CD100/Plexin-B1 interactions sustain proliferation and survival of normal and leukemic CD5+ B lymphocytes

Blood ◽  
2003 ◽  
Vol 101 (5) ◽  
pp. 1962-1969 ◽  
Author(s):  
Luisa Granziero ◽  
Paola Circosta ◽  
Cristina Scielzo ◽  
Elisa Frisaldi ◽  
Stefania Stella ◽  
...  
Keyword(s):  
B Cells ◽  
B Lymphocytes ◽  
Lymphocytic Leukemia ◽  
Leukemic Cells ◽  
Follicular Dendritic Cells ◽  
Growth And Survival ◽  
High Affinity Receptor ◽  
Activated T Lymphocytes ◽  
Affinity Receptor

Growth and survival of chronic B-cell tumors are favored by the malignant cell's capacity to respond to selected microenvironmental stimuli provided by nontumoral bystander cells. To investigate which mechanisms operate in these crosstalks and whether they are malignancy-related or reproduce the mechanisms used by normal B cells we have studied the expression and functional role of semaphorin CD100 (now called Sema4D) in chronic lymphocytic leukemia (CLL) cells and normal CD5+ B cells. We demonstrate here that (1) leukemic and normal CD5+ B lymphocytes uniformly express CD100; (2) the CD100 high-affinity receptor Plexin-B1 is expressed by bone marrow stromal cells, follicular dendritic cells, and activated T lymphocytes, and is thus available to CD100+ lymphocytes in different specific microenvironments; and (3) upon interaction between CD100 and Plexin-B1 both CLL and normal CD5+ B cells increase their proliferative activity and extend their life span. These findings establish that Plexin-B1 is an easily accessible receptor for CD100 within the immune system. The encounter of CD100+ leukemic cells with Plexin-B1 may promote the proliferation and survival of malignant cells. The crosstalk operated by the CD100/Plexin-B1 interaction is not malignancy related but reproduces a mechanism used by normal CD5+ B cells.

scholarly journals Interleukin-15 promotes the growth of leukemic cells of patients with B- cell chronic lymphoproliferative disorders

Blood ◽  
1996 ◽  
Vol 87 (8) ◽  
pp. 3327-3335 ◽  
Author(s):  
L Trentin ◽  
A Cerutti ◽  
R Zambello ◽  
R Sancretta ◽  
C Tassinari ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
B Lymphocytes ◽  
Nk Cell ◽  
Lymphoproliferative Disorders ◽  
Lymphocytic Leukemia ◽  
Leukemic Cells ◽  
Interleukin 15

The recently discovered cytokine, interleukin-15 (IL-15), has been demonstrated to share several biologic properties with IL-2 in different cell systems, including T-cell and natural killer (NK) cell compartments. As for B lymphocytes, IL-15 has been shown to provide stimulatory activities in normal preactivated B cells that are mainly transduced through IL- 2 receptor (IL-2R) complex components. Since leukemic B cells from patients with chronic lymphoproliferative disorders (CLD) bear IL-2R and grow in response to IL-2, we investigated whether IL-15 triggers the proliferation of malignant B cells obtained from 12 patients with B-cell chronic lymphocytic leukemia (B-CLL) and five patients with hairy cell leukemia (HCL). Enriched B cells recovered from five healthy subjects were also studied as controls. IL-15 stimulated the proliferation of freshly isolated leukemic B cells, but not resting normal B lymphocytes, the latter being able to grow in the presence of IL-15 only after in vitro preactivation with phorbol myristate acetate. The proliferation elicited by IL-2 on leukemic cells was comparable to that determined by IL-15. Following addition of graded concentrations of IL-15 to low/intermediate-dose IL-2, resting leukemic B cells showed a higher stimulatory rate than that observed using the two cytokines separately. In normal resting B lymphocytes, this cumulative effect was not observed. The role of different IL-2R subunits in IL-15-driven growth of malignant B cells was investigated both by their expression on leukemic cells and by the block of different IL-2R subunits (p55, p75, and p64) with specific monoclonal antibodies (MoAbs). Using flow cytometry and reverse transcriptase-polymerase chain reaction (RT-PCR) analyses we demonstrated that both B-CLL and HCL leukemic B cells express the beta and gamma chains of the IL-2R system. The stimulatory activity achieved by IL-15 decreased significantly, blocking the beta and gamma chains of the IL-2R. Taken together, these findings demonstrate that IL-15 triggers the growth of leukemic B cells through IL-2R system subunits, pointing to the role of this novel cytokine in regulating the neoplastic proliferation in CLD.

