Associations Between Serum Transferrin Receptor Concentrations and Erythropoietic Activities According to Body Iron Status

Transcription

1 Annals of Clinical & Laboratory Science, vol. 33, no. 3, Associations Between Serum Transferrin Receptor Concentrations and Erythropoietic Activities According to Body Iron Status Jong Weon Choi and Soo Hwan Pai Department of Laboratory Medicine, College of Medicine, Inha University, Inchon, South Korea Abstract. This study investigated the associations between serum transferrin receptor (stfr) concentrations and erythropoietic activities during 3 stages of iron deficiency in humans. Serum iron markers, fluorescent intensity of reticulocytes, and stfr concentrations were measured in 227 prepubescent children, age 9 to 12 yr. Reticulocyte subpopulations were analyzed by flow cytometry and stfr concentrations were measured by enzyme immunoassay. Mean values of middle-fluorescence reticulocytes (MFR), reticulocyte maturity index (RMI), and stfr concentrations were significantly higher in iron-deficiency anemia subjects than in healthy controls. Reticulocyte subpopulations increased gradually, as body iron status diminished; the mean values of MFR and RMI in subjects with serum ferritin concentrations <4.0 µg/l were 3-fold higher than those in healthy controls (p <0.01). Correlation coefficients of MFR and RMI vs log ferritin values (r = 0.43 and r = 0.42) were higher than those of MFR and RMI vs stfr concentrations (r = 0.24 and r = 0.27) in iron-deficiency anemia subjects. In summary, iron deficiency leads to increased production of immature reticulocytes. Erythropoietic activity is more closely associated with log ferritin values than with stfr concentrations in iron-deficiency anemia. (received 30 April 2003; accepted 15 May 2003) Keywords: serum transferrin receptor, erythropoietic activity, ferritin, reticulocyte subpopulations. Introduction Transferrin receptor (TfR) is a transmembrane protein that mediates iron delivery from the extracellular pool into erythroblasts by receptormediated endocytosis [1]. Nearly all mammalian cells have TfR on their surfaces, but TfR is mostly located in the erythroid precursors in bone marrow [2]. Serum TfR (stfr) concentration is proportional to cellular expression of the membrane-associated TfR and increases with elevated cellular iron needs and cellular proliferation [3]. Thus, measurement of stfr concentration can be used as a diagnostic tool for evaluating body iron status and provides a new laboratory parameter of erythropoiesis [4-6]. Erythropoiesis can be monitored precisely by quantitative measurement of reticulocytes using flow cytometry. The degree of reticulocyte maturation is Address correspondence to Jong Weon Choi, MD, PhD, Dept. of Laboratory Medicine, Inha University Hospital, 7-206, 3- ga, Shinheung-dong, Jung-gu, Inchon, , South Korea; tel ; fax ; jwchoi@inha. ac.kr. determined by the relative concentration of residual RNA, to which the fluorescent intensity of reticulocytes is directly proportional. Reticulocytes can be divided into 3 subpopulations based on fluorescent intensity: low-, middle-, and high-fluorescence reticulocytes (LFR, MFR, and HFR, respectively) [7]. A reticulocyte maturity index (RMI), calculated from the proportion of reticulocyte subpopulations, appears to be the earliest and most sensitive predictor of erythropoiesis [8]. Iron deficiency develops in sequential stages during a period of negative iron balance. These stages include the iron-depletion phase (stage I), irondeficient erythropoiesis (stage II), and irondeficiency anemia (IDA, stage III) [9]. During the iron-depletion phase, iron stores are exhausted; but anemia or decrease of serum iron is not present. In the stage of iron-deficient erythropoiesis, serum iron and serum ferritin levels are decreased; but anemia and hypochromia are still undemonstrable. The relationship between stfr concentrations and iron parameters in IDA or anemia of chronic diseases has been extensively studied, but few studies have closely examined associations between stfr /03/0300/0279 $ by the Association of Clinical Scientists, Inc.

