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Hemophilia A arises from mutations in the F8 gene, affecting ~ 1/5000 males. Treatment options include frequent intravenous factor and subcutaneous non-factor therapies. While these approaches have been widely used, they have significant limitations, such as breakthrough bleeds and joint disease due to suboptimal adherence, non-ideal factor kinetics, inhibitor generation, (Weyand, Blood 2018) as well as risk of thrombotic events and coagulation test interference with newer non-factor therapies. (Peters, Nat Rev Drug Discov 2018) Alternative modalities such as cell therapies with genetically modified, ready-made human cells are being investigated. To avoid a cytotoxic immune response by the host, allogeneic cells either need to be physically shielded and/or the host immunosuppressed. Various biomaterials, e.g. hydrogels, could serve as the physical barrier that prevents host immune cells from accessing the allogeneic cells, avoiding the need for immunosuppression altogether. However, the host can still activate a foreign body response (FBR), targeting the biomaterial, which significantly limits cell survival and durability of cell therapies. (Anderson, Semin Immunol 2008) We have successfully identified a library of proprietary small molecules, which when conjugated to alginate used to create encapsulating spheres, limit the FBR (Bochenek, Nat Biomed Eng 2018). In addition, we further reduced the FBR using two-compartment design, 1.5 mm diameter spheres, in which the cells are encapsulated in an inner compartment surrounded by an outer, acellular compartment. Using this innovative technology, we aimed to create a novel product that will deliver long-term, sustained human coagulation factor VIII (hFVIII) in vivo. First, we selected a human epithelial cell line with optimal properties for encapsulation within the spheres; considerations included safety, contact inhibition and longevity. We genetically modified this cell line using a non-viral vector and an optimized the coding sequence for a B-domain deleted hFVIII to create a proprietary engineered cell line that constitutively expresses high levels of this protein. Second, we optimized the inner compartment matrix by modulating cell density/sphere and by the addition of a novel modified alginate; these changes maximized cell viability and protein production in vivo. Finally, we further optimized the acellular outer compartment with a proprietary mixture of small-molecule-modified and unmodified alginates. The resulting SIG-001 product candidate consists of two-compartment, 1.5 mm spheres encapsulating hFVIII-expressing human cells. The spheres are sufficiently porous to allow gasses, nutrients, and secreted proteins to freely diffuse, while limiting FBR and prohibiting cell contact with the host's tissues including immune cells. Our in vitro studies demonstrated similar secretion of hFVIII protein by non-encapsulated and encapsulated engineered cells, along with viability of the same cell line after encapsulation. Several doses of SIG-001 were administered intraperitoneally to mice. Stable hFVIII production and good cell viability was shown for spheres retrieved after long-term placement in immunocompromised mice (up to 6 months). Furthermore, our data showed dose-responsive hFVIII activity and efficacious correction of the bleeding phenotype in immunocompetent Hemophilia A mice (Carmona, ISTH 2019). In conclusion, SIG-001 can deliver sustained therapeutic plasma levels of hFVIII in vivo. Our technology could eliminate the need for regular factor injections, lowering the patient burden and providing consistent factor levels without the peaks and troughs observed with factor and non-factor therapies. It also has the potential for expanded use in pediatric patients, and allows for re-dosing if needed. Additionally, there is no concern about the pre-existing antibodies to viral capsids which limit eligibility for gene therapies. We aim to use our technology platform to develop a new category of medicines for severe chronic diseases including rare blood disorders such as Hemophilia A, and to advance their development into clinical testing. Disclosures Carmona: Sigilon Therapeutics: Employment. Barney:Sigilon Therapeutics: Employment. Sewell:Sigilon Therapeutics: Employment. Newman:Sigilon Therapeutics: Employment. Carroll:Sigilon Therapeutics: Employment. Beauregard:Sigilon Therapeutics: Employment. Huang:Sigilon Therapeutics: Employment. Heidebrecht:Sigilon Therapeutics: Employment. Corzo:Sigilon Therapeutics: Employment. Moller:Sigilon Therapeutics: Employment. Smith:Sigilon Therapeutics: Employment. Peritt:Sigilon Therapeutics: Employment. Vivaldi:Sigilon Therapeutics: Employment.

