Stromal Factor Research Articles

The effects of stromal cel1-derived factor 1 on the migration of neural stem cells and the recovery of lower limb function after spinal cord injury

Objective To assess the effect of stromal cel1-derived factor 1 (SDF-1) on the migration of neural stem cells and recovery of lower limb function after spinal cord injury (SCI). Methods A modified version of Allen's method was used to establish a SCI model in each of 96 adult male Sprague-Dawley rats. They were then randomly divided into group A which received an injection of phosphate buffer solution, group B which received an injection of neural stem cells, group C which received a combination of SDF-1 and neural stem cells, and group D which received AMD3100 and neural stem cells 7 days after the modeling. The functioning of the hind limbs of all of the rats was assessed on the 7th, 14th, 21st and 28th day after the modeling. The rats were then sacrificed and frozen sections of their spinal cords were stained with hematoxylin-eosin (HE) and marked with CM-Dil under fluorescent light. Results An accumulation of fluorescing cells were observed in spine cords from both group B and C, with the counts of group B [(23.6±3.7), (18.9±5.6)and(15.2±4.3)respectively] at the day 14, 21 and 28 after modeling significantly smaller than those of group C [(27.4±4.7), (20.4±5.2) and (18.3±3.9) respectively]. During the same period of time, the average BBB scores of groups B and C were significantly better than those of groups A and D. Moreover, the average score of group C was significantly higher than group B's at all time points. Conclusions Transplanted neural stem cells can migrate to the injured site, surviving and differentiating to promote the recovery of limb function in rats with spinal cord injury. SDF-1 can promote that migration and proliferation. Moreover, CXCR4 receptor's antagonist AMD3100 can significantly hinder neural stem cells' migration to the injured site on the spinal cord. Key words: Neural stem cells; Spinal cord injury; Stromal cel1 derived factor

A cross-talk network that facilitates tumor virotherapy.

To target and kill cancer cells, oncolytic viruses exploit signaling pathways that promote tumor growth. A new study shows that the cancer stromal fibroblast factor FGF2 is a crucial component of a tumor–TGF-β axis that renders cancer cells sensitive to virus infection.

Attenuated store-operated Ca2+ entry in mesenteric artery endothelial cells of aged rat

Objective To observe attenuated store-operated Ca2+ entry in superior mesenteric artery endothelial cells of aged rat. Methods The pressure myograph was applied to measure the vasodilation of superior mesenteric artery induced by bradykinin (BK) in aged versus non-elder rats. Primarily cultured MAECs were treated by thapsigargin (TG) and BK to deplete intracellular Ca2+ stores for comparing SOCE. SOCE of MAECs was detected by calcium imaging technology and was compared between aged and non-elder rats. The expression levels of Orai 1 and stromal interaction factor 1 (STIM 1) were observed by immunofluorescence method. Results Compared with the non-elder rats (70.0±4.1%), BK-induced vasodilation in aged rats (27.0±4.9)% was declined by (60.7±4.3)%. SOCE induced by BK and TG in primarily cultured MAECs was attenuated by 37.6% and 39.2% in aged rats (1.71±0.18 and 1.06±0.03) respectively as compared with non-elder rats (2.72±0.39 and 1.75±0.06). The expressions of SOCE's components Orai 1 and STIM 1 in MAECs were decreased obviously in aged rats than in non-elder rats. Conclusions SOCE of MAECs and SOCE-induced vasodilation are significantly attenuated in aged rats. The decreased expression level of Orai 1 and STIM 1 may contribute to this alteration. Key words: Mesenteric artery superior; Endothelial cells; Calcium

Matrix stiffening and β1 integrin drive subtype-specific fibroblast accumulation in lung cancer.

