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ORIGINAL ARTICLE
Year : 2021 | Volume
: 46
| Issue : 2 | Page : 92-98
Evaluation of serum hepcidin and its relationship to iron homeostasis in obese anemic patients
Shaza A Alkourashy, Walaa A El-Salakawy, Alia M Saeed MD
Department of Internal Medicine, Clinical Haematology and Oncology Unit, Faculty of Medicine, Ain Shams University, Cairo, Egypt
Correspondence Address:
Alia M Saeed
Faculty of Medicine, Ain Shams University, 38 Abbasia, Next to Al-Nour Mosque, Al Waili, Cairo Governorate 11711
Egypt
Source of Support: None, Conflict of Interest: None
CheckDOI: 10.4103/ejh.ejh_44_20
Context Obesity is an increasingly recognized medical problem in both high-income and low-income countries owing to poor dietary habits and physical inactivity. This is frequently associated with dysregulated iron homeostasis. Iron deficiency in obese patients might be attributable to imbalanced diet or increased bodily demands. However, a role of enhanced hepcidin expression as a result of proinflammatory milieu in this setting has been contemplated in the recent years.
Objectives First, we aimed at the measurement of serum hepcidin levels in obese anemic individuals. Second, we aimed at the assessment of the presence or absence of a relationship between its levels and deranged iron homeostatic profile.
Patients and methods A total of 90 adult participants have been enrolled from the Clinical Hematology and Oncology Unit, Internal Medicine Department, Center ‘X,’ City ‘Z.’ Country ‘Y’. They have been divided into group I: 30 obese anemic patients, group II: 30 obese nonanemic patients, and group III: 30 healthy age-matched and sex-matched controls. Serum hepcidin levels were assayed using hepcidin ELISA kits.
Results Group I enjoyed much higher serum hepcidin values as compared with either group II or III. Hepcidin level was associated with significantly lower mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, serum iron, and serum ferritin levels. However, it was remarkably associated with higher BMI, red cell distribution width, erythrocyte sedimentation rate, and high-sensitivity C-reactive protein levels.
Conclusion There is a low state of chronic inflammation in obese anemic individuals associated with higher serum hepcidin levels. This results into repressed iron absorption and release by the liver and phagocytic system, contributing largely to obesity-associated iron deficiency.
Keywords: anemia, ferritin, hepcidin
Obesity has emerged as a chronic public health problem in the industrialized world in the last century. Nowadays, its prevalence is rising in the developing countries as well. This is attributable to the increasing consumption of energy-dense diet and sedentary lifestyle [1]. This complex chronic condition has serious multisystem repercussions. It increases the risk of type 2 diabetes mellitus, systemic hypertension, cardiovascular disorders, dyslipidemia, osteoarthritis, obstructive sleep apnea syndrome as well as some types of cancers. Moreover, it enhances the susceptibility of an individual to infections and impairs postoperative wound healing [2].
It is frequently associated with dysregulated iron homeostasis either in the form of iron deficiency or dysmetabolic iron overload syndrome [1]. Several mechanisms have been proposed to contribute to obesity-associated iron deficiency. These include imbalanced diet, increased iron demands owing to increased body size and blood volume, and menstrual disturbances. The most recent observations underscore the influence of obesity-associated low-grade inflammation and hepcidin overexpression on systemic iron metabolism [3].
This is mediated by the proinflammatory cytokine interleukin 6, which enhances the release of the principal iron-regulatory peptide, hepcidin [4]. Hepcidin is a small cysteine-rich hormone secreted by the liver into the plasma and gets cleared by the kidney. Its secretion is enhanced by elevated iron stores and inflammation, whereas it gets repressed by anemia and hypoxia [5].
Hepcidin causes internalization and degradation of the sole iron cellular exporter, ferroprotein. Thus, it limits iron absorption through the basolateral membranes of enterocytes and reduces iron mobilization from macrophages and hepatic iron stores [5]. In the current study, hepcidin levels are examined in obese anemic patients to identify its relationship to anemia and the coexistence of iron deficiency.
