Stromal-derived Factor-1 Receptor Research Articles

MiRNA-126a plays important role in myoblast and endothelial cell interaction

Muscle satellite cells (SCs) are stem cells and the main players in skeletal muscle reconstruction. Since satellite cells are located near or in direct contact with blood vessels their niche is formed, inter alia, by endothelial cells. The cross-talk between satellite cells and endothelial cells determines quiescence or proliferation of these cells. However, little is known about the role of miRNA in these interactions. In the present study we identified miRNA that were up-regulated in SC-derived myoblasts treated with stromal derived factor-1 (SDF-1) and/or down-regulated in cells in which the expression of CXCR4 or CXCR7, that is, SDF-1 receptors, was silenced. SDF-1 is one of the important regulators of cell migration, mobilization, skeletal muscle regeneration, and angiogenesis. We hypothesized that selected miRNAs affect SC-derived myoblast fate and interactions with endothelial cells. We showed that miR-126a-3p inhibited both, myoblast migration and fusion. Moreover, the levels of Cxcl12, encoding SDF-1 and Ackr3, encoding CXCR7, were reduced by miR-126a-3p mimic. Interestingly, the miR-126a-3p mimic significantly decreased the level of numerous factors involved in myogenesis and the miR-126a-5p mimic increased the level of Vefga. Importantly, the treatment of endothelial cells with medium conditioned by miR-126-5p mimic transfected SC-derived myoblasts promoted tubulogenesis.

CXCR4 (CD184) expression on stem cell harvest and CD34+ cells post-transplant

Objectives/backgroundCXCR4 is a receptor for stromal-derived factor-1 (SDF-1), a molecule that has a chemotactic activity for lymphocytes and is important in homing of hematopoietic stem cells to their adult marrow. We evaluated the CXCR4 (CD184) expression in the harvest cells and in the post-transplant bone marrow (BM) and its relation to engraftment, as determined by the consensus criteria and chimerism. MethodsThis is a prospective study which included 30 patients undergoing hematopoietic stem cell transplantation; 15 patients received autograft and 15 patients received allograft on dates between January 2012 and May 2014. We assessed CD184 (CXCR4) using flow cytometry in the harvest cells together with post-transplant BM assessment on Day 28 and Day 90 for complete morphologic, molecular studies, and detection of CD184 expression on CD34+ cells with chimerism studies on total peripheral blood mononuclear cells. ResultsDiagnoses of the enrolled patients were as follows: seven (24.1%) with acute myeloid leukemia, eight (27.6%) with multiple myeloma, four (13.8%) with acute lymphoblastic leukemia, three (10.3%) with non-Hodgkin lymphoma, two (6.9%) with myelodysplastic syndromes, two (6.9%) with aplastic anemia, two (6.9%) with chronic myeloid leukemia, one (3.4%) with Hodgkin lymphoma, and one (3.4%) with plasmacytomas. One patient died and was excluded from the study because there were not enough data about engraftment. There was no statistical significance between the level of CD184 in stem cell harvest and the prediction of successful engraftment (p>0.05) as well as in Day 28 BM sample (p>0.05), whereas there was a statistical significance between the level of CD184 in Day 90 BM sample and the occurrence of successful engraftment (p=0.002). ConclusionSDF-1/CXCR4 axis plays a crucial role in engraftment; however, more studies are warranted to assess their expression post-transplant. Evaluating the ligand (chemokine, SDF-1) or its receptor (CXCR4) may serve as potential surrogate markers for assessment of engraftment.

A novel biphenyl urea derivate inhibits the invasion of breast cancer through the modulation of CXCR4.

The increased migration and invasion of breast carcinoma cells are key events in the development of metastasis to the lymph nodes and distant organs. CXCR4, the receptor for stromal-derived factor-1, is reportedly involved in breast carcinogenesis and invasion. In this study, we investigated a novel biphenyl urea derivate, TPD7 for its ability to affect CXCR4 expression as well as function in breast cancer cells. We demonstrated that TPD7 inhibited the breast cancer proliferation and down-regulated the CXCR4 expression on breast cancer cells both over-expressing and low-expressing HER2, an oncogene known to induce the chemokine receptor. Treatments with pharmacological proteasome inhibitors partial suppressed TPD7-induced decrease in CXCR4 expression. Real-time PCR analysis revealed that down-regulation of CXCR4 by TPD7 also occurred at the translational level. Inhibition of CXCR4 expression by TPD7 further correlated with the suppression of SDF-1α-induced migration and invasion in breast tumour cells, knockdown of CXCR4 attenuated TPD7-inhibitory effects. In addition, TPD7 treatment significantly suppressed matrix metalloproteinase (MMP)-2 and MMP-9 expression, the downstream targets of CXCR4, perhaps via inactivation of the ERK signaling pathway. Overall, our results showed that TPD7 exerted its anti-invasive effect through the down-regulation of CXCR4 expression and thus had the potential for the treatment of breast cancer.

