Increase In Capillary Density Research Articles

Non-invasive Assessment of Microvascular and Endothelial Function

The authors have utilized capillaroscopy and forearm blood flow techniques to investigate the role of microvascular dysfunction in pathogenesis of cardiovascular disease. Capillaroscopy is a non-invasive, relatively inexpensive methodology for directly visualizing the microcirculation. Percent capillary recruitment is assessed by dividing the increase in capillary density induced by postocclusive reactive hyperemia (postocclusive reactive hyperemia capillary density minus baseline capillary density), by the maximal capillary density (observed during passive venous occlusion). Percent perfused capillaries represents the proportion of all capillaries present that are perfused (functionally active), and is calculated by dividing postocclusive reactive hyperemia capillary density by the maximal capillary density. Both percent capillary recruitment and percent perfused capillaries reflect the number of functional capillaries. The forearm blood flow (FBF) technique provides accepted non-invasive measures of endothelial function: The ratio FBF(max)/FBF(base) is computed as an estimate of vasodilation, by dividing the mean of the four FBF(max) values by the mean of the four FBFbase values. Forearm vascular resistance at maximal vasodilation (FVR(max)) is calculated as the mean arterial pressure (MAP) divided by FBF(max). Both the capillaroscopy and forearm techniques are readily acceptable to patients and can be learned quickly. The microvascular and endothelial function measures obtained using the methodologies described in this paper may have future utility in clinical patient cardiovascular risk-reduction strategies. As we have published reports demonstrating that microvascular and endothelial dysfunction are found in initial stages of hypertension including prehypertension, microvascular and endothelial function measures may eventually aid in early identification, risk-stratification and prevention of end-stage vascular pathology, with its potentially fatal consequences.

Muscle-derived Stem Cell Sheets Support Pump Function and Prevent Cardiac Arrhythmias in a Model of Chronic Myocardial Infarction

Direct intracardiac cell injection for heart repair is hindered by numerous limitations including: cell death, poor spreading of the injected cells, arrhythmia, needle injury, etc. Tissue-engineered cell sheet implantation has the potential to overcome some of these limitations. We evaluated whether the transplantation of a muscle-derived stem cell (MDSC) sheet could improve the regenerative capacity of MDSCs in a chronic model of myocardial infarction. MDSC sheet-implanted mice displayed a reduction in left ventricle (LV) dilation and sustained LV contraction compared with the other groups. The MDSC sheet formed aligned myotubes and produced a significant increase in capillary density and a reduction of myocardial fibrosis compared with the other groups. Hearts transplanted with the MDSC sheets did not display any significant arrhythmias and the donor MDSC survival rate was higher than the direct myocardial MDSC injection group. MDSC sheet implantation yielded better functional recovery of chronic infarcted myocardium without any significant arrhythmic events compared with direct MDSC injection, suggesting this cell sheet delivery system could significantly improve the myocardial regenerative potential of the MDSCs.

VDAC: old protein with new roles in diabetes

A decrease in capillary density due to an increase in endothelial cell apoptosis in the heart is implicated in cardiac ischemia in diabetes. The voltage-dependent anion channel (VDAC) plays a crucial role in the regulation of mitochondrial metabolic function and mitochondria-mediated apoptosis. This study is designed to examine the role of VDAC in coronary endothelial dysfunction in diabetes. Endothelial cells (ECs) were more apoptotic in diabetic left ventricle of diabetic mice and mouse coronary ECs (MCECs) isolated from diabetic mice exhibited significantly higher mitochondrial Ca(2+) concentration and VDAC protein levels than control MCECs. The expression of VDAC-short hairpin RNA (shRNA) not only decreased the resting mitochondrial Ca(2+) concentration but also attenuated mitochondrial Ca(2+) uptake in diabetic MCECs. Furthermore, the downregulation of VDAC in diabetic MCECs significantly decreased mitochondrial superoxide anion (O(2)(-)) production and the activity of the mitochondrial permeability transition pore (mPTP) opening (an indirect indicator of cell apoptosis) toward control levels. These data suggest that the increased VDAC level in diabetic MCECs is responsible for increased mitochondrial Ca(2+) concentration, mitochondrial O(2)(-) production, and mPTP opening activity. Normalizing VDAC protein level may help to decrease endothelial cell apoptosis, increase capillary density in the heart, and subsequently decrease the incidence of cardiac ischemia in diabetes.

