Background
Methods
Blood, plasma and urine sampling
DNA isolation and qPCR
Leukocyte mitochondrial DNA copy number
Quantification of urine and plasma cell-free DNA by real-time PCR
Plasma neutrophil gelatinase-associated lipocalin (NGAL)
Statistical analysis
Results
The demographic and clinical characteristics of the varying stages of chronic kidney diseases
Variables | Stage 1,2a | Stage 3Ab | Stage 3Bc | Stage 4,5d | p value | Post hoc |
---|---|---|---|---|---|---|
n = 23 | n = 28 | n = 31 | n = 42 | |||
Age (year) | 47.1 ± 12.7 | 53.6 ± 9.4 | 54.9 ± 10.5 | 58.7 ± 6.1 | 0.002* | a < c,d |
Gender (M/F) | 3/20 | 8/20 | 22/9 | 29/13 | 0.212 | |
Hypertension (%) | 42% | 37% | 59% | 54% | 0.191 | |
Diabetes mellitus (%) | 12% | 11% | 23% | 54% | 0.160 | |
Systolic blood pressure (mm Hg) | 138 ± 18 | 144 ± 28 | 138 ± 24 | 142 ± 15 | 0.943 | |
Diastolic blood pressure (mm Hg) | 90 ± 15 | 91 ± 4 | 84 ± 18 | 84 ± 12 | 0.784 | |
BMI (kg/m2) | 27.8 ± 5.4 | 28.2 ± 6.1 | 26.2 ± 6.1 | 24.8 ± 4.3 | 0.172 | |
BUN (mg/dL) | 14.3 ± 4.4 | 17.7 ± 3.8 | 31.4 ± 13.0 | 64.5 ± 27.1 | <0.001** | a,b < c < d |
eGFR (ml/min/1.73 m^2) | 84.4 ± 20.2 | 52.1 ± 4.4 | 31 ± 9.6 | 11.5 ± 7.3 | <0.001** | a > b > c > d |
Phosphate (mg/dL) | 3.5 ± 0.4 | 3.5 ± 0.6 | 4.0 ± 0.7 | 5.0 ± 1.1 | <0.001** | a,b,c < d |
Cholesterol (mg/dL) | 198.3 ± 50.4 | 171.5 ± 31.8 | 196.6 ± 47.3 | 153.3 ± 44.5 | 0.001** | a,c > d |
HDL-C (mg/dL) | 47.8 ± 14.3 | 43.3 ± 9.8 | 46.1 ± 12.6 | 46.7 ± 21.2 | 0.677 | |
LDL-C (mg/dL) | 118.2 ± 45.3 | 104.6 ± 31.4 | 114.9 ± 41.3 | 80.8 ± 26.7 | 0.040* | c > d |
Fasting Glucose (mg/dL) | 118.3 ± 46.1 | 106 ± 21.8 | 112.6 ± 43.5 | 94.9 ± 24.6 | 0.263 | |
HbA1C (%) | 6.3 ± 1.4 | 6.0 ± 0.6 | 6.2 ± 1.1 | 6.1 ± 0.9 | 0.778 | |
Urine A/C ratio (mg/g) | 438.0 ± 826.4 | 535.7 ± 1018.4 | 2121.3 ± 2540.7 | 1548.0 ± 2377.2 | 0.058 | |
WBC (×103/μL) | 7.1 ± 2.4 | 6.2 ± 1.9 | 6.4 ± 2.1 | 6.8 ± 2.0 | 0.323 | |
Uric Acid (mg/dL) | 7.1 ± 1.2 | 7.0 ± 1.4 | 7.5 ± 2.6 | 7.0 ± 1.2 | 0.629 | |
Platelet(×103/μL) | 230.2 ± 72.6 | 209.2 ± 59.5 | 201.9 ± 51.1 | 195.1 ± 58.2 | 0.159 |
The levels of cell-free mitochondrial (cf-mtDNA) and nuclear deoxyribonucleic acid (cf-nDNA), both urine and plasma, in varying stages of CKD patients
Stage 1,2 a | Stage 3 b | Stage 3 c | Stage 4,5 d | P value | Post hoc tests | |
---|---|---|---|---|---|---|
Plasma cf-mtDNA (GE/mL) | 523 ± 710 | 373 ± 360 | 420 ± 664 | 112 ± 134 | 0.209 | |
Plasma cf-nDNA (GE/mL) | 1269 ± 1195 | 698 ± 527 | 439 ± 569 | 104 ± 70.01 | <0.001** | a > b,c,d |
Urine cf-mtDNA (GE/mL) | 0.43 ± 0.69 | 0.98 ± 3.10 | 2.78 ± 10.12 | 6.49 ± 17.85 | 0.220 | |
Urine cf-nDNA (GE/mL) | 2.83 ± 4.60 | 4.49 ± 12.09 | 15.39 ± 44.01 | 8.01 ± 12.08 | 0.351 | |
Plasma NGAL (ng/mL) | 394 ± 175 | 489.6 ± 174.9 | 744.2 ± 272.1 | 1096 ± 488.7 | <0.001** | a,b,<c,d |
MCN (per cell) | 90.7 ± 66.9 | 94.9 ± 77.01 | 92.9 ± 89.9 | 110.5 ± 112.7 | 0.901 | |