The crucial role of tumor-associated fibroblasts (TAF) in cancer progression is now clear in non-small cell lung cancer (NSCLC). However, therapies against TAFs are limited due to a lack of understanding in the subtype-specific mechanisms underlying their accumulation. Here, the mechanical (i.e., matrix rigidity) and soluble mitogenic cues that drive the accumulation of TAFs from major NSCLC subtypes: adenocarcinoma (ADC) and squamous cell carcinoma (SCC) were dissected. Fibroblasts were cultured on substrata engineered to exhibit normal- or tumor-like stiffnesses at different serum concentrations, and critical regulatory processes were elucidated. In control fibroblasts from nonmalignant tissue, matrix stiffening alone increased fibroblast accumulation, and this mechanical effect was dominant or comparable with that of soluble growth factors up to 0.5% serum. The stimulatory cues of matrix rigidity were driven by β1 integrin mechano-sensing through FAK (pY397), and were associated with a posttranscriptionally driven rise in β1 integrin expression. The latter mechano-regulatory circuit was also observed in TAFs but in a subtype-specific fashion, because SCC-TAFs exhibited higher FAK (pY397), β1 expression, and ERK1/2 (pT202/Y204) than ADC-TAFs. Moreover, matrix stiffening induced a larger TAF accumulation in SCC-TAFs (>50%) compared with ADC-TAFs (10%-20%). In contrast, SCC-TAFs were largely serum desensitized, whereas ADC-TAFs responded to high serum concentration only. These findings provide the first evidence of subtype-specific regulation of NSCLC-TAF accumulation. Furthermore, these data support that therapies aiming to restore normal lung elasticity and/or β1 integrin-dependent mechano regulation may be effective against SCC-TAFs, whereas inhibiting stromal growth factor signaling may be effective against ADC-TAFs. This study reveals distinct mechanisms underlying the abnormal accumulation of tumor-supporting fibroblasts in two major subtypes of lung cancer, which will assist the development of personalized therapies against these cells.

Unmet Medical Needs in Non-Small-Cell Lung Cancer Treatment: How to Design Pre-Emptive Combination Therapies

The rapidly expanding catalogue of human oncogenic mutations, coupled with difficulties in identifying the cellular targets of active compounds in phenotypic screens, has refocused drug discovery efforts on inhibitors of specific cellular proteins. This new ‘target-based’ approach has enjoyed some spectacular successes in several types of tumours, including non-small-cell lung cancer (NSCLC). Epidermal growth factor receptor (EGFR) mutations occur in 17% of NSCLC patients, with notable response to single agent therapy. Unfortunately, all patients eventually develop acquired resistance to EGFR tyrosine kinase inhibitors (TKIs), while complete remission rate to EGFR TKIs monotherapy is low. Priming BIM, a proapoptotic signalling BH3-only protein, induces sensitivity to erlotinib [Tarceva®] in EGFR-mutant cell lines. Synthetic lethal approaches and pre-emptive therapies based on the initial expression of BIM may significantly improve treatment outcomes. EGFR mutations result in transient pro-death imbalance of survival and apoptotic signalling in response to EGFR inhibition. Src homology 2 domain-containing phosphatase 2 is essential to the balance between extracellular signal-regulated kinase, phosphoinositide- 3-kinase/protein kinase B and signal transducer and activator of transcription 3 activity. Furthermore, stromal hepatocyte growth factor confers EGFR TKI resistance and induces inter-receptor crosstalk with Ephrin Type-A receptor 2, CDCP1, AXL, and JAK1. A better understanding of the complex cancer molecular biology of EGFR mutant lung cancer is crucial for development of effective treatment and design of successful future clinical studies.

Three-dimensional power Doppler indices of ovarian stromal blood flow and serum vascular endothelial growth factor after laparoscopic ovarian drilling in women with polycystic ovary syndrome

Abstract Objective To evaluate the effects of laparoscopic ovarian drilling (LOD) on three-dimensional (3D) power Doppler indices of ovarian stromal blood flow and serum vascular endothelial growth factor (VEGF) levels in women with polycystic ovary syndrome (PCOS). Design Prospective controlled study. Setting Minia University Hospital, Minia, Egypt. Material and methods 30 clomiphene citrate resistant women with PCOS undergoing LOD and 30 fertile women with normal ovaries were recruited in this study. Hormonal profile, Doppler indices of ovarian stromal blood flow and serum VEGF assays before and after LOD were evaluated and compared between the two groups. Evaluation was done at the beginning of the study, 1 week after LOD and at 3 and 6 month follow up periods. Main outcome measures 3-D power Doppler indices of both ovaries and serum VEGF concentration. Results Before LOD, serum levels of VEGF, total testosterone (T), free androgen index (FAI), LH, LH:FSH ratio, total antral follicle count (AFC), total ovarian volume (OV) and the 3D power Doppler blood flow indices were significantly higher in the PCOS group than in the control group. After LOD, there was a significant reduction in the serum levels of VEGF, T, sex-hormone binding globulin (SHBG), FAI, LH, LH:FSH ratio, AFC, OV and the 3D power Doppler indices and remained all low at 3 and 6 month follow up. There were significant positive correlations between power Doppler flow indices (VI, FI, and VFI) with serum VEGF, total T, and LH before and after LOD. Conclusions Serum VEGF and ovarian blood flow indices were higher in women with PCOS than in normal women. LOD reduced serum levels of VEGF, in addition to ovarian blood flow indices, in women with PCOS.

