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NEOPLASIA
From the Department of Haematology, University of
Liverpool, Liverpool, United Kingdom.
Malignant lymphocyte migration into lymph nodes is an important
aspect of chronic lymphocytic leukemia (CLL), yet little is known about
the processes involved. Here we demonstrate that CLL cells migrate
across vascular endothelium in response to at least 3 chemokines,
namely, CCL21, CCL19, and CXCL12. Moreover, transendothelial cell
migration (TEM) in response to CCL21 and CCL19 was significantly higher
for the malignant B cells of patients who had clinical lymph node
involvement as compared with those of patients lacking such
organomegaly. Furthermore, the expression of CCR7, the receptor for
both CCL21 and CCL19, correlated with clinical lymphadenopathy, and
blocking of CCR7 inhibited CLL cell TEM. By using immunohistochemistry we demonstrated that CCL21 and CCL19, but not CXCL12, are located in
high endothelial venules and are, therefore, in an appropriate location
to induce TEM. Regarding the adhesion receptors involved in TEM, Malignant cell adhesion and migration are central
aspects of the pathophysiology of chronic lymphoproliferative
disorders.1-6 In chronic lymphocytic leukemia (CLL), these
processes must determine the pattern and extent of organ involvement,
including that of lymph nodes, but the receptors and ligands involved
are still undefined.
The study is concerned with the mechanism of CLL cell entry into lymph
nodes. This mechanism is important because substantial lymphadenopathy
is a feature of more advanced disease that, in turn, is associated with
a poor prognosis.7 In CLL, the normal architecture of
lymph nodes is completely obliterated by infiltrating malignant cells,
and this obliteration contributes to the immune defect commonly
present in the disease.8 Furthermore, it is likely that
the node provides an environment favoring the growth and survival of
the malignant lymphocytes.9,10 There is, therefore, the
prospect that interfering with CLL cell migration into nodes might have
therapeutic potential.
Although little is known about the mechanisms determining lymph node
enlargement in CLL, this process clearly involves a complex set of
steps, including malignant cell movement into, and within, nodes and
either their enhanced accumulation or reduced exit into lymphatic
channels.11,12 In normal lymphoreticular tissues, these
processes are controlled by lymphocyte responses to distinct chemokines, stromal cells, and extracellular matrix13-15
and by maturation-induced changes in the cells' response to these
microenvironmental signals.11,16,17 High endothelial
venules (HEVs) are an important route of entry of lymphocytes into
lymph nodes. Although HEVs are prominent in CLL nodes and malignant
cells can be observed migrating through these vessels,18
little is known about the chemokines, adhesion molecules, or their
respective receptors involved in this migration process.
Regarding chemokines potentially relevant for CLL cell transendothelial
migration (TEM), we examine the effects of 3 chemokines: CCL21, CCL19,
and CXCL12. CCL21 (also known as secondary lymphoid tissue chemokine,
6Ckine, Exodus 2, and TCA-4) seemed relevant because it is a potent
B-cell chemoattractant19 and is associated with
HEV,20 and because knockout mice lacking receptor (CCR7) for this chemokine display defective B-cell entry into
nodes.21 CCL19 (also known as Epstein-Barr virus-induced
receptor ligand chemokine and macrophage inflammatory protein-3 Other chemokine receptors implicated in B-cell migration are CXCR5,
CXCR3, and CCR6,29-31 but they did not seem relevant for the following reasons. Although CLL cells express CXCR5,32
its ligand CXCL13 (also known as B-lymphocyte chemoattractant and BCA-1) was not investigated because the chemokine is involved in the
formation of follicles rather than in entry of B cells into
nodes.29,33 CXCR3 is also expressed by CLL
cells.30,32 However, although CLL cells migrate in
response to its ligands CXCL9 (also known as monokine induced by Here we present data suggesting that the CCR7 ligands CCL21 and CCL19
are likely to be important for CLL cell transmigration across HEVs into
nodes. Furthermore, the expression levels of CCR7, and also those of
the Patients
Clinical details, including the presence or absence of lymph node
enlargement (> 1cm at 2 or more sites as detected by clinical examination) and VH gene status, are listed in Table
1. This table also lists the relevant
phenotypic data obtained in the present study to allow assessment of
their relationships to clinical disease.
