- Email: [email protected]
Urinary albumin excretion and chronic kidney disease in children with vesicoureteral reflux
Accepted Manuscript Urinary albumin excretion and chronic kidney disease in children with vesicoureteral reflux Romaine de Sépibus, François Cachat, Blaise J. Meyrat, Gezim Dushi, Ariane Boubaker, Mohamed Faouzi, Eric Girardin, Hassib Chehade PII:
S1477-5131(17)30178-X
DOI:
10.1016/j.jpurol.2017.04.004
Reference:
JPUROL 2527
To appear in:
Journal of Pediatric Urology
Received Date: 22 November 2016 Accepted Date: 6 April 2017
Please cite this article as: de Sépibus R, Cachat F, Meyrat BJ, Dushi G, Boubaker A, Faouzi M, Girardin E, Chehade H, Urinary albumin excretion and chronic kidney disease in children with vesicoureteral reflux, Journal of Pediatric Urology (2017), doi: 10.1016/j.jpurol.2017.04.004. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
ACCEPTED MANUSCRIPT Urinary albumin excretion and chronic kidney disease in children with vesicoureteral reflux Romaine de Sépibus1,MD, François Cachat1,MD, Blaise J. Meyrat2,MD, Gezim Dushi2,MD, Ariane Boubaker3,MD, Mohamed Faouzi4,MD, Eric Girardin1,MD, Hassib Chehade1,MD
RI PT
Affiliations: 1Peadiatric Nephrology Unit, University Hospital of Lausanne (CHUV),
Lausanne, Switzerland, 2Department of Peadiatric surgery, University Hospital of Lausanne (CHUV), Lausanne, Switzerland, 3Institute of Radiology, Clinique de La Source, Lausanne,
Corresponding author's address:
M AN U
Hospital of Lausanne (CHUV), Lausanne, Switzerland.
SC
Switzerland, 4Institute of Social and Preventive Medicine, Biostatistics Unit, University
Hassib Chehade, MD, Peadiatric Nephrology Unit, Service of Peadiatrics, University Hospital of
Lausanne
(CHUV),
Rue
Bugnon
46,
1011
Lausanne,
Switzerland,
TE D
[email protected], Tel: +41 21 314 18 11; Fax: +41 21 314 36 26.
EP
Short title: Albuminuria in children with vesicoureteral reflux. Abstract word count: 300
AC C
Text word count: 2534
E-mail:
ACCEPTED MANUSCRIPT 1
Urinary albumin excretion and chronic kidney disease in children with vesicoureteral reflux
2 3
Romaine de Sépibusa, François Cachata, Blaise J. Meyratb, Gezim Dushib, Ariane Boubakerc,
4
Mohamed Faouzid, Eric Girardina, and Hassib Chehadea,*
5 6
a
7
Switzerland
8
b
9
Switzerland
RI PT
Paediatric Nephrology Unit, University Hospital of Lausanne (CHUV), Lausanne,
Department of Paediatric surgery, University Hospital of Lausanne (CHUV), Lausanne,
10
c
11
d
12
Lausanne (CHUV), Lausanne, Switzerland
SC
Institute of Radiology, Clinique de La Source, Lausanne, Switzerland
Institute of Social and Preventive Medicine, Biostatistics Unit, University Hospital of
M AN U
13 14
* Corresponding author. Paediatric Nephrology Unit, Service of Paediatrics, University
15
Hospital of Lausanne (CHUV), Rue Bugnon 46, 1011 Lausanne, Switzerland.
16
E-mail address: [email protected]
17
Summary Background: Albuminuria is a potential biomarker of chronic kidney disease
19
(CKD) in various glomerular diseases. Vesicoureteral reflux (VUR) often progresses to CKD,
20
and study is required of use of albuminuria as a biomarker for this condition. The aim of this
21
study was to evaluate the association between albuminuria and glomerular filtration rate
22
(GFR) or filtration fraction (FF) in children with VUR.
23
Study design: In this retrospective study, renal parameters of 141 children with VUR were
24
investigated, using inulin clearance, FF, and albuminuria. The association between urinary
25
albumin to creatinine ratio (ACR), GFR, and FF was analyzed in a continuous manner by
26
calculating the β coefficient, and also in a binary manner by calculating the OR.
27
Results: Using both continuous and binary analyses, ACR values were negatively and
28
significantly associated to GFR values in patients with low, normal, or high FF values (Table).
29
It was also positively and significantly associated with FF values in patients with low, normal
30
or high GFR values (Table). No association was found between ACR and gender, VUR stages
31
or laterality, number of urinary tract infection, presence of a single functional kidney, history
32
of reflux surgery, or renal scars or hypertension.
33
Discussion: ACR is associated with CKD in patients with VUR. In addition, increased urinary
34
albumin excretion cannot be completely and solely explained by decreased GFR and/or
AC C
EP
TE D
18
1
ACCEPTED MANUSCRIPT 35
increased FF values. The two main limitations of this study are the crude assessment of renal
36
scarring, which prevented finer analysis between albuminuria and renal scarring surface area,
37
and that the study cohort may not be representative of the true VUR population.
38
Conclusion: This study shows that albuminuria is associated with decreased renal function in
39
patients with VUR and that it could be used to monitor renal function in this condition.
RI PT
40 Table. Association results between ACR values, GFR values, and FF values using continuous
42
and binary methods
43
Association β coefficient = – p = 0.01 OR 0.94 p ≤ 0.001 between ACR 0.02 and GFR values Association β coefficient = p = 0.01 OR 1.11 p = 0.036 between ACR 0.07 and FF values ACR, albumin to creatinine ratio; FF, filtration fraction; GFR, glomerular filtration rate.
M AN U
SC
41
44 KEYWORDS
46
Vesicoureteral reflux;
47
Chronic kidney disease;
48
Albuminuria;
49
Child
50
TE D
45
Introduction
52
Vesicoureteral reflux (VUR) is a common urological abnormality in children with an
53
estimated prevalence of 1-6% [1,2]. Up to 30% of children with febrile urinary tract infection
54
(UTI) exhibit VUR and 15-30% will develop permanent renal scarring [3-6]. A substantial
55
number of children with reflux nephropathy (RN) will develop chronic renal failure, arterial
56
hypertension (HTN), and eventually end-stage renal disease [7]. According to the North
57
American Pediatric Renal Trials and Collaborative Studies, RN is the fourth leading cause of
58
chronic kidney disease (CKD), dialysis, and paediatric renal transplantation [8]. For this
59
reason, renal function of children with VUR should be monitored periodically, to identify the
60
group of patients most at risk of developing RN [9,10].
AC C
EP
51
61
It has been demonstrated that albuminuria is not only associated with decreased
62
glomerular filtration rate (GFR) but could also and by itself contribute to renal damage by
63
inducing tubular apoptosis and promoting interstitial fibrosis and inflammation [11,12].
64
Several studies have shown a close association between albuminuria and kidney diseases in 2
ACCEPTED MANUSCRIPT paediatric patients. Karlen et al. [13] found increased urine albumin excretion (>20
66
micrograms/min per 100 mL GFR) in more than 50% of children with pyelonephritic scarring.
67
Cachat et al. [14] showed that albuminuria predicted, to some extent, filtration fraction (FF)
68
in children with a single functioning kidney. However, studies analyzing the association
69
between albuminuria and GFR in children with VUR are scarce. Basic et al. [15] showed
70
significantly higher albuminuria and lower GFR in children with high grade VUR (IV and V)
71
compared with children with lower grade VUR (I, II, III). Similarly, Bell et al. [16] studied a
72
group of 36 children and found that albuminuria was significantly higher in high grade VUR
73
when compared with low and moderate grade VUR, but they did not evaluate GFR. To the
74
best of our knowledge it has not been clearly established whether albuminuria is a reliable
75
biomarker of kidney dysfunction in patients with VUR. This study aims to evaluate the
76
association between albuminuria and GFR using inulin clearance (Clin), and also between
77
albuminuria and FF in children with VUR.
78
Patients and methods
79
The study was conducted in the laboratory of the Paediatric Nephrology unit of Lausanne
80
University Hospital. It was approved by the local research ethics committee (Lausanne Ethic
81
Committee Protocol number: 215/13).
M AN U
SC
RI PT
65
One-hundred and forty-six paediatric patients with VUR were recruited. Patients who
83
were under treatment with angiotensin-converting enzyme inhibitors or angiotensin receptor
84
blockers were excluded (n=5). All included patients had a VUR confirmed by VCUG, and
85
GFR and FF measured using Clin and para-aminohippurate clearances (ClPAH), respectively,
86
together with a single morning urinary albumin excretion measurement. Patients were all
87
referred from treating physicians, nephrologists, or urologists. Indications for GFR evaluation
88
were left at the discretion of the treating physicians.