Activated hypoxia‐inducible factor‐1α pathway modulates early events in stretch‐induced skin neovascularization via stromal cell‐derived factor‐1 and vascular endothelial growth factor

Tissue expansion promotes skin regeneration. These responses occur only early after mechanical load and are then replaced by apoptosis-related events during stress relaxation. The mechanism modulating this transient process remains unknown. To elucidate key phenomena that drive early regenerative events after tissue expansion. Intraoperative tissue expansion was performed on 25 patients undergoing facial reconstruction. Paired skin biopsies were obtained from an expanded and unexpanded site from each patient. Differentially expressed genes were inspected by microarray and bioinformatic analysis, and dissected by quantitative polymerase chain reaction, Western blot and immunostaining. Paired biopsies from another nine patients undergoing cyclical expansion for 3months were also investigated. A total of 124 upregulated and 282 downregulated genes were identified. Among them, the biological terms of extracellular matrix organization and blood vessel growth were most significantly enriched, as shown by Gene Ontology analysis. GeneMANIA dissection demonstrated an interactive network highlighted by the canonical hypoxia-inducible factor-1α (HIF-1α) pathway with stromal cell-derived factor-1 (SDF-1) and vascular endothelial growth factor (VEGFA) being the hub genes. Levels of the HIF-1α protein and its targets SDF-1 and VEGFA were elevated in expanded skin, and CD31 and Ki67 expression increased, indicating augmented vascularity and cell proliferation. Trafficking of CD34(+) CD133(+) endothelial progenitor cells was enhanced in skin undergoing long-time cyclical expansion, a phenomenon that was usually modulated by the HIF-1α pathway. The HIF-1α pathway is quickly activated and modulates early events in stretch-induced skin neovascularization. The effect may be augmented through enhanced endothelial progenitor cells recruitment into the expanded skin.

Tumor stroma-derived factors skew monocyte to dendritic cell differentiation toward a suppressive CD14+ PD-L1+ phenotype in prostate cancer

Tumor-associated stromal myofibroblasts are essential for the progression and metastatic spread of solid tumors. Corresponding myeloid cell infiltration into primary tumors is a negative prognostic factor in some malignancies. The aim of this study was to define the exact role of stromal myofibroblasts and stromal factors in early prostate carcinoma (PCa) regulating monocyte infiltration and differentiation into dendritic cells (DCs). Epithelial and stromal primary cultures were generated from PCa biopsies and their purity confirmed. Stromal cells produced significantly more of the (C‒C) motif chemokine ligand 2 (CCL2), interleukin 6 (IL-6) and transforming growth factor β (TGFβ) than epithelial cells. Monocyte chemoattraction was predominantly due to stromal-derived factors, mainly CCL2. DCs generated in the presence of stromal (but not epithelial) factors upregulated CD209, but failed to downregulate the monocyte marker CD14 in a signal transducer and activator of transcription 3 (STAT3)-dependent manner. Monocytes exposed to stromal factors did not produce detectable amounts of IL-10, however, upon lipopolysaccharide stimulation, stromal factor generated dendritic cells (sDC) produced significantly more IL-10 and less IL-12 than their conventional DC counterparts. sDC failed to cross-present tumor-antigen to CD8+ T cells and suppressed T-cell proliferation. Most importantly, sDC expressed significantly elevated levels of programmed cell death ligand-1 (PD-L1) in a primarily STAT3 and IL-6-dependent manner. In parallel with our findings in vitro, tumor-infiltrating CD14+ cells in situ were found to express both PD-L1 and CD209, and a higher percentage of tumor-associated CD3+ T cells expressed programmed cell death-1 (PD-1) molecules compared to T cells in blood. These results demonstrate a hitherto undescribed, fundamental contribution of tumor-associated stromal myofibroblasts to the development of an immunosuppressive microenvironment in early PCa.

