Stromal-derived Factor-1alpha Research Articles

An Acellular Scaffold Facilitates Endometrial Regeneration and Fertility Restoration via Recruiting Endogenous Mesenchymal Stem Cells.

Severe intrauterine adhesions (IUAs), characterized by inadequate endometrial repair and fibrosis, can lead to infertility. Stem cell-based therapies, which deliver mesenchymal stem cells (MSCs) to the wound site, hold a considerable promise for endometrium regeneration. However, some notable hurdles, such as stemness loss, immunogenicity, low retention and survival rate, limit their clinical application. Evidence shows a strategy of mobilizing endogenous MSCs recruitment can overcome the traditional limitations of exogenous stem cell-based therapies. Here, an acellular biomaterial named stromal derived factor-1 alpha (SDF-1α)/E7-modified collagen scaffold (CES) is explored. CES based on harnessing the innate regenerative potential of the body enables near-complete endometrium regeneration and fertility restoration both in a rat endometrium acute damage model and a rat IUA model. Mechanistically, the CES implantation promotes endogenous MSCs recruitment via a macrophage-coordinated strategy; then the homing MSCs exert the function of immunomodulation and altered local microenvironments toward regeneration. To conclude, CES, which can harness endogenous MSCs and overcome the traditional limitations of cell-based therapies, can serve as a clinically feasible and cell-free strategy with high therapeutic efficiency for IUA treatment.

Measurement of multiple cytokines for discrimination and risk stratification in patients with Chagas' disease and idiopathic dilated cardiomyopathy.

Chagas’ disease (CD), caused by the hemoflagellate protozoan, Trypanosoma cruzi, is endemic in most countries of Latin America. Heart failure (HF) is often a late manifestation of chronic CD, and is associated with high morbidity and mortality. Inflammatory processes mediated by cytokines play a key role in the pathogenesis and progression of CD. Keeping in view the inflammatory nature of CD, this study investigated the possible role of 21 different inflammatory cytokines as biomarkers for prediction and prognosis of CD. The plasma concentration of these cytokines was measured in a group of patients with CD (n = 94), and then compared with those measured in patients with dilated cardiomyopathy (DCM) from idiopathic causes (n = 48), and with control subjects (n = 25). Monovariately, plasma levels of cytokines such as stem cell growth factor beta (SCGF beta), hepatocyte growth factor (HGF), monokine induced by interferon gamma (CXCL9), and macrophage inhibitory factor (MIF) were significantly increased in CD patients with advanced HF compared to control group. None of the cytokines could demonstrate any prognostic potency in CD patients, and only MIF and stromal derived factor-1 alpha (CXCL12) showed significance in predicting mortality and necessity for heart transplant in DCM patients. However, multivariate analysis prognosticated a large proportion of CD and DCM patients. In CD patients, HGF and Interleukin-12p40 (IL-12p40) together separated 81.9% of 3-year survivors from the deceased, while in DCM patients, CXCL12, stem cell factor (SCF), and CXCL9 together discriminated 77.1% of survivors from the deceased. The significant increase in plasma concentrations of cytokines such as HGF and CXCL9 in CD patients, and the ability of these cytokines to prognosticate a large proportion of CD and DCM patients multivariately, encourages further studies to clarify the diagnostic and prognostic potential of cytokines in such patients.

SDF-1\u03b1 gene-activated collagen scaffold enhances provasculogenic response in a coculture of human endothelial cells with human adipose-derived stromal cells

Novel biomaterials can be used to provide a better environment for cross talk between vessel forming endothelial cells and wound healing instructor stem cells for tissue regeneration. This study seeks to investigate if a collagen scaffold containing a proangiogenic gene encoding for the chemokine stromal-derived factor-1 alpha (SDF-1α GAS) could be used to enhance functional responses in a coculture of human umbilical vein endothelial cells (HUVECs) and human adipose-derived stem/stromal cells (ADSCs). Functional responses were determined by (1) monitoring the amount of junctional adhesion molecule VE-cadherin released during 14 days culture, (2) expression of provasculogenic genes on the 14th day, and (3) the bioactivity of secreted factors on neurogenic human Schwann cells. When we compared our SDF-1α GAS with a gene-free scaffold, the results showed positive proangiogenic determination characterized by a transient yet controlled release of the VE-cadherin. On the 14th day, the coculture on the SDF-1α GAS showed enhanced maturation than its gene-free equivalent through the elevation of provasculogenic genes (SDF-1α—7.4-fold, CXCR4—1.5-fold, eNOS—1.5-fold). Furthermore, we also found that the coculture on SDF-1α GAS secretes bioactive factors that significantly (p < 0.01) enhanced human Schwann cells’ clustering to develop toward Bünger band-like structures. Conclusively, this study reports that SDF-1α GAS could be used to produce a bioactive vascularized construct through the enhancement of the cooperative effects between endothelial cells and ADSCs.

