| Abstract|| |
Background: CD71 or Transferrin receptor is expressed on the surface of erythroid lineage cells. CD71 expression has been found to be significantly increased in rapidly proliferating cells. Methods: This cross-sectional study included 37 bone marrow samples of acute leukemia cases diagnosed between October 2016 to April 2018. The samples were analysed on BD FACS Canto II. We evaluated the expression of CD71 on leukemic blasts and compared median fluorescent intensities (MFI) of blasts in different types of acute leukemias. Results: The 37 cases comprised of 21 Acute Myeloid Leukemia (AML), 13 B-Acute Lymphoblastic Leukemia (B-ALL), 2 T- Acute Lymphoblastic Leukemia (T-ALL) and 1 mixed phenotypic acute leukemia (MPAL), T/Myeloid. CD 71 expression was noted in 70.3% (n= 26/37) of acute leukemia cases. CD71 expression was most commonly observed in AML (n= 15/21;71.4%), followed by B-ALL (n= 9/13;69.2%) and T-ALL (n= 1/2;50%). Single case of MPAL revealed blasts positive for CD71. MFI of leukemic blasts of single CD71 positive T-ALL was found to be highest, followed by AML, MPAL (T/Myeloid) and least in B ALL. Of the AML cases, the blasts of AML-M6, acute promyelocytic leukemia and AML-M1 showed higher CD71 expression in terms of MFI. Conclusions: Surface CD71 expression is not only found in erythroid lineage cells, but also in proliferating cells. CD71 MFI is highest in T lymphoblasts followed by leukemic erythroblasts, myeloblasts and least in B lymphoblasts.
Keywords: Acute leukemia, acute lymphoblastic leukemia, acute myeloid leukemia, CD71, median fluorescent intensity, mixed phenotypic acute leukemia
|How to cite this article:|
Acharya S, Kala PS. Role of CD71 in acute leukemia– An immunophenotypic marker for erythroid lineage or proliferation?. Indian J Pathol Microbiol 2019;62:418-22
|How to cite this URL:|
Acharya S, Kala PS. Role of CD71 in acute leukemia– An immunophenotypic marker for erythroid lineage or proliferation?. Indian J Pathol Microbiol [serial online] 2019 [cited 2019 Aug 21];62:418-22. Available from: http://www.ijpmonline.org/text.asp?2019/62/3/418/263492
| Introduction|| |
CD71 or Transferrin receptor (TfR) is a transmembrane homodimeric glycoprotein with 760 amino acids, expressed on the surface of erythroid lineage cells, where it mediates the uptake of transferrin-iron complex by endocytosis. Two TfRs have been identified- TfR1 and TfR2. Of the two, TfR1 is the main protein responsible for iron uptake because of its greater affinity. CD71 expression is highest in early erythroblasts and tends to decrease after intermediate erythroblasts through reticulocyte stage. It is significantly downregulated in dyserythropoiesis. CD71 expression has been found to be significantly increased in rapidly proliferating cells, hence, its role in defining leukemic blasts on flow cytometry. The role of CD71 expression by flow cytometry has been widely studied in diverse hematological diseases ranging from Myelodysplastic syndrome (MDS) to leukemias and lymphomas.,, We evaluated the expression of CD71 on leukemic blasts and compared median fluorescent intensities (MFI) of blasts in different types of acute leukemia.
| Materials and Methods|| |
This is a cross-sectional retrospective study conducted in a tertiary care hospital located in Uttarakhand, India. The ethical committee of the institute permitted the study. An informed consent was taken from all patients. A total of 37 bone marrow samples of acute leukemia cases diagnosed between October 2016 and April 2018, where flow cytometry analysis was available, were included in the study. All cases of acute leukemia under treatment were excluded from the study. For flow cytometric immunophenotyping, a minimum of 3 ml bone marrow aspirate sample was collected with EDTA as anticoagulant. The processing was done within 24 h with stain-lyse-wash method. The antibody panel for flow cytometric analysis of acute leukemia included the following.