Cell preparation and culture CLL cells. CLL cells were isolated from peripheral blood by Ficoll-Hypaque density gradient centrifugation and stored in liquid nitrogen before use. To ensure high CLL cell purity, only patients with counts of more than 50 × 109/L were studied. Frozen cells were rapidly thawed at 37°C and slowly reconstituted in RPMI containing 1% bovine serum albumin (BSA), 2 mM L-glutamine, 100 U/mL penicillin, and 100 µg/mL streptomycin (Life Technologies, Paisley, United Kingdom). Cells were then allowed to recover for 60 minutes at 37°C in 5% CO2 in air (recovery times of 30, 60, and 120 minutes were tested and transmigration was similar at 60 and 120 minutes and greater than at 30 minutes). In selected patients (n = 3), both fresh and frozen cells were studied. Very similar transmigration was observed (data not shown). Human umbilical vein endothelial cells. Endothelial cells were stripped from the vein with trypsin and were cultured to confluence in Iscoves modified minimal essential medium containing 20% newborn calf serum, 2 mM L-glutamine, 100 µ/mL penicillin, 100 µg/mL streptomycin, and 15 µg/mL endothelial cell growth factor (Life Technologies). Human umbilical vein endothelial cells (HUVECs) that had been passaged up to 3 times were used in the transmigration assays. Chemokines and antibodies The following chemokines were used: CCL21, CCL19, CXCL12, and CXCL8 (interleukin-8) (all from R&D Systems, Oxford, United Kingdom).Antibodies against CCL21 (goat polyclonal), CCL19, and CXCL12 (monoclonal antibody [mAb]; immunoglobulin G [IgG]2b and IgG1, respectively), and vascular cell adhesion molecule 1 (VCAM-1; goat polyclonal) (all from R&D Systems) were used for tissue staining. The following biotinylated second-layer antibodies were used: rabbit antigoat (Vector Laboratories, Peterborough, United Kingdom) and goat antimouse (Zymed, Cambridge, United Kingdom). Nonspecific goat immunoglobulin and mouse IgG1 and IgG2b were used as controls (all from R&D Systems). A mAb (IgM; Pharmingen, Oxford, United Kingdom) was used to examine CLL
cell expression of CCR7, the receptor for both CCL21 and CCL19;
antibody staining was detected with fluorescein isothiocyanate (FITC)-conjugated goat antimouse immunoglobulin. An FITC-conjugated anti-CD19 (IgG1; Becton Dickinson, Oxford, United Kingdom) was used to
determine both CLL cell purity and the nature of cells migrating in the
TEM system (because only very high count cases were used,
proportionally very few normal B cells were present in the cell
preparations). For both mAbs, fluorescence was measured by
fluorescence-activated cell sorter (FACS), with inclusion of nonspecific IgM and IgG1 (Becton Dickinson) as class-specific controls.
