune
responses. Inflammatory cell infiltration into the renal
allografts was probably the driving force for BKVN
progression. Eosinophil and neutrophil involvement in
the pathophysiological mechanisms of BKVN requires
further study.
Disclosure statement
No potential conflicts of interest were reported by
the authors.
Funding
This study was funded by the National Natural Science
Foundation and National Key Research and Development
Program of China.
We observed eosinophil and neutrophil infiltration
in moderate to advanced BKVN, consistent with
previous research. Eosinophils and neutrophils are
multifunctional pro-inflammatory leukocytes that are
involved in the pathogenesis of immune disorders
through releasing cytokines, chemokines and toxic
cytoplasmic granule constituents [33,34]. Some stud-
ies have indicated that interstitial eosinophil and
neutrophil infiltration in renal allografts is signifi-
cantly associated with acute renal allograft rejection
[35–37]. Nevertheless, the role of eosinophil and neu-
trophil infiltration in the pathophysiology of BKVN
is unclear.
References
[1] Ramos E, Drachenberg CB, Wali R, et al. The decade of
polyomavirus BK-associated nephropathy: state of
affairs. Transplantation. 2009;87:621–630.
[2] Pham PT, Schaenman J, Pham PC. BK virus infection
following kidney transplantation: an overview of risk
factors, screening strategies, and therapeutic interven-
tions. Curr Opin Organ Transplant. 2014;19:401–412.
[3] Manitpisitkul W, Drachenberg C, Ramos E, et al.
Maintenance immunosuppressive agents as risk fac-
tors for BK virus nephropathy: a case-control study.
Transplantation. 2009;88:83–88.
[4] Lamarche C, Orio J, Collette S, et al. BK polyomavirus
and the transplanted kidney: immunopathology and
therapeutic approaches. Transplantation. 2016;100:
2276–2287.
Virus-specific T cells can recognize cells presenting
viral epitopes on the cell surface via major histocom-
patibility complex (MHC) molecules [38]. Classically,
CD8þ T cells recognize and destroy infected cells dis-
playing viral peptides presented by MHC class I
[5] Saad ER, Bresnahan BA, Cohen EP, et al. Successful
treatment of BK viremia using reduction in

RENAL FAILURE
861
immunosuppression
Transplantation. 2008;85:850–854.
without
antiviral
therapy.
[21] Li X, Sun Q, Chen J, et al. Immunophenotyping in BK
virus allograft nephropathy distinct from acute rejec-
tion. Clin Dev Immunol. 2013;2013:1.
[22] Mannon RB, Hoffmann SC, Kampen RL, et al.
Molecular evaluation of BK polyomavirus nephrop-
athy. Am J Transplant. 2005;5:2883–2893.
[23] Comoli P, Hirsch HH, Ginevri F. Cellular immune
responses to BK virus. Curr Opin Organ Transplant.
2008;13:569–574.
[24] Schmidt T, Adam C, Hirsch HH, et al. BK polyomavirus-
specific cellular immune responses are age-dependent
and strongly correlate with phases of virus replication.
Am J Transplant. 2014;14:1334–1345.
[25] Rodriguez B, Valdez H, Freimuth W, et al. Plasma lev-
els of B-lymphocyte stimulator increase with HIV dis-
ease progression. AIDS. 2003;17:1983–1985.
[26] Toubi E, Gordon S, Kessel A, et al. Elevated serum B-
Lymphocyte activating factor (BAFF) in chronic hepa-
titis C virus infection: association with autoimmunity.
J Autoimmun. 2006;27:134–139.
[27] Dalrymple NA, Mackow ER. Endothelial cells elicit
immune-enhancing responses to dengue virus infec-
tion. J Virol. 2012;86:6408–6415.
[28] Vincent FB, Saulep-Easton D, Figgett WA, et al. The
BAFF/APRIL system: emerging functions beyond B cell
biology and autoimmunity. Cytokine Growth Factor
Rev. 2013;24:203–215.
[29] Lu B, Zhang B, Wang L, et al. Hepatitis B virus e anti-
gen regulates monocyte function and promotes B
lymphocyte activation. Viral Immunol. 2017;30:35–44.
[30] Bose S, Cho J. Role of chemokine CCL2 and its recep-
tor CCR2 in neurodegenerative diseases. Arch Pharm
Res. 2013;36:1039–1050.
[31] Covino DA, Sabbatucci M, Fantuzzi L. The CCL2/CCR2
axis in the pathogenesis of HIV-1 infection: a new
cellular target for therapy. Curr Drug Targets. 2016;
17:76–110.