Immunohistochemical expression of MMP1 and TIMP1 as markers of migration in Hodgkin’s and non-Hodgkin’s lymphoma of the head and neck region (A comparative study)

Background: Malignant lymphoma is a term that describes primary tumors of the lymphoreticular system, almost all of which arise from lymphocytes.MMP-1 is the most ubiquitously expressed interstitial collagenase, a subfamily of MMPs that cleaves stromal collagens. It is also called collagenase-1.TIMPs which inhibits MMP activity and thereby restrict extracellular matrix breakdown, TIMP-1 is a stromal factor that has a wide spectrum of functions in different tissues. Material and Methods: This study was performed on (68) formalin-fixed, paraffin-embedded blocks, histopathologically diagnosed as lymphoma (head and neck lesions). Immunohistochemical staining of MMP1and TIMP1 was performed on each case of the study sample. Results: The expression of MMP1was cytoplasmic, the study cases showed a 98.5% positive reaction to MMP1, score 3 was the most common and found in 60.3% of all cases.The expression of TIMP1was cytoplasmic, 92.6% of cases expressed positive reactions to TIMP1, score 1 was the most common and found in 57.4% of all cases. Conclusion: This study showed for the first time the effect of MMP-1 in HL, which is considered to be as an invasive and migratory cell marker. A significant difference was found among the subtypes of NHL in relation to TIMP1, TIMP1inhibits the effect of MMP1 and as MMP1 is elevated the TIMP1 will be elevated too.

Distinct Stromal Cell Factor Combinations Can Separately Control Hematopoietic Stem Cell Survival, Proliferation, and Self-Renewal

SummaryHematopoietic stem cells (HSCs) are identified by their ability to sustain prolonged blood cell production in vivo, although recent evidence suggests that durable self-renewal (DSR) is shared by HSC subtypes with distinct self-perpetuating differentiation programs. Net expansions of DSR-HSCs occur in vivo, but molecularly defined conditions that support similar responses in vitro are lacking. We hypothesized that this might require a combination of factors that differentially promote HSC viability, proliferation, and self-renewal. We now demonstrate that HSC survival and maintenance of DSR potential are variably supported by different Steel factor (SF)-containing cocktails with similar HSC-mitogenic activities. In addition, stromal cells produce other factors, including nerve growth factor and collagen 1, that can antagonize the apoptosis of initially quiescent adult HSCs and, in combination with SF and interleukin-11, produce >15-fold net expansions of DSR-HSCs ex vivo within 7 days. These findings point to the molecular basis of HSC control and expansion.

Predicting resistance by selection of signaling pathways.

Epidermal growth factor receptor (EGFR) mutations occur in 17% of non-small-cell lung cancer (NSCLC) patients with notable response to single agent therapy but with low complete remission rate and, eventually, disease progression. Priming BIM, a pro-apoptotic signaling BH3-only protein, induces sensitivity to erlotinib in EGFR-mutant cell lines. Synthetic lethal approaches and preemptive therapies based on the initial expression of BIM may significantly improve the treatment outcome. EGFR mutations result in transient pro-death imbalance of survival and apoptotic signaling in response to EGFR inhibition. SHP2 is essential to the balance between ERK and the phosphoinositide-3-kinase (PI3K)/AKT and signal transducer activator of transcription (STAT) activity, while mTOR can be an additional marker for patients with high BIM expression. Furthermore, stromal hepatocyte growth factor (HGF) confers EGFR tyrosine kinase inhibitor (TKI) resistance and induces interreceptor crosstalk with integrin-b4, Eph2, CUB domain-containing protein-1 (CDCP1), AXL and JAK1. Only by understanding better, and in more depth, complex cancer molecular biology will we have the information that will help us to design strategies to augment efficacy of EGFR TKIs and offer our patients the best, most correct therapeutic option.

