Effect of shock wave-facilitated intracoronary cell therapy on LVEF in patients with chronic heart failure: the CELLWAVE randomized clinical trial.

IMPORTANCE: The modest effects of clinical studies using intracoronary administration of autologous bone marrow-derived mononuclear cells (BMCs) in patients with chronic postinfarction heart failure may be attributed to impaired homing of BMCs to the target area. Extracorporeal shock wave treatment has been experimentally shown to increase homing factors in the target tissue, resulting in enhanced retention of applied BMCs. OBJECTIVE: To test the hypothesis that targeted cardiac shock wave pretreatment with subsequent application of BMCs improves recovery of left ventricular ejection fraction (LVEF) in patients with chronic heart failure. DESIGN, SETTING, AND PARTICIPANTS: The CELLWAVE double-blind, randomized, placebo-controlled trial conducted among patients with chronic heart failure treated at Goethe University Frankfurt, Germany, between 2006 and 2011. INTERVENTIONS: Single-blind low-dose (n = 42), high-dose (n = 40), or placebo (n = 21) shock wave pretreatment targeted to the left ventricular anterior wall. Twenty-four hours later, patients receiving shock wave pretreatment were randomized to receive double-blind intracoronary infusion of BMCs or placebo, and patients receiving placebo shock wave received intracoronary infusion of BMCs. MAIN OUTCOMES AND MEASURES: Primary end point was change in LVEF from baseline to 4 months in the pooled groups shock wave + placebo infusion vs shock wave + BMCs; secondary end points included regional left ventricular function assessed by magnetic resonance imaging and clinical events. RESULTS: The primary end point was significantly improved in the shock wave + BMCs group (absolute change in LVEF, 3.2% [95% CI, 2.0% to 4.4%]), compared with the shock wave + placebo infusion group (1.0% [95% CI, -0.3% to 2.2%]) (P = .02). Regional wall thickening improved significantly in the shock wave + BMCs group (3.6% [95% CI, 2.0% to 5.2%]) but not in the shock wave + placebo infusion group (0.5% [95% CI, -1.2% to 2.1%]) (P = .01). Overall occurrence of major adverse cardiac events was significantly less frequent in the shock wave + BMCs group (n = 32 events) compared with the placebo shock wave + BMCs (n = 18) and shock wave + placebo infusion (n = 61) groups (hazard ratio, 0.58 [95% CI, 0.40-0.85]; P = .02). CONCLUSIONS AND RELEVANCE: Among patients with postinfarction chronic heart failure, shock wave-facilitated intracoronary administration of BMCs vs shock wave treatment alone resulted in a significant, albeit modest, improvement in LVEF at 4 months. Determining whether the increase in contractile function will translate into improved clinical outcomes requires confirmation in larger clinical end point trials. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT00326989.

Sphingosine-1-phosphate stimulates the functional capacity of progenitor cells by activation of the CXCR4-dependent signaling pathway via the S1P3 receptor.

OBJECTIVE: Sphingosine-1-phosphate (S1P) is a bioactive lipid, which influences migration and proliferation of endothelial cells through activation of S1P receptors and has been shown to support SDF-1 induced migration and bone marrow homing of CD34+ progenitors. METHODS AND RESULTS: Here, we show that incubation of patient-derived endothelial progenitor cells (EPCs) with S1P or its synthetic analog FTY720 improved blood flow recovery in ischemic hind limbs. Likewise, recovery of blood flow was dramatically reduced after induction of hindlimb ischemia in mice deficient for the S1P receptor 3 (S1P3). S1P3-/- bone marrow-derived mononuclear cells (BMCs) failed to augment neovascularization after hind limb ischemia. Of note, treatment of BMCs derived from S1P3-/- mice with S1P did not rescue blood flow recovery. Mechanistically, S1P and FTY720 induced phosphorylation of CXCR4, activated the Src kinase, and stimulated phosphorylation of JAK2. The contribution of CXCR4 for S1P-mediated effects was further supported by the findings that S1P preincubation failed to stimulate invasion capacity and in vivo blood flow recovery of BMCs from CXCR4+/- mice. The activation of CXCR4 was dependent on the Src kinase family as demonstrated by preincubation with the Src inhibitor PP2. The activation of the CXCR4 signaling by S1P is mediated via the S1P3 receptor, since S1P-induced Src phosphorylation was abrogated in EPC from S1P3-/- mice. CONCLUSIONS: S1P agonists might serve as sensitizers of CXCR4-mediated signaling and may be applied in clinical progenitor cell therapy to improve EPC or BMC function in patients with coronary artery disease.