[32] Braga TT, Correa-Costa M, Guise YF, et al. MyD88 sig-
naling pathway is involved in renal fibrosis by favoring
a TH2 immune response and activating alternative M2
macrophages. Mol Med. 2012;18:1231–1239.
[33] Gleich GJ, Adolphson CR. The eosinophilic leukocyte:
structure and function. Adv Immunol. 1986;39:177–253.
[34] Naumenko V, Turk M, Jenne CN, et al. Neutrophils
in viral infection. Cell Tissue Res. 2018;371:505–516.
[35] Weir MR, Hall-Craggs M, Shen SY, et al. The prognostic
value of the eosinophil in acute renal allograft rejec-
tion. Transplantation. 1986;41:709–712.
[36] Saisu K, Morozumi K, Suzuki K, et al. Significance of
interstitial lesions as the early indicator for acute
vascular rejection in human renal allografts. Clin
Transplant. 1999;13:17–23.
[37] Wen T, Rothenberg ME. The regulatory function of
eosinophils. Microbiol Spectr. 2016;4:1–12.
[38] Sester M, Leboeuf C, Schmidt T, et al. The “ABC” of
virus-specific T cell immunity in solid organ trans-
plantation. Am J Transplant. 2016;16:1697–1706.
[6] Drachenberg CB, Papadimitriou JC, Chaudhry MR,
et al. Histological evolution of BK virus-associated
nephropathy: importance of integrating clinical
and pathological findings. Am J Transplant. 2017;17:
2078–2091.
[7] Nankivell BJ, Renthawa J, Sharma RN, et al. BK virus
nephropathy: histological evolution by sequential
pathology. Am J Transplant. 2017;17:2065–2077.
[8] Hirsch HH, Knowles W, Dickenmann M, et al.
Prospective study of polyomavirus type BK replication
and nephropathy in renal-transplant recipients. N Engl
J Med. 2002;347:488–496.
[9] Drachenberg CB, Papadimitriou JC, Ramos E.
Histologic versus molecular diagnosis of BK polyoma-
virus-associated nephropathy:
CJASN. 2006;1:374–379.
[10] Drachenberg CB, Papadimitriou JC, Hirsch HH, et al.
Histological patterns of polyomavirus nephropathy:
correlation with graft outcome and viral load. Am J
Transplant. 2004;4:2082–2092.
a shifting paradigm.
€
[11] Buettner M, Xu H, Bohme R, et al. Predominance of
TH2 cells and plasma cells in polyoma virus nephrop-
athy: a role for humoral immunity. Hum Pathol. 2012;
43:1453–1462.
[12] Sawinski D, Goral S. BK virus infection: an update on
diagnosis and treatment. Nephrol Dial Transplant.
2015;30:209–217.
[13] Hirsch HH, Randhawa P. BK polyomavirus in solid
organ transplantation. Am J Transplant. 2013;13(Suppl
4):179–188.
[14] Dekeyser M, Franc¸ois H, Beaudreuil S, et al.
Polyomavirus-specific cellular immunity: from BK-virus-
specific cellular immunity to BK-virus-associated
nephropathy. Front Immunol. 2015;6:307.
[15] Haas M, Loupy A, Lefaucheur C, et al. The Banff 2017
kidney meeting report: revised diagnostic criteria for
chronic active T cell-mediated rejection, antibody-
mediated rejection, and prospects for integrative
endpoints for next-generation clinical trials. Am J
Transplant. 2018;18:293–307.
[16] El AM, Abd ES, Saadi G, et al. Prevalence of polyoma
BK virus infection among living-donor renal transplant
recipients. Transpl Infect Dis. 2016;18:529–537.
[17] Costa C, Cavallo R. Polyomavirus-associated nephrop-
athy. World J Transplant. 2012;2:84–94.
[18] Drachenberg CB, Papadimitriou JC. Polyomavirus-asso-
ciated nephropathy: update in diagnosis. Transplant
Infect Dis. 2006;8:68–75.
[19] Comoli P, Cioni M, Basso S, et al. Immunity to polyo-
mavirus BK infection: immune monitoring to regulate
the balance between risk of BKV nephropathy and
induction of alloimmunity. Clin Dev Immunol. 2013;
2013:256923.
[20] Binggeli S, Egli A, Schaub S, et al. Polyomavirus BK-
specific cellular immune response to VP1 and large
T-antigen in kidney transplant recipients. Am
Transplant. 2007;7:1131–1139.
J