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Year : 2023  |  Volume : 9  |  Issue : 1  |  Page : 50-56

Assessment of Vitamin A Status in Patients with Iron Deficiency Anemia

1 Department of Internal Medicine, Maulana Azad Medical College and Associated Hospitals, Delhi, India
2 Department Cardiology, Govind Ballabh Pant Institute of Postgraduate Medical Education and Research, Delhi, India
3 Department of Biochemistry, Maulana Azad Medical College and Associated Hospitals, Delhi, India

Date of Submission24-Nov-2022
Date of Decision09-Dec-2022
Date of Acceptance26-Jan-2023
Date of Web Publication28-Apr-2023

Correspondence Address:
MD Abhishek Verma
Department of Internal Medicine, Maulana Azad Medical College and Associated Hospitals, F-190h, Laxmi Nagar, Delhi-110092
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/mamcjms.mamcjms_70_22

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Introduction: Iron and vitamin A deficiency are two very prevalent and easily preventable nutrient deficiencies. This study was conducted to assess vitamin A status in patients with iron deficiency anemia and to further study the correlation of vitamin A status with biochemical markers of iron deficiency. Materials and Method: Eighty patients with iron deficiency anemia were enrolled and investigated for a complete blood count, an iron profile, liver, and kidney function tests and plasma retinol binding protein levels. The mean age of patients was 31.14 ± 11.33 years, with a range of 16 to 62 years. Results: Mean hemoglobin was 7.19 ± 2.1 g/dL. Serum iron, ferritin, and transferrin saturation were low in all patients, while total iron binding capacity (TIBC) was elevated in only 74 patients (94.81%). Nineteen patients (23.8%) had vitamin A deficiency, with a mean retinol binding protein (RBP) 0.53 ± 0.13 µmol/L. Vitamin A deficient patients had a mean hemoglobin of 6.8±2.14 gm/dL, mean corpuscular volume (MCV) 71.35 ± 8.86 fL, a mean corpuscular hemoglobin (MCH) 19.43±4.36 pg, mean corpuscular hemoglobin concentration (MCHC) 25.44±4.92 gm/dL, serum iron of 28.21 ± 9.73 mcg/dL, serum ferritin 13.04 ± 12.41 ng/mL, transferrin saturation 6.81 ± 3.07%, and TIBC 427.85 ± 78.57 mcg/dL. Among vitamin A deficient patients, RBP had positive correlation with serum iron and transferrin saturation; while, simultaneously showing negative correlation with serum ferritin and TIBC. Conclusion: Vitamin A deficiency affects iron metabolism, causing abnormal iron trapping and systemic iron deficiency, thus worsening the clinical profile of iron deficiency anemia. This study guides us to screen iron deficiency anemia patients for the concomitant vitamin A deficiency for efficient treatment of such patients.

Keywords: ferritin, iron deficiency anemia, retinol binding protein, vitamin A deficiency

How to cite this article:
Aggarwal S, Verma A, Tiwari S, Kaushik S, Garg S, Kumar S. Assessment of Vitamin A Status in Patients with Iron Deficiency Anemia. MAMC J Med Sci 2023;9:50-6

How to cite this URL:
Aggarwal S, Verma A, Tiwari S, Kaushik S, Garg S, Kumar S. Assessment of Vitamin A Status in Patients with Iron Deficiency Anemia. MAMC J Med Sci [serial online] 2023 [cited 2023 Jun 6];9:50-6. Available from: https://www.mamcjms.in/text.asp?2023/9/1/50/375338

Key Message: Iron and vitamin A deficiency are common nutritional deficiencies in our country. Vitamin A deficiency modulates iron metabolism and worsens iron deficiency. Vitamin A status should be assessed in iron deficiency anemia patients for effective management of both diseases.

