The Role of ADMA as an Indicator of Progression in Early Stage of CKD

Abstract

Several studies have shown an association of fibroblast growth factor-23 (FGF-23), 25-hydroxyvitamin D (25(OH)D), and asymmetric dimethylarginine (ADMA) with the pathogenesis of albuminuria. However, the direct relationship of these biomarkers with albuminuria independent of other risk factors for chronic kidney disease (CKD) remains controversial. FGF-23 and ADMA levels were associated with the progression of CKD, with a cutoff value of ≥100 RU/mL for FGF-23 and 0.69 μmol/L for ADMA. Background/Objectives: To analyze the correlation between FGF-23, 25(OH)D, and ADMA levels and albuminuria. Methods: This was an observational analytic study with a cross-sectional design conducted in patients with CKD with various disease stages (non-dialysis). The output is albuminuria. Statistical analysis was performed using multivariate logistic regression analysis. Results: This study included 107 patients with CKD stages 2–5 with an average age of 57.32 years. Their average FGF-23, vitamin D, ADMA, and uACR levels were 197.75 RU/mL, 23.44 ng/mL, 0.719 µmol/L, and 940 mg/g, respectively. FGF-23 was weakly correlated with uACR (r = 0.252; p = 0.009). Vitamin D was weakly correlated with uACR (r = −0.375; p = 0.000). ADMA was strongly correlated with uACR (r = 0.687; p = 0.00). Multivariate analysis showed an association of ADMA ≥ 0.69 µmol/L (p = 0.000) with albuminuria ≥ 300 mg/g (p = 0.003). Conclusions: ADMA was correlated with the presence of macroalbuminuria, strongly indicating its role in the progression of CKD.

1. Introduction

Chronic kidney disease (CKD) represents a critical public health issue with its prevalence increasing globally. It puts a strain on healthcare systems worldwide in terms of expenses and resources. In the absence of timely therapeutic intervention, patients with CKD demonstrate accelerated deterioration of renal function, which adversely affects quality of life parameters and significantly increases morbidity and mortality risk, particularly from cardiovascular events [1]. Albuminuria constitutes a factor that determines the advancement of CKD [2,3]. Diseases related to the metabolism of minerals and bones are risk factors for the progression of CKD and cardiovascular consequences [4]. Phosphate retention and a decline in renal excretory function go hand in hand, causing the phosphaturic hormone fibroblast growth factor-23 (FGF-23) to be synthesized. This hormone is synthesized by osteocytes with the main stimulators other than phosphates, including 1,25-dihydroxy vitamin D (1,25(OH)2D3) and parathyroid hormone (PTH).

In the kidney, the function of FGF-23 is mediated by binding to receptor complexes (FGFRs) and its specific coreceptor, klotho. When FGF-23 cannot maintain phosphate homeostasis, a decrease in klotho expression and FGF-23 resistance occur. Lower 1,25(OH)2D3 levels are likely to result from elevated FGF-23 levels, which will ultimately lead to secondary hyperparathyroidism. This process of failure to compensate will lead to hyperphosphatemia [5]. A cohort study called the Prevention of Renal and Vascular End-stage Disease study found that the levels of FGF-23 were separately correlated with increased urine albumin-to-creatinine ratio (albuminuria) and reduced estimated glomerular filtration rate (eGFR), reflecting CKD progression and mortality risk [6]. Additionally, reduced concentrations of 25-hydroxyvitamin D (25(OH)D) were strongly correlated with increased albuminuria levels [7]. The inflammatory process is a potential mediator linking vitamin D deficiency with albuminuria [8].

Natural amino acid analogs of L-arginine include symmetric dimethylarginine (SDMA) and its enantiomer, asymmetric dimethylarginine (ADMA), which were first isolated from human urine by Kakimoto and Akazawa [9]. Increased ADMA levels occur from the early phase of CKD and will continue to increase as kidney function decreases [10]. Research has shown that nitric oxide (NO) synthesis is inhibited in CKD, and ADMA is a potent inhibitor of NO production in vivo and in vitro, thereby impacting vasoconstriction, hypertension, immune dysfunction, and the risk of cardiovascular complications [11]. The renal resistive index (RRI), measured by Doppler ultrasound, reflects intrarenal vascular resistance and compliance. High RRI values indicate increased resistance within renal blood vessels. Higher ADMA concentrations correlate independently with increased RRI (p < 0.001). This suggests that ADMA-mediated endothelial dysfunction and reduced vasodilation contribute to intrarenal vascular resistance, which the RRI measures noninvasively as an indicator of kidney microvascular injury and disease progression [12].

