The Involvement of NF-κB Subunits in the Interaction Between CLL-B Cells and Bone Marrow Stromal Cells

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4590-4590
Author(s):  
Aining Sun ◽  
Guo Feng ◽  
Jingjing Xu ◽  
Wenjuan Wang ◽  
Wu Depei
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
Mrna Expression ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Family Members ◽  
Proteasome Inhibitors ◽  
Marrow Stromal Cells ◽  
Protein Levels ◽  
Relative Mrna Expression

Abstract Abstract 4590 Objective: To investigate the function of NF-κB signaling pathway in the interaction between chronic lymphocytic leukemia B cells and bone marrow stromal cells. Method: The expression of NF-κB family members at mRNA and protein levels were examined by quantitative RT-PCR and Western bloting analyses individualy, and the differences between CLL-human bone marrow stromal cell (hBMSC) and non-CLL-hBMSC were investigated with the methods above. Cell death was measured by flow cytometry analysis after B-CLL cells were co-cultured with hBMSC and treated with proteasome inhibitors. The changes of NF-κB expression at the protein levels were examined by Western bloting after co-culture experiment. Result: The expression of NF-κB family members turned out to be heterogeneous at both mRNA and protein levels in B-CLL cells, and the members demonstrated a different κB-DNA binding activities. The mRNA expression of NF-κB family members in B-CLL cells was shown at a remarkable higher level than that of the controls. The relative mRNA expression of relA was 0.0214±0.012, whereas it was 0.0130±0.012 for the controls of CD19 positive cells. The relative mRNA expression of p50 and p52 were 66.0860±21.649 and 0.0208±0.011 respectively, and the corresponding expression of CD19 positive cells were 24.8440±9.749 and 0.0065±0.002. The differences were statistically significant. There was no notable difference between CLL-hBMSC and Non-CLL-hBMSC. hBMSC protected B-CLL cells against the proteasome inhibitors and facilitated the survival of B-CLL cells. Conclusion: NF-κB expression is heterogeneous in bone marrow B-CLL cells. There is no significant difference between CLL-hBMSC and non-CLL-hBMSC. hBMSC can protect the survival of B-CLL cells dependending on the endogenous NF-κB activity. hBMSC can increase the drug resistance of B-CLL cells to proteasome inhibitors. Disclosures: No relevant conflicts of interest to declare.

Crosstalk between Chronic Lymphocytic Leukemia (CLL) B-Cells and Marrow Stromal Cells: Implication for CLL B-Cell Activation and Survival.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 337-337
Author(s):  
Wei Ding ◽  
Grzegorz S. Nowakowski ◽  
Jennifer L. Abrahamzon ◽  
Linda E. Wellik ◽  
Asish K. Ghosh ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
Growth Factor ◽  
B Cell ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Marrow Stromal Cells ◽  
Drug Induced ◽  
Cd38 Expression ◽  
The Mean