Different Expression of FcgammaRIIb in Chronic Lymphocytic Leukemia and Human Normal B Lymphocytes

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3134-3134
Author(s):  
Carol Moreno ◽  
Rajendra Damle ◽  
Sonia Jansa ◽  
Gerardo Ferrer ◽  
Pau Abrisqueta ◽  
...  
Keyword(s):  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
B Lymphocytes ◽  
Surface Membrane ◽  
Memory B Cells ◽  
Lymphocytic Leukemia ◽  
Cd38 Expression ◽  
B Cell Subsets

Abstract The Fcgamma receptors (FcγRs) are a family of molecules that modulate immune responses. FcγRIIb is an inhibitory FcγR that bears immunoreceptor tyrosine-based inhibitory motifs which transduce inhibitory signals on coligation with the surface membrane Ig of the B-cell antigen receptor (BCR). The role of FcγRIIb in controlling B cell activation through inhibition of BCR signaling has been extensively studied in animal models. Nevertheless, data on FcγRIIb are scant in human normal and neoplastic B cells, this being due to the lack of a specific antibody for human FcγRIIb. Consequently, there is little information on this receptor in chronic lymphocytic leukemia (CLL). Considering the activated nature of CLL cells and the central role of the BCR in the biology of the disease, studies of FcγRs are warranted. We used a novel specific mAb directly conjugated with Alexa 488 fluorophore that solely reacts with the human FcγRIIb (MacroGenics, Inc.) to investigate the receptors expression on CLL and normal human B cells. The study population included 84 patients with CLL and 24 age- and sex-matched controls. FcγRIIb expression was assessed as the mean fluorescence intensity (MFI) of surface membrane staining. In CLL cells, FcγRIIb was measured on CD19+CD5+ cells in combination with CD38, CD49d or CD69. Normal B cells were immunostained for CD19, CD5, IgD and CD38 expression and B cell subsets: naïve (IgD+CD38−), activated (IgD+CD38+) and memory B cells (IgD−CD38−) were studied for their relative expression of FcγRIIb. FcγRIIb expression was found significantly higher in naïve B cells compared to activated and memory B cells [median MFI: 17420 (11960–21180) vs. 11.140 (7899–16970) and 11.830 (6984–17100); p<0.001]. Significant differences were also observed between CD5− and CD5+ normal B cells. In contrast, FcγRIIb expression was lower in CLL cells than in CD5+ and CD5− normal B lymphocytes [median MFI: 6901(1034–42600), 10180 (5856–14820) and 12120 (7776–16040); p<0.05)]. Interestingly, FcγRIIb expression was variable within individual CLL clones, this being higher in CD38+ and CD49d+ cells than in CD38− and CD49d− cells (p<0.05). Furthermore, the highest density of FcγRIIb was observed on those cells which coexpressed CD38 and CD49d. In contrast, no significant differences were observed between FcγRIIb and the expression of the activation antigen CD69. Although CD69 and CD38 expression was significantly higher on unmutated IGHV cases, no correlation was found between FcγRIIb levels and IGHV mutational status. Similarly, there was no correlation between FcγRIIb and other poor prognostic variables such as ZAP-70 (≥20%), CD38 (≥ 30%) or high risk cytogenetics. Nevertheless, cases with ≥ 30% CD49d+ cells had higher FcγRIIb expression than those with <30% CD49d+ cells (p=0.006). The findings presented in this study suggest a hierarchy of FcγRIIb expression in normal B-cells, CLL cells and their subpopulations: circulating normal CD5− B cells > circulating normal CD5+ B cells > circulating CD5+ CLL B cells. In addition, although FcγRIIb is present on all normal B cell subsets its expression is higher in naïve B cells. Furthermore, in CLL FcγRIIb density is greater in CD38+ and CD49d+ cells within the clone. Although CD49d and FcγRIIb on CLL clones is linked in a direct manner, there is no relationship with FcγRIIb density and IGHV mutations, ZAP-70, CD38 and unfavorable cytogenetic markers. Finally, the relationship between FcγRIIb expression on CLL cells and functional responses to BCR and other receptor-mediated signals deserve further investigation.

scholarly journals Interleukin 10 (IL-10) upregulates functional high affinity IL-2 receptors on normal and leukemic B lymphocytes.

