Artículo Original / Original Article Calcium and phosphorus parameters and their association with serum parathormone in chronic kidney patients on hemodialysis

The objective of this study was to investigate the association between calcium and phosphorus parameters with serum parathormone concentrations in patients with Chronic Kidney Disease (CKD) undergoing hemodialysis. It is descriptive quantitative cross-sectional study. The sample was composed of 50 individuals distributed in: group with up to 5 years of hemodialysis and group with more than 5 years of hemodialysis. Food consumption was assessed using 24-hour dietary recall. Descriptive analysis of quantitative variables was presented as mean and standard deviation. Data analysis was performed using the SPSS program; Student “T” test, Fisher’s exact test and Pearson’s chi-square test were used, with significance level p <0.05. The groups presented adequate serum calcium concentrations and high concentrations of phosphorus and parathormone. Patients with more than 5 years of hemodialysis had a statistically higher serum parathormone level (p= 0.034); there was an association between longer hemodialysis time and higher serum phosphorus concentrations (p= 0.039). There was a moderate positive correlation between serum parathormone and phosphorus in the group with up to 5 years of hemodialysis (p= 0.012). It was concluded that the progression of CKD increases serum phosphorus and parathormone concentrations, reflecting the pathophysiological events and altered metabolic demand inherent to the pathology.


INTRODUCTION
Chronic Kidney Disease (CKD) develops when the kidneys lose the ability to perform its functions, with a progressive functional decline, culminating in the accumulation of urea and nitrogenous toxins in the blood 1 . In the last decade, the number of CKD cases has increased in the world and in Brazil, due to the demographic transition and the aging of the population 2 . CKD affects 5 to 10% of the global population 3 . In Piauí 1,674 patients are on chronic dialysis, with a prevalence of 521 cases per million of the population 4 .
As patients are diagnosed with renal failure there is a gradual loss of glomerular function, with complications such as metabolic acidosis, malnutrition, anemia and changes in the metabolism and serum profile of calcium and phosphorus, or even death 5 . Changes in micronutrient profiles in CKD can be caused by numerous situations, such as: dietary restrictions, use of diuretics, renal replacement therapy and absorptive disorders 6 .
The reduction in urinary phosphorus excretion induces an increase in the concentration of serum phosphorus, with a consequent increase in serum levels of parathormone (PTH), bone mobilization and hypocalcemia 3 . This research aims to verify the existence of an association between calcium and phosphorus with serum concentrations of PTH, according to the prolongation of hemodialysis (HD).

METHODS
The study is descriptive, quantitative and cross-sectional among patients with CKD on HD. The sample size was calculated considering a 95% confidence interval, a margin of error of 5% and a prevalence of 0.048% of HD patients in the state of Piauí in 2016, reaching a sample size of 50. The selection of patients took place by simple selection.
The inclusion criteria were: patients of both sexes, clinically stable, who had been on HD for at least 6 months, aged over 18 years; non-smokers, who do not use alcohol in a chronic way; without severe liver dysfunction (jaundice or cirrhosis); no recent infections (less than three months ago), cancer, tuberculosis, acquired immunodeficiency syndrome, chronic obstructive pulmonary disease, previously diagnosed cardiovascular disease, cerebrovascular disease and symptomatic heart failure. The established exclusion criteria: pregnant women, nursing mothers and patients in situations that make anthropometric assessment impossible, such as: advanced bone disease, stroke sequelae, people with physical disabilities or amputations.
Age and HD time were extracted from medical records. The chemiluminescence method (Liaison XL) was used to detect PTH. Serum calcium and phosphorus were determined by dry chemistry (Vitros 4600). Weight and height were measured after HD session; body mass index (BMI) was derived from the ratio between weight and height squared, according to the classification of the World Health Organization 7 .
The reference values used as the cutoff point for laboratory data: serum calcium (8.4 to 10.2 mg / dL), serum phosphorus (2.5 to 4.5 mg/dL) and PTH (≤110 pg/mL) 8 .
Food consumption was assessed using 24-hour dietary recall (R24h), applied for three non-consecutive days, with two alternating days during the week and one weekend (one day on HD, one without HD and one day on the weekend). When completing forms, participants were instructed on the correct way to discriminate meal types, preparations, consumption times, and portion size with the help of a photo album of portions and quantities.
The consumption of energy, macronutrients, calcium and phosphorus were calculated using the NutWin ® Software, version 1.5 of the Department of Health Informatics at the Federal University of São Paulo. Nutritional information of foods and preparations consumed by the participants were extracted from the Brazilian Food Composition Table 9 and from the Food Composition Table of the Brazilian Institute of Geography and Statistics 10 , and added to the program's database.
As for minerals, the Food Composition Table of the Brazilian Institute of Geography and Statistics 10 and the United States Department of Agriculture's National Nutrient Database for Standard Reference were used. Home measurements were converted to grams or milliliters 11,12 .
After analyzing the 3 days of the R24h, the distribution of macronutrients and minerals (calcium and phosphorus) was found to be normal, which were adjusted for intrapersonal and interpersonal variability, avoiding to changes generated by differences in energy consumption 18,19,20 . First, the data for this research was organized in Excel ® spreadsheets. Next, data were transferred to SPSS program (Windows ® version 22.0) for statistical analysis of the results. The descriptive analysis of quantitative variables was presented as mean and standard deviation of the mean.
To verify normality, the Kolmogorov-Smirnov test was applied. Student T test was used to compare the averages for parametric variables. In the analysis of correlations, considering the normal distribution of data, Pearson's linear correlation coefficient was used. Chi-square test, Fisher's exact test and Phi coefficients were used to study categorical variables. In all tests, the difference was considered statistically significant when the p value was <0.05, with a 95% confidence interval. The results were presented in tables and graphs.
The project was approved by the Research Ethics Committee of the Federal University of Piauí, number 2,527,329 and CAAE number 82702617.8.0000.5214. The study was carried out according to research ethics legislation for human beings (Resolution 466/2012). To perform data collection, consent was requested from the center chosen as the research site. By agreeing to participate in the research, participants signed an informed consent form and had the benefit of receiving their mineral calcium and phosphorus profile, in order to adopt intervention measures to curb the negative impact of mineral imbalance induced by CKD.

