Abstract
In Japan, the standard method for measuring plasma aldosterone concentration (PAC) for primary aldosteronism (PA) diagnosis was changed from radioimmunoassay (RIA) to a novel chemiluminescent enzyme immunoassay (CLEIA). The purpose of this study is to simulate the possible impact of the change on PA diagnosis. This retrospective study assessed 2,289 PA patients. PACs measured by conventional RIA were transformed to estimated PACs (CLEIA) as follows: RIA (pg/mL) = 1.174 × CLEIA (pg/mL) + 42.3. We applied the estimated PAC (CLEIA) to the conventional cut-off of aldosterone-to-renin activity ratio ≥200 for screening and captopril challenge test (CCT) and PAC ≥60 pg/mL for saline infusion test (SIT). Application of the estimated PAC to screening and confirmatory tests decreased the number of PA diagnoses by 36% (743/2,065) on CCT and 52% (578/1,104) on SIT (discrepant cases). Among the discrepant cases, 87% (548/628) of CCT and 87% (452/522) of SIT were bilateral on adrenal venous sampling (AVS). Surgically treatable aldosterone-producing adenomas (APAs) were observed in 6% (36/579) and 5% (23/472) of discrepant cases on CCT and SIT, respectively; most were characterized by hypokalemia and/or adrenal nodule on CT imaging. Application of the PAC measured by the novel CLEIA to conventional cut-offs decreases the number of PA diagnoses. Although most discrepant cases were bilateral on AVS, there are some APA cases that were characterized by hypokalemia and/or adrenal tumor on CT. Further studies which evaluate PACs measured by both RIA and CLEIA for each patient are needed to identify new cut-offs for PAC measured by CLEIA.
PRIMARY ALDOSTERONISM (PA) is characterized by autonomous aldosterone secretion from the adrenal glands, which causes hypertension, and hypokalemia. The number of PA diagnoses has dramatically increased since the establishment of a method for measuring the plasma aldosterone concentration (PAC) and screening for PA by the aldosterone-to-renin activity ratio (ARR). Currently, PA is a major cause of secondary hypertension, with a prevalence of 5%–10% among hypertensive patients [1-5].
Regarding aldosterone measurement, a number of changes to the method and standard approach for measuring the PAC has affected the diagnosis of PA in the clinical setting. The measurement of the PAC by liquid chromatography/tandem mass spectrometry (LC-MS/MS) [6] is not indicated in Japan because the national health insurance system designates the cost of each laboratory investigation. Thus, a radioimmunoassay (RIA) has been used for measuring the PAC and plasma renin activity (PRA) in clinical practice. However, there were various issues for RIA, such as the use and disposal of radioisotopic material, the complexity of the manual assay, the poor traceability of certified reference materials, and a low detection sensitivity at lower concentrations [7, 8]. This has led to the development of a two-site sandwich chemiluminescent enzyme immunoassay (CLEIA) to measure the PAC using monoclonal antibodies immobilized onto ferrite particles, which was introduced in Japan in April 2021. The CLEIA for the PAC can be calibrated in-house using certified reference material for aldosterone [9]. Thus, the novel two-site sandwich CLEIA demonstrates good traceability to the certified reference material of aldosterone and good correlation with the values measured by LC-MS/MS [10].
On the other hand, the values obtained by CLEIA are much lower than those obtained by RIA, and it is highly possible that application of the PAC measured by CLEIA to the conventional cut-off levels of screening and confirmatory tests for PA would decrease the number of cases diagnosed with PA as essential hypertension.
Thus, in the current study, we evaluated the impact of a change from the conventional RIA to the novel CLEIA for PAC measurement on the diagnosis of PA. To do so, we used a large multicenter Japan PA registry based on the PAC measured by conventional RIA. PACs measured by conventional RIA were transformed to estimated PACs (CLEIA) using a conversion formula. We applied the estimated PAC (CLEIA) to the conventional cut-offs of the screening test, captopril challenge test (CCT), and saline infusion test (SIT) for the diagnosis of PA and evaluated the number of patients who were not diagnosed with PA by the estimated PAC (CLEIA) but considered discrepant cases. In addition, we assessed the clinical characteristics of the discrepant cases.