2 280 Annals of Clinical & Laboratory Science concentrations and reticulocyte subpopulations during the 3 stages of iron deficiency, especially as compared to log ferritin values. In the present study, we determined which index most accurately reflects erythropoietic activities during iron deficiency, based on the correlation coefficients of stfr, log ferritin, and stfr/log ferritin ratio (stfr-f index), in respect to each reticulocyte subpopulation. Materials and Methods Complete blood cell count (CBC), iron profiles, reticulocyte subpopulations, and stfr concentrations were measured in 227 prepubescent children, aged 9 to 12 yr (male 51.7%, mean 10.8 yr). The subjects included were all volunteers and were South Korean children from middle-class families. This survey was explained to and approved by the parents and by the directors at each participating school. The subjects were divided into 4 groups based on their body iron status: iron-depletion phase (stage I, n = 41), iron-deficient erythropoiesis (stage II, n = 25), IDA (stage III, n = 72), and healthy controls (n = 89). Non-anemic subjects with a normal serum iron level (>50 µg/dl), but with decreased serum ferritin concentration (<12 µg/l), were classified in the iron-depletion phase (stage I). Iron-deficient erythropoiesis (stage II) was defined as serum ferritin concentration <12 µg/l and serum iron level <50 µg/dl without overt anemia. Subjects showing a decreased serum ferritin concentration, decreased serum iron level, and decreased blood hemoglobin level (<12 g/dl) were considered to have IDA (stage III). The IDA subjects and healthy controls were divided into 4 or 3 subgroups, respectively according to serum ferritin levels: IDA subjects (serum ferritin <4.0 µg/l, n = 16; µg/l, n = 19; µg/ L, n = 17; µg/l, n = 20) and healthy controls (serum ferritin 30.0 µg/l, n = 34; µg/l, n = 31; µg/l, n = 24). Venous blood was drawn in iron-free evacuated tubes. CBC and reticulocyte subpopulations were measured with EDTA-anticoagulated blood within 3 hr after collection. CBC and red cell indices were determined with an electronic counter (SE 9000; Sysmex, Kobe, Japan). Reticulocytes and their subpopulations were analyzed by flow cytometry (R- 3000; Sysmex). The corrected reticulocyte count was calculated, based on a normal hematocrit of 45%, from the following formula: corrected reticulocyte count (%) = (subject s hematocrit/45) x reticulocyte count (%). RMI was calculated from the equation, RMI = [(MFR + HFR) x 100]/LFR and was expressed as the percentage [10]. Serum iron and total ironbinding capacity (TIBC) were assayed with a chemical analyzer (Hitachi 747; Hitachi, Tokyo, Japan) and ferritin was measured by the chemiluminescence method (ACS 180; Chiron, MA, USA). The stfr concentrations were measured by an immunoenzymometric method (IDeA TM stfr, Orion Diagnostica, Espoo, Finland). The intraassay coefficients of variation (CVs, n = 15) for 3 samples (mean stfr, mg/l) were %; the interassay CVs calculated from duplicate results in 10 consecutive runs were %. Data analysis was performed with the SAS 6.12 software package (SAS Institute, Cary, NC). Nonparametric tests were used because the distributions of most variables were non-gaussian by the Kolmogorov-Smirnov test. The Mann-Whitney U test was used to evaluate the differences in mean values between 2 groups. Correlation coefficients were analyzed by Spearman s method. All p values 0.01 were considered statistically significant. Results Mean values of reticulocyte subpopulations and iron parameters based on three stages of iron deficiency are summarized in Table 1. There were no significant differences in reticulocyte parameters between the iron-depletion phase (stage I) and the control group, nor between the iron-depletion phase (stage I) and iron-deficient erythropoiesis (stage II). However, mean proportions of MFR and RMI in IDA (stage III) were 2.3 ± 1.1% and 2.5 ± 1.3%, which were significantly higher than those of the iron-depletion phase (stage I, 0.9 ± 0.6% and 1.0 ± 0.6%), or irondeficient erythropoiesis (stage II, 1.1 ± 0.8% and 1.2 ± 0.7%, p <0.01, respectively). The mean stfr concentrations began to increase significantly at iron-deficient erythropoiesis (stage II) and reached the maximal values at IDA (stage III).