The antibodies, Flourescence-activated cell sorting (FACS) lyse solution, and Perm II permeabilisation buffer were obtained from Becton Dickinson Biosciences (BD), USA. The samples were analyzed on BD FACS Canto II, 8 color flow cytometer using FACS Diva software. The gating of leukemic blasts, lymphocytes, and erythroblasts was done on CD45/Side scatter(A) plots. Leukemic blasts were deemed positive for CD 71, if 20% of cells showed its expression. The MFIs of CD71 APC (allophycocyanin) H7 of leukemic blasts, lymphocytes, and erythroblasts were recorded. The statistical analysis was performed with SPSS (version 18.0, SPSS, Chicago, IL).
| Results|| |
Of the 37 cases, 21 were acute myeloid leukemia (AML), 13 were B-acute lymphoblastic leukemia (B-ALL), two were T-acute lymphoblastic leukemia (T-ALL), and one was mixed phenotypic acute leukemia (MPAL), T/Myeloid [Table 1] and [Figure 1], [Figure 2], [Figure 3]. CD 71 expression was noted in 70.3% (n = 26/37) of all cases of acute leukemia. CD71 expression was most commonly observed in AML (n = 15/21;71.4%), followed by B-ALL (n = 9/13;69.2%) and T-ALL (n = 1/2;50%). A single case of MPAL revealed blasts positive for CD71.
|Table 1: Flow cytometric diagnosis in acute leukemia and corresponding Mean MFI (Median fluorescent intensity) of blasts for CD71|
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|Figure 1: Acute myeloid leukemia. Dim CD45 and low to moderate SSC cells (Blasts) are positive for cytoplasmic MPO (heterogenous), CD34 (heterogenous), and CD71. In other plots, these cells showed positive expression for CD13, CD33, CD117, and did not express any of the B and T lineage markers|
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|Figure 2: B-acute lymphoblastic leukemia. Dim CD45 and low SSC cells (Blasts) expressing CD19 (bright), CD34, and CD71. In other plots, blasts expressed CD10 (heterogenous) and were negative for MPO and T lineage markers|
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|Figure 3: T-acute lymphoblastic leukemia. Dim CD45 and low SSC cells (Blasts) express cytoplasmic CD3, CD5, and CD71. In other plots, blasts expressed CD7 and CD34 (dim)|
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However, the MFI of leukemic blasts of single CD71 positive T-ALL was found to be highest, followed by AML and MPAL (T/Myeloid). B-ALL blasts showed the least MFI for CD71.
Among the AML cases, the blasts of AML-M6, acute promyelocytic leukemia. and AML-M1 showed higher CD71 expression in terms of MFI. The expression pattern of CD 71 by leukemic blasts in AML subtypes is tabulated in [Table 2].
|Table 2: Acute myeloid leukemia, CD71 expression with respect to subtypes|
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The mean MFI of lymphocytes and normal erythroblasts was 159.9 and 43,056.3, respectively.
| Discussion|| |
CD71 or TfR binds to diferric transferrin, which is then internalized, and diferric iron is released in presence of endosomal acidification., The high expression of TfRs is present on placental syncytiotrophoblasts, myocytes, basal keratinocytes, hepatocytes, pancreas (endocrine), spermatocytes, and erythroid precursors. In the erythroid series, the highest expression is noted in early erythroid precursors. The expression declines during the maturation process, in parallel with the decline in the capacity for hemoglobin synthesis. CD71 is an excellent marker for erythroid series in bone marrow and has been used in the immunophenotyping in leukemia as a lineage marker for the diagnosis of erythroid leukemia. The expression of CD71 in erythroid series reduces in cases of MDS with dysplastic erythroblasts.,
CD71 is over expressed in acute leukemia. In the present study, 70.4% of all cases of acute leukemia demonstrated CD71 positivity. CD71 plays an important role in controlling cell proliferation because iron is required for ribonucleotide reductase activity. Latter is the sole enzyme catalyzing the conversion of ribonucleotides to deoxyribonucleotides.