Expression of In addition, mAbs were used to determine which integrins and chemokine
receptors might be involved in TEM. Blocking mAbs against the following
were used: TEM assay HUVECs were grown to confluence on the inserts of Transwell plates (5-µm pore size; Corning Costar, High Wycombe, United Kingdom). The HUVECs were washed in RPMI, and 5 × 105 CLL cells (in RPMI containing 0.1% BSA, 2 mM L-glutamine, 100 U/mL penicillin, and 100 µg/mL streptomycin) were added to the inserts. Chemokines were then added to the bottom wells at the following concentrations: CCL21 and CCL19 at 10, 100, 1000, and 2000 ng/mL; CXCL12 at 10, 100, and 1000 ng/mL; and CXCL8 at 0.5, 5, and 50 ng/mL (all in RPMI containing 0.1% BSA, 2 mM L-glutamine, 100 U/mL penicillin, and 100 µg/mL streptomycin). It was shown in preliminary experiments that maximal migration indices (MIs; more to follow) were observed at CCL21 and CCL19 concentrations of 1000 ng/mL and at a CXCL12 concentration of 100 ng/mL. These concentrations of chemokine were, therefore, used in all subsequent experiments. No TEM was observed at any of the concentrations of CXCL8 (previous work from this department1 has shown that CXCL8 at 5 ng/mL induces maximal CLL cell movement on the extracellular matrix component hyaluronan). A range of incubation times was also tested (2, 4, 6, 8, and 24 hours); the MI was maximal at 6 hours and remained constant until 8 hours; therefore, an incubation time of 6 hours was routinely used. All assays were performed in triplicate.After incubation, the undersides of the inserts were scraped to remove any cells that had recently transmigrated, and the cells were then harvested from the bottom wells. As some CLL cells had adhered to the bottom wells, EDTA (0.2%; Sigma, Poole, United Kingdom) was added for 5 minutes at 37°C before harvesting. These transmigrated cells were then counted in a hemocytometer. To determine the CLL cell content of the transmigrated cells, an aliquot was always stained for CD19 and analyzed by FACS. The MI (no. of CD19+ cells transmigrating with chemokine divided by no. of cells transmigrating in the absence of chemokine) was then calculated. Student t test was used to determine the statistical significance of the results. Inhibition of TEM CLL cells were incubated at 4°C for 30 minutes with blocking mAbs (concentrations) to the following integrin chains: 1 (1 and 10 µg/mL), 2 (1 and 10 µg/mL), 7 (1 and 10 µg/mL),  4 (0.5 and 5µg/mL), 5 (0.2 and 2 µg/mL), and L (1 and 10 µg/mL).
Also, in certain experiments a blocking mAb to CCR7 was used (50 and 100 µg/mL). These amounts of mAbs were chosen to cover the range of
concentrations that have been reported to block adhesion in other
systems (manufacturers' recommendations and M. Lipp, written personal
communication, October 2001). CLL cells were then added to the
Transwell inserts, and the TEM assay was performed as above.
In addition, CLL cells were incubated for 2 hours at 37°C with 1 µg/mL pertussis toxin (Sigma) to block chemokine receptor signaling and were washed. TEM measurements were performed as above. Tissue staining CLL nodes (n = 5) were diagnostic samples. Normal nodes (n = 5) were obtained from axillary clearance for breast cancer and were macroscopically and microscopically normal.Formalin-fixed paraffin-embedded tissue was stained for chemokines as follows. After clearing and rehydration, slides were boiled in 10 mM sodium citrate buffer (pH 6) for 10 minutes and blocked with 10 mg/mL BSA before overnight incubation with antibodies to CCL21 (20 µg/mL) and VCAM-1 (1:200) at 4°C, or with mAbs to CCL19 (5 µg/mL) and CXCL12 (50 µg/mL) at 25°C. These amounts of the antibodies were chosen after titration over a range of concentrations. Sections were then incubated with biotinylated rabbit antigoat (CCL21 and VCAM-1) or with biotinylated goat antimouse (CCL19 and CXCL12) antibodies, followed by ExtraAvidin alkaline phosphatase (Sigma) and exposure to substrate (Fast Red/Naphthol AS MX phosphatase/levamisole; Sigma). Slides were counterstained with hematoxylin (Sigma). VH hypermutation analysis For each case, total RNA was extracted from CLL cells by using Trizol reagent (Life Technologies) and 1-µg aliquots were reverse transcribed with M-MuLV reverse transcriptase (Promega, Southampton, United Kingdom) and an oliog(dT)15 primer. Aliquots of the resulting complimentary DNAs were used to isolate clonally expressed VHDHJH sequences in 2 sets of VH gene family-specific polymerase chain reaction (PCR) reactions. In the first set of 7 reactions, the sense primers were consensus sequences derived from the framework region 1 (FWR1)36 of each of the 7 families. In the second set of 6 reactions, the sense primers were from the leader regions (families 1 and 7 with a common primer37). In all reactions, the same combination of 3 constant region-specific (,  , µ) antisense primers were used.36 The 50-µL reactions (20 pmol of
each primer, 1.5 mM MgCl2, 100 µM of each dNTP, and 2.5 U
Taq Polymerase [in supplied buffer; Promega]) were cycled 30 times by
using a touch-down protocol,38 with the annealing
temperature reducing from 63°C to 57°C over the first 12 cycles and
then analyzed by electrophoresis. Products for a given family from 2 separate PCR reactions were cloned. Between 2 and 4 clones derived from
each PCR (ie, a total of 4 to 8 cases from each patient) were sequenced
commercially with a vector-specific primer. The sequences were accepted
as representing the CLL clone if there was identity between and within FWR1 and the leader sequence-derived PCR clones.