1993 ◽  
Vol 178 (5) ◽  
pp. 1473-1481 ◽  
Author(s):  
A C Fluckiger ◽  
P Garrone ◽  
I Durand ◽  
J P Galizzi ◽  
J Banchereau
Keyword(s):  
B Cells ◽  
B Cell ◽  
B Lymphocytes ◽  
Flow Cytometric Analysis ◽  
Interleukin 10 ◽  
Lymphocytic Leukemia ◽  
High Affinity ◽  
Cell Proliferation And Differentiation ◽  
Proliferation And Differentiation ◽  
High Affinity Receptor

Interleukin 10 (IL-10) has recently been shown to induce normal human B lymphocytes to proliferate and differentiate into immunoglobulin (Ig)-secreting cells. Herein, we show that IL-10 also promotes DNA synthesis and IgM production by anti-CD40 activated B cell chronic lymphocytic leukemia (B-CLL). Most strikingly, IL-2 and IL-10 were found to synergize to induce the proliferation and differentiation of B-CLL cells. This synergy between IL-2 and IL-10 was also observed with normal B cells which proliferated strongly and secreted large amounts of IgM, IgG, and IgA. The observed synergy is likely to be due to the IL-10-induced increase of high affinity IL-2 receptors on both normal and leukemic B cells. This increase of high affinity receptor is associated to an increase of Tac/CD25 expression that can be detected by flow cytometric analysis. Taken together, these results indicate that IL-10 permits anti-CD40 activated B cells to respond to IL-2 through an induction of high affinity IL-2 receptors. This effect of IL-10 may partly explain how T cells, which activate B cells in a CD40-dependent fashion, induce B cell proliferation and differentiation mostly through IL-2.

Telomere length and telomerase activity delineate distinctive replicative features of the B-CLL subgroups defined by immunoglobulin V gene mutations

Blood ◽  
2004 ◽  
Vol 103 (2) ◽  
pp. 375-382 ◽  
Author(s):  
Rajendra N. Damle ◽  
Franak M. Batliwalla ◽  
Fabio Ghiotto ◽  
Angelo Valetto ◽  
Emilia Albesiano ◽  
...  
Keyword(s):  
B Cells ◽  
Telomere Length ◽  
B Lymphocytes ◽  
Gene Mutations ◽  
Telomerase Activity ◽  
Lymphocytic Leukemia ◽  
Leukemic Cells ◽  
Survival Times ◽  
V Gene ◽  
Cellular Replication

Abstract Patients with B-cell chronic lymphocytic leukemia (B-CLL) segregate into subgroups with very different survival times. Because clinical observations suggest that leukemic cells accumulate at different rates, we measured telomere length and telomerase activity in B-CLL cells to distinguish differences in cellular replication. Our data indicate that the telomeres of B-CLL cells are shorter than telomeres of B cells from healthy subjects, indicating that the leukemic cells have a prolonged proliferative history. Leukemic cells of the immunoglobulin V gene mutation subgroups differ in telomere length and telomerase activity. B lymphocytes from the subgroup with poor outcome and with limited IgV gene mutations have uniformly shorter telomeres and more telomerase activity than those from the subgroup with better outcome and with considerable mutations. Differences in telomere length appear to largely reflect the proliferative histories of precursors of the leukemic cells, although differences in cell division, masked by the action of telomerase, cannot be excluded. These results may provide insight into the stages of maturation and the activation pathways of the cells that give rise to B-CLL. In addition, they reinforce the concept that B-CLL is not simply an accumulative disease of slowly dividing B lymphocytes but possibly one of B cells with extensive proliferative histories.