EP
VUR was classified according to the international reflux classification [17]. Bilateral
AC C
89
TE D
82
90
VUR was defined as bilateral high-grade VUR when grade III or IV or V VUR were found on
91
both sides. The highest grade of VUR was then recorded for statistical analysis. All other
92
conditions with grade I or II VUR on one side and grade III or IV or V VUR on the other side
93
were included in the group with unilateral VUR. History of VUR surgery was recorded. The
94
indication for surgery was left at the discretion of the treating physicians. The number of
95
symptomatic urinary tract infections (UTI) was recorded based on medical chart review, and
96
children were classified into three groups according to the number of UTIs (0; 1 or 2; >2
97
UTIs). The presence or absence of renal scarring was evaluated by two nuclear medicine
98
physicians using iodine-123 hippuran dynamic renal scintigraphy done at least 6 months after 3
ACCEPTED MANUSCRIPT 99 100
the last documented UTI. Renal scarring was defined as cortical thinning, flattening, or wedge-shaped defect. Urinary albumin excretion, GFR, and FF were measured simultaneously. All patients
102
fasted overnight before the procedure. Blood pressure was measured and hypertension (HTN)
103
was defined according to the National High Blood Pressure Education Program Working
104
Group on Children and Adolescents [18].
105
Urinary albumin excretion was measured by the immunoturbidimetric method, using a second
106
urine specimen in the morning, and was expressed as the urinary albumin to creatinine ratio
107
(ACR).
RI PT
101
Clin was used to measure GFR. Intravenous catheters were inserted into both of a
109
patient’s arms and an intravenous loading dose of 30 mg/kg of inulin (Inutest 25%, Fresenius,
110
Austria) was administered. A continuous infusion of inulin was applied for 3 hours to obtain a
111
constant plasmatic concentration between 200 and 500 mg/L. Diuresis was induced by oral
112
administration of 20 mL/kg of water (maximum 1,200 mL) during the first hour, followed by
113
maintenance hydration at a rate of 3 mL/kg/h. Concomitant intravenous hydration (NaCl
114
0.9%, maximum 300 mL) was given every 30 minutes. Urine and blood were collected every
115
30 minutes beginning after 90 minutes. Levels of inulin were measured, using the automated
116
anthrone method according to Wright and Gann. Intra-assay variability coefficients in serum
117
were 2.44% at 10 mg/100 mL, 1.47% at 30 mg/100 mL, and 0.94% at 40 mg/100 mL. Intra-
118
assay variability coefficients in urine were 1.71% at 10 mg/100 mL, 1.22% at 30 mg/100 mL,
119
and 1.07% at 50 mg/100 mL. GFR was calculated as the average of the three clearance
120
periods [19].
M AN U
TE D
EP
121
SC
108
Renal plasmatic flow (RPF) was measured using the ClPAH (Bratton-Marshall reaction). FF was defined as the GFR to RPF ratio (Clin/ClPAH).
123
Statistical analysis
124
All statistical analyses were performed by the Institute of Social and Preventive Medicine at
125
Lausanne University Hospital, using Stata statistical software (StataCorp. 2011. Stata
126
Statistical Software: Release 12. College Station, TX: StataCorp LP). The variables were
127
summarized by their median and interquartile range (IQR) and by numbers and percentages
128
for categorical variables.
AC C
122
129
Log-transformed ACR was used for expression of urinary albumin excretion. To avoid
130
any bias in the definition of pathological ACR, we studied this outcome as a continuous
131
variable and as a binary variable (coded 0/1). For the binary outcome, the patients were
132
divided into two groups, according to the absence or presence of pathological ACR (<2.5 4
ACCEPTED MANUSCRIPT 133
mg/mmol coded by 1 and ≥2.5 mg/mmol coded by 0) [20]. For the continuous outcome, the associations between log-transformed ACR and
135
several variables (GFR, FF, gender, VUR grade and laterality, age at diagnosis, interval
136
between diagnosis of VUR and surgery, history of urological surgery, single functioning
137
kidney, history of UTIs, presence of unilateral or bilateral renal scarring, and presence of
138
HTN) were examined using a univariable linear regression model. The degree of mutual
139
association was determined by the β coefficient (β) and the calculated p value. The binary
140
outcome was analyzed using a univariable logistic regression model to identify the associated
141
factors with the absence or presence of pathological ACR and OR and 95% CI were
142
calculated to set apart the differences between the two groups. A p-value <0.05 was
143
considered to be significant.
SC
RI PT
134
For both outcomes (continue and binary) associated factors to the outcome (p-value
145
<20%) were used in a backward-selection procedure to fit a multivariable model. Models
146
diagnostic (residual, influence statistics) were performed to examine how well the model fits
147
the data.
148
Results
149
Patient characteristics
150
One-hundred and forty-one children (68 females and 73 males) were included in this
151
retrospective study. Details of the patients’ characteristics are summarized in Table 1. Median
152
(IQR) follow-up was 10.21 (6.37) years. Median interval between GFR measurement and
153
renal scintigraphy was 0.24 months (IQR 18.48 months). Median (IQR) age at urinary
154
albumin excretion measurement, GFR, and FF measurements was 12.2 (7.54) years.
EP
TE D
M AN U
144
Median (IQR) GFR was 92.0 (19.50) mL/min per 1.73m2. Median (IQR) GFR in the
156
group with normal log-transformed ACR was 94.0 (18.0) mL/min per 1.73m2 and median
157
(IQR) GFR in the group with abnormal log-transformed ACR was 82.0 (30.0) mL/min per
158
1.73m2. Median (IQR) FF in all patients was 20.0% (14.0-34.0%). Median (IQR) FF in the
159
group with normal log-transformed ACR was 20.0% (4.0%), and median (IQR) FF in the
160
group with abnormal log-transformed ACR was 21% (5.0%).
161
ACR analysis as a continuous variable
162
In a linear regression model, using a univariable analysis, we showed a significant negative
163
association between log-transformed ACR and GFR (β = –0.02; p = 0.010) and a significant
164
positive association between log-transformed ACR and FF (β = 0.07; p = 0.010). This has
165
been confirmed in a multivariable analysis (Table 2).
166
AC C
155
A linear regression model analyzing the association between the log-transformed ACR 5
ACCEPTED MANUSCRIPT and other factors, showed no significant association between the log-transformed ACR and
168
different stages of VUR (ref. = VUR II) (VUR II versus VUR III (β = 0.07; p = 0.866); VUR
169
II versus VUR IV (β = 0.25; p = 0.504)) or VUR laterality (reference (ref.) = unilateral; β =
170
0.44; p = 0.067). There was no association between log-transformed ACR and all other
171
independent factors (gender, presence of a functional single kidney, history of reflux surgery,
172
number of UTI, renal scars, and hypertension). All results are summarized in Table 2.
RI PT
167
Constructing the ROC curve (Fig. 1), we found that an ACR value of 5.09 mg/mmol
174
had the best sensitivity and specificity of 69% and 91%, respectively, to detect inulin
175
clearance below 70 mL/mn x1.73m2.
176
ACR analysis as a binary variable
177
Thirty-one of the 141 patients (22%) exhibited pathological ACR (i.e. ACR ≥2.5 mg/mmol).
178
We found a significant association between the presence of abnormal log-transformed ACR
179
and decreased GFR (OR 0.94; 95% CI 0.91–0.97; p ≤ 0.001) and between abnormal log-
180
transformed ACR and increased FF (OR 1.11; 95% CI 1.01–2.23; p = 0.036). In multivariate
181
analysis, decreased GFR and increased FF were significantly and independently associated
182
with pathological log-transformed ACR with OR of 0.94 (95% CI 0.91–0.97; p ≤ 0.001) and
183
1.12 (95 % CI 1.01–1.25; p = 0.036), respectively.
M AN U
SC
173
There is no association between abnormal log-transformed ACR and VUR stages
185
(VUR II (ref.) versus VUR III (OR 1.54; 95% CI 0.29–8.07; p = 0.610), VUR II versus VUR
186
IV (OR 2.20; 95% CI 0.46–10. 62, p = 0.325)), and uni- or bilaterality of VUR (unilateral
187
VUR = ref.; OR 2.21; 95% CI 0.95–5.13; p = 0.064). Logistic regression model analysis also
188
showed no association between abnormal log-transformed ACR and other independent factors
189
(i.e. gender, the presence of a functional single kidney, previous reflux surgery, number of
190
UTI, renal scars, or hypertension). Distribution of patients according to ACR as a binary
191
variable is summarized in Table 3. The fitted multivariable logistic model was well calibrated
192
as shown by the Hosmer-Lemeshow goodness of fit test (chi-square (8) = 4.20, p = 0.83).
193
Discussion
194
We found in a paediatric population with VUR followed over a median period of 10.2 years a
195
significant negative association between ACR and GFR levels, independently of FF values.