Patients and methodsParticipants
The current case–control study was conducted on 90 adult participants. All the patients were recruited from the outpatient clinic of the Clinical Hematology and Oncology Unit, Internal Medicine Department, Faculty of Medicine, Center ‘X,’ City ‘Z,’ Country ‘Y’ in the period extending between November 2017 and December 2018. A written informed consent has been taken from all the participants participating in the present study. The study conformed to the stipulations of the local ethical and scientific committees of Center ‘X’ and was conducted according to the declaration of Helsinki. Data were collected and retrieved from the patients’ files. Study participants were grouped into three different groups:
Group I: 30 obese anemic patients.Group II: 30 obese nonanemic patients.Group III: 30 healthy age-matched and sex-matched controls.Inclusion criteria
The following were the inclusion criteria:
Age 18–70 years.Overweight and obese adults with BMI more than 25.Anemic patients as per the WHO definition [6].Exclusion criteria
The following were the inclusion criteria:
Age less than 18 and more than 70 years old.Pregnant or lactating women or those taking oral contraceptive pills.Patients with chronic inflammatory diseases or chronic infections.Patients with macrocytic anemias.Patients with autoimmune hemolytic anemias either primary or secondary.Patients with congenital chronic hemolytic anemias.Patients with bone marrow failure syndromes.Patients with deranged thyroid profile.Patients with chronic kidney disease.Methods
First, all the study participants underwent the assessment of their anthropometric measures including weight and height. Weight measurement has been performed using digital scale adjusted to the nearest 0.1 kg, whereas height was measured using a fixed stadiometer. BMI was then calculated as weight in kilograms divided by the square of height in meters. This was followed by the assessment of their iron status by measuring serum ferritin, total iron-binding capacity (TIBC), serum iron, and transferrin saturation. Then, the assessment of the inflammatory status using erythrocyte sedimentation rate (ESR) and high-sensitivity C-reactive protein (hs-CRP) has been performed. Finally, the measurement of serum hepcidin level using ELISA has been done.
For each participant, ∼5 ml of venous blood was drawn under complete aseptic conditions and dispensed into a labeled vacutainer containing gel and clot activator, and serum was separated by centrifugation for 5 min, then stored at temperature −20°C till the time of assay. Serum hepcidin levels were determined by ELISA assay using a commercially available ELISA kit (ABIN415120 Hepcidin ELISA Kit; Schloss Rahe GmbH Company, Germany). The values were expressed in μg/l.
Statistical analysis
The collected data were coded, tabulated, and statistically analyzed using IBM SPSS statistics software (version 18.0, 2009; IBM Corp., Chicago, Illinois, USA). Descriptive statistics were done for quantitative data using minimum and maximum of the range as well as mean±SD for quantitative normally distributed data, whereas it was done for qualitative data as frequencies and percentages. Inferential analyses were done for quantitative variables using Shapiro–Wilk test for normality testing, and analysis of variance test with post-hoc Tukey test for more than two independent groups with normally distributed data. In qualitative data, inferential analyses for independent variables were done using χ2 test for differences between proportions. However, correlations were done using Pearson correlation for numerical normally distributed data. The level of significance was taken at P value less than 0.05 being significant.
ResultsA total of 90 adult participants were recruited from the outpatient clinic of the Clinical Hematology and Oncology Unit, Internal Medicine Department, Center ‘X,’ City ‘Z,’ Country ‘Y.’ They were divided into three equal groups as per their BMI and hemoglobin percentages as illustrated in the patients and methods section. [Table 1] shows demographic and laboratory characteristics of the three groups. The three groups exhibited almost the same mean age with an average age of 49.8±12.1, 49.7±12.1, and 49.2±11.4 years for groups I, II, and III; respectively. Group I had 12 (40%) males and 18 (60%) females, group II contained 13 men and 17 women, whereas group III had equal sex distribution, with comparable sex distribution among the three study cohorts. Their mean BMIs are 30.1±2.9, 30.2±1.9, and 22.6±1.9 kg/m2 for groups I, II, and III, respectively. Group I had a mean hemoglobin percentage that was far less than that of either groups II or III, being 9.5±1.1, 13.5±0.8, and 13.9±0.8 g/dl, consecutively. The same is replicated for hematocrit, mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), and mean corpuscular hemoglobin concentration (MCHC), with figures well-illustrated in [Table 1]. In spite of all of these differences demonstrable between group I from one side and groups II and III from the other side, it is obvious that the three groups were comparable in terms of their platelet and total leukocytic counts. A great deal of anisocytosis is observable in group I in comparison with groups II and III as shown by red cell distribution width (RDW). RDW was 18.0±1.6 in group I as compared with groups II and III, whose RDWs were 13.7±1.2 and 13.3±1.3, respectively.