Role of Hematopoietic-Specific Protein 1 (HS1) in Apoptosis in B-Chronic Lymphocytic Leukemia.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2809-2809
Author(s):  
Livio Trentin ◽  
Antonella Contri ◽  
Anna Maria Brunati ◽  
Federica Frezzato ◽  
Martina Frasson ◽  
...  
Keyword(s):  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
Cell Survival ◽  
B Lymphocytes ◽  
Lymphocytic Leukemia ◽  
Leukemic Cells ◽  
Cell Fractionation ◽  
Normal Peripheral Blood ◽  
Protein Levels ◽  
The Mean

Abstract B-cell chronic lymphocytic leukemia (B-CLL) is the most common leukemia in adults and is characterized by the accumulation of clonal CD5+ B lymphocytes. Several protein kinase pathways have been claimed to be involved in the regulation of apoptosis and cell survival. We previously demonstrated that Src kinase Lyn is overexpressed at the protein level in leukemic cells as compared to normal B lymphocytes with substantial amount of the kinase anomalously present in the cytosol. Moreover, most of Lyn is constitutively active in resting leukemic cells and is poorly responsive to BCR engagement. The finding that B CLL cells contained cytosolic Lyn fraction and are defective in programmed cell death suggest that the tyrosine phosphorlation of specific cytosolic targets might account, at least in part, for cell resistance to apoptosis. The 75 KDa HS1 protein is one of the major substrate of Lyn kinase upon BCR cross-linking that plays a crucial role in BCR- induced apoptosis in the mouse B lymphoma cell line WEHI-231. A recent study demonstrates that most HS1 protein was constitutively phosphorylated in B CLL patients with poor prognosis whereas only a fraction was phosphorylated in patients with good prognoses. In the present study, the relative HS1 protein levels were measured by Western blot analysis in 50 CLL patients belonging to different clinical stages. The relative HS1 protein levels were compared with corresponding levels in normal peripheral blood and with Jurkat cells. For normal B cells, the mean ± SD for HS1: actin ratio was 0,88 ± 0,10. There was considerable variation in the levels of HS1/actin ratio in CLL cells, which ranged from 0,49 to 2,50. Thus, compared to normal B cells, 15 CLL patients had a HS1 level which fell within the mean ± 1SD HS1 levels for normal B cells, while 9 patients had lower levels and 26 patients had higher levels. When assessed by flow cytometry, HS1 expression was normally distributed among CLL cells in individual patients and the mean levels correlated with those obtained by Western blotting. A difference in the levels of HS1 was also observed between mutated and unmutated patients. Using confocal microscopy and subcellular cell fractionation, we observed that HS1 protein was abnormally distributed in malignant cells as compare with normal B cells: a 4–7% aliquot of HS1 was anomalously present in the nucleus of leukemic cells. When primary CLL cells were in vitro treated whith dexamethazone, cyclosporin A, chlorambucil, or fludarabine the HS1 levels decreased correlating with the sensitivity of these cells to undergo apoptosis. Using a polyclonal antiserum against HS1 a major cleavage product of the apparent molecular weight of 64 KDa and one minor product of approximately 46 Kda was detected in B CLL cells cultured for 24 hours whith drugs. These findings suggest that HS1 plays a pivotal role in the regulation of cell survival of leukemic B cells and suggest that HS1 might represent a target for the development of new drugs to be used in vivo in these patients.

High Expression of Haematopoietic Cell Specyfic Lyn Substrate-1 (HS1) Predicts Survival of B-Cell Chronic Lymphocytic Leukemia Patients

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2853-2853
Author(s):  
Aleksandra Butrym ◽  
Miroslaw Majewski ◽  
Justyna Dzietczenia ◽  
Tomasz Wrobel ◽  
Kazimierz Kuliczkowski ◽  
...  
Keyword(s):  
Prognostic Factors ◽  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
Patient Survival ◽  
Clinical Course ◽  
Lymphocytic Leukemia ◽  
Leukemic Cells ◽  
Mutational Status ◽  
Cell Chronic Lymphocytic Leukemia