Methods
Setting and participants
We performed a retrospective observational cohort study of patients enrolled in the Japan Rare/Intractable Adrenal Diseases Study (JRAS), a nationwide registry of patients from 41 referral centers. The JRAS included patients with PA aged 20–90 years who underwent adrenal venous sampling (AVS). We used the JRAS dataset made available in May 2019, which included patients registered between January 2006 and December 2018. Due to the retrospective nature of the present study, the indication for adrenalectomy was determined in each patient by physicians at each center, based on the AVS results and other clinical characteristics. We included patients with PA who had an ARR ≥200 and PAC ≥120 pg/mL in screening tests and were diagnosed by CCT or SIT. Patients with PA diagnosed by the furosemide upright test alone and patients with missing data on important clinical variables were excluded from the analysis.
Diagnosis of primary aldosteronism
PA was diagnosed based on the guidelines of the Japan Endocrine Society and the Japan Society of Hypertension [11, 12]. Hypertensive patients with an elevated plasma ARR (≥200 with a PAC expressed in pg/mL and PRA in ng/mL/h) underwent at least one of the confirmatory tests, either CCT or SIT. If possible, antihypertensive medications were replaced with calcium channel blockers or α-blockers before testing. The CCT was considered positive if the ARR 60 or 90 minutes after 50 mg captopril intake was ≥200. The SIT was considered positive if the aldosterone level after 4 h of a 2-L saline infusion was ≥60.0 pg/mL. The diagnosis of PA was based on at least one positive confirmatory test result.
Determination of clinical characteristics
PAC was determined using a RIA (SPAC-S Aldosterone kits; Fuji Rebio Co., Ltd., Tokyo, Japan). PRA was measured using a RIA (PRA-FR RIA kits, Fuji Rebio Co., Ltd.; PRA RIA kits, Yamasa Co., Ltd., Chiba, Japan) and an enzyme immunoassay (PRA EIA kits; Yamasa Co., Ltd.). Hypokalemia was defined by receipt of potassium supplements or a serum potassium level <3.5 mEq/L. An adrenal nodule detected during CT was defined as a nodule with a diameter of at least 10.0 mm.
Estimation of the PAC measured by CLEIA
The purpose of this study was to evaluate the impact of a change from the conventional RIA to CLEIA for measuring the PAC in the diagnosis of PA. Thus, we estimated the value of the PAC measured by CLEIA by using PAC values measured by RIA (SPAC-S Aldosterone kits; Fuji Rebio Co., Ltd., Tokyo, Japan). The estimated PAC (CLEIA) was calculated using the following formula: CLEIA (pg/mL) = [RIA (pg/mL) – 42.3] /1.174. This formula was made based on the relationship between RIA-measured PAC and LC-MS/MS-measured PAC which should be equivalent value to CLEIA-measured PAC [7]. Because of the detection limit of the PAC on CLEIA (LUMIPULSE® Presto II; Fuji Rebio Co.), an estimated PAC (CLEIA) <4 pg/mL was set at 4 pg/mL.
Outcome assessment of adrenalectomy
The Primary Aldosteronism Surgical Outcome (PASO) investigators developed an international consensus for the assessment of outcomes after unilateral adrenalectomy for PA [13]. In this consensus, clinical and biochemical outcomes were classified as complete, partial, or absent based on the surgical results. In the current study, biochemical benefit was defined as complete or partial in the biochemical outcome of the PASO consensus. Detailed information on the outcome criteria has previously been provided [13]. Outcome assessment was performed 6 to 12 months after surgery.
Adrenal venous sampling
The AVS procedure has been described elsewhere [14, 15]. Briefly, adrenal blood samples were obtained via sequential cannulation by the percutaneous femoral vein approach. Cosyntropin was administered by bolus injection, by bolus injection followed by continuous infusion, or by continuous infusion only, throughout the procedure. Adrenal vein cannulation was defined as successful if the selectivity index was ≥5 after adrenocorticotropic hormone (ACTH) stimulation [16]. Unilateral hyperaldosteronism on AVS was defined as a lateralization index (LI) ≥4 after ACTH use, which is the most commonly used cut-off value [16, 17]. The selectivity index was defined as the ratio of the cortisol level in the adrenal vein to that in the inferior vena cava. The LI was calculated by dividing the aldosterone/cortisol ratio on the dominant side by that on the non-dominant side.
Definition of aldosterone-producing adenoma and idiopathic hyperaldosteronism
PA patients were diagnosed with aldosterone-producing adenoma (APA) if they underwent adrenalectomy and had pathological evidence of adenoma with a biochemical benefit on PASO criteria. PA patients were diagnosed with idiopathic hyperaldosteronism (IHA) if they did not undergo adrenalectomy and their LI was less than 4 on successful AVS or if they underwent adrenalectomy and did not have pathological evidence of adenoma or had biochemical absence on PASO criteria.