Mesenchymal Stem Cells Pretreatment With Stromal-Derived Factor-1 Alpha Augments Cardiac Function and Angiogenesis in Infarcted Myocardium

BackgroundStem cell therapy is among the novel approaches for the treatment of post-myocardial infarction cardiomyopathy. This study aims to compare the effect of stromal-derived factor 1 α (SDF1α), mesenchymal stem cells (MSCs) in combination with the lentiviral production of vascular endothelial growth factor (VEGF) on infarct area, vascularization and eventually cardiac function in a rat model of myocardial infarction (MI). MethodsThe influence of SDf1α on MSCs survival was investigated. MSCs were transduced via a lentiviral vector containing VEGF. After that, the effect of mesenchymal stem cell transfection of VEGF-A165 and SDf1α preconditioning on cardiac function and scar size was investigated in five groups of MI rat models. The MSC survival, cardiac function, scar size, angiogenesis, and lymphocyte count were assessed 72 hours and 6 weeks after cell transplantation. ResultsSDF1α decreased the lactate dehydrogenase release in MSCs significantly. Also, the number of viable cells in the SDF1α-pretreated group was meaningfully more than the control. The left ventricular systolic function significantly enhanced in groups with p240MSC, SDF1αMSC, and VEGF-A165MSC in comparison to the control group. ConclusionsThese findings suggest that SDF1α pretreatment and overexpressing VEGF in MSCs could augment the MSCs' survival in the infarcted myocardium, reduce the scar size, and improve the cardiac systolic function.

SDF-1α Gene-Activated Collagen Scaffold Restores Pro-Angiogenic Wound Healing Features in Human Diabetic Adipose-Derived Stem Cells.

Non-healing diabetic foot ulcers (DFUs) can lead to leg amputation in diabetic patients. Autologous stem cell therapy holds some potential to solve this problem; however, diabetic stem cells are relatively dysfunctional and restrictive in their wound healing abilities. This study sought to explore if a novel collagen–chondroitin sulfate (coll–CS) scaffold, functionalized with polyplex nanoparticles carrying the gene encoding for stromal-derived factor-1 alpha (SDF-1α gene-activated scaffold), can enhance the regenerative functionality of human diabetic adipose-derived stem cells (ADSCs). We assessed the impact of the gene-activated scaffold on diabetic ADSCs by comparing their response against healthy ADSCs cultured on a gene-free scaffold over two weeks. Overall, we found that the gene-activated scaffold could restore the pro-angiogenic regenerative response in the human diabetic ADSCs similar to the healthy ADSCs on the gene-free scaffold. Gene and protein expression analysis revealed that the gene-activated scaffold induced the overexpression of SDF-1α in diabetic ADSCs and engaged the receptor CXCR7, causing downstream β-arrestin signaling, as effectively as the transfected healthy ADSCs. The transfected diabetic ADSCs also exhibited pro-wound healing features characterized by active matrix remodeling of the provisional fibronectin matrix and basement membrane protein collagen IV. The gene-activated scaffold also induced a controlled pro-healing response in the healthy ADSCs by disabling early developmental factors signaling while promoting the expression of tissue remodeling components. Conclusively, we show that the SDF-1α gene-activated scaffold can overcome the deficiencies associated with diabetic ADSCs, paving the way for autologous stem cell therapies combined with novel biomaterials to treat DFUs.

Erythropoetin can partially restore cigarette smoke induced effects on Adipose derived Stem Cells.

Adipose derived Stem Cells (ASCs) have been proven to play a key role in tissue regeneration. However, exposure to large amounts of cigarette smoke can drastically diminish their function. Erythropoetin (EPO), can modulate cellular response to injury. Therefore, we investigated the ability of EPO to restore the regenerative function and differentiation capacity of ASCs. Human ASCs were isolated from abdominoplasty samples using standard isolation procedures. Cell identity was established by means of Fluorescence Activated Cell Scanning. Subsequently, isolated ASCs were cultivated with cigarette smoke extract both with and without EPO. Parameters investigated included cellular metabolic activity, adipogenic and osteogenic differentiation capacity, and in vitro wound closure capacity. For further enhancing wound closure, EPO was combined with Granulocyte Macrophage Colony Stimulating Factor (GM-CSF) or Stromal Derived Factor-1 alpha (SDF-1 a). Cigarette smoke reduces adipogenic differentiation, the osteogenic differentiation capacity as well as the in vitro wound healing ability of human derived ASCs. EPO did not change metabolic activity of ASCs significantly. The addition of EPO could partially restore their function. The combination of EPO with GM-CSF or SDF-1 did not result in a synergistic effect regarding wound healing ability. Exposure to cigarette smoke significantly reduced the regenerative potential of ASCs. Treatment of ASCs exposed to cigarette smoke with EPO has the potential to partially restore their function.