Role of simvastatin and/or antioxidant vitamins in therapeutic angiogenesis in experimental diabetic hindlimb ischemia: effects on capillary density, angiogenesis markers, and oxidative stress

Therapeutic angiogenesis has emerged as an attractive approach for the management of peripheral arterial disease in diabetic patients. Oxidative stress generated and aggravated by prolonged hyperglycemia may interfere with and destroy the newly formed blood vessels. Angiogenic effect of simvastatin has been reported; however, its exact mechanism is yet to be evaluated. In addition, the exact role of antioxidant vitamins in diabetic peripheral arterial disease is still controversial. The present study was undertaken to investigate the therapeutic potential of simvastatin and antioxidant vitamins (E and C) and their combined effects on angiogenesis in diabetic hind-limb ischemia. Streptozotocin diabetic rats were treated for 6 weeks with simvastatin either alone or in combination with vitamin E or vitamin C. Parameters of angiogenesis, nitric oxide, heme oxygenase-1 (HO-1), and oxidative stress markers were evaluated. CD31 immunostaining revealed an increased capillary density in ischemic gastrocnemious tissue of diabetic rats treated with either simvastatin or its combination with vitamin C. This effect was accompanied by up-regulated plasma levels of HO-1, nitric oxide, vascular endothelial growth factor (VEGF) and its intra-muscular receptor type-2 (Flk-1). Tissue reduced glutathione and antioxidant enzymes activities were normalized in groups treated with antioxidant vitamins or their combination with simvastatin with concomitant blunting of lipid peroxidation. Vitamins E and C, through their antioxidant effects, evidently enhanced the angiogenic effect of simvastatin in ischemic diabetic muscle. Hence, the use of antioxidant vitamins combined with statins to induce therapeutic angiogenesis is a promising strategy in the management of diabetes-associated peripheral arterial disease.

Can increases in capillarization explain the early adaptations in metabolic regulation in human muscle to short-term training?

To investigate the hypothesis that increases in fibre capillary density would precede increases in oxidative potential following training onset, tissue was extracted from the vastus lateralis prior to (0 days) and following 3 and 6 consecutive days of submaximal cycle exercise (2 h·day(-1)). Participants were untrained males (age = 21.4 ± 0.58 years; peak oxygen consumption = 46.2 ± 1.6 mL·kg(-1)·min(-1); mean ± standard error (SE)). Tissue was assessed for succinic dehydrogenase activity (SDH) by microphotometry and indices of capillarization based on histochemically assessed area and capillary counts (CC) in specific fibre types. Three days of training (n = 13) resulted in a generalized decrease (p < 0.05) in fibre area (-14.2% ± 3.0%; mean ± SE) and increase (p < 0.05) in CC/Area (20.4% ± 2.7%) and no change in either CC or SDH activity. Following 6 days of treatment (n = 6), increases (p < 0.05) in CC (18.2% ± 4.2%), CC/Area (28.9% ± 3.2%), and SDH activity (22.9% ± 6.0%) occurred that was not specific to major fibre type. No changes in either fibre area or fibre-type distribution were observed with additional training. We conclude that increases in angiogenic-based capillary density and oxidative potential occur coincidentally following training onset, while increases in capillary density, mediated by reductions in fibre area, represent an initial isolated response, the significance of which may be linked to the metabolic alterations that also result.

Aerobic Training (T) Improves Cerebral Circulation of Normotensive (WKY) and Spontaneously Hypertensive Rats (SHR)

We compared the effects of hypertension and T in brain macro and microcirculation. Rats were submitted to treadmill T (55% of maximum capacity, 1h/day, 5 d/week) or kept sedentary (S) for 3 months. After hemodynamic measurements at rest, rats were anesthetized for iv. administration of FITC‐dextran (capillary density) or 4% PFA perfusion (morphometry of cerebral arteries). Brains were removed, post‐fixed and sliced for analysis of the capillary density within the NTS (25 μm) and of the basilar (BA), middle (MCA) and posterior cerebral (PCA) arteries (10 μm) (Image ProPlus). Compared to WKY, SHR exhibited elevated MAP and HR (180±2 mmHg, 359±6 b/min), increased wall/lumen ratio (+40%, +100% and +80% for BA, MCA and PCA, respectively), but no NTS capillary rarefaction. T was equally effective to improve treadmill performance and to cause resting bradycardia (−9%) in both groups; pressure fall (−8%) was only observed in the SHR. T reduced wall/lumen ratio only in the SHR (BA= −40%, MCA= −47%; PCA= −37%) and caused marked increase in capillary density in medial, intermediate and commissural NTS in both groups (on average 7.05 ±0.18 and 7.07±0.88%, corresponding to changes of +55% and +43%, WKY and SHR, respectively). Data showed that T improves brain perfusion by causing eutrophic outward remodeling in SHR brain arteries and capillary angiogenesis in both groups. Financial Support: CAPES, FAPESP