Construction of recombinant adenovirus vector co-expressing VEGF165 and SDF-1 genes and its expression in ischemic cerebral tissue of rats

To construct a recombinant adenoviral vector carrying and co-expressing vascular endothelial growth factor 165 (VEGF165) and stromal cell derived factor 1 (SDF-1) and explore its co-expression in ischemic brain tissue in rats. The VEGF165 and SDF-1 genes were directionally connected with internal ribosome entry site (IRES). And the double gene co-expression recombinant shuttle plasmid pDC316-VEGF165-IRES-SDF-1 was built with homologous recombination. The resultant plasmid pDC316-VEGF165-IRES-SDF-1 and backbone plasmid pBHGlox_E1, 3Cre were transfected into HEK293 cells by liposome and the recombinant adenoviral particles capable of infection were acquired. With the rounds of amplification, the purified adenoviral vector Ad5-VEGF165-IRES-SDF-1 was obtained with a titer of up to 1×10(10) IU/ml. The rat model of middle cerebral artery occlusion (MCAO) was established by intra-luminal suturing. And the viral vectors were transfused into the lateral ventricle by a stereotactic microinjection. The expressions of VEGF165 and SDF-1 in ischemic brain tissue were examined by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. The results of PCR, double enzyme digestion and gene sequencing showed that both the recombinant plasmid and the constructed adenoviral vector were expressed. And the adenoviral vector Ad5-VEGF165-IRES-SDF-1 could mediate a co-expression of VEGF165 and SDF-1 in ischemic cerebral tissue. The recombinant adenoviral vector carrying VEGF165 and SDF-1 are successfully constructed. And Ad5-VEGF165-IRES-SDF-1 may mediate a co-expression of VEGF165 and SDF-1 in ischemic cerebral tissue of rats.

Stromal derived factor-1 (SDF-1) and its receptors CXCR4 and CXCR7 in endometrial cancer patients.

PurposeOne of the most important function of stromal derived factor-1 (SDF-1) and its receptors, is regulating the process of metastasis formation. The aim of our study was to investigate the correlation between SDF-1, CXCR4 and CXCR7 protein levels measured by immunohistochemistry with the clinicopathological features and the survival of endometrial cancer patients.Materials and Methods92 patients aged 37–84 (mean 65.1±9.5) were enrolled to our study between January 2000 and December 2007. After the diagnosis of endometrial cancer, all women underwent total abdominal hysterectomy, with bilateral salpingoophorectomy and pelvic lymph node dissection. In all patients clinical stage (according to FIGO classification), histologic grade, myometrial invasion, lymph node and distant metastases were determined.Furthermore, the survival time was assessed. Immunohistochemical analyses of SDF-1, CXCR4 and CXCR7 were performed on archive formalin fixed paraffin embedded tissue sections.ResultsStatistically significant correlations (p<0.01) were reported between SDF-1 and the clinical stage of disease, lymph node metastases, distant metastases, deep myometrial invasion (≥50%), cervical involvement, involvement of adnexa. Statistically significant correlation (p<0.01) was found between SDF-1 expression and the risk of the recurrence. Higher SDF-1 expression was associated with ahigher risk of recurrence (p = 0.0001). The results of CXCR4and CXCR7 expression didn't reveal any significant differences(p>0.05) between the proteins expression in the primary tumor cells and the clinicopathological features. Moreover, the Kaplan-Meier analyses demonstrated a stepwise impairment of cancer overall survival (OS) with increasing SDF-1 expression.ConclusionThe important role of SDF-1 as a predictor of negative clinicopathological characteristics of atumor suggests that the expression of this stromal factor should be included in the panel of accessory pathomorphological tests and could be helpful in establishing a more accurate prognosis in endometrial cancer patients.