The present study demonstrates that CD71 expression is seen more commonly in cases of AML followed by B-ALL and T-ALL. Pande et al. reported that CD 71 expression is seen most commonly in AML and least in B-ALL [
[Table 3]. Wei et al. reported expression rates to be highest in T ALL followed by AML- M1 and M2, B-ALL, APML, and AML-M5. Hence, among the acute myeloid leukemias, the higher expression of CD71 is seen in the minimally differentiated AML, AML without maturation, and acute myelomonocytic leukemia as compared to the partially differentiated ones, including acute promyelocytic and acute monocytic leukemia., Present study also revealed similar results. It was observed that highest percentage positive CD71 expression was noted in AML with minimal differentiation, AML without maturation, and acute erythroid leukemia followed by AML with maturation and acute myelomonocytic leukemia. Acute monocytic leukemia and acute promyelocytic leukemia showed minimum percentage positivity. However, this finding was not statistically significant, possibly because of lesser number of cases in present study (P > 0.05).
The present finding of higher MFI for CD71 in T-lymphoblasts than the B-lymphoblasts correlates with the earlier studies. In addition, MFI for CD71 expression was found to be highest in T-ALL followed by AML and least in B-ALL in the present study. Similar observations have been reported in previous studies.,
The erythroblasts in erythroleukemia usually lack myeloid associated markers including myeloperoxidase. Markers commonly used for erythroleukemia are glycophorin A, CD 71, and CD 36. Blasts in some of the cases of erythroleukemia can be negative for glycophorin A. CD 36 is a non-specific marker for AML-M6. In erythroleukemia, the erythroblasts usually express CD71, but occasionally CD71 expression can be aberrantly dim. In the present study, the blasts of pure erythroid leukemia expressed CD71, but the MFI of these leukemic blasts (MFI = 3897) was lower in comparison with the overall mean MFI of the normal erythroblasts (mean MFI = 43,056.3). Thus, CD71 expression level, in terms of MFI, is lower in leukemic erythroblasts.
It is well-known that the erythroid cells express a higher level of CD71 than other lineage cells. We found the CD71 expression of the admixed normal erythroid population to be higher than all the leukemic blasts, which is in concordance with the earlier studies.
El-Menshawy et al. reported that CD71 expression in ALL is associated with an adverse clinical outcome and a higher relapse rate. CD71 can serve as a surrogate marker for cell proliferation activity in hematological malignancies and can replace Ki-67. The former can be easily and rapidly determined by flow cytometry. The expression of CD71 by leukemic blasts may be useful in detection of minimal residual disease. However, this needs to be evaluated by more studies.
The TfR1 has been demonstrated to be a direct transcriptional activator of c-Myc proto-oncogene. It has been demonstrated that monoclonal antibody against TfR1 can induce apoptosis in T-lymphocytes in CD71 positive T-ALL cases. Hence, it has been proposed that CD71 can also serve as a therapeutic target in acute leukemias, which needs to be investigated.
Apart from its role in precursor cell neoplasms, the role of CD71 expression in certain lymphomas is being evaluated. Glasova et al. reported that the proliferation index of CD71 positive subpopulation of Burkitt's lymphoma cells was higher than CD71 negative subpopulation.
| Conclusion|| |
In summary, surface CD71 expression is not only found in erythroid lineage cells but also in proliferating cells. Hence, CD71 is also an immunophenotypic marker for cell proliferation. CD71 MFI is highest in T-lymphoblasts followed by leukemic erythroblasts, myeloblasts, and least in B-lymphoblasts.
The authors want to acknowledge Mr Hemant Saini and Ms Kavita Chamoli for technical support throughout the study.
Contributors and authorship
PSK and SA have contributed equally in conception of idea, drafting of manuscript, literature search, data analysis, statistical analysis, manuscript editing, and review.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Nakahata T, Okumura N. Cell surface antigen expression in human erythroid progenitors: Erythroid and megakaryocytic markers. Leuk Lymphoma 1994;13:401-9.
Sargent PJ, Farnaud S, Evans RW. Structure/function overview of proteins involved in iron storage and transport. Curr Med Chem 2005;12:2683-93.
Yan ZY, Tian X, Li Y, Yang MR, Zhang S, Wang XM,et al
. Changes of CD34(+) and CD71(+) CD45(-) cell levels in bone marrow of MDS and AA patients. Zhongguo Shi Yan Xue Ye Xue Za Zhi2014;22:382-6.