Defining an optimal stromal derived factor-1 presentation for effective recruitment of mesenchymal stem cells in 3D.

In "situ" tissue engineering is a promising approach in regenerative medicine, envisaging to potentiate the physiological tissue repair processes by recruiting the host's own cellular progenitors at the lesion site by means of bioactive materials. Despite numerous works focused the attention in characterizing novel chemoattractant molecules, only few studied the optimal way to present signal in the microenvironment, in order to recruit cells more effectively. In this work, we have analyzed the effects of gradients of stromal derived factor-1 (SDF-1) on the migratory behavior of human mesenchymal stem cells (MSCs). We have characterized the expression of the chemokine-associated receptor, CXCR4, using cytofluorimetric and real-time PCR analyses. Gradients of SDF-1 were created in 3D collagen gels in a chemotaxis chamber. Migration parameters were evaluated using different chemoattractant concentrations. Our results show that cell motion is strongly affected by the spatio-temporal features of SDF-1 gradients. In particular, we demonstrated that the presence of SDF-1 not only influences cell motility but alters the cell state in terms of SDF-1 receptor expression and productions, thus modifying the way cells perceive the signal itself. Our observations highlight the importance of a correct stimulation of MSCs by means of SDF-1 in order to implement on effective cell recruitment. Our results could be useful for the creation of a "cell instructive material" that is capable to communicate with the cells and control and direct tissue regeneration. Biotechnol. Bioeng. 2014;111: 2303-2316. © 2014 Wiley Periodicals, Inc.

SDF-1/CXCR7 axis enhances ovarian cancer cell invasion by MMP-9 expression through p38 MAPK pathway.

Ovarian cancer is an aggressive gynecological malignancy with high metastatic potential. Recently, the CXC receptor (CXCR7) has been identified as a new receptor for stromal-derived factor-1 (SDF-1), and exerts important roles in cancer development. However, its effect on ovarian cancer and the underlying mechanism remain unknown. In this study, we detected abundant CXCR7 expression in ovarian cancer tissues and cells. Moreover, SDF-1 induced dramatically upregulation of CXCR7 mRNA and protein levels, indicating that the SDF-1/CXCR7 axis existed in ovarian cancer. Further analysis confirmed that SDF-1 enhanced cell adhesion and subsequent invasion, which were significantly attenuated when pretreated with CXCR7 small interference RNA (siRNA), indicating the critical function of SDF-1/CXCR7 in cell invasion. Further mechanistic analysis indicated that SDF-1/CXCR7 enhanced cell invasion by matrix metalloproteinase (MMP)-9, as pretreatment with MMP-9 siRNA significantly abrogated a number of invading cells. Additionally, SDF-1/CXCR7 induced phosphorylation of the p38 MAPK pathway, which was accounted for MMP-9 expression as preconditioning with the p38 MAPK inhibitor SB203580 obviously decreased MMP-9 expression. Together, our data implied that SDF-1/CXCR7 enhanced ovarian cancer cell invasion by MMP-9 expression through the p38 MAPK pathway. Thus, these findings confirmed the critical role of SDF-1/CXCR7 during the pathological processes of ovarian cancer and supported its potential targets for further development of antiovarian cancer therapy.

A Fine Line Between Healing and Fibrosis

The liver’s microvascular endothelium lies at the crux of regeneration and fibrosis.