3 Serum transferrin receptor and erythropoiesis 281 Table 1. Mean values of reticulocyte and iron parameters based on 3 stages of iron deficiency. Mean values (mean ± SD; [median]) Iron-depletion Iron-deficient Iron-deficiency Healthy controls phase erythropoiesis anemia (Stage I, n = 41) (Stage II, n = 25) (Stage III, n = 72) (n = 89) Reticulocyte parameters Reticulocytes (%) 0.88 ± 0.23 [0.88] 0.86 ± 0.29 [0.85] 0.91 ± 0.28 [0.91] 0.84 ± 0.25 [0.83] Corrected reticulocytes (%) 0.78 ± 0.22 [0.79] 0.75 ± 0.28 [0.75] 0.64 ± 0.27 [0.65] 0.78 ± 0.23 [0.79] Low-fluorescence reticulocytes (%) 98.9 ± 0.7 [99.1] 98.8 ± 0.8 [98.7] 97.6 ± 2.1 [97.5] 98.9 ± 0.9 [99.1] Middle-fluorescence reticulocytes (%) 0.9 ± 0.6 [0.7] 1.1 ± 0.8 [1.3] 2.3 ± 1.1 [2.2] a,c 0.9 ± 0.8 [0.9] High-fluorescence reticulocytes (%) 0.1 ± 0.2 [0.1] 0.1 ± 0.2 [0.1] 0.1 ± 0.2 [0.1] 0.1 ± 0.1 [0.1] Reticulocyte maturity index (%) 1.0 ± 0.6 [0.9] 1.2 ± 0.7 [1.3] 2.5 ± 1.3 [2.4] a,c 1.1 ± 0.7 [0.9] Iron parameters Serum iron (µg/dl) 117 ± 39 [110] 36 ± 13 [41] a,b 28 ± 12 [32] a 122 ± 49 [119] Total iron-binding capacity (µg/dl) 375 ± 47 [379] 379 ± 49 [373] 451 ± 46 [448] a,c 376 ± 52 [376] Serum ferritin (µg/l) 9.2 ± 4.8 [9.2] a 8.8 ± 3.9 [8.5] a 6.7 ± 2.6 [6.5] a 28.8 ± 13.8 [29.0] Serum transferrin receptor (mg/l) 2.2± 0.8 [2.1] 2.9 ± 1.0 [3.2] a 5.1 ± 1.6 [5.1] a,c 1.8 ± 0.7 [1.5] Blood hemoglobin levels (g/dl) 13.5 ± 0.9 [13.3] 13.1 ± 0.8 [13.4] 10.3 ± 0.9 [10.4] a,c 13.9 ± 0.9 [13.9] a p <0.01 vs healthy controls, computed by Mann-Whitney U test. b p <0.01 vs iron-depletion phase (stage I), computed by Mann-Whitney U test. c p <0.01 vs iron-deficient erythropoiesis (stage II), computed by Mann-Whitney U test. Table 2. Reticulocyte and iron parameters in 72 iron-deficiency anemia subjects, based on serum ferritin concentrations. Serum ferritin concentrations in iron-deficiency anemic subjects (mean ± SD; [median]) <4.0 µg/l µg/l µg/l µg/l (n = 16) (n = 19) (n = 17) (n = 20) Reticulocyte parameters Reticulocytes (%) 0.93 ± 0.29 [0.91] 0.92 ± 0.31 [0.92] 0.89 ± 0.26 [0.89] 0.91 ± 0.24 [0.89] Corrected reticulocytes (%) 0.62 ± 0.21 [0.62] 0.65 ± 0.19 [0.64] 0.64 ± 0.27 [0.65] 0.68 ± 0.13 [0.69] Low-fluorescence reticulocytes (%) 96.7 ±2.4 [96.8] 97.8 ± 1.4 [98.0] 97.9 ± 1.4 [97.8] 98.7 ± 0.8 [98.6] Middle-fluorescence reticulocytes (%) 3.1 ± 1.8 [2.9] a,b 2.1 ± 1.2 [2.0] a 1.9 ± 0.8 [1.9] a 1.1 ± 0.6 [1.1] High-fluorescence reticulocytes (%) 0.2 ± 0.2 [0.2] 0.1 ± 0.2 [0.1] 0.2 ± 0.1 [0.2] 0.2 ± 0.1 [0.3] Reticulocyte maturity index (%) 3.4 ± 1.6 [3.3] a,b 2.3 ± 1.4 [2.3] a 2.2 ± 1.0 [2.2] a 1.3 ± 0.6 [1.3] Iron parameters Serum iron (µg/dl) 23 ± 11 [25] a,b 27 ± 10 [29] 32 ± 12 [36] 33± 12 [34] Total iron-binding capacity (µg/dl) 464 ± 39 [461] 455 ± 43 [453] 451 ± 47 [448] 435 ± 50 [431] Serum ferritin (µg/l) 3.5 ± 0.3 [3.7] a,c 5.2 ± 0.4 [5.1] a,b 7.3 ± 0.5 [6.9] a 10.4 ± 1.2 [10.5] Serum transferrin receptor (mg/l) 7.2± 2.1 [7.1] a,c 4.9 ± 1.8 [4.7] a 4.7 ± 1.6 [4.5] a 3.4 ± 1.5 [3.2] Blood hemoglobin levels (g/dl) 9.6 ± 0.9 [9.2] a,c 10.6 ± 0.8 [10.8] a 10.8 ± 0.9 [11.1] 11.2 ± 0.5 [11.3] a p <0.01 vs anemic subjects with ferritin levels of µg/l, computed by Mann-Whitney U test. b p <0.01 vs anemic subjects with ferritin levels of µg/l, computed by Mann-Whitney U test. c p <0.01 vs anemic subjects with ferritin levels of µg/l, computed by Mann-Whitney U test.