Abstract Abstract 2853 B-cell chronic lymphocytic leukemia (B-CLL) is the most common leukemia in adults in western countries. It is characterized by B lymphocyte accumulation in peripheral blood, bone marrow, lymph nodes and other lymphatic organs. Leukemic cells derive most commonly from B lymphocytes, rarely from T or NK cells. B-CLL is known from its heterogeneous clinical course from indolent to very aggressive. In spite of many known prognostic factors (such as immunoglobulin heavy chain gene mutational status – IgVH, expression of ZAP70 and CD38), is still difficult to classify a single patient to particular risk group and to predict CLL clinical course. That is why new prognostic factors are still needed. HS1 (hematopoietic cell specific Lyn substrate-1) is an intracellular protein, which expression occurs mainly in hematopoietic cells. HS1 plays an important role in regulating T cell immune synapse and affects many functions of NK cells, including the lysis of target cells, adhesion, chemotaxis and clustering of actin in the lytic synapse. The role of HS1 in B cells is poorly understood. This protein was identified in B cells as the primary receptor substrate for phosphorylation by BCR after antigenic stimulation. Other studies have confirmed the role of HS1 in the process of clonal expansion and deletion induced by antigen-receptor interaction in B cells and T. HS1 is rapidly phosphorylated in B cells in the vicinity of tyrosine residues and is a substrate for tyrosine kinases: the Src family and Syk, including Lyn, FGR, Fyn and Lck. It has been shown that HS1 interacts with the cell cytoskeleton in both: normal and leukemic B cells. HS1 protein is an important regulator of motility, migration and adhesion of leukemic cells and is involved in cytoskeleton rearrangement. HS1 can have impact on homing and migration of CLL cells. It can indirectly promote disease progression and influence patient survival. The aim of this study was to evaluate HS1 expression in CLL patients in connection with other known prognostic factors and patient survival. Material and methods: 92 untreated CLL patients (45 women and 47 men), aged between 42 and 88 years (median age 67 years), were included into the study. Diagnosis was made basing on typical clinical, hematological and immunophenotypical picture. The control group was consist of 28 healthy matched people (11 men and 17 women), aged between 36 and 79 years (median age 59 years). HS1 protein expression was determined by western blot. Comparative semi-quantitative indication of the degree of saturation of the bands analyzed by densitometry using the gel documentation system Gel-Doc (Bio-Rad) and a computer program to analyze the 1-D Quantity One (Bio-Rad). Assuming conventional units [AU - arbitrary units], depending on the saturation band, patients were divided into four groups with the expression of HS1 protein expressed in value from 0 to 3. Lack of expression was expressed as 0 [AU], and expression of the strongest, with the highest saturation band measured as 3 [AU]. Mutational status of IgVH, as well as CD38 and ZAP70 expression were also analyzed. Results: HS1 expression was significantly higher in CLL patients comparing to controls. Positive correlation was shown between HS1 and: age (p=0.0454), Rai stage (p=0.0412), leukocytosis (p=0.0129) and β2-microglobulin (p=0.0342). There was negative correlation between HS1 and hemoglobin level (p=0.0464) and platelet count (p=0.0310). Patients with lymphocyte doubling time shorter or equal to 6 months had higher expression of HS1. Expression of HS1 significantly influenced survival of CLL patients. Patients with higher HS1 expression had shorter survival than those with lower HS1 expression (p=0.0329). Conclusions: 1. Higher HS1 expression is observed in more advanced CLL stages. 2. Expression of HS1 in CLL cells is matched with shorter patient survival The relationship between expression of HS1 and survival of patients with B-CLL. Disclosures: No relevant conflicts of interest to declare.

Leukotriene B4 plays a pivotal role in CD40-dependent activation of chronic B lymphocytic leukemia cells

Blood ◽  
2005 ◽  
Vol 105 (3) ◽  
pp. 1274-1279 ◽  
Author(s):  
Gudmundur Runarsson ◽  
Anquan Liu ◽  
Yilmaz Mahshid ◽  
Stina Feltenmark ◽  
Annika Pettersson ◽  
...  
Keyword(s):  
Dna Synthesis ◽  
B Lymphocytes ◽  
Antigen Expression ◽  
Leukotriene B4 ◽  
Lymphocytic Leukemia ◽  
Flow Cytometry Analysis ◽  
Lipoxygenase Inhibitor ◽  
Biosynthesis Inhibitors ◽  
High Affinity Receptor ◽  
Affinity Receptor

