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 Table of Contents  
ORIGINAL ARTICLE
Year : 2019  |  Volume : 19  |  Issue : 1  |  Page : 13-18

Serum hepcidin levels and erythropoietin resistance in hemodialysis patients


1 Nephrology Unit, Department of Internal Medicine, Zagazig University, Zagazig, Egypt
2 Clinical Pathology Department, Zagazig University, Zagazig, Egypt

Date of Submission10-Dec-2018
Date of Acceptance18-Feb-2019
Date of Web Publication01-Apr-2019

Correspondence Address:
Dr. Walid M Afifi
Nephrology Unit, Department of Internal Medicine, Faculty of Medicine, Zagazig University, Zagazig
Egypt
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/jesnt.jesnt_33_18

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  Abstract 


Introduction Erythropoietin (EPO) resistance is an important cause of anemia in patients with chronic kidney disease. Enhancement of erythropoiesis by EPO requires intact EPO signaling and effective mobilization of iron stores. Enhanced synthesis of hepcidin leads to reduction of iron absorption in the small intestine and sequestering of iron in macrophages. Hepcidin may contribute to EPO resistance through a direct inhibitory effect on erythroid progenitor proliferation and survival.
Objective To assess serum hepcidin levels in patients undergoing hemodialysis (HD) treatment and to distinguish its analogous association with anemia parameters and EPO resistance.
Patients and methods A cross-sectional study was carried out among HD patients at Nephrology Unit of the Zagazig University Hospital from 2017 to 2018. The study included 90 participants, meeting our inclusion criteria, who had end stage renal disease (ESRD) and were on regular HD three times weekly. All patients were subjected to full history taking and clinical and laboratory assessment. It included routine investigations, with complete blood picture, calcium, phosphorus, parathyroid hormone (PTH), lipid profile, iron panel, and c reactive peptide (CRP). Specific investigations included the EPO resistance index and serum hepcidin measurement.
Results Hepcidin is positively related to EPO resistance index. EPO resistance index as well as ferritin were significant determinants of hepcidin in HD patients.
Conclusion Hepcidin is associated with anemia, iron status, and microinflammation in HD patients. If used as a diagnostic tool, it might improve iron therapy during periods of reticuloendothelial blockage of iron transport. This is important to avoid iron overload and to improve EPO response in patients with ESRD.

Keywords: anemia, erythropoietin resistance, hemodialysis, hepcidin


How to cite this article:
Afifi WM, Mostafa E, Elsaid M, Elakad G, Ebian HF. Serum hepcidin levels and erythropoietin resistance in hemodialysis patients. J Egypt Soc Nephrol Transplant 2019;19:13-8

How to cite this URL:
Afifi WM, Mostafa E, Elsaid M, Elakad G, Ebian HF. Serum hepcidin levels and erythropoietin resistance in hemodialysis patients. J Egypt Soc Nephrol Transplant [serial online] 2019 [cited 2019 Jun 25];19:13-8. Available from: http://www.jesnt.eg.net/text.asp?2019/19/1/13/255244




  Introduction Top


Hepcidin, is a 25-aa cysteine-rich peptide, found in human serum and urine and represents the key peptide hormone, isolated as a novel peptide from plasma.

As it is produced by the liver and had antimicrobial properties, it has been named liver-expressed antimicrobial peptide-1. It is isolated from human urine and is designated as hepcidin (hepatic bactericidal protein), which modulates iron homeostasis in the body [1].

Data imply that an excess in hepcidin may account for the insignificant absorption of iron and the presence of reticuloendothelial cell iron blockade in many patients with chronic kidney disease (CKD) [2].

Hepcidin, is the principal hormone involved in iron homeostasis, which is manufactured by the liver and secreted into the circulation. Hepcidin binds and instigates destruction of the iron exporter, ferroportin, on duodenal enterocytes, reticuloendothelial macrophages, and hepatocytes, preventing the entry of iron into the plasma [3].

Inflammatory cytokines directly stimulate hepcidin transcription, to isolate iron from attacking pathogens, thus resulting in iron sequestration, hypoferremia, and anemia, which are trademarks of many chronic illnesses, including CKD [4].

The use of erythropoiesis-stimulating agents (ESAs) among dialysis patients is supported by many retrospective population-based studies in a very large number of hemodialysis (HD) patients and some prospective controlled studies that have explored outcomes associated with various hemoglobin (Hb) or hematocrit levels. Although there is some variability in results among studies, the most consistent observation has been that better outcomes in terms of quality of life, without an increase in adverse reactions, are associated with Hb values between 10 and 12 g/dl, compared with values below this level. ESAs should not be started until iron parameters have been evaluated. Among patients with evidence of iron deficiency, iron therapy should be given first before initiating ESAs [5].