Evaluation of PLGA Nanoparticles Carrying Leukaemia Inhibitory Factor for Stromal-Like Support of Rat Fetal Dopaminergic Cells

Evaluation of PLGA Nanoparticles Carrying Leukaemia Inhibitory Factor for Stromal-Like Support of Rat Fetal Dopaminergic Cells After Alzheimer’s disease, Parkinson’s Disease (PD) is the second most common progressive neurodegenerative disease of the CNS. This currently incurable condition is characterised by the loss of dopaminergic neurons in the substantia nigra of the ventral midbrain. New cell-based treatments for PD aim to reintroduce healthy dopaminergic (DA) cells to replace the lost dopaminergic input to the striatum which underlies many of the motor, and some of the cognitive, features of this disease. However, a critical step in this process is the survival and integration of the grafted dopaminergic neurons, and by virtue of the process involved in this therapeutic approach, the cells lack both stromal and growth factor support and are acutely vulnerable to ischaemic stress and an innate immune response: furthermore, endogenous immune tolerance needs to be guided to accept the graft as “self” if rejection is to be avoided. In all these respects, nanotechnologies offer a unique solution and here we outline some fundamental biological considerations relevant to cell based therapies for PD.

High Lactate Dehydrogenase 5 Expression Correlates with High Tumoral and Stromal Vascular Endothelial Growth Factor Expression in Gastric Cancer

Background: Lactate dehydrogenase 5 (LDH5) is a major lactate dehydrogenase isoenzyme catalyzing the transformation of pyruvate to lactate to provide anaerobic energy. Vascular endothelial growth factor (VEGF) is expressed in both tumor and stromal cells in gastric cancer. Our aim was to study the prognostic effect of LDH5, and tumoral and stromal expression of the angiogenic factor VEGF in gastric cancer, and the intercorrelation of tissue expression of both factors. Methods: Tissue microarray analysis of 382 consecutive gastric cancer resection specimens was used for immunohistochemistry of LDH5 and VEGF, and expression of LDH5, tumoral VEGF, and stromal VEGF was categorized into low and high groups. Results: High expression was observed for LDH5 in 57.9% (219/378), tumoral VEGF in 35.7% (136/381), and stromal VEGF in 58.5% (223/381) of the specimens. Regarding high expression of LDH5 and VEGF, significant associations with intestinal type, advanced gastric cancer, lymph node metastasis, higher TNM stage, and upper-third location were noted. Positive intercorrelations occurred among the expression of LDH5 and VEGF. Results of survival analyses revealed a significant association of high expression of LDH5 and VEGF with lower survival (overall and disease-free survival). Five-year survival rates were significantly lower in tumors with high LDH5 and tumoral VEGF expression in diffuse- or mixed-type cancers and high expression of stromal VEGF in intestinal-type cancer. Conclusion: The results of our study showed that high LDH5 and VEGF expression in both tumor and stroma was a prognostic factor for patients with gastric cancers, especially diffuse- or mixed-type cancers. Therefore, LDH5 expression may play a role in the regulation of tumoral and stromal VEGF expression in gastric cancer. Our results suggest the potential use of LDH5 expression as a biomarker for response to VEGF-targeted therapy.

Does plerixafor destroy the hematopoietic progenitor cell/mesenchymal stromal cell niche?

Tissue homeostasis is dependent on replacement of damaged and aged cells, and stem cells are the primary source of new cell generation. Most tissue and organs of the body contain stromal cells that are part of the stem cell niche and are essential for the maintenance, proliferation and differentiation of stem cells. Hematopoiesis is a well-characterized tissue system in which interactions between hematopoietic stem cells (HSCs) and stromal cells in the bone marrow (BM) have been clearly established (1e3). The BM environment is subject to complex regulation that is critical for hematopoiesis. Many studies have demonstrated the production of both stimulating and inhibitory factors by BM stromal cells (mesenchymal stromal cells, MSCs) (4e6). However, the full potential of MSCs and their role in control of the HSC proliferation and differentiation remains unclear. Critical to sustained hematopoiesis is themaintenance of stem cell quiescence. Without a niche in which a stem cell pool can reside unstimulated by proliferative and differentiation signals, hematopoiesis cannot be sustained life-long. There is evidence that HSC quiescence is at least in part regulated by direct contact between HSCs and MSCs. This contact is mediated through chemokine receptor 4 (CXCR4) on HSCs engaging stromal cellederived factor (SDF)-1 on MSCs (7). With the advent of plerixafor—a CXCR4 antagonist used to mobilize HSCs for stem cell transplantation—it is pertinent to ask how this agent interferes with HSC/MSC interactions. In this issue of Cytotherapy, Ludwig et al. (8) cocultured HSCs and MSCs with and without plerixafor, studying cell division kinetics and stem cell surface marker changes to explore the relationship between HSCs and MSCs and the role of the CXCR4/SDF-1 axis. They found that co-culture of