During tumor progression, alternative splicing gives rise to different Mena protein isoforms. We analyzed how Mena11a, an isoform enriched in epithelia and epithelial-like cells, affects Mena-dependent regulation of actin dynamics and cell behavior. While other Mena isoforms promote actin polymerization and drive membrane protrusion, we find that Mena11a decreases actin polymerization and growth factor-stimulated membrane protrusion at lamellipodia. Ectopic Mena11a expression slows mesenchymal-like cell motility, while isoform-specific depletion of endogenous Mena11a in epithelial-like tumor cells perturbs cell:cell junctions and increases membrane protrusion and overall cell motility. Mena11a can dampen membrane protrusion and reduce actin polymerization in the absence of other Mena isoforms, indicating that it is not simply an inactive Mena isoform. We identify a phosphorylation site within 11a that is required for some Mena11a-specific functions. RNA-seq data analysis from patient cohorts demonstrates that the difference between mRNAs encoding constitutive Mena sequences and those containing the 11a exon correlates with metastasis in colorectal cancer, suggesting that 11a exon exclusion contributes to invasive phenotypes and leads to poor clinical outcomes., Virginia and D.K. Ludwig Fund for Cancer Research (Graduate Student Fellowship), National Institutes of Health (U.S.) (GM58801), Massachusetts Institute of Technology. Ludwig Center for Molecular Oncology, David H. Koch Institute for Integrative Cancer Research at MIT (NCI Core Grant P30-CA14051)

International audience; Protocadherins are a group of transmembrane proteins with homophilic binding activity, members of the cadherin superfamily. Apart from their role in adhesion, the cellular functions of protocadherins are essentially unknown. Protocadherin (PCDH)12 was previously identified in invasive trophoblasts and endothelial and mesangial cells in the mouse. Invalidation studies revealed that the protein was required for optimal placental development. In this article, we show that its human homolog is abundantly expressed in various trophoblast subtypes of the human placenta and at lower levels in endothelial cells. We demonstrate that PCDH12 is shed at high rates in vitro. The shedding mechanism depends on ADAM10 and results in reduced cellular adhesion in a cell migration assay. PCDH12 is subsequently cleaved by the γ-secretase complex, and its cytoplasmic domain is rapidly degraded by the proteasome. PCDH12 shedding is regulated by interlinked intracellular pathways, including those involving protein kinase C, PI3K, and cAMP, that either increase or inhibit cleavage. In endothelial cells, VEGF, prostaglandin E(2), or histamine regulates PCDH12 shedding. The extracellular domain of PCDH12 was also detected in human serum and urine, thus providing evidence of PCDH12 shedding in vivo. Importantly, we observed an increase in circulating PCDH12 in pregnant women who later developed a pre-eclampsia, a frequent pregnancy syndrome and a major cause of maternal and fetal morbidity and mortality. In conclusion, we speculate that, like in mice, PCDH12 may play an important role in human placental development and that proteolytic cleavage in response to external factors, such as cytokines and pathological settings, regulates its activity.