196
This could indicate that factors other than haemodynamic disturbances play a significant role
197
in generation of increased albuminuria in CKD progression. Indeed, chronic interstitial
198
inflammation, fibrosis, tubular dilatation, and atrophy have been described in RN [21]. These
199
reported alterations may partially explain the pathophysiological mechanism of the negative
200
association between albuminuria and GFR independently of hemodynamic factors (e.g. FF),
AC C
EP
TE D
184
6
ACCEPTED MANUSCRIPT through impaired proximal tubular albumin reabsorption [22]. The ACR measurement in
202
patients with CKD has been evaluated in several clinical studies. It has been shown that
203
albuminuria can predict CKD and/or cardiovascular events in conditions such as diabetes
204
mellitus or IgA nephropathy [23,24]. Recently, the KDIGO guidelines added to their CKD
205
classification ACR as a marker for all-cause mortality in patients with CKD [25].
206
Furthermore, patients with proteinuric nephropathies greatly benefit from renin-angiotensin-
207
aldosterone system blockade [26]. This study allowed definition of an ACR threshold of 5.09
208
mg/mmol to best detect inulin clearances below 70 mL/mn x 1.73 m2 in children with
209
vesicoureteral reflux. The validity of this new ACR cutoff requires assessment in prospective
210
randomized controlled trials, and also the benefits of nephroprotection targeting renin-
211
angiotensin should be assessed according to ACR thresholds. Given the paucity of studies
212
analyzing the association between albuminuria and GFR in patients with VUR, we did not
213
calculate the sample size. However, in a post-hoc analysis, the study has a power of 98.52%
214
to detect a significant association between ACR and GFR.
M AN U
SC
RI PT
201
We also found a significant positive association between ACR and FF levels,
216
independently of GFR values. These results, using both continuous and binary analyses,
217
emphasize the role of renal perfusion in association with interstitial changes in this complex
218
process of increased albuminuria, CKD progression, and hyperfiltration. Finally, and in
219
contrast to the findings of Basic et al. [15] and Bell et al. [16], we did not find any association
220
between ACR and VUR stages. Furthermore, we did not find any association between ACR
221
and gender, VUR laterality, the number of UTI, the presence of a single functioning kidney,
222
the history of reflux surgery, and the presence of renal scars or HTN.
EP
TE D
215
All these observations emphasize the potential role of urinary albumin excretion in
224
monitoring of renal function (decreased GFR and/or hyperfiltration) in patients with VUR,
225
and suggest that increased urinary albumin excretion cannot be completely and solely
226
explained by decreased GFR and/or increased FF values.
227
AC C
223
As previously reported by Lahdes-Vasama et al. [27], it is also interesting to note that,
228
in both the linear and the logistic regression analysis, there was a tendency towards an
229
association between ACR and bilateral VUR or more than two UTIs.
230
The strength of this study was the long term follow-up of children over 10 years, the
231
use of inulin for GFR evaluation, and the analysis of the association among ACR, GFR, and
232
FF in both continuous and binary manners.
233
This study has several limitations. Although albumin excretion is often evaluated in
234
the literature with 24-hour urine collection or repeated samples collections, we used a single 7
ACCEPTED MANUSCRIPT random urine sample. However, the second morning urine ACR sample has been shown in
236
other studies to have a good association with the 24-hour urine albumin excretion rate [28]. In
237
addition, a small number of patients might still have a persistent VUR at the time of renal
238
function evaluation; however, we did not find any association between ACR and a history of
239
reflux surgery. Furthermore, the crude assessment of renal scarring prevented us from finer
240
analysis between albuminuria and renal scarring surface area. Finally, the study sample may
241
not be representative of the VUR population, as there were no patients with grade V VUR in
242
our cohort. In addition, all patients referred to our laboratory unit for inulin clearance were
243
included; however, we cannot exclude an inclusion bias because some patients with VUR
244
were not referred by their treating physicians for GFR evaluation. Referral was not based on a
245
particular ACR threshold.
SC
RI PT
235
In conclusion, this study showed an association between ACR and CKD. It also
247
defined a new ACR cutoff of 5.09 mg/mmol to accurately detect a GFR < 70 mL/mn x 1.73
248
m2 in patients with VUR.
249
Conflict of interest
250
None.
251
Funding
252
None.
253
References
254
[1] Toka HR, Toka O, Hariri A, Nguyen HT. Congenital anomalies of kidney and urinary
255
tract. Semin Nephrol 2010; 30:374–86.
256
[2] Guignard JP, Maherzi M, Torrado A. Screening for reflux. Lancet 1978; 2:316.
257
[3] Sargent MA. What is the normal prevalence of vesicoureteral reflux? Pediatr Radiol 2000;
258
30:587–93.
259
[4] Wadie GM, Moriarty KP. The impact of vesicoureteral reflux treatment on the incidence
260
of urinary tract infection. Pediatr Nephrol 2012; 27:529–38.
261
[5] Ismaili K, Wissing KM, Lolin K, Le PQ, Christophe C, Lepage P, Hall M. Characteristics
262
of first urinary tract infection with fever in children: a prospective clinical and imaging study.
263
Pediatr Infect Dis J 2011; 30:371–4.
264
[6] Coulthard MG, Lambert HJ, Vernon SJ, Hunter EW, Keir MJ, Matthews JN. Does prompt
265
treatment of urinary tract infection in preschool children prevent renal scarring: mixed
266
retrospective and prospective audits. Arch Dis Child 2014; 99:342–7.
267
[7] Ardissino G, Avolio L, Dacco V, Testa S, Marra G, Viganò S, et al. Long-term outcome
268
of vesicoureteral reflux associated chronic renal failure in children. Data from the ItalKid
AC C
EP
TE D
M AN U
246
8
ACCEPTED MANUSCRIPT Project. J Urol. 2004; 172: 305–10.
270
[8] Chesney RW, Brewer E, Moxey-Mims M, Watkins S, Furth SL, Harmon WE, et al.
271
Report of an NIH task force on research priorities in chronic kidney disease in children.
272
Pediatr Nephrol 2006; 21:14–25.
273
[9] Jodal U, Smellie JM, Lax H, Hoyer PF. Ten-year results of randomized treatment of
274
children with severe vesicoureteral reflux. Final report of the International Reflux Study in
275
Children. Pediatr Nephrol. 2006; 21:785–92.
276
[10] Sanna-Cherchi S, Ravani P, Corbani V, Parodi S, Haupt R, Piaggio G, et al. Renal
277
outcome in patients with congenital anomalies of the kidney and urinary tract. Kidney Int.
278
2009;76:528–33.
279
[11] Turin TC, Ahmed SB, Tonelli M, Manns B, Ravani P, James M, et al. Kidney function,
280
albuminuria and life expectancy. Can J Kidney Health Dis 2014; 1:33.
281
[12] Thomas ME, Brunskill NJ, Harris KP, Bailey E, Pringle JH, Furness PN, Walls J.
282
Proteinuria induces cell turnover: a potential mechanism for tubular atrophy. Kidney Int 1999;
283
55:890–898.
284
[13] Karlen J, Linné T, Wikstad I, Aperia A. Incidence of microalbuminuria in children with
285
pyelonephritis scarring. Pediatr Nephrol 1996; 10:705–8.
286
[14] Cachat F, Combescure C, Chehade H, Zeier G, Mosig D, Meyrat B, et al.
287
Microalbuminuria and hyperfiltration in subjects with nephro-urological disorders. Nephrol
288
Dial Transplant 2013; 28:386–91.
289
[15] Basic J, Golubovic E, Miljkovic P, Bjelakovic G, Cvetkovic T, Milosevic V.
290
Microalbuminuria in children with vesicoureteral reflux. Ren Fail 2008; 30:639–43.
291
[16] Bell FG, Wilkin TJ, Atwell JD. Microproteinuria in children with vesicoureteric reflux.
292
Br J Urol 1986; 58:605–9.
293
[17] Lebowitz RL, Olbing H, Parkkulainen KV, Smellie JM, Tamminen-Möbius TE.
294
International system of radiographic grading of vesicoureteric reflux: International Reflux
295
Study in Children. Pediatr Radiol 1985; 15:105–9.
296
[18] National High Blood Pressure Education Program Working Group on High Blood
297
Pressure in Children and Adolescents. The Fourth Report on the Diagnosis, Evaluation, and
298
Treatment of High Blood Pressure in Children and Adolescents. Pediatrics 2004; 114:555–76.
299
[19] Gao A, Cachat F, Faouzi M, Bardy D, Mosig D, Meyrat BJ, et al. Comparison of the
300
glomerular filtration rate in children by the new revised Schwartz formula and a new
301
generalized formula. Kidney Int 2013; 83:524–30.