Heterogeneity Of CXCR4 Expression In Pediatric B-Cell Precursor Acute Lymphoblastic Leukemia

Background CXCR4 (CD184) is a receptor specific to the Stromal Derived Factor 1 (SDF-1), a ligand also known as CXCL12. The ligand-receptor interaction has a pleiotropic effect on hematopoietic cell proliferation, migration and activation through several signaling pathways. CXCR4 expression on neoplastic cells might be responsible for their dissemination to particular organs with cells expressing CXCL12 (e.g. lymph nodes, bones, and within bone marrow). In B-cell precursor acute lymphoblastic leukemia (BCP-ALL), expression of CXCR4 was associated with higher capacity of leukemic blasts to seed into bone marrow niches. Aim of the study The study aimed at thorough analysis of CXCR4 expression on BCP-ALL blasts and correlation of CXCR4 expression with the expression of other antigens such as CD66c, CD34, CD10, CD38, CD20 and CD45 as well as with the levels of minimal residual disease on day 15. Patients and methods The study group consisted of 198 consecutive children aged 0-18 years (median 4.4 years) treated for BCP-ALL in the centers of the Polish Pediatric Leukemia/Lymphoma Study Group. Bone marrow samples obtained at initial diagnosis were stained with monoclonal antibodies (CD58, CD66c, CD34, CD19, CD10, CD38, CD20, CD45, CXCR4) in two 8-color tubes and analyzed with multiparameter flow cytometry (BD FACS Canto II, Becton Dickinson, San Jose, CA, USA) according to the EuroFlow standard protocols. The expression of particular antigens on BCP-ALL blasts was defined by median fluorescence. In 177 patients the samples from day 15 were available and analyzed for the presence of minimal residual disease (MRD) with multicolor flow cytometry. Infinicyt software (Cytognos, Salamanca, Spain) was used for more detailed analyses of the flow cytometric data. Results The expression of CXCR4 in BCP-ALL was highly variable with median fluorescence ranging from 252 to 24 388 (median 4011). There was no obvious correlation of CXCR4 expression with immunophenotype and with the expression of other analyzed markers (CD66, CD34, CD10, CD38 i CD45). The only borderline significant correlation found was between CXCR4 and CD20 expression. On day 15, 70 children (39%) demonstrated MRD levels below 0.1%, which is consistent with MRD-based low-risk group. Among these patients, 41 children had undetectable MRD already at this time point. In contrary, MRD levels &gt; 10% were recorded in 21 patients (12%), who were stratified to high-risk group, accordingly. Maximal MRD levels recorded at day 15 were 85.6%. In remaining 86 children (49%), MRD levels at day 15 were in-between 0.1% and 10%, which reflects intermediate response to the treatment. There was no correlation between CXCR4 expression and MRD levels at day 15. Conclusion CXCR4 expression on BCP-ALL blasts is highly heterogeneous and is not associated with particular leukemia immunophenotype. Further analyses should characterize clinical features of leukemia and treatment response with regard to CXCR4 expression. The study was supported by Polish National Center of Science grant N N407 687040. Disclosures: No relevant conflicts of interest to declare.

Ultrasound Targeted Microbubble Destruction Promotes Angiogenesis and Heart Function by Inducing Myocardial Microenvironment Change