Objective— Endothelial progenitor cells (EPCs) comprise a heterogeneous population of cells, which improve therapeutic neovascularization after ischemia. The neovascularization-promoting potential of progenitor cells depends on survival and retention of the infused cells to the tissue. Caspases mediate apoptosis but are also involved in other critical biological processes. Therefore, we aimed to address the role of caspases in proangiogenic cells. Methods and Results— The caspase-8 inhibitor zIETD abrogated the ex vivo formation of EPCs, inhibited EPC adhesion and migration, and reduced their capacity to improve neovascularization in vivo. Consistently, cells isolated from caspase-8-deficient mice exhibited a reduced capacity for enhancing neovascularization when transplanted into mice after hindlimb ischemia. Because inhibition of Caspase-8 reduced the adhesion and homing functions of EPCs, we further determined the surface expression of integrins and receptors involved in cell recruitment to ischemic tissues. Pharmacological inhibition of caspase-8 and genetic depletion of caspase-8 reduced the expression of the fibronectin receptor subunits α5 and β1 and the SDF-1 receptor CXCR4. Moreover, we identified the E3 ubiquitin ligase Cbl-b, which negatively regulates integrin and receptor-mediated signaling, as a potential Caspase-8 substrate. Conclusion— In summary, our data demonstrate a novel apoptosis-unrelated role of caspase-8 in proangiogenic cells.

Circulating blood–derived vasculogenic cells improve neovascularization of ischemic tissue by a broad repertoire of potential therapeutic actions. Whereas initial studies documented that the cells incorporate and differentiate to cardiovascular cells, other studies suggested that short-time paracrine mechanisms mediate the beneficial effects. The question remains to what extent a physical incorporation is contributing to the beneficial effects of cell therapy. By using the inducible suicide gene thymidine kinase to deplete transplanted cells, we determined the contribution of physical incorporation in 3 animal models. After acute myocardial infarction, depletion of cells 14 days after infusion resulted in a reduction of capillary density and a substantial deterioration of heart function. Likewise, neovascularization of Matrigel plugs and ischemic limbs was significantly suppressed when infused cells were depleted 7 days after infusion. Induction of cell death in the previously transplanted cells reduced perfusion and led to vascular leakage as evidenced by Evans blue extravasation. These results indicate that physical incorporation and persistence of cells contribute to cell-mediated improvement of neovascularization and cardiac function. Long-term paracrine activities and/or cell intrinsic mechanisms may have contributed to the maintenance of functional improvement.

Cell therapy is a novel promising option for treatment of ischemic diseases. Administered endothelial progenitor cells (EPCs) are recruited to ischemic regions and improve neovascularization. However, the number of cells that home to ischemic tissues is restricted. The GTPase Rap1 plays an important role in the regulation of adhesion and chemotaxis. We investigated whether pharmacologic activation of Epac1, a nucleotide exchange protein for Rap1, which is directly activated by cAMP, can improve the adhesive and migratory capacity of distinct progenitor cell populations. Stimulation of Epac by a cAMP-analog increased Rap1 activity and stimulated the adhesion of human EPCs, CD34(+) hematopoietic progenitor cells, and mesenchymal stem cells (MSCs). Specifically, short-term stimulation with a specific Epac activator increased the beta2-integrin-dependent adhesion of EPCs to endothelial cell monolayers, and of EPC and CD34(+) cells to ICAM-1. Furthermore, the Epac activator enhanced the beta1-integrin-dependent adhesion of EPCs and MSCs to the matrix protein fibronectin. In addition, Epac1 activation induced the beta1- and beta2-integrin-dependent migration of EPCs on fibronectin and fibrinogen. Interestingly, activation of Epac rapidly increased lateral mobility of beta1- and beta2-integrins, thereby inducing integrin polarization, and stimulated beta1-integrin affinity, whereas the beta2-integrin affinity was not increased. Furthermore, prestimulation of EPCs with the Epac activator increased homing to ischemic muscles and neovascularization-promoting capacity of intravenously injected EPCs in the model of hind limb ischemia. These data demonstrate that activation of Epac1 increases integrin activity and integrin-dependent homing functions of progenitor cells and enhances their in vivo therapeutic potential. These results may provide a platform for the development of novel therapeutic approaches to improve progenitor cell homing.

