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  
J. Pathol. 188: 139–145 (1999)
HODGKIN AND REED–STERNBERG CELLS OF
CLASSICAL HODGKIN’S DISEASE OVEREXPRESS THE
TELOMERASE RNA TEMPLATE (hTR)
 ,  *,     
Institute for Pathology, Universitätsklinikum Benjamin Franklin, Free University Berlin, 12200 Berlin, Germany
SUMMARY
There is accumulating evidence to suggest that Hodgkin and Reed–Sternberg (HRS) cells represent the malignant cell population in
Hodgkin’s disease (HD). A recent report that HD tissue is in most instances devoid of telomerase activity was therefore unexpected.
Since telomerase activity was determined in whole tissue extracts and HRS cells comprise only a small minority of the cells in the
affected tissue, the telomerase activity of the HRS cells might have escaped detection. To test this possibility and to clarify whether HRS
cells contain the enzyme telomerase, 13 cases of classical HD were analysed by three different methods. The presence of telomerase was
studied at the single cell level by a sensitive radioactive in situ hybridization method employing a probe specific for the telomerase RNA
template (hTR). In addition, tissue extracts were studied for telomerase activity by a modified TRAP assay and for hTR by reverse
transcription-polymerase chain reaction (RT-PCR). The extractive methods revealed telomerase activity in eight and hTR in all of the
13 HD cases studied. In situ hybridization located large amounts of hTR in the HRS cells of all 13 HD cases and low to medium amounts
in some of the non-malignant lymphoid bystander cells. These results indicate that HRS cells constitutively overexpress telomerase and
thus use this enzyme for stabilizing their telomeres. This substantiates the malignant nature of HRS cells. Furthermore, the results
confirm that normal lymphoid cells can express telomerase. In consequence, methods of measuring telomerase in tissue extracts are not
suitable for determining the presence of this molecule in lymphoma cells, since the vast majority of lymphoid neoplasms contain
significant amounts of non-neoplastic lymphoid cells. Copyright 1999 John Wiley & Sons, Ltd.
KEY WORDS—Hodgkin’s
disease; HRS cells; telomerase; lymphocytes
INTRODUCTION
For a long time, the nature of the Hodgkin and
Reed–Sternberg (HRS) cells of Hodgkin’s disease (HD)
was an enigma. Recent molecular biological studies with
single HRS cells revealed that they represent a clonal
expansion of late B cells in most instances.1,2 From this
finding, it follows that the HRS cells start their proliferation from a single transformed B lymphocyte and thus
must have the capacity of unlimited growth. A prerequisite for this property is the activity of the ribonucleoprotein telomerase.3–13 Without this enzyme, the
chromosomal ends (telomeres) are shortened by each
mitotic division. When the telomeres are exhausted,
the cells stop proliferating and undergo senescence.
Malignant cells consistently harbour telomerase activity
to stabilize their telomeres. It was therefore a surprise
when Brousset et al.14 recently reported that in 13 of 14
HD cases, telomerase was not detectable by the TRAP
assay (telomeric repeat amplification protocol). In five of
these 13 cases, signs of degradation were found. For the
other eight cases, however, a true absence of telomerase
activity was assumed. The authors therefore concluded
that HRS cells use, in part, a telomerase-independent
mechanism for immortalization and that telomerase
activity (TA) might define new prognostic subcategories
*Correspondence to: Michael Hummel, PhD, Institute of
Pathology, Klinikum Benjamin Franklin, Free University Berlin,
Hindenburgdamm 30, 12200 Berlin, Germany. E-mail: hummel@
ukbf.fuberlin.de.
Contract/grant sponsor: Deutsche Forschungsgemeinschaft (DFG);
Contract/grant number: Hu 557/2-1.
CCC 0022–3417/99/070139–07$17.50
Copyright 1999 John Wiley & Sons, Ltd.
in HD. However, there are alternative interpretations
for the negative results of the TRAP assays reported by
Brousset et al. Negative results can easily be obtained,
for example, when telomerase is degraded by prolonged
tissue processing times, or through a TRAP assay with
low sensitivity. Another problem of the TRAP assay is
that its signal cannot be assigned to single cells. The
TRAP assay, like other extractive procedures, is thus
inappropriate for the investigation of HD, in which
HRS cells usually comprise less than 1 per cent of the
total cellular mass.