Abstract It is believed that malignant cells “condition” the microenvironment to facilitate tumor cell survival. We hypothesized that crosstalk between CLL B-cells and marrow stromal cells impacts both cell types bi-directionally and ultimately contributes to leukemic cell apoptotic resistance. To test this hypotheses, bone marrow stromal cells from core bone biopsies from CLL patients were isolated and cultured using methods we have previously described (Leuk Res 2007 31(7):899). Subsequently, we determined the impact of co-culture on CLL B-cell features including apoptosis and CD38 expression. In addition, we evaluated the release of angiogenic cytokines on co-culture and signal events in the stromal cells. Immunophenotyping demonstrated that cultured bone biopsy derived stromal cells were CD73+, CD105+, CD146+, CD14−, CD45−, CD34−, HLA-DR-, suggesting they were mesenchymal stem cells (MSC). Co-culture of these MSC with CLL B-cells protected CLL B-cells from both spontaneous apoptosis (SA) and drug-induced (fludarabine and chlorambucil) apoptosis (DA). For SA, the mean survival of CLL B-cells with or without co-culture of MSC for 5 days were 56.9 ± 10.0 and 7.7 ±3.7 (p<0.05), respectively. When CLL B cells were treated with fludarabine or chlorambucil, the fraction of CLL cells tightly adherent to MSC (TA-CLL) showed higher survival than a less adherent but viable fraction of CLL B-cells. The mean survival of TA-CLL cells treated with 10 μM of fludarabine for 48 hours in the presence of MSC were 67.5 ± 3.6 vs 29.8 ± 11.1 without MSC (P<0.05), respectively. When CLL cells with evidence for CD38 expression were co-cultured with MSC, both the percentage of CD38 positive cells and level of expression of CD38 per cell were up-regulated (mean fold change: CD38 percentage, 2.7, p<0.05; CD38 MFI, 1.9, p<0.05) after 2 weeks. In contrast, the CD38 percentage and expression were not changed in cells with minimal CD38 expression when these CLL B-cells were co-cultured with MSC. In addition, co-culture of MSC with CLL cells induced rapid ERK and AKT phosphorylation (within 30 min) in the MSC on immunoblot analysis. When CLL B cells and MSCs were cultured in transwells, the activation of ERK and AKT in MSC occurred at similar levels, indicating that activation of MSC was mediated by soluble factors. In addition, co-culture led to increased secretion of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) as well as a decrease of thrombospondin-1 (TSP-1) in the culture medium. These findings confirm that co-culture of CLL B-cells and MSC culminates in “angiogenic switch.” Taken together, these results strongly suggest interactions between MSC and CLL B cells are a bi-directional process. In leukemic cells, the interaction not only protects against spontaneous and drug induced apoptosis but also leads to an increase in CD38 expression consistent with an activated status. In MSC, the interaction leads to activation of ERK and AKT. Co-culture also facilitates angiogenic switching. These results underscore the dynamic and complex nature of the interactions between bone marrow stromal cells and CLL B-cells. Further studies are needed to dissect how crosstalk between CLL B-cells and MSC relates to disease progression, and determines whether these interactions can be targeted with therapeutic intent.

scholarly journals Induction of Poly(ADP-ribose) Polymerase in Mouse Bone Marrow Stromal Cells Exposed to 900 MHz Radiofrequency Fields: Preliminary Observations

2016 ◽  
Vol 2016 ◽  
pp. 1-7 ◽  
Author(s):  
Qina He ◽  
Yulong Sun ◽  
Lin Zong ◽  
Jian Tong ◽  
Yi Cao
Keyword(s):  
Bone Marrow ◽  
Ionizing Radiation ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Messenger Rna ◽  
Positive Control ◽  
Marrow Stromal Cells ◽  
Mouse Bone Marrow ◽  
Protein Levels ◽  
Parp 1

Background. Several investigators have reported increased levels of poly(ADP-ribose) polymerase-1 (PARP-1), a nuclear enzyme which plays an important role in the repair of damaged DNA, in cells exposed to extremely low dose ionizing radiation which does not cause measurable DNA damage.Objective. To examine whether exposure of the cells to nonionizing radiofrequency fields (RF) is capable of increasing messenger RNA of PARP-1 and its protein levels in mouse bone marrow stromal cells (BMSCs).Methods. BMSCs were exposed to 900 MHz RF at 120 μW/cm2power intensity for 3 hours/day for 5 days. PARP-1 mRNA and its protein levels were examined at 0, 0.5, 1, 2, 4, 6, 8, and 10 hours after exposure using RT-PCR and Western blot analyses. Sham-exposed (SH) cells and those exposed to ionizing radiation were used as unexposed and positive control cells.Results. BMSCs exposed to RF showed significantly increased expression of PARP-1 mRNA and its protein levels after exposure to RF while such changes were not observed in SH-exposed cells.Conclusion. Nonionizing RF exposure is capable of inducing PARP-1.

scholarly journals CHIP regulates bone mass by targeting multiple TRAF family members in bone marrow stromal cells

Bone Research ◽  
2018 ◽  
Vol 6 (1) ◽  
Author(s):  
Tingyu Wang ◽  
Shan Li ◽  
Dan Yi ◽  
Guang-Qian Zhou ◽  
Zhijie Chang ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Bone Mass ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Family Members ◽  
Marrow Stromal Cells

scholarly journals Bone marrow stromal cells modulate both kappa light chain and Ly1 antigen expression on Ly1+ pre-B cell lines in vitro