Therapeutic angiogenesis inhibits or rescues chemotherapy-induced peripheral neuropathy: taxol- and thalidomide-induced injury of vasa nervorum is ameliorated by VEGF.

Toxic neuropathy represents an important clinical problem in the use of the chemotherapeutic substances Taxol and thalidomide. Sensory neuropathy has a high incidence, lacks an effective treatment and is the dose-limiting factor for these drugs. The pathogenic basis of these neuropathies is unknown. We investigated the hypothesis that the experimental toxic neuropathies from Taxol and thalidomide results from destruction of vasa nervorum and can be reversed by the administration of an angiogenic cytokine. In animal models of Taxol- and thalidomide-induced neuropathy, nerve blood flow has been attenuated and the number of vasa nervorum has been reduced. Intramuscular gene transfer of naked plasmid DNA encoding VEGF-1 administered in parallel with Taxol injections completely inhibited deterioration of nerve function and diminution of the peripheral nerve vasculature. Gene therapy in animals with established Taxol- or thalidomide-induced neuropathies resulted in recovery of vascularity and improved nerve electrophysiology. These findings implicate microvascular damage as the basis for toxic neuropathy and suggest that angiogenic growth factors may constitute a novel treatment for this disorder.

Impaired CXCR4 signaling contributes to the reduced neovascularization capacity of endothelial progenitor cells from patients with coronary artery disease.

Transplantation of bone marrow cells as well as circulating endothelial progenitor cells (EPC) enhances neovascularization after ischemia. The chemokine receptor CXCR4 is essential for migration and homing of hematopoietic stem cells. Therefore, we investigated the role of CXCR4 and its downstream signaling cascade for the angiogenic capacity of cultured human EPC. Ex vivo, differentiated EPC derived from peripheral blood abundantly expressed CXCR4. Incubation of EPC from healthy volunteers with neutralizing antibodies against CXCR4 profoundly inhibited vascular endothelial growth factor- and stromal-derived factor-1-induced migration as well as EPC-induced angiogenesis in an ex vivo assay. Preincubation of transplanted EPC with CXCR4 antibody reduced EPC incorporation and impaired blood-flow recovery in ischemic hindlimbs of nude mice (57+/-4% of normal perfusion versus untreated EPC: 80+/-11%, P<0.001). Bone marrow mononuclear cells (BM-MNC) or EPC of heterozygous CXCR4(+/-) mice displayed reduced CXCR4 expression and disclosed impaired in vivo capacity to enhance recovery of ischemic blood flow in nude mice (blood flow 27+/-11% versus 66+/-25% using wild-type cells, P<0.01). Importantly, impaired blood flow in ischemic CXCR4(+/-) mice was rescued by injection of wild-type BM-MNC. Next, we investigated the role of CXCR4 for functional capacities of EPC from patients with coronary artery disease (CAD). Surface expression of CXCR4 was similar in EPC from patients with CAD compared with healthy controls. However, basal Janus kinase (JAK)-2 phosphorylation was significantly reduced and less responsive to stromal-derived factor-1 in EPC from patients with CAD compared with healthy volunteers, indicating that CXCR4-mediated JAK-2 signaling is dysregulated in EPC from patients with CAD. The CXCR4 receptor signaling profoundly modulates the angiogenic activity and homing capacity of cultured human EPC. Disturbance of CXCR4 signaling, as demonstrated by reduced JAK-2 phosphorylation, may contribute to functional impairment of EPC from patients with CAD. Stimulating CXCR4 signaling might improve functional properties of EPC and may rescue impaired neovascularization capacity of EPC derived from patients with CAD.

Effects of granulocyte colony simulating factor on functional activities of endothelial progenitor cells in patients with chronic ischemic heart disease.