AC C
EP
TE D
M AN U
SC
RI PT
269
9
ACCEPTED MANUSCRIPT [20] Gibb DM, Shah V, Preece M, Barratt TM. Variability of urine albumin excretion in
303
normal and diabetic children. Pediatr Nephrol 1989; 3:414-9.
304
[21] Chertin B, Solari V, Reen DJ, Farkas A, Puri P. Up-regulation of angiotensin-converting
305
enzyme (ACE) gene expression induces tubulointerstitial injury in reflux nephropathy. Pediatr
306
Surg Int 2002; 18:635–9.
307
[22] Birn H, Christensen EI. Renal albumin absorption in physiology and pathology. Kidney
308
Int 2006; 69:440–9.
309
[23] Wada T, Haneda M, Furuichi K, Babazono T, Yokoyama H, Iseki K, et al. Clinical
310
impact of albuminuria and glomerular filtration rate on renal and cardiovascular events, and
311
all-cause mortality in Japanese patients with type 2 diabetes. Clin Exp Nephrol. 2014;18:
312
613–20.
313
[24] Zhao YF, Zhu L, Liu LJ, Shi SF, Lv JC, Zhang H. Measures of Urinary Protein and
314
Albumin in the Prediction of Progression of IgA Nephropathy. Clin J Am Soc Nephrol.
315
2016;11: 947–55.
316
[25] Kidney Disease: Improving Global Outcomes (KDIGO) CKD Work Group. KDIGO
317
2012 clinical practice guideline for the evaluation and management of chronic kidney disease.
318
Kidney Int Suppl 2013; 3: 1–150.
319
[26] Tylicki L, Lizakowski S, Rutkowski B. Renin-angiotensin-aldosterone system blockade
320
for nephroprotection: current evidence and future directions. J Nephrol. 2012; 25: 900–10.
321
[27] Lahdes-Vasama T, Niskanen K, Rönnholm K. Outcome of kidneys in patients treated for
322
vesicoureteral reflux (VUR) during childhood. Nephrol Dial Transplant. 2006; 21:2491–7
323
[28] Zelmanovitz T, Gross JL, Oliveira JR, Paggi A, Tatsch M, Azevedo MJ. The receiver
324
operating characteristics curve in the evaluation of a random urine specimen as a screening
325
test for diabetic nephropathy. Diabetes Care 1997; 20:516–19.
SC
M AN U
TE D
EP
AC C
326
RI PT
302
327
Figure 1. ROC curve demonstrating the sensitivity and specificity of albumin to creatinine
328
ratio in detecting glomerular filtration rate below 70 mL/mn x1.73m2.
329 330
Figure 2. Association between albumin to creatinine ratio and glomerular filtration rate and
331
between albumin to creatinine ratio and filtration fraction.
332 333
Table 1. Summary of sociodemographic and clinical data n (%) Patient gender (= 141) 10
ACCEPTED MANUSCRIPT
334
M AN U
SC
RI PT
Female 68 (48) Male 73 (52) Age at the diagnosis of VUR in years, mean (SD) 2.0 (2.8) VUR grades (n = 141) VUR II 15 (11) VUR III 47 (33) VUR IV 79 (56) Bilateral high-grade VUR 42 (30) Single functioning kidney 15 (11) Interval between diagnosis of VUR and reflux surgery in years, mean (SD) 0.7 (1.3) Reflux surgery 113 (80) UTI (n = 123) 0 28 (23) 1–2 56 (45) >2 39 (32) Renal scintigraphy (n = 140) Scarring 97 (69) Unilateral 75 Bilateral 22 Hypertension 10 (7) UTI, urinary tract infection; VTR, vesicoureteral reflux.
335 336
Table 2. Linear regression model analysis to examine the association between the log-
337
transformed ACR and the explanatory factors
TE D
GFR FF
EP
Variables Gender (female = ref.)
Univariable analysis pβ coefficient (CI) value 0.19 (-0.62-0.25) 0.40 - 0.02 (-0.03 to 0.004) 0.01 0.07 (0.02-0.13) 0.01 - 0.03 (0.11-0.06) 0.55
AC C
Age at the diagnosis of VUR Grade of VUR VUR grade II (= ref.) VUR grade III VUR grade IV Bilateral high-grade VUR (unilateral VUR= ref.) Single functional kidney (two functional kidneys = ref.) Interval between diagnosis of VUR and reflux surgery History of reflux surgery (absence of reflux surgery = ref.) Number of UTI 0 (= ref.)
Multivariable analysis β coefficient p(CI) value -0.02 (-0.03 to 0.004) 0.01 0.07 (0.02-0.13) 0.01 -
-
0.07 (-0.70-0.84) 0.25 (-0.48-0.98)
0.87 0.50
-
-
0.44 (-0.03-0.92)
0.07
-
-
0.49 (-0.21-1.20)
0.17
-
-
0.01 (-0.21-0.23)
0.95
-
-
0.09 (-0.46-0.63)
0.76
-
-
-
-
-
11
ACCEPTED MANUSCRIPT
339
urinary tract infection; VUR, vesicoureteral reflux. The degree of mutual association with
340
factors was determined by the β coefficient (β) and the calculated p-value.
RI PT
338
1–2 0.18 (-0.43-0.79) 0.56 >2 0.58 (-0.07-1.23) 0.08 Presence of renal scarring (absence of renal scarring = ref.) 0.13 (-0.35-0.60) 0.60 Bilateral renal scarring (unilateral renal scarring = ref.) 0.34 (-0.29-0.97) 0.29 Presence of hypertension (absence of hypertension = ref.) 0.49 (-0.35-1.34) 0.25 ACR, albumin to creatinine ratio; FF, filtration fraction; GFR, glomerular filtration rate; UTI,
341
Table 3. Logistic regression model analysis to identify associated factors to the absence or
343
presence of pathological ACR
Male GFR, mean (SD)
17 (55) 14 (45) 81.7 (16.8) 22.0 (4.7) 2.1 (2.9)
AC C
EP
FF, mean (SD) Age at the diagnosis of VUR, mean (SD) Grades of VUR VUR grade II (Ref)
51 (46) 59 (54) 95.4 (14.5) 20.3 (3.6) 2.0 (2.9)
TE D
Female (ref.)
ACR <2.5 N (%) 110 (78)
Univariable analysis ACR ≥2.5 N (%) OR (95% CI) 31 (22)
VUR grade III
VUR grade IV Bilateral high-grade or unilateral VUR
Unilateral (ref.) Bilateral highgrade Two or single functional kidneys
M AN U
N Gender
SC
342
13 (12) 38 (34) 59 (54)
81 (74) 28 (26)
0.71 (0.32-1.59) 0.94 (0.91-0.97) 1.11 (1.01-2.23) 1.02 (0.89-1.18)
Multivariable analysis
pvalue
OR (95% CI)
pvalue
0.41
<0.001
0.04
0.94 (0.91-0.97) 1.12 (1.01-2.25)
0.77
-
-
-
-
-
-
<0.001
2 (6) 9 (29) 20 (65)
1.54 (0.29-8.07) 2.20 (0.46-10.62)
0.33
17 (55) 14 (45)
2.21 (0.95-5.13)
0.06
0.04
0.61
12
ACCEPTED MANUSCRIPT
Yes Number of UTI 0 (ref.) 1–2 >2 Renal scarring No (ref.)
Unilateral renal scarring (ref.) Bilateral renal scarring Hypertension No (ref.)
4 (13)
1.33 [0.39, 4.52]
0.64
0.7 (1.3)
0.7 (1.2)
1.00 (0.69-1.45)
1.00
22 (20) 88 (80)
6 (19) 25 (81)
1.04 (0.38-2.85)
25 (26) 42 (44) 28 (30)
3 (11) 14 (50) 11 (39)
2.78 (0.73-10.63) 3.27 (0.82-13.09)
0.09
35 (32) 74 (68)
8 (26) 23 (74)
1.36 (0.55-3.34)
0.50
60
15
2.29 (0.81-6.45)
0.12
TE D
Yes
11 (10)
14
8
102 (93)
29 (94)
-
-
-
-
RI PT
No (ref.)
27 (87)
-
-
-
-
-
-
0.94
SC
One Interval between diagnosis of VUR and reflux surgery in years History of reflux surgery
99 (90)
0.14
M AN U
Two (ref.)
345
urinary tract infection; VUR, vesicoureteral reflux.
AC C
346
EP
344
0.88 Yes 8 (7) 2 (6) (0.18-4.37) 0.88 ACR, albumin to creatinine ratio; FF, filtration fraction; GFR, glomerular filtration rate; UTI,
13
ACCEPTED MANUSCRIPT Figures legends: Figure 1: ROC Curve demonstrating the sensitivity and specificity of ACR in detecting GFR below 70 ml/mn x 1.73m2.
AC C
EP
TE D
M AN U
SC
RI PT
Figure 2: Association between ACR and GFR and between ACR and FF.
