Association between mineral and bone disorder in patients with acute kidney injury following cardiac surgery and adverse outcomes

We conducted a single-center, prospective cohort study and enrolled patients who underwent cardiac surgery and developed incident AKI at a comprehensive hospital between June 2014 and January 2016. This study was approved by the hospital ethics committee of Shanghai Ninth People’s Hospital, School of Medicine, Shanghai Jiaotong University (approval number: [2014]45). All of the experimental protocols were implemented in accordance with the relevant guidelines and regulations. In accordance with the Declaration of Helsinki, patients or their legal guardians had obtained written informed consent prior to participation,. This study was registered at www.​clinicaltrials.​gov with the identification number NCT00953992.

The inclusion criteria were an age > 18 years and the development of incident AKI following cardiac surgery. AKI was defined as an increase in serum creatinine of ≥0.3 mg/dL within 48 h or that of ≥50% within 7 days, which conforme with the criteria established by the Kidney Disease Improving Global Outcomes (KDIGO) work group [17].

The exclusion criteria were: (1) preoperative AKI (defined as a 0.3 mg/dL rise in the serum creatinine level over 24 h or a 0.5 mg/dL rise over 48 h) [18], (2) pre-existing CKD (defined as an estimated glomerular filtration rate (eGFR) of < 30 mL/min per 1.73 m²) or ESRD requiring dialysis, (3) post-renal obstruction or rapid progressive glomerulonephritis as the main cause of AKI, (4) renal transplantation, (5) malignancy, and (6) pregnancy.

The serum creatinine data of the participants were monitored daily in the hospital’s information system (HIS) to rapidly identify patients with incident AKI. We collected and stored the serum samples within 12 h after establishing the clinical diagnosis. The samples were centrifuged, aliquoted, and stored at − 80 °C within 2 h of collection.

We adjudicated all of the outcomes by reviewing the electronic medical records from the HIS. The pre-specified primary endpoint was the 28-day all-cause mortality.

The secondary endpoints were Renal Replacement Therapy or in-hospital mortality (RRT/death). Additional endpoints included the ventilator-free days and hospital-free days in 28 days. The patients who died before 28 days were graded a score of zero.

The bone-specific alkaline phosphatase and tartrate-resistant acid phosphatase 5b levels were measured using a standard ELISA kit (Immunodiagnostic Systems Limited, UK). The C-terminal FGF-23 levels were measured using a standard ELISA kit (Immutopics Inc., USA). The serum calcium and phosphate levels were measured on an automated chemistry analyzer (AU5800, Beckman Coulter, USA). The iPTH level was measured on an automated immunoassay analyzer (UniCel DxI800, Beckman Coulter, USA). The 25-hydroxyvitamin D level was measured on an automated immunoassay analyzer (LIAISON, DiaSorin, Italy).

The data are reported as the median and interquartile range (IQR: 25th percentile–75th percentile) for continuous variables and as the n (%) for categorical variables. The baseline and operative characteristics were compared in patients with different stages of AKI using the Kruskal-Wallis test for continuous variables with a non-normal distribution and using the chi-square test for category variables. The correlation between the enrollment mineral metabolite levels and the other variables was analyzed using the Spearman rank correlation coefficient. The comparison of the mineral metabolite levels among the patients who died and those of the patients who did not die was assessed using the Mann-Whitney U test.

Univariate Cox regression was used to identify potential confounding variables, including demographics (sex and age), preoperative renal function (serum creatintine and eGFR), comorbidities (hypertension, chronic heart failure and diabetes mellitus), operative characteristics (operation type, whether cardiopulmonary bypass or not), postoperative characteristics (low blood pressure, applying diuretics), Acute Physiology and Chronic Health Evaluation (APACHE) II score and biochemical parameters. After that, we used multivariate Cox regression to adjust for covariates using two different models: model 1 was adjusted for age, sex, baseline eGFR, hypertension, chronic heart failure (New York Heart Association functional class [NYHA] ≥ 2) and diabetes mellitus; model 2 was further adjusted for the operation type and APACHE II score on enrollment, according to P value in the univariable regression (P < 0.1) or clinical importance. Logistic and linear regression models were used to assess the association between the mineral metabolite levels and secondary endpoints, adjusting for the same covariates as above. Receiver-operating characteristics (ROC) curves and areas under the curves (AUCs) were calculated using the survival and time ROC packages to compare the predictability of the mineral and bone metabolite levels for the 28-day mortality, Furthermore, the C-index was calculated using the survival package. The evaluation criterion of the C-index was similar to that of the AUC.

We used linear mixed models for the repeated measures to test for significant differences in the mineral and bone metabolite levels over time between the patients with different severities of AKI. The comparison of the mineral and bone metabolite levels at individual time points was assessed using the Spearman rank correlation coefficient (for the correlation between the severity of AKI and the mineral and bone metabolite levels).

All of the comparisons were two-tailed, with P < 0.05 considered as being statistically significant. The statistical analysis was performed using the software package R version 3.4.0 (www.​r-project.​org).

We enrolled 158 patients who developed incident AKI following cardiac surgery but had no evidence of AKI at baseline into a prospective cohort study.

Among the 158 patients enrolled, 57 (36.1%) had AKI stage 1, 53 (33.5%) had AKI stage 2, and 48 (30.4%) had AKI stage 3. The median (interquartile range [IQR]) age of the patients was 57 years (49–66 years), and 74.7% of them were men. The most common comorbidities were congestive heart failure [NYHA≥2] (62.0%), hypertension (53.8%), and diabetes mellitus (10.1%). The median (IQR) APACHE II score was 11 (9–13). The additional baseline characteristics are shown in Table 1.