A cohort study has indicated that ADMA levels and their interaction with FGF-23 in NO system inhibition are predictors of CKD progression [13,14]. To date, reducing albuminuria has primarily involved renin–angiotensin–aldosterone system (RAAS) inhibitor therapy as the main choice. High FGF-23 levels reduce RAAS inhibitor effectiveness in albuminuria treatment associated with impaired antiproteinuric responses. The RAAS inhibitor helps reduce albuminuria related to ADMA-mediated vascular damage through significantly increasing urinary NO metabolite excretion and decreasing oxidative stress [15,16]. Vitamin D primarily acts as an inhibitor of intrarenal RAAS activation. Moreover, low vitamin D levels are thought to be linked to elevated ADMA levels and might impair RAAS regulation and worsen albuminuria in CKD patients [17]. However, renal function deterioration persists even with RAAS inhibitor therapy. Therefore, to identify potential therapeutic targets, further studies are needed regarding the pathophysiology underlying albuminuria and CKD progression [18].

This study aimed to analyze the correlation of FGF-23, 25(OH)D, and ADMA levels with albuminuria in patients with CKD not undergoing dialysis. Several previous studies have stated that the direct relationship of FGF-23, 25(OH)D, and ADMA levels with albuminuria independent of other risk factors for CKD remains controversial.

2. Material and Methods

2.1. Design

This cross-sectional observational analytical investigation was carried out on CKD patients with various disease stages (non-dialysis) in an outpatient setting of Airlangga University Hospital from May 2022 to August 2022. Informed consent was obtained from all participants before participating in this study.

2.2. Inclusion and Exclusion

The inclusion requirements were as follows: CKD patients with various disease stages (non-dialysis), aged > 21 years, who entered the community with normal creatinine values ranging between 0.5 and 0.9 mg/dL. The exclusion criteria were as follows: patients with CKD who had undergone dialysis, been hospitalized in the last 2 weeks, and shown signs of infections.

2.3. Sample Collection and Assay

Here, FGF-23, 25(OH)D, ADMA serum, and urinary albumin levels as risk factors for CKD progression were evaluated. Blood samples were taken from all patients during their follow-up appointment. Overall, 10 mL of blood was put into a serum separator tube and 20 mL of urine was put into a nonsterile urine container. Subsequently, blood and urine samples were sent to the laboratory at Airlangga University Hospital, Surabaya.

Serum levels of FGF-23 were assessed using the sandwich enzyme-linked immunosorbent assay (Bio-Rad Laboratories Inc., Hercules, CA, USA) with a 100 RU/mL cutoff value. FGF-23 elevation was operationally defined as concentrations ≥100 RU/mL, a cutoff value established at 2.5 times the median FGF-23 level and corresponding to the 90th percentile of FGF-23 distribution observed in prior research involving cohorts where approximately 80% of participants demonstrated preserved renal function (eGFR > 60 mL/min/1.73 m²) [19]. Utilizing chemiluminescent microparticle immunoassay (Architect 1000i; Abbott Diagnostic, Abbott Park, IL, USA), serum levels of 25(OH)D were determined with a 30 ng/mL cutoff value. ADMA levels were measured using ultrahigh-performance liquid chromatography (Agilent 6460 Triple Quad With 1290 UPLC; Agilent Technology, Santa Clara, CA, USA) with a 0.69 μmol/L cutoff value. An ADMA level above 0.69 μmol/L was associated with increased cardiovascular events and mortality risks [20]. Urinary albumin levels were measured using Cobas c-501 (Roche Diagnostics, Mannheim, Germany) with a 300 mg/g cutoff value.

eGFR was calculated using the CKD–EPI formula with the following five stages: CKD stage 1, ≥90 mL/min/1.73 m²; CKD stage 2, 60–89 mL/min/1.73 m²; CKD stage 3, 30–59 mL/min/1.73 m²; CKD stage 4, 15–20 mL/min/1.73 m²; and CKD stage 5, <15 mL/min/1.73 m².

2.4. Sample Size

The sample size in this study was calculated applying the correlation formula [21]:

N = [(Zα + Zβ)/C]² + 3 = 85 participants

N = number of participants.