OBJECTIVE: Bone marrow-derived circulating endothelial progenitor cells (EPCs) may contribute to regeneration of infarcted myocardium and enhance neovascularization. Granulocyte colony-stimulating factor (G-CSF) is well-established to mobilize hematopoietic stem cells (HSCs) and might, thereby, also increase the pool of endogenously circulating EPC. Therefore, we investigated the effects of G-CSF administration on mobilization and functional activities of blood-derived EPC in patients with chronic ischemic heart disease (CIHD). METHODS AND RESULTS: Sixteen patients with CIHD received 10 microg/kg per day subcutaneous G-CSF injection for 5 days. Leukocyte counts, the number of HSCs and EPCs, and the migratory response to VEGF and SDF-1 were analyzed before and after G-CSF-therapy. At day 5 of G-CSF treatment, the number of circulating leukocytes, CD34+ CD45+ and CD34+ CD133+ cells was significantly increased. Likewise, G-CSF treatment augmented the numbers of colony forming units with endothelial cell morphology (EC-CFU). However, the functional activity of the EPC as assessed by the migratory response to VEGF and SDF-1 was significantly reduced after G-CSF treatment (P<0.01). Because G-CSF was previously shown to cleave the CXCR4 receptor, we determined the surface expression of the 6H8 epitope of the CXCR4 receptor by fluorescence-activated cell sorter (FACS) analysis. Consistent with the reduced migratory capacity, the surface expression of the functionally active CXCR4 receptor was significantly reduced. To test the functional activity of the cultivated EPCs in vivo, cells were intravenously infused in nude mice after hind limb ischemia. EPCs, which were cultivated before G-CSF administration, increased blood flow recovery and prevented limb necrosis. However, infusion of EPCs, which were isolated 5 days after G-CSF treatment from the same patient, showed a reduced capacity to augment blood flow recovery and to prevent necrosis by 27%. CONCLUSIONS: G-CSF treatment effectively mobilizes HSCs and EPCs. However, the migratory response to SDF-1 and in vivo capacity of G-CSF-mobilized EPCs was significantly reduced.

p38 mitogen-activated protein kinase downregulates endothelial progenitor cells.

BACKGROUND: Transplantation of endothelial progenitor cells (EPCs) improves neovascularization after ischemia, but patients with coronary artery disease (CAD) or diabetes mellitus show a reduced number of EPCs and impaired functional activity. Therefore, we investigated the effects of risk factors, such as glucose and TNF-alpha, on the number of EPCs in vitro to elucidate the underlying mechanisms. METHODS AND RESULTS: EPCs of patients or healthy subjects were isolated from peripheral blood. Incubation with glucose or TNF-alpha dose-dependently reduced the number of EPCs (79.9+/-1.3% and 74.3+/-8.1% of control; P<0.05, respectively). This reduction was not caused by apoptosis. TNF-alpha and glucose induced a dose- and time-dependent activation of the p38 MAP kinase, the downstream kinase mitogen- and stress-activated kinase 1, and the transcription factor cAMP-responsive element-binding protein (CREB), in EPCs. Moreover, EPCs from CAD patients had significantly higher basal p38-phosphorylation levels (1.83+/-0.2-fold increase; P<0.05) compared with healthy subjects. The inhibition of the p38-kinase by SB203580 or infection with a dominant negative p38 kinase adenovirus significantly increased basal number of EPCs (136.7+/-6.3% and 142.9+/-18% versus control, respectively). Likewise, ex vivo cultivation of EPCs from patients with CAD with SB203580 significantly increased the number of EPCs and partially reversed the impaired capacity for neovascularization of EPCs in vivo (relative blood flow: 0.40+/-0.03 versus 0.64+/-0.08, P<0.05). The increased numbers of EPCs by SB203580 were associated with an augmentation of EPC proliferation and a reduction of cells expressing the monocytic marker proteins CD14 and CD64, suggesting that p38 regulates proliferation and differentiation events. CONCLUSIONS: These results demonstrate that p38 MAP kinase plays a pivotal role in the signal transduction pathways regulating the number of EPCs ex vivo. SB203580 can prevent the negative effects of TNF-alpha and glucose on the number of EPCs and may be useful to improve the number of EPCs for potential cell therapy.