Molecular Response of Pulp Fibroblasts after Stimulation with Pulp Capping Materials.

This in vitro study evaluated cell viability and metabolism, nitric oxide release and production of two chemokines and one cytokine by cultured human dental pulp fibroblasts (HDPF) in contact with two glass ionomer cements (Ketac Molar-KM and Vitrebond-VB), Single Bond (SB) and calcium hydroxide (Dycal-DY). Cultures of HDPF were established by means of an explant technique. The specimens were prepared under sterile conditions and in disks measuring 5 mm x 2 mm obtained from a prefabricated mold and placed on a permeable membrane to avoid direct contact with the cells. Cytotoxicity was assessed by Trypan Blue exclusion method and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Nitric oxide release in cell supernatant was detected by the Griess Method whereas stromal derived factor-1 alpha (SDF-1α or CXCL12), chemokine (C-X-C motif) ligand 8 [Interleukin 8 (IL-8 or CXCL8)] and interleukin-6 (IL-6) were detected by ELISA. RT-qPCR was employed for gene expression analysis. Statistical analyses were performed by One-way ANOVA followed by Tukey's post hoc test for materials independent of the time, and Two-way ANOVA followed by Bonferroni correction test for the comparisons between materials and experimental time (p<0.05). Cytotoxic tests showed significant differences only for DY. Protein levels and mRNA expression were significantly increased for IL-8 for both periods of time. IL-6 production increased when fibroblasts were stimulated by KM. SDF-1α protein production and mRNA expression were not affected by any of the materials. There was a decrease in nitrate/nitrite levels only for KM. Although DY caused intense cell death and did not stimulate the production of the inflammatory mediators evaluated in this work, it is known that this event seems to be fundamental for the process of repair of the pulp tissue and formation of mineralized barrier. KM and VB increased production of proteins related to the inflammatory process, thus favoring tissue repair. Therefore, although these glass ionomer cements did not lead to large cell death, they should be used with caution.

1096-P: DPP-4 Inhibitor Teneligliptin Improves the Vascular Endothelial Function via Mechanism Different from Changes of EPCs Levels and Blood Glucose Metabolism

Background: DPP-4 inhibitors have been reported to have beneficial effects on endothelial function as sitagliptin increases circulating endothelial progenitor cells (EPCs) via inhibition of plasma stromal derived factor-1 alpha (SDF-1α). However, its mechanisms are unclear. We evaluated whether DPP-4 inhibitor teneligliptin increases the number of circulating EPCs through inhibition of SDF-1α metabolism and improves flow mediated vascular dilatation (FMD) in type 2 diabetes mellitus (T2DM) patients with acute coronary syndrome (ACS) or its risk factors. Methods: This study was a prospective, single-center, open-label, randomized controlled trial. A total of 16 patients with present or past history of ACS or multiple cardiovascular risk factors were enrolled, and who take any medicine for diabetes, whose HbA1c &amp;gt; 7.5% and/or peak CPK &amp;gt; 2000 IU/ml, were excluded. Variable of glucose and lipids, the number of EPCs, activity levels of DPP-4 and SDF-1α and FMD were evaluated at baseline and at 28±4 weeks after enrollment in all patients. They were randomly assigned into 2 groups either 20mg per day teneligliptin or control. Finally, 8 patients were included in teneligliptin group and 8 in control. All data were analyzed by paired t-test, and p &amp;lt; 0.05 was considered statistically significant. Results: DPP-4 activity (∆-509.5±105.7 vs. ∆32.8±53.4 μU/ml) and SDF-1α levels (∆-695.6±443.2 vs. ∆11.1±193.7 pg/ml) were significantly decreased after 28 weeks in teneligliptin group compared to control, respectively. Number of EPCs, and variables of glucose and lipids were not significantly different between two groups before and after 28 weeks. However, FMD was significantly improved in the teneligliptin group compared to control (∆3.8±2.1 vs. ∆-0.3±2.9%, p = 0.006) after 28 weeks treatment. Conclusion: These results indicate that teneligliptin improves FMD by different mechanisms from the increase of blood EPC number via SDF-1α. Disclosure N. Akashi: None. T. Umemoto: None. Y. Hodaka: None. T. Fujiwara: None. K. Yamamoto: None. Y. Taniguchi: None. K. Sakakura: None. H. Wada: None. S. Momomura: None. H. Fujita: None.