Statistical analyses
Continuous variables are expressed as the mean ± standard error or median (interquartile range) and were compared using the Mann–Whitney U test. Fisher’s exact test was used to compare categorical variables. Statistical significance was set at p < 0.05.
To identify the best overall set of predictors of APA among borderline PA cases, a logistic regression model with backward variable elimination at p = 0.1 was performed for those clinical variables that were significantly different between APA and IHA in the univariate analysis. Then, receiver operating curve analysis with C-statistics was performed to predict APA. Statistical analyses were performed with SPSS version 22 (IBM Corp., Armonk, NY, USA).
Ethical considerations
The study was conducted according to the tenets of the Declaration of Helsinki (1964) and its later amendments and the clinical study guidelines published by the Japanese Ministry of Health, Labour and Welfare. The study protocol was approved by the Ethics Committee of the National Hospital Organization Kyoto Medical Center (Kyoto, Japan) (the lead center) and the institutional ethics committees of the participating centers. The study was registered with the University Hospital Medical Information Network (UMIN ID: 18756). Informed consent was obtained from each patient.
Results
Study population
Of the 3,689 patients with information in the JRAS dataset, we excluded patients with missing or incomplete information regarding screening PAC and PRA data and patients with negative results on both CCT and SIT. A total of 2,289 patients were thus included in the current study. To evaluate the possible impact of a change in the method for measuring the PAC on the cut-off values of CCT and SIT, we divided the patients into two datasets, a CCT dataset comprising PA patients diagnosed by CCT (n = 2,065) and a SIT dataset comprising PA patients diagnosed through a positive result on SIT (n = 1,104) (Fig. 1).
Impact of estimated ARR and PAC (CLEIA) on the diagnosis of PA using the conventional cut-offs for screening and confirmatory tests
In analysis 1, we evaluated the impact of estimated ARR and PAC (CLEIA) on the diagnosis of PA using the conventional cut-offs for screening and confirmatory tests (Fig. 1).
In analysis 1 for CCT, application of the estimated PAC (CLEIA) to the conventional cut-off of 200 for both the screening and post-CCT ARR decreased the number of PA diagnoses by 36% (743 of 2,065) (discrepant cases). Compared with discrepant cases (Table 1), PAC, ARR, and post-CCT ARR were significantly higher and PRA and BMI were significantly lower in patients with PA who had positive screening and CCT results using the estimated ARR (CLEIA) (confirmed PA). In addition, participants with confirmed PA were more likely to be male and have hypokalemia and an adrenal nodule on CT imaging. Importantly, 87% (548 of 628) of discrepant cases were diagnosed with bilateral hyperaldosteronism on AVS and 6.2% (36 of 579) had surgically treatable APA. These proportions were 45% (502 of 1,113) and 45% (392 of 866), respectively, in the confirmed PA group.
Table 1
Baseline clinical characteristics of discrepant cases and patients with confirmed primary aldosteronism in analysis 1
|
Positive result of captopril challenge test
(n = 2,065) |
Positive result of saline infusion test
(n = 1,104) |
Discrepant cases
(n = 743) |
Confirmed PA
(n = 1,322) |
p |
Discrepant cases
(n = 578) |
Confirmed PA
(n = 526) |
p |
| Age (years) |
52.7 (± 11.1) |
52.1 (± 11.9) |
0.29 |
51.7 (± 11.5) |
50.6 (± 12.1) |