Saxagliptin Improves Cardiac Function in the Post MI Diabetes Setting: Potential Role of Non GLP1 Mediated Cardioprotection Via Stromal Derived Factor 1 α

Di-peptidyl peptidase 4 (DPP-4) is a membrane spanning exopeptidase that cleaves dipeptides from the N terminus of proteins/peptides. Recent studies demonstrate circulating DPP-4 activity is increased in diabetes, which may in part explain the poor prognosis of diabetic patients with ischemic heart disease. In addition to degrading glucagon-like peptide-1 (GLP-1), DPP4 is also known to degrade stromal derived factor-1 (SDF-1), a pro-angiogenic and cardiomyocyte protective protein currently in early phase clinical trials for patients with ischemic heart disease. We hypothesized that DPP-4 inhibition in the diabetic post myocardial infarction (MI) setting may confer a benefit as a result of enhanced SDF-1 availability rather than potentiating GLP-1. We therefore compared DPP-4 inhibition with GLP-1 agonism, and used the specific SDF-1 receptor (CXCR4) antagonist, AMD3100 to demonstrate SDF-1 specificity.

The CXCR4 Antagonist AMD3465 Regulates Oncogenic Signaling and Invasiveness In Vitro and Prevents Breast Cancer Growth and Metastasis In Vivo

CXCR4, the receptor for stromal-derived factor-1, is reportedly involved in breast carcinogenesis. However, the mechanisms through which CXCR4 contributes to breast cancer cell growth and metastases are poorly understood. In this study, we examined the putative in vitro and in vivo anti-cancer effects of the specific CXCR4 inhibitor AMD3465. Here, we report that AMD3465 triggers a reduction in breast cancer cell invasiveness in vitro, and promotes marked changes in oncogenic signaling proteins including a reduction in STAT3, JAK2, AKT, and CXCR4 phosphorylation and the reduced expression of GSK3 and cMYC. Using three breast cancer cell lines as murine syngeneic immunocompetent breast cancer models, we found that AMD3465 inhibited breast tumor formation and reduced tumor cell metastases to the lung and liver. Furthermore, treatment with AMD3465 significantly reduced the infiltration of myeloid CD11b positive cells at the aforementioned metastatic sites as well as the spleen implying this agent could regulate the formation of the tumor microenvironment and conceivably the premetastatic niche. In conclusion, our studies suggest that AMD3465 inhibits breast cancer growth and metastases by acting on tumor cells as well as immune cells that constitute the tumor microenvironment. This process appears to be regulated, at least in part, through the modulation of oncogenic signaling that includes the STAT3 pathway. Thus, CXCR4 could be a novel target for breast cancer therapy.

Genetic Modification of Mesenchymal Stem Cells to Overexpress CXCR4 and CXCR7 Does Not Improve the Homing and Therapeutic Potentials of These Cells in Experimental Acute Kidney Injury

The therapeutic potential of bone marrow mesenchymal stem cells (MSCs) in kidney failure has been examined in some studies. However, recent findings indicate that after transplantation, these cells home to kidneys at very low levels. Interaction of stromal derived factor-1 (SDF-1) with its receptor, CXCR4, is of pivotal importance in migration and homing. Recently, CXCR7 has also been recognized as another SDF-1 receptor that interacts with CXCR4 and modulates its functions. In this study, CXCR4 and CXCR7 were separately and simultaneously overexpressed in BALB/c bone marrow MSCs by using a lentiviral vector system and the homing and renoprotective potentials of these cells were evaluated in a mouse model of cisplatin-induced acute kidney injury. Using flow cytometry, immunohistochemistry, and real-time PCR methods for detection of GFP-labeled MSCs, we found that although considerably entrapped in lungs, native MSCs home very rarely to kidneys and bone marrow and this rate cannot be significantly affected by CXCR4 and/or CXCR7 upregulation. Transplantation of neither native nor genetically engineered MSCs ameliorated kidney failure. We concluded that overexpression of CXCR4 and CXCR7 receptors in murine MSCs cannot improve the homing and therapeutic potentials of these cells and it can be due to severe chromosomal abnormalities that these cells bear during ex vivo expansion.