Antiangiogenesis mediates cisplatin-induced peripheral neuropathy: attenuation or reversal by local vascular endothelial growth factor gene therapy without augmenting tumor growth.

BACKGROUND: Toxic neuropathies induced by cisplatin and other chemotherapeutic agents are important clinical problems because of their high incidence, their lack of effective treatment, and the fact that neuropathy represents a dose-limiting factor for these therapies. The pathogenic basis for toxic neuropathies induced by chemotherapeutic agents has not been completely elucidated. METHODS AND RESULTS: We investigated the hypothesis that experimental toxic neuropathy results from an antiangiogenic effect of these drugs, resulting in destruction of the vasa nervorum, and accordingly that the neuropathy could be prevented or reversed by locally administered VEGF gene transfer without augmenting tumor growth. In an animal model of cisplatin-induced neuropathy, nerve blood flow was markedly attenuated, and there was a profound reduction in the number of vasa nervorum associated with marked endothelial cell apoptosis, resulting in a severe peripheral neuropathy with focal axonal degeneration characteristic of ischemic neuropathy. After intramuscular gene transfer of naked plasmid DNA encoding VEGF-1 in animals with an established neuropathy, vascularity and blood flow returned to levels similar to those of control rats, peripheral nerve function was restored, and histological nerve architecture was normalized. Gene therapy administered in parallel with cisplatin chemotherapy completely attenuated endothelial cell apoptosis and inhibited destruction of nerve vasculature, deterioration of nerve function, and axonal degeneration. In a rat tumor model, VEGF gene transfer administered locally did not alter tumor growth or vascularity. CONCLUSIONS: These findings implicate microvascular damage as the basis for toxic neuropathy induced by cisplatin and suggest that local angiogenic gene therapy may constitute a novel prevention or treatment for this disorder without augmenting tumor growth or vascularization.

FOXO-dependent expression of the proapoptotic protein Bim: pivotal role for apoptosis signaling in endothelial progenitor cells.

Endothelial progenitor cells (EPCs) contribute to postnatal neovascularization. Risk factors for coronary artery disease reduce the number of EPCs in humans. Since EPC apoptosis might be a potential mechanism to regulate the number of EPCs, we investigated the effects of oxidative stress and HMG-CoA-reductase inhibitors (statins) on EPC apoptosis. Atorvastatin, mevastatin, or VEGF prevented EPC apoptosis induced by H2O2. The antiapoptotic effect was reversed by inhibition of the PI3K/Akt pathway. Forkhead transcription factors (FOXO1, FOXO3a, FOXO4) exert proapoptotic effects and are phosphorylated and, thereby, inactivated by Akt. Therefore, we elucidated the involvement of forkhead transcription factors. Atorvastatin induced the phosphorylation of the predominant forkhead factor FOXO4 in EPCs. In addition, atorvastatin reduced the expression of the proapoptotic forkhead-regulated protein Bim in a PI3K-dependent manner. Consistently, overexpression of FOXO4 activated the Bim promoter as determined by reporter gene expression and stimulated the expression of Bim, resulting in an increased EPC apoptosis. Statins failed to prevent EPC apoptosis induced by overexpression of Bim or nonphosphorylatable FOXO4, suggesting that the protective effects of statins depend on this pathway. In summary, our results show that FOXO-dependent expression of Bim plays a pivotal role for EPC apoptosis. Statins reduce oxidative stress-induced EPC apoptosis, inactivate FOXO4, and down-regulate Bim.

Profoundly reduced neovascularization capacity of bone marrow mononuclear cells derived from patients with chronic ischemic heart disease.