In this paper, we report the results obtained by the
investigation of 13 cases of classical HD for the expression of telomerase using three different methods. The
TRAP assay was performed to determine the TA and
RT-PCR was used for the detection of hTR in tissue
RNA extracts. In addition, in situ hybridization was
applied for the localization of hTR at the single cell
level, in order to demonstrate which cells in HD give rise
to the signals of the extractive methods.
MATERIALS AND METHODS
Cell line and tissue samples
Cell lines (K562, L428, L540, L591) were cultured in
RPMI medium supplemented with 10 per cent FCS.
Peripheral blood mononuclear cells were used from
healthy donors and were separated through a Ficoll
gradient. Cell suspensions were centrifuged at 1000 g
for 10 min and washed twice with phosphate-buffered
Received 2 July 1998
Revised 7 October 1998
Accepted 25 January 1999
140
B. HEINE ET AL.
saline. Tissue samples were surgically obtained for
diagnostic or therapeutic reasons. To investigate tissue
samples for the presence of TA, 15 ìm thick tissue
sections were produced from fresh frozen tissue blocks
and controlled by histological examination of 4–7 ìm
sections directly above and below the extracted tissue.
TRAP assay
TA was determined according to Kim et al.10
with minor modifications using the non-radioactive
GeneScan method as described previously.15 Briefly, an
aliquot of 6 ìg, 0·6 ìg, and 0·06 ìg of total protein from
each sample was investigated. Two TRAP assays were
independently performed from each sample. The supernatants of TRAP-negative samples were mixed with
TRAP-positive cell line supernatants (5:1; 1:5) in order
to detect telomerase inhibitors.
employing primers deduced from published sequence.
TEL-UP:
5-GGTGGCCATTTTTTGTCTAAC-3;
TEL-LOW: 5-TGCATGTGTGAGCCGAGT-3. Total
RNA from phytohaemagglutinin (PHA)-activated
peripheral blood lymphocytes (1 ìg) was used for
reverse transcription and amplification as described
above. The resulting 417 bp amplificates were subcloned
in pAMP1 (GibcoBRL, Gaithersburg, U.S.A.) and
sequenced (Model 377A, Perkin-Elmer/Applied Biosystems, Weiterstadt, Germany). Hybridization was carried
out with 35S-labelled run-off transcript as described
elsewhere.16 In brief, dewaxed and rehydrated paraffin
sections were exposed to 0·2  HCl and 0·125 mg/ml
pronase (Boehringer-Mannheim, Germany), followed
by acetylation with 0·1  triethanolamine, pH 8·0/0·25
per cent (v/v) acetic anhydride and dehydration through
graded ethanol. Slides were hybridized to 2–4105 cpm
of labelled probes overnight at 50C.
Analysis and quantification of the TRAP assay
TRAP-positive samples were semi-quantified employing the peak areas between 37 and 150 bp of the TRAP
amplification products as described elsewhere.15 The
sum of corresponding peak areas of a positive control
(K562; 0·6 ìg of protein) obtained from the same
GeneScan run was set to 100 units and the peaks of the
negative control (lysis buffer) were set to 0 units. The
relative TA (rTA) of a given sample X (0·6 ìg of protein)
was calculated using the following formula:
rTAX =[((X1nc)/(pcnc)+
(X2nc)/(pcnc))/200] units
where X1 and X2 are the peak areas of sample X
obtained from two different TRAP assays; pc is the peak
area, positive control; and nc is the peak area, negative
control.
RT-PCR for hTR
RT-PCR for the detection of the telomerase
RNA component was performed using the GeneAmp
RNA-PCR kit (Perkin-Elmer/Applied Biosystems,
Weiterstadt, Germany) as described previously. In brief,
random hexamer primed reverse transcription was performed with 2 ìl of lysate prepared for TRAP assay in
a 20 ìl reaction volume. PCR for amplification of the
RNA component was carried out employing the following primers: HTR-UP: 5-ACCCTAACTGAGAAGG
GCGT-3; HTR-LOW: 5-GCCAGCAGCTGACATTT
TTT-3. Forty cycles (96C for 15 s, 55C for 30 s, and
72C for 30 s) were used. The sequence of the resulting
142 bp amplificate was determined by automated DNA
sequencing and confirmed to comprise the expected
proportion of hTR.
In situ hybridization (ISH)
ISH was carried out as described previously.16 In
brief, the probe for detection of the human telomerase
RNA component by ISH was generated by RT-PCR
Copyright 1999 John Wiley & Sons, Ltd.