Enhanced external counterpulsation (EECP) increases GLUT‐4 protein expression, capillary density and glucose tolerance in patients with abnormal glucose tolerance

We sought to evaluate the effects of EECP therapy on outcomes of glucose tolerance and skeletal muscle morphology in patients with abnormal glucose tolerance (AGT).MethodsEighteen patients with AGT were randomly assigned (2:1 ratio) to receive either 7 weeks (35 1‐hour sessions) of EECP (n = 12) or 7‐weeks of standard care (control; n = 6). Biochemical assays, oral glucose tolerance, and skeletal muscle morphology were assessed before and after EECP or control.ResultsFollowing EECP, plasma vascular endothelial growth factor (VEGF) was increased (9.74 ± 1.59 to 17.04 ± 2.65 pg/mL, P&lt;0.05). Vastus lateralis skeletal muscle biopsies demonstrated improvements in capillary density (454 ± 21 to 497 ± 19 mm−2, P&lt;0.05) following EECP. There was no change in glucose uptake signaling pathways, but total GLUT‐4 protein expression was elevated (+47%, P&lt;0.05) following EECP. Fasting (−16.9 ± 5.4 mg/dL) and 120‐min (−28.3 ± 7.3 mg/dL) plasma glucose from oral glucose tolerance testing were also improved following EECP (P&lt;0.05). Further, the Matsuda index, a whole‐body composite insulin sensitivity index, increased (+21%, P&lt;0.05).ConclusionsEECP mediated increases in capillary density and GLUT‐4 in patients with AGT paralleled improvements in surrogates of insulin sensitivity and glucose tolerance.This research was supported by the Department of Applied Physiology and Kinesiology at the University of Florida.

Microcirculation and left-ventricular hypertrophy

Microcirculation and left-ventricular hypertrophy

Periodontal microcirculation in diabetics: An in vivo non-invasive analysis by means of videocapillaroscopy

SummaryBackgroundDiabetes mellitus is today considered a society-wide disease of a chronic/degenerative nature. Among the secondary effects of diabetes, the one that interests the dental surgeon most is diabetic parodontopathy. The aim of this study was to underline and objectify microcirculatory variations at a periodontal mucous level in type 2 diabetics.Material/MethodsThe study enrolled 80 subjects: 40 subjects with a diagnosis of diabetes mellitus type II (18 males and 22 females, between 44 and 85 years of age); and 40 healthy subjects (17 males and 23 females, between 44 and 78 years of age). All the subjects, both diabetic and healthy, were submitted to a videocapillaroscopic examination of the mucosa of the oral cavity.ResultsThe measurements concerning the density (expressed in the number of loops/mm2) of the capillary loops presented differences between the healthy subjects and the diabetic subjects. The average periodontal capillary density (DC-P) was clearly superior in diabetic subjects (35.62±10.40 n°loop/mm2) compared to healthy subjects (17.55±3.88 n°loop/mm2). The statistical analysis was performed by means of the Mann Whitney test. The value of P (p=0.000000986), well below the level of significance, demonstrates the high significance of the results obtained.ConclusionsThe increase in capillary density could suggest the presence of active inflammatory phenomena or, more probably, a tendency to a greater susceptibility to inflammatory phenomena. Ultimately, this study shows that there is some peripheral damage to microcirculation at the masticatory mucous level in diabetic subjects and that such alterations can be instrumentally objectified and quantified through the videocapillaroscopic method.

Angiopoietin-1-expressing adipose stem cells genetically modified with baculovirus nanocomplex: investigation in rat heart with acute infarction.