The extent of VH gene hypermutation was determined by using VBASE.39 For the purpose of assigning prognostic significance to the extent of VH hypermutation, previous studies have used a threshold of either 2% or 5% divergence from the nearest germline IgVH gene to define hypermutation.36,40 Therefore, for our studies of the relationship between VH gene hypermutation and TEM/lymphadenopathy, we have used both the 2% and 5% hypermutation thresholds for our data analyses.
CCL21, CCL19, and CXCL12 all induce CLL cell TEM In the absence of chemokine, CLL cells showed little TEM (< 0.2%) in the Transwell migration assay. Variable CLL cell TEM (0%-12% of input cells) was seen in response to CCL21, CCL19, and CXCL12 (n = 8 cases; Figure 1). When CLL cell migration was observed, CCL21 and CCL19 produced comparable MIs, which were usually higher than those induced by CXCL12. As expected from our previous work showing that CXCL8 stimulates CLL cell movement within, but not into nodes,1 this chemokine failed to induce CLL cell TEM in any of the cases studied (Figure 1).
Because the TEM induced by CCR7 stimulation seemed of potential in vivo relevance (more to follow), the reproducibility of cell migration in response to CCL21 was studied on 2 or more occasions in all 8 cases. When the MI was low (< 10; n = 4), it remained low on repeated testing. Similarly, when the MI was high (> 20; n = 4), it remained high but varied at different times of study (eg, in case 7 studied 5 times, the MI was between 48 and 120). CCL21-induced CLL cell TEM was completely abrogated by preincubation of CLL cells with pertussis toxin (MI = 0.7 ± 0.3; n = 3), an inhibitor of chemokine-receptor signaling. Clinical analysis of the patients studied in Figure 1 indicated that the 4 patients whose cells repeatedly migrated in response to CCL21 and CCL19 all had lymph node enlargement, whereas those cases whose cells displayed little or no transmigration had no lymphadenopathy. Because at the onset of the present study CCL21, known to be concentrated in HEV,20 seemed to be the more likely candidate chemokine to induce CLL cell TEM in vivo, we examined the relationship between migration to CCL21 and clinical lymphadenopathy in a larger number of patients. CCL21-mediated TEM of CLL cells is related to clinical lymphadenopathy When an additional 22 CLL patients were studied, the mean MI was significantly higher in patients with nodal enlargement compared with those without lymphadenopathy (31 versus 10; P < .01; Figure 2A). This finding demonstrates that the ability of CLL cells to cross endothelium is related to the presence of clinical lymphadenopathy. As expected, because lymphadenopathy is one of the features used in clinical staging, there was also an association between high MI and later stages of the disease (stage 0 versus I-IV = 8.4 ± 0.2 versus 27 ± 6, P = .002).