Endothelial progenitor cells (EPCs) are recruited to ischemic regions and improve neovascularization. Integrins contribute to EPC homing. High-mobility group box 1 (HMGB1) is a nuclear protein that is released extracellularly on cell necrosis and tissue damage, eliciting a proinflammatory response and stimulating tissue repair. In the present study, we investigated the effects of HMGB1 on EPC homing. EPCs express the HMGB1 receptors RAGE (receptor for advanced glycation end products) and TLR2 (Toll-like receptor 2). EPC migration was stimulated by HMGB1 in a RAGE-dependent manner. In addition, the HMGB1-induced migration of EPCs on fibronectin and fibrinogen was significantly inhibited by antibodies against β 1 and β 2 integrins, respectively. Short-term prestimulation of EPCs with HMGB1 also increased EPC adhesion to endothelial cell monolayers, and this effect was blocked by antibodies to β 2 integrins or RAGE. HMGB1 increased EPC adhesion to the immobilized integrin ligands intercellular adhesion molecule-1 and fibronectin in a RAGE-dependent manner. Strikingly, HMGB1 rapidly increased integrin affinity and induced integrin polarization. Using intravital microscopy in a tumor model of neovascularization, prestimulation of EPCs with HMGB1 enhanced the initial in vivo adhesion of EPCs to microvessels and the recruitment of EPCs in the tumor tissue. In addition, prestimulation of EPCs with HMGB1 increased the homing of EPCs to ischemic muscles. In conclusion, these data represent a link between HMGB1 and integrin functions of EPCs and demonstrate that HMGB1 stimulates EPC homing to ischemic tissues. These results may provide a platform for the development of novel therapeutic approaches to improve EPC homing.

Wiscott Aldrich Syndrome protein (WASP) deficiency results in defects in calcium ion signaling, cytoskeletal regulation, gene transcription and overall T cell activation. The activation of WASP constitutes a key pathway for actin filament nucleation. Yet, when WASP function is eliminated there is negligible effect on actin polymerization at the immunological synapse, leading to gaps in our understanding of the events connecting WASP and calcium ion signaling. Here, we identify a fraction of total synaptic F-actin selectively generated by WASP in the form of distinct F-actin ‘foci’. These foci are polymerized de novo as a result of the T cell receptor (TCR) proximal tyrosine kinase cascade, and facilitate distal signaling events including PLCγ1 activation and subsequent cytoplasmic calcium ion elevation. We conclude that WASP generates a dynamic F-actin architecture in the context of the immunological synapse, which then amplifies the downstream signals required for an optimal immune response., Cancer Research Institute (New York, N.Y.) (Postdoctoral Fellowship), National Institutes of Health (U.S.). Nanomedicine Development Center (PN2EY016586)

β1-Integrins are essential for angiogenesis. The mechanisms regulating integrin function in endothelial cells (EC) and their contribution to angiogenesis remain elusive. Brag2 is a guanine nucleotide exchange factor for the small Arf-GTPases Arf5 and Arf6. The role of Brag2 in EC and angiogenesis and the underlying molecular mechanisms remain unclear. siRNA-mediated Brag2-silencing reduced EC angiogenic sprouting and migration. Brag2-siRNA transfection differentially affected α5β1- and αVβ3-integrin function: specifically, Brag2-silencing increased focal/fibrillar adhesions and adhesion on β1-integrin ligands (fibronectin and collagen), while reducing the adhesion on the αVβ3-integrin ligand, vitronectin. Consistent with these results, Brag2-silencing enhanced surface expression of α5β1-integrin, while reducing surface expression of αVβ3-integrin. Mechanistically, Brag2-mediated αVβ3-integrin-recycling and β1-integrin endocytosis and specifically of the active/matrix-bound α5β1-integrin present in fibrillar/focal adhesions (FA), suggesting that Brag2 contributes to the disassembly of FA via β1-integrin endocytosis. Arf5 and Arf6 are promoting downstream of Brag2 angiogenic sprouting, β1-integrin endocytosis and the regulation of FA. In vivo silencing of the Brag2-orthologues in zebrafish embryos using morpholinos perturbed vascular development. Furthermore, in vivo intravitreal injection of plasmids containing Brag2-shRNA reduced pathological ischemia-induced retinal and choroidal neovascularization. These data reveal that Brag2 is essential for developmental and pathological angiogenesis by promoting EC sprouting through regulation of adhesion by mediating β1-integrin internalization and link for the first time the process of β1-integrin endocytosis with angiogenesis.