α = type 1 error rate. The probability limit of the null hypothesis is rejected.

β = type 2 error rate. It is possible that the null hypothesis is rejected.

r = correlation coefficient.

C = A constant based on the expected correlation coefficient (r).

We recruited 85 participants to detect correlations of r ≥ 0.3 with a power of 80%.

2.5. Statistical Analysis

Data were processed using Statistical Product and Service Solution version 24.0 for Windows. The Kolmogorov–Smirnov test was used to determine whether the data was normal. To establish the relationship between each FGF-23, 25(OH)D, and ADMA level and albuminuria, the Pearson correlation test was used if the parametric test requirements were met; if not, the Spearman test was used. The chi-square test and multivariate logistic regression were used in the bivariate analysis to observe the relationship between clinical features as well as the relationship of FGF-23, 25(OH)D, and ADMA levels with albuminuria.

2.6. Ethical Clearance

The ethical board of Airlangga Hospital approved this study (certificate number: 049/KEP/2022, approval date 25 May 2022).

3. Results

3.1. Patients Characteristics

A total of 107 non-dialysis patients with CKD stages 1–4 at the Kidney and Internal Medicine Hypertension Polyclinic of Airlangga University Hospital, Surabaya, were involved in this study. No participants dropped out or died during the study period. In this study, 54.2% of the patients were male (n = 58), with 57.32 years as the average age. The average eGFR was 30 mL/min/1.73 m². The mean level of urinary albumin-to-creatinine ratio (albuminuria) was 940.92 mg/g, with the highest and lowest albuminuria values of 5849.50 and 2.29 mg/g, respectively. The mean FGF-23 level of the participants was 197.75 RU/mL, with the highest and lowest FGF-23 levels of 1054.70 and 21.59 RU/mL, respectively. Of the participants, the average vitamin D level was 23.44 ng/mL, with the highest and lowest vitamin D levels being 64.27 and 3.11 ng/mL, respectively. The average ADMA level was 719.63 µmol/L, with the highest and lowest ADMA levels being 4345.01 and 40.07 µmol/L, respectively (

Table 1. Participants’ characteristics.

Characteristics	Results
Age (years)	Median (min–max)	59 (32–70)
Gender
Male	% (n)	54.2% (n = 58)
Female	% (n)	45.8% (n = 49)
Blood pressure (mmHg)
Systole	Mean ± SD	145.59 ± 23.31
Diastole	Median (min–max)	81 (51−121)
Risk factor
Diabetes mellitus type 2	% (n)	72% (n = 77)
Hypertension	% (n)	86.9% (n = 93)
Smoking	% (n)	27.1% (n = 29)
Obesity	% (n)	16.8% (n = 18)
Renal function
Creatinine (mg/dL)	Median (min–max)	2.12 (1.15–11.82)
eGFR (mL/min/1.73 m2)	Median (min–max)	31 (4–68)
Albuminuria (mg/g)	Median (min–max)	289.7 (2.29–5849.50)
FGF-23 (RU/mL)	Median (min–max)	136 (21.59–1054.70)
Vitamin D (ng/mL)	Mean ± SD	23.44 ± 11.95
ADMA (µmol/L)	Median (min–max)	0.626 (0.040–4.345)
CKD stage
Stage 2	% (n)	4.7% (n = 5)
Stage 3	% (n)	47.7% (n = 51)
Stage 4	% (n)	27.1% (n = 29)
Stage 5	% (n)	20.6% (n = 22)

).

3.2. Correlation Between FGF-23, 25(OH)D, and ADMA Levels with the Degree of Albuminuria

In this study, the correlations between FGF-23, 25(OH)D, and ADMA levels and the degree of albuminuria were analyzed using Spearman’s rank correlation test, as these variables were not normally distributed. The results of the correlation test between FGF-23 levels and the degree of albuminuria showed a Spearman’s r-value of 0.252 with a p-value of 0.009 (<0.05), indicating a weak correlation. The results of the correlation test between the degree of albuminuria and 25(OH)D levels showed a Spearman’s r-value of −0.375 with a p-value of 0.000 (<0.05), indicating a weak correlation. Vitamin D concentrations show negative correlation with the degree of albuminuria, indicating that as the degree of albuminuria increases, the vitamin D levels subsequently decrease. The results of the correlation test between ADMA levels and the degree of albuminuria showed a Spearman’s r-value of 0.687 with a p-value of 0.000 (<0.05), demonstrating a strong association (

Table 2. Correlation of FGF-23, 25(OH)D, and ADMA level with the degree of albuminuria.

