Open Access

VSTM-v1, a potential myeloid differentiation antigen that is downregulated in bone marrow cells from myeloid leukemia patients

  • Min Xie1,
  • Ting Li2,
  • Ning Li1,
  • Jinlan Li1,
  • Qiumei Yao1,
  • Wenling Han2 and
  • Guorui Ruan1Email author
Contributed equally
Journal of Hematology & Oncology20158:25

https://doi.org/10.1186/s13045-015-0118-4

Received: 27 January 2015

Accepted: 10 February 2015

Published: 15 March 2015

Abstract

Leukocyte differentiation antigens often represent important markers for the diagnosis, classification, prognosis, and therapeutic targeting of myeloid leukemia. Herein, we report a potential leukocyte differentiation antigen gene VSTM1 (V-set and transmembrane domain-containing 1) that was downregulated in bone marrow cells from leukemia patients and exhibited a higher degree of promoter methylation. The expression level of its predominant encoded product, VSTM1-v1, was positively correlated with myeloid cell maturation state. Restoration of VSTM1-v1 expression inhibited myeloid leukemia cells’ growth. Therefore, VSTM1-v1 might represent an important myeloid leukocyte differentiation antigen and provide a potential target for the diagnosis and treatment of leukemia.

Keywords

Acute myeloid leukemiaLeukocyte differentiation antigenVSTM1MethylationBiomarker

To the Editor

Acute myeloid leukemia (AML) and chronic myeloid leukemia (CML) are myeloid blood cell malignancies that show great heterogeneity. VSTM1 (V-set and transmembrane domain-containing 1) encodes a potential leukocyte differentiation antigen that is highly expressed in myeloid cells, but silenced in multiple leukemia cell lines [1]. To determine whether it plays a role in leukemogenesis, we characterized its expression pattern and function in bone marrow cells from AML/CML patients and myeloid leukemia cell lines.

We measured VSTM1 expression in leukemia cell lines and bone marrow biopsies from leukemia patients using qRT-PCR. VSTM1 was downregulated or silenced in all cell lines tested (Additional file 1: Table S1). Compared to healthy donors (HDs), VSTM1 was downregulated in AML (Table 1). Additionally, in CML-AP/BC (accelerated phase/blast crisis), which clinically behaves like AML, VSTM1 expression levels were much lower than those in CML-CP (chronic phase, P = 0.003, Table 1). Analogous protein expression differences were found by Western blotting. Compared to HD bone marrow, VSTM1 was similarly expressed in CML-CP patients, whereas it was barely detectable in untreated AML patients. However, in AML patients who achieved complete remission, VSTM1 expression was completely restored (Additional file 2: Figure S1). Higher levels of VSTM1 promoter methylation in bone marrow cells from AML patients compared to those from HDs were observed, which might contribute to its downregulation (Additional file 3: Figure S2).
Table 1

The expression level of VSTM1 in bone marrow cells from leukemia patients and healthy donors

Groups

MIC

Sample size

Mean ratio, VSTM1 / ABL

P value a

HD

 

36

17.358 ± 17.904

 

Untreated AML

 

145

4.333 ± 7.895

<0.001

 

M1

3

0.374 ± 0.619

n.a.

 

M2

72

5.292 ± 10.171

<0.001

 

M3

29

2.884 ± 3.960

<0.001

 

M4

24

4.814 ± 5.293

0.001

 

M5

14

3.052 ± 4.867

<0.001

 

M6

3

1.417 ± 2.205

n.a.

Untreated ALL

 

40

0.381 ± 0.755

<0.001

Untreated CML

 

57

5.479 ± 8.266

<0.001

 

CP

38

7.743 ± 9.312

0.001

 

AP/BC

19

0.950 ± 1.367

<0.001

MIC morphological, immunological, and cytogenetic classifications, n.a. not available due to a small sample size.

aThe P value was calculated using Wilcoxon signed ranks test as compared to the HD group.

VSTM1-v1 is the most abundantly expressed gene product encoded by VSTM1 [2] and is an ITIM-bearing immune receptor that negatively regulates neutrophil activity [3-5]. We used flow cytometry to divide bone marrow cells into subpopulations based on CD45 levels and SSC [6], and found that VSTM1-v1 was highly expressed in mature granulocytes and monocytes from HDs (94.01 ± 6.80% positive with a mean fluorescence intensity (MFI) of 167.61 ± 90.95), but was much less abundant in naive cells (37.14 ± 14.47% with a MFI of 46.70 ± 23.51, n = 27; P <0.001). Moreover, the percentage of VSTM1-v1-positive cells among naive cells from AML patients (19.60 ± 21.09%, n = 52, including 4 M0/M1, 22 M2, 4 M3, 14 M4, and 8 M5) was even lower than that in HDs (P = 0.047). Therefore, we speculated that VSTM1-v1 expression might be associated with the maturity of myeloid cells. A combination of CD34/CD117/CD13/CD16 staining showed that VSTM1-v1 expression in myeloid cells was positively correlated with cell maturation state. Differences between any two continuous stages were significant (n = 11; P < 0.001; Table 2 and Additional file 4: Figure S3). Using CD16 and CD14 as phenotypic markers for mature granulocytes and monocytes, respectively, we found a similar result (Additional file 1: Table S2). This correlation was subsequently confirmed by increased VSTM1 expression in bone marrow cells from APL patients and in NB4 cells after ATRA treatment in vitro (Additional file 5: Figure S4). These findings provide a potential reason why VSTM1 expression levels were reduced so markedly in AML and CML-AP/BC patients.
Table 2

The expression of VSTM1-v1 at various stages of myelocytic differentiation

 

Myeloblasts

Promyelocytes

Myelocytes

Metamyelocytes

CD markers

CD34+

CD34, CD117+, CD16

CD34, CD117, CD16, CD13+

CD16+

VSTM1-v1-positive cells (%)

5.69 ± 2.69

11.48 ± 4.58

35.02 ± 11.58

61.63 ± 8.53

P valuea

<0.001

<0.001

<0.001

 

aThe P value was calculated using a Wilcoxon signed ranks test as compared with the next stage.