Of Life, Limb, and a Small RNA Gene expression in mammals is controlled not only by proteins but by small noncoding RNAs called microRNAs. The involvement of these RNAs provides powerful clues about the molecular origins of human diseases and how they might be treated. Ischemic diseases arise from an inadequate blood supply. Bonauer et al. (p. 1710 , published online 21 May) find that a specific microRNA that is expressed in the cells lining blood vessels (called miR-92a) functions to repress the growth of new blood vessels. MiR-92a probably acts through effects on expression of integrins, proteins involved in cell adhesion and migration. In mouse models in which an inadequate blood supply had caused damage either to heart or limb muscle, therapeutic inhibition of miR-92a led to an increase in blood vessel density in the damaged tissues and enhanced functional recovery.

Progenitor cells (PC) are recruited to ischemic tissues and improve neovascularization. Beta2-integrins are essential for adhesion, transmigration and homing of PC to ischemic tissues. Developmental Endothelial Locus-1 (Del-1) is an extracellular matrix protein, that binds alphaVbeta3- and alphaVbeta5-integrins and is up-regulated during ischemia. Therefore, we investigated the role of endogenous Del-1 for angiogenesis and homing functions of PC. The Del-1-deficient mice (Del-1 −/− ) displayed a significantly increased angiogenic response in ischemic muscles in comparison to the wild type (WT) mice in the model of hind limb ischemia. However, when we assessed the role of Del-1 in HUVEC in vitro , silencing of Del-1 by siRNA did not affect angiogenic sprouting. Moreover, the ischemic muscles of Del-1 −/− mice displayed a higher infiltration with CD45 + hematopoietic cells than WT mice, suggesting that Del-1 may have an inhibitory effect on homing of PC and inflammatory cells to ischemic tissues. Interestingly, in adhesion assays human endothelial progenitor cells (EPC) and murine Lin − progenitor cells bound to Del-1 via beta2-integrins, but not via the alphaVbeta3- and alphaVbeta5- integrins. Furthermore, soluble Del-1 significantly inhibited the adhesion of EPC to HUVEC monolayers and to the major beta2-integrin-ligand, ICAM-1, raising the possibility that Del-1 is a beta2-integrin-inhibitor. Indeed, WT murine bone marrow mononuclear cells displayed higher adhesion rates on Del-1-deficient murine lung endothelial cells (LEC) than on WT LEC. In order to investigate the role of Del-1 for in vivo homing of PC, we intravenously injected murine fluorescence-labeled WT Lin − bone marrow PC in WT and Del-1 −/− mice 2 days after the induction of hind limb ischemia. Interestingly, the homing of injected Lin − cells to ischemic muscles was significantly increased in Del-1 −/− in comparison to WT mice (200± 30 % increase). Taken together, endogenous Del-1 is a new beta2-integrin ligand, which blocks beta2-integrin-dependent adhesion and homing of PC to ischemic tissues. It is conceivable, that endogenous Del-1 may reduce ischemia-induced neovascularization through an inhibitory effect on the beta2-integrins of progenitor and inflammatory cells.