Mechanical Cyclic Stretch Regulate Angiogenic Abilities of Endothelial Progenitor cells

Background & Aim Human endothelial progenitor cells (EPCs) are adult stem cells, located in the bone marrow (BM), peripheral blood, placenta and others. EPCs have important role in tissue regeneration via angiogenesis and vasculogenesis. In response to tissue injury, BM-EPCs mobilize to injury site where various growth factors such as vascular endothelial growth factors (VEGF), stromal derived factor 1 alpha (SDF-1a) are released. At the injured site, EPCs can differentiate into mature endothelial cells (ECs) and then directly incorporate into the blood vessel through the stimulation of growth factors and change of cell to cell contact. Because of immunogenicity, autologous EPC therapy is preferable than allograft. However, the number and function of EPCs derived from old and chronic patients are insufficient for successful treatment. In order to retrieve sufficient numbers of EPC from old or chronic patients and restore its function for clinical application, suitable cell preconditioning such as cell stretch, shear stress, and mechanical pressure given by blood flow in vivo may be required before transplantation. Our previous studies have shown that exposure of ECs to mechanical stretch in vitro increased the abilities of migration, proliferation, and tube formation, which could accelerate angiogenesis in impaired tissue. Methods, Results & Conclusion In this study, we explored whether mechanical stretch can improve the survival and proliferation of EPC derived from old or chronic patients by applying cyclic stretch in the range of 5%-15% with use of computerized cell strain system Flexcell-6000. In response to cyclic strain, EPCs increased cell viability, migration and tube formation ability based on matrigel tube assay. Effect of cyclic stretch on the expression of a variety of paracrine factors and extracellular matrix and also their efficacy on vasculogenesis in vivo using limb ischemia model will be estimated.

Hypoxic Preconditioning Effects of Bone Marrow-derived Culture Mesenchymal Stem Cells on CD31+ Expression, Vascular Endothelial Growth Factors-a (VEGF-A) and Stromal-derived Sactors-1 Alpha (SDF-1α)

Increasing the ability of survival stem cells can be determined by assessing the state of hypoxic preconditioning (HPC) in stimulated cells. The assessment is carried out through bone marrow-derived stem (BMSCs) cultures obtained from the femoral bone marrow aspiration procedure (Wismar Rat). This study aimed to prove the hypothesis that there is CD31+ expression on HPC cultures; also, the concentration of VEGF-A and SDF-1α were higher in HPC than control. This research is designed as a laboratory experiment for a period of three months. Male Wistar rat (n = 16) was divided into 2 groups: P0 controls (21% [O2]) and P1 treatments (1% [O2]). Each sample was run into eight repetitions and observed for 48 hours. The stages in this study include isolation and culture of BMSCs, identification of phenotypic BMSCs based on the expression of cell-surface markers (CD105+, CD34−) and hypoxic exposure. After that, it was observed that the potential of angiogenesis in samples was based on CD31+ expression, the paracrine activity of vascular endothelial growth factor-A (VEGF-A) and stromal-derived factor-1 Alpha (SDF-1α). The results showed that there is no CD31+ expression on P1. A of VEGF-A and SDF-1⌑ were higher in P1 than P0.

Role of Acute Myeloid Leukemia Derived-Extracellular Vesicles in Dysregulating Normal Hematopoiesis