Della Porta MG, Malcovati L, Invernizzi R, Travaglino E, Pascutto C, Maffioli M,et al
. Flow cytometry evaluation of erythroid dysplasia in patients with myelodysplastic syndrome. Leukemia 2006;20:549-55.
Liu Q, Wang M, Hu Y, Xing H, Chen X, Zhang Y,et al
. Significance of CD71 expression by flow cytometry in diagnosis of acute leukemia. Leuk Lymphoma 2014;55:892-8.
Lawrence CM, Ray S, Babyonyshev M, Galluser R, Borhani DW, Harrison SC. Crystal structure of the ectodomain of human transferrin receptor. Science 1999;286:779-82.
Marsee DK, Pinkus GS, Yu H. CD71 (Transferrin Receptor) An effective marker for erythroid precursors in bone marrow biopsy specimens. Am J Clin Pathol 2010;134:429-35.
Ponka P, Lok CN. The transferrin receptor: Role in health and disease. Int J Biochem Cell Biol1999;31:1111-37.
Davis BH, Holden JT, Bene MC, Borowitz MJ, Braylan RC, Cornfield D,et al
. 2006 Bethesda International Consensus recommendations on the flow cytometric immunophenotypic analysis of hematolymphoid neoplasia: Medical indications. Cytometry B Clin Cytom 2007;72:S5-13.
Malcovati L, Della Porta MG, Lunghi M, Pascutto C, Vanelli L, Travaglino E,et al
. Flow cytometry evaluation of erythroid and myeloid dysplasia in patients with myelodysplastic syndrome. Leukemia 2005;19:776-83.
El-Menshawy N, Abd-Aziz SM, Ebrahim MA, Nader El-Malky. Impact of cellular CD71 (transferrin receptor 1) expression in Egyptian acute leukemia: Correlation with clinicopathologic features. Comp Clin Pathol 2016;25:511.
Testa U, Pelosi E, Peschle C. The transferrin receptor. Crit Rev Oncog1993;4:241-76.
Pande A, Dorwal P, Jain D, Tyagi N, Mehra S, Sachdev R, et al
. Expression of CD71 by flow cytometry in acute leukemias: More often seen in acute myeloid leukemia. Indian J Pathol Microbiol 2016;59:310-3.
] [Full text]
Wei YY, Zhang XZ, Zhang F, Li ZB, Wang NN, Liu S,et al
. Expression of CD71 on cell proliferation in hematologic malignancy and its correlation with Ki-67. Zhongguo Shi Yan Xue Ye Xue Za Zhi 2015;23:234-40.
Koehler M, Behm F, Hancock M, Pui CH. Expression of activation antigens CD38 and CD71 is not clinically important in childhood acute lymphoblastic leukemia. Leukemia 1993;7:41-5.
Masood A, Holkova B, Chanan-Khan A. Erythroleukemia: Clinical course and management. Clin Adv Hematol Oncol 2010;8:288-90.
Zuo Z, Polski JM, Kasyan A, Medeiros LJ. Acute Erythroid Leukemia. Arch Pathol Lab Med2010;134:1261-70.
O'Donnell KA, Yu D, Zeller KI, Racke F, Thomas-Tikhonenko A, Dang CV,et al
. Activation of transferrin receptor 1 by c-Myc enhances cellular proliferation and tumorigenesis. Mol Cell Biol 2006;26:2373-86.
Moura IC, Lepelletier Y, Arnulf B, England P, Baude C, Beaumont C,et al
. A neutralizing monoclonal antibody (mAb A24) directed against the transferrin receptor induces apoptosis of tumor T lymphocytes from ATL patients. Blood 2003;103:1838-45.
Glasová M, Koníková E, Stasáková J, Babusíková O. The relationship of HLA-DR, CD38 and CD71 markers to activation, proliferation and differentiation of some human leukemia and lymphoma cells. Neoplasma 1998;45:88-95.
Pooja Sharma Kala
Department of Pathology, Sri Guru Ram Rai Institute of Medical and Health Sciences, Dehradun, Uttarakhand
Source of Support: None, Conflict of Interest: None
[Figure 1], [Figure 2], [Figure 3]
[Table 1], [Table 2], [Table 3]