Variable 1	Variable 2	r Spearman	p-Value
FGF-23	Albuminuria	0.252	0.009
25(OH)D	Albuminuria	−0.375	0.000
ADMA	Albuminuria	0.687	0.000

).

3.3. Multivariate Analysis of Albuminuria with FGF-23, 25(OH)D, ADMA Levels Along with Concomitant Factors

We use multivariate regression analysis to determine the interactions between albuminuria and different variables, which included 25(OH)D, ADMA, age ≥ 60 years, gender, diabetes mellitus, and hypertension. Multivariate analysis showed an association of ADMA ≥ 0.69 µmol/L (p = 0.000), male gender (p = 0.041), diabetes mellitus (p = 0.031), and hypertension (p = 0.003) with albuminuria ≥ 300 mg/g (p = 0.003). In this study, patients with chronic kidney disease, ADMA and FGF-23 levels progressively increase with worsening CKD stages, while 25(OH)D levels demonstrate an inverse correlation with disease progression (Figure 1).

4. Discussion

Anomalies in kidney structure and function that last for >3 months and have clinical consequences are the hallmarks of CKD. In individuals with CKD, GFR and albuminuria are required to characterize the extent and course of kidney damage. The mechanism of CKD is highly dependent on the etiology, and the process is highly specific according to the underlying cause, that is, structural renal abnormalities, immune complex-mediated glomerulonephritis, or nephrotoxic injury to tubular and interstitial compartment. Following a prolonged decrease in kidney function, injured nephrons will be replaced by functional nephrons, thereby leading to hypertrophy and hyperfiltration of the remaining nephrons as an overall effect after a prolonged decrease in kidney mass, and causing a further decline in kidney function [22].

Clinically, albuminuria can indicate the risks of cardiovascular disease and mortality in addition to the development of CKD to end-stage renal disease (ESRD). A study conducted in South Korea determined the prevalence of microalbuminuria on the basis of some selected risk variables. The results showed that of 5202 participants (mean age, 45.6 years; 2337 males, 2865 females), 5.2% (95% CI: 4.4–6.1) had microalbuminuria. In addition to increasing the likelihood of albuminuria, one of the risk factors for CKD is renal injury. Some research studies have stated the relationship between the following factors and an increased risk of albuminuria: older age; gender (males are diagnosed with albuminuria at a higher rate than females); and Asian race (53.9%), African–American (18.2%), non-Hispanic Caucasian (13.8%), Hispanic (9.2%), and others (4.9%). Comorbidities, including diabetes and insulin resistance, dyslipidemia, obesity, hypertension, lack of physical activity, cardiovascular disease, and smoking, are risk factors for albuminuria in individuals with endothelial damage [23,24,25]. In the present study, male gender (p = 0.041), diabetes mellitus (p = 0.031), hypertension (p = 0.003), and CKD stage (p = 0.003) were associated with albuminuria ≥300 mg/g (p = 0.003) (

Table 3. Multivariate logistic regression analysis of albuminuria with FGF-23, vitamin D, and ADMA levels along with concomitant factors.

Variable	Albuminuria		p-Value Bivariate	OR (95% CI)	p-Value Multivariate	Adjusted OR (95% CI)
	≥300 mg/g	<300 mg/g
FGF-23 (RU/mL)
≥100	40	39	0.702	0.92 (0.58–1.44)
<100	13	15
25(OH)D (ng/mL)
≥30	14	28	0.007 *	1.67 (1.16–2.41)
<30	39	26
ADMA (µmol/L)
≥0.69	44	19	0.00 *	2.64 (1.76–3.95)	0.000 **	9.49 (3.18–28.25)
<0.69	9	35
Age (years)
≥60	15	31	0.002 *	2.03 (1.25–3.29)
<60	38	23
Gender
Male	24	34	0.066 *	1.39 (0.97–1.99)	0.041 **	3.13 (1.05–9.32)
Female	29	20
Diabetes mellitus
Yes	44	33	0.012 *	2.29 (1.16–4.53)	0.031 **	3.73 (1.12–12.38)
No	9	21
Hypertension
Yes	40	24	0.001 *	2.27 (1.33–3.84)	0.003 **	5.29 (1.75–15.99)
No	13	30
Obesity
Yes	10	8	0.56	1.05 (0.89–1.25)
No	43	46
Smoking
Yes	14	15	0.87	0.98 (0.78–1.24)
No	39	39

* p-value bivariate < 0.25 for next to multivariate analysis, ** p-value multivariate analysis < 0.05.