Urine 8-OH dG/Cr | 3.73 ± 1.81 | 3.78 ± 2.93 | 4.15 ± 2.65 | 2.99 ± 1.53 | 0.416 |
The difference of baseline parameters between different (favorable vs. unfavorable) renal outcome at 6 months (Table 3)
Favorable renal outcome (n = 53) | Unfavorable renal outcome (n = 70) | P value | |
---|---|---|---|
Plasma cf-mtDNA (GE/mL) | 441.1 ± 525.6 | 370.6 ± 641.9 | 0.538 |
Plasma cf-nDNA (GE/mL) | 760.9 ± 732.9 | 554.1 ± 850.9 | 0.184 |
Urine cf-mtDNA (GE/mL) | 0.489 ± 0.751 | 3.745 ± 12.197 | 0.027* |
Urine cf-nDNA (GE/mL) | 1.764 ± 2.388 | 15.119 ± 40.791 | 0.009* |
MCN (per cell) | 87.21 ± 69.40 | 101.55 ± 95.75 | 0.356 |
Urine 8-OH dG/Cr | 3.62 ± 2.44 | 4.03 ± 2.44 | 0.346 |
NGAL (ng/mL) | 585.24 ± 251.62 | 780.85 ± 391.39 | 0.032* |
The correlation between urinary cf-nDNA, cf-mtDNA and variables
Variable | Rho correlation coefficient | P value |
---|---|---|
Urine A/C ratio (mg/g) | 0.408 | 0.005* |
Urine P/C ratio (mg/g) | 0.314 | 0.007* |
Urine cf-mtDNA (GE/mL) | 0.448 | <0.001** |
Urine protein (mg/dL) | 0.246 | 0.029* |
NGAL (ng/mL) | 0.322 | 0.001** |
Variable | Rho correlation coefficient | P value |
---|---|---|
Urine A/C ratio (mg/g) | 0.375 | 0.008* |
Urine P/C ratio (mg/g) | 0.349 | 0.002** |
Urine cf-nDNA (GE/mL) | 0.448 | <0.001** |
Urine protein (mg/dL) | 0.167 | 0.125 |
NGAL (ng/mL) | 0.168 | 0.066 |
Predictors | Unstandardized Coefficients | Standardized Coefficients | p value | |
---|---|---|---|---|
B | Std. Error | Beta | ||
(Constant) | −14.839 | 24.831 | 0.576 | |
NGAL | −0.017 | 0.024 | −0.071 | 0.502 |
Urine protein | −0.144 | 0.150 | −0.463 | 0.380 |
Urine A/C ratio | −0.074 | 0.019 | −2.214 | 0.013* |
Urine P/C ratio | 0.085 | 0.014 | 3.526 | 0.002** |
Predictors | Unstandardized Coefficients | Standardized Coefficients | p value | |
---|---|---|---|---|
B | Std. Error | Beta | ||
(Constant) | −1.790 | 2.295 | 0.471 | |
NGAL | −0.001 | 0.002 | −0.049 | 0.597 |
Urine protein | −0.018 | 0.014 | −0.537 | 0.261 |
Urine A/C ratio | −0.007 | 0.002 | −1.867 | 0.015* |
Urine P/C ratio | 0.008 | 0.001 | 3.277 | 0.002** |
Evaluation of urine cf-mtDNA, and urine cf-nDNA levels as predictors of CKD patient outcomes after 6 months
B | S.E. | P value | Odds ratio | 95% C.I. | ||
---|---|---|---|---|---|---|
Module 1 | ||||||
Baseline eGFR | .010 | .011 | .377 | 1.010 | .988 | 1.031 |
Gender (male) | .044 | .494 | .929 | 1.045 | .396 | 2.753 |
Age | .023 | .020 | .249 | 1.023 | 0.984 | 1.063 |
NGAL | .001 | .001 | .152 | 1.001 | 1.000 | 1.003 |
Urine cf-mtDNA | .596 | .266 | .025* | 1.815 | 1.076 | 3.059 |
Constant | −2.543 | 1.632 | .119 | .079 | ||
Module 2 | ||||||
Baseline eGFR | .027 | .014 | .061 | 1.027 | .999 | 1.056 |
Gender (male) | .278 | .587 | .636 | 1.320 | .417 | 4.175 |
Age | .054 | .024 | .027* | 1.056 | 1.006 | 1.107 |
NGAL | .002 | .001 | .110 | 1.002 | 1.000 | 1.004 |
Urine cf-nDNA | .255 | .102 | .012* | 1.290 | 1.057 | 1.575 |
Constant | −5.688 | 2.182 | .009 | .003 |