  Introduction Top

Iron deficiency anemia (IDA) is one of the commonest forms of nutritional anemia worldwide. World Health Organization (WHO) reported that iron deficiency is responsible for approximately 50% of all cases of anemia among nonpregnant and pregnant women and for 42% of cases in children aged under 5 years.[1] As per National Family Health Survey − 5 (2019–2021), in India, prevalence of anemia is 67.1% in children below 5 years, 57% in women aged 15 to 49 years, and 25% in men aged 15 to 49 years. Iron deficiency anemia causes significant loss of productivity in adults.[2] It shows the significant economic and medical impact of anemia on middle- and low-income countries. To deal with that, American Academy of Pediatrics (AAP) recommends assessing risk factors for iron deficiency annually in all adolescents, followed by laboratory screening in those with positive risk factors.[3] Clinical features commonly seen in IDA are fatigue, weakness, headache, pica, irritability, exercise intolerance, exertional dyspnoea, pallor, dryskin, atrophic glossitis, cheilosis, koilonychia, tachycardia, and flow murmur.[4] IDA in pregnancy is associated with increased maternal mortality, preterm labor, low birth weight, and increased infant mortality. IDA in children leads to poor cognitive and motor development and increases susceptibility to infections.[5]

Vitamin A is an essential fat-soluble vitamin needed in small amounts for the normal functioning of the visual system, maintenance of cell function for growth, epithelial integrity, red blood cell production, immunity, and reproduction. Vitamin A deficiency (VAD) is a major nutritional concern in poor societies and lower income countries. Inadequate vitamin A intake is a major cause of VAD. Deficiency of sufficient duration or severity can lead to night blindness, xerophthalmia, corneal ulcers, anemia, and compromised host resistance to infection. VAD-related ocular pathologies are one of the leading causes of preventable childhood blindness.[6]

Circulating retinol is the most commonly used indicator for vitamin A status. Low level of serum retinol is associated with functional outcomes of VAD, and it reflects vitamin depleted liver stores. Recently, retinol binding protein has also been used to measure vitamin A status and has produced similar results to serum retinol.[7] VAD affects iron metabolism and systemic iron availability, [Figure 1] for which, the following mechanisms have been proposed:
  1. Modulation of hematopoiesis − vitamin A stimulates erythroid precursor cells in marrow and also induces erythropoietin production.[8],[9]
  2. Prevention from infection − During infections, synthesis of transferrin and RBP decreases, causing entrapment of iron and vitamin A in the liver and spleen. Suppression of infection by vitamin A would lead to the resumption of transferrin and RBP synthesis; therefore, releasing trapped iron and vitamin A.[10],[11]
  3. Iron metabolism and transport − It has been suggested that during VAD, iron is trapped in the liver and spleen and not effectively released and used for erythropoiesis, leading to the development of anemia and abnormal iron indices. It is found that iron absorption is increased during VAD, but iron utilization in bone marrow is impaired.[12],[13]
Figure 1 Relation between vitamin A, iron metabolism and erythropoiesis.

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It is also proposed in studies that vitamin A status may influence the metabolism of proteins involved in hepatic iron storage and mobilization.[14],[15]

IDA and VAD, both are treatable major public health problems in India. Deficiency of these are important problems in adolescence, pregnancy, lactation, and in women of child-bearing age. Hence, these nutritional deficiencies are a matter of serious concern and should be managed on priority. Various studies have documented association and co-existence of these two. Majority of such studies were conducted in children, adolescents, and pregnant women. Further, there is a paucity of data in India regarding the adult population with IDA. Hence, we planned a study among population (15–60 years age) with iron deficiency anemia to assess vitamin A status in them and to further study the correlation of vitamin A status with biochemical markers of iron deficiency, so that future treatment can be planned accordingly to efficiently manage both these problems.