AbstractBiosynthesis of leukotrienes (LTs) occurs in human myeloid cells and B lymphocytes. However, the function of leukotrienes in B lymphocytes is unclear. Here, we report that B-cell chronic lymphocytic leukemia (B-CLL) cells produce leukotriene B4, and that specific leukotriene biosynthesis inhibitors counteracted CD40-dependent activation of B-CLL cells. Studies on the expression of the high-affinity receptor for LTB4 (BLT1) by flow cytometry analysis showed that the receptor was expressed, to a varying degree, in all investigated B-CLL clones. At a concentration of 100 nM, the drugs BWA4C (a specific 5-lipoxygenase inhibitor) and MK-886 (a specific 5-lipoxygenase activating protein inhibitor) markedly inhibited CD40-induced DNA synthesis (45% and 38%, respectively) and CD40-induced expression of CD23, CD54, and CD150. Addition of exogenous LTB4 (150 nM) almost completely reversed the effect of the inhibitors on DNA synthesis and antigen expression. Taken together, the results of the present study suggest that leukotriene biosynthesis inhibitors may have a therapeutic role in B-CLL.

Leukotriene B4 Plays a Pivotal Role in CD40 Dependent Activation of Chronic B Lymphocytic Leukemia Cells.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4808-4808
Author(s):  
Hans-Erik Claesson ◽  
Gudmundur Runarsson ◽  
Anquan Liu ◽  
Yilmaz Mahshid ◽  
Stina Feltenmark ◽  
...  
Keyword(s):  
Dna Synthesis ◽  
B Lymphocytes ◽  
Antigen Expression ◽  
Leukotriene B4 ◽  
Lymphocytic Leukemia ◽  
Flow Cytometry Analysis ◽  
Lipoxygenase Inhibitor ◽  
Biosynthesis Inhibitors ◽  
High Affinity Receptor ◽  
Affinity Receptor

Abstract Biosynthesis of leukotrienes occurs in human myeloid cells and B lymphocytes. However, the function of leukotrienes in B lymphocytes is unclear. Here we report that B-cell chronic lymphocytic leukemia (B-CLL) cells produce leukotriene (LT) B4 and that specific leukotriene biosynthesis inhibitors counteracted CD40-dependent activation of B-CLL cells. Studies on the expression of the high affinity receptor for LTB4 (BLT1) by flow cytometry analysis showed that the receptor was expressed, to a varying degree, in all investigated B-CLL clones. The drugs BWA4C (a specific 5-lipoxygenase inhibitor) and MK-886 (a specific 5-lipoxygenase activating protein inhibitor), at a concentration of 100 nM, markedly inhibited CD40-induced DNA synthesis (45% and 38%, respectively) and CD40-induced expression of CD23, CD54 and CD150. Addition of exogenous LTB4 (150 nM) almost completely reversed the effect of the inhibitors on DNA synthesis and antigen expression. Taken together, the results of the present study suggest that leukotriene biosynthesis inhibitors may have a therapeutic role in B-CLL.

Overexpression of the CXCR5 Chemokine Receptor, and Its Ligand, B Cell-Activating Chemokine-1 (BCA-1/CXCL13) in B Cell Chronic Lymphocytic Leukemia.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2948-2948
Author(s):  
Andrea Bürkle ◽  
Jan A. Burger
Keyword(s):  
Dendritic Cells ◽  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Healthy Volunteers ◽  
B Lymphocytes ◽  
Surface Expression ◽  
Lymphocytic Leukemia ◽  
Lymphoid Tissues ◽  
Follicular Dendritic Cells