A novel perspective on stem cell homing and mobilization: review on bioactive lipids as potent chemoattractants and cationic peptides as underappreciated modulators of responsiveness to SDF-1 gradients

Hematopoietic stem progenitor cells (HSPCs) respond robustly to α-chemokine stromal-derived factor-1 (SDF-1) gradients, and blockage of CXCR4, a seven-transmembrane-spanning G(αI)-protein-coupled SDF-1 receptor, mobilizes HSPCs into peripheral blood. Although the SDF-1-CXCR4 axis has an unquestionably important role in the retention of HSPCs in bone marrow (BM), new evidence shows that, in addition to SDF-1, the migration of HSPCs is directed by gradients of the bioactive lipids sphingosine-1 phosphate and ceramide-1 phosphate. Furthermore, the SDF-1 gradient may be positively primed/modulated by cationic peptides (C3a anaphylatoxin and cathelicidin) and, as previously demonstrated, HSPCs respond robustly even to very low SDF-1 gradients in the presence of priming factors. In this review, we discuss the role of bioactive lipids in stem cell trafficking and the consequences of HSPC priming by cationic peptides. Together, these phenomena support a picture in which the SDF-1-CXCR4 axis modulates homing, BM retention and mobilization of HSPCs in a more complex way than previously envisioned.

700 A novel mechanism of action of transmyocardial revascularization: Induction of mesenchymal stem cell transmigration and engraftment via upregulation of stem cell factor and stromal derived factor-1

Transmyocardial revascularization (TMR) prior to mesenchymal stem cell (MSC) transplantation augments angiogenesis and functional recovery of infarcted hearts through unknown mechanisms. One potential mechanism could be via signaling to circulating progenitor cells. We evaluated the effect of TMR on homing and engraftment of circulating MSC and potential mediators of this effect. Female rats underwent LAD ligation to create a myocardial infarct. After 3 weeks, 10-channel needle TMR was performed in the infarct zone, followed by daily IV injections of 1 million GFP-positive male MSC for 5 days. Control rats (Ctrl) had MSC infusions without TMR (N = 13/group). Three days and 1 week later, rats were sacrificed and hearts were evaluated for male donor MSC survival by quantitative PCR for sry and GFP in the scar, border zone and normal myocardium, and expression of stem cell factor (SCF), stromal derived factor-1 (SDF-1) and their receptors c-kit and CXCR4. Vascular densities were evaluated by quantitative histology and LV function by echocardiography. Male MSC engraftment was greater in TMR hearts at 3 days (scar: TMR 520 ± 153, Ctrl 126 ± 72; border zone TMR 658 ± 192, Ctrl 325 ± 117 cells/mg tissue, P < 0.05) and 1 week (scar: TMR 411 ± 62, Ctrl 302 ± 91; border zone TMR 511 ± 104, Ctrl 365 ± 96 cells/mg tissue, P < 0.05). SCF was upregulated by TMR at 3 days (TMR 5.7 ± 2.2x109, Ctrl 2.7 ± 1.8x109 copies/ug RNA, P < 0.05) but levels fell at 1 week and were similar between groups (P = NS). In contrast, SDF-1 levels were higher in TMR hearts at both 3 days and 1 week, with a continuing rise over this interval (P < 0.05). C-kit levels did not differ between groups or timepoints. CXCR4 was upregulated early by TMR (3 days: TMR 9.4 ± 3.2x1010, Ctrl 3.1 ± 5.1x1010 copies/ug RNA, P < 0.05) but levels dropped dramatically (1 week: TMR 6.5 ± 3.5x109, Ctrl 2.4 ± 2.2x109 copies/ug RNA, P < 0.05). TMR induces transmigration and engraftment of circulating MSCs. Both SCF and SDF-1 are upregulated by TMR, but uptranscription of SCF is rapid and corresponds temporally to MSC engraftment, while SDF-1 levels rise more slowly. CXCR4, the SDF-1 receptor, is also upregulated, while expression of the SCF receptor c-kit is unaffected. TMR-augmented repair of infarcted hearts by stem cell transplantation may be mediated by a novel mechanism: transmigration and engraftment of circulating progenitor cells. The effect of TMR on angina in humans may be due in part to similar recruitment of endogenous cell-mediated repair mechanisms.