BACKGROUND: Cell therapy with bone marrow-derived stem/progenitor cells is a novel option for improving neovascularization and cardiac function in ischemic heart disease. Circulating endothelial progenitor cells in patients with coronary heart disease are impaired with respect to number and functional activity. However, whether this impairment also extends to bone marrow-derived mononuclear cells (BM-MNCs) in patients with chronic ischemic cardiomyopathy (ICMP) is unclear. METHODS AND RESULTS: BM-MNCs were isolated from bone marrow aspirates in 18 patients with ICMP (ejection fraction, 38+/-11%) and 8 healthy control subjects (controls). The number of hematopoietic stem/progenitor cells (CD34+/CD133+), CD49d(+) (VLA-4) cells, and CXCR4+ cells did not differ between the 2 groups. However, the colony-forming capacity of BM-MNCs from patients with ICMP was significantly lower compared with BM-MNCs from healthy controls (37.3+/-25.0 versus 113.8+/-70.4 granulocyte-macrophage colony-forming units; P=0.009). Likewise, the migratory response to stromal cell-derived factor 1 (SDF-1) and vascular endothelial growth factor (VEGF) was significantly reduced in BM-MNCs derived from patients with ICMP compared with BM-MNCs from healthy controls (SDF-1, 46.3+/-26.2 versus 108.6+/-40.4 cells/microscopic field, P<0.001; VEGF, 34+/-24.2 versus 54.8+/-29.3 cells/microscopic field, P=0.027). To assess the in vivo relevance of these findings, we tested the functional activity of BM-MNCs to improve neovascularization in a hindlimb animal model using nude mice. Two weeks after ligation of the femoral artery and intravenous injection of 5x10(5) BM-MNCs, laser Doppler-derived relative limb blood flow in mice treated with BM-MNCs from patients with ICMP was significantly lower compared with mice treated with BM-MNCs from healthy controls (0.45+/-0.14 versus 0.68+/-0.15; P<0.001). The in vivo neovascularization capacity of BM-MNCs closely correlated with the in vitro assessment of SDF-1-induced migration (r=0.78; P<0.001) and colony-forming capacity (r=0.74; P<0.001). CONCLUSIONS: BM-MNCs isolated from patients with ICMP have a significantly reduced migratory and colony-forming activity in vitro and a reduced neovascularization capacity in vivo despite similar content of hematopoietic stem cells. This functional impairment of BM-MNCs from patients with ICMP may limit their therapeutic potential for clinical cell therapy.

Statin therapy accelerates reendothelialization: a novel effect involving mobilization and incorporation of bone marrow-derived endothelial progenitor cells.

BACKGROUND: Primary and secondary prevention trials suggest that statins possess favorable effects independent of cholesterol reduction. We investigated whether statin therapy may also accelerate reendothelialization after carotid balloon injury. METHODS AND RESULTS: Simvastatin treatment in 34 male Sprague-Dawley rats accelerated reendothelialization of the balloon-injured arterial segments (reendothelialized area at 2 weeks, 12.3+/-1.8 versus 5.4+/-1.1 mm2, P< 0.01) and resulted in a dose-dependent (0.2 or 1 mg/kg IP) significant reduction in neointimal thickening at 2, 3, and 4 weeks compared with saline-injected controls (n=18). To elucidate the mechanism, we investigated the contribution of bone marrow-derived endothelial progenitor cells (EPCs) by bone marrow transplantation from Tie2/lacZ mice to background mice or nude rats. X-gal staining of mouse carotid artery specimens revealed a 2.9-fold increase in the number of beta-gal-positive cells per square millimeter appearing on the carotid artery luminal surface at 2 weeks, and double-fluorescence immunohistochemistry disclosed a significant 5-fold increase in the number of double-positive cells (beta-gal, isolectin B4) on the luminal surface in carotid arteries of statin-treated nude rats (20+/-3 versus 4+/-1 cells/mm surface length, P<0.005). Statins increased circulating rat EPCs (2.4-fold at 2 weeks and 2.5-fold at 4 weeks, P<0.001) and induced adhesiveness of cultured human EPCs by upregulation of the integrin subunits alpha5, beta1, alpha(v), and beta5 of human EPCs as shown by reverse transcription-polymerase chain reaction and fluorescence-activated cell sorting. CONCLUSIONS: These findings establish additional mechanisms by which statins may specifically preempt disordered vascular wall pathology and constitute physiological evidence that EPC mobilization represents a functionally relevant consequence of statin therapy.

Secretoneurin, an angiogenic neuropeptide, induces postnatal vasculogenesis.

