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Spironolactone to Prevent Peritoneal Fibrosis in Peritoneal Dialysis Patients: A Randomized Controlled Trial
Correspondence RESEARCH LETTER Spironolactone to Prevent Peritoneal Fibrosis in Peritoneal Dialysis Patients: A Randomized Controlled Trial To the Editor: One of the greatest challenges in the management of long-term peritoneal dialysis (PD) patients is ultrafiltration failure (UFF), which occurs as a result of a change in peritoneal transport characteristics.1 Approximately 4%-12% of PD patients have UFF, and 6 years after initiation of PD therapy, it affects up to 30% of patients.2 UFF is a consequence of chronic peritoneal damage due to chronic inflammation. These peritoneal membrane changes appear to be related to factors such as infections and exposure to hypertonic dialyzing solutions.3 In experimental models, activation of the renin-angiotensin-aldosterone system (RAAS) leads to peritoneal fibrosis (PF) by activation of NF-kB, TGF-b, and fibroblast and vascular endothelial growth factors, among others. Although irbesartan and spironolactone can prevent PF in rats,4 to our knowledge, no studies of humans have evaluated the role of spironolactone on inflammation and PF assessed by peritoneal biopsies. This was a randomized double-blind controlled trial including incident PD patients at our institution. Patients were eligible if they were 18 years or older, were not on RAAS blockade, and had signed informed consent. Exclusion criteria included pregnancy or potassium level . 5.5 mEq/L. One week after catheter placement, patients were randomly assigned to spironolactone 25 mg daily or placebo for 6 months. Peritoneal biopsies were performed when the PD catheter was placed and at the end of follow-up. Clinical and laboratory data were assessed monthly by the nephrologist.
Peritoneal biopsies were evaluated for PF (pixels in low-power field) and thickness (lineal mm) and by immunohistochemistry for collagen IV, CD20, and CD3 (detailed methods in Item S1). Sample size was calculated from the peritoneal biopsy study in spironolactone-treated rats,4 assuming a 5 0.05, b 5 0.2, and a peritoneal thickness difference of 124 (standard deviation, 34 mm), calculation yielded 6 patients per group; we aimed to recruit 20 patients per group. The primary outcome was PF and thickness. Secondary outcomes were markers of inflammation, episodes of peritonitis, and hyperkalemia. In 2008-2011, 38 patients signed informed consent; 7 missed the first visit, 5 were lost to follow-up (3 and 2 in the placebo and spironolactone groups, respectively), and 6 underwent transplantation before the final biopsy could be obtained. Due to recruitment difficulties, the study was terminated after completing 50% of the trial. Twenty patients completed the protocol, but 2 were eliminated (1 from each group) due to inadequate tissue sample, leaving 18 patients for analysis (Table 1). There were no significant differences between groups. At the end of follow-up, there was a significant increase in PF and thickness in both the treatment and placebo groups (P , 0.01). Although spironolactone had no effect on PF, it attenuated the increase in collagen IV and CD20 (Table 2; Fig S1). Extreme potassium values (.6 mEq/L) were not different between groups (mean potassium levels during follow-up of 4.8 6 0.4 and 4.4 6 0.5 [P 5 0.2] in the treatment and placebo groups, respectively). In this small cohort of PD patients treated with spironolactone, RAAS blockade resulted in less inflammation and collagen deposition, but no difference in peritoneal thickness or fibrosis. Compared to baseline, there was a significant increase in PF and thickness in both groups. Our results demonstrate the natural progression of fibrosis and thickness of the peritoneal membrane
Table 1. Baseline Characteristics of the Cohort All (N 5 18)
Spironolactone (n 5 9)
Placebo (n 5 9)
P
45.1 6 19.5 11 (61) 7 (39)
40.2 6 18.8 6 (67) 4 (44)
50 6 20 5 (56) 3 (33)
0.3 0.5 0.5
Kidney disease cause Diabetic nephropathy Unknown Lupus nephritis Nephrolithiasis Polycystic kidney disease ANCA GN Hypertensive nephropathy Membranoproliferative GN
6 (33) 6 (33) 1 (6) 1 (6) 1 (6) 1 (6) 1 (6) 1 (6)
3 (33) 5 (56) 0 (0) 0 (0) 0 (0) 0 (0) 0 (0) 1 (11)
3 (33) 1 (11) 1 (11) 1 (11) 1 (11) 1 (11) 1 (11) 0 (0)
Peritoneal equilibrium test High High-average Low-average Low
3 (17) 11 (61) 4 (22) 0 (0)
3 (33) 4 (44) 2 (22) 0 (0)
0 (0) 7 (78) 2 (22) 0 (0)
General characteristics Age (y) Male sex Diabetes
0.3
0.1
262.1 6 49.4
254.1 6 56.6
270 6 42.9
0.5
Dextrose exposure (g/dL)
6,094.5 6 1,738.7
5,898 6 2,398.9
6,291 6 762.9
0.6
Baseline peritoneal biopsy Fibrosis (pixel units) Thickness (mm) CD20 (pixel units) CD3 (pixel units) Collagen IV (pixel units)
17,055 [9,677-19,431] 1,049 [765-1,431] 12.5 [9-17] 7 [5-12] 108 [99-132]
18,451 [11,451-19,431] 865 [762-1,238] 11 [9-15] 7 [5-12] 108 [101-121]
12,771 [9,541-17,911] 1,110 [911-1,654] 13 [11-17] 8 [5-11] 123 [98-162]
0.3 0.3 0.8 0.6 0.7
Follow-up (d)
Note: Data shown as n (%), mean 6 SD, or median [25th-75th percentile].
Am J Kidney Dis. 2014;-(-):---
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Correspondence Table 2. Peritoneal Characteristics Between Groups at Follow-up Spironolactone (n 5 9)
Catheter Placement Fibrosis Thickness
End of Follow-up
Placebo (n 5 9)
P
18,451 [11,451-19,431] 19,561 [15,761-24,123] 0.07
Catheter Placement
P Between Groups
End of Follow-up
P
12,771 [9,541-17,911] 17,887 [14,998-21,611] 0.03
0.9
0.3
0.9
0.8
865 [762-1,238]
1,342 [1,281-1,781]
0.05
1,110 [911-1,654]
1,712 [1,441-1,981]
CD20
11 [9-15]
29 [28-39]
0.008
13 [11-17]
65 [50-86]
0.008
0.01
0.03
CD3
7 [5-12]
16 [12-25]
0.008
8 [5-11]
31 [27-57]
0.01
0.02
0.1
108 [101-121]
147 [146-156]
0.008
123 [98-162]
299 [276-320]
0.008
,0.001
,0.001
Collagen IV
0.1
Absolute Percent Change Change
Note: Data shown as median [25th-75th percentile]. Values are given in pixel units, except for thickness, which is given in mm.
over a relatively short time. In theory, RAAS blockade has potential benefits to reverse these processes. It is known that high glucose concentrations in peritoneal dialysate are associated with the increase in RAAS in human peritoneal mesothelial cells (in turn increasing fibronectin expression) and that the addition of angiotensin-converting enzyme inhibitors inhibits the production of TGF-b and fibronectin, preserves ultrafiltration, and decreases PF.5,6 In addition, infectious processes are associated with significantly higher angiotensin II levels in peritoneal fluid.7 In animal models, RAAS blockade shows a reduction of the damage to the peritoneum, with less thickness, inflammation, and PF and lower circulating TGF-1 levels.4 In our study, there was a significant decrease in inflammation and collagen deposition in the treatment group. There are several potential reasons why PF did not change between groups. First, the sample obtained is not always homogeneous, with areas having different muscle thickness resulting in discrepant Masson trichrome pixel determination. Second, markers of collagen and inflammation may not reflect all pathways to fibrosis. Third, the relative increased incidence in peritonitis might have decreased the effect of spironolactone on peritoneal architecture. Fourth, the dose and duration of spironolactone could have been suboptimal. Fifth, the study may not be powered to see a difference in PF or thickness. In conclusion, spironolactone decreased CD20 and collagen IV levels in peritoneal biopsy specimens from PD patients. The clinical benefits of these results should be evaluated in larger trials. Armando Vazquez-Rangel, MD, MS, Virgilia Soto, MD Marco Escalona, MD, Rafael G. Toledo, MD Edgar A. Castillo, MD, Nasser Abdel Polanco Flores, MD Ilse Falcon-Chavez, BS, Magdalena Madero, MD National Heart Institute, México City, México Corresponding author: [email protected]
Supplementary Material Figure S1: Representative images of collagen IV and CD20 at baseline and follow up. Item S1: Detailed methods. Note: The supplementary material accompanying this article (http://dx.doi.org/10.1053/j.ajkd.2014.01.426) is available at www. ajkd.org
References 1. Mujais S, Nolph K, Gokal R, et al. Evaluation and management of ultrafiltration problems in peritoneal dialysis. International Society for Peritoneal Dialysis Ad Hoc Committee on Ultrafiltration Management in Peritoneal Dialysis. Perit Dial Int. 2000;20(suppl 4):S5-S21. 2. Heimburger O, Waniewski J, Werynski A, et al. Peritoneal transport in CAPD patients with permanent loss of ultrafiltration capacity. Kidney Int. 1990;38(3):495-506. 3. Williams JD, Craig KJ, von Ruhland C, et al. The natural course of peritoneal membrane biology during peritoneal dialysis. Kidney Int Suppl. 2003;88:S43-S49. 4. Ersoy R, Celik A, Yilmaz O, et al. The effects of irbesartan and spironolactone in prevention of peritoneal fibrosis in rats. Perit Dial Int. 2007;27(4):424-431. 5. Noh H, Ha H, Yu MR, et al. Angiotensin II mediates high glucose-induced TGF-beta1 and fibronectin upregulation in HPMC through reactive oxygen species. Perit Dial Int. 2005;25(1):38-47. 6. Kiribayashi K, Masaki T, Naito T, et al. Angiotensin II induces fibronectin expression in human peritoneal mesothelial cells via ERK1/2 and p38 MAPK. Kidney Int. 2005;67(3):1126-1135. 7. Metsarinne K, Fyhrquist F, Gronhagen-Riska C. Increased levels of immunoreactive plasma renin substrate during infectious peritonitis in patients on continuous ambulatory peritoneal dialysis. Clin Sci (Lond). 1988;75(4):411-414.
Acknowledgements Support: The study was funded by the Mexican Kidney Foundation and the Miguel Aleman Foundation. Financial Disclosure: The authors declare that they have no relevant financial interests.
2
Received October 10, 2013. Accepted in revised form January 10, 2014. Ó 2014 by the National Kidney Foundation, Inc. http://dx.doi.org/10.1053/j.ajkd.2014.01.426
Am J Kidney Dis. 2014;-(-):---
Peritoneal biopsies were evaluated for PF (pixels in low-power field) and thickness (lineal mm) and by immunohistochemistry for collagen IV, CD20, and CD3 (detailed methods in Item S1). Sample size was calculated from the peritoneal biopsy study in spironolactone-treated rats,4 assuming a 5 0.05, b 5 0.2, and a peritoneal thickness difference of 124 (standard deviation, 34 mm), calculation yielded 6 patients per group; we aimed to recruit 20 patients per group. The primary outcome was PF and thickness. Secondary outcomes were markers of inflammation, episodes of peritonitis, and hyperkalemia. In 2008-2011, 38 patients signed informed consent; 7 missed the first visit, 5 were lost to follow-up (3 and 2 in the placebo and spironolactone groups, respectively), and 6 underwent transplantation before the final biopsy could be obtained. Due to recruitment difficulties, the study was terminated after completing 50% of the trial. Twenty patients completed the protocol, but 2 were eliminated (1 from each group) due to inadequate tissue sample, leaving 18 patients for analysis (Table 1). There were no significant differences between groups. At the end of follow-up, there was a significant increase in PF and thickness in both the treatment and placebo groups (P , 0.01). Although spironolactone had no effect on PF, it attenuated the increase in collagen IV and CD20 (Table 2; Fig S1). Extreme potassium values (.6 mEq/L) were not different between groups (mean potassium levels during follow-up of 4.8 6 0.4 and 4.4 6 0.5 [P 5 0.2] in the treatment and placebo groups, respectively). In this small cohort of PD patients treated with spironolactone, RAAS blockade resulted in less inflammation and collagen deposition, but no difference in peritoneal thickness or fibrosis. Compared to baseline, there was a significant increase in PF and thickness in both groups. Our results demonstrate the natural progression of fibrosis and thickness of the peritoneal membrane
Table 1. Baseline Characteristics of the Cohort All (N 5 18)
Spironolactone (n 5 9)
Placebo (n 5 9)
P
45.1 6 19.5 11 (61) 7 (39)
40.2 6 18.8 6 (67) 4 (44)
50 6 20 5 (56) 3 (33)
0.3 0.5 0.5
Kidney disease cause Diabetic nephropathy Unknown Lupus nephritis Nephrolithiasis Polycystic kidney disease ANCA GN Hypertensive nephropathy Membranoproliferative GN
6 (33) 6 (33) 1 (6) 1 (6) 1 (6) 1 (6) 1 (6) 1 (6)
3 (33) 5 (56) 0 (0) 0 (0) 0 (0) 0 (0) 0 (0) 1 (11)
3 (33) 1 (11) 1 (11) 1 (11) 1 (11) 1 (11) 1 (11) 0 (0)
Peritoneal equilibrium test High High-average Low-average Low
3 (17) 11 (61) 4 (22) 0 (0)
3 (33) 4 (44) 2 (22) 0 (0)
0 (0) 7 (78) 2 (22) 0 (0)
General characteristics Age (y) Male sex Diabetes
0.3
0.1
262.1 6 49.4
254.1 6 56.6
270 6 42.9
0.5
Dextrose exposure (g/dL)
6,094.5 6 1,738.7
5,898 6 2,398.9
6,291 6 762.9
0.6
Baseline peritoneal biopsy Fibrosis (pixel units) Thickness (mm) CD20 (pixel units) CD3 (pixel units) Collagen IV (pixel units)
17,055 [9,677-19,431] 1,049 [765-1,431] 12.5 [9-17] 7 [5-12] 108 [99-132]
18,451 [11,451-19,431] 865 [762-1,238] 11 [9-15] 7 [5-12] 108 [101-121]
12,771 [9,541-17,911] 1,110 [911-1,654] 13 [11-17] 8 [5-11] 123 [98-162]
0.3 0.3 0.8 0.6 0.7
Follow-up (d)
Note: Data shown as n (%), mean 6 SD, or median [25th-75th percentile].
Am J Kidney Dis. 2014;-(-):---
1
Correspondence Table 2. Peritoneal Characteristics Between Groups at Follow-up Spironolactone (n 5 9)
Catheter Placement Fibrosis Thickness
End of Follow-up
Placebo (n 5 9)
P
18,451 [11,451-19,431] 19,561 [15,761-24,123] 0.07
Catheter Placement
P Between Groups
End of Follow-up
P
12,771 [9,541-17,911] 17,887 [14,998-21,611] 0.03
0.9
0.3
0.9
0.8
865 [762-1,238]
1,342 [1,281-1,781]
0.05
1,110 [911-1,654]
1,712 [1,441-1,981]
CD20
11 [9-15]
29 [28-39]
0.008
13 [11-17]
65 [50-86]
0.008
0.01
0.03
CD3
7 [5-12]
16 [12-25]
0.008
8 [5-11]
31 [27-57]
0.01
0.02
0.1
108 [101-121]
147 [146-156]
0.008
123 [98-162]
299 [276-320]
0.008
,0.001
,0.001
Collagen IV
0.1
Absolute Percent Change Change
Note: Data shown as median [25th-75th percentile]. Values are given in pixel units, except for thickness, which is given in mm.
over a relatively short time. In theory, RAAS blockade has potential benefits to reverse these processes. It is known that high glucose concentrations in peritoneal dialysate are associated with the increase in RAAS in human peritoneal mesothelial cells (in turn increasing fibronectin expression) and that the addition of angiotensin-converting enzyme inhibitors inhibits the production of TGF-b and fibronectin, preserves ultrafiltration, and decreases PF.5,6 In addition, infectious processes are associated with significantly higher angiotensin II levels in peritoneal fluid.7 In animal models, RAAS blockade shows a reduction of the damage to the peritoneum, with less thickness, inflammation, and PF and lower circulating TGF-1 levels.4 In our study, there was a significant decrease in inflammation and collagen deposition in the treatment group. There are several potential reasons why PF did not change between groups. First, the sample obtained is not always homogeneous, with areas having different muscle thickness resulting in discrepant Masson trichrome pixel determination. Second, markers of collagen and inflammation may not reflect all pathways to fibrosis. Third, the relative increased incidence in peritonitis might have decreased the effect of spironolactone on peritoneal architecture. Fourth, the dose and duration of spironolactone could have been suboptimal. Fifth, the study may not be powered to see a difference in PF or thickness. In conclusion, spironolactone decreased CD20 and collagen IV levels in peritoneal biopsy specimens from PD patients. The clinical benefits of these results should be evaluated in larger trials. Armando Vazquez-Rangel, MD, MS, Virgilia Soto, MD Marco Escalona, MD, Rafael G. Toledo, MD Edgar A. Castillo, MD, Nasser Abdel Polanco Flores, MD Ilse Falcon-Chavez, BS, Magdalena Madero, MD National Heart Institute, México City, México Corresponding author: [email protected]
Supplementary Material Figure S1: Representative images of collagen IV and CD20 at baseline and follow up. Item S1: Detailed methods. Note: The supplementary material accompanying this article (http://dx.doi.org/10.1053/j.ajkd.2014.01.426) is available at www. ajkd.org
References 1. Mujais S, Nolph K, Gokal R, et al. Evaluation and management of ultrafiltration problems in peritoneal dialysis. International Society for Peritoneal Dialysis Ad Hoc Committee on Ultrafiltration Management in Peritoneal Dialysis. Perit Dial Int. 2000;20(suppl 4):S5-S21. 2. Heimburger O, Waniewski J, Werynski A, et al. Peritoneal transport in CAPD patients with permanent loss of ultrafiltration capacity. Kidney Int. 1990;38(3):495-506. 3. Williams JD, Craig KJ, von Ruhland C, et al. The natural course of peritoneal membrane biology during peritoneal dialysis. Kidney Int Suppl. 2003;88:S43-S49. 4. Ersoy R, Celik A, Yilmaz O, et al. The effects of irbesartan and spironolactone in prevention of peritoneal fibrosis in rats. Perit Dial Int. 2007;27(4):424-431. 5. Noh H, Ha H, Yu MR, et al. Angiotensin II mediates high glucose-induced TGF-beta1 and fibronectin upregulation in HPMC through reactive oxygen species. Perit Dial Int. 2005;25(1):38-47. 6. Kiribayashi K, Masaki T, Naito T, et al. Angiotensin II induces fibronectin expression in human peritoneal mesothelial cells via ERK1/2 and p38 MAPK. Kidney Int. 2005;67(3):1126-1135. 7. Metsarinne K, Fyhrquist F, Gronhagen-Riska C. Increased levels of immunoreactive plasma renin substrate during infectious peritonitis in patients on continuous ambulatory peritoneal dialysis. Clin Sci (Lond). 1988;75(4):411-414.
Acknowledgements Support: The study was funded by the Mexican Kidney Foundation and the Miguel Aleman Foundation. Financial Disclosure: The authors declare that they have no relevant financial interests.
2
Received October 10, 2013. Accepted in revised form January 10, 2014. Ó 2014 by the National Kidney Foundation, Inc. http://dx.doi.org/10.1053/j.ajkd.2014.01.426
Am J Kidney Dis. 2014;-(-):---