0.28 |
| Sex (male, %) |
43% (317/743) |
51% (679/1,322) |
<0.001 |
51% (292/578) |
57% (298/526) |
0.046 |
| Body mass index (kg/m2) |
25.2 (± 4.2)
(n = 742) |
24.6 (± 4.2)
(n = 1,320) |
<0.001 |
25.6 (± 4.2) |
24.7 (± 4.4)
(n = 525) |
<0.001 |
| Duration of hypertension (years) |
4 (1–10) |
7 (3–15) |
<0.001 |
5 (1–10) |
8 (3–15) |
<0.001 |
| Number of antihypertensive medications |
1 (1–1) |
1 (1–2) |
<0.001 |
1 (1–1) |
1 (1–2) |
<0.001 |
| Systolic blood pressure (mmHg) |
140.7 (± 18.8)
(n = 734) |
141.6 (± 17.7)
(n = 1,307) |
0.21 |
141.3 (± 17.7)
(n = 569) |
141.8 (± 17.3)
(n = 517) |
0.82 |
| Diastolic blood pressure (mmHg) |
87.0 (± 13.3)
(n = 734) |
86.8 (± 12.9)
(n = 1,307) |
0.74 |
88.3 (± 13.4)
(n = 569) |
87.2 (± 12.5)
(n = 517) |
0.36 |
| Serum potassium level (mmol/L) |
3.9 (± 0.40)
(n = 740) |
3.5 (± 0.57)
(n = 1,320) |
<0.001 |
3.9 (± 0.40)
(n = 576) |
3.4 (± 0.52)
(n = 524) |
<0.001 |
| Prevalence of hypokalemia (%) |
15% (113/740)
(n = 740) |
60% (790/1,320)
(n = 1,320) |
<0.001 |
19% (110/576)
(n = 576) |
66% (347/524)
(n = 524) |
<0.001 |
| eGFR (mL/min/1.73 m2) |
79.2 (± 17.7) |
79.6 (± 21.5) |
0.97 |
78.8 (± 18.0) |
80.4 (± 21.0) |
0.32 |
| PAC (RIA) (pg/mL) |
168 (142–215) |
265 (183–407) |
<0.001 |
179 (150–230) |
296 (213–429) |
<0.001 |
| Estimated PAC (CLEIA) (pg/mL) |
107 (85–147) |
190 (120–311) |
<0.001 |
116 (92–160) |
216 (145–329) |
<0.001 |
| Plasma renin activity (ng/mL/h) |
0.40 (0.30–0.60) |
0.20 (0.10–0.40) |
<0.001 |
0.40 (0.20–0.60) |
0.20 (0.10–0.40) |
<0.001 |
| ARR (RIA) |
430 (305–655) |
1,183 (656–2,111) |
<0.001 |
451 (281–756) |
1,175 (635–2,302) |
<0.001 |
| Estimated ARR (CLEIA) |
279 (188–425) |
791 (438–1,528) |
<0.001 |
297 (181–494) |
836 (433–1,736) |
<0.001 |
| Post-CCT ARR (RIA) |
274 (233–334) |
940 (568–1,891) |
<0.001 |
|
|
|
| Estimated post-CCT ARR (CLEIA) |
136 (108–169) |
607 (339–1,341) |
<0.001 |
|
|
|
| Post-SIT PAC (RIA) (pg/mL) |
|
|
|
84 (71–101) |
220 (149–340) |
<0.001 |
| Estimated post-SIT PAC (CLEIA) (pg/mL) |
|
|
|
36 (24–50) |
151 (91–254) |
<0.001 |
| Prevalence of adrenal nodule on CT imaging (%) |
32% (240/743) |
62% (815/1,322) |
<0.001 |
28% (159/578) |
65% (340/526) |
<0.001 |
| Prevalence of bilateral hyperaldosteronism on AVS (%) |
87% (548/628)
(n = 628) |
45% (502/1,113)
(n = 1,113) |
<0.001 |
87% (452/522)
(n = 522) |
38% (177/469)
(n = 469) |
<0.001 |
| Prevalence of clinical success defined by PASO criteria (%) |
70% (50/71)
(n = 71) |
78% (409/524)
(n = 524) |
0.18 |
63% (29/46)
(n = 46) |
79% (167/212)
(n = 212) |
0.035 |
| Prevalence of biochemical success defined by PASO criteria (%) |
79% (44/56)
(n = 56) |
93% (415/444)
(n = 444) |
<0.001 |
84% (31/37)
(n = 37) |
95% (170/179)
(n = 179) |
<0.001 |
| Prevalence of aldosterone-producing adenoma (%) |
6.2% (36/579)
(n = 579) |
45% (392/866)
(n = 866) |
<0.001 |
4.9% (23/472)
(n = 472) |
49% (163/331)
(n = 331) |
<0.001 |
An adrenal nodule on CT imaging was defined as a nodule with a diameter of at least 10.0 mm.
Bilateral hyperaldosteronism was defined by a lateralization index ≤4.0 on adrenal venous sampling. In patients with positive result of captopril challenge test, a total of 90 patients in discrepant cases and 694 in confirmed PA cases undertook adrenalectomy. In patients with positive result of saline infusion test, a total of 62 patients in discrepant cases and 283 in confirmed PA cases undertook adrenalectomy.
Aldosterone-producing adenoma was defined by both pathological evidence of adenoma on adrenalectomy and biochemical success defined by PASO criteria.