Blood ◽  
1990 ◽  
Vol 76 (2) ◽  
pp. 383-392 ◽  
Author(s):  
MG Kruger ◽  
RL Riley ◽  
EA Riley ◽  
JM Elia
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Cell ◽  
Cell Lines ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Marrow Stromal Cells ◽  
Light Chains ◽  
Bone Marrow Cultures ◽  
Marrow Cultures

Abstract Murine Ly1+ pre-B cell lines, including 70Z/3 and three pre-B cell lines derived from long-term bone marrow cultures, exhibited selective adherence to bone marrow stromal cells. In contrast, splenic B cells, the A20 B-cell lymphoma, and four Ly1- B cell lines derived from long- term bone marrow cultures failed to adhere substiantially to bone marrow cultures failed to adhere substiantially to bone marrow stroma. Ly1+ pre-B cell lines were induced to express kappa light chains by exposure to either lipopolysaccharide (LPS), recombinant interleukin-1 (IL-1), or stromal cells. However, induction of kappa light chains failed to prevent pre-B cell adherence to stromal cells. Supernatants derived from primary bone marrow stromal cells decreased Ly1 expression on the Ly1+ pre-B cell lines. These experiments suggest that (1) expression of immunoglobulin light chains by developing Ly1+ pre-B cells is mediated by bone marrow stromal cells; (2) loss of specific adherence to stroma is progressive and occurs post-light chain induction; and (3) soluble products of stromal cells may downregulate expression of surface Ly1 on otherwise Ly1+ pre-B cells. The importance of these observations to the development of both the Ly1- and Ly1+ B cell lineages in the mouse is discussed.

Rooting into the Soil, a Model for the Role of Sox4 in Leukemia

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3794-3794
Author(s):  
Saradhi Mallampati ◽  
Baohua Sun ◽  
Yun Gong ◽  
Enze Wang ◽  
M. James You ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Cell ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Leukemic Cell ◽  
Marrow Stromal Cells ◽  
Leukemic Cells ◽  
Cell Stage

Abstract Development and progression of leukemia requires interaction of leukemia-initiating cells with their bone marrow niches. The niches serve as the nursery and shelter for the leukemic cells, which can result in drug resistance, disease recurrence, and minimal residual disease, the most important causes for the death of patients with leukemia. Therefore, obliteration of the interaction between the leukemic cells and their niches is of utmost importance in eradicating leukemic cells during therapy to cure the disease. However, little is currently known of the molecular mechanisms underlying the interaction of the two types of cells. Sox4, a SRY-related HMG-box containing transcription factor that is vital during development, plays an important role in leukemia. Published mouse studies demonstrated that increased expression of Sox4 was associated with leukemogenesis. We determined the expression levels of Sox4 by real-time RT-PCR in 100 human leukemic samples and found high levels of expression in B- and T-ALL, but not in AML, CML, CLL, Sezary syndrome, or T cell prolymphocytic leukemia. In accordance, 7 of the 8 ALL cell lines (the exception was 697) we tested showed high expression levels of Sox4, but AML cell lines, normal mature B cells, T cells, and bone marrow CD34+ cells had low levels of expression. Since the majority of clinical B-ALL cases correspond to the pre-B cell stage, we investigated the role of Sox4 in a pre-B cell line (Nalm6) by lentivirus-mediated RNAi. Remarkably, knockdown of Sox4 in Nalm6 cells caused 70% reduction in the formation of leukemic cell clusters under the monolayer of co-cultured M2-10B4 bone marrow stromal cells, a phenomenon known as pseudo-emperipolesis. Similar results were obtained with ex vivo cultured bone marrow cells from conditional Sox4 knockout mice that displayed B cell developmental arrest at the transition from pro-B to pre-B cell stage and an absence of pre-B cells. These findings suggested that Sox4 is required for the interaction of the developing B cells or leukemic cells with bone marrow stromal cells, a component of the bone marrow niche. Since CXCR4/SDF1-mediated “homing” is known to be required for pseudo-emperipolesis, we tested the effect of Sox4 on Nalm6 cell migration toward SDF1 gradient and found that Sox4 did not affect the migration, suggesting that Sox4 is not acting through “homing”. Instead, our data indicated that the role of Sox4 in the interaction of leukemic cells with stromal cells is most likely mediated by its ability in enhancing the adhesion of the leukemic cells because we found that lentivirus-medicated overexpression of Sox4 in the 697 B cell line caused the suspension cells to display a spindle and adhesive morphology. In addition, 21% of the putative Sox4 downstream genes that we identified by multiple sets of gene expression microarray experiments are known to be involved in cell adhesion. Moreover, we found that the changes in gene expression profile of leukemic cells upon Sox4 knockdown or overexpression significantly overlap with the changes in response to the presence of bone marrow stromal cells in co-culture, indicating that Sox4 pathways are involved in leukemic cell response to stromal cell signaling. Based on these findings we hypothesize that deletion of Sox4 abolishes the interaction between the developing lymphocytes and their niches during lymphopoiesis. Conversely, overexpression of Sox4 may enforce these cells to over-interact with the niches so that they are overexposed to local growth factor stimuli. If superimposed with other genetic and/or epigenetic changes in the developing lymphocytes, such over-interaction may result in the development of leukemia. In case of established leukemia, such over-interaction may lead to the enhanced protection of leukemic cells by their niches. Therefore, the role of Sox4 in the interaction of developing lymphocytes or leukemic cells with their niches is like “rooting into the soil” of a growing tree, abbreviated as “rooting”.