AC C
EP
TE D
M AN U
SC
RI PT
ACCEPTED MANUSCRIPT
AC C
EP
TE D
M AN U
SC
RI PT
ACCEPTED MANUSCRIPT
S1477-5131(17)30178-X
DOI:
10.1016/j.jpurol.2017.04.004
Reference:
JPUROL 2527
To appear in:
Journal of Pediatric Urology
Received Date: 22 November 2016 Accepted Date: 6 April 2017
Please cite this article as: de Sépibus R, Cachat F, Meyrat BJ, Dushi G, Boubaker A, Faouzi M, Girardin E, Chehade H, Urinary albumin excretion and chronic kidney disease in children with vesicoureteral reflux, Journal of Pediatric Urology (2017), doi: 10.1016/j.jpurol.2017.04.004. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
ACCEPTED MANUSCRIPT Urinary albumin excretion and chronic kidney disease in children with vesicoureteral reflux Romaine de Sépibus1,MD, François Cachat1,MD, Blaise J. Meyrat2,MD, Gezim Dushi2,MD, Ariane Boubaker3,MD, Mohamed Faouzi4,MD, Eric Girardin1,MD, Hassib Chehade1,MD
RI PT
Affiliations: 1Peadiatric Nephrology Unit, University Hospital of Lausanne (CHUV),
Lausanne, Switzerland, 2Department of Peadiatric surgery, University Hospital of Lausanne (CHUV), Lausanne, Switzerland, 3Institute of Radiology, Clinique de La Source, Lausanne,
Corresponding author's address:
M AN U
Hospital of Lausanne (CHUV), Lausanne, Switzerland.
SC
Switzerland, 4Institute of Social and Preventive Medicine, Biostatistics Unit, University
Hassib Chehade, MD, Peadiatric Nephrology Unit, Service of Peadiatrics, University Hospital of
Lausanne
(CHUV),
Rue
Bugnon
46,
1011
Lausanne,
Switzerland,
TE D
[email protected], Tel: +41 21 314 18 11; Fax: +41 21 314 36 26.
EP
Short title: Albuminuria in children with vesicoureteral reflux. Abstract word count: 300
AC C
Text word count: 2534
E-mail:
ACCEPTED MANUSCRIPT 1
Urinary albumin excretion and chronic kidney disease in children with vesicoureteral reflux
2 3
Romaine de Sépibusa, François Cachata, Blaise J. Meyratb, Gezim Dushib, Ariane Boubakerc,
4
Mohamed Faouzid, Eric Girardina, and Hassib Chehadea,*
5 6
a
7
Switzerland
8
b
9
Switzerland
RI PT
Paediatric Nephrology Unit, University Hospital of Lausanne (CHUV), Lausanne,
Department of Paediatric surgery, University Hospital of Lausanne (CHUV), Lausanne,
10
c
11
d
12
Lausanne (CHUV), Lausanne, Switzerland
SC
Institute of Radiology, Clinique de La Source, Lausanne, Switzerland
Institute of Social and Preventive Medicine, Biostatistics Unit, University Hospital of
M AN U
13 14
* Corresponding author. Paediatric Nephrology Unit, Service of Paediatrics, University
15
Hospital of Lausanne (CHUV), Rue Bugnon 46, 1011 Lausanne, Switzerland.
16
E-mail address: [email protected]
17
Summary Background: Albuminuria is a potential biomarker of chronic kidney disease
19
(CKD) in various glomerular diseases. Vesicoureteral reflux (VUR) often progresses to CKD,
20
and study is required of use of albuminuria as a biomarker for this condition. The aim of this
21
study was to evaluate the association between albuminuria and glomerular filtration rate
22
(GFR) or filtration fraction (FF) in children with VUR.
23
Study design: In this retrospective study, renal parameters of 141 children with VUR were
24
investigated, using inulin clearance, FF, and albuminuria. The association between urinary
25
albumin to creatinine ratio (ACR), GFR, and FF was analyzed in a continuous manner by
26
calculating the β coefficient, and also in a binary manner by calculating the OR.
27
Results: Using both continuous and binary analyses, ACR values were negatively and
28
significantly associated to GFR values in patients with low, normal, or high FF values (Table).
29
It was also positively and significantly associated with FF values in patients with low, normal
30
or high GFR values (Table). No association was found between ACR and gender, VUR stages
31
or laterality, number of urinary tract infection, presence of a single functional kidney, history
32
of reflux surgery, or renal scars or hypertension.
33
Discussion: ACR is associated with CKD in patients with VUR. In addition, increased urinary
34
albumin excretion cannot be completely and solely explained by decreased GFR and/or
AC C
EP
TE D
18
1
ACCEPTED MANUSCRIPT 35
increased FF values. The two main limitations of this study are the crude assessment of renal
36
scarring, which prevented finer analysis between albuminuria and renal scarring surface area,
37
and that the study cohort may not be representative of the true VUR population.
38
Conclusion: This study shows that albuminuria is associated with decreased renal function in
39
patients with VUR and that it could be used to monitor renal function in this condition.
RI PT
40 Table. Association results between ACR values, GFR values, and FF values using continuous
42
and binary methods
43
Association β coefficient = – p = 0.01 OR 0.94 p ≤ 0.001 between ACR 0.02 and GFR values Association β coefficient = p = 0.01 OR 1.11 p = 0.036 between ACR 0.07 and FF values ACR, albumin to creatinine ratio; FF, filtration fraction; GFR, glomerular filtration rate.
M AN U
SC
41
44 KEYWORDS
46
Vesicoureteral reflux;
47
Chronic kidney disease;
48
Albuminuria;
49
Child
50
TE D
45
Introduction
52
Vesicoureteral reflux (VUR) is a common urological abnormality in children with an
53
estimated prevalence of 1-6% [1,2]. Up to 30% of children with febrile urinary tract infection
54
(UTI) exhibit VUR and 15-30% will develop permanent renal scarring [3-6]. A substantial
55
number of children with reflux nephropathy (RN) will develop chronic renal failure, arterial
56
hypertension (HTN), and eventually end-stage renal disease [7]. According to the North
57
American Pediatric Renal Trials and Collaborative Studies, RN is the fourth leading cause of
58
chronic kidney disease (CKD), dialysis, and paediatric renal transplantation [8]. For this
59
reason, renal function of children with VUR should be monitored periodically, to identify the
60
group of patients most at risk of developing RN [9,10].
AC C
EP
51
61
It has been demonstrated that albuminuria is not only associated with decreased
62
glomerular filtration rate (GFR) but could also and by itself contribute to renal damage by
63
inducing tubular apoptosis and promoting interstitial fibrosis and inflammation [11,12].
64
Several studies have shown a close association between albuminuria and kidney diseases in 2
ACCEPTED MANUSCRIPT paediatric patients. Karlen et al. [13] found increased urine albumin excretion (>20
66
micrograms/min per 100 mL GFR) in more than 50% of children with pyelonephritic scarring.
67
Cachat et al. [14] showed that albuminuria predicted, to some extent, filtration fraction (FF)
68
in children with a single functioning kidney. However, studies analyzing the association
69
between albuminuria and GFR in children with VUR are scarce. Basic et al. [15] showed
70
significantly higher albuminuria and lower GFR in children with high grade VUR (IV and V)
71
compared with children with lower grade VUR (I, II, III). Similarly, Bell et al. [16] studied a
72
group of 36 children and found that albuminuria was significantly higher in high grade VUR
73
when compared with low and moderate grade VUR, but they did not evaluate GFR. To the
74
best of our knowledge it has not been clearly established whether albuminuria is a reliable
75
biomarker of kidney dysfunction in patients with VUR. This study aims to evaluate the
76
association between albuminuria and GFR using inulin clearance (Clin), and also between
77
albuminuria and FF in children with VUR.
78
Patients and methods
79
The study was conducted in the laboratory of the Paediatric Nephrology unit of Lausanne
80
University Hospital. It was approved by the local research ethics committee (Lausanne Ethic
81
Committee Protocol number: 215/13).
M AN U
SC
RI PT
65
One-hundred and forty-six paediatric patients with VUR were recruited. Patients who
83
were under treatment with angiotensin-converting enzyme inhibitors or angiotensin receptor
84
blockers were excluded (n=5). All included patients had a VUR confirmed by VCUG, and
85
GFR and FF measured using Clin and para-aminohippurate clearances (ClPAH), respectively,
86
together with a single morning urinary albumin excretion measurement. Patients were all
87
referred from treating physicians, nephrologists, or urologists. Indications for GFR evaluation
88
were left at the discretion of the treating physicians.