BACKGROUND: Induction of postnatal vasculogenesis, the mobilization of bone marrow-derived endothelial progenitor cells and incorporation of these cells into sites of blood vessel formation, is a well-known feature of angiogenic cytokines such as vascular endothelial growth factor. We hypothesized that the angiogenic neuropeptide secretoneurin induces this kind of neovascularization. METHODS AND RESULTS: Secretoneurin induced mobilization of endothelial progenitor cells to sites of vasculogenesis in vivo in the cornea neovascularization assay. Progenitor cells were incorporated into vascular structures or were located adjacent to them. Systemic injection of secretoneurin led to increase of circulating stem cells and endothelial progenitor cells. In vitro secretoneurin induced migration, exerted antiapoptotic effects, and increased the number of these cells. Furthermore, secretoneurin stimulated the mitogen-activated protein kinase system, as shown by phosphorylation of extracellular signal-regulated kinase, and activated the protein kinase B/Akt pathway. Activation of mitogen-activated protein kinase was necessary for increase of cell number and migration, whereas Akt seemed to play a role in migration of endothelial progenitor cells. CONCLUSIONS: These data show that the angiogenic neuropeptide secretoneurin stimulates postnatal vasculogenesis by mobilization, migration, and incorporation of endothelial progenitor cells.

Local gene transfer of phVEGF-2 plasmid by gene-eluting stents: an alternative strategy for inhibition of restenosis.

BACKGROUND: Drug-eluting stents represent a useful strategy for the prevention of restenosis using various antiproliferative drugs. These strategies share the liability of impairing endothelial recovery, thereby altering the natural biology of the vessel wall and increasing the associated risk of stent thrombosis. Accordingly, we tested the hypothesis that local delivery via gene-eluting stent of naked plasmid DNA encoding for human vascular endothelial growth factor (VEGF)-2 could achieve similar reductions in neointima formation while accelerating, rather than inhibiting, reendothelialization. METHODS AND RESULTS: phVEGF 2-plasmid (100 or 200 microg per stent)-coated BiodivYsio phosphorylcholine polymer stents versus uncoated stents were deployed in a randomized, blinded fashion in iliac arteries of 40 normocholesterolemic and 16 hypercholesterolemic rabbits. Reendothelialization was nearly complete in the VEGF stent group after 10 days and was significantly greater than in control stents (98.7+/-1% versus 79.0+/-6%, P<0.01). At 3 months, intravascular ultrasound analysis revealed that lumen cross-sectional area (4.2+/-0.4 versus 2.27+/-0.3 mm(2), P<0.001) was significantly greater and percent cross-sectional narrowing was significantly lower (23.4+/-6 versus 51.2+/-10, P<0.001) in VEGF stents compared with control stents implanted in hypercholesterolemic rabbits. Transgene expression was detectable in the vessel wall along with improved functional recovery of stented segments, resulting in a 2.4-fold increase in NO production. CONCLUSIONS: Acceleration of reendothelialization via VEGF-2 gene-eluting stents provides an alternative treatment strategy for the prevention of restenosis. VEGF-2 gene-eluting stents may be considered as a stand-alone or combination therapy.

Insights into early and rapid effects of statin therapy after coronary interventions.

Recent trials provide a wealth of data documenting the benefit of cholesterol-lowering therapy with statins in both primary and secondary prevention. A growing body of evidence indicates that statins possess pleiotropic effects - independent of, or at least in addition to their lipid-lowering capacity, including inhibition of smooth muscle cell proliferation and subsequently neointimal proliferation, platelet aggregation as well as antiinflammatory and direct beneficial effects on endothelial function. The current review summarizes recent findings in non-randomized and randomized trials assessing the efficacy of statin therapy following coronary interventions with or without stent implantation. It highlights the efficacy of statins in the settings of acute coronary syndromes. Current management of acute coronary syndromes requires early administration in combination with acute interventions. High risk patients which receive the greatest benefit could be identified by measuring serum markers such as preprocedural C-reactive protein levels. The present article also describes important novel actions of statins supporting the early initiation of statins post intervention, such as the stimulation of reendothelialization - a crucial step in the healing process of the vascular wall - which is mediated at least in part by mobilization of bone-marrow derived endothelial progenitor cells.

Benefits of immediate initiation of statin therapy following successful coronary stent implantation in patients with stable and unstable angina pectoris and Q-wave acute myocardial infarction.