RESULTS
TRAP assay
The TRAP assay performed on all three HD-derived
cell lines tested (L428, L540, L591) displayed high TA
(Table I). Extracts from normal lymphoid tissue samples
(three spleens and two tonsils) demonstrated low to
medium TA, ranging from 5 to 45 units (rTA) (Table I).
When applied to 13 cases of HD, the TRAP assay
revealed in eight instances medium to high activity
ranging from 30 to 200 units (rTA) (Figs 1 and 2), with
lymphocyte-depleted HD cases tending to show the
highest activity. For two of the five TRAP assaynegative HD cases, the non-detection of TA proved to
be due to inhibitory effects, as revealed by mixing these
tissue extracts with TRAP assay-positive control lysates
(Fig. 2). No correlation was found between TA and age,
sex or Epstein–Barr virus infection.
RT-PCR for hTR
RT-PCR for the detection of hTR yielded positive
results in all HD cases as well as in all specimens derived
from non-neoplastic lymphoid tissues (Table I). There
were no significant differences between the amount of
PCR products in HD specimens and those obtained
from non-malignant lymphoid tissues.
ISH for hTR
ISH for the detection of hTR produced very strong
signals over HRS cells in all 13 HD cases (Fig. 3B–3D).
In addition, weak to moderate signals were seen over a
proportion of lymphoid bystander cells, most of which
had the morphology of activated cells. ISH performed
on an immortalized cell line (K562) mixed with peripheral blood cells from healthy donors led to comparable
results, i.e. a high accumulation of hTR signals over the
leukaemia cell line cells, whereas there were no grains
over the non-neoplastic peripheral blood cells, with the
exception of a few lymphoid cells of larger size (Fig. 3A).
J. Pathol. 188: 139–145 (1999)
141
HRS CELLS OF CLASSICAL HD OVEREXPRESS hTR
Table I—Detection of telomerase activity and the telomerase RNA component in Hodgkin’s disease (HD)
Case
No.
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
Organ
Diagnosis
Typing
EBV
Lymph node
Lymph node
Lymph node
Lymph node
Lymph node
Lymph node
Lymph node
Lymph node
Lymph node
Lymph node
Lymph node
Lymph node
Lymph node
Spleen
Spleen
Spleen
Tonsil
Tonsil
L428
L540
L591
HD
HD
HD
HD
HD
HD
HD
HD
HD
HD
HD
HD
HD
Trauma
Morbus Werlhof
Gastric carcinoma
Chronic inflammation
Chronic inflammation
HD cell line
HD cell line
HD cell line
Nodular-sclerosing
Nodular-sclerosing
Nodular-sclerosing
Nodular-sclerosing
Nodular-sclerosing
Mixed cellularity
Mixed cellularity
Mixed cellularity
Mixed cellularity
Lymphocyte-depleted
Lymphocyte-depleted
Lymphocyte-depleted
Lymphocyte-depleted
EBV
EBV
EBV
EBV
EBV
EBV
EBV
Telomerase RNA
component
Age
(years)
Sex
rTA
RT-PCR
ISH
81
27
24
54
90
57
26
64
56
87
60
34
27
25
69
63
10
12
F
M
F
M
M
M
M
F
F
M
M
M
M
F
F
F
M
M
50
30
Inhib.
190
50
Inhib.
30
180
150
200
5
15
15
45
35
140
140
105
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
n.d.
n.d.
n.d.
+*
+*
+*
+*
+*
+*
+*
+*
+*
+*
+*
+*
+*
†
†
†
†
†
n.d.
n.d.
n.d.
rTA=relative telomerase activity deduced from the TRAP assay in units; Inhib.=telomerase inhibition; RT-PCR=reverse transcription
polymerase chain reaction; ISH=in situ hybridization; EBV=Epstein–Barr virus; immunohistological visualization of the EBV-encoded latent
membrane antigen (LMP-1) in tumour cells; EBV=EBV detection by ISH for the EBV-encoded nuclear RNAs (EBER1 and 2) in some lymphoid
cells.
*Strong expression in Hodgkin and Reed–Sternberg cells.
†Weak to moderate signals in activated lymphocytes.
n.d.=not determined.
The application of our ISH to normal or hyperplastic
tonsil and spleen specimens generated only some weak
to moderately strong signals over activated lymphoid
cells (not shown). The hTR sense probe gave only very
weak randomly distributed background signals, without
any specific accumulation over cells.