Certain patients display low sensitivity to ESAs, thus requiring a higher dose that probably correlates with increase in patient mortality, an outcome that persists after modification for the usually lower hematocrit in such patients [6]. Our quest is to quantify serum hepcidin levels in HD patients and to establish the linkage between hepcidin and anemia parameters and erythropoietin (EPO) resistance.


  Patients and methods Top


A cross-sectional study was carried out at Nephrology Unit of the Zagazig University Hospital from 2017 to 2018. A total of 90 HD patients were included, comprising 54 females and 36 males, with mean±SD age of 47.86±15.05 years. Patients were dialyzed on Fresenius 4008B (fresenius) dialysis machine (Bad Homburg, Germany) at blood flow rate of 300–350 ml/min, dialysate flow rate of 600 ml/min, using polysulfone hollow fiber dialyzers suitable for the surface area of the patients (Fresenius FX8=1.4 m2, FX10=1.8 m2 and F6=1.2 m2) and using 1.5 mmol/l dialysate calcium. Patients were dialyzed three sessions per week, 4 h for each session. Protocols were not changed during the study, with adequate dialysis treatment (Kt/V>1.4) in more than 90% of the patients.

EPO resistance is defined according to the European Best Practice Guidelines [7] as a failure to achieve target Hb levels with doses of rhEPO more than 300 IU/kg/week of epoetin or 1.5 µg/kg/week of darbepoetin-α.

Inclusion criteria

Age 18–75 years, having end stage renal disease (ESRD), on regular HD, with good patient compliance, and less missed treatments with anemia on recombinant EPO therapy for at least 8 weeks were the inclusion criteria.

Exclusion criteria

Uncontrolled hypertension or diabetes; liver dysfunction; malignancy; hemoglobinopathies; hepatitis C and hepatitis B, active or subclinical infection; blood transfusion within the past 3 months; or treatment with steroids or other immunosuppressive drugs.

Written informed consent was taken from the patients to participate in the study. Approval for performing the study was obtained from Internal Medicine and Clinical Pathology Departments, Zagazig University Hospitals, after taking Institutional Review Board approval.

All patients were subjected to full history taking and clinical and laboratory assessment. It included routine investigations, with complete blood picture, kidney function tests, calcium, phosphorus, parathyroid hormone (PTH), lipid profile, iron panel, virology [hepatitis b surface antigen (HBsAg), hepatitis c virus antibody (HCV ab)], and c reactive peptide (CRP). Specific investigations included the EPO resistance index and serum hepcidin.

The erythropoietin resistance index (ERI) was calculated as the weekly weight-adjusted dose of EPO (IU/kg/week) divided by Hb concentration (g/dl) and has also been considered useful to assess the EPO resistance (32).

Serum hepcidin using enzyme-linked immunosorbent assay. The assay was carried out by hepcidin enzyme-linked immunosorbent assay kit by New Test Company (DaXing Industry Zone, Beijing, China).

Statistical analysis

Data were presented and analyzed by using computerized software statistical packages by ‘statistical package for the social sciences’ (SPSS version 19.0; SPSS Inc., Chicago, IL, USA). Mean±SD with range were used to describe quantitative data, whereas numbers with percentages were used to describe qualitative data. χ2-Test and Fisher exact were used to compare proportions, whereas independent sample (t) test was used to compare means between the main groups (P≤0.05 was considered to be statistically significant at 95% confidence interval). We considered positive sign as indication for direct correlation ‘increase frequency of independent lead to increase frequency of dependent’ and negative sign as indication for inverse correlation ‘increase frequency of independent lead to decrease frequency of dependent’. Moreover, we considered values near to (1) as strong correlation and values near (0) as weak correlation. The significance level for all aforementioned statistical tests done. Multiple regression analysis using the stepwise method was used to detect independent factors affecting hepcidin and ERI, with P value less than 0.05 in univariate analysis. Receiver operating characteristic curve was constructed to permit selection of threshold values for test results and comparison of different testing strategies.


  Results Top


[Table 1],[Table 2],[Table 3],[Table 4],[Table 5],[Table 6],[Table 7].
Table 1 Demographic and clinical data according to erythropoietin resistance index in the hemodialysis group

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Table 2 Laboratory parameters according to erythropoietin resistance index in the hemodialysis group

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Table 3 Correlations between hepcidin and other parameters in the dialysis group

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Table 4 Correlations between erythropoietin resistance index and other parameters in the dialysis group

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Table 5 Multivariate analysis of factors affecting hepcidin in hemodialysis patients

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Table 6 Multivariate linear regression analysis of factors affecting erythropoietin resistance index in hemodialysis patients

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Table 7 Validity of hepcidin as an indicator of erythropoietin resistance in hemodialysis patients