4 282 Annals of Clinical & Laboratory Science Table 3. Mean values of reticulocyte and iron parameters in 89 healthy controls based on serum ferritin concentrations. Serum ferritin concentrations in healthy controls (mean ± SD; [median]) µg/l µg/l 30.0 µg/l (n = 24) (n = 31) (n = 34) Reticulocyte parameters Reticulocytes (%) 0.83 ± 0.23 [0.82] 0.82 ± 0.27 [0.82] 0.84 ± 0.28 [0.84] Corrected reticulocytes (%) 0.75 ± 0.22 [0.74] 0.77 ± 0.21 [0.76] 0.81 ± 0.25 [0.82] Low-fluorescence reticulocytes (%) 99.1 ±0.8 [99.2] 98.9 ± 0.9 [98.9] 99.1 ± 0.9 [99.2] Middle-fluorescence reticulocytes (%) 1.0 ± 0.7 [0.9] 1.1 ± 0.7 [1.0] 0.9 ± 0.6 [0.9] High-fluorescence reticulocytes (%) 0.1 ± 0.2 [0.1] 0.1 ± 0.3 [0.1] 0.1 ± 0.2 [0.1] Reticulocyte maturity index (%) 1.1 ± 0.6 [1.1] 1.1 ± 0.5 [1.0] 1.0 ± 0.7 [1.1] Iron parameters Serum iron (µg/dl) 113 ± 38 [109] 120 ± 37 [118] 139 ± 45 [134] Total iron-binding capacity (µg/dl) 397 ± 50 [392] 378 ± 34 [380] 356 ± 43 [351] Serum ferritin (µg/l) 16.1 ± 4.4 [16.2] a 25.1 ± 7.2 [25.6] a 45.3 ± 12.4 [42.7] Serum transferrin receptor (mg/l) 2.2± 1.0 [1.9] a 1.7 ± 0.9 [1.6] 1.3 ± 0.9 [1.1] Blood hemoglobin levels (g/dl) 13.5 ± 0.9 [13.4] 14.0 ± 0.9 [13.8] 14.4 ± 1.1 [14.6] a p <0.01 vs healthy controls with serum ferritin 30.0 µg/l, computed by Mann-Whitney U test. Table 4. Correlation coefficients of serum ferritin, log ferritin, stfr, and stfr-f index vs reticulocyte parameters in iron-deficiency anemia subjects and healthy controls. Correlation coefficients (r) in prepubescent children Ferritin Log ferritin a stfr b stfr-f index c Iron-deficiency anemic subjects (n = 72) Corrected reticulocytes (%) d 0.21 d 0.22 d Low-fluorescence reticulocytes (%) d d d d Middle-fluorescence reticulocytes (%) 0.39 d 0.43 d 0.24 d 0.32 d High-fluorescence reticulocytes (%) 0.17 d 0.19 d Reticulocyte maturity index (%) 0.37 d 0.42 d 0.27 d 0.31 d Healthy controls (n = 89) Corrected reticulocytes (%) Low-fluorescence reticulocytes (%) Middle-fluorescence reticulocytes (%) High-fluorescence reticulocytes (%) Reticulocyte maturity index (%) a log ferritin = logarithmic values for serum ferritin concentrations. b stfr = serum transferrin receptor concentrations. c stfr-f index = stfr/log ferritin ratio. d statistically significant (p <0.01), analyzed by Spearman s method. Among the iron-deficiency anemia subjects, the children with serum ferritin concentrations of µg/l showed no elevation in the values of reticulocyte subpopulations, compared to the control group. However, when the serum ferritin fell to the level of µg/l, MFR and RMI reached 1.9 ± 0.8% and 2.2 ± 1.0%, which were significantly higher than those (1.1 ± 0.6% and 1.3 ± 0.6%) of the subjects with serum ferritin levels of µg/l (p <0.01, respectively).