Abstract The chemokine B cell-activating chemokine-1 (BCA-1/CXCL13) is an important homing factor for lymphocytes to B cell zones of secondary lymphoid tissues. CXCL13 acts through its cognate receptor, CXCR5. Normal, mature B cells and a subset of memory T cells express CXCR5 chemokine receptors and migrate in response to BCA-1. However, BCA-1 displays a preferential chemotactic activity for B1 B cells when compared to “normal” B2 B cells. Because B lymphocytes from patients with Chronic Lymphocytic Leukemia (B-CLL) are in several aspects comparable to murine B1 cells, we hypothesized that the CXCR5-CXCL13 axis may be highly active in CLL. Initially, we noticed that CLL cells express functional CXCR5 receptors that induce actin polymerization, CXCR5 endocytosis, chemotaxis, and a prolonged activation of p44/42 MAP kinases. In addition, we examined CXCR5 surface expression in a series of CLL patients by flow cytometry and compared the results with normal B cells, or other leukemic B cell lymphoma. In CLL, leukemia B cells expressed significantly higher surface expression of CXCR5 (mean fluorescence intensity ratio/MFIR: 121 ± 9 (±SEM), n = 26) than circulating, CD19 positive B cells from healthy volunteers (CXCR5-MFIR: 69.9 ± 5.4, n = 11, p = 0.002). Neoplastic B cells from other leukemic B cell lymphomas displayed low surface CXCR5 expression (MFIR 19.7 ± 5.9, n = 11). Serum levels of CXCL13 were evaluated by ELISA. Sera from CLL patients displayed significantly higher levels of CXCL13 (mean ± SEM: 170.1 ± 21.5 pg/ml, n = 22) when compared to sera from healthy volunteers (mean ± SEM: 70.7 ± 5.2 pg/ml, n = 10, p = 0.004). Follicular dendritic cells (FDC) have been considered the main source of CXCL13 in secondary lymphoid tissues, thereby attracting T and B lymphocytes for cognate interactions. Surprisingly, we did not detect significant levels of CXCL13 in supernatants of HK follicular dendritic cells, that previously were demonstrated to protect CLL cells from apoptosis (Pedersen &Reed, Blood.2002;100:1795–801). In contrast, high levels of CXCL13 were detected in supernatants of CLL cell cultures in the presence of nurselike cells (NLC). In NLC cultures, CXCL13 levels were 610 ± 129.8 pg/ml (mean ± SEM, n = 4), whereas FDC supernatants contained 0.22 ± 0 pg/ml CXCL13 (mean ± SEM, n = 2). Because of these high CXCL13 levels in NLC cultures, we examined CXCR5 downregulation on CLL B cells in NLC co-cultures. When compared to freshly isolated CLL B cells, CLL cells from NLC cultures express significantly lower surface CXCR5. CXCR5 MFIR of CLL cells from NLC co-cultures was 7 ± 0.9, n = 4, compared to a CXCR5 MFIR of 91.6 ± 12 for freshly isolated CLL cells the same patients (mean ± SEM, n = 4, p = 0.000). These data indicate that high levels of bioactive CXCL13 are released in NLC cultures that stimulate cognate CXCR5 receptors on CLL B cells and induce signaling cascades, such as p44/42 MAPK, that induce prolonged survival. As such, this study provides a novel insight into interactions between CLL cells and their microenvironment within lymphoid tissues.

scholarly journals Palmitoylation of Cytoskeleton Associated Protein 4 by DHHC2 Regulates Antiproliferative Factor-mediated Signaling

2009 ◽  
Vol 20 (5) ◽  
pp. 1454-1463 ◽  
Author(s):  
Sonia L. Planey ◽  
Susan K. Keay ◽  
Chen-Ou Zhang ◽  
David A. Zacharias
Keyword(s):  
Epithelial Cells ◽  
Cellular Proliferation ◽  
Phosphorylated Protein ◽  
Normal Bladder ◽  
High Affinity Receptor ◽  
Palmitoyl Acyltransferase ◽  
Cellular Behaviors ◽  
Affinity Receptor ◽  
Induced Changes

Previously, we identified cytoskeleton-associated protein 4 (CKAP4) as a major substrate of the palmitoyl acyltransferase, DHHC2, using a novel proteomic method called palmitoyl-cysteine identification, capture and analysis (PICA). CKAP4 is a reversibly palmitoylated and phosphorylated protein that links the ER to the cytoskeleton. It is also a high-affinity receptor for antiproliferative factor (APF), a small sialoglycopeptide secreted from bladder epithelial cells of patients with interstitial cystitis (IC). The role of DHHC2-mediated palmitoylation of CKAP4 in the antiproliferative response of HeLa and normal bladder epithelial cells to APF was investigated. Our data show that siRNA-mediated knockdown of DHHC2 and consequent suppression of CKAP4 palmitoylation inhibited the ability of APF to regulate cellular proliferation and blocked APF-induced changes in the expression of E-cadherin, vimentin, and ZO-1 (genes known to play a role in cellular proliferation and tumorigenesis). Immunocytochemistry revealed that CKAP4 palmitoylation by DHHC2 is required for its trafficking from the ER to the plasma membrane and for its nuclear localization. These data suggest an important role for DHHC2-mediated palmitoylation of CKAP4 in IC and in opposing cancer-related cellular behaviors and support the idea that DHHC2 is a tumor suppressor.

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