Role of CCL5 and Interleukin-6 in the Biology of Waldenström Macroglobulinemia.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 688-688
Author(s):  
Sherine F. Elsawa ◽  
Anne J. Novak ◽  
Steven C. Ziesmer ◽  
Thomas E. Witzig ◽  
Steven P. Treon ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Line ◽  
Stromal Cells ◽  
Gene Array ◽  
Serum Levels ◽  
Mitogen Activated Protein Kinase ◽  
Dependent Manner ◽  
Malignant Cells ◽  
Monoclonal Protein

Abstract Waldenström macroglobulinemia (WM) is a B-cell malignancy that is characterized by the production of a monoclonal IgM protein, a lymphoplasmacytic infiltrate in the bone marrow, and associated symptoms including anemia, lymphadenopathy and hyperviscosity. The aberrant production of a monoclonal IgM in the serum is a major factor causing significant morbidity in patients with this disease, yet little is known about the mechanisms that regulate monoclonal protein synthesis. While recent gene array studies and serum analysis have shown that IL-6 is elevated in WM patients suggesting an important role for this cytokine in this disease, the precise role played by IL-6 in WM is unknown. Using a multiplex ELISA approach to screen sera from WM patients, we confirmed that IL-6 was significantly elevated (p<0.0019) in patients (n=20) compared to controls (n=20). Serum levels of IL-6 in WM patients correlated with elevated levels of β2-microglobulin (p<0.0019). Additionally, we also found that serum levels of CCL5 (Rantes) were significantly elevated in WM patients (p<0.0001). CCL5 has been shown to regulate IL-6 secretion, and we therefore wanted to determine if CCL5 influenced IL-6 expression in WM and what the subsequent consequence of IL-6 stimulation was on WM cells. To define the source of IL-6 in the tumor microenvironment, we used stromal cells from the bone marrow of healthy donors, malignant cells from patients with WM, and the BCWM.1 WM cell line, and tested their ability to secrete IL-6 by ELISA. All cell types secreted IL-6, with stromal cells secreting the most. We then tested the ability of CCL5 to induce IL-6 secretion by WM and stromal cells. CCL5 significantly increased IL-6 secretion by stromal cells (p<0.03) and also increased IL-6 secretion by fresh CD19+ CD138+ cells from WM patients (p<0.02). Using fresh patient WM cells and the BCWM.1 WM cell line as a model, we then determined the effect of IL-6 on growth of WM cells. We found that IL-6 had a modest effect (mean=20% increase, range=5–41%) on cell proliferation (p<0.0039) but had no effect on cell viability. In contrast, when we addressed the role of IL-6 on IgM secretion, we found that IL-6 increased IgM secretion by BCWM.1 cells in a dose-dependent manner. The IL-6 mediated increase in IgM secretion was abolished in the presence of neutralizing antibodies to IL-6. When we analyzed the downstream signaling events activated by IL-6 in WM cells we found that stimulation of BCWM.1 cells, which express the IL-6R, resulted in phosphorylation of Stat1, Stat3 and Erk1/2, but not Akt. Using a mitogen activated protein kinase (MAPK) inhibitor, we could inhibit the IL-6-mediated phosphorylation of Erk1/2. Similarly, using a JAK1 Inhibitor, we could inhibit IL-6 mediated signaling through Stat1 and Stat3. In summary, we have clearly shown that IL-6 significantly upregulates IgM secretion by WM cells and increases their proliferation. We have also demonstrated the ability of both the malignant cells and the stromal cells to secrete IL-6, and that this secretion is regulated in part by CCL5. We have found that WM cells express IL-6R, and that IL-6 induced signaling is through both the MAPK and Jak/Stat pathways. Therapies targeting IL-6 secretion or the IL-6 signaling pathways may therefore provide clinical benefit to patients with WM; not just by inhibiting the malignant cells but by down regulating the production of the monoclonal protein.

scholarly journals The Key Role of PML in IFN-α Induced Cellular Senescence of Human Mesenchymal Stromal Cells

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 3611-3611
Author(s):  
Shan Fu ◽  
Jieping Wei ◽  
Binsheng Wang ◽  
He Huang
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Real Time ◽  
Mesenchymal Stromal Cells ◽  
Cellular Senescence ◽  
Stromal Cells ◽  
Real Time Pcr ◽  
Dependent Manner ◽  
P53 Pathway

