Blood, 15 July 2001, Vol. 98, No. 2, pp. 486-488
BRIEF REPORT
Low frequency of BCL-2/JH translocation in
peripheral blood lymphocytes of healthy Japanese
individuals
Masaki Yasukawa,
Shiro Bando,
Gottfried Dölken,
Eiji Sada,
Yoshihiro Yakushijin,
Shigeru Fujita, and
Hideichi Makino
From the First Department of Internal Medicine, and
Department of Clinical Laboratory Medicine, Ehime University School of
Medicine, Ehime, Japan; Klinik und Poliklinik für Innere
Medizin C, Hämatologie und internistische Onkologie,
Ernst-Moritz-Arndt-Universität, Greifswald, Germany; and
Department of Medical Technology, Ehime Medical College of Health
Science, Ehime, Japan.
 |
Abstract |
The incidence of follicular lymphoma differs significantly between
white and Japanese individuals. Translocation between the BCL-2 and immunoglobulin heavy chain genes is detected in
85% to 90% of all follicular lymphomas in whites. Recently,
BCL-2/JH translocation was detected in
peripheral blood lymphocytes from more than 50% of healthy white
individuals. To clarify the reason for the difference in incidence of
follicular lymphoma between whites and Japanese, the frequency of
BCL-2/JH translocation in peripheral blood
lymphocytes of healthy Japanese individuals was compared with that of
German individuals. The prevalence of BCL-2/JH translocation in Japanese adults appeared to be significantly lower than that in German adults. The present data suggest that the low frequency of BCL-2/JH translocation in
the Japanese general population may be one of the major reasons for the
difference in incidence of follicular lymphoma between whites and Japanese.
(Blood. 2001;98:486-488)
© 2001 by The American Society of Hematology.
| |
Introduction |
The incidence of follicular lymphoma differs
between white and Japanese individuals, accounting for about 40% of
all non-Hodgkin lymphomas in whites, compared with less than 10% in
Japanese.1-5 The age-adjusted incidence of follicular
lymphoma (per 100 000) is reported to be 3.8 in the United States and
0.5 in Japan.2 The reason for this difference between the
2 populations is unknown.
The translocation t(14;18)(q32;q21) between the BCL-2
proto-oncogene and the JH immunoglobulin gene
region is detected in 85% to 90% and approximately 50% of all
follicular lymphomas in whites and Japanese,
respectively.5-10 This leads to overexpression of BCL-2,
conferring a growth advantage on the neoplastic cells.11 Recently, it has been reported that BCL-2/JH
translocation is frequently detected in peripheral blood lymphocytes of
healthy white individuals.12-17 In the multihit theory of
tumorigenesis, translocation of the BCL-2 gene is considered
to be the first somatic mutation, and additional mutations are needed
for development of follicular lymphoma. From this viewpoint, the low
incidence of follicular lymphoma in Japanese individuals may be due to
the low frequency of BCL-2/JH translocation in
the general population or other factors affecting lymphomagenesis.
Accordingly, we compared the frequency of
BCL-2/JH translocation in peripheral blood
lymphocytes of Japanese individuals with that of German individuals.
The results showed that the incidence of
BCL-2/JH translocation in healthy Japanese
individuals is significantly lower than that in German individuals.