Acute myeloid leukemia (AML) is an aggressive and heterogeneous clonal disorder of hematopoietic stem/progenitor cells (HSPCs). Leukemogenesis is characterized by rapid growth and impaired differentiation of leukemic cells, which progressively occupy and likely alter bone marrow (BM) niche where normal HSPCs reside [Kumar B., et al. Leukemia. 2018]. The homing and adhesion of AML cells to the BM niche is critical for its pathogenesis and involves many adhesion molecules, chemokine receptors, their ligands and signaling cascades. In the healthy context, the CXC chemokine receptor type 4 (CXCR4) expressed on HSPCs and its ligand, stromal derived factor-1 alpha (SDF-1α) produced by BM niche, are key mediators of HSPC-BM niche interaction inducing the homing, quiescence and survival of HSPCs, and the regulation of the HSC pool amount [Sánchez-Aguilera A., et al. Cell Mol Life Sci. 2017]. In AML context, where CXCR4/SDF-1α axis is deregulated, growth, BM niche-homing and adhesion of leukemic cells are altered also inducing AML relapse/resistance [Sánchez-Aguilera A., et al. Cell Mol Life Sci. 2017]. Despite recent improvements in therapeutic approaches, only a fraction of AML patients is cured. Therefore, it is crucial to identify new molecular mechanisms driving the AML pathogenesis. Extracellular vesicles (EVs) are lipidic bi-layer membrane particles released by normal and neoplastic cells. They shuttle a complex molecular cargo which reflects that of the origin cell [Caivano A., et al. Tumor Biol 2015; Caivano A., et al. Cell Oncol 2017] and which can be transferred to target cells altering their biological functions [De Luca L., et al. Oncotarget 2016; De Luca L., et al. Int J Mol Sci. 2017]. In particular, neoplasm derived EVs promote a tumor-supporting environment in non-malignant cells favoring cancer proliferation [Laurenzana I., et al. Stem Cells Int. 2018; De Luca L., et al. Expert Rev Mol Diagn. 2019]. We studied the effect of leukemia derived-EVs (LEVs) on healthy umbilical cord blood HSPCs to explore their potential role in dysregulating normal hematopoiesis. Specifically, the proliferation and differentiation, the quantitative and qualitative colony-forming features and the SDF-1α /CXCR4 mediated-migration of HSPCs induced by LEVs were investigated. EVs were isolated from AML cell lines by centrifugation steps and analyzed for AML surface markers by flow cytometry. Next, they were co-incubated with healthy CD34+ sorted cells for 24 hours. After treatment, cell count, apoptosis, HSPC-specific cluster of differentiation marker (CD10, CD34, CD38, CD45RA, CD90, CD123) expression [Karamitros D., et al. Nat Immunol. 2018], in vitro colony-forming unit cell assay, CXCR4 levels and SDF-1α mediated-migration assay were assessed. Our data showed: i) LEVs displaying specific AML surface markers; ii) a similar number of CD34+ cells in LEV treated-HSPCs, including HSCs, compared with controls; iii) no differences in apoptosis between the two groups; iv) an increase of multipotent progenitor (MPP, p=0.01) associated with a reduction of lymphoid-primed multipotent progenitors (LMPP, p=0.01) in treated cells; v) an increase of common myeloid progenitors (CMP, p=0.01), together with reduction of granulocyte macrophage progenitors (GMP, p=0.01) and megakaryocyte erythroid progenitors (MEP) in treated group; vi) a reduction of colony-forming ability, including erythroid, granulocytic and megakaryocytic colonies (p=0.03), in treated cells; vii) decreased levels of CXCR4 in LEV treated-HSPCs, that were accompanied by reduced HSPC migration mediated by SDF-1α. Altogether, this study suggests that AML derived-EVs can dysregulate the normal hematopoiesis promoting an increase of less differentiated precursors (MPP and CMP) which is associated with a reduction of more committed myeloid progenitors (GMP and MEP) (Figure 1). Furthermore, AML derived-EVs seem to induce less attraction of HSPCs in BM niche by interfering with SDF-1α/CXCR4 axis. Strategies to block EV production and secretion, as well as EV induced-reprogramming, could be novel exciting therapeutic approaches in AML. Disclosures Musto: Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees.

In Vitro Angiogenic Properties of Plasmid DNA Encoding SDF-1α and VEGF165 Genes.

The stromal-derived factor-1 alpha (SDF-1α) and vascular endothelial growth factor (VEGF) play an important role in angiogenesis and exert a significant trophic function. SDF-1α is a chemoattractant for endothelial progenitor cells derived from bone marrow and promotes new blood vessel formation. VEGF regulates all types of vascular growth, stimulates angiogenesis, and is involved in the induction of lymphangiogenesis. The possibility of using these growth factors for regenerative medicine is currently under investigation. The angiogenic potential of a pBud-SDF-1α-VEGF165 bicistronic plasmid construct which simultaneously encodes VEGF165 and SDF-1α genes cDNA was evaluated in this study. The conditioned medium collected from HEK293T cells transfected with the pBud-SDF-1α-VEGF165 plasmid was shown to stimulate the formation of capillary-like structures by human umbilical vein-derived endothelial cells (HUVEC) on Matrigel and to increase the proliferative activity of these cells in vitro. Thus, the pBud-SDF-1α-VEGF165 plasmid exhibits angiogenic properties in cell cultures in vitro. As interest in the development of non-viral techniques for regenerative medicine increases, this plasmid which simultaneously expresses VEGF165 and SDF-1α may provide a platform for advanced methods of stimulating therapeutic angiogenesis.