DISCUSSION
Recent attempts by Brousset et al.14 to demonstrate
telomerase activity (TA) by TRAP in tissue extracts
from HD cases led to unexpected results, since only one
out of 14 cases was positive. In five of the negative cases,
TA was not detectable, due to enzymatic inhibition. In
the remaining eight cases, the authors regarded the
negative TRAP assay as evidence for a true absence of
TA. Based on these data, it was concluded that HD
might represent a special human malignancy that
frequently reaches immortality in the absence of TA,
which might therefore serve as a new prognostic
indicator.
To test these hypotheses, we analysed 13 cases of HD
for the expression of telomerase with three different
methods. Eight of these 13 cases showed TA, as revealed
by a TRAP assay employing fluorescence labelled oligonucleotides.15 Two cases were not interpretable, due to
inhibitory factors, leaving only three cases (28 per cent)
Copyright 1999 John Wiley & Sons, Ltd.
negative. This is in sharp contrast to the study of
Brousset et al., in which 88 per cent of the HD cases
were found to be devoid of TA.14 The most likely
explanation for this discrepancy lies in technical artefacts inherent to the TRAP assay. The protein and RNA
component of the telomerase are highly accessible for
degradation, as a result of which the TRAP assay might
easily become negative, due, for example, to delayed
sample preparation. The possibility that degradation is
responsible for the negative results in the abovementioned study is underscored by the investigation
of non-malignant lymphoid tissue. All such samples
analysed by Brousset et al. and our group were TRAPpositive, indicating expression of telomerase in some
reactive lymphoid cells. This finding is in harmony with
the results of other research groups.17–23 In addition, the
granulocytic infiltrate in HD might contribute to falsenegative outcomes of the TRAP assay, as recently
shown by Norrback et al.24
Our assumption that telomerase molecules are present
in all lymphoid tissues, including those affected by HD,
is further supported by the application of RT-PCR for
the demonstration of the RNA component (hTR) of
telomerase. This method demonstrated hTR in all HD
specimens, as well as in all benign lymphoid tissues.
In TRAP assay, as well as other PCR techniques for
the demonstration of hTR, or the recently discovered
protein moiety of telomerase, measures whole tissue
J. Pathol. 188: 139–145 (1999)
142
B. HEINE ET AL.
Fig. 1—Non-radioactive TRAP for the detection of telomerase activity (TA). (A–D) Serial dilution of lysates of cell line K562. (A) 6 ìg, (B) 0·6 ìg,
(C) 0·06 ìg, and (D) 0·006 ìg of protein. Note the partial inhibition of the TA when 6 ìg of protein was applied. (E–G) Serial dilution of lysates
obtained from case 11 (HD, lymphocyte-depleted). (E) 6 ìg, (F) 0·6 ìg, and (G) 0·06 ìg of protein. (H) 6 ìg of protein from the same case (11) after
pretreatment with RNase. Note the complete disappearance of specific products
Copyright 1999 John Wiley & Sons, Ltd.
J. Pathol. 188: 139–145 (1999)
HRS CELLS OF CLASSICAL HD OVEREXPRESS hTR
143
Fig. 2—Non-radioactive TRAP for the detection of telomerase activity. (A–C) Serial dilution of lysates obtained from case 4 (HD, nodulatsclerosing). (A) 6 ìg, (B) 0·6 ìg, and (C) 0·06 ìg of protein. (E) 5 ìg of protein (case 4) mixed with lysate (1 ìg of protein) obtained from cell line
K562. (F) 1 ìg of protein (case 4) mixed with lysate (5 ìg of protein) obtained from cell line K562. Note the inhibition of the signals due to inhibitory
effects of the tissue lysate. (D) Negative control (lysis buffer). (G, H) Cell line K562. (G) 5 ìg of protein; (H) 1 ìg of protein
Copyright 1999 John Wiley & Sons, Ltd.