5 Serum transferrin receptor and erythropoiesis 283 The mean values of MFR and RMI in the subjects with serum ferritin concentrations <4.0 µg/ L were 3-fold higher than those in healthy controls (p <0.01) (Table 2). Reticulocyte parameters of healthy controls according to serum ferritin concentrations are summarized in Table 3. There were no significant differences in reticulocyte subpopulations between the subjects with serum ferritin 30.0 µg/l and with serum ferritin concentrations of µg/l. As shown in Table 4, no significant correlations were observed between reticulocyte parameters and the values of ferritin, log ferritin, stfr, and stfr-f index in healthy controls. In iron-deficiency anemia subjects, however, the correlation coefficients of LFR (r = , p <0.01), MFR (r = 0.43, p <0.01), and RMI (r = 0.42, p <0.01) vs log ferritin values were higher than those of corresponding reticulocyte subpopulations vs stfr or stfr-f index. Discussion In this study, we investigated the changes in reticulocyte subpopulations and RMI as the body iron store was depleted and we also evaluated the associations between reticulocyte production and stfr concentrations during negative iron balance. We found that erythropoietic activity more strongly correlates with log ferritin values than with stfr concentrations or stfr-f index in iron-deficiency anemic children. Our data for relationships between reticulocyte subpopulations and iron deficiency are in accordance with the results of some investigators, who demonstrated that reticulocyte mean channel fluorescence was elevated in patients with IDA [11]. In a previous study, we reported that immature reticulocyte fractions began to increase from the time that serum iron and ferritin levels both decreased, reaching their peak when the subjects acquired overt IDA [12]. In the present study, no significant elevation of reticulocyte subpopulations was noted in the iron-deficient erythropoietic group. Furthermore, among the subgroups of IDA populations, there was no increase in the mean values of RMI even in subjects with serum ferritin of µg/l; however, RMI began to increase significantly when the serum ferritin fell to the range of µg/l. These results suggest that a serum ferritin concentration <9.0 µg/l may be a critical level, which induces elevated production of immature reticulocytes in children with irondeficiency anemia. The discrepancies between the results of the present and the previous studies may be due to differences in body iron requirement according to subject s age during a period of growth spurt: in the previous work we investigated only female adolescents, whereas in the present study we evaluated prepubescent children. In normal erythropoiesis, reticulocytes become gradually mature red blood cells in the peripheral blood, losing both RNA and TfR [13,14]. Reticulocytes continue to synthesize hemoglobin, provided there is a sufficient supply of both iron and mrna. Iron deficiency restricts hemoglobin synthesis and increases TfR production rate [14]. In the present study, stfr concentrations in iron-deficient erythropoiesis (stage II) were significantly higher than those in healthy controls. However, no significant changes in immature reticulocyte fractions were observed in iron-deficient erythropoiesis (stage II), compared to healthy controls. Similar findings were observed in healthy controls who had normal hemoglobin levels: mean values of stfr concentrations were significantly higher in the subjects with serum ferritin levels of µg/ L than in the subjects with serum ferritin 30.0 µg/ L; however, there were no significant differences in reticulocyte parameters between the 2 groups. These results suggest stfr reflects more sensitively the body iron status than do the reticulocyte subpopulations. Our data imply that decreased ferritin concentration in non-anemic subjects does not affect reticulocyte production; however, once a subject has attained a state of frank anemia, decreased ferritin levels may influence erythropoietic activity. Because serum ferritin reflects the storage iron compartment and stfr reflects the functional iron compartment, the stfr-f index, based on these two values, has been suggested as a good estimate of body iron [15]. Some investigators have used the stfr-f index as an additional biochemical marker for identification of iron-deficient erythropoiesis [16]. In this study, we investigated to what extent stfr concentrations correlate with reticulocyte subpopulations, compared to the values of ferritin, log ferritin, and stfr-f index. In healthy controls,