PA, primary aldosteronism; eGFR, estimated glomerular filtration rate; PAC, plasma aldosterone concentration; RIA, radioimmunoassay; CLEIA, chemiluminescent enzyme immunoassay; ARR, aldosterone-to-renin ratio; CCT, captopril challenge test; SIT, saline infusion test; CT, computed tomography; AVS, adrenal venous sampling; PASO, primary aldosteronism surgical outcome.
In analysis 1 for SIT, application of the estimated PAC (CLEIA) to the conventional cut-off of 200 for screening ARR and the conventional cut-off of 60 for post-SIT PAC decreased the number of PA diagnoses by 52% (578 of 1,104) (discrepant cases). Compared with discrepant cases (Table 1), PAC, ARR, and post-SIT PAC were significantly higher, and PRA and BMI were significantly lower in patients with PA who had positive screening and SIT results using the estimated PAC (CLEIA) (confirmed PA). In addition, patients with confirmed PA were more likely to be male and to have hypokalemia and an adrenal nodule on CT imaging. Importantly, 87% (452 of 522) of discrepant cases were diagnosed with bilateral hyperaldosteronism on AVS and 4.9% (23 of 472) had surgically treatable APA. These proportions were 38% (177 of 469) and 49% (163 of 331), respectively, in patients with confirmed PA.
Distribution of APA and IHA cases by the estimated ARR and PAC (CLEIA) on CCT and SIT
To evaluate the prevalence of APA and IHA in discrepant PA cases, we excluded patients who did not fulfill the criteria of either APA or IHA from the dataset for analysis 1 (Fig. 1). First, we evaluated the prevalence of APA in each group categorized by the quartile of the estimated screening ARR (CLEIA), post-CCT ARR (CLEIA), and post-SIT PAC (CLEIA). As shown in Fig. 2, the higher the estimated screening ARR (CLEIA), post-CCT ARR (CLEIA), and post-SIT PAC (CLEIA), the higher the prevalence of APA. Next, we evaluated the distribution of APA and IHA cases by the estimated ARR on screening and CCT (CLEIA), and PAC (CLEIA) on SIT (Fig. 3A and B). Importantly, 8.4% (36 of 428) of APAs were in discrepant cases among patients diagnosed by CCT. Then, to reduce the number of cases overlooked with the conventional cut-off of the ARR for screening and CCT, we defined the PA patients with estimated ARRs (CLEIA) on screening or CCT between 100 and 200 as borderline cases. Of the borderline cases, 7.2% (34 of 473) were APA cases; they also had a higher prevalence of hypokalemia and an adrenal nodule on CT imaging and a higher PAC and post-CCT ARR (CLEIA) (two APA cases had a post-CCT ARR <100) (Table 2). Multivariable logistic regression with backward elimination for the prevalence of hypokalemia, an adrenal nodule on CT imaging, estimated PAC (CLEIA), and post-CCT ARR (CLEIA) demonstrated that prevalence of hypokalemia and an adrenal nodule on CT imaging were independent predictors of APA among borderline cases, with C-statistics of 0.833 (95% confidence interval 0.780–0.886) (Fig. 4).
Table 2
Baseline clinical characteristics of APA and IHA in borderline PA cases
|
Positive result of captopril challenge test
(n = 473) |
Positive result of saline infusion test
(n = 472) |
| APA (n = 34) |
IHA (n = 439) |
p |
APA (n = 23) |
IHA (n = 449) |
p |
| Age (years) |
50.6 (± 12.2) |
51.9 (± 10.8) |
0.38 |
48.7 (± 10.9) |
51.1 (± 11.3) |
0.23 |
| Sex (male, %) |
56% (19/34) |
44% (194/439) |
0.21 |
57% (13/23) |
49% (222/449) |
0.53 |
| Body mass index (kg/m2) |
25.0 (± 3.6) |
25.4 (± 4.1)
(n = 438) |
0.62 |
24.9 (± 4.1) |
25.7 (± 4.1) |
0.23 |
| Duration of hypertension (years) |
9 (2–19) |
4 (1–10) |
0.036 |
5 (1–16) |
5 (1–10) |
0.55 |
| Number of antihypertensive medications |