Pro-angiogenic impact of SDF-1α gene-activated collagen-based scaffolds in stem cell driven angiogenesis

Ensuring an adequate angiogenic response during wound healing is a prevailing clinical challenge in biomaterials science. To address this, we aimed to develop a pro-angiogenic gene-activated scaffold (GAS) that could activate MSCs to produce paracrine factors and influence angiogenesis and wound repair. A non-viral polyethyleneimine (PEI) nanoparticles carrying a gene encoding for stromal derived factor-1 alpha (SDF-1α) was combined with a collagen-chondroitin sulfate scaffold to produce the GAS. The ability of this platform to enhance the angiogenic potential of mesenchymal stem cells (MSCs) was then assessed. We found that the MSCs on GAS exhibited early over-expression of SDF-1α mRNA with the activation of angiogenic markers VEGF and CXCR4. Exposing endothelial cells to conditioned media collected from GAS supported MSCs promoted a 20% increase in viability and 33% increase in tubule formation (p < 0.05). Furthermore, the conditioned media promoted a 50% increase in endothelial cell migration and wound closure (p < 0.005). Gene expression analysis of the endothelial cells revealed that the functional response was associated with up-regulation of angiogenic genes; VEGF, CXCR4, eNOS and SDF-1α. Overall, this study shows collagen-based scaffolds combined with SDF-1α gene therapy can provide enhanced pro-angiogenic response, suggesting a promising approach to overcome poor vasculature during wound healing.

TCT-592 Primary Left Ventricular Unloading Infarct Size by Increasing Myocardial Levels of Stromal Derived Factor One Alpha (SDF1a) in Acute Myocardial Infarction

Stromal derived factor 1 alpha (SDF1a) is a constitutively expressed cardioprotective chemokine that is rapidly degraded by proteases. We reported that compared to Primary Reperfusion (PR), first reducing myocardial demand by activating a trans-valvular pump while delaying coronary reperfusion (

Abstract 452: Primary Left Ventricular Unloading Reduces Infarct Size by Increasing Myocardial Levels of Stromal Derived Factor One Alpha (SDF1a) in Acute Myocardial Infarction

Stromal derived factor 1 alpha (SDF1a) is a constitutively expressed cardioprotective chemokine that is rapidly degraded by proteases. We reported that compared to Primary Reperfusion (PR), first reducing myocardial oxygen demand by activating a trans-valvular pump (Impella CP) while delaying coronary reperfusion (Primary Unloading; PU) reduces infarct size in acute myocardial infarction (AMI). We now hypothesize that PU reduces proteolytic degradation of SDF1a thereby increasing SDF1a signaling via the receptor CXCR4 in AMI. Methods: AMI was induced by occlusion of the left anterior descending artery (LAD) for 90 min in male swine (n=4/group). In the PR group, the LAD was reperfused for 120 min. In the PU group, after 90 min of ischemia an Impella CP was activated and the LAD left occluded for an additional 30 min, followed by 120 min of reperfusion. Whole-transcript expression analysis was performed on RNA from the infarct zone using Porcine 1.0 ST microarrays and ConsensusPathDB programs. Quantitative polymerase chain reaction, western blots, and activity assays determined expression and activity of the SDF1a signaling pathway. Sham operated LV samples served as controls. Results: Compared to PR, PU reduced fibrotic and inflammatory gene expression including reduced transcript levels of matrix-metalloprotease-2 (MMP2), MMP9 and dipeptidyl peptidase-4 (DPP4). Compared to PR, PU increased SDF1a protein levels within the infarct zone. Gel zymography confirmed reduced activity levels of MMP2 and MMP9 within the infarct zone after PU, not PR. Compared to PR, PU attenuated DPP4 protein levels and activity and protein levels of CXCR7, an SDF1a sequestration receptor, within the infarct zone. To explore a functional role for SDF1a in PU, adult male swine received intra-coronary injections of AMD3100, a CXCR4 receptor antagonist, after Impella CP activation. Loss of CXCR4 activity attenuated cardioprotective signaling via Akt, Erk and GSK3b and increased infarct size compared to vehicle treated controls. Conclusion: We introduce a novel mechanism by which PU limits proteolytic degradation and CXCR7 mediated sequestration of SDF1a, thereby increasing cardioprotective signaling via SDF1a and overcoming a critical barrier to SDF1a therapeutics in AMI.