The objective of this study was to develop angiopoietin-1 (Ang1)-expressing genetically modified human adipose tissue derived stem cells (hASCs) for myocardial therapy. For this, an efficient gene delivery system using recombinant baculovirus complexed with cell penetrating transactivating transcriptional activator TAT peptide/deoxyribonucleic acid nanoparticles (Bac-NP), through ionic interactions, was used. It was hypothesized that the hybrid Bac- NPAng1 system can efficiently transduce hASCs and induces favorable therapeutic effects when transplanted in vivo. To evaluate this hypothesis, a rat model with acute myocardial infarction and intramyocardially transplanted Ang1-expressing hASCs (hASC-Ang1), genetically modified by Bac-NPAng1, was used. Ang1 is a crucial pro-angiogenic factor for vascular maturation and neovasculogenesis. The released hAng1 from hASC-Ang1 demonstrated profound mitotic and anti-apoptotic activities on endothelial cells and cardiomyocytes. The transplanted hASC-Ang1 group showed higher cell retention compared to hASC and control groups. A significant increase in capillary density and reduction in infarct sizes were noted in the infarcted hearts with hASC-Ang1 treatment compared to infarcted hearts treated with hASC or the untreated group. Furthermore, the hASC-Ang1 group showed significantly higher cardiac performance in echocardiography (ejection fraction 46.28% ± 6.3%, P < 0.001 versus control, n = 8) than the hASC group (36.35% ± 5.7%, P < 0.01, n = 8), 28 days post-infarction. The study identified Bac-NP complex as an advanced gene delivery vehicle for stem cells and demonstrated its potential to treat ischemic heart disease with high therapeutic index for combined stem cell-gene therapy strategy.

Akt1–Mediated Skeletal Muscle Growth Attenuates Cardiac Dysfunction and Remodeling After Experimental Myocardial Infarction

It is appreciated that aerobic endurance exercise can attenuate unfavorable myocardial remodeling following myocardial infarction. In contrast, little is known about the effects of increasing skeletal muscle mass, typically achieved through resistance training, on this process. Here, we utilized transgenic (TG) mice that can induce the growth of functional skeletal muscle by switching Akt1 signaling in muscle fibers to assess the impact of glycolytic muscle growth on post-myocardial infarction cardiac remodeling. Male-noninduced TG mice and their nontransgenic littermates (control) were subjected to left anterior coronary artery ligation. Two days after surgery, mice were provided doxycycline in their drinking water to activate Akt1 transgene expression in a skeletal muscle-specific manner. Myogenic Akt1 activation led to diminished left ventricular dilation and reduced contractile dysfunction compared with control mice. Improved cardiac function in Akt1 TG mice was coupled to diminished myocyte hypertrophy, decreased interstitial fibrosis, and increased capillary density. ELISA and protein array analyses demonstrated that serum levels of proangiogenic growth factors were upregulated in Akt1 TG mice compared with control mice. Cardiac eNOS was activated in Akt1 TG mice after myocardial infarction. The protective effect of skeletal muscle Akt activation on cardiac remodeling and systolic function was abolished by treatment with the eNOS inhibitor l-NAME. Akt1-mediated skeletal muscle growth attenuates cardiac remodeling after myocardial infarction and is associated with an increased capillary density in the heart. This improvement appears to be mediated by skeletal muscle to cardiac communication, leading to activation of eNOS-signaling in the heart.

Adipose-derived stem cells accelerate neovascularization in ischaemic diabetic skin flap via expression of hypoxia-inducible factor-1α

Skin flaps are frequently performed for diabetic patients in spite of countless detrimental effects of diabetes on flap survival, most of which may result from a defective response of the tissues to low oxygen tension. In this study, the authors explored the feasibility of applying human adipose-derived stem cells (ASCs) to increase the viability of random-patterned skin flaps in streptozotocin-induced diabetic mice. ASCs were isolated from the fresh human lipoaspirates and expanded ex vivo for three passages. After the elevation of caudally based random-patterned skin flaps (3 cm long and 1 cm wide), ASCs suspensions were then injected into the flap (group A). Media containing no ASCs were similarly injected as a control (group B), although nothing was injected into the flap base of mice in control group C. Flap assessments were carried out at post-operative day 7 for evaluation of flap viability. The flap survival rate of group A was significantly higher than those of groups B and C, whereas no difference was observed between groups B and C. Histological examination also demonstrated a statistically significant increase in capillary density in group A over both groups B and C. Furthermore, it was found that ASCs not only augmented the expression of vascular endothelial growth factor and hypoxia-inducible factor-1α (HIF-1α) in flap tissues from dermis of diabetes mice, but also promoted their expression in dermal fibroblasts from diabetic mice. Thus, ASCs could enhance the survival of random-patterned skin flaps in streptozotocin-induced diabetic mice via elevated expression of HIF-1α.