In view of the recent interest in the fact that CLL can be divided into 2 very different prognostic groups according to the extent of VH gene hypermutation, we next examined the relationship between such hypermutation and TEM. In the patients (n = 26) for whom VH data were available, there was no significant difference (P > .05; Figure 2B) between CLL cell TEM in patients with or without hypermutation regardless of whether a threshold of 2% or 5% deviation from the germline was used in the analysis. However, more CLL patients with clinical lymphadenopathy had nonhypermutated VH genes (10 of 15 at < 2% VH mutation and 13 of 15 at < 5%VH mutation) than did those without nodes (3 of 11 at < 2% VH mutation and 4 of 11 at < 5% VH mutation). When the proportions were analyzed by Fisher exact test, the association between lymphadenopathy and VH nonhypermutation was significant when the 5% mutation threshold was used (P = .01) but not at the 2% threshold level (P = .11). We, therefore, concluded that the ability of CLL cells to migrate into, or accumulate in, nodes may be related to their VH hypermutational status. Because expression of CD38 is a poor prognostic indicator in CLL40-42 and is often associated with the absence of VH hypermutation,40 we also examined the relationship between CD38 expression, TEM, and nodal disease. We found that, although there was a trend toward an association between high CD38 expression and lymphadenopathy (mean ± SE for patients with lymphadenopathy = 20.8% ± 6.3% CD38+ cells; for those without enlarged lymph nodes 6.8% ± 4.6%), it did not reach statistical significance (P = .08). There was no association between high CD38 expression (> 30% positive cells) and TEM (Figure 2C; n = 27; P = .29), reflecting that not all patients with lymphadenopathy displayed high CD38 expression, and yet in all cases the malignant cells migrated in response to CCL21. In conclusion, the data show that clinical lymphadenopathy is clearly related to the ability of CCL21-stimulated CLL cells to undergo TEM. In addition, it seems that less mature non-VH hypermutated and CD38+ CLL cells have a greater propensity to accumulate in nodes than do more mature VH hypermutated and CD38 cells. We next investigated the mechanism(s) responsible for the different CLL cell migratory responses to CCL21 observed in patients with lymphadenopathy, compared with those lacking lymph node enlargement. It has been shown in adult T-cell leukemia that CCL21 receptor (CCR7) expression is related both to the malignant cell chemotactic response to CCL21 and to organ involvement in the disease.43 We, therefore, examined the CCR7 expression of the CLL cells in our series of patients and related receptor levels to the presence or absence of lymphadenopathy. CCR7 expression is higher in patients with lymphadenopathy Variable CCR7 expression was detected on the CLL cells from all of the patients examined. In most cases, a distinct positive peak consisting of more than 95% of cells was observed. In 5 cases in which the cells had a mean fluorescence intensity (MFI) less than 20, a peak shift was seen, indicating that virtually all (> 95%) of the cells were weakly positive.The intensity of CCR7 expression was significantly higher on the CLL
cells of patients with lymphadenopathy compared with those without
lymph node enlargement (Figure 3 and
Table 1). This finding strongly suggests that transmigration to lymph
nodes involves stimulation of CCR7. We, therefore, used an anti-CCR7 blocking mAb in TEM experiments and showed that this reagent strongly reduced CLL cell migration to CCL21 (Figure 3B).
Because both CCL21 and CCL19 are ligands of CCR7, we next examined the expression of these 2 chemokines in CLL and normal nodes. In addition, because CXCL12 also stimulated CLL cell TEM, we examined the distribution of this CXCR4- binding chemokine. CCL21 and CCL19, but not CXCL12, are associated with HEVs In CLL nodes, the normal architecture is completely replaced by infiltrating leukemic cells among which a variety of vascular and lymphatic channels are visible.8In both CLL and normal nodes (Figure 4),
weak staining for both CCL21 and CCL19 was found in HEVs; in contrast,
these vessels were negative for CXCL12. The stroma was reactive for all
these chemokines with some stromal cells, including the peri-HEV
fibroblasts, showing strong staining for CCL21 (Figure 4).