Abstract 215: Allogeneic MSCs Improve Endothelial Function in Patients with Dilated Cardiomyopathy via an SDF-1α-mediated Mechanism and the Suppression of Pathologic Cytokines

Endothelial dysfunction is central to the pathophysiology of heart failure, including dilated cardiomyopathy (DCM). Current drug therapies are unable to halt the progression of DCM, compelling the emergence of novel stem cell therapy approaches. Mesenchymal stem cells (MSCs) are pro-angiogenic, immunomodulatory, antifibrotic, and stimulate endogenous endothelial progenitor (EPC) proliferation and function, thus having the potential to ameliorate endothelial dysfunction. We demonstrated that patients with DCM who received allogeneic MSCs had a significant improvement in endothelial function 3-months post treatment, whereas patients who received autologous MSCs had no improvement. Therefore, we hypothesized that allogeneic MSCs preferentially improve endothelial function via a mechanism involving the suppression of pathologic levels of vascular endothelial growth factor (VEGF), stromal derived factor-1 alpha (SDF-1α), and tumor necrosis factor alpha (TNFα). Accordingly, patient serum VEGF and TNFα were measured at baseline and 3 months post MSC treatment. In vitro, MSC secretion of SDF-1α and TNFα was also measured. Our results show that patients with DCM had elevated levels of VEGF (n=21, 581.2±812.2 pg/mL) and TNFα (n=15, 22±9.4 pg/mL) at baseline, and that only allogeneic MSCs were able to restore these levels toward normal (VEGF: n=10, Δ-267.1±252.1, P=0.01; TNFα: n=8, Δ-7.1±3.1 pg/mL, P=0.0005). While there was no difference in TNFα secretion by autologous or allogeneic MSCs (0.01±0.14 vs. 0.4±0.6 pg/mL), autologous MSCs secreted significantly higher levels of SDF-1α compared to allogeneic MSCs (n=12, 79.3±16.7 vs. 14.2±9.4 pg/mL, P=0.0001). In vitro secreted SDF-1α and serum VEGF and TNFα levels correlated with EPC bioactivity (ΔSDF-1α to ΔEPC-CFUs, R=-0.9, P&lt;0.0001; ΔVEGF to ΔEPC-CFUs, R=-0.7, P=0.001; ΔTNFα to ΔEPC-CFUs, R=-0.6, P=0.01). These findings reveal a novel mechanism by which allogeneic MSCs secrete physiologic levels of SDF-1α resulting in physiologic levels of VEGF signaling, reduced TNFα, increased EPC bioactivity, and improved endothelial function. These findings have important clinical and biological implications for the use of MSCs in patients with DCM.

Role of soluble and membrane-bound dipeptidyl peptidase-4 in diabetic nephropathy.

Diabetic nephropathy is one of the most frequent, devastating and costly complications of diabetes. The available therapeutic approaches are limited. Dipeptidyl peptidase type 4 (DPP-4) inhibitors represent a new class of glucose-lowering drugs that might also have reno-protective properties. DPP-4 exists in two forms: a plasma membrane-bound form and a soluble form, and can exert many biological actions mainly through its peptidase activity and interaction with extracellular matrix components. The kidneys have the highest DPP-4 expression level in mammalians. DPP-4 expression and urinary activity are up-regulated in diabetic nephropathy, highlighting its role as a potential target to manage diabetic nephropathy. Preclinical animal studies and some clinical data suggest that DPP-4 inhibitors decrease the progression of diabetic nephropathy in a blood pressure- and glucose-independent manner. Many studies reported that these reno-protective effects could be due to increased half-life of DPP-4 substrates such as glucagon-like peptide-1 (GLP-1) and stromal derived factor-1 alpha (SDF-1a). However, the underlying mechanisms are far from being completely understood and clearly need further investigations.

Hypoxia modulates cell migration and proliferation in placenta-derived mesenchymal stem cells