Expression of CXCR4 and CXCR7 Receptors for Stromal Derived Factor-1 (SDF- 1) Is Differently Regulated in Human Rhabdomyosarcomas (RMS): The Biological Consequences

Rhabdomyosarcoma (RMS) is the most common soft-tissue sarcoma of adolescents and children that frequently infiltrates bone marrow (BM). RMS is divided in two sub-types, alveolar-RMS (ARMS) and embryonal-RMS (ERMS). The ARMS subtype is often associated with a more aggressive phenotype, poorer clinical prognosis, and expression of Pax3-FKHR and Pax7-FKHR fusion proteins that function as potent transcriptional activators with enhanced activity as compared to normal Pax3 or Pax7 transcription factors. We reported that the stromal derived factor-1 (SDF-1)-CXCR4 receptor axis plays a crucial role in RMS metastasis to BM (Blood 2002; 100:2597, Cancer Res 2003; 63:7926, Cancer Res. 2007; 67:213). After the recent identification of CXCR7, a new receptor for SDF-1, we became interested in its potential role in metastases of RMS cells. We found that CXCR7 was expressed and functional on all 10 human RMS cell lines investigated in this study. Furthermore, we noticed that while CXCR4 was more involved in regulating RMS chemotaxis, CXCR7 primarily regulated adhesion of RMS cells. Based on this observation, we assessed the mechanisms that regulate expression of these receptors. We noticed that CXCR4 is highly expressed on the more metastatic ARMS while CXCR7 is expressed at higher levels on the less metastatic ERMS cell lines. However, both receptors express several hypoxia-inducible factor (HIF)-1a binding sites, expression of CXCR4 was not affected by exposure to hypoxia, and expression of CXCR7 was even paradoxically downregulated in hypoxic conditions. We also observed that ERMS cells transduced with Pax3-FKHR, a gene typical for ARMS fusion that enhances RMS metastatic behavior, highly upregulated expression of CXCR4. In contrast, expression of CXCR7 was again down-regulated. To learn more on the role of Pax-3-FKHR in the expression of CXCR4, we performed a functional analysis of promoter fragments subcloned into luciferase vector. To our surprise, expression of CXCR4 did not depend on direct binding of Pax3-FKHR to classical Pax3 binding sites in promoter sequences but on direct protein-to-protein interaction with nuclear respiratory factor (NRF)-1 transcription factor. These results suggest that the Pax3-FKHR-NRF-1 complex binds to the NRF-1 binding site and aberrantly activates transcription. In conclusion, both of the SDF-1 binding receptors (CXCR4 and CXCR7) are differently regulated in RMS cells. Since CXCR7 is more important in adhesion and becomes downregulated during hypoxia, this may give an advantage for SDF-1 to signal through CXCR4 receptor to activate pathways critical for chemotactic responses. As a biological consequence, this may increase the metastatic potential of RMS by “mobilizing” RMS cells from the primary tumor to migrate and metastasize. In addition, the novel transcriptional mechanism for the Pax3-FKHR-NRF-1 complex in regulating CXCR4 receptors in RMS cells has been identified.

Plasmatic Levels of FLT-3 Ligand and SDF-1 and Expression of FLT-3 and CXCR4 in CD34+ Cells Population Prior and Post Mobilization of Peripheral Blood Stem Cell.

Introduction: Peripheral blood stem cells (PBSC) became the main source of cells for autologous and allogeneic marrow transplantation. FLT3L is capable to inducing multi-lineage hematopoietic cell differentiation in vivo and in vitro. It was postulated that an increase in FLT-3 levels correlated with a compensatory mechanism to boost the amount of PBSC. The stromal-derived- factor-1 (SDF-1) is a member of the family of α-chemokines that binds to the seven-span transmembrane G-protein-couple CXCR4. Mobilized human CD34+ progenitors cells express reduced levels of the SDF-1 receptor of CXCR4, which correlates with improved mobilization, suggesting the involvement of SDF-1/CXCR4 interactions in the trafficking of CD34+ cells from marrow to peripheral blood. Objective: To determine the behavior of the of FLT-3-L and SDF-1 plasma concentration, prior and post mobilization of PBSC. The cellular expression of the FLT-3 and CXCR4 in the CD34+ cell population in the bone marrow (BM) prior and post mobilization and in the apheresis product and to evaluate its association with the good and bad yield of CD34+cells.Material and Methods: The plasma concentration of SDF-1 and FLT3-L was determined by ELISA, prior and after PBSC mobilization in 48 individuals, being 12 donors and 36 patients. The cellular expression of CXCR4 and FLT-3 in the CD34+ cell population, was estimated as the mean fluorescence intensity (MFI), in material of bone marrow (BM), prior and after mobilization and in the apheresis product. It was considered success of mobilization who had reached in the peripheral blood (PB), 15 or more CD34+cells and in the apheresis product ≥5x 106/Kg CD34+cells.Results: Considering globally the total of individuals, it was observed a plasma concentration variation of FLT3L and SDF-1, prior and post mobilization of PBSC (p=0.001 and 0,012, respectively), even this, did not have influenced the good yield of CD34+ cells. When evaluated the cellular expression of the CXCR4, it was observed variations in the BM, prior and post mobilization of PBSC, for the myeloma group (p<0.001), however when the total of individuals was considered, the behavior was different. There was a variation between the measures prior and post mobilization, only to the success mobilization group(p=0.036). For the FLT-3, we find variations between BM mobilized and not mobilized for all studied groups, however it did not have relation with the yield of CD34+ cells.Conclusion: Although, FLT-3L and SDF-1 prior and post mobilization showed variations in the plasmatic concentration in this study, the only variable that had association with the good yield of CD34+ cells was the cellular expression of the CXCR4.