Among these three groups, the patients with aortic dissection and those with low blood pressure after the operation presented with higher ratios of severe kidney injury (P = 0.002 and P < 0.001, respectively). Meanwhile, the ratios of AKI stage 3 were lowest in the patients with coronary artery bypass alone (P = 0.009). Moreover, patients with more severe kidney injury had higher scores of the APACHE II (P = 0.007). However, there were no significant differences among three groups in the demographics, preoperative renal function, comorbidities, cardiopulmonary bypass during the operation, and applying diuretics after the operation (P > 0.05).

The serum iPTH level was inversely correlated with the corrected calcium level (rs = − 0.379, P < 0.001) and directly correlated with the cFGF23 level (rs =0.330, P < 0.001). The serum TRACP-5b level was directly correlated with the 25D level (rs =0.189, P < 0.05) and BAP level (rs =0.179, P < 0.05). The additional correlations are shown in Additional file 1: Table S1.

Among the 158 patients enrolled in this study, 18 (18.8%) had died within 28 days, and the median time from the clinical diagnosis to death was 7 days (IQR: 2–17 days). The causes of death included multiple organ dysfunction syndromes (n = 7), heart failure (n = 4), adult respiratory distress syndrome (n = 3), acute cerebral infarction (n = 2), cute gangrenous cholecystitis (n = 1) and circulatory failure of unknown disease (n = 1). The serum mineral metabolite levels in patients who died and in those who did not die are shown in Table 2. The patients who died had higher levels of serum cFGF23 (P = 0.007), as well as lower levels of 25D (slightly insignificant, P = 0.072), compared with the patients who were alive. The levels of the other mineral metabolites were similar between the two groups (Table 2).

Then, we analyzed the association between each mineral metabolite biomarker and the risk of death after AKI. Table 3 shows the unadjusted and adjusted hazard ratios for the 28-day mortality risk according to the levels of the mineral metabolites. In the unadjusted analyses, the serum iPTH and cFGF23 levels were both significantly associated with the 28-day mortality (iPTH: HR = 1.045, 95% CI: 1.010 to 1.080, p = 0.009; cFGF23: HR = 1.258, 95% CI: 1.149 to 1.378, p < 0.001). These associations were slightly attenuated but remained significant when the model was adjusted for age, sex, preoperative eGFR, hypertension, congestive heart failure, and diabetes mellitus (Model 1), as well as when it was further adjusted for operation type and APACHE II score (Model 2; iPTH: HR = 1.044, 95% CI: 1.001 to 1.090, p = 0.046; cFGF23: HR = 1.367, 95% CI: 1.168 to 1.599, p < 0.001). The serum phosphate level was also associated with the 28-day mortality in the adjusted analyses (HR = 2.620, 95% CI: 1.083 to 6.338, p = 0.035). The other biomarkers, including the corrected calcium, 25D, BAP, and TRACP-5b, were not associated with the 28-day mortality. Additionally, Table 4 shows the area under the receiver operating characteristic for the 28-day mortality risk according to the levels of the mineral metabolites. The strengths of the AUC appeared to be greater for cFGF23 than for the other mineral biomarkers that were tested (P = 0.007).

In an exploratory analysis, we evaluated whether the phosphate, iPTH, and cFGF23 levels remained associated with 28-day mortality after the adjustment for each. As shown in Additional file 1: Table S2, only the cFGF23 level, and not the iPTH or phosphate levels, remained as being significantly associated with the 28-day mortality even after the adjustment for each (HR = 1.380, 95% CI: 1.162 to 1.639, p < 0.001).

Table 5 shows the adjusted logistic and linear regression models of the secondary endpoints according to the levels of the mineral metabolites. The serum cFGF23 levels were also associated with the following secondary endpoint in the adjusted analyses: RRT/death (OR = 1.469, 95% CI: 1.055 to 2.045, p = 0.029). Additionally, the serum cFGF23 and BAP levels were inversely associated with the hospital-free days and ventilator-free days, implying that they were correlated with longer durations of hospitalization and mechanical ventilation (hospital-free days: cFGF23, β [SEM]: − 0.951 [0.264], p < 0.001; BAP, β [SEM]: − 0.299 [0.087], p < 0.001; ventilator-free days: cFGF23, β [SEM]: − 1.681 [0.365], p < 0.001; BAP, β [SEM]: − 0.268 [0.126], p = 0.027).

To compare the time course of changes in the mineral and bone metabolite biomarkers in patients with AKI after cardiosurgery, we also measured the levels in a subgroup of 71 patients, and these results are shown in Fig. 1. We found that there was a hierarchical association between the AKI severity and the levels of cFGF23 over time (P < 0.001). Among the AKI stage 3 patients, we found rapid and significant increases in the cFGF23 levels like that the median levels at enrollment were ~ 18-fold higher than the baseline (preoperative) levels (P < 0.001) and the Day 5 levels were ~ 14-fold higher (P < 0.001) than the baseline levels. Among the AKI stage 1 patients, the median cFGF23 levels increased by ~ 4.5-fold at the time of enrollment and by ~ 1.6-fold on Day 5. The AKI stage 2 group of patientshad medium rises in the cFGF23 levels that were between those of the AKI stage 1 group and those of the AKI stage 3 group. The serum creatinine and iPTH levels showed a similar hierarchical association, which higher levels were associated with a greater severity of AKI. Additionally, when the phosphate and 25D levels were evaluated at individual time points, only the Day 5 levels were significantly different between the groups. The longitudinal calcium levels, as well as the BAP and TRACP-5b levels, were similar in these three groups.