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  Discussion Top


Under diverse conditions, EPO can act as a hepcidin-inhibitory hormone once it downregulates hepcidin expression. Anemia also appears to be an additional influence that affects hepcidin expression. Moreover, anemia is associated with a decline in hepcidin expression, prompting an increase in iron absorption in the intestines and iron release by the macrophages supplementing an improvement in the obtainability of iron for EPO [8]. The function of hepcidin as a cardiovascular (CV) indicator reaped the awareness in the CKD population. Latest statistics corroborate that hepcidin plays an encompassing role in the instigation of atherosclerosis, and by its quantitative value, it may succor in distinguishing individual risk of patients [9]. Patients with CKD who have resistance to ESAs exemplify a sizeable percentage and are also bracketed with augmented CV morbidity and all-cause mortality. ESAs are known to convalesce the quality of life and also ease the demand of blood transfusion; nonetheless, there is no concrete statistics to show upturn on other sequels related with anemia of CKD, such as CV disease and mortality [10]. It is our quest to quantify serum hepcidin levels in HD patients so as to establish the linkage between hepcidin and anemia parameters and eventually to resolve its involvement with EPO resistance.

Sezgin and Zumrutdal [11] reported that the hepcidin levels in HD patients who received no EPO were lower when compared with the levels in those patients and receiving the maximum dose of EPO, and this consistent with our finding. When we divided the HD patients according to ERI into responders (ERI<15) group and nonresponders (ERI>15) group, we observed that ferritin, CRP, and hepcidin were significantly higher in nonresponders, but Hb was found to be lower in the nonresponders. Our data are consistent with Rubab et al. [12] who found significantly higher hepcidin and lower Hb levels in nonresponder HD patients. However, Costa et al. [13] and Do Sameiro-Faria et al. [14] determined that, in contrary to our results, hepcidin serum levels between nonresponders were suggestively lower than those of the responders.

It has been established that EPO downregulates liver hepcidin expression, acting as a hepcidin inhibitory hormone. Equated to responders, as nonresponders were treated with higher doses of ESA, the lower hepcidin levels among nonresponders could be explicated by this structure. In agreement with Petrulienė et al. [10] who found that significantly higher CRP and lower Hb in nonresponders. However, Petrulienė et al. [10] found higher ferritin and hepcidin concentrations, but without statistical significance unlike our findings. This may be owing to different regimes of anemia management and HD, as they also found a significant difference regarding EPO doses, which is unlike our findings, where we found no significant difference in EPO doses between responders and nonresponders.

Nonresponders group had significantly higher ERI than the rest of the patients in this study, as did most of the studies, such as Petrulienė et al. [10]. In this study, we found a nonsignificant correlation between age and hepcidin levels among our HD patients. In agreement with our findings, (26) demonstrated that there was no statistically significant correlation between hepcidin and patient age. We also found a significant positive correlation between serum hepcidin levels and ferritin in HD patients. Findings consistent with ours have been seen in the studied by Al-Amir et al. [15], Petrulienė et al. [10], and Zhang et al. [24]. In addition, Caliskan et al. [25] reported that serum prohepcidin levels were positively correlated with ferritin.

Serum ferritin is a marker of iron stores in the liver and reticuloendothelial system, as well as being an acute-phase protein. This suggests that hepcidin plays a major role in regulating iron homeostasis in HD patients [15]. Multivariate analyses validated that hepcidin levels in patients with CKD have a major correlation with serum ferritin and are affected by inflammation, iron therapy, estimated glomerular filtration rate, ESA dose, dialysis adequacy, and Hb [16],[17],[18],[19]. This result showed a negative significant correlation between hepcidin and Hb in HD groups. These findings are consistent with Petrulienė et al. [10].

We found a positive significant correlation between ERI and hepcidin in HD patients. These findings are consistent with Petrulienė et al. [10]. This result showed a positive significant correlation between hepcidin and CRP in HD patients. This is in line with Rubab et al. [12] and Malyszko et al. [20]. However, hepcidin did not correlate with CRP but ERI correlated with CRP according to Petrulienė et al. [10]. We also found a significant negative correlation between ERI and BMI. This is in agreement with Petrulienė et al. [10] and Kotanko et al. [21] who found that ERI was inversely related to BMI. In agreement with Petrulienė et al. [10], we also found a significant positive correlation between hepcidin and EPO dose in HD patients.

Multiple variable analysis was executed to explore the predictors of ERI in dialysis patients among the variables that correlated with ERI in simple regression analysis. Included in these variables are CRP and HD duration, which were found to be significant independent determinant of ERI. In addition, we found ERI to be an independent factor that influences hepcidin. It was reported by Petrulienė et al. [10] that ERI and ferritin were observed to be significant determinants of hepcidin in HD patients and CRP was found to be significant independent determinant of ERI. These remarks are harmonious with our results. The said conclusion is in conformity with Ibrahim et al [22], [23],[26] who found that CRP can be an independent factor influencing ERI.

One of study’s conceivable constraint is that nutritional deficiencies that can create circumstances that limit the production of erythrocytes, such as vitamin B12 and folate, were not evaluated.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
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    Tables

  [Table 1], [Table 2], [Table 3], [Table 4], [Table 5], [Table 6], [Table 7]



 

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