The In Vitro Migration Capacity of Human Bone Marrow Mesenchymal Stem Cells: Comparison of Chemokine and Growth Factor Chemotactic Activities

Adult bone marrow (BM)-derived stem cells, including hematopoietic stem cells (HSCs) and MSCs, represent an important source of cells for the repair of a number of damaged tissues. In contrast to HSCs, the soluble factors able to induce MSC migration have not been extensively studied. In the present work, we compared the in vitro migration capacity of human BM-derived MSCs, preincubated or not with the inflammatory cytokines interleukin 1beta (IL1beta) and tumor necrosis factor alpha (TNFalpha), in response to 16 growth factors (GFs) and chemokines. We show that BM MSCs migrate in response to many chemotactic factors. The GFs platelet-derived growth factor-AB (PDGF-AB) and insulin-like growth factor 1 (IGF-1) are the most potent, whereas the chemokines RANTES, macrophage-derived chemokine (MDC), and stromal-derived factor-1 (SDF-1) have limited effect. Remarkably, preincubation with TNFalpha leads to increased MSC migration toward chemokines, whereas migration toward most GFs is unchanged. Consistent with these results, BM MSCs express the tyrosine kinase receptors PDGF-receptor (R) alpha, PDGF-Rbeta, and IGF-R, as well as the RANTES and MDC receptors CCR2, CCR3, and CCR4 and the SDF-1 receptor CXCR4. TNFalpha increases CCR2, CCR3, and CCR4 expression (as opposed to that of CXCR4), together with RANTES membrane binding. These data indicate that the migration capacity of BM MSCs is under the control of a large range of receptor tyrosine kinase GFs and CC and CXC chemokines. Most chemokines are more effective on TNFalpha-primed cells. Our results suggest that the mobilization of MSCs and their subsequent homing to injured tissues may depend on the systemic and local inflammatory state. Disclosure of potential conflicts of interest is found at the end of this article.

Pulmonary Stromal-Derived Factor-1 Expression and Effect on Neutrophil Recruitment during Acute Lung Injury

The severe and protracted inflammation that characterizes acute lung injury (ALI) is driven by the ongoing recruitment of neutrophils to the lung. Although much of the cytokine signaling responsible for the initial phase of ALI has been elaborated, relatively little is known about the mechanisms governing the recruitment of neutrophils from the bone marrow to the lung in the later period of this disease. Given its previously described chemoattractant effects on marrow neutrophils, we investigated whether stromal-derived factor-1 (SDF-1) (CXCL12) might participate in this later phase of recruitment. Using immunohistochemistry to examine both banked human lung specimens from patients with ALI and lungs from mice with LPS-induced pneumonitis, we found that pulmonary SDF-1 expression increases during ALI. We further determined that both lung SDF-1 protein expression and mRNA expression rise in a delayed but sustained pattern in this mouse model and that the major source of the increase in expression appears to be the lung epithelium. Lastly, we found that expression of the SDF-1 receptor CXCR4 rises in a similar temporal pattern on neutrophils in both the blood and airspace of LPS-injured mice and that Ab-mediated SDF-1 blockade significantly attenuates late but not early pulmonary neutrophilia in this model. These results implicate SDF-1 in neutrophil recruitment to the lung in the later period of acute lung injury and suggest a novel role for this cytokine in coordinating the transition from the inflammatory response to the initiation of tissue repair.