Table 1 Demographic and laboratory characteristics of the studied groups with comparison of different study variables among themThe assessment of iron status in the three groups showed that group I had an iron-deficient profile with lower serum iron and ferritin and high TIBC as compared with groups II and III. This group exhibited increased proinflammatory markers including ESR and hs-CRP in reference to the other two groups, as demonstrable in [Table 2].
Table 2 Comparison of the levels of different inflammatory biomarkers among the studied groupsGroup I exhibited the highest mean serum hepcidin level, whereas group III had the lowest levels and group II fell in between. Mean hepcidin levels were 28.8±3.4, 25.1±4.5, and 21.7±3.6 μg/l in groups I, II, and III, respectively. This difference has reached high statistical significance, with a P value less than 0.001.
Serum hepcidin levels in the three studied groups has been correlated to the other quantitative study variables, as shown in [Table 3]. It is evident that there has been a strong positive correlation between serum hepcidin levels on one side and the BMI of the three study groups on the other side, with P values of less than 0.001 0.006, and 0.004 for groups I, II, and III, respectively. This is well-illustrated in [Figure 1] and [Figure 2]. Moreover, serum hepcidin levels exhibited positive correlation with RDW and TIBC. Patients with higher serum hepcidin had higher degree of anisocytosis, with P values of 0.002, less than 0.001, and less than 0.001 in groups I, II, and III, respectively. Moreover, the higher is the serum hepcidin level, the higher is the capacity of transferrin to bind iron, with P values of 0.004, less than 0.001, and less than 0.001 in groups I, II, and III in turn. Inflammatory biomarkers, ESR and hs-CRP, also demonstrated positive correlations with serum hepcidin values, with P values less than 0.05 in the three studied groups. [Figure 3] depicts the direct relationship between serum hepcidin and hs-CRP.
Table 3 Correlations of serum hepcidin levels with other quantitative variables among the studied groups
Figure 3 Correlation between serum hepcidin levels and hs-CRP. hs-CRP, high-sensitivity C-reactive protein.By sharp contrast to this, serum hepcidin levels were negatively correlated with hemoglobin percentages, hematocrit, MCV, MCH, MCHC, serum iron, and serum ferritin in all the three study groups, with P values less than 0.05.
[Figure 4] portrays the inverse relationship between serum hepcidin and hemoglobin percentages.
DiscussionThis case–control study has been conducted on 90 adult participants, comprising 30 obese anemic patients, 30 obese nonanemic patients, and 30 lean healthy age-matched and sex-matched controls.
In our study, it has been observable that there were more females than males in the obese groups either anemic or nonanemic, but this discrepancy did not exhibit statistical significance. This observation is compatible with a systematic review conducted by Kanter and Caballero [7] comparing the rates of obesity by sex in 105 countries. It was clear that the prevalence of female obesity surpassed that of males in all income groups of countries, but the situation was reversed in a subgroup analysis of high-income countries. Moreover, female obesity was significantly higher than male obesity in MENA region as compared with Europe and Central Asia, where the reverse is true. This was in part explained by the higher physical activity exerted by males in MENA region as compared with females due to the nature of their occupations. Moreover, the deeply seated sociocultural beliefs in this region link obesity in women to maternity, nurturing, fertility, high socioeconomic status, and prosperity, discouraging females to have leisure time physical activity. In our study, this difference did not reach a statistical significance owing to the low numbers of participants in each group.