Statin therapy reduces clinical events in patients with stable coronary artery disease. Recent data indicate that the beneficial effects of statin therapy may also extend to patients experiencing an acute ischemic coronary event. However, the potential role of statins to further modify clinical outcome in patients undergoing coronary stent implantation has not been addressed. Therefore, we investigated whether the initiation of statin therapy immediately after successful coronary stent implantation improves short-term clinical outcome in 704 patients (335 patients with stable angina pectoris [AP], 224 patients with unstable AP, and 145 patients with Q-wave acute myocardial infarction [AMI]). Compared with the lowest risk group (patients with stable AP receiving statin therapy), patients with unstable AP (RR 6.9, 95% confidence interval [CI] 1.5 to 31, p = 0.004) and patients with Q-wave AMI (RR 7.6, 95% CI 1.5 to 37, p = 0.004) experienced an increased risk for the occurrence of the primary combined end point of cardiac death and AMI. Importantly, initiation of statin therapy abrogated the increased risk in patients with unstable AP to the level of patients with stable AP receiving statin therapy (RR 1.5, 95% CI 0.2 to 11, p = 0.7). In contrast, statin therapy did not affect the RR in patients with Q-wave AMI during 6-month follow-up (RR 7.9, 95% CI 1.6 to 39 vs RR 7.6, 95% CI 1.5 to 37, p = NS). The beneficial effects of statin therapy after successful coronary stent implantation in unstable AP were most prominent during the first 4 weeks after the ischemic episode. Statins appear to contribute to the rapid transformation of unstable coronary artery disease into a stable condition with a very low event rate over the forthcoming 6 months in patients with unstable AP undergoing successful coronary stent implantation.

Effects of statins on endothelium and their contribution to neovascularization by mobilization of endothelial progenitor cells.

Statins are potent drugs with a variety of cardiovascular protective effects which appear to occur independent of cholesterol reduction. The vasculoprotective effects of statins might be due to their direct effect on endothelial cells leading to improved nitric oxide (NO) bioavailability. Mechanistically, statins induce endothelial nitric oxide synthesis (eNOS) mRNA stability in endothelial cells and promote eNOS activity through a PI3K/Akt dependent pathway. Novel targets of statins are pro-angiogenic actions including the mobilization and differentiation of bone marrow derived endothelial progenitor cells, which accelerate angiogenesis or re-endothelialization. The functional improvement and increased homing capacity of endothelial progenitor cells induced by statin treatment might reverse impaired functional regeneration capacities seen in patients with risk factors for coronary artery disease or documented active coronary artery disease.

Intraarterial administration of bone marrow mononuclear cells in patients with critical limb ischemia: a randomized-start, placebo-controlled pilot trial (PROVASA).

BACKGROUND: Critical limb ischemia due to peripheral arterial occlusive disease is associated with a severely increased morbidity and mortality. There is no effective pharmacological therapy available. Injection of autologous bone marrow-derived mononuclear cells (BM-MNC) is a promising therapeutic option in patients with critical limb ischemia, but double-blind, randomized trials are lacking. METHODS AND RESULTS: Forty patients with critical limb ischemia were included in a multicenter, phase II, double-blind, randomized-start trial to receive either intraarterial administration of BM-MNC or placebo followed by active treatment with BM-MNC (open label) after 3 months. Intraarterial administration of BM-MNC did not significantly increase ankle-brachial index and, thus, the trial missed its primary end point. However, cell therapy was associated with significantly improved ulcer healing (ulcer area, 3.2±4.7 cm(2) to 1.89±3.5 cm(2) [P=0.014] versus placebo, 2.92±3.5 cm(2) to 2.89±4.1 cm(2) [P=0.5]) and reduced rest pain (5.2±1.8 to 2.2±1.3 [P=0.009] versus placebo, 4.5±2.4 to 3.9±2.6 [P=0.3]) within 3 months. Limb salvage and amputation-free survival rates did not differ between the groups. Repeated BM-MNC administration and higher BM-MNC numbers and functionality were the only independent predictors of improved ulcer healing. Ulcer healing induced by repeated BM-MNC administration significantly correlated with limb salvage (r=0.8; P<0.001). CONCLUSIONS: Intraarterial administration of BM-MNC is safe and feasible and accelerates wound healing in patients without extensive gangrene and impending amputation. These exploratory findings of this pilot trial need to be confirmed in a larger randomized trial in patients with critical limb ischemia and stable ulcers.