Plasma level of stromal derived factor-1 (SDF-1) is increased in disseminated intravascular coagulation patients who have poor outcomes: In vitro effect of SDF-1 on coagulopathy

Stromal cell-derived factor-1 (SDF-1) is a CXC chemokine that activates and directs the migration of leukocytes that have CXCR4, which is the unique receptor for SDF-1. Although SDF-1/CXCR4 interaction has been implicated in various inflammatory conditions, its role in modulating coagulation has not been determined. We studied the plasma SDF-1 levels in 90 patients with suspected disseminated intravascular coagulation (DIC) and we found that circulating SDF-1 was significantly increased in the overt DIC patients and was also increased in overt DIC patients who have a poor outcome. We then tested in vitro whether SDF-1 can affect the expression of monocyte tissue factor (TF) and endothelial thrombomodulin (TM), and both of these play important roles in coagulopathy. SDF-1 did not affect the expression of surface TF protein and its function and the TF mRNA level in both monocytes and the monocytic leukemia cell line THP-1. SDF-1 also did not change the surface TM expression of endothelial cells. SDF-1 could enhance low-dose ADP induced platelet aggregation, although it failed by itself to induce aggregation. These findings suggest that plasma SDF-1 might be closely associated with hypercoagulability though its action as a platelet activator.

Stromal-Derived Factor-1 (CXCL12) Regulates Laminar Position of Cajal-Retzius Cells in Normal and Dysplastic Brains

Normal brain development requires a series of highly complex and interrelated steps. This process presents many opportunities for errors to occur, which could result in developmental defects in the brain, clinically referred to as malformations of cortical development. The marginal zone and Cajal-Retzius cells are key players in cortical development and are established early, yet there is little understanding of the factors resulting in the disruption of the marginal zone in many types of cortical malformation syndromes. We showed previously that treatment with methylazoxymethanol in rats causes marginal zone dysplasia with displacement of Cajal-Retzius cells to deeper cortical layers. Here we establish that loss of activity of the chemokine stromal-derived factor-1 (SDF1) (CXCL12), which is expressed by the leptomeninges, is necessary and sufficient to cause marginal zone disorganization in this widely used teratogenic animal model. We also found that mice with mutations in the main receptor for SDF1 (CXCR4) have Cajal-Retzius cells displaced to deeper cortical layers. Furthermore, by inhibiting SDF1 signaling in utero by intraventricular injection of a receptor antagonist, we establish that SDF1 signaling is required for the maintenance of Cajal-Retzius cell position in the marginal zone during normal cortical development. Our data imply that cortical layering is not a static process, but rather requires input from locally produced molecular cues for maintenance, and that complex syndromes of cortical malformation as a result of environmental insults may still be amenable to explanation by interruption of specific molecular signaling pathways.

Nitric Oxide Cytoskeletal–Induced Alterations Reverse the Endothelial Progenitor Cell Migratory Defect Associated With Diabetes

Stromal-derived factor-1 (SDF-1) is a critical chemokine for endothelial progenitor cell (EPC) recruitment to areas of ischemia, allowing these cells to participate in compensatory angiogenesis. The SDF-1 receptor, CXCR4, is expressed in developing blood vessels as well as on CD34+ EPCs. We describe that picomolar and nanomolar concentrations of SDF-1 differentially influence neovascularization, inducing CD34+ cell migration and EPC tube formation. CD34+ cells isolated from diabetic patients demonstrate a marked defect in migration to SDF-1. This defect is associated, in some but not all patients, with a cell surface activity of CD26/dipeptidyl peptidase IV, an enzyme that inactivates SDF-1. Diabetic CD34+ cells also do not migrate in response to vascular endothelial growth factor and are structurally rigid. However, incubating CD34+ cells with a nitric oxide (NO) donor corrects this migration defect and corrects the cell deformability. In addition, exogenous NO alters vasodilator-stimulated phosphoprotein and mammalian-enabled distribution in EPCs. These data support a common downstream cytoskeletal alteration in diabetic CD34+ cells that is independent of growth factor receptor activation and is correctable with exogenous NO. This inability of diabetic EPCs to respond to SDF-1 may contribute to aberrant tissue vascularization and endothelial repair in diabetic patients.

Megakaryocyte-Active Chemokines: Dysregulation in the SDF-1a/CXCR4 Axis in Patients with Essential Thrombocythemia.