Group III exhibited significantly lower mean BMI as compared with groups I and II. This is attributable to the selection and grouping criteria defining each group, whereas group I demonstrated remarkably lower mean hemoglobin percentage and hematocrit in relation to groups II and III, which are attributable to the specifications that were set for the allocation of patients in each group to investigate the role of iron-restricted erythropoiesis in the evolvement of anemia in obese individuals.
The work of Vuong et al. [8] published in 2014 contradicts this finding. Their study showed that waist circumference, used as a surrogate marker for obesity, has been positively correlated with hemoglobin percentage and hematocrit values. They explained that observation by several mechanisms. The first is the occurrence of insulin resistance with the development of hyperinsulinemia in obese individuals. This in turn raises insulin-like growth factor I levels serving as a stimulus for erythropoiesis. The second is the development of chronic hypoxia thanks to obstructive sleep apnea syndrome, and the last is the build-up of glycated hemoglobin levels shifting hemoglobin oxygen dissociation curve to the left hindering the delivery of oxygen to the tissues, which again contributes to hypoxia. Hypoxia stimulates the expression of hypoxia inducible factor-1, which in turn stimulates erythropoietin production in the liver and kidney [8].
Another point of comparison of the three cohorts was the red blood corpuscle indices, namely, MCV, MCH, MCHC, and RDW. It is noticeable that group I had substantially lower MCV, MCH, and MCHC and higher RDW in relation to groups II and III. This goes in line with the previous study held by Vuong et al [8], exhibiting an inverse relationship between waist circumference from one side and MCV, MCH, and MCHC from the other side. By contrast, a direct relationship has been observed between waist circumference and RDW. Again, these findings were reproducible in another study conducted in 2017 by El-kerdany et al. [9].
However, in our study it is observed that the mean white blood cell and platelet counts were comparable among the three studied groups. This is discordant with the findings of Sal et al. [10]. They observed that obese children had higher mean white cell as well as platelet counts. They ascribed this to inflammatory status prevalent in obese individuals with these serving as acute-phase reactants whose elevations are associated with the occurrence of metabolic syndrome. The lack of difference in these two parameters in our study may be ascribed to the fact that obesity creates a low state of inflammation that causes no gross alteration of white cell as well as platelet populations.
Concerning iron parameters examined in the study population, it is discernible that group I had significantly lower serum iron and ferritin levels as well as higher TIBC as compared with the other two groups. In contrast to this, a study by Tussing-Humphreys et al. [3] has been done on 40 premenopausal women segregating them equally by their BMIs into obese and nonobese ones. In this study, there was no statistically significant difference between both groups in terms of their serum iron, ferritin, TIBC, and transferrin saturation. Nevertheless, a remarkable discrepancy has been exhibited between both groups with respect to their soluble transferrin receptors considered as the most accurate indicator of iron status. This makes their findings compatible with ours. The difference noticed between both studies is related to their lower sample size, precluding them from reaching statistical significance. In addition to that, their sampling characteristics were different; they recruited obese nonanemic patients.When comparing the inflammatory status among the three groups under study, it is noticeable that the three inflammatory markers examined, namely ESR, hs-CRP, and serum hepcidin, mirrored each other with the highest levels obtained in obese anemic group and the lowest yielded in the lean healthy cohort. The differences among the groups were statistically significant. Furthermore, correlating serum hepcidin to other study variables demonstrated that it had a strong direct relationship to BMI, serum ferritin, TIBC, as well as to other inflammatory biomarkers such as hs-CRP and ESR. However, it had a strong negative correlation to hemoglobin and hematocrit percentages as well as to MCV, MCH, and MCHC.
This imitated the work done by Tussing-Humphreys et al. [3], which showed significantly higher hepcidin values in obese participants in relation to nonobese ones. Moreover, they demonstrated the presence of a direct relationship between serum hepcidin levels on one side and BMI, serum ferritin, and CRP from the other side.
Acknowledgements
Authors’ contributions: S.A.A.: designed the study and analyzed the data. W.A.E. contributed to analyzing data, writing the manuscript, and revising it. A.M.S. collected the data, made major contribution to analysis of the data, drafted the manuscript, and revised it.
Financial support and sponsorship
Nil.
Conflicts of interest
There are no conflicts of interest.
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