Intracoronary infusion of progenitor cells is not associated with aggravated restenosis development or atherosclerotic disease progression in patients with acute myocardial infarction.

AIMS: Experimental and clinical pilot studies suggest that intracoronary progenitor cell infusion can improve left ventricular function and remodelling after acute myocardial infarction (AMI). Since progenitor cells are also known to be involved in restenosis development and atherosclerosis progression, an increased restenosis rate may be a risk of intracoronary cell therapy. METHODS: We performed a retrospective study to compare quantitative angiographic measurements of the infarct target vessel in 83 patients with AMI treated with bare metal stent PCI (matched control) and in 83 patients receiving additional intracoronary progenitor cell infusion at a mean of 5 days post-AMI stent PCI and after 4 months. RESULTS: The late loss as a measure of neointima formation was similar between the control and the cell-treated group at follow-up (0.9+/-0.8 vs. 0.9+/-0.7 mm, P=0.9). Moreover, restenosis rate was comparable in both groups (35% control vs. 27% cell-treated group, P=0.2). Multivariable analysis excluded cell therapy as an independent significant predictor of increased late loss (P=0.4), whereas acute gain (P=0.012) and diabetes mellitus (P=0.002) were independent predictors of late loss. Finally, in the cell-treated group, target vessel revascularization rate remained at 28.9% during a median of >3 years of follow-up, thus excluding an effect on atherosclerotic disease progression. CONCLUSION: In patients with AMI successfully treated with bare metal stent PCI, additional intracoronary progenitor cell infusion does not lead to an increased neointima formation within the implanted stent within 4 months or aggravation of atherosclerotic disease progression.

Endothelial progenitor cells: regulation and contribution to adult neovascularization.

BACKGROUND: Recent studies suggest that circulating progenitor cells contribute to the formation of new blood vessels in adults after tissue ischemia. The infusion of these progenitor cells was used as a therapeutic approach to increase vascularization. In several animal models, progenitor cells improved vascularization and capillary density after peripheral or myocardial ischemia. Moreover, transplantation or progenitor cells increased cardiac function after myocardial ischemia. These studies suggest a potential use of progenitor cells for improvement of therapeutic vasculogenesis in patients with ischemic heart disease. PRESENT STATE: The present article will summarize these findings gained in experimental models. Moreover, novel approaches to increase the function and the number of circulating progenitor cells by pharmacological modulation (cytokines, statins) or gene therapy (VEGF) will be highlighted. CONCLUSION: Identification of mediators and cellular mechanisms that promote organ-specific recruitment of bone-marrow derived circulating progenitor cells as well as modulation of progenitor cell engraftment will lead to new strategies in order to improve neovascularization and cardiac regeneration in patients with ischemic heart disease.

Effects of statins on endothelium and endothelial progenitor cell recruitment.

Statins appear to be potent drugs with a variety of pleiotropic effects with vasculoprotective and cardioprotective activity. The beneficial effects of statins on endothelial cells as well as on endothelial cell function appear to be related to improved nitric oxide bioavailability. Mechanistically, statins induce endothelial nitric oxide synthase mRNA stability in endothelial cells and promote endothelial nitric oxide synthase activity through a PI3K/Akt dependent pathway, which is a common signal transduction pathway shared by growth factors such as vascular endothelial growth factors or fibroblast growth factors (FGFs), estrogens, or statins. Furthermore, statins have potent antiinflammatory capacities by potently interfering with the generation of reactive oxygen species or activating scavenging systems for free radicals such as the thioredoxin system. These mechanisms might all contribute to improved NO bioavailability and confer the beneficial actions of statins. The proangiogenic properties of statins and their effects on reendothelialization following vessel injury include novel actions such as the mobilization, differentiation, and improved survival of endothelial progenitor cells. Statin therapy might reverse the impaired functional regeneration capacities seen in patients with risk factors for coronary artery disease or documented active coronary artery disease by specifically interacting with progenitor cell function. Accordingly, augmentation of functionally active endothelial progenitor cells with improved homing capacity will be a critical step in advancing therapeutic neovascularization as well as reendothelialization in patients with coronary artery disease.