The myocardial microenvironment plays a decisive role in the survival, migration and differentiation of stem cells. We studied myocardial micro-environmental changes induced by ultrasound-targeted microbubble destruction (UTMD) and their influence on the transplantation of mesenchymal stem cells (MSCs). Various intensities of ultrasound were applied to the anterior chest in canines with myocardial infarction after intravenous injection of microbubbles. The expression of cytokines and adhesion molecules in the infarcted area of the myocardium was detected after three sessions of UTMD in 1wk. Real-time quantitative reverse transcription polymerase chain reaction (RTQ-PCR) showed that the expression of vascular cell adhesion molecule-1 (VCAM-1), stromal cell-derived factor-1 (SDF-1) and vascular endothelial growth factor (VEGF) in the 1.5W/cm(2) and 1W/cm(2) groups was markedly increased compared with the 0.5W/cm(2) or the control groups (3.8- to 4.7-fold, p<0.01), and the expression of interleukin-1β (IL-1β) in the 1.5W/cm(2) group was increased twofold over the 1.0W/cm(2) group, whereas the 0.5W/cm(2) group experienced no significant changes. UTMD at 1.0W/cm(2) was performed as previously described before mesenchymal stem cell (MSC) transplantation. Myocardial perfusion, angiogenesis and heart function were investigated before and 1month after MSC transplantation. Coronary angiography and 99mTc-tetrofosmin scintigraphy revealed that myocardial perfusion was markedly improved after UTMD+MSCs treatment (p<0.05). At echocardiographic analysis, heart function and the wall motion score index were significantly improved by UTMD+MSCs treatment compared with MSCs or UTMD alone and the control. In a canine model of myocardial infarction, therapeutic effects were markedly enhanced by MSC transplantation after the myocardial micro-environmental changes induced by UTMD; therefore, this novel method may be useful as an efficient approach for cellular therapy.

Interleukin-1 beta is an irradiation-induced stromal growth factor

Gamma irradiation of tissues and organs leads to many pathological consequences due to the formation of reactive oxygen species, DNA damage and the subsequent massive death of cells. The therapeutic use of gamma irradiation in the treatment of cancer is based on its penetrating power and damaging effects on tumor cells. Other effects from the irradiation are unnoticeable in comparison. Moreover, the long-term consequences of gamma irradiation are still poorly understood. When a donor bone marrow plug is implanted under the renal capsule of a syngeneic animal, а hematopoietic ectopic focus is formed. The size of the focus is increased in mice that received irradiation compared to non-irradiated ones, regardless of the amount of time between irradiation and bone marrow plug implantation. Long-term repetitive injections of blood serum from irradiated mice given to syngeneic non-irradiated recipients of bone marrow plugs also lead to the formation of enlarged foci. Hence, the blood of irradiated animals must contain an activity that induces the growth of a hematopoietic microenvironment. It was previously shown that the bones of irradiated animals secrete a growth factor required to create stromal microenvironments. The identity of this factor has, until now, been difficult to obtain. We demonstrated that interleukin 1 beta (IL-1) stimulates the growth of murine bone marrow stromal cells in vitro and in vivo. It was shown that the expression of the Il1b gene and the secretion of its product, IL-1, were activated in bone cells long after total body gamma irradiation. Hence, IL-1, or proteins regulated by this cytokine, appears to be the same stromal growth factor previously observed in the serum of irradiated animals. Our data demonstrate several non-canonical functions of IL-1. In addition, the presence of up-regulated levels of IL-1 long after irradiation points to an unknown mechanism governing its gene expression.

Tumor-associated macrophages and stromal TNF-α regulate collagen structure in a breast tumor model as visualized by second harmonic generation

Collagen fibers can be imaged with second harmonic generation (SHG) and are associated with efficient tumor cell locomotion. Preferential locomotion along these fibers correlates with a more aggressively metastatic phenotype, and changes in SHG emission properties accompany changes in metastatic outcome. We therefore attempted to elucidate the cellular and molecular machinery that influences SHG in order to understand how the microstructure of tumor collagen fibers is regulated. By quantifying SHG and immunofluorescence (IF) from tumors grown in mice with and without stromal tumor necrosis factor (TNF)-α and in the presence or absence of tumor-associated macrophages (TAMs), we determined that depletion of TAMs alters tumor collagen fibrillar microstructure as quantified by SHG and IF. Furthermore, we determined that abrogation of TNF-α expression by tumor stromal cells also alters fibrillar microstructure and that subsequent depletion of TAMs has no further effect. In each case, metastatic burden correlated with optical readouts of collagen microstructure. Our results implicate TAMs and stromal TNF-α as regulators of breast tumor collagen microstructure and suggest that this regulation plays a role in tumor metastasis. Furthermore, these results indicate that quantification of SHG represents a useful strategy for evaluating the cells and molecular pathways responsible for manipulating fibrillar collagen in breast tumor models.