Abstract As immunomodulatory cytokines, Type 1 interferons (IFNs) have a long history of efficacy in treating chronic myeloid leukemia (CML). Recently, many research reported the combination of IFN-α and imatinib significantly increased the rates of molecular responses, comparing to single imatinib treatment. Related molecular mechanism may be the direct effect of IFN-alpha on stem cells. Therefore, IFN-α was renewed to be a vital candidate for CML treatment. Bone marrow mesenchymal stem cells (MSCs), which also be defined as mesenchymal stromal cells, are important to hematopoiesis. IFN-α was indicated as a potential inhibitor of MSCs; however the exact mechanism remains unclear. PML is known as a tumor suppressor, which locates at the downstream of IFN-α pathway. In our previous research, we have proved that PML stably expressed in human MSCs (hMSCs), which was important in maintaining the normal function of hMSCs. To our knowledge, although PML has been extensively studied in tumor cells, little is known about PML gene regulation in MSCs. In this study, we investigated the effect of IFN-α on hMSCs and the role of PML involved in this process. After approval by institutional review board, hMSCs were isolated from the bone marrow of volunteers and confirmed by flow cytometry. Cells were treated with different concentration of IFN-α up to 14 days. We found that IFN-α treated cells were growing slowly, and had a dramatically decreased number of colone in a dose dependent manner (Fig A). However, IFN-α did not induce significant cell apoptosis. Then a variety of senescence-associated detection was measured. hMSCs senescence induced by IFN-α had a dose and time dependent manner (Fig B). After treated with IFN-α at 1000 U/ml for 7 or 14 days, we found that up to 18% ± 1.1 or 27.56% ± 1.33 of hMSCs became SA-b-gal-positive as compared with 7.53% ± 0.55 or 6.47% ± 2.5 of untreated cells (P<0.05 for both). Real time PCR analysis proved this process by an increase in production of the senescence marker p53 and p21. Expression of PML was detected by real-time PCR and immunofluorescence in hMSCs treated with IFN-α. Consistent with other studies, mRNA expression of PML can be up-regulated by IFN-α in hMSCs. When cells were treated with IFN-α at 1000 U/ml for 7 or 14 days, PML gene expression in hMSCs was increased by more than 2 fold. At the same time, both the number and size of PML-NBs were increased markedly and had a concentration dependent manner. These results indicate that PML protein can be up regulated by IFN-α in hMSCs. Then, PML expression was inhibited using an RNAi-mediated PML knockdown system. After treated with IFN-α at 1000 U/ml for 7 days, hMSCs senescence can be rescued by the knocking down of PML. The percentage of SA-b-gal positive cells in PML knocking down hMSCs has a significant decrease as compared with cell transfected with control-RNAi (4.49% ±1.27 vs. 17.26% ± 1.44, P < 0.05) (Fig C). To further characterize the effect of PML on cellular senescence in hMSCs, PML-overexpressed hMSCs were used. 7 days post-transfection, PML overexpressing hMSCs were strongly positive for SA-b-gal activity (47.43%±3.8), as compared with normal and empty vector transfected cells (4.9%±0.7, 5.97%±0.75) (P< 0.001) (Fig D). mRNA levels of P53 and P21 were also enhanced in PML-overexpressed hMSCs. P53 pathway contributes to cell senescence and the role of PML has been proved in the regulation of P53 activity, we wondered whether upregulation of PML induced by IFN-α has relationship with P53 pathway in hMSCs. In the process of IFN-α induced hMSCs senescence, an increasing co-localization of PML and P53 was observed in IFN-α treated cells (1000U/ml, 7 days) as compared with untreated cells (Fig E). To further confirm whether or not the change of P53 location was mediated through the upregulation of PML, we knocked-down the expression of PML in hMSCs. Treated with IFN-α (1000U/ml, 7 days), we did not found significant location of P53 in PML-knocking down cells as compared with control. Taken together, our results suggested that hMSCs incurred senescence upon IFN-α stimulation, while PML levels were observed significant increase. By knocking-down and overexpressing PML, we demonstrated that PML was indispensable to IFN-α mediated hMSCs senescence. The molecular mechanisms underlying this process may be an increased co-localization of PML and p53 induced by IFN-α. These findings provided a novel insight into the role of IFN-α on hMSCs. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

scholarly journals Hemonectin mediates adhesion of engrafted murine progenitors to a clonal bone marrow stromal cell line from Sl/Sld mice

Blood ◽  
1991 ◽  
Vol 77 (8) ◽  
pp. 1691-1698 ◽  
Author(s):  
P Anklesaria ◽  
JS Greenberger ◽  
TJ Fitzgerald ◽  
B Sullenbarger ◽  
M Wicha ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Line ◽  
Cell Lines ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Cumulative Number ◽  
Specific Staining ◽  
Marrow Stromal Cell