J. Pathol. 188: 139–145 (1999)
144
B. HEINE ET AL.
Fig. 3—Radioactive in situ hybridization for the detection of the telomerase RNA component (hTR). (A) Mixture (1:1) of cell line cells (K562) and
normal peripheral mononuclear cells. (B) TRAP-negative HD case (case 3); (C) TRAP-negative HD case due to inhibition (case 4); (D)
TRAP-positive HD case (case 2). Note that all cases display strong expression of hTR irrespective of the results of the TRAP assay
extracts.12,13,25,26 As already demonstrated in previous
studies on HD, extractive procedures performed with
whole tissue DNA or RNA extracts cannot determine
the cellular source of the signals obtained.1,2,27–29 This
also holds true in terms of telomerase, since the
achieved signals might be derived from HRS cells or
from other cells, such as activated non-neoplastic
bystander cells. We therefore extended our study to the
single cell level, by applying ISH with radioactively
labelled probe specific for hTR. With this method, we
found in a previously published study,30 using gastric
carcinoma as an example, a striking difference in the
expression of hTR between malignant cells and most
normal cells. The gastric carcinoma cells showed a
strong hTR signal, whereas all non-malignant bystander cells proved to be negative, except for some
activated lymphocytes and physiologically regenerating
epithelial cells displaying a low expression of hTR.30
The distribution of hTR in mixed lesions therefore
corresponds very well to the in vitro results of the
TRAP assay,10 although the detection of hTR is an
indirect demonstration of TA. Until now, no method
has been described to demonstrate TA directly in tissue
sections at the single cell level.
Copyright 1999 John Wiley & Sons, Ltd.
ISH applied to our HD cases revealed a high load of
hTR only in the HRS cells of all our cases, indicating
that HRS cells overexpress telomerase to an extent similar to gastric cancer cells. As in non-neoplastic lymphoid
tissue, some reactive lymphoid cells in the HD tissue
samples showed a low level of hTR-specific signals.
The high load of hTR in the HRS cells of all HD cases
studied disproves the hypothesis of Brousset et al.,14 that
HRS cells acquire their potential for unlimited growth
mostly by mechanisms other than TA and indicates that
in terms of telomerase, HD is indistinguishable from
other malignant disorders. Up-regulated telomerase is
therefore not a prognostic indicator in HD, but a constant feature of HRS cells. In conjunction with the recent
finding that HRS cells are derived from a single transformed B cell,1,2,31 the overexpression of hTR by HRS
cells shows that HD represents a clonal expansion of B
cells that are immortalized by the enzyme telomerase.
ACKNOWLEDGEMENTS
We are indebted to Elisabeth Oker for her excellent
technical assistance and to Leslie Udvarhely for editorial
J. Pathol. 188: 139–145 (1999)
HRS CELLS OF CLASSICAL HD OVEREXPRESS hTR
help with the preparation of the manuscript. This work
was supported by the Deutsche Forschungsgemeinschaft
(DFG; Hu 557/2-1).
REFERENCES
1. Kuppers R, Rajewsky K, Zhao M, et al. Hodgkin disease: Hodgkin and
Reed–Sternberg cells picked from histological sections show clonal
immunoglobulin gene rearrangements and appear to be derived from B cells
at various stages of development. Proc Natl Acad Sci U S A 1994; 91:
10 962–10 966.
2. Hummel M, Marafioti T, Ziemann K, Stein H. Hodgkin’s disease with
monoclonal and polyclonal populations of Reed–Sternberg cells. N Engl J
Med 1995; 333: 901–906.
3. Watson ID. Origin of concatemeri T7 DNA. Nature New Biol 1972; 239:
197–201.
4. Olovnikov A. A theory of marginotomy. The incomplete copying of
template margin in enzymic synthesis of polynucleotides and biological
significance of the phenomenon. Theor Biol 1973; 41: 181–190.
5. Moyzis RK, Buckingham JM, Cram IS, et al. A highly conserved
repetitive DNA sequence (TTAGGG)n, present at the telomeres of human
chromosomes. Proc Natl Acad Sci U S A 1988; 85: 6622–6626.
6. Harley CB, Futcher AB, Greider CW. Telomeres shorten during aging of
human fibroblasts. Nature 1990; 345: 458–460.
7. Lindsey J, McGill NI, Lindsey LA, Green DK, Kooke HJ. In vivo loss of
telomeric repeats with age in humans. Mutat Res 1991; 256: 45–48.
8. Greider CW, Blackburn EH. Identification of a specific telomere terminal
transferase activity in tetrahymena extracts. Cell 1985; 43: 405–413.
9. Morin GB. The human telomere terminal transferase enzyme is a
ribonucleoprotein that synthesizes TTAGGG repeats. Cell 1989; 59:
521–529.
10. Kim NW, Piatyszek MA, Prowse KR, et al. Specific association of human
telomerase activity with immortal cells and cancer. Science 1994; 266:
2011–2015.
11. Feng J, Funk W, Wang S, et al. The RNA component of human telomerase.
Science 1995; 269: 1236–1241.
12. Lingner J, Hughes T, Shevchenko A, Mann M, Lundblad V, Cech T.
Reverse transcriptase motifs in the catalytic subunit of telomerase. Science
1997; 276: 561–567.