  Materials and Methods Top

It was an observational, cross-sectional study. Based on previous literature research on the prevalence of iron deficiency anemia and keeping a level of significance of 95%, the power of the study at 80%, and an allowable error of 5% as acceptable, the sample size was calculated as 384. The sample size was calculated by using the formula 3.84p (1−p)/n2, where p is prevalence taken as 50% and n is allowable error taken as 0.05. However, due to financial constraints, we kept the convenience sample size in the study at 80. The study was conducted after being approved by the ethical committee of the institute and was in accordance with the guidelines provided by the World Medical Association Declaration of Helsinki on ethical principles for medical research involving humans. A well-informed consent was taken from all patients above 15 years and hemoglobin <12 gm/dL. A detailed history was taken, and a physical examination was performed on these patients. All patients were subjected to the following investigations: complete blood count with red blood cell (RBC) indices, liver function test (by spectrophotometry), kidney function test (by spectrophotometry), serum ferritin levels (by electrochemiluminescence immuno assay), serum iron levels (by electrochemiluminescence immuno assay), total iron binding capacity (TIBC) (by electrochemiluminescence immuno assay), transferrin saturation (by electrochemiluminescence immuno assay), chest X ray, plasma retinol binding protein (RBP) levels (by Enzyme Linked ImmunoSorbent Assay [ELISA]). Eighty patients of IDA with decreased serum iron and decreased serum ferritin were enrolled in the study. Patients with chronic diseases (e.g., chronic kidney disease, chronic liver disease, etc.), dimorphic anemia, and autoimmune diseases were excluded from the study.

In our study, we assessed vitamin A status by measuring plasma RBP. Plasma RBP occurs in 1:1 molar complex with retinol. Also, assessment of RBP is easier than plasma retinol. Because RBP is a protein, it can be detected with an immunologic assay, which is simpler and less expensive than High Performance Liquid Chromatography (HPLC) analysis of plasma retinol. RBP is more stable than retinol with respect to heat and light. Hence RBP is a sensitive and specific surrogate marker for serum retinol.[16]

RBP level estimation was done by the direct ELISA method with wells precoated with purified human RBP antibodies. The detection range of RBP with the used ELISA kit is 0.5 to 500 µg/mL. VAD is defined by plasma retinol level <0.7 µmol/L. As RBP and retinol exist in a 1:1 molar complex, patients with RBP levels <0.7 µmol/L (14.7 µg/mL) were considered vitamin A deficient.[16]

For statistical analysis, data were entered in MS-excel and analyzed using Statistical Package for Social Sciences (SPSS) version 24.0 for windows. Qualitative data were expressed in proportion/percentage and difference between proportions was expressed in chi-square test or Fisher exact test. Quantitative data were expressed in mean ± SD or median ± interquartile range and differences were analyzed using students t-test. Statistical analysis was carried out to assess iron study parameters like serum iron, ferritin, TIBC, transferrin saturation, and plasma retinol levels in patients enrolled in the study. P value of <0.05 was kept as significant.

Each patient was given a consent form to fill. Patients were explained the purpose of the study and their right to quit at any time without having to give reason. Patient information was dealt with confidentiality. Any abnormality detected during screening was appropriately managed.

  Results Top

Patients in study were in between 16 and 62 years, with mean age of 31.14 ± 11.33 years. Maximum number of patients, 25 (31.2%) each were in the age group 21 to 30 years and 31 to 40 years, followed by 14 (17.5%) in 16 to 20 years, 10 (12.5%) in 41 to 50 years and 6 (7.5%) in 51 to 62 years. Twenty five (31%) patients were male and 55 (69%) females. Patients had overlapping symptoms of IDA, among which, weakness was most common, in 30 (37.5%) patients, followed by dyspnoea on exertion in 23 (28.8%) patients. Pallor was the most common sign, in 69 (86.2%) patients, followed by splenomegaly in 15 (18.8%) patients. Eleven patients had no signs of IDA [Table 1]. All patients in this study had normal renal function test, liver function test, and chest X-ray.
Table 1 Prevalence of symptoms and signs of IDA in study patients