Abstract Mutant Sl/Sld mice exhibit decreased marrow hematopoiesis. The defect is known to reside in the marrow microenvironment of these animals, which is reproduced in vitro by primary marrow explants as well as by cloned marrow stromal cell lines. Bone marrow progenitor cells are incapable of adhering to primary Sl/Sld stromal cells or cloned stromal cell lines derived from them to form cobblestone-islands and proliferate. The role of hemonectin, a marrow-specific adhesion protein in the defective hematopoiesis of the Sl/Sld mice, was studied. Indirect immunoperoxidase staining of marrow in situ from Sl/Sld mice showed little specific staining while specific staining was seen in a pericellular distribution in marrow from +/+ mice. Hemonectin expression in several cloned stromal cell lines from Sl/Sld mice was compared by immunoblotting with that in cloned stromal cell lines from normal +/+ littermates. Cell line Sld3, which has the least hematopoiesis supportive capacity in vitro, showed no detectable hemonectin by immunoblotting, while Sld1 and Sld2 showed detectable but greatly reduced amounts compared with normal +/+ 2.4, GBI/6, and D2XRII. Confluent cultures incubated with purified hemonectin and engrafted with enriched progenitors showed a significant increase in the cumulative number of cobbleston-islands and day 14 spleen colony- forming units (CFU-s) forming progenitors (39.15 +/- 3.6/dish; 16.3 +/- 3.1/dish, respectively), compared with untreated Sld3 cultures (cobblestone-islands 8.1 +/- 3.6/dish; CFU-s forming progenitors 8.8 +/- 0.05/dish). Hemonectin-mediated progenitor cell binding to the Sld3 stromal cells was specifically inhibited by antihemonectin but not by preimmune serum. These data support the role of hemonectin in early progenitor-stromal cell interactions.

scholarly journals Differential roles of stromal cells, interleukin-7, and kit-ligand in the regulation of B lymphopoiesis

Blood ◽  
1992 ◽  
Vol 79 (5) ◽  
pp. 1185-1192 ◽  
Author(s):  
LG Billips ◽  
D Petitte ◽  
K Dorshkind ◽  
R Narayanan ◽  
CP Chiu ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cell ◽  
Cell Line ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Kit Ligand ◽  
Interleukin 7 ◽  
Stromal Cell Line ◽  
Differentiation And Proliferation

Abstract Newly formed B lymphocytes are a population of rapidly renewed cells in the bone marrow of mammals and their steady state production presumably depends on a cascade of regulatory cells and cytokines. Although considerable information has been forthcoming about the role of interleukin-7 (IL-7) in potentiating pre-B-cell proliferation, few studies have addressed the possibility that multiple cytokines are involved in the progression of early events in cellular differentiation and proliferation in this hematopoietic lineage. Our laboratory previously described pre-B-cell differentiation mediated by the bone marrow stromal cell line S17. In this study, we further delineate the role of stromal cells in differentiation and proliferation of pre-B cells. These experiments show that the stromal cell line S17 potentiates the proliferative effect of IL-7 on B-lineage cells and that this S17-derived potentiator can be replaced with recombinant kit- ligand (KL). Our results further show that pre-B-cell formation from B220-, Ig- progenitor cells and expression of mu heavy chain of immunoglobulin is uniquely dependent on the presence of S17 stromal cells and cannot be reproduced with IL-7, KL, or costimulation with both IL-7 and KL. These data contribute to a rapidly evolving model of stromal cell regulation of B-cell production in the marrow and suggest unique roles for IL-7, KL, and as yet uncharacterized stromal cell- derived lymphokines in this process.

scholarly journals Differential roles of stromal cells, interleukin-7, and kit-ligand in the regulation of B lymphopoiesis

Blood ◽  
1992 ◽  
Vol 79 (5) ◽  
pp. 1185-1192 ◽  
Author(s):  
LG Billips ◽  
D Petitte ◽  
K Dorshkind ◽  
R Narayanan ◽  
CP Chiu ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cell ◽  
Cell Line ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Kit Ligand ◽  
Interleukin 7 ◽  
Stromal Cell Line ◽  
Differentiation And Proliferation

Newly formed B lymphocytes are a population of rapidly renewed cells in the bone marrow of mammals and their steady state production presumably depends on a cascade of regulatory cells and cytokines. Although considerable information has been forthcoming about the role of interleukin-7 (IL-7) in potentiating pre-B-cell proliferation, few studies have addressed the possibility that multiple cytokines are involved in the progression of early events in cellular differentiation and proliferation in this hematopoietic lineage. Our laboratory previously described pre-B-cell differentiation mediated by the bone marrow stromal cell line S17. In this study, we further delineate the role of stromal cells in differentiation and proliferation of pre-B cells. These experiments show that the stromal cell line S17 potentiates the proliferative effect of IL-7 on B-lineage cells and that this S17-derived potentiator can be replaced with recombinant kit- ligand (KL). Our results further show that pre-B-cell formation from B220-, Ig- progenitor cells and expression of mu heavy chain of immunoglobulin is uniquely dependent on the presence of S17 stromal cells and cannot be reproduced with IL-7, KL, or costimulation with both IL-7 and KL. These data contribute to a rapidly evolving model of stromal cell regulation of B-cell production in the marrow and suggest unique roles for IL-7, KL, and as yet uncharacterized stromal cell- derived lymphokines in this process.