Abstract 9481: Whole Body Periodic Acceleration, Novel Passive Exercise Devices, Enhances Blood Supply to Ischemic Hindlimb in Mice and Human

Introduction: Whole body periodic acceleration (WBPA) is a non-invasive, passive exercise device that has been shown to improve endothelial function by increasing shear stress by head-to-foot direction movements of the body. However, the impact of WBPA on blood flow to ischemic legs has not been investigated. Hypothesis: WBPA enhances blood supply in a mouse model of hindlimb ischemia and in patients with peripheral arterial disease (PAD). Methods and Results: After unilateral femoral artery excision, wild type mice were assigned to either the WBPA (n=15) or control (n=13) group. WBPA was applied at 150 cpm for 45 min under anesthesia once a day. WBPA significantly increased blood flow recovery at 7 and 14 days after surgery as determined by laser doppler perfusion imaging. Sections of ischemic adductor muscle stained with anti-CD31 antibody showed a significant increase in capillary density in WBPA-treated mice compared with control mice, suggesting that WBPA promotes angiogenesis at a microcirculatory level. WBPA increased the phosphorylation of endothelial nitric oxide synthase (eNOS) in adductor muscle. The pro-angiogenic effect of WBPA on ischemic limb was blunted in eNOS-deficient mice, suggesting that the stimulatory effects of WBPA on revascularization are eNOS-dependent. Quantitative real-time PCR analysis showed significant increases in molecules involved in angiogenesis, such as VEGF, FGF2, SDF-1, PDGF-B in ischemic hindlimb by WBPA. Protein array analysis showed that WBPA induced changes in the expression profile of serum angiogenesis-related proteins. Facilitated blood flow recovery was observed in a mouse model of diabetes despite no changes in glycemic control. Clinically, 10 patients (age, 69±7 years, 9 men) with PAD underwent single session of WBPA treatment at 140 cpm for 45 min. WBPA significantly improved the ischemic/non-ischemic ratio of blood flow in the lower extremity from 0.74±0.21 to 0.87±0.21 (p&lt;0.05). Conclusions: WBPA increased blood supply to ischemic lower extremities through activation of eNOS signaling and upregulation of pro-angiogenic growth factor in ischemic skeletal muscle. WBPA is a potentially suitable non-invasive intervention to facilitate therapeutic angiogenesis.

Endothelial Cells Derived From Human iPSCS Increase Capillary Density and Improve Perfusion in a Mouse Model of Peripheral Arterial Disease

Stem cell therapy for angiogenesis and vascular regeneration has been investigated using adult or embryonic stem cells. In the present study, we investigated the potential of endothelial cells (ECs) derived from human induced pluripotent stem cells (hiPSCs) to promote the perfusion of ischemic tissue in a murine model of peripheral arterial disease. Endothelial differentiation was initiated by culturing hiPSCs for 14 days in differentiation media supplemented with BMP-4 and vascular endothelial growth factor. The hiPSC-ECs exhibited endothelial characteristics by forming capillary-like structures in matrigel and incorporating acetylated-LDL. They stained positively for EC markers such as KDR, CD31, CD144, and eNOS. In vitro exposure of hiPSC-ECs to hypoxia resulted in increased expression of various angiogenic related cytokines and growth factors. hiPSC-ECs were stably transduced with a double fusion construct encoded by the ubiquitin promoter, firefly luciferase for bioluminescence imaging and green fluorescence protein for fluorescent detection. The hiPSC-ECs (5×10(5)) were delivered by intramuscular injection into the ischemic hindlimb of SCID mice at day 0 and again on day 7 after femoral artery ligation (n=8). Bioluminescence imaging showed that hiPSC-ECs survived in the ischemic limb for at least 2 weeks. In addition, laser Doppler imaging showed that the ratio of blood perfusion was increased by hiPSC-EC treatment by comparison to the saline-treated group (0.58±0.12 versus 0.44±0.04; P=0.005). The total number of capillaries in the ischemic limb of mice receiving hiPSC-EC injections was greater than those in the saline-treated group (1284±155 versus 797±206 capillaries/mm(2)) (P<0.002). This study is a first step toward development of a regenerative strategy for peripheral arterial disease based on the use of ECs derived from hiPSCs.

Passive Exercise Using Whole-Body Periodic Acceleration Enhances Blood Supply to Ischemic Hindlimb

Whole-body periodic acceleration (WBPA) has been developed as a passive exercise technique to improve endothelial function by increasing shear stress through repetitive movements in spinal