1 (1–1) |
1 (1–2) |
0.012 |
1 (1–1) |
1 (1–2) |
0.027 |
| Systolic blood pressure (mmHg) |
141.1 (± 19.0) |
141.1 (± 21.5)
(n = 434) |
0.79 |
145.7 (± 17.5) |
141.0 (± 17.6)
(n = 442) |
0.18 |
| Diastolic blood pressure (mmHg) |
88.2 (± 14.6) |
87.6 (± 13.5)
(n = 434) |
0.72 |
90.4 (± 15.1) |
88.5 (± 13.5)
(n = 442) |
0.59 |
| Serum potassium level (mmol/L) |
3.7 (± 0.57)
(n = 33) |
3.9 (± 0.38)
(n = 438) |
0.007 |
3.7 (± 0.45)
(n = 22) |
3.9 (± 0.39)
(n = 448) |
0.12 |
| Prevalence of hypokalemia (%) |
45% (15/33)
(n = 33) |
11% (50/438)
(n = 438) |
<0.001 |
41% (9/22)
(n = 22) |
16% (70/448)
(n = 448) |
0.005 |
| eGFR (mL/min/1.73 m2) |
82.2 (± 20.2) |
79.6 (± 17.3) |
0.46 |
83.2 (± 18.1) |
79.5 (± 18.0) |
0.46 |
| PAC (RIA) (pg/mL) |
206 (156–298) |
169 (143–211) |
0.013 |
191 (160–215) |
179 (150–228) |
0.54 |
| Estimated PAC (CLEIA) (pg/mL) |
139 (97–217) |
108 (86–144) |
0.013 |
127 (100–147) |
116 (92–158) |
0.54 |
| Plasma renin activity (ng/mL/h) |
0.50 (0.30–0.80) |
0.40 (0.30–0.60) |
0.11 |
0.30 (0.20–0.80) |
0.40 (0.30–0.60) |
0.87 |
| ARR (RIA) |
406 (275–799) |
417 (302–627) |
0.90 |
533 (249–835) |
438 (281–732) |
0.80 |
| Estimated ARR (CLEIA) |
252 (182–543) |
270 (187–402) |
0.59 |
353 (179–531) |
287 (181–477) |
0.67 |
| Post-CCT ARR (RIA) |
305 (243–354) |
276 (238–340) |
0.36 |
|
|
|
| Estimated post-CCT ARR (CLEIA) |
166 (127–205) |
145 (123–172) |
0.014 |
|
|
|
| Post-SIT PAC (RIA) (pg/mL) |
|
|
|
83 (70–139) |
82 (71–99) |
0.57 |
| Estimated post-SIT PAC (CLEIA) (pg/mL) |
|
|
|
35 (24–82) |
34 (24–48) |
0.57 |
| Prevalence of adrenal nodule on CT imaging (%) |
65% (22/34) |
26% (112/439) |
<0.001 |
43% (10/23) |
23% (105/449) |
0.043 |
| Prevalence of bilateral hyperaldosteronism on AVS (%) |
38% (11/29)
(n = 29) |
97% (424/437)
(n = 437) |
<0.001 |
35% (8/23) |
97% (436/449) |
<0.001 |
APA was defined by both pathological evidence of adenoma on adrenalectomy and biochemical success defined by PASO criteria.
IHA was defined either by patients who did not undergo adrenalectomy and had a confirmed laterality ratio below 4.0 on adrenal venous sampling or by patients who underwent adrenalectomy and did not have pathological evidence of adenoma or had biochemical absence on PASO criteria.
An adrenal nodule on CT imaging was defined as a nodule with a diameter of at least 10.0 mm.
Bilateral hyperaldosteronism was defined by a lateralization index ≤4.0 on adrenal venous sampling.
PA, primary aldosteronism; APA, aldosterone-producing adenoma; IHA, idiopathic hyperaldosteronism; eGFR, estimated glomerular filtration rate; PAC, plasma aldosterone concentration; RIA, radioimmunoassay; CLEIA, chemiluminescent enzyme immunoassay; ARR, aldosterone-to-renin ratio; CCT, captopril challenge test; SIT, saline infusion test; CT, computed tomography; AVS, adrenal venous sampling.
Regarding SIT, 12% (23 of 186) of APAs were in discrepant cases on SIT. To reduce the number of cases overlooked with the conventional cut-off of the ARR on screening and PAC on SIT, we defined PA patients whose estimated ARR (CLEIA) on screening was between 100 and 200 or whose estimated post-SIT PAC (CLEIA) was between 15 and 60 pg/mL as borderline cases. Among the borderline cases, 4.9% (23 of 472) had APA; they also had a higher prevalence of hypokalemia and an adrenal nodule on CT imaging (Table 2). Multivariable logistic regression with backward elimination for the prevalence of hypokalemia and an adrenal nodule on CT imaging demonstrated that both variables were independent predictors of APA in borderline PA, with C-statistics of 0.676 (95% confidence interval 0.551–0.801) (Fig. 4).