ObjectivesFor more than a decade, stem cells isolated from different tissues have been evaluated in cell therapy. Among them, the human bone marrow–derived mesenchymal stem cells (hBM-MSCs) were investigated extensively in the treatment of myocardial infarction. Recently, the human placenta–derived mesenchymal stem cells (hPD-MSCs), which are readily available from a biological waste, appear to be a viable alternative to hBM-MSCs. MethodsC-X-C chemokine receptor type 4 (CXCR4) gene expression and localization were detected and validated in hPD-MSCs and hBM-MSCs via polymerase chain reaction and immunofluorescence. Subsequently, cell culture conditions for CXCR4 expression were optimized in stromal-derived factor-1 alpha (SDF1-α), glucose, and cobalt chloride (CoCl2) by the use of cell viability, proliferation, and migration assays. To elucidate the cell signaling pathway, protein expression of CXCR4, hypoxia-inducible factor-1α, interleukin-6, Akt, and extracellular signal-regulated kinase were analyzed by Western blot. CXCR4-positive cells were sorted and analyzed by florescence-activated cell sorting. ResultsCXCR4 was expressed on both hPD-MSCs and hBM-MSCs at the basal level. HPD-MSCs were shown to have a greater sensitivity to SDF-1α–dependent cell migration compared with hBM-MSCs. In addition, CXCR4 expression was significantly greater in both hPD-MSCs and hBM-MSCs with SDF-1α or CoCl2-induced hypoxia treatment. However, CXCR4+ hPD-MSCs population increased by 10-fold in CoCl2-induced hypoxia. In contrast, only a 2-fold increase was observed in the CXCR4+ hBM-MSCs population in similar conditions. After CoCl2-induced hypoxia, the CXCR4/mitogen-activated protein kinase kinase/extracellular signal-regulated kinase signaling pathway was activated prominently in hPD-MSCs, whereas in hBM-MSCs, the CXCR4/phosphatidylinositol 3-kinase/Akt pathway was triggered. ConclusionsOur current results suggest that hPD-MSCs could represent a viable and effective alternative to hBM-MSCs for translational studies in cardiocellular repair.

Tumor-associated stromal cells as key contributors to the tumor microenvironment.

The tumor microenvironment is a heterogeneous population of cells consisting of the tumor bulk plus supporting cells. It is becoming increasingly evident that these supporting cells are recruited by cancer cells from nearby endogenous host stroma and promote events such as tumor angiogenesis, proliferation, invasion, and metastasis, as well as mediate mechanisms of therapeutic resistance. In addition, recruited stromal cells range in type and include vascular endothelial cells, pericytes, adipocytes, fibroblasts, and bone-marrow mesenchymal stromal cells. During normal wound healing and inflammatory processes, local stromal cells change their phenotype to become that of reactive stroma. Under certain conditions, however, tumor cells can co-opt these reactive stromal cells and further transition them into tumor-associated stromal cells (TASCs). These TASCs express higher levels of proteins, including alpha-smooth muscle actin, fibroblast activating protein, and matrix metalloproteinases, compared with their normal, non-reactive counterparts. TASCs are also known to secrete many pro-tumorigenic factors, including IL-6, IL-8, stromal-derived factor-1 alpha, vascular endothelial growth factor, tenascin-C, and matrix metalloproteinases, among others, which recruit additional tumor and pro-tumorigenic cells to the developing microenvironment. Here, we review the current literature pertaining to the origins of recruited host stroma, contributions toward tumor progression, tumor-associated stromal cells, and mechanisms of crosstalk between endogenous host stroma and tumor cells.

Vasculogenesis and Diabetic Erectile Dysfunction: How Relevant Is Glycemic Control?

Erectile dysfunction (ED) is a complication of diabetes, condition responsible for causing endothelial dysfunction (EDys) and hampering repair mechanisms. However, scarce information is available linking vasculogenesis mediated by Endothelial Progenitor Cells (EPCs) and diabetes-associated ED. Furthermore, it remains to be elucidated if glycemic control plays a role on EPCs functions, EPCs modulators, and penile vascular health. We evaluated the effects of diabetes and insulin therapy on bone marrow (BM) and circulating EPCs, testosterone, and systemic/penile Stromal Derived Factor-1 alpha (SDF-1α) expression. Male Wistar rats were divided into groups: age-matched controls, 8-weeks streptozotocin-induced type 1 diabetics, and insulin-treated 8-weeks diabetics. EPCs were identified by flow cytometry for CD34/CD133/VEGFR2/CXCR4 antigens. Systemic SDF-1α and testosterone levels were evaluated by ELISA. Penile SDF-1α protein expression was assessed, in experimental and human diabetic cavernosal samples, by immunohistochemical techniques. Diabetic animals presented a reduction of BM-derived EPCs and an increase in putative circulating endothelial cells (CECs) sloughed from vessels wall. These alterations were rescued by insulin therapy. In addition, glycemic control promoted an increase in systemic testosterone and SDF-1α levels, which were significantly decreased in animals with diabetes. SDF-1α protein expression was reduced in experimental and human cavernosal diabetic samples, an effect prevented by insulin in treated animals. Insulin administration rescued the effects of diabetes on BM function, CECs levels, testosterone, and plasmatic/penile SDF-1α protein expression. This emphasizes the importance of glycemic control in the prevention of diabetes-induced systemic and penile EDys, by the amelioration of endothelial damage, and increase in protective pathways. J. Cell. Biochem. 118: 82-91, 2017. © 2016 Wiley Periodicals, Inc.