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Hematological and iron profile of study patients

All study patients had anemia, with a mean hemoglobin 7.19 ± 2.1 gm/dL. One (1.2%) patient had thrombocytopenia. All patients had a normal leucocyte count. Microcytosis was found in 74 (94.81%) patients with mean corpuscular volume (MCV) < 82 fL, low mean corpuscular hemoglobin (MCH), and low mean corpuscular hemoglobin concentration (MCHC). Mean serum iron, ferritin, and transferrin saturation were 24.49 ± 6.95 mcg/dL, 17.18 ± 11.62 ng/mL, and 6.62± 2.56%, respectively. These biochemical markers were low in all patients. Mean TIBC was 404.63 ± 69.52 mcg/dL. Seventy four patients (94.81%) had elevated TIBC.

Vitamin A

The mean RBP level of all study patients was 1.04 ± 0.51 µmol/L. Out of 80 IDA patients, 19 (23.8%) had vitamin A deficiency (plasma RBP < 0.7µmol/L) with mean plasma RBP of 0.53 ± 0.13 µmol/L. Among retinol deficient patients, 6 (31.6%) patients each, were in 21 to 30 and 31 to 40 years age, followed by 4 (21%) patients in 16 to 20 years, 2 (10.5%) patients in 41 to 50 years, and 1 (5.3%) patient in 51 to 62 years. Retinol deficient patients consisted of 15 females and 4 males. Twelve patients among these had symptoms of VAD, consisting of 9 (47.3%) patients with dry eyes, 2 (10.5%) patients with dry skin, and 1 (5.2%) patient with both dry eyes and dry skin. Ten (52.6%) patients had conjunctival xerosis, followed by 4 (21.1%) patients with corneal xerosis [Table 2]. No patient had bitot spots and keratomalacia.
Table 2 Prevalence of symptoms and signs in 19 patients of vitamin A deficiency

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Co-relation between vitamin A status, hematological, and iron profile

Among 19 patients with vitamin A deficiency, mean hemoglobin was 6.8 ± 2.14 gm/dL. Mean MCV was 71.35± 8.86 fL. Seventeen patients (98%) had microcytosis with MCV < 82 fL. Mean MCH was 19.43 ± 4.36 pg. Seventeen patients (98%) had low MCH. Mean MCHC was 25.44 ± 4.92 gm/dL. Seventeen patients (98%) had low MCHC. Mean serum iron was 28.21 ± 9.73 mcg/dL. Mean serum ferritin was 13.04 ± 12.41 ng/mL. Mean transferrin saturation was 6.81± 3.07%. Mean TIBC was 427.85 ± 78.57 mcg/dL. Four patients (21%) had normal TIBC level and 15 (79%) had elevated TIBC level [Table 3].
Table 3 Hematological parameters and iron profile in 19 vitamin A deficient patients

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Among these vitamin A deficient patients, RBP levels were correlated with serum iron, serum ferritin, TIBC, and transferrin saturation using scatter plot and Pearson correlation coefficient [Figure 2]. RBP levels (or Retinol) had positive correlation with serum iron and transferrin saturation, with r value of 0.37 and 0.23, respectively [Table 4]. It shows that lower retinol levels were associated with lower serum iron and transferrin saturation. While, negative correlation was seen with serum ferritin and TIBC, with r value of −0.05 and −0.03, respectively. It shows that lower retinol levels were associated with a relatively higher serum ferritin and TIBC levels.
Figure 2 Scatter plots showing correlation of (a) serum iron with RBP (r = 0.37), (b) serum ferritin with RBP (r = −0.05), (c) TIBC with RBP (r = −0.03), (d) transferrin saturation with RBP (r = 0.23).

