**2. Results**

#### *2.1. Characteristics of the Study Population*

The study included 100 KTRs, all Caucasian, 55 men and 45 women. The mean age was 55 ± 11 years (range 19 to 79 years). The average time from transplantation was 10 ± 7 years (range 2 to 28 years). The cohort included in the analysis had chronic kidney disease of transplanted kidney (CKD-T) of all five stages, including patients just before starting renal replacement therapy. The data presenting parameters of GFR are shown in Figure 1.

For investigation of uromodulin, the study included also 15 patients with non-kidney diseases, all Caucasian, 7 men and 8 women. The mean age was 43 ± 13 years (range 20 to 58 years).

**Figure 1.** Boxplots for each parameter of glomerular filtration rate. Boxplots for serum creatinine concentration (s-Creatinine), serum cystatin C concentration (s-CysC), serum urea concentration (s-Urea), serum uromodulin concentration (s-Uromodulin), estimated glomerular filtration rate with Chronic Kidney Disease Epidemiology Collaboration study formula with s-Creatinine (eGFR CKD EPI creatinine), estimated glomerular filtration rate with Chronic Kidney Disease Epidemiology Collaboration study formula with s-CyC (eGFR CKD EPI CysC), estimated glomerular filtration rate with Chronic Kidney Disease Epidemiology Collaboration study formula with s-Creatinine and s-CysC (eGFR CKD EPI creatinine CysC), and measured glomerular filtration rate with Chromium-51-ethylenediaminetetraacetic acid clearance (mGFR 51CrEDTA).

#### *2.2. Relation between Kidney Function Parameters and s-Uromodulin*

In the control group, the mean level of s-Uromodulin was 291 ± 71 ng/mL. S-Uromodulin levels decreased significantly in the group of KTRs (*p* < 0.001), in which the mean s-Uromodulin was 74 ± 53 ng/mL. We further analyzed s-Uromodulin in different stages of CKD-T (1–5, based on measured GFR with 51CrEDTA (mGFR 51CrEDTA)), in relation to the s-Uromodulin in the control group. We found that already in stage 1 of CKD-T, the s-Uromodulin significantly dropped compared to the control group (*p* = 0.013). In the other four stages (2–4), the significance of reduced s-Uromodulin compared to that in the control group was even lower (*p* < 0.001). Between CKD-T stage 1 and 2, the drop of s-Uromodulin showed borderline significance (*p* = 0.067), while s-Uromodulin was significantly lower in CKD-T stage 2 compared to CKD-T stage 3 (*p* < 0.05) and in CKD-T stage 3 compared to CKD-T stage 4 (*p* < 0.05). There were no significant differences between stages CKD-T 4 and 5 (Figure 2A).

According to receiver operating characteristics (ROC), the area under the curve (AUC) represents the probability that a randomly selected patient will have a lower or higher test result than a randomly selected control. The ROC curve of s-Uromodulin in KTRs demonstrated an AUC of 0.991 (SE-0.007) (95% Cl 0.977–1.00, *p* < 0.001) at an optimal cut-off of 191.5 ng/mL with 97% sensitivity and 100% specificity (Figure 2B).

Analyzing bivariate correlations using Spearman's correlation coefficient, s-Uromodulin was significantly associated with all parameters of kidney graft function. The highest correlation coefficient was noted for estimated glomerular filtration rate (eGFR) with Chronic Kidney Disease Epidemiology Collaboration study formula with serum creatinine concentration (s-Creatinine) and serum cystatin C concentration (s-CysC) (eGFR CKD EPI creatinine CysC) (Rho = 0.758, *p* < 0.001), followed by serum urea concentration (s-Urea) (Rho = −0.740, *p* < 0.001), eGFR with Chronic Kidney Disease Epidemiology Collaboration study formula with s-Creatinine (eGFR CKD EPI creatinine) (Rho = −0.736, *p* < 0.001), s-CysC (Rho = −0.720, *p* < 0.001), eGFR with Chronic Kidney Disease Epidemiology Collaboration study formula with s-CysC (eGFR CKD EPI CysC) (Rho = 0.718, *p* < 0.001), s-Creatinine (Rho = −0.698, *p* < 0.001), and mGFR 51CrEDTA (Rho = 0.669, *p* < 0.001) (Table 1).

**Figure 2.** Uromodulin concentration in serum. (**A**) The boxplots for serum uromodulin concentration (s-Uromodulin) in chronic kidney disease of transplanted kidney (CKD-T) stages 1–5 based on measured GFR with 51CrEDTA (mGFR 51CrEDTA) (stage 1, *n* = 5; stage 2, *n* = 21; stage 3, *n* = 43; stage 4, *n* = 17; and stage 5, *n* = 11, with mGFR 51CrEDTA not possible to perform for 3 patients) in comparison to the control group (Ctrl); (**B**) ROC curve and AUC distinguishing CKD-T from the control group without any renal diseases. Legend: \* *p* < 0.05, \*\*\* *p* < 0.001 (Ctrl versus Stage 1–5).

**Table 1.** Bivariate correlations between serum uromodulin concentration (s-Uromodulin) and parameters of kidney graft function: serum creatinine concentration (s-Creatinine), serum cystatin C concentration (s-CysC), serum urea concentration (s-Urea), estimated glomerular filtration rate with Chronic Kidney Disease Epidemiology Collaboration study formula with s-Creatinine (eGFR CKD EPI creatinine), estimated glomerular filtration rate with Chronic Kidney Disease Epidemiology Collaboration study formula with s-CysC (eGFR CKD EPI CysC), estimated glomerular filtration rate with Chronic Kidney Disease Epidemiology Collaboration study formula with s-Creatinine and s-CysC (eGFR CKD EPI creatinine CysC) and measured GFR with Chromium-51-ethylenediaminetetraacetic acid (mGFR 51CrEDTA).