Chemokines belong to a large family of molecules that are implicated in the localization and production of blood cells. Some of them, such as Interleukin 8 (IL-8) and GRO-a, participate in the regulation of megakaryopoiesis, mostly by exerting an inhibitory action. Recently, the involvement of stromal derived factor 1 (SDF-1) in synergizing the stimulatory effect of thrombopoietin on megakaryopoiesis and its participation in megakaryocyte transendothelial migration has been described. The aim of the present study was the evaluation of the plasma levels of IL-8, GRO- a and SDF-1 in patients with essential thrombocythemia (ET), a myeloproliferative disorder characterized by megakaryocytic hyperproliferation with increased circulating platelet count. Besides, the corresponding chemokine receptors were assayed on platelet membrane from ET patients. A cohort of 27 patients diagnosed according to the Polycitemia Vera Study Group were enrolled in the study (mean age, 45; 21 women). Twenty-seven normal subjects matched by sex and age were taken as the control group. The Ethic Committee from IDIM A. Lanari approved the study and all patients and normal controls signed the informed consent. Plasma levels of the chemokines were measured by ELISA technique (R&D Systems) according to the manufacturer. Expression levels of IL-8 receptors (CXCR1 and CXCR2), GRO-a receptors (CXCR2) and SDF-1 receptors (CXCR4) on platelet membrane were evaluated by flow cytometry using specific MoAbs and the corresponding isotype controls (B-D Pharmingen). Flow cytometry results were expressed as relative fluorescence intensity (RFI, the relationship between the mean fluorescence intensity from the specific antibody and the isotype control). Results were expressed as median and range. Statistic analysis was carried out using Mann-Whitney Wilcoxon rank sum test and Wilcoxon signed rank test. Plasma levels of the chemokines measured in 19 ET patients were similar to that found in normal controls, IL-8 2.5, pg/ml (0.8–28.2) and 2.8 pg/ml (1.1–16.5), GRO-a, 30.0 pg/ml (7.4–463.1) and 23.9 pg/ml (9.6–148.0), SDF-1, 1895.0 pg/ml (1246.0–2719.0) and 1915.0 pg/ml (822–2424.0), respectively. The expression levels of CXCR4 receptor was found diminished in platelets from ET patients, RIF 16.94 (1.3–31.3) compared to normal controls, 27.4 (2.4–58.4); p=0.0059, n=10. However, the expression of CXCR1 and CXCR2 in platelets from ET patients was normal. In conclusion, although plasma levels of the chemoquines IL-8, GRO-a and SDF-1 were normal in these patients, the decreased level of CXCR4 on platelet membrane suggests a dysregulation in the SDF-1a/CXCR4 axis in patients with essential thrombocythemia.

Stromal-derived factor-1 promotes the growth, survival, and development of human bone marrow stromal stem cells

AbstractThe maintenance of bone marrow stromal stem cells (BMSSCs) is tightly controlled by the local microenvironment and by autocrine regulatory factors secreted by BMSSCs. To identify such factors, a cDNA subtraction library was generated from purified BMSSCs, based on their high expression of the STRO-1 antigen. Stromal-derived factor-1 (SDF-1) was one differentially expressed gene highly expressed by purified BMSSCs prior to culture. In vitro, immature preosteogenic cells expressed greater levels of SDF-1 when compared with mature cell types representative of osteoblasts and osteocytes/bone lining cells. Furthermore, SDF-1 expression was rapidly down-regulated when BMSSCs were cultured under osteoinductive conditions. BMSSCs were also shown to express functional cell surface SDF-1 receptors (CXCR4). Transduced BMSSC lines, secreting high SDF-1 levels, displayed an enhanced ability to form ectopic bone in vivo, in comparison with control BMSSC lines. Moreover, high SDF-1–expressing BMSSCs displayed an increased capacity for cellular growth and protection against interleukin-4–induced apoptosis. Similarly, fibroblast colony-forming units (CFU-Fs) also displayed increased growth and resistance to α-interferon-2a–induced apoptosis, in synergy with platelet-derived growth factor BB (PDGF-BB) and SDF-1 in vitro. These studies indicate that the chemokine, SDF-1, may play a role in the maintenance, survival, and osteogenic capacity of immature BMSSC populations.