13. Nakamura J, Saito M, Nakamura H, Matsuura A, Ishikawa F. TLP1. A
gene encoding a protein component of mammalian telomerase is a novel
member of WD repeats family. Cell 1997; 88: 875–884.
14. Brousset P, Al Saati T, Chaouche N, Schlaifer D, Chittal S, Delsol G.
Telomerase activity in reactive and neoplastic lymphoid tissue: infrequent
detection of activity in Hodgkin’s disease. Blood 1997; 89: 26–31.
15. Heine B, Hummel M, Müller M, Heicappell R, Miller K, Stein H.
Non-radioactive measurement of telomerase activity in human bladder
cancer, bladder washings, and in urine. J Pathol 1998; 184: 71–76.
Copyright 1999 John Wiley & Sons, Ltd.
145
16. Foss HD, Herbst H, Oelmann E, et al. Lymphotoxin, tumour necrosis
factor and interleukin-6 gene transcripts are present in Hodgkin and
Reed–Sternberg cells of most Hodgkin’s disease cases. Br J Haematol 1993;
84: 627–635.
17. Counter CM, Gupta J, Harley CB, Leber B, Bacchetti S. Telomerase
activity in normal leukocytes and in hematologic malignancies. Blood 1995;
85: 2315–2320.
18. Avilion AA, Piatyszek MA, Gupta J, Shay JW, Bacchetti S, Greider CW.
Human telomerase RNA and telomerase activity in immortal cell lines and
tumor tissues. Cancer Res 1996; 56: 645–650.
19. Norrback KF, Dahlenborg K, Carlsson R, Roos G. Telomerase activation
in normal B lymphocytes and non-Hodgkin’s lymphomas. Blood 1996; 88:
222–229.
20. Weng NP, Granger L, Hodes RJ. Telomere lengthening and telomerase
activation during human B cell differentiation. Proc Natl Acad Sci U S A
1997; 94: 10 827–10 832.
21. Hu BT, Lee SC, Marin E, Ryan DH, Insel RA. Telomerase is upregulated
in human germinal cancer B cells in vivo and can be re-expressed in memory
B cells activated in vitro. J Immunol 1997; 159: 1068–1071.
22. Weng N, Levine BL, June CH, Hodes RJ. Regulation of telomerase RNA
template expression in human T lymphocyte development and activation.
J Immunol 1997; 158: 3215–3220.
23. Yashima K, Piatyszek MA, Saboorian HM, et al. Telomerase activity and in
situ telomerase RNA expression in malignant and non-malignant lymph
nodes. J Clin Pathol 1997; 50: 110–117.
24. Norrback K, Enblad G, Erlanson M, Sundstrom C, Roos G. Telomerase
activity in Hodgkin’s disease. Blood 1998; 92: 567–573.
25. Nakayama J, Tahara H, Tahara E, et al. Telomerase activation by hTRT in
human normal fibroblasts and hepatocellular carcinomas. Nature Genet
1998; 18: 65–68.
26. Bodnar A, Ouellett M, Frolkis M, Holt S, Chiu C, Morin G. Extension of
life-span by introduction of telomerase into normal human cells. Science
1998; 279: 349–352.
27. Herbst H, Niedobitek G, Kneba M, et al. High incidence of Epstein–Barr
virus genomes in Hodgkin’s disease. Am J Pathol 1990; 137: 13–18.
28. Hummel M, Anagnostopoulos I, Dallenbach F, Korbjuhn P, Dimmler C,
Stein H. EBV infection patterns in Hodgkin’s disease and normal lymphoid
tissue: expression and cellular localization of EBV gene products. Br J
Haematol 1992; 82: 689–694.
29. Korbjuhn P, Anagnostopoulos I, Hummel M, et al. Frequent latent
Epstein–Barr virus infection of neoplastic T cells and bystander B cells in
human immunodeficiency virus-negative European peripheral pleomorphic
T-cell lymphomas. Blood 1993; 82: 217–223.
30. Heine B, Hummel M, Demel G, Stein H. Demonstration of constant
upregulation of telomerase RNA component in human gastric carcinomas
using in situ hybridisation. J Pathol 1998; 185: 139–144.
31. Stein H, Hummel M, Marafioti T, Anagnostopoulos I, Foss H. Molecular
biology of Hodgkin’s disease. Cancer Surv 1997; 30: 107–123.
J. Pathol. 188: 139–145 (1999)
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