We, therefore, concluded that the location of CCL21 and CCL19, but not CXCL12, indicated that these chemokines can mediate CLL cell entry into lymph nodes. Thus, these histologic findings add relevance to our in vitro observations that migration to CCL21 and CCL19, and expression of the receptor (CCR7) for these chemokines, is related to the presence of lymphadenopathy in CLL. However, although the levels of CCR7 were significantly different between the patients with and without lymphadenopathy, there was some overlap in expression between the 2 groups. We, therefore, examined other factors that might be involved in determining the entry of CLL cells into lymph nodes in vivo. We first examined the expression of L-selectin by CLL cells because this molecule is essential for the initial tethering of lymphocytes to HEVs before integrin-mediated arrest and subsequent diapedesis across endothelium. L-selectin expression by CLL cells is not correlated with clinical lymphadenopathy FACS analysis showed that L-selectin expression by CLL cells varies greatly from case to case. In all patients studied, at least a proportion (16%-82%) of cells expressed variable levels of L-selectin (MFI, 30-176). However, there was no correlation between the percentage positivity (P = .3), or MFI (P = .5), and the presence or absence of lymphadenopathy (data not shown).As regards integrin involvement in lymphocyte TEM, the most important
receptors are CLL cell TEM depends on 1 or 2 integrins (data not shown). As expected, in all 3 cases,
inhibition of CLL cell TEM was seen with anti-4 or anti-L mAbs,
whereas anti-5 mAb and isotypic control antibodies had no effect
(Figure 5). Blocking with a combination
of antibodies (anti-1 + anti-2 or anti-L + anti-4) was not more effective than blocking each integrin chain
alone (data not shown; n = 3). Because a study46
implicated 47, as well as 41, in normal lymphocyte
migration into peripheral lymph nodes, we also tested the effect of a
blocking anti-7 mAb on CLL cell TEM. Inhibition was only seen in one
of the 3 cases studied (data not shown).
Having shown that
4 as detected by FACS analysis with the use of a
sensitive triple-layer technique (Figure
6; Table 1). Whereas, in patients without
lymphadenopathy, no 4 expression was observed in 14 of 15 patients
studied (Figure 6; Table 1); in all 14 cases, the MFI of the test cells
was identical to that of cells stained with isotypic control antibody.
In the single patient without nodal enlargement whose cells expressed
4, CCR7 expression was low (MFI = 18 compared with a MFI of 7 for
cells stained with isotypic control mAb).
The Regarding Vascular endothelial cells in CLL node express VCAM-1 In CLL nodes, VCAM-1 staining was observed in HEVs and other vascular endothelial cells (Figure 7). Essentially similar staining was observed in normal lymph nodes (Figure 7).