EP
VUR was classified according to the international reflux classification [17]. Bilateral
AC C
89
TE D
82
90
VUR was defined as bilateral high-grade VUR when grade III or IV or V VUR were found on
91
both sides. The highest grade of VUR was then recorded for statistical analysis. All other
92
conditions with grade I or II VUR on one side and grade III or IV or V VUR on the other side
93
were included in the group with unilateral VUR. History of VUR surgery was recorded. The
94
indication for surgery was left at the discretion of the treating physicians. The number of
95
symptomatic urinary tract infections (UTI) was recorded based on medical chart review, and
96
children were classified into three groups according to the number of UTIs (0; 1 or 2; >2
97
UTIs). The presence or absence of renal scarring was evaluated by two nuclear medicine
98
physicians using iodine-123 hippuran dynamic renal scintigraphy done at least 6 months after 3
ACCEPTED MANUSCRIPT 99 100
the last documented UTI. Renal scarring was defined as cortical thinning, flattening, or wedge-shaped defect. Urinary albumin excretion, GFR, and FF were measured simultaneously. All patients
102
fasted overnight before the procedure. Blood pressure was measured and hypertension (HTN)
103
was defined according to the National High Blood Pressure Education Program Working
104
Group on Children and Adolescents [18].
105
Urinary albumin excretion was measured by the immunoturbidimetric method, using a second
106
urine specimen in the morning, and was expressed as the urinary albumin to creatinine ratio
107
(ACR).
RI PT
101
Clin was used to measure GFR. Intravenous catheters were inserted into both of a
109
patient’s arms and an intravenous loading dose of 30 mg/kg of inulin (Inutest 25%, Fresenius,
110
Austria) was administered. A continuous infusion of inulin was applied for 3 hours to obtain a
111
constant plasmatic concentration between 200 and 500 mg/L. Diuresis was induced by oral
112
administration of 20 mL/kg of water (maximum 1,200 mL) during the first hour, followed by
113
maintenance hydration at a rate of 3 mL/kg/h. Concomitant intravenous hydration (NaCl
114
0.9%, maximum 300 mL) was given every 30 minutes. Urine and blood were collected every
115
30 minutes beginning after 90 minutes. Levels of inulin were measured, using the automated
116
anthrone method according to Wright and Gann. Intra-assay variability coefficients in serum
117
were 2.44% at 10 mg/100 mL, 1.47% at 30 mg/100 mL, and 0.94% at 40 mg/100 mL. Intra-
118
assay variability coefficients in urine were 1.71% at 10 mg/100 mL, 1.22% at 30 mg/100 mL,
119
and 1.07% at 50 mg/100 mL. GFR was calculated as the average of the three clearance
120
periods [19].
M AN U
TE D
EP
121
SC
108
Renal plasmatic flow (RPF) was measured using the ClPAH (Bratton-Marshall reaction). FF was defined as the GFR to RPF ratio (Clin/ClPAH).
123
Statistical analysis
124
All statistical analyses were performed by the Institute of Social and Preventive Medicine at
125
Lausanne University Hospital, using Stata statistical software (StataCorp. 2011. Stata
126
Statistical Software: Release 12. College Station, TX: StataCorp LP). The variables were
127
summarized by their median and interquartile range (IQR) and by numbers and percentages
128
for categorical variables.
AC C
122
129
Log-transformed ACR was used for expression of urinary albumin excretion. To avoid
130
any bias in the definition of pathological ACR, we studied this outcome as a continuous
131
variable and as a binary variable (coded 0/1). For the binary outcome, the patients were
132
divided into two groups, according to the absence or presence of pathological ACR (<2.5 4
ACCEPTED MANUSCRIPT 133
mg/mmol coded by 1 and ≥2.5 mg/mmol coded by 0) [20]. For the continuous outcome, the associations between log-transformed ACR and
135
several variables (GFR, FF, gender, VUR grade and laterality, age at diagnosis, interval
136
between diagnosis of VUR and surgery, history of urological surgery, single functioning
137
kidney, history of UTIs, presence of unilateral or bilateral renal scarring, and presence of
138
HTN) were examined using a univariable linear regression model. The degree of mutual
139
association was determined by the β coefficient (β) and the calculated p value. The binary
140
outcome was analyzed using a univariable logistic regression model to identify the associated
141
factors with the absence or presence of pathological ACR and OR and 95% CI were
142
calculated to set apart the differences between the two groups. A p-value <0.05 was
143
considered to be significant.
SC
RI PT
134
For both outcomes (continue and binary) associated factors to the outcome (p-value
145
<20%) were used in a backward-selection procedure to fit a multivariable model. Models
146
diagnostic (residual, influence statistics) were performed to examine how well the model fits
147
the data.
148
Results
149
Patient characteristics
150
One-hundred and forty-one children (68 females and 73 males) were included in this
151
retrospective study. Details of the patients’ characteristics are summarized in Table 1. Median
152
(IQR) follow-up was 10.21 (6.37) years. Median interval between GFR measurement and
153
renal scintigraphy was 0.24 months (IQR 18.48 months). Median (IQR) age at urinary
154
albumin excretion measurement, GFR, and FF measurements was 12.2 (7.54) years.
EP
TE D
M AN U
144
Median (IQR) GFR was 92.0 (19.50) mL/min per 1.73m2. Median (IQR) GFR in the
156
group with normal log-transformed ACR was 94.0 (18.0) mL/min per 1.73m2 and median
157
(IQR) GFR in the group with abnormal log-transformed ACR was 82.0 (30.0) mL/min per
158
1.73m2. Median (IQR) FF in all patients was 20.0% (14.0-34.0%). Median (IQR) FF in the
159
group with normal log-transformed ACR was 20.0% (4.0%), and median (IQR) FF in the
160
group with abnormal log-transformed ACR was 21% (5.0%).
161
ACR analysis as a continuous variable
162
In a linear regression model, using a univariable analysis, we showed a significant negative
163
association between log-transformed ACR and GFR (β = –0.02; p = 0.010) and a significant
164
positive association between log-transformed ACR and FF (β = 0.07; p = 0.010). This has
165
been confirmed in a multivariable analysis (Table 2).
166
AC C
155
A linear regression model analyzing the association between the log-transformed ACR 5
ACCEPTED MANUSCRIPT and other factors, showed no significant association between the log-transformed ACR and
168
different stages of VUR (ref. = VUR II) (VUR II versus VUR III (β = 0.07; p = 0.866); VUR
169
II versus VUR IV (β = 0.25; p = 0.504)) or VUR laterality (reference (ref.) = unilateral; β =
170
0.44; p = 0.067). There was no association between log-transformed ACR and all other
171
independent factors (gender, presence of a functional single kidney, history of reflux surgery,
172
number of UTI, renal scars, and hypertension). All results are summarized in Table 2.
RI PT
167
Constructing the ROC curve (Fig. 1), we found that an ACR value of 5.09 mg/mmol
174
had the best sensitivity and specificity of 69% and 91%, respectively, to detect inulin
175
clearance below 70 mL/mn x1.73m2.
176
ACR analysis as a binary variable
177
Thirty-one of the 141 patients (22%) exhibited pathological ACR (i.e. ACR ≥2.5 mg/mmol).
178
We found a significant association between the presence of abnormal log-transformed ACR
179
and decreased GFR (OR 0.94; 95% CI 0.91–0.97; p ≤ 0.001) and between abnormal log-
180
transformed ACR and increased FF (OR 1.11; 95% CI 1.01–2.23; p = 0.036). In multivariate
181
analysis, decreased GFR and increased FF were significantly and independently associated
182
with pathological log-transformed ACR with OR of 0.94 (95% CI 0.91–0.97; p ≤ 0.001) and
183
1.12 (95 % CI 1.01–1.25; p = 0.036), respectively.
M AN U
SC
173
There is no association between abnormal log-transformed ACR and VUR stages
185
(VUR II (ref.) versus VUR III (OR 1.54; 95% CI 0.29–8.07; p = 0.610), VUR II versus VUR
186
IV (OR 2.20; 95% CI 0.46–10. 62, p = 0.325)), and uni- or bilaterality of VUR (unilateral
187
VUR = ref.; OR 2.21; 95% CI 0.95–5.13; p = 0.064). Logistic regression model analysis also
188
showed no association between abnormal log-transformed ACR and other independent factors
189
(i.e. gender, the presence of a functional single kidney, previous reflux surgery, number of
190
UTI, renal scars, or hypertension). Distribution of patients according to ACR as a binary
191
variable is summarized in Table 3. The fitted multivariable logistic model was well calibrated
192
as shown by the Hosmer-Lemeshow goodness of fit test (chi-square (8) = 4.20, p = 0.83).
193
Discussion
194
We found in a paediatric population with VUR followed over a median period of 10.2 years a
195
significant negative association between ACR and GFR levels, independently of FF values.