scholarly journals Hemonectin mediates adhesion of engrafted murine progenitors to a clonal bone marrow stromal cell line from Sl/Sld mice

Blood ◽  
1991 ◽  
Vol 77 (8) ◽  
pp. 1691-1698
Author(s):  
P Anklesaria ◽  
JS Greenberger ◽  
TJ Fitzgerald ◽  
B Sullenbarger ◽  
M Wicha ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Line ◽  
Cell Lines ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Cumulative Number ◽  
Specific Staining ◽  
Marrow Stromal Cell

Mutant Sl/Sld mice exhibit decreased marrow hematopoiesis. The defect is known to reside in the marrow microenvironment of these animals, which is reproduced in vitro by primary marrow explants as well as by cloned marrow stromal cell lines. Bone marrow progenitor cells are incapable of adhering to primary Sl/Sld stromal cells or cloned stromal cell lines derived from them to form cobblestone-islands and proliferate. The role of hemonectin, a marrow-specific adhesion protein in the defective hematopoiesis of the Sl/Sld mice, was studied. Indirect immunoperoxidase staining of marrow in situ from Sl/Sld mice showed little specific staining while specific staining was seen in a pericellular distribution in marrow from +/+ mice. Hemonectin expression in several cloned stromal cell lines from Sl/Sld mice was compared by immunoblotting with that in cloned stromal cell lines from normal +/+ littermates. Cell line Sld3, which has the least hematopoiesis supportive capacity in vitro, showed no detectable hemonectin by immunoblotting, while Sld1 and Sld2 showed detectable but greatly reduced amounts compared with normal +/+ 2.4, GBI/6, and D2XRII. Confluent cultures incubated with purified hemonectin and engrafted with enriched progenitors showed a significant increase in the cumulative number of cobbleston-islands and day 14 spleen colony- forming units (CFU-s) forming progenitors (39.15 +/- 3.6/dish; 16.3 +/- 3.1/dish, respectively), compared with untreated Sld3 cultures (cobblestone-islands 8.1 +/- 3.6/dish; CFU-s forming progenitors 8.8 +/- 0.05/dish). Hemonectin-mediated progenitor cell binding to the Sld3 stromal cells was specifically inhibited by antihemonectin but not by preimmune serum. These data support the role of hemonectin in early progenitor-stromal cell interactions.

scholarly journals Leukocyte immunoglobulin-like receptor B4 deficiency exacerbates acute lung injury via NF-κB signaling in bone marrow-derived macrophages

2019 ◽  
Vol 39 (6) ◽  
Author(s):  
Tao Qiu ◽  
Jiangqiao Zhou ◽  
Tianyu Wang ◽  
Zhongbao Chen ◽  
Xiaoxiong Ma ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Lung Injury ◽  
Lung Injury ◽  
Stromal Cells ◽  
Marrow Cell ◽  
Marrow Transplant ◽  
Cell Source ◽  
Transplant Model ◽  
Acute Inflammatory Disease

AbstractAcute lung injury (ALI) is an acute inflammatory disease. Leukocyte immunoglobulin-like receptor B4 (LILRB4) is an immunoreceptor tyrosine-based inhibitory motif (ITIM)-bearing inhibitory receptor that is implicated in various pathological processes. However, the function of LILRB4 in ALI remains largely unknown. The aim of the present study was to explore the role of LILRB4 in ALI. LILRB4 knockout mice (LILRB4 KO) were used to construct a model of ALI. Bone marrow cell transplantation was used to identify the cell source of the LILRB4 deficiency-aggravated inflammatory response in ALI. The effect on ALI was analyzed by pathological and molecular analyses. Our results indicated that LILRB4 KO exacerbated ALI triggered by LPS. Additionally, LILRB4 deficiency can enhance lung inflammation. According to the results of our bone marrow transplant model, LILRB4 regulates the occurrence and development of ALI by bone marrow-derived macrophages (BMDMs) rather than by stromal cells in the lung. The observed inflammation was mainly due to BMDM-induced NF-κB signaling. In conclusion, our study demonstrates that LILRB4 deficiency plays a detrimental role in ALI-associated BMDM activation by prompting the NF-κB signal pathway.

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