RESULTS
This study included 50 patients in the age group between 23 and 76 years old, who were separated into: group with up to 5 years of HD (24 patients) and group with more than 5 years of HD (26 patients). Among patients with less than 5 years of HD, 66.7% (n= 16) were male and 33.3% (n= 8) female. In relation to those with more than 5 years, 73.1% (n= 19) were male and 26.9% (n= 7) female.
Mean values and standard deviations of age and anthropometric parameters used in the assessment of nutritional status of the participants are shown in table 1. Patients with up to 5 years of HD had body weight and BMI statistically higher than those who had HD more than 5 years (p= 0.005 and p= 0.010, respectively).
Mean values and standard deviations of food consumption: energy, macronutrient, phosphorus and calcium intake of study participants is shown in table 2. There was no statistically significant difference between groups regarding the intake of energy, protein, carbohydrate, lipid, calcium and phosphorus. Table 3 shows the serum concentrations of calcium, phosphorus and PTH in both groups. Patients with more than 5 years of HD had a statistically higher serum PTH level, compared to those who in the up to 5 years HD group (p= 0.034). Table 4 shows the percent adequacy of participant calcium and phosphorus intake, according to HD group. There was no statistically significant difference between groups (p= 0.340) for calcium. There was an association between longer HD time and high serum phosphorus concentrations (p= 0.039). Table 5 shows the results of the linear correlation analysis between serum and dietary calcium and phosphorus concentrations and serum PTH in patients with CKD on HD. There was a moderate and significant positive correlation between the serum concentration of phosphorus and PTH, in the group of patients with up to 5 years of HD (p= 0.012).