| |
Study design |
Peripheral blood mononuclear cells were obtained with
informed consent from Japanese and German (white) individuals who had no serious diseases, and the DNA was extracted using standard procedures. The DNA samples from German individuals were sent to Japan
and used for experiments. The presence of the
BCL-2/JH translocation in the major breakpoint
region (MBR) of the BCL-2 gene was examined using a nested
polymerase chain reaction (PCR), as described previously in a study of
white individuals.12,14 Briefly, 10 µg DNA corresponding
to approximately 1 to 2 × 106 cells in 100 µL PCR
mixture was used for each PCR analysis. The first round of
amplification was performed for 30 cycles using the outer primers
5'-ACCTGAGGAGACGGTGACCAGGGT-3' for the JH region and 5'-CAGCCTTGAAACATTGATGG-3' for BCL-2 (MBR). The second
round was performed for 30 cycles using the inner primers
5'-CAGGGTCCCTTGGCCCCAG-3' for the JH region and
5'-TATGGTGGTTTGACCTTTAG-3' for BCL-2. Positive and negative
controls were included in all experiments. The PCR products were loaded
on 2% agarose electrophoresis gel containing ethidium bromide and
visualized under UV light. Southern blot analysis was also performed
using an internal oligonucleotide (5'-CACAGACCCACCCAGAGCCCTCCTG-3') as
a probe. The PCR analysis of DNAs from both Japanese and German
individuals was performed by a single investigator. To confirm the
specificity of the PCR products and to determine the breakpoints, the
nucleotide sequences of the DNA fragments amplified from the samples
were analyzed by direct sequencing of the PCR products using an ABI
PRISM 310 Genetics Analyzer (PerkinElmer, Norwalk, CT).
The sensitivity of this nested PCR method, determined using the
t(14;18)-positive lymphoma cell lines TK,18 FL218, and
FL318,19 appeared to be approximately 1/106
cells, and was almost the same as that reported previously in studies
of white individuals.12-17 Statistical analysis between the 2 groups was performed using the 
2 test.
| |
Results and discussion |
The frequencies of BCL-2/JH translocation
examined using peripheral blood lymphocytes of 241 healthy Japanese and
75 healthy German individuals were 16.2% and 52.0%, respectively
(Table 1). The frequency of
BCL-2/JH translocation in Japanese individuals aged 20 to 79 years appeared to be significantly lower than that in
age-matched German individuals (P < .0001). In Japanese
individuals, the frequency of BCL-2/JH
translocation increased dramatically at around 20 years of age. In
addition, in contrast to previous reports that the frequency of
BCL-2/JH translocation rises with age in white
individuals,12,14 no significant difference was detected
between young, middle-aged, and elderly Japanese adult groups. On the
other hand, the frequency of BCL-2/JH
translocation in German individuals examined in the present study was
almost as high as that reported previously in studies of white
individuals, including Germans.12-17
Another noteworthy finding in the present study was that more
than one distinguishable BCL-2/JH amplified DNA
fragment, indicating the presence of oligoclonal lymphocytes with
t(14;18) in the samples, was detected in only 6 of 39 (15.4%) Japanese
positive samples and 17 of 39 (43.6%) German positive samples (Table 1
and Figure 1). Figure 1 gives data for
ethidium bromide staining of the agarose electrophoresis gel of
positive samples. Southern blot analysis and direct sequencing of the
PCR products confirmed that the amplified DNA fragments were the fusion
gene of BCL-2 and the immunoglobulin JH region, and that breakpoints in the
BCL-2 MBR region tended to fall within 3 microclusters, as
reported previously in whites (data not shown). The frequency of
oligoclonal t(14;18)-positive lymphocytes in the Japanese samples was
significantly lower than that in the German samples
(P < .01). This finding also indicates that the frequency
of BCL-2/JH translocation in peripheral blood lymphocytes of Japanese individuals is lower than that in German individuals.
View larger version (55K):
[in this window]
[in a new window]
| Figure 1.
Agarose gel electrophoresis of positive samples.
The nested PCR was performed as described in "Study design," and
the PCR products were loaded on a 2% agarose electrophoresis gel
containing ethidium bromide and visualized under UV light. (A) Japanese
samples showing a single band (lanes 1-33). (B) Japanese samples
showing multiple bands (lanes 1-6). (C) German samples showing a single
band (lanes 1-22). (D) German samples showing multiple bands (lanes
1-17). Lane M shows 100-bp ladder markers.
|
|
To our knowledge, only one previous report has compared
BCL-2/JH translocation in white and Japanese
individuals. That study showed no significant difference between the
frequencies of BCL-2/JH translocation in
hyperplastic lymphoid tissues obtained from 15 American and 10 Japanese
patients.20 The discrepancy between that study and the
present one may be due to the difference in the sources of materials
and the numbers of samples examined.