).

Bones secrete a hormone called FGF-23, which regulates the metabolism of phosphate and vitamin D balance. In mineral metabolism, FGF-23 plays a role in suppressing phosphate reabsorption in urine and suppressing the synthesis of 1α,25-dihydroxyvitamin D-3 [1,25(OH)2D3], the hormone of vitamin D in kidneys. In a normal kidney function, excess FGF-23 levels can result in renal phosphate-wasting syndrome and low 1,25(OH)2D3 levels in the circulation. In patients with CKD, circulating FGF-23 can reach 1000 times the normal limit. Elevated levels of FGF-23 in the early phases of CKD can help maintain normophosphatemia; however, in advanced CKD, serum phosphate levels increase despite FGF-23 levels having increased from an early age [26]. In this research, FGF-23 levels were weakly correlated with the degree of albuminuria (r = 0.252; p = 0.009). Cohort research conducted in Korea by Kim et al. reported that elevated levels of FGF-23 were found associated with proteinuria, indicating CKD progression [4]. However, a study by Drew et al. stated that in older adult patients, elevated FGF-23 levels were not always associated with the incidence of CKD or impaired renal function.

The roles of 1,25(OH)2D3 and FGF-23 extend beyond the metabolism of minerals. Both hormones are also closely associated with the inflammatory process. Several studies have suggested that the stimulation of FGF-23 expression by 1,25(OH)2D3 necessitates a signal on polymerized actin filaments, which are regulated by the proinflammatory transcription factor, ras-related C3 botulinum substrate 1 (Rac1), nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), and p21 protein-activated kinase (PAK1). Continuously increasing NF-κB activity increases the expression of the calcium channel protein Orai1 and its stimulator (STIM), enabling more calcium ions to enter the cell; this signaling cascade is triggered by store-operated Ca²⁺ entry (SOCE). In this manner, SOCE stimulation will increase FGF-23 production. NF-κB activity ultimately triggers the release of TGFβ, TNFα, and other proinflammatory cytokines, which is consistent with FGF-23 production. Meanwhile, 1,25(OH)2D3 not only functions in mineral metabolism but can also function as an anti-inflammatory hormone. Considering the effect of FGF-23 in suppressing 1,25(OH)2D3 production, cells will increasingly lose their anti-inflammatory effects while proinflammatory cytokines increase. This inflammatory process is believed to be one of the mechanisms by which increased FGF-23 levels trigger the worsening of CKD prognosis [27,28].

CKD and ESRD patients frequently have a lack of vitamin D, which is correlated with an albuminuria condition that is usually found in further kidney damage. Vitamin D administration in diabetic nephropathy patients can improve podocyte function, thereby reducing the risk of albuminuria. Another study has shown that vitamin D administration can suppress tissue fibrosis, inflammation, and apoptosis through various pathways, including the signaling pathways of RAAS, NF-κB, Wnt/β-catenin, and TGF-β/Smad [29]. A study involving experimental animals has reported that 25(OH)D, or calcidiol, suppresses RAAS activity, whereas low 25(OH)D levels can activate RAAS (in the present study, RAAS was measured on the basis of increased basal aldosterone levels, which is inversely proportional to 25(OH)D levels, thereby causing hyperfiltration; the impacts are quite detrimental [30]. In the current study, albuminuria and lower vitamin D levels were associated (r = −0.375; p = 0.000). Decreased levels of 25(OH)D levels are associated with albuminuria, in patients with diabetic nephropathy [31]. Isakova et al. stated that patients with albuminuria are more susceptible to decreased 25(OH)D levels and increased levels of inflammatory markers than those with albuminuria < 30 mg/g [9]. Echida et al. indicated that lack of 25(OH)D is frequently observed in pre-dialysis patients with CKD and is an accompanying risk factor for both obesity and diabetes mellitus [32].