196
This could indicate that factors other than haemodynamic disturbances play a significant role
197
in generation of increased albuminuria in CKD progression. Indeed, chronic interstitial
198
inflammation, fibrosis, tubular dilatation, and atrophy have been described in RN [21]. These
199
reported alterations may partially explain the pathophysiological mechanism of the negative
200
association between albuminuria and GFR independently of hemodynamic factors (e.g. FF),
AC C
EP
TE D
184
6
ACCEPTED MANUSCRIPT through impaired proximal tubular albumin reabsorption [22]. The ACR measurement in
202
patients with CKD has been evaluated in several clinical studies. It has been shown that
203
albuminuria can predict CKD and/or cardiovascular events in conditions such as diabetes
204
mellitus or IgA nephropathy [23,24]. Recently, the KDIGO guidelines added to their CKD
205
classification ACR as a marker for all-cause mortality in patients with CKD [25].
206
Furthermore, patients with proteinuric nephropathies greatly benefit from renin-angiotensin-
207
aldosterone system blockade [26]. This study allowed definition of an ACR threshold of 5.09
208
mg/mmol to best detect inulin clearances below 70 mL/mn x 1.73 m2 in children with
209
vesicoureteral reflux. The validity of this new ACR cutoff requires assessment in prospective
210
randomized controlled trials, and also the benefits of nephroprotection targeting renin-
211
angiotensin should be assessed according to ACR thresholds. Given the paucity of studies
212
analyzing the association between albuminuria and GFR in patients with VUR, we did not
213
calculate the sample size. However, in a post-hoc analysis, the study has a power of 98.52%
214
to detect a significant association between ACR and GFR.
M AN U
SC
RI PT
201
We also found a significant positive association between ACR and FF levels,
216
independently of GFR values. These results, using both continuous and binary analyses,
217
emphasize the role of renal perfusion in association with interstitial changes in this complex
218
process of increased albuminuria, CKD progression, and hyperfiltration. Finally, and in
219
contrast to the findings of Basic et al. [15] and Bell et al. [16], we did not find any association
220
between ACR and VUR stages. Furthermore, we did not find any association between ACR
221
and gender, VUR laterality, the number of UTI, the presence of a single functioning kidney,
222
the history of reflux surgery, and the presence of renal scars or HTN.
EP
TE D
215
All these observations emphasize the potential role of urinary albumin excretion in
224
monitoring of renal function (decreased GFR and/or hyperfiltration) in patients with VUR,
225
and suggest that increased urinary albumin excretion cannot be completely and solely
226
explained by decreased GFR and/or increased FF values.
227
AC C
223
As previously reported by Lahdes-Vasama et al. [27], it is also interesting to note that,
228
in both the linear and the logistic regression analysis, there was a tendency towards an
229
association between ACR and bilateral VUR or more than two UTIs.
230
The strength of this study was the long term follow-up of children over 10 years, the
231
use of inulin for GFR evaluation, and the analysis of the association among ACR, GFR, and
232
FF in both continuous and binary manners.
233
This study has several limitations. Although albumin excretion is often evaluated in
234
the literature with 24-hour urine collection or repeated samples collections, we used a single 7
ACCEPTED MANUSCRIPT random urine sample. However, the second morning urine ACR sample has been shown in
236
other studies to have a good association with the 24-hour urine albumin excretion rate [28]. In
237
addition, a small number of patients might still have a persistent VUR at the time of renal
238
function evaluation; however, we did not find any association between ACR and a history of
239
reflux surgery. Furthermore, the crude assessment of renal scarring prevented us from finer
240
analysis between albuminuria and renal scarring surface area. Finally, the study sample may
241
not be representative of the VUR population, as there were no patients with grade V VUR in
242
our cohort. In addition, all patients referred to our laboratory unit for inulin clearance were
243
included; however, we cannot exclude an inclusion bias because some patients with VUR
244
were not referred by their treating physicians for GFR evaluation. Referral was not based on a
245
particular ACR threshold.
SC
RI PT
235
In conclusion, this study showed an association between ACR and CKD. It also
247
defined a new ACR cutoff of 5.09 mg/mmol to accurately detect a GFR < 70 mL/mn x 1.73
248
m2 in patients with VUR.
249
Conflict of interest
250
None.
251
Funding
252
None.
253
References
254
[1] Toka HR, Toka O, Hariri A, Nguyen HT. Congenital anomalies of kidney and urinary
255
tract. Semin Nephrol 2010; 30:374–86.
256
[2] Guignard JP, Maherzi M, Torrado A. Screening for reflux. Lancet 1978; 2:316.
257
[3] Sargent MA. What is the normal prevalence of vesicoureteral reflux? Pediatr Radiol 2000;
258
30:587–93.
259
[4] Wadie GM, Moriarty KP. The impact of vesicoureteral reflux treatment on the incidence
260
of urinary tract infection. Pediatr Nephrol 2012; 27:529–38.
261
[5] Ismaili K, Wissing KM, Lolin K, Le PQ, Christophe C, Lepage P, Hall M. Characteristics
262
of first urinary tract infection with fever in children: a prospective clinical and imaging study.
263
Pediatr Infect Dis J 2011; 30:371–4.
264
[6] Coulthard MG, Lambert HJ, Vernon SJ, Hunter EW, Keir MJ, Matthews JN. Does prompt
265
treatment of urinary tract infection in preschool children prevent renal scarring: mixed
266
retrospective and prospective audits. Arch Dis Child 2014; 99:342–7.
267
[7] Ardissino G, Avolio L, Dacco V, Testa S, Marra G, Viganò S, et al. Long-term outcome
268
of vesicoureteral reflux associated chronic renal failure in children. Data from the ItalKid
AC C
EP
TE D
M AN U
246
8
ACCEPTED MANUSCRIPT Project. J Urol. 2004; 172: 305–10.
270
[8] Chesney RW, Brewer E, Moxey-Mims M, Watkins S, Furth SL, Harmon WE, et al.
271
Report of an NIH task force on research priorities in chronic kidney disease in children.
272
Pediatr Nephrol 2006; 21:14–25.
273
[9] Jodal U, Smellie JM, Lax H, Hoyer PF. Ten-year results of randomized treatment of
274
children with severe vesicoureteral reflux. Final report of the International Reflux Study in
275
Children. Pediatr Nephrol. 2006; 21:785–92.
276
[10] Sanna-Cherchi S, Ravani P, Corbani V, Parodi S, Haupt R, Piaggio G, et al. Renal
277
outcome in patients with congenital anomalies of the kidney and urinary tract. Kidney Int.
278
2009;76:528–33.
279
[11] Turin TC, Ahmed SB, Tonelli M, Manns B, Ravani P, James M, et al. Kidney function,
280
albuminuria and life expectancy. Can J Kidney Health Dis 2014; 1:33.
281
[12] Thomas ME, Brunskill NJ, Harris KP, Bailey E, Pringle JH, Furness PN, Walls J.
282
Proteinuria induces cell turnover: a potential mechanism for tubular atrophy. Kidney Int 1999;
283
55:890–898.
284
[13] Karlen J, Linné T, Wikstad I, Aperia A. Incidence of microalbuminuria in children with
285
pyelonephritis scarring. Pediatr Nephrol 1996; 10:705–8.
286
[14] Cachat F, Combescure C, Chehade H, Zeier G, Mosig D, Meyrat B, et al.
287
Microalbuminuria and hyperfiltration in subjects with nephro-urological disorders. Nephrol
288
Dial Transplant 2013; 28:386–91.
289
[15] Basic J, Golubovic E, Miljkovic P, Bjelakovic G, Cvetkovic T, Milosevic V.
290
Microalbuminuria in children with vesicoureteral reflux. Ren Fail 2008; 30:639–43.
291
[16] Bell FG, Wilkin TJ, Atwell JD. Microproteinuria in children with vesicoureteric reflux.
292
Br J Urol 1986; 58:605–9.
293
[17] Lebowitz RL, Olbing H, Parkkulainen KV, Smellie JM, Tamminen-Möbius TE.
294
International system of radiographic grading of vesicoureteric reflux: International Reflux
295
Study in Children. Pediatr Radiol 1985; 15:105–9.
296
[18] National High Blood Pressure Education Program Working Group on High Blood
297
Pressure in Children and Adolescents. The Fourth Report on the Diagnosis, Evaluation, and
298
Treatment of High Blood Pressure in Children and Adolescents. Pediatrics 2004; 114:555–76.
299
[19] Gao A, Cachat F, Faouzi M, Bardy D, Mosig D, Meyrat BJ, et al. Comparison of the
300
glomerular filtration rate in children by the new revised Schwartz formula and a new
301
generalized formula. Kidney Int 2013; 83:524–30.
AC C
EP
TE D
M AN U
SC
RI PT
269
9
ACCEPTED MANUSCRIPT [20] Gibb DM, Shah V, Preece M, Barratt TM. Variability of urine albumin excretion in
303
normal and diabetic children. Pediatr Nephrol 1989; 3:414-9.