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Table 4 Correlation of RBP level with iron profile

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  Discussion Top

Deficiency of iron and vitamin A are major public health problems in developing countries, especially affecting children, adolescents, pregnant women, and other females in reproductive age group. Vitamin A deficiency, itself, have also been emerging as a potential cause of iron deficiency, in previous years, as evident by various studies. This study was conducted to assess vitamin A status in patients of IDA and to observe correlation between vitamin A stores and serum biomarkers of iron metabolism in such patients. Eighty patients were enrolled in study with a range of 16 to 62 years and mean age of 31.14 ± 11.33 years. Maximum number of patients (62.4%) were in the age group of 20 to 40 years. Female to male patient ratio was 2.2:1. Higher number of female patients in study is associated with higher prevalence of IDA and VAD in them.[18] All patients had anemia with mean hemoglobin 7.19 ± 2.08 gm/dL. Seventy four patients (94.81%) had low RBC indices with microcytosis and hypochromia. All patients had low values of all serum iron biomarkers except for TIBC, which was elevated in only 74 (94.81%) patients. Sixty six patients had symptoms of IDA, among which, weakness was most common in 30 (37.5%) patients. Pallor was most common sign, present in 69(86.2%) patients.

Mean RBP level in patients was 1.04 ± 0.51 µmol/L. Nineteen (23.8%) out of 80 IDA patients were vitamin A deficient (serum retinol binding protein <0.7 µmol/L). Maximum number of retinol deficient patients, 12 (63.2%) were in 21 to 40 years. Among these, 12 were symptomatic, with dry eye being most common symptom, in 9 (47.3%) patients. Conjunctival xerosis, seen in 10 (52.6%) patients, was most common sign, followed by corneal xerosis. Prevalence of vitamin A deficiency in the study is comparable with the study by Saraiva et al.[19] However, prevalence of vitamin A deficiency is variable in various other studies, based on variations in geographic areas, nutritional status, dietary habits, and age group of study patients.[20],[21],[22],[23],[24]

In our study, plasma RBP level was positively correlated with serum iron and transferrin saturation, whereas it was negatively correlated with serum ferritin and TIBC. Patients who were retinol deficient had lower serum iron and transferrin saturation but elevated serum ferritin and TIBC level, compared to remaining patients with normal vitamin A level. Similar correlation was also seen in various observational studies conducted on patients of different age groups and pregnant females also. Further, various clinical trials and interventional studies have also shown similar correlation and improvement in serum iron and decrease in serum ferritin levels with vitamin A supplementation.[19],[23],[25],[26],[27],[28],[29],[30] This correlation seen in this study is further supportive of the hypothesis that vitamin A deficiency is associated with abnormal trapping of iron in form of ferritin, impaired erythropoiesis, and impaired utilization of iron by erythroblasts in bone marrow. Correlation of vitamin A with serum iron and serum ferritin seen in this study is also evidence towards possible role of vitamin A supplementation in the treatment of iron-deficiency anemia.

Our study had some limitations too. Small sample size being one. Also, effects of vitamin A supplementation on biochemical markers of iron deficiency were not studied.

  Conclusion Top

Iron and vitamin A are two very essential nutrients, needed for a healthy living. Deficiency of these two is seen in all age groups, with predominant distribution in younger population including pregnant females. Deficient patients have variety of symptoms with severe forms including blindness, preterm labor, growth retardation, etc. However, availability of oral supplements of iron and vitamin A helps in treating deficient patients, before they develop irreversible and disabling complications. Vitamin A deficiency is found to be associated with iron deficiency, which makes it necessary to study their inter-relationship, for better management of deficient patients. From this study, we can conclude that vitamin A deficiency affects iron metabolism, causing abnormal iron trapping and systemic iron deficiency, thus worsening clinical profile in IDA. So, from this, we can recommend routine screening of vitamin A status in patients of iron deficiency, so that, such patients can be treated efficiently.

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Conflicts of interest

The authors report no conflicts of interest.

  References Top

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  [Figure 1], [Figure 2]

  [Table 1], [Table 2], [Table 3], [Table 4]


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