Ras associated protein 1 (Rap1), a small GTPase of the Ras family, has attracted much attention because of its involvement in several aspects of cell adhesion, including integrin- and cadherin-mediated adhesion. Yet, the role of Rap1 genes for integrin signaling in endothelial cells (EC) and angiogenesis has not been investigated. Human umbilical vein endothelial cells (HUVEC) express Rap1a and Rap1b mRNA as assessed by RT-PCR. In order to determine the contribution of Rap1 activity for angiogenesis, we overexpressed Rap1GAP1, a GTPase activating protein, which specifically inhibits the activity of both Rap1a and Rap1b. Overexpression of Rap1GAP1 led to a significant inhibition of angiogenic sprouting of HUVEC under basal conditions and bFGF stimulation by 44 ± 5 % in a 3-dimensional spheroidal system and blocked tube formation in a matrigel assay, migration and adhesion. In order to separately investigate the role of Rap1a and Rap1b genes in angiogenesis, we performed gene silencing with siRNA. Silencing of either Rap1a or Rap1b significantly and additively blocked the sprouting of HUVEC under basal and bFGF-stimulated conditions (Rap1a-siRNA: 55 ± 5 %, Rap1b-siRNA: 61 ± 9 % and Rap1a+Rap1b siRNA: 73 ± 5% inhibition) and significantly reduced HUVEC migration and adhesion on fibronectin and collagen. Moreover, silencing of Rap1a and Rap1b reduced beta1-integrin affinity in HUVEC, suggesting the importance of Rap1a and Rap1b for inside-out integrin activation in EC. In addition, silencing of Rap1a and Rap1b prevented VEGF-induced PKB/Akt1 activation. These data prompted us to investigate the in vivo role of Rap1a using Rap1a-deficient mice. Interestingly, Rap1a −/− mice are born with a substantially reduced mendelian ratio. Rap1a +/− heterozygote mice displayed decreased microvessel density in comparison to wild-type mice (Rap1a +/+ ) in a matrigel plug assay. Moreover Rap1a +/− and Rap1a −/− displayed significantly reduced microvessel density in ischemic muscles in the model of hind limb ischemia in comparison to wild-type mice (Rap1a +/− : 32 ± 3 % ; Rap1a −/− : 43 ± 3 % inhibition). Thus, our data demonstrated a critical role of Rap1 in the regulation of β1-integrin signaling in endothelial cells and for postnatal neovascularization.

Ras-associated protein 1 (Rap1), a small GTPase, attracted attention because of its involvement in several aspects of cell adhesion, including integrin- and cadherin-mediated adhesion. Yet, the role of Rap1 genes and of Rap1 effectors for angiogenesis has not been investigated. Human umbilical vein endothelial cells (HUVECs) express Rap1a and Rap1b mRNA. To determine the contribution of Rap1 activity for angiogenesis, we overexpressed Rap1GAP1, a GTPase-activating protein that inhibits Rap1 activity. Overexpression of Rap1GAP1 significantly blocked angiogenic sprouting and tube-forming activity of HUVECs as well as migration and integrin-dependent adhesion. Silencing of Rap1a, Rap1b, or both significantly blocked HUVECs sprouting under basal and basic fibroblast growth factor-stimulated conditions and reduced HUVEC migration and integrin-dependent adhesion. We found that Rap1a and Rap1b are essential for the conformational activation of β1-integrins in endothelial cells. Furthermore, silencing of Rap1a and Rap1b prevented phosphorylation of tyrosine 397 in focal adhesion kinase (FAK) and vascular endothelial growth factor-induced Akt1-activation. Rap1a−/−-deficient and Rap1a+/− heterozygote mice displayed reduced neovascularization after hind limb ischemia compared with wild-type mice. Silencing of RAPL significantly blocked the Rap1-induced sprouting of HUVECs, suggesting that the angiogenic activity of Rap1 is partly mediated by RAPL. Our data demonstrate a critical role of Rap1 in the regulation of β1-integrin affinity, adhesion, and migration in endothelial cells and in postnatal neovascularization.