Possible algorithm for the management of PA
Fig. 5 shows the number of patients in each group in the current study. PA patients who were diagnosed by the conventional cut-offs of screening and confirmatory tests with the PAC measured by CLEIA are recommended to undergo AVS and should be considered for adrenalectomy based on the results of AVS and other clinical characteristics because of their high prevalence of surgically treatable APA. On the other hand, not all borderline cases are recommended to undergo AVS because of their high prevalence of IHA. Among borderline cases, evaluation of clinical characteristics, especially the serum potassium level, and CT imaging for adrenal nodules are important to determine the indication for AVS and adrenalectomy. For example, 170 of the 471 borderline PAs on CCT had either hypokalemia or an adrenal nodule on CT imaging, and these patients comprised 88% (29 of 33) of the APA cases in the borderline PA group. A total of 162 of the 470 borderline PAs on SIT had either hypokalemia or an adrenal nodule on CT imaging, and these patients comprised 64% (14 of 22) of the APA cases in the borderline PA group.
Discussion
Although PA is the most common form of secondary hypertension and the prevalence reaches 5%–10% among hypertensive patients, there have been methodological issues with the RIAs used to measure the PAC, which are critical for the screening and diagnosis of PA. For example, this assay has demonstrated limitations in sensitivity and reproducibility, particularly at lower concentration ranges [7], as well as at higher concentration ranges in adrenal vein samples. In this regard, the novel two-site sandwich CLEIAs has demonstrated good traceability for the certified reference material of aldosterone, good linearity over a wide range of concentrations, including lower concentrations, and good correlation with LC-MS/MS results [10, 18-20]. Furthermore, the introduction of the CLEIA method has enabled international comparison of PA diagnosis because of the comparability between CLEIA-measured and LC-MS/MS-measured PAC [7]. Thus, CLEIA is applicable as a standard assay method for measuring the PAC in daily clinical practice.
On the other hand, the PAC measured by CLEIA was significantly lower than the value obtained with RIA, and the new cut-offs of screening and confirmatory tests using CLEIA for the diagnosis of PA remain to be resolved. In the current study, we evaluated the impact of a change from the conventional RIA to the novel CLEIA for measuring the PAC on the diagnosis of PA. Importantly, application of the PAC measured by the novel CLEIA assay to the conventional cut-offs of screening and confirmatory tests dramatically decreased the number of diagnoses, especially those of bilateral hyperaldosteronism. On the other hand, there were some misdiagnoses of surgically treatable APAs as essential hypertension. These results bring an issue of what is PA, how should we diagnose PA, and how should we treat PA?
Typical PA with phenotype of hypertension, hypokalemia, and unilateral adrenal nodule have been firstly described by Conn [21]. The pathological findings of these PAs are adenoma, and recent reports demonstrated that these unilateral APAs can be treated by adrenalectomy better than by medications [22-24]. On the other hand, the introduction of the ARR and the widespread screening of patients with hypertension led to a 5- to 15-fold increase in the diagnosis of milder form of PA [2]. Most of these mild PAs have normokalaemia with bilateral hyperaldosteronism by AVS, and can be treated by mineralocorticoid receptor antagonists.
Thus, AVS is strongly recommended as the standard criterion in the clinical guidelines for the subtype diagnosis of PA to decide surgical indication. However, despite significant increases in the number of PA diagnoses, as well as the implementation of AVS and its technical development over the past decade, the overall rate of adrenalectomy corresponds to only one-third of patients with PA who undergo AVS [25]. Given the various issues associated with AVS, such as its invasive and technically demanding nature, cost, and radiation exposure, its diagnostic efficiency needs to be improved. The identification of those with unilateral PA who would benefit most from adrenalectomy remains essential, and improvements in the noninvasive pre-AVS prediction of the bilateral subtype are required, especially in Japan. In this regard, PA diagnosed by the conventional cut-off using the PAC measured by CLEIA in the current study is a good indication for AVS because of its high prevalence of surgically treatable APA. On the other hand, most of the borderline cases were ultimately diagnosed with IHA (bilateral hyperaldosteronism). Thus, is there any point to diagnose these borderline cases as PA?