In comparison with the frequency found in German samples and the
previous reports indicating that BCL-2/JH
translocation was detected in more than 50% of
whites,12-17 the prevalence in Japanese individuals
detected in the present study is significantly low. This low frequency
of BCL-2/JH translocation in the Japanese
general population may be one of the major reasons for the difference in incidence of follicular lymphoma between whites and Japanese. The
present findings also suggest that the differences in instability of
certain chromosome regions may account for the differences in incidence
of various cancers between races. The reason for the different
incidence of BCL-2/JH translocation between
Japanese and white individuals is unknown. A recent study has shown
that the incidence of t(14;18) in healthy individuals might be affected by smoking.21 These data suggest that the differing
incidence of BCL-2/JH translocation between
whites and Japanese may be due to environmental as well as genetic
factors, as has been discussed in the case of inv(7), which is detected
at a higher incidence in agriculture workers.22 An
increased incidence of inv(7) or t(14;18) in a population or a subgroup
may be a measure of not only genomic instability, but also the V(D)J
recombinase activity possibly generating these chromosomal translocations.
| |
Acknowledgments |
We thank Drs K. Inokuchi (Nippon Medical School) and H. Ohno (Kyoto
University) for providing t(14;18)-positive cell lines. We also thank
Dr J. Torii (Ehime Medical College of Health Science) for statistical analyses.
| |
Footnotes |
Submitted December 28, 2000; accepted March 21, 2001.
Supported by grants from the Ministry of Education, Science, Sports,
and Culture of Japan; the Sagawa Cancer Research Foundation; and the
Daiwa-Shoken Health Foundation.
The publication costs of this
article were defrayed in part by
page charge payment. Therefore,
and solely to indicate this fact,
this article is hereby marked
"advertisement"
in accordance with 18 U.S.C.
section 1734.
Reprints: Masaki Yasukawa, The First Department of Internal
Medicine, Ehime University School of Medicine, Shigenobu, Ehime
791-0295, Japan; e-mail: yasukawa{at}m.ehime-u.ac.jp.
| |
References |
1.
Jones SE, Fuks Z, Bull M, et al.
Non-Hodgkin's lymphomas, IV: clinicopathologic correlation in 405 cases.
Cancer.
1973;31:806-823[CrossRef][Medline]
[Order article via Infotrieve].
2.
Kadin ME, Berald CW, Nanba K, Wakasa H.
Lymphoproliferative diseases in Japan and Western countries.
Hum Pathol.
1983;14:745-772[Medline]
[Order article via Infotrieve].
3.
Ohsawa M, Aozasa K, Tsujimura T, Ikeda H, Tsujimura T.
Non-Hodgkin's lymphomas of follicular center type in Osaka, Japan.
Eur J Cancer Clin Oncol.
1989;25:1875-1878[CrossRef][Medline]
[Order article via Infotrieve].
4.
Amakawa R, Fukuhara S, Ohno H, et al.
Involvement of bcl-2 gene in Japanese follicular lymphoma.
Blood.
1989;73:787-791[Abstract/Free Full Text].
5.
Matsuyama F, Fukuhara S, Oguma S, et al.
Geographical aspects of bcl-2 gene involvement in Japanese patients with non-Hodgkin's B-cell lymphomas.
Int J Hematol.
1992;55:71-79[Medline]
[Order article via Infotrieve].
6.
Tsujimoto Y, Gorham J, Cossman J, Jaffe E, Croce CM.
The t(14;18) chromosomal translocation involved in the B-cell neoplasms results from mistakes in VDJ joining.
Science.
1985;229:963-967[Free Full Text].
7.
Bakhshi A, Jensen JP, Goldman P, et al.
Cloning the chromosomal breakpoint of t(14;18) human lymphomas: clustering around JH on chromosome 14 and near a transcriptional unit on 18.
Cell.
1985;41:899-906[CrossRef][Medline]
[Order article via Infotrieve].
8.
Cleary ML, Sklar J.