A Clinical Study of Bone Marrow Stromal Cell Therapy in Patients with Refractory Cgvhd by Down-Regulating the Jagged2 Gene

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 4673-4673
Author(s):  
Jianyu Weng ◽  
Xin Huang ◽  
Suxia Geng ◽  
Chengwei Luo ◽  
Suijing Wu ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mrna Expression ◽  
Stromal Cells ◽  
Peripheral Blood ◽  
Bone Marrow Stromal Cells ◽  
Marrow Stromal Cells ◽  
Control Group ◽  
Expression Levels ◽  
Mrna Expression Levels

Abstract Abstract 4673 Refractory chronic GVHD (cGVHD) is an important complication after allogeneic hematopoietic SCT and is prognostic of poor outcome. Bone Marrow Stromal Cells (MSCs) are involved in tissue repair and modulating immune responses in vitro and in vivo. MSCs as salvage treatment for refractory cGVHD have been reported in our previous study, however, the possible mechanism have yet not to be determined. Between November 2006 and November 2010, 18 patients were diagnosed with refractory cGVHD, 8 patients were treated with in vitro expanded BM-derived MSCs as a compassionate treatment for refractory cGVHD, 10 patients that did not receive BMSCs treatment were control group. The median MSC dose given was 0.6×106/kg body weight. MSCs were harvested fresh from culture and administered to the patients by intravenous infusions over 30 minutes. The median time of MSC administrations was 3 (range, 2–6). The response was assessed monthly after BMSCs treatment, and the total follow-up period was 6 months. The organ response and the overall response were used to determine the therapeutic efficacies of MSC for refractory cGVHD. The expression of the Jagged2 gene of peripheral blood mononuclear cells in patients at the assessment points were analyzed using the TaqMan real-time polymerase chain reaction, with ABL mRNA expression levels as an internal reference. After BMSCs treatment, a total of 6 patients (75%) had an overall response (PR n=6), and 2 patients had a minor partial response (mPR n=2). The expression levels of Jagged2 mRNA in these cases at the diagnosis of refractory cGVHD were significantly increased, compared with none cGVHD patients (23.94%±18.68% vs 3.76%±1.50%, P < 0.05), and the copies of Jagged2 mRNA in BMSC treatment responsed patients' peripheral blood were significantly reduced (5.15%±3.25%, P <0.05), while Jagged2 mRNA expression levels of the control group were no significant difference (P> 0.05). Our pilot study showed that Jagged2 gene reproduction upregulated when the cGVHD is active, so, dynamic monitoring of Jagged2 mRNA expression may have the potential effect on predicting the activity of chronic graft-versus-host disease. Mechanism of Bone marrow stromal cells to treat refractory cGVHD may be related to down-regulation of donor T cells Notch ligand Jagged2 gene expression, which suppression of T cell Notch signaling pathway activation, thus inducing immune tolerance. Disclosures: No relevant conflicts of interest to declare.

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