Importantly, Sartoli et al. defined hypertensive patients with a positive screening test and negative confirmatory test as having aldosterone-associated hypertension [26]. They also found that a similar number of patients with aldosterone-associated hypertension and patients with definitive PA developed resistant hypertension over a 22-month follow-up. Shibata et al. [27]. defined hypertensive patients with hyperaldosteronemia who did not satisfy the diagnostic criteria for PA as a subtype of patients with mineralocorticoid receptor-associated hypertension, probably caused by pathophysiological activation of the aldosterone–mineralocorticoid receptor axis [28]. In addition, a recent report demonstrated that, beyond the categorical definition of PA, there is a prevalent continuum of renin independent aldosterone production that parallels the severity of hypertension, implying that PA is a borderline disease [29]. Thus, although it may be less necessary to perform AVS for these borderline cases because of the low prevalence of APA, it is possible that blockade of the mineralocorticoid receptor axis in these patients improves prognosis. Further studies are needed to elucidate the possible importance of the diagnosis of borderline cases with PA in terms of medication and natural course.
In the current study, we also found that about 5% of borderline cases had surgically treatable APAs. These APAs were confirmed by both pathological findings on adrenalectomy and by biochemical benefit on PASO criteria. Adrenalectomy theoretically cures the disease through the removal of the tumor-producing aldosterone [30]. Importantly, accumulating evidence indicates that medical therapy leads to less favorable outcomes than surgical treatment for APAs [31]. Thus, it would be unfavorable for endocrinologists to misdiagnose these APAs in the clinical setting. It is important to note that, in the current study, these APAs could be differentiated among borderline cases by clinical characteristics, including hypokalemia and an adrenal nodule on CT imaging. These two clinical factors are also important for differentiating APA not only in borderline cases, but also in those with confirmed PA (data are not shown). These results imply that detailed evaluation by AVS may be necessary even in borderline cases if they have hypokalemia or an adrenal nodule on CT imaging.
The strength of the present study is that we used a large multicenter dataset of patients with PA who underwent AVS. Importantly, APAs in the current study were defined by both pathological findings and surgical outcome, which strengthen the interpretation. On the other hand, an important limitation of this study is that we did not directly measure the PAC by CLEIA. PAC measured by RIA tends to have intra-patient variability, which may limit the interpretation of the results [32]. In addition, we used the formula, CLEIA (pg/mL) = [RIA (pg/mL) – 42.3] /1.174, to estimate the CLEIA-measured PAC. This formula is based on the relationship between RIA-measured PAC and LC-MS/MS-measured PAC which should be equivalent value to CLEIA-measured PAC. Thus, the current study is based on the speculation, and direct comparison between CLEIA-measured PAC and RIA-measured PAC would be needed [10, 33].
Additionally, we included only patients with PA who underwent AVS and this could increase the proportion of APA among PA compared with the true proportion as a selection bias. Finally, PA patients in the current study were diagnosed by Japanese clinical practice guideline, there must be a limitation that may limit the application to other countries clinical practices.
Acknowledgments
We thank Yoshiyu Takeda (Kanazawa University), Kenji Oki (Hiroshima University), Ryuichi Sakamoto (Kyushu University), Dr. Michio Otsuki (Tokyo Women’s Medical University), Shintaro Okamura (Tenri Hospital), Dr. Tomikazu Fukuoka (Matsuyama Red Cross Hospital), Yuichi Fujii (JR Hiroshima Hospital), Dr. Shozo Miyauchi (Ehime Prefectural Central Hospital), Yoshiro Chiba (Mito Saiseikai General Hospital), Masanobu Yamada (Gunma University), Yoichi Ohno (Kyoto University), Yuichi Matsuda (Sanda City Hospital), Megumi Fujita (Tokyo University), Minemori Watanabe (Okazaki City Hospital), Kohei Kamamura (Shinko Hospital) for collecting the clinical data.
This study was supported by the JPAS and JRAS from the Japan Agency for Medical Research and Development (AMED) under Grant Number JP17ek0109122 and JP20ek0109352 and by a Research Grant and the National Center for Global Health and Medicine, Japan (27-1402, 30-1008). The data that support the findings of this study are available from the corresponding author upon reasonable request.
Conflicts of Interest
The authors declare no conflicts of interest.
Author Contributions
Study Concept: H.K., M.N. Writing manuscript: H.K. Statistical analysis: H.K. Acquisition of data: H.K., Y.N., M.A., A.T., M.S., T.K., I.K., T.I., M.T., S.I., N.W., T.Y., K.T., K.T., Y.O., N.I., K.Y., H.R., M.N., and contributors from the JPAS/JRAS study group. Critical revision of the manuscript for important intellectual content: All authors.
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