Similar to its function in Jurkat cells [1], restoration of VSTM1-v1 expression in the myeloid leukemia cell lines K562 and MEG-01 also inhibited cell growth (Additional file 6: Figure S5). Moreover, when searching for clinical features that could be related to VSTM1 expression in AML patients, we detected a higher expression level of VSTM1 in AML1-ETO-positive patients (Additional file 1: Table S3 and Additional file 7: Figure S6). This chimeric oncogene in AML is often associated with a relatively favorable prognosis [7,8]. Considering the inhibitory effect of VSTM1-v1 on leukemia cell growth, we can hypothesize that VSTM1-v1 might enhance the cytotoxic effects of chemotherapeutics in patients with this genetic abnormality.

In conclusion, our findings suggest that VSTM1-v1 might be an important myeloid leukocyte differentiation antigen. Our elucidation of its expression pattern throughout myeloid cell differentiation and its effect on leukemia cell growth could help to establish it as a novel target for the development of diagnostics and treatments for myeloid leukemia.

Notes

Abbreviations

AML: 

Acute myeloid leukemia

CML: 

Chronic myeloid leukemia

CP: 

Chronic phase

AP: 

Accelerated phase

BC: 

Blast crisis

ALL: 

Acute lymphocytic leukemia

APL: 

Acute promyelocytic leukemia

ATRA: 

All-trans retinoic acid

WBC: 

White blood cell

Hb: 

Hemoglobin

Plt: 

Blood platelet count

Declarations

Acknowledgements

This work was supported by grants from the Beijing Municipal Natural Science Foundation [grant number 7122199], the Specialized Research Fund for the Doctoral Program of Higher Education of China [grant number 20130001110079], the National Basic Research Program of China [grant number 2013CB733701], and the National Natural Science Foundation of China [grant number 81170484].

Authors’ Affiliations

(1)
Peking University People’s Hospital and Institute of Hematology
(2)
Peking University Center for Human Disease Genomics, Department of Immunology, Key Laboratory of Medical Immunology, Ministry of Health, School of Basic Medical Sciences, Peking University Health Science Center

References

  1. Li T, Guo XH, Wang WY, Mo XN, Wang PZ, Han WL. V-set and transmembrane domain-containing 1 is silenced in human hematopoietic malignancy cell lines with promoter methylation and has inhibitory effects on cell growth. Mol Med Rep. 2015;11:1344–51.PubMedGoogle Scholar
  2. Guo X, Zhang Y, Wang P, Li T, Fu W, Mo X, et al. VSTM1-v2, a novel soluble glycoprotein, promotes the differentiation and activation of Th17 cells. Cell Immunol. 2012;278:136–42.View ArticlePubMedGoogle Scholar
  3. Steevels TA, Lebbink RJ, Westerlaken GH, Coffer PJ, Meyaard L. Signal inhibitory receptor on leukocytes-1 is a novel functional inhibitory immune receptor expressed on human phagocytes. J Immunol. 2010;184:4741–8.View ArticlePubMedGoogle Scholar
  4. Steevels TA, van Avondt K, Westerlaken GH, Stalpers F, Walk J, Bont L, et al. Signal inhibitory receptor on leukocytes-1 (SIRL-1) negatively regulates the oxidative burst in human phagocytes. Eur J Immunol. 2013;43:1297–308.View ArticlePubMedGoogle Scholar
  5. Van Avondt K, Fritsch-Stork R, Derksen RH, Meyaard L. Ligation of signal inhibitory receptor on leukocytes-1 suppresses the release of neutrophil extracellular traps in systemic lupus erythematosus. PLoS One. 2013;8:e78459.View ArticlePubMed CentralPubMedGoogle Scholar
  6. Borowitz MJ, Guenther KL, Shults KE, Stelzer GT. Immunophenotyping of acute leukemia by flow cytometric analysis. Use of CD45 and right-angle light scatter to gate on leukemic blasts in three-color analysis. Am J Clin Pathol. 1993;100:534–40.PubMedGoogle Scholar
  7. Grimwade D, Walker H, Harrison G, Oliver F, Chatters S, Harrison CJ, et al. The predictive value of hierarchical cytogenetic classification in older adults with acute myeloid leukemia (AML): analysis of 1065 patients entered into the United Kingdom Medical Research Council AML11 trial. Blood. 2001;98:1312–20.View ArticlePubMedGoogle Scholar
  8. Schnittger S, Weisser M, Schoch C, Hiddemann W, Haferlach T, Kern W. New score predicting for prognosis in PML-RARA+, AML1-ETO+, or CBFBMYH11+ acute myeloid leukemia based on quantification of fusion transcripts. Blood. 2003;102:2746–55.View ArticlePubMedGoogle Scholar

Copyright

© Xie et al.; licensee BioMed Central. 2015

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0) which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

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