The present study is concerned with the process by which CLL cells enter lymph nodes. We demonstrated that a number of relevant chemokines (CCL21, CCL19, and CXCL12) can induce the malignant cells of a proportion of cases of CLL to migrate through vascular endothelium. However, we found that CCL21 and CCL19 induce more CLL cells to transmigrate than does CXCL12. Furthermore, we found that the ability of CLL cells to respond to CCL21 and CCL19 in vitro strongly correlates with the presence of clinical lymphadenopathy. We then went on to show that, although all CLL cells possess the receptor for these chemokines (CCR7), levels of expression are higher on the cells of patients with lymph node enlargement. These results are entirely compatible with a study in adult T-cell leukemia43 which found that the ability of the malignant cells to respond to CCL21 correlated with CCR7 expression and lymph node enlargement. Moreover, by using a blocking antibody, we now directly demonstrate involvement of this receptor in CLL cell TEM in response to CCL21. In view of the current interest in the pathogenetic and prognostic significance of VH hypermutational status and CD38 expression in CLL,40,41,47,48 and because lymph node enlargement is a feature of more advanced disease and poor prognosis, we also examined whether these parameters are related to the degree of in vitro TEM and clinical lymphadenopathy. We found that there was no correlation between VH hypermutation and TEM, but more patients with lymphadenopathy had nonmutated VH genes than did patients without nodes. These findings are compatible with previous reports that lymphadenopathy can be found in patients with or without VH hypermutation.40,48 As for CD38 expression, the cells of patients with enlarged lymph nodes expressed more CD38 and had higher levels of TEM than did patients without lymphadenopathy. However, only the relationship between VH nonhypermutation (< 5%) and lymphadenopathy achieved statistical significance. With regard to CD38 expression and lymphadenopathy, it is interesting that a recent report of a very large number of CLL cases showed a significant positive correlation between these 2 parameters.42 By using immunohistochemistry, we next investigated the location of CCL21, CCL19, and CXCL12 in CLL and normal lymph node tissue. CCL21 and CCL19 were found in both HEVs and the surrounding stroma, whereas CXCL12 was found in stroma only. These findings, together with our demonstration of the correlation between CCR7 expression and lymphadenopathy, suggest that CCL21 and/or CCL19 are likely to be involved in the stimulation of CLL cell entry into lymph nodes. These conclusions are in accord with studies in knockout mice, suggesting that CCR7 is important for the migration of B cells into nodes.21 Furthermore, our immunohistochemical observations are compatible with previous studies in mice showing that, of the 2 chemokines, CCL21 is produced in HEV20 and that CCL19 produced by stromal cells is transcytosed through endothelium to the luminal surface of HEVs.49 Although our data suggest that CCR7 ligand(s) are important in CLL cell TEM, they do not rule out the possibility that other chemokines such as CXCL13 (for which CLL cells express receptor CXCR532) may be involved in the lymphadenopathy of the disease. However, this possibility seems unlikely because CLL nodes lack follicles, the formation of which critically depends on the presence of CXCL13.29,33 It has previously been reported that CXCL12 stimulates in the TEM of CLL cells, and it was proposed that this chemokine may be involved in the migration of CLL cells into the bone marrow.27,28 Our results confirm that CXCL12 is able to induce CLL cell TEM, but our immunohistochemical demonstration that the chemokine is not present in HEVs suggests that CXCL12 is not involved in CLL cell entry into nodes. It, therefore, seems likely that different chemokines direct CLL cells into distinct lymphoreticular tissues. By showing that CCR7 stimulation is likely to be important for CLL cell
entry into lymph nodes and in the generation of clinical lymphadenopathy, we next focused on the adhesive interactions involved.
This focus seemed all the more important, given that there was some
overlap in the levels of CCR7 expression in patients with and without
lymphadenopathy. After demonstrating that L-selectin expression,
despite being variable, did not correlate with clinical lymphadenopathy, we next concentrated on integrins. Both There have been a number of previous studies of integrin expression on
CLL cells.5,45,53 Most demonstrate In conclusion, then, the present study suggests that CCR7 engagement by
CCL21 and/or CCL19 stimulates CLL cell entry into lymph nodes by HEV
and that CCR7 and
We thank Dr M. Lipp of the Max Delbruck Centrum, Berlin, for his kind gift of a blocking CCR7 antibody (3D12).
Submitted July 19, 2001; accepted November 30, 2001.
Supported by the Leukaemia Research Fund (United Kingdom) (M.Z. and J.C.C.).
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: Kathleen J. Till, Department of Haematology, Royal Liverpool University Hospital, Daulby Street, Liverpool, L69 3GA, United Kingdom; e-mail: k.j.till{at}liv.ac.uk.
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| Copyright © 2002 by American Society of Hematology Online ISSN: 1528-0020 | |||||||||
4 integrin are important for
migration of chronic lymphocytic leukemia cells into lymph
nodes
Top
Abstract
Introduction
n = 20). The bar represents the average
MI of each patient group. VH