Endothelial progenitor cells (EPCs) and hematopoietic progenitor cells are recruited to ischemic regions, improving neovascularization. β1 and β2 integrins play a crucial role for progenitor cell homing to ischemic tissues. Integrin activity is regulated by chemokines and their respective G protein–coupled receptors. The phosphatidylinositol-3-kinase catalytic subunit γ (PI3Kγ) is the PI3K isoform that selectively transduces signals from G protein–coupled receptors. Here, we investigated the role of PI3Kγ as a signaling intermediate in the chemokine-induced integrin-dependent homing functions of progenitor cells. A pharmacological PI3Kγ inhibitor significantly reduced chemokine-induced chemotaxis and stromal cell–derived factor (SDF)1α-induced transmigration of human EPCs. Moreover, the PI3Kγ inhibitor significantly reduced SDF1α-induced adhesion of EPCs to intercellular adhesion molecule-1 and human umbilical vein endothelial cell monolayers. These findings were corroborated with Lin−bone marrow–derived progenitor cells from PI3Kγ-deficient mice that displayed reduced SDF1α-induced migration and intercellular adhesion molecule-1 adhesion as compared with wild-type cells. Pharmacological inhibition or genetic ablation of PI3Kγ reduced SDF1α-induced integrin activation in human EPCs and in murine Lin−BM-derived progenitor cells, respectively. In vivo, the homing of PI3Kγ-deficient Lin−progenitor cells to ischemic muscles after intravenous infusion in the model of hindlimb ischemia and their neovascularization-promoting capacity was reduced as compared with wild-type cells. In conclusion, PI3Kγ is integral to the integrin-dependent homing of progenitor cells.

Endothelial progenitor cells (EPC) are recruited to ischemic regions to improve neovascularization. β1- and β2-integrins play a crucial role for progenitor cell homing to ischemic tissues. Chemokines and their respective G-protein coupled receptors (GPCRs) are involved in the EPC homing to ischemic tissues. The phosphatidylinositol-3-kinase catalytic subunit gamma (PI3Kγ) is the PI3K isoform, which selectively transduces signals from GPCRs. Here, we investigated the role of PI3Kγ for integrin-dependent homing functions of progenitor cells. As assessed by western blot, EPC express the catalytic subunit PI3Kγ. We then studied the role of PI3Kγ for EPC migration. AS-605240 (100 nM), a selective PI3Kγ-inhibitor (Camps M, Nat. Med., 2005), significantly reduced the SDF1- and the IL-8-induced migration and the SDF1-induced transendothelial migration of human EPC. Adhesion is a further essential step during EPC homing to ischemic tissues. In this regard, the PI3Kγ-inhibitor significantly reduced the SDF1-induced adhesion of EPC on HUVEC monolayers by 69 ± 8 % and on ICAM-1, a β2-integrin ligand. However, the PI3Kγ-inhibitor did not affect the SDF1-induced adhesion of EPC on fibronectin, a β1-integrin ligand, suggesting that PI3Kγ in EPC is involved in the regulation of β2-, but not of β 1-integrin-dependent adhesion. In line with these results, inhibition PI3Kγ blocked the SDF1-induced increase of β2-, but not of β1-integrin-affinity in EPC. Beside EPC, the SDF1-induced migration and adhesion on ICAM-1 of murine bone marrow (BM)-derived Lin − progenitor cells from PI3Kγ-deficient mice (PI3Kγ − / − ) were reduced in comparison to wild type (WT) cells. In addition, PI3Kγ-deficiency led to a significant reduction of homing of murine BM-Lin − progenitor cells to ischemic muscles after intravenous infusion in the model of hind limb ischemia in comparison to WT cells (48 ± 8 % inhibition). In conclusion, these data demonstrate that PI3Kγ plays an essential role for the integrin-dependent homing of progenitor cells in vitro and in vivo. The understanding of the molecular mechanisms of progenitor cell homing is essential for the development of new therapeutic strategies in order to improve the efficacy of cell-based therapies in patients with ischemic disorders.