304
[21] Chertin B, Solari V, Reen DJ, Farkas A, Puri P. Up-regulation of angiotensin-converting
305
enzyme (ACE) gene expression induces tubulointerstitial injury in reflux nephropathy. Pediatr
306
Surg Int 2002; 18:635–9.
307
[22] Birn H, Christensen EI. Renal albumin absorption in physiology and pathology. Kidney
308
Int 2006; 69:440–9.
309
[23] Wada T, Haneda M, Furuichi K, Babazono T, Yokoyama H, Iseki K, et al. Clinical
310
impact of albuminuria and glomerular filtration rate on renal and cardiovascular events, and
311
all-cause mortality in Japanese patients with type 2 diabetes. Clin Exp Nephrol. 2014;18:
312
613–20.
313
[24] Zhao YF, Zhu L, Liu LJ, Shi SF, Lv JC, Zhang H. Measures of Urinary Protein and
314
Albumin in the Prediction of Progression of IgA Nephropathy. Clin J Am Soc Nephrol.
315
2016;11: 947–55.
316
[25] Kidney Disease: Improving Global Outcomes (KDIGO) CKD Work Group. KDIGO
317
2012 clinical practice guideline for the evaluation and management of chronic kidney disease.
318
Kidney Int Suppl 2013; 3: 1–150.
319
[26] Tylicki L, Lizakowski S, Rutkowski B. Renin-angiotensin-aldosterone system blockade
320
for nephroprotection: current evidence and future directions. J Nephrol. 2012; 25: 900–10.
321
[27] Lahdes-Vasama T, Niskanen K, Rönnholm K. Outcome of kidneys in patients treated for
322
vesicoureteral reflux (VUR) during childhood. Nephrol Dial Transplant. 2006; 21:2491–7
323
[28] Zelmanovitz T, Gross JL, Oliveira JR, Paggi A, Tatsch M, Azevedo MJ. The receiver
324
operating characteristics curve in the evaluation of a random urine specimen as a screening
325
test for diabetic nephropathy. Diabetes Care 1997; 20:516–19.
SC
M AN U
TE D
EP
AC C
326
RI PT
302
327
Figure 1. ROC curve demonstrating the sensitivity and specificity of albumin to creatinine
328
ratio in detecting glomerular filtration rate below 70 mL/mn x1.73m2.
329 330
Figure 2. Association between albumin to creatinine ratio and glomerular filtration rate and
331
between albumin to creatinine ratio and filtration fraction.
332 333
Table 1. Summary of sociodemographic and clinical data n (%) Patient gender (= 141) 10
ACCEPTED MANUSCRIPT
334
M AN U
SC
RI PT
Female 68 (48) Male 73 (52) Age at the diagnosis of VUR in years, mean (SD) 2.0 (2.8) VUR grades (n = 141) VUR II 15 (11) VUR III 47 (33) VUR IV 79 (56) Bilateral high-grade VUR 42 (30) Single functioning kidney 15 (11) Interval between diagnosis of VUR and reflux surgery in years, mean (SD) 0.7 (1.3) Reflux surgery 113 (80) UTI (n = 123) 0 28 (23) 1–2 56 (45) >2 39 (32) Renal scintigraphy (n = 140) Scarring 97 (69) Unilateral 75 Bilateral 22 Hypertension 10 (7) UTI, urinary tract infection; VTR, vesicoureteral reflux.
335 336
Table 2. Linear regression model analysis to examine the association between the log-
337
transformed ACR and the explanatory factors
TE D
GFR FF
EP
Variables Gender (female = ref.)
Univariable analysis pβ coefficient (CI) value 0.19 (-0.62-0.25) 0.40 - 0.02 (-0.03 to 0.004) 0.01 0.07 (0.02-0.13) 0.01 - 0.03 (0.11-0.06) 0.55
AC C
Age at the diagnosis of VUR Grade of VUR VUR grade II (= ref.) VUR grade III VUR grade IV Bilateral high-grade VUR (unilateral VUR= ref.) Single functional kidney (two functional kidneys = ref.) Interval between diagnosis of VUR and reflux surgery History of reflux surgery (absence of reflux surgery = ref.) Number of UTI 0 (= ref.)
Multivariable analysis β coefficient p(CI) value -0.02 (-0.03 to 0.004) 0.01 0.07 (0.02-0.13) 0.01 -
-
0.07 (-0.70-0.84) 0.25 (-0.48-0.98)
0.87 0.50
-
-
0.44 (-0.03-0.92)
0.07
-
-
0.49 (-0.21-1.20)
0.17
-
-
0.01 (-0.21-0.23)
0.95
-
-
0.09 (-0.46-0.63)
0.76
-
-
-
-
-
11
ACCEPTED MANUSCRIPT
339
urinary tract infection; VUR, vesicoureteral reflux. The degree of mutual association with
340
factors was determined by the β coefficient (β) and the calculated p-value.
RI PT
338
1–2 0.18 (-0.43-0.79) 0.56 >2 0.58 (-0.07-1.23) 0.08 Presence of renal scarring (absence of renal scarring = ref.) 0.13 (-0.35-0.60) 0.60 Bilateral renal scarring (unilateral renal scarring = ref.) 0.34 (-0.29-0.97) 0.29 Presence of hypertension (absence of hypertension = ref.) 0.49 (-0.35-1.34) 0.25 ACR, albumin to creatinine ratio; FF, filtration fraction; GFR, glomerular filtration rate; UTI,
341
Table 3. Logistic regression model analysis to identify associated factors to the absence or
343
presence of pathological ACR
Male GFR, mean (SD)
17 (55) 14 (45) 81.7 (16.8) 22.0 (4.7) 2.1 (2.9)
AC C
EP
FF, mean (SD) Age at the diagnosis of VUR, mean (SD) Grades of VUR VUR grade II (Ref)
51 (46) 59 (54) 95.4 (14.5) 20.3 (3.6) 2.0 (2.9)
TE D
Female (ref.)
ACR <2.5 N (%) 110 (78)
Univariable analysis ACR ≥2.5 N (%) OR (95% CI) 31 (22)
VUR grade III
VUR grade IV Bilateral high-grade or unilateral VUR
Unilateral (ref.) Bilateral highgrade Two or single functional kidneys
M AN U
N Gender
SC
342
13 (12) 38 (34) 59 (54)
81 (74) 28 (26)
0.71 (0.32-1.59) 0.94 (0.91-0.97) 1.11 (1.01-2.23) 1.02 (0.89-1.18)
Multivariable analysis
pvalue
OR (95% CI)
pvalue
0.41
<0.001
0.04
0.94 (0.91-0.97) 1.12 (1.01-2.25)
0.77
-
-
-
-
-
-
<0.001
2 (6) 9 (29) 20 (65)
1.54 (0.29-8.07) 2.20 (0.46-10.62)
0.33
17 (55) 14 (45)
2.21 (0.95-5.13)
0.06
0.04
0.61
12
ACCEPTED MANUSCRIPT
Yes Number of UTI 0 (ref.) 1–2 >2 Renal scarring No (ref.)
Unilateral renal scarring (ref.) Bilateral renal scarring Hypertension No (ref.)
4 (13)
1.33 [0.39, 4.52]
0.64
0.7 (1.3)
0.7 (1.2)
1.00 (0.69-1.45)
1.00
22 (20) 88 (80)
6 (19) 25 (81)
1.04 (0.38-2.85)
25 (26) 42 (44) 28 (30)
3 (11) 14 (50) 11 (39)
2.78 (0.73-10.63) 3.27 (0.82-13.09)
0.09
35 (32) 74 (68)
8 (26) 23 (74)
1.36 (0.55-3.34)
0.50
60
15
2.29 (0.81-6.45)
0.12
TE D
Yes
11 (10)
14
8
102 (93)
29 (94)
-
-
-
-
RI PT
No (ref.)
27 (87)
-
-
-
-
-
-
0.94
SC
One Interval between diagnosis of VUR and reflux surgery in years History of reflux surgery
99 (90)
0.14
M AN U
Two (ref.)
345
urinary tract infection; VUR, vesicoureteral reflux.
AC C
346
EP
344
0.88 Yes 8 (7) 2 (6) (0.18-4.37) 0.88 ACR, albumin to creatinine ratio; FF, filtration fraction; GFR, glomerular filtration rate; UTI,
13
ACCEPTED MANUSCRIPT Figures legends: Figure 1: ROC Curve demonstrating the sensitivity and specificity of ACR in detecting GFR below 70 ml/mn x 1.73m2.
AC C
EP
TE D
M AN U
SC
RI PT
Figure 2: Association between ACR and GFR and between ACR and FF.
AC C
EP
TE D
M AN U
SC
RI PT
ACCEPTED MANUSCRIPT
AC C
EP
TE D
M AN U
SC
RI PT
ACCEPTED MANUSCRIPT