The 1α-hydroxylase enzyme is regulated by calcium directly or via PTH and phosphate metabolism (directly or via FGF-23). Osteocytes produce FGF-23 stimulated by excess levels of vitamin D and phosphate. FGF-23, along with klotho, can reduce the expression of the CYP27b1 gene, which codes for 1α-hydroxylase synthesis, thereby leading to calcitriol synthesis inhibition, reduced phosphate absorption in the intestine, phosphate reabsorption in the renal tubules, and increased Cyp24 gene expression. Cyp24 encodes 24-hydroxylase to inactivate calcitriol, thereby leading to reduced klotho expression. The klotho–FGF-23 axis via CD4 directly influences the blood vessel wall and induces endothelial dysfunction [33,34].

There remains no evidence of a direct influence of RAAS on VDR or 1α-hydroxylase; however, Angiotensin II’s effects on 1α-hydroxylase inhibition through downregulating klotho expression and FGF-23 resistance are believed to contribute to kidney injury. In typical circumstances, 1,25(OH)2D3 suppresses renin production. When angiotensin II is low, klotho levels are adequate to maintain normal levels of FGF-23. The balance between RAAS, klotho, FGF-23, and vitamin D in CKD is disturbed. When nephron damage results from decreased 1α-hydroxylase activity, FGF-23 levels are increased in the kidneys, thereby reducing 1,25(OH)2D3 production. When renin and angiotensin production increases, klotho production is inhibited, and FGF-23 upregulation occurs. CKD is worsened by RAAS activation, lack of vitamin D, elevated levels of FGF-23, and low levels of klotho. A study on patients undergoing hemodialysis exhibited elevated levels of 1,25-dihydroxivitamin D following vitamin D supplementation. In the 158 patients undergoing hemodialysis who were administered cholecalciferol, albumin, 25(OH)D, and 1,25-dihydroxyvitamin D levels increased, whereas serum levels of brain natriuretic peptide, calcium, PTH, active vitamin D, erythropoietin-stimulating agents, and the left ventricular mass index decreased [35]. Vitamin D activation can inhibit TGF-β activation and reduce proinflammatory cytokine expression, including TNF, IL-8, and IL-6, in podocytes and renal tubular cells. Some studies are being conducted with small sample sizes; therefore, they require further clinical trials [33].

ADMA and SDMA are potent NO synthase inhibitors and are excreted in the urine. Patients with CKD undergoing hemodialysis have higher serum ADMA and SDMA levels than those with CKD who are not receiving hemodialysis. Additionally, increasing serum ADMA and SDMA levels increases the levels of oxidative stress markers, including serum malondialdehyde, as well as the urinary excretion of 8-Oxo-2′-deoxy guanosine [10,36]. Increased ADMA levels in CKD demonstrate the significant role of the kidneys in ADMA metabolism, particularly in the ADMA elimination process, which involves dimethylarginine–dimethylamino–hydrolase (DDAH), a catalyst in ADMA metabolism. Studies conducted on diabetic mice with streptozotocin-induced tubulointerstitial ischemia revealed that DDAH overexpression resulted in decreased ADMA levels, decreased urinary albumin excretion, and decreased tubulointerstitial hypoxia. Tubular DDAH activity can be inactivated by oxidative stress due to proteinuria, thereby leading to increased ADMA levels along with worsening of CKD severity and albuminuria [37,38,39]. Our study found that ADMA levels were associated with the degree of albuminuria (r = 0.687; p = 0.000). Research by Oliva-Damaso et al. stated that ADMA is a risk marker for mortality for cardiovascular disease in patients with CKD and is related to high ADMA levels accompanied by high proteinuria. This process occurs owing to glomerular filtration leakage, which causes impaired ADMA elimination [10]. In CKD patients, there is a 37% higher chance of mortality for every increase of 0.1 μmol/L in plasma ADMA levels. ADMA levels were associated with low eGFR levels and albuminuria in patients with stages 3 and 4 of CKD [40].

Our study provides preliminary evidence supporting ADMA as a biomarker for disease progression in the early stage of CKD. To our knowledge, our study was the first to investigate the associations between ADMA, FGF-23, and 25(OH)D with albuminuria in an adequate sample size and relatively homogeneous study population in Indonesia that minimized major confounding factors for these biomarker measurements. To determine ADMA as a novel biomarker in CKD progression is a new challenge; thus further research is warranted.

5. Conclusions

Our findings demonstrate that ADMA with the presence of additional risk factors, holds a significant role in the progression of CKD. This conclusion is supported by a strong positive association between ADMA and albuminuria, which positions ADMA as a potential early biomarker of declining renal function.