6 284 Annals of Clinical & Laboratory Science who had no evidence of anemia or iron depletion, there were no significant correlations between reticulocyte parameters and the values of stfr, ferritin, log ferritin, and stfr-f index. Interestingly, in IDA subjects, log ferritin values were more strongly correlated with reticulocyte subpopulations and RMI than serum ferritin, stfr, or the stfr-f index. These results suggest that the stfr or stfr- F index are not superior to log ferritin values for the evaluation of erythropoietic activity during iron deficiency. The main sources of the stfr are known to be the erythroblasts and reticulocytes that eventually shed their receptors into the circulating blood during maturation sequence [17]. In our study, however, significant elevation of stfr concentrations was observed in a considerable numbers of subjects with no increase of reticulocytes or their subpopulations. Therefore, it is conceivable that elevated stfr concentrations in subjects without an increase of reticulocytes may be derived from erythroblasts in the bone marrow. In contrast, one group of investigators has reported that the turnover of erythroblasts was markedly reduced in iron deficiency and the reduction was caused by a progressively decreasing rate of erythroblast proliferation and maturation [18]. Because the present study measured only stfr concentrations and reticulocyte subpopulations in the circulating blood, it does not provide direct evidence of a relationship between intramedullary erythroblasts and stfr concentrations. In conclusion, reticulocytes and their subpopulations appear to be more closely associated with log ferritin values than with stfr concentrations or the stfr-f index. Serum ferritin concentration <9.0 µg/l seems to be a critical level to induce to elevate immature reticulocyte production in iron-deficiency anemic children. To verify the relationships of stfr concentrations and reticulocyte production to intramedullary erythroid precursors, further studies are needed, especially in regard to examination of bone marrow samples. References 1. Feelders RA, Kuiper-Kramer EP, van Eijk HG. Structure, function and clinical significance of transferrin receptors. Clin Chem Lab Med 1999;37: Mast AE, Blinder MA, Gronowski AM, Chumley C, Scott MG. Clinical utility of the soluble transferrin receptor and comparison with serum ferritin in several population. Clin Chem 1998;44: R zik S, Beguin Y. Serum soluble transferrin receptor concentration is an accurate estimating of the mass of tissue receptors. Exp Hematol 2001;29: Cook JD, Dassenko S, Skikne BS. Serum transferrin receptor as an index of iron absorption. Br J Haematol 1990;75: Hubers HA, Beguin Y, Pootrakul P, Einspahr D, Finch CA. Intact transferrin receptors in human plasma and their relation to erythropoiesis. Blood 1990;75: Kohgo Y, Niitsu Y, Kondo H, Kato J, Tsushima N, Sakaki K, Kirayama N, Numata T, Nishisato T, Urushizaki I. Serum transferrin receptor as a new index of erythropoiesis. Blood 1987;70: Chang CC, Kass L. Clinical significance of immature reticulocyte fraction determined by automated reticulocyte counting. Am J Clin Pathol 1997;108: Davis BH, Ornvold K, Bigelow NC. Flow cytometric reticulocyte maturity index: a useful laboratory parameter of erythropoietic activity in anemia. Cytometry 1995;22: Suominen P, Punnonen K, Rajamaki A, Irjala K. Serum transferrin receptor and transferrin receptor-ferritin index identify healthy subjects with subclinical iron deficits. Blood 1998;8: Ataulfo Gonzalez F, Bermejo A, Sanchez J, Anguita E, Villegas A. Value of analyzing reticulocyte subpopulations in polycythemia. Sangre 1997;42: Wells DA, Daigneault-Creech CA, Simrell CR. Effect of iron status on reticulocyte mean channel fluorescence. Am J Clin Pathol 1992;97: Choi JW, Pai SH. Reticulocyte subpopulations and reticulocyte maturity index (RMI) rise as body iron status falls. Am J Hematol 2001;67: Rouault T, Rao K, Harford J, Mattia E, Klausner R. Hemin, chelatable iron, and the regulation of transferrin receptors. J Biol Chem 1985;260: Rao K. Transcriptional regulation by iron of the gene for the transferrin receptor. Mol Cell Biol 1986;6: Punnonen K, Irjala K, Rajamaki A. Serum transferrin receptor and its ratio to serum ferritin in the diagnosis of iron deficiency. Blood 1997;89: Thomas C, Thomas L. Biochemical markers and hematologic indices in the diagnosis of functional iron deficiency. Clin Chem 2002;48: Iacopetta BJ, Morgan EH, Yeoh GCT. Transferrin receptors and iron uptake during erythroid cell development. Biochim Biophys Acta 1982;687: Dormer P, Lau B. Erythropoiesis in iron deficiency. Blut 1977;34: