The STAT5 transcription factor in B-cells of patients with chronic lymphocytic leukemia
Aim. To find out the cause of inhibition of the IL2-STAT5 signaling pathway in chronic lymphocytic leukemia (CLL) cells. Methods.CLL cells were isolated from peripheral blood, using gradient centrifugation on a ficoll-verografin mixture. Expression of the STAT1-6 genes at the mRNA level was analyzed...
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irk-123456789-1543822019-07-07T12:56:25Z The STAT5 transcription factor in B-cells of patients with chronic lymphocytic leukemia Matvieieva, A.S. Kovalevska, L.M. Ivanivska, T.S. Klein, E. Kashuba, E.V. Biomedicine Aim. To find out the cause of inhibition of the IL2-STAT5 signaling pathway in chronic lymphocytic leukemia (CLL) cells. Methods.CLL cells were isolated from peripheral blood, using gradient centrifugation on a ficoll-verografin mixture. Expression of the STAT1-6 genes at the mRNA level was analyzed, using the Oncomine database. Expression, phosphorylation status and cellular localization of the STAT5 protein were studied by fluorescence microscopy, using specific antibodies. Results.Unlike B-cells of healthy donors, expression of the STAT5A protein was low in the patient CLL cells. As we have previously shown, the IL-2-STAT5 (JAK-STAT5) signaling pathway is inhibited in CLL cells. Now we demonstrated a low level of phosphorylation of the STAT5 protein, or a complete lack of phosphorylation in CLL cells. The STAT5A protein shows cytoplasmic localization, indicating the absence of complexes in the nucleus that activate/repress transcription of the STAT5-dependent genes. Conclusions. Inhibition of the IL-2-STAT5 pathway in CLL cells is caused by a lack of the STAT5 proteins phosphorylation and/or the absence of the active STAT5A transcription complexes in the nucleus of CLL cells. Мета. Встановити причину блокування сигнального шляху IL2-STAT5 у клітинах крові хворих на хронічний ліфмолейкоз (ХЛЛ). Методи. Клітини ХЛЛ виділяли з периферичної крові пацієнтів, хворих на ХЛЛ, за допомогою центрифугування у градієнтіфікол-верографін. Експресію генів STAT1-6 на рівні мРНК аналізували за допомогою бази даних Oncomine. Експресію, статус фосфорилювання і клітинну локалізацію білка STAT5 вивчали методом флуоресцентної мікроскопії з використаннім специфічних антитіл. Результати. Навідмінувід В-клітин здорових людей, експресія білка STAT5А була низькою у клітинах хворих на ХЛЛ. Як нами було встановлено раніше, сигнальний шлях IL-2-STAT5 (JAK-STAT5) інгібовано у клітинах ХЛЛ. Нами було показано низький рівень фосфорилювання білків STAT5, або повну відсутність фосфорильованої форми протеїнів в лейкемічних клітинах. Протеїн STAT5А показує цитоплазматичну локалізацію, що вказує н авідсутність у ядрікомплексів, активуючих транскрипцію генів, залежних від фактора транскрипції STAT5. Висновки. Інгибування сигнального клітинного шляху IL-2-STAT5 в клітинах крові хворих на ХЛЛ реалізується за рахунок гіпофосфорилювання протеїнів STAT5 та/або відсутності активних комплексів транскрипції STAT5А у ядрі лейкемичних клітин. Цель. Установить причину блокирования сигнального пути IL2-STAT5 в клетках крови больных хроническим лимфолейкозом (ХЛЛ). Методы. Лейкемические клетки выделяли из периферической крови с помощью центрифугирования в градиенте плотности фиколл-верографин. Экспрессию генов STAT1-6 на уровне мРНК анализировали с помощью базы данных Oncomine. Экспрессию, статус фосфорилирования и клеточную локализацию белка STAT5 изучали методом флуоресцентной микроскопии с использованием специфических антител. Результаты. В отличие от В-клеток здоровых людей, экспрессия белка STAT5А была низкой в клетках больных ХЛЛ. Как нами было установлено ранее, сигнальный путь IL-2-STAT5 (JAK-STAT5) ингибирован в клетках ХЛЛ. Намибылпоказаннизкийуровеньфосфорилированиябелков STAT5 или полное отсутствие фосфорилированной формы протеинов в лейкемических клетках. Протеин STAT5А показывает цитоплазматическую локализацию, что указывает на отсутствие в ядре комплексов, активирующих транскрипцию генов, зависимых от фактора транскрипции STAT5. Выводы. Ингибирование сигнального клеточного пути IL-2-STAT5 в клетках крови больных ХЛЛ происходит за счёт гипофосфорилирования белков STAT5 и/или отсутствия активных комплексов транскрипции STAT5А в ядре лейкемических клеток. 2019 Article The STAT5 transcription factor in B-cells of patients with chronic lymphocytic leukemia / A.S. Matvieieva, L.M. Kovalevska, T.S. Ivanivska, E. Klein, E.V. Kashuba // Вiopolymers and Cell. — 2019. — Т. 35, № 1. — С. 30-38. — Бібліогр.: 21 назв. — англ. 0233-7657 DOI: http://dx.doi.org/10.7124/bc.000993 http://dspace.nbuv.gov.ua/handle/123456789/154382 576.3 + 577.2+ 616.006.4 en Вiopolymers and Cell Інститут молекулярної біології і генетики НАН України |
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Biomedicine Biomedicine Matvieieva, A.S. Kovalevska, L.M. Ivanivska, T.S. Klein, E. Kashuba, E.V. The STAT5 transcription factor in B-cells of patients with chronic lymphocytic leukemia Вiopolymers and Cell |
| description |
Aim. To find out the cause of inhibition of the IL2-STAT5 signaling pathway in chronic lymphocytic leukemia (CLL) cells. Methods.CLL cells were isolated from peripheral blood, using gradient centrifugation on a ficoll-verografin mixture. Expression of the STAT1-6 genes at the mRNA level was analyzed, using the Oncomine database. Expression, phosphorylation status and cellular localization of the STAT5 protein were studied by fluorescence microscopy, using specific antibodies. Results.Unlike B-cells of healthy donors, expression of the STAT5A protein was low in the patient CLL cells. As we have previously shown, the IL-2-STAT5 (JAK-STAT5) signaling pathway is inhibited in CLL cells. Now we demonstrated a low level of phosphorylation of the STAT5 protein, or a complete lack of phosphorylation in CLL cells. The STAT5A protein shows cytoplasmic localization, indicating the absence of complexes in the nucleus that activate/repress transcription of the STAT5-dependent genes. Conclusions. Inhibition of the IL-2-STAT5 pathway in CLL cells is caused by a lack of the STAT5 proteins phosphorylation and/or the absence of the active STAT5A transcription complexes in the nucleus of CLL cells. |
| format |
Article |
| author |
Matvieieva, A.S. Kovalevska, L.M. Ivanivska, T.S. Klein, E. Kashuba, E.V. |
| author_facet |
Matvieieva, A.S. Kovalevska, L.M. Ivanivska, T.S. Klein, E. Kashuba, E.V. |
| author_sort |
Matvieieva, A.S. |
| title |
The STAT5 transcription factor in B-cells of patients with chronic lymphocytic leukemia |
| title_short |
The STAT5 transcription factor in B-cells of patients with chronic lymphocytic leukemia |
| title_full |
The STAT5 transcription factor in B-cells of patients with chronic lymphocytic leukemia |
| title_fullStr |
The STAT5 transcription factor in B-cells of patients with chronic lymphocytic leukemia |
| title_full_unstemmed |
The STAT5 transcription factor in B-cells of patients with chronic lymphocytic leukemia |
| title_sort |
stat5 transcription factor in b-cells of patients with chronic lymphocytic leukemia |
| publisher |
Інститут молекулярної біології і генетики НАН України |
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2019 |
| topic_facet |
Biomedicine |
| url |
http://dspace.nbuv.gov.ua/handle/123456789/154382 |
| citation_txt |
The STAT5 transcription factor in B-cells of patients with chronic lymphocytic leukemia / A.S. Matvieieva, L.M. Kovalevska, T.S. Ivanivska, E. Klein, E.V. Kashuba // Вiopolymers and Cell. — 2019. — Т. 35, № 1. — С. 30-38. — Бібліогр.: 21 назв. — англ. |
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Вiopolymers and Cell |
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30
A. S. Matvieieva, L. M. Kovalevska, T. S. Ivanivska
© 2019 A. S. Matvieieva et al.; Published by the Institute of Molecular Biology and Genetics, NAS of Ukraine on behalf of Bio-
polymers and Cell. 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 reuse, distribution, and reproduction in any medium,
provided the original work is properly cited
ISSN 0233-7657
Biopolymers and Cell. 2019. Vol. 35. N 1. P 30–38
doi: http://dx.doi.org/10.7124/bc.000993
UDC576.3 + 577.2+ 616.006.4
The STAT5 transcription factor in B-cells of patients with chronic
lymphocytic leukemia
A. S. Matvieieva1, L. M. Kovalevska1, T. S. Ivanivska1, E. Klein2, E. V. Kashuba1,2
1 R. E. Kavetsky Institute of Experimental Pathology, Oncology and Radiobiology, NAS of Ukraine
45, Vasilkivska Str., Kyiv, Ukraine, 01022
2 Karolinska Institutet
Stockholm SE-171 77, Sweden
Kashuba@nas.gov.ua, lenakash@yahoo.com
Aim. To find out the cause of inhibition of the IL2-STAT5 signaling pathway in chronic lym-
phocytic leukemia (CLL) cells. Methods. CLL cells were isolated from peripheral blood,
using gradient centrifugation on a ficoll-verografin mixture. Expression of the STAT1-6 genes
at the mRNA level was analyzed, using the Oncomine database. Expression, phosphorylation
status and cellular localization of the STAT5 protein were studied by fluorescence microscopy,
using specific antibodies. Results. Unlike in B-cells of healthy donors, expression of the
STAT5A protein was low in the patient CLL cells. As we have previously shown, the
IL-2-STAT5 (JAK-STAT5) signaling pathway is inhibited in CLL cells. Now we demon-
strated a low level of phosphorylation of the STAT5 protein, or a complete lack of phospho-
rylation in CLL cells. The STAT5A protein shows cytoplasmic localization, indicating the
absence of complexes in the nucleus that activate/repress transcription of the STAT5-dependent
genes. Conclusions. Inhibition of the IL-2-STAT5 pathway in CLL cells is caused by a lack
of the STAT5 proteins phosphorylation and/or the absence of the active STAT5A transcription
complexes in the nucleus of CLL cells.
K e y w o r d s: Chronic lymphocytic leukemia (CLL), B-peripheral blood cells, STAT5,
STAT5A, STAT5B, IL2-STAT5 signaling pathway.
Introduction
Chronic lymphocytic leukemia (CLL) [1] is one
of the most common forms of leukemia in
Europe and the United States. The incidence rate
is approximately 3.5 per 100,000 population (5.0
for men and 2.5 for women) [2, 3]. Prior to the
onset of CLL, 5–10% of patients develop mono-
clonal B cell lymphocytosis (MBL), when the
number of B-lymphocytes in the peripheral
blood counts 5·104 - 2·106/ml. MBL occurs in
individuals over 40. With a frequency of about
1% per year, MBL can progress to CLL, when
the number of B cells is 5–10·106/ml [4].
https://teacode.com/online/udc/57/577.2.html
mailto:Kashuba@nas.gov.ua
31
The STAT5 transcription factor in B-cells of patients with chronic lymphocytic leukemia
CLL develops due to the slow accumulation
of the long-lived, but immunologically incom-
petent B-lymphocytes. Such cells are often
referred to as “immuno-senescent”, that de-
scribes their inability to differentiate into plas-
ma cells, producing antibodies [4]. CLL cells
do not proliferate, can not be activated by
ligands and do not undergo apoptosis.
Importantly, the transforming Epstein-Barr
virus (EBV) could infect CLL cells, but in-
fected cells do not proliferate even in vitro. It
was shown, that one of the important viral
proteins, i.e. LMP1, is not expressed in CLL
cells [5, 6]. Obviously, that several transcrip-
tion factors, ATF-2/c-Jun, for example [5], are
missing or not activated in CLL cells.
Of note, CLL cells express interleukin recep-
tors (IL2R, IL4R, IL6R, IL10R, IL13R), TNFα,
interferon alpha (INFA) and gamma (INFG),
and tumor growth factor beta (TGFB) [ 7].
The activation of the canonical TGFB path-
way in B cells usually leads to induction of
pro-apoptotic BMF, BIM, BAX and, and as a
consequence, to apoptosis [7]. In CLL cells,
the level of BCL2 does not differ from the
expression of this gene in the peripheral blood
B-cells of healthy individuals. TGFB receptors
(TGFBRs) are expressed approximately equal-
ly in B-CLL and peripheral blood
B-lymphocytes [8, 9]. However, most of the
genes that are usually induced by activation of
the TGFB-SMAD2/3 pathway, namely
BCL2L1 (BCL-XL), CCND2 (cyclin D2), ID1,
MYC, ATF3, TGIF1 andKLF10 (TIEG) are
basically not expressed in CLL cells [9].
In the leukemic cells, the IL-2-STAT5 path-
way is blocked [10]. The genes, usually in-
duced by the homo (hetero) dimers of the
STAT5 (Signal Transducer and Activator of
Transcription 5) proteins, namely BCL2L1
(BCL-XL), CCND2 (cyclin D2), HIF1A, ID1,
MCL1 and MYC, are barely detected.
Previously, we have shown that the TGFB-
SMAD2/3 pathway is inhibited in CLL cells,
due to retention of the SMAD3 and SMAD4
proteins in cytoplasm [11].Partially, the TGFB-
SMAD2/3 and IL-2-STAT5 pathways overlap.
Anyway, both pathways are blocked in B-CLL-
cells.
In the present paper, the expression levels
of the STAT1-6 genes, the phosphorylation
status of the STAT5 protein and STAT5 cel-
lular localization were studied in CLL cells
and in the peripheral blood B-cells of healthy
donors, with the aim to find a cause of inhibi-
tion of the IL-2-STAT5 pathways upon CLL.
Materials and Methods
The samples of the peripheral blood of 9 pa-
tients with CLL and one patient with B-cell
prolymphocytic leukemia (BCPL) were ob-
tained from the staff of the Department of
Onco-hematology (headed by Professor
D.F.Gluzman) at the R.E. Kavetsky Institute
of Experimental Pathology, Oncology and
Radiobiology (IEPOR) of National Academy
of Sciences of Ukraine. In order to verify the
CLL diagnosis, the immunocytochemical
methods were used utilizing anti-alkaline phos-
phatase (APAAP) labeled streptavidin-biotin
and alkaline phosphatase (LSAB-AP) and a
wide panel of monoclonal antibodies. As a
control, B-cells were isolated from peripheral
blood of two healthy donors in the ficoll-vero-
grafin gradient. T cells were removed by form-
ing rosettes with erythrocytes of sheep, fol-
lowed by centrifugation. The experimental
protocol was approved by a Committee on
32
A. S. Matvieieva, L. M. Kovalevska, T. S. Ivanivska et al.
Bioethics at R.E. Kavetsky IEPOR of National
Academy of Sciences of Ukraine.
The CLL cells were isolated from periph-
eral blood in the ficoll-verografin gradient.
50000 cells were attached to a glass slide for
the further immune- fluorescent analysis.
Double staining of cells was performed,
according to the following scheme: rabbit
anti-phosphorylated STAT5 (recognizing both,
STAT5A and STAT5B) (Cell Signaling, USA);
secondary anti-rabbit antibodies produced in
swine and conjugated with fluorescein-5-iso-
A
B
C
D
E
F
G
Fig. 1. A bioinformatic analysis of expression of STAT genes, using the Oncomine database. STAT2 (B), STAT5A (E)
and STAT5B (F) genes are expressed at a lower level in peripheral blood mononuclear cells and in CLL cells, than
STAT1 (A), STAT3 (C), STAT4 (D), and STAT6 (G). The lowest relative values are registered for STAT5B. The output
data for STAT2 are published in a study [14], in which 8603 genes were analyzed. For other STAT genes, the initial
analysis of 19574 genes is described in a work [15]. Conditions of the analysis: change of expression two-folds at
least, only 5% of the best genes was chosen, p = 0.0067.
33
The STAT5 transcription factor in B-cells of patients with chronic lymphocytic leukemia
thiocyanate (FITC, DAKO, Denmark); then
mouse monoclonal antibody against STAT5A
(Cell Signaling, USA); and the secondary
anti-mouse antibodies produced in horse and
conjugated with Texas red (TR, DAKO,
Denmark). The DNA was stained with Hoechst
3321 (Sigma-Aldrich, USA).
The images were captured by a CCD cam-
era (Hamamatsu, Japan), assembled and ana-
lyzed in Photoshop.
To analyze the STAT gene expression at the
mRNA level, a public database Oncomine was
used. Oncomine contains the published data
that was collected, standardized, annotated and
statistically analyzed by Compendia Bioscience
(www.oncomine.com, October 2018, Thermo
Fisher Scientific, Ann-Arbor, MI, USA).
Results and Discussion
The bioinformatics analysis of the Oncomine
database showed that the STAT genes at the
mRNA level were expressed at approximately
the same level in CLL cells, as in peripheral
blood mononuclear cells (Fig. 1). Of note, the
STAT2, STAT5А and STAT5В genes are ex-
pressed at lower levels in peripheral blood
mononuclear cells and in CLL cells, compared
with the STAT1, 3, 4 and 6 genes. The lowest
relative values of expression in CLL were
shown for STAT5B. The obtained results con-
firm our previously published data [10, 12, 13].
Subsequently, the levels of phosphorylation
of the STAT5 proteins (A and B isoforms) were
assessed as well as the STAT5 cellular localiza-
tion, using immunostaining. The control double
staining (excluding one of the primary antibo-
dies) did not show any background signals.
Noteworthy, the STAT5 protein (isoforms A
and B) showed basal levels of phosphorylation
in the control samples, i.e. B cells of healthy
donors (Fig. 2, green signal). The phosphory-
lated protein was observed almost exclusively
in the nucleus. Moreover, STAT5 formed large
nuclear inclusions (indicated by green arrows
in Fig. 2). The STAT5A protein was also local-
ized mainly in the nucleus in B-cells of healthy
donors (red signal in Figure 2), but a propor-
tion of protein was observed in the cytoplasm
as well (the red arrows in Fig. 2). Of note, the
signals of phosphorylated STAT5 and STAT5A
Fig. 2. Pattern of phosphorylation of
STAT5 proteins (isoforms A and B)
and their cellular localization. Phos-
phorylated form of proteins is shown
in green. Red — a signal of the STA-
T5A protein. DNA is shown in blue.
34
A. S. Matvieieva, L. M. Kovalevska, T. S. Ivanivska et al.
were partially co-localized in the nucleus (Fig.
2, marked with asterisks), indicating activation
of the IL-2-STAT5 (JAK-STAT5) pathway in
B cells of healthy donors. Previously, the con-
stitutively active STAT5 was found in the
nucleus of rapidly proliferating malignant he-
matopoetic cells [16].
In contrast to the pattern observed in B-cells
of healthy individuals, in CLL cells a very low
signal of the phosphorylated STAT5 proteins
was observed (Fig. 3, in green). The expression
levels of the STAT5A protein were quite low
as well (Fig. 3, red signal). Noteworthy, when
the STAT5A signal was rather high, phos-
phorylation was not detected (Fig. 3, samples
102884 and 97570, in green). Moreover, the
phosphorylated form was localized almost
exclusively in the cytoplasm.
Fig. 3. Pattern of phosphorylation of STAT5 proteins (isoforms A and B) and their cellular localization in CLL cells.
Phosphorylated form of proteins is shown in green. Red — a signal of the STAT5A protein. DNA is shown in blue.
35
The STAT5 transcription factor in B-cells of patients with chronic lymphocytic leukemia
In several patients the STAT5A protein was
practically absent (Fig. 3, samples 103963 and
89819). Also, the expression levels of both
isoforms could be very low (Fig. 3, sample
103063).
We have to emphasize that the STAT5A
protein and the STAT5 phosphorylated iso-
forms were localized exclusively in the cyto-
plasm of peripheral blood mononuclear cells
of CLL patients, in contrast to the pattern
observed in B cells of healthy individual.
In B cell of the BCPL patient, the phos-
phorylated form of the STAT5 proteins was
not detected. However, a high expression le vel
of STAT5A was found, and a large proportion
of the protein was localized in the nucleus
(Fig. 4, red signal).
It is known, that the phosphorylated STAT5
proteins are activated constitutively in indi-
viduals with onco-hematological diseases
[17–19].
The obtained results allow us to speculate
that the inhibition of the IL-2-STAT5 (JAK-
STAT5) pathway in CLL cells might be due to
the low levels of STAT5 phosphorylation, or
the complete lack of the STAT5 phosphoryla-
tion. Noteworthy also, the STAT5A protein is
found mainly in cytoplasm, which suggests
that the protein complexes, activating tran-
scription of the STAT5-depending genes are
absent in the nucleus. It was demonstrated
earlier that the STAT5 functions as transcrip-
tion factor exclusively in the nucleus [20].
Actually, the reason for such cellular local-
ization of the STAT5 proteins I CLL cells re-
mains an open question. No doubt, that local-
ization of proteins depends on their phosphor-
ylation status. Moreover, activation of the
JAK-STAT pathway is also regulated by the
formation of homo(hetero)dimers of the vari-
ous phosphorylated STAT proteins [21]. It is
important, to study the phosphorylation status
of other STAT proteins, namely, STAT2, 3 and
6, and their cellular localization before and
after the interaction of surface IL2R with the
corresponding ligand (IL2) in CLL cells.
Fig. 4. Pattern of phosphorylation of
STAT5 proteins (isoforms A and B)
and their cellular localization in
blood cells of BCPL patient. Phos-
phorylated form of proteins is shown
in green. Red — a signal of the STA-
T5A protein. DNA is shown in blue.
36
A. S. Matvieieva, L. M. Kovalevska, T. S. Ivanivska et al.
Conclusions
As we have shown previously, the IL-2-STAT5
(JAK-STAT5) cellular signaling pathway is
inhibited in CLL cells. In the present research
we found a low level of the STAT5 phos-
phorylation, or even the complete absence of
the phosphorylated protein in leukemic cells.
The STAT5A protein is localized mainly in
cytoplasm, indicating the absence of active
transcriptional complexes in the nucleus, i.e.
the STAT5 dependent genes are not induced.
Acknowledgments
We thank Professor Danilo F. Gluzman for a
fruitful discussion and the critical reading of
this manuscript. This work was supported by
the National Academy of Science of Ukraine
(grant 0116U007817) and Cancer Foundation
(Sweden).
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Фактор транскрипції STAT5 у В-клітинах крові
при хронічному лімфолейкозі
А. С. Матвєєва, Л. М. Ковалевська,
Т. С. Іванівська, Е. Клейн, О. В. Кашуба
Мета. Встановити причину блокування сигнального
шляху IL2-STAT5 у клітинах крові хворих на хроніч-
ний ліфмолейкоз (ХЛЛ). Методи. Клітини ХЛЛ виді-
ляли з периферичної крові пацієнтів, хворихна ХЛЛ,
за допомогою центрифугування у градієнті фікол-ве-
рографін. Експресію генів STAT1-6 на рівні мРНК
аналізували за допомогою бази даних Oncomine.
Експресію, статус фосфорилювання і клітинну лока-
лізацію білка STAT5 вивчали методом флуоресцентної
мікроскопії з використанням специфічних антитіл.
Результати. На відміну від В-клітин здорових людей,
експресія білка STAT5А була низькою у клітинах хво-
рих на ХЛЛ. Як нами було встановлено раніше, сиг-
нальний шлях IL-2-STAT5 (JAK-STAT5) інгібовано у
клітинах ХЛЛ. Нами було показано низький рівень
фосфорилювання білків STAT5, або повну відсутність
фосфорильованої форми протеїнів в лейкемічних клі-
тинах. Протеїн STAT5А показує цитоплазматичну
локалізацію, що вказує на відсутність у ядрі комплек-
сів, активуючих транскрипцію генів, залежних від
фактора транскрипції STAT5. Висновки. Інгибування
сигнального клітинного шляху IL-2-STAT5 в клітинах
крові хворих на ХЛЛ реалізується за рахунок гіпофос-
форилювання протеїнів STAT5 та/або відсутності ак-
тивних комплексів транскрипції STAT5А у ядрі лей-
кемичних клітин.
К л юч ов і с л ов а: Хронічний лімфолейкоз (ХЛЛ),
В-клітини периферичної крові, STAT5, STAT5A,
STAT5B, сигнальний шлях IL2-STAT5.
Фактор транскрипции STAT5 в В-клетках
крови при хроническом лимфолейкозе
А. С. Матвеева, Л. Ковалевская, Т. C. Ивановская,
Е. Клейн, E. В. Кашуба
Цель. Установить причину блокирования сигнального
пути IL2-STAT5 в клетках крови больных хроническим
лимфолейкозом (ХЛЛ). Методы. Лейкемические клет-
ки выделяли из периферической крови с помощью
центрифугирования в градиенте плотности фиколл-ве-
38
A. S. Matvieieva, L. M. Kovalevska, T. S. Ivanivska et al.
рографин. Экспрессию генов STAT1-6 на уровне мРНК
анализировали с помощью базы данных Oncomine.
Экспрессию, статус фосфорилирования и клеточную
локализацию белка STAT5 изучали методом флуорес-
центной микроскопии с использованием специфиче-
ских антител. Результаты. В отличие от В-клеток
здоровых людей, экспрессия белка STAT5А была низ-
кой в клетках больных ХЛЛ. Как нами было установ-
лено ранее, сигнальный путь IL-2-STAT5 (JAK-STAT5)
ингибирован в клетках ХЛЛ. Нами был показан низкий
уровень фосфорилирования белков STAT5 или полное
отсутствие фосфорилированной формы протеинов в
лейкемических клетках. Протеин STAT5А показывает
цитоплазматическую локализацию, что указывает на
отсутствие в ядре комплексов, активирующих транс-
крипцию генов, зависимых от фактора транскрипции
STAT5. Выводы. Ингибирование сигнального клеточ-
ного пути IL-2-STAT5 в клетках кровибольных ХЛЛ
происходит за счёт гипофосфорилирования белков
STAT5 и/или отсутствия активных комплексов транс-
крипции STAT5А в ядре лейкемических клеток.
К л юч е в ы е с л ов а: Хронический лимфолейкоз
(ХЛЛ), В-клетки периферической крови, STAT5,
STAT5A, STAT5B, сигнальный путь IL2-STAT5.
Received 01.10.2018
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