DISCUSSION
The predominance of males in this research was similar to some studies found 21,22 , suggesting that men are more vulnerable to the progression of kidney disease and the need for HD, as they reveal a more negligent behavior regarding looking for health services in early stages of symptoms, present higher intake of caloric foods rich in fats and a higher frequency of alcohol consumption 23 .
By stratifying the sample according to the duration of HD treatment, it can be seen that patients with a shorter duration of HD had a statistically higher weight and BMI than those who had been on HD for a longer time (Table  1), signaling more significant nutritional wear as therapy extends over time, as nutrient plunder and metabolic changes intensify that cause protein catabolism and depletion of lean and fat mass, as well as uremia, causing weight loss and lack of appetite 24 .
The evaluation of food consumption indicates that all nutrients (except carbohydrates) had a high probability of nutritional inadequacy, which may suggest that the present sample did not follow the specific dietary guidelines for patients with CKD on HD, or the impact of the pathology interfered in food consumption. In this perspective, the loss of appetite caused by the disease and HD, hormonal changes and inflammation contribute to the deficit in food consumption of these patients 21,25,26 .
Inadequate consumption of energy and protein ( Table  2) corroborates the results of some research 25,26,27 . Rodrigues et al. 21 obtained adequate consumption of phosphorus, protein, carbohydrate and lipid, but inadequate consumption of calcium and energy.
According to Rouhani et al. 28 the energy density of the diet may indicate the risk of developing a higher stage of CKD. It is noteworthy that in patients undergoing HD, sufficient energy consumption can promote protein balance and consequently improve the performance of HD 24 .
It is appropriate to call attention to the inadequacy in dietary intake of calcium and phosphorus in both groups, with no significant difference; which is with some studies 29,30 . Low dietary calcium intake can be attributed to the insufficient consumption of milk and dairy products and other foods that are sources of calcium 14 . A study has shown that insufficient calcium consumption may be due to the limitation of consumption of phosphorus-based foods, which are also rich in calcium and protein 30 .
It should be noted that, despite the inadequate dietary intake of phosphorus, most patients in both groups analyzed presented hyperphosphatemia, which suggests the ineffectiveness of renal phosphorus excretion due to renal function failure. Furthermore, it reveals that the HD procedure does not efficiently remove serum phosphorus, due to its high molecular weight 31,30 . In this context, the low dietary intake of phosphorus found can benefit the homeostasis of calcium and phosphorus in the conservation of bone mass, by compensating the competitive mechanism existing between these minerals and contributing to maintain the level of serum phosphorus up to the limit of 4.5 mg / dL 8,13,15. . As an effective strategy, it is recommended to use phosphate binders, calcium supplements and vitamin D to minimize the effects of phosphorus retention on bone integrity 17,32 . Fifty-two percent (52%) of the sample had been undergoing HD for more than 5 years; some participants in this study used calcium supplements (1 mcg/mL calcitriol) and phosphorus chelator (sevelamer hydrochloride active ingredient 800 mg on anhydrous basis). The adequacy of dialysis by means of fractional urea clearance (Kt/V) reached values of 1.5 to 2.0. Venous access was performed through arteriovenous fistula, and among the catheters, permcath in the jugular and double lumen were the most used 13 .
Food consumption related to minerals, calcium and phosphorus, does not seem to have influenced respective serum levels, since there was no significant correlation between mineral content evaluated in the diet and serum concentrations. It is known that there are factors that affect the bioavailability of minerals, such as fiber consumption, dietary energy density, physiological states and the endogenous status of the minerals 14 . Thus, analyzing the influence of diet on serum mineral levels is very complex, and the disease in question can precipitate profound changes in mineral levels, due to its pathophysiological events and altered metabolic demand 33,15 .
The groups showed adequate mean values for calcium and high values for phosphorus and PTH (Table  3). Lehmkuhl et al. 34 obtained similar results. The study by Barros et al. 35 elevated serum levels of calcium, PTH and phosphorus in patients with a longer duration of HD, corroborating with this study only in terms of phosphorus and PTH levels.
Furthermore, hypocalcemia manifested itself in 12.5% and 3.8% of the sample, in the group with up to 5 years of HD and in the group with more than 5 years in HD, respectively, with no significant difference between groups; and hyperphosphatemia was statistically superior in the group with the longest treatment (p= 0.039) ( Table 4). This result is attributed to the ineffectiveness of the treatment in removing serum phosphate and greater phosphorus retention, due to the progression of the disease and the bone remodeling condition 36,37 . Another study 37 found adequate serum phosphorus concentration, contradicting the results of this research.
The average level of PTH was inadequate in both groups, being higher in the group with longer duration of HD (Table 3), since the disease progression aggravates the disorders inherent to this pathology. Tentori et al. 38 also observed a concentration of PTH above the recommended in the studied population, corroborating this research.
There was a moderate and significant positive correlation between serum PTH and phosphorus in the group with up to 5 years HD (Table 5), showing that the higher the serum phosphorus concentrations, the higher the PTH concentrations. This result is explained by the fact that the minerals, phosphorus and calcium, exercise competitive mechanisms; high phosphorus rates culminate in the excretion of calcium, that is, hyperphosphatemia results in the development of hypocalcemia, which is perceived by the parathyroid, which secretes PTH in an attempt to regulate the serum concentrations of this mineral. This mechanism is a risk to bone metabolism because it promotes mineral and bone disorders of chronic kidney disease (CKD-BMD) in the future; since, PTH in high amounts acts on the activation of 1α-hydroxylase and calcium reabsorption. Thus, high levels of serum phosphorus seem to cause an increase in PTH 31,38,39 .
As CKD worsens, phosphate excretion remains active, causing a fall in tubular phosphate reabsorption, which occurs due to FGF-23 and PTH. However, in view of the proximal deficiency of tubular klotho, the participation of FGF-23 to control phosphate levels is insufficient; and PTH becomes responsible for performing phosphorus homeostasis. However, in the final stages of kidney disease, PTH does not support this adaptation and hyperphosphatemia sets in, even with high levels of PTH and FGF-23 40 .

CONCLUSIONS
The study shows higher concentrations of phosphorus and PTH with the prolongation of HD, suggesting that therapy is not effective in preventing metabolic endocrine changes that favor mineral and bone disorders in CKD. Despite resistance to the action of PTH with disease progression, the results showed a significant positive correlation between serum phosphorus and PTH only in patients with less HD time, suggesting that the prolongation of dialysis therapy may be effective to remove excess phosphorus and thereby delay the progression of CKD-BMD compared to the start of treatment.