Nucleotide sequence of a t(14;18) chromosomal breakpoint in follicular lymphoma and demonstration of a breakpoint-cluster region near a transcriptionally active locus on chromosome 18.
Proc Natl Acad Sci U S A.
1985;82:7439-7443[Abstract/Free Full Text].
9.
Yunis JJ, Oken MM, Kaplan ME, Ensrud KE, Howe RR, Theologides A.
Distinctive chromosomal abnormalities in histologic subtypes of non-Hodgkin's lymphomas.
N Engl J Med.
1982;307:1231-1236[Abstract].
10.
Weiss LM, Wamke RA, Sklar J, Cleary ML.
Molecular analysis of the t(14;18) chromosomal translocation in malignant lymphomas.
N Engl J Med.
1987;317:1185-1189[Abstract].
11.
Korsmeyer SJ.
Bcl-2 initiates a new category of oncogenes: regulators of cell death.
Blood.
1992;80:879-886[Free Full Text].
12.
Liu Y, Hernandez AM, Shibata D, Cortopassi GA.
BCL2 translocation frequency rises with age in humans.
Proc Natl Acad Sci U S A.
1994;91:8910-8914[Abstract/Free Full Text].
13.
Limpens J, Stad R, Vos C, et al.
Lymphoma-associated translocation t(14;18) in blood B cells of normal individuals.
Blood.
1995;85:2528-2536[Abstract/Free Full Text].
14.
Ji W, Qu G, Ye P, Zhang X-Y, Halabi S, Ehrlich M.
Frequent detection of bcl-2/JH translocations in human blood and organ samples by a quantitative polymerase chain reaction assay.
Cancer Res.
1995;55:2876-2882[Abstract/Free Full Text].
15.
Dölken G, Illerhaus G, Hirt C, Mertelsmann R.
BCL-2/JH rearrangements in circulating B cells of healthy blood donors and patients with nonmalignant diseases.
J Clin Oncol.
1996;14:1333-1344[Abstract/Free Full Text].
16.
Fuscoe JC, Setzer RW, Collard DD, Moore MM.
Quantification of t(14;18) in the lymphocytes of healthy adult humans as a possible biomarker for environmental exposures to carcinogens.
Carcinogenesis.
1996;17:1013-1020[Abstract/Free Full Text].
17.
Dölken L, Schüler F, Dölken G.
Quantitative detection of t(14;18)-positive cells by real-time quantitative PCR using fluorogenic probes.
Biotechniques.
1998;25:1058-1064[Medline]
[Order article via Infotrieve].
18.
Yamaguchi H, Inokuchi K, Hanawa H, et al.
Establishment of a near-triploid human B-cell lymphoma cell line with t(14;18) and a p53 gene point mutation.
Br J Haematol.
1999;105:764-767[CrossRef][Medline]
[Order article via Infotrieve].
19.
Hamada T, Yonetani N, Ueda C, et al.
Expression of the PAX5/BSAP transcription factor in haematological tumour cells and further molecular characterization of the t(9;14)(p13;q32) translocation in B-cell non-Hodgkin's lymphoma.
Br J Haematol.
1998;102:691-700[CrossRef][Medline]
[Order article via Infotrieve].
20.
Aster JC, Kobayashi Y, Shiota M, Mori S, Sklar J.
Detection of the t(14;18) at similar frequencies in hyperplastic lymphoid tissues from American and Japanese patients.
Am J Pathol.
1992;141:291-299[Abstract].
21.
Bell DA, Liu Y, Cortopassi GA.
Occurrence of bcl-2 oncogene translocation with increased frequency in the peripheral blood of heavy smokers.
J Natl Cancer Inst.
1995;87:223-224[Free Full Text].
22.
Lipkowitz S, Garry VF, Kirsch IR.
Interlocus V-J recombination measures genomic instability in agriculture workers at risk for lymphoid malignancies.
Proc Natl Acad Sci U S A.
1992;89:5301-5305[Abstract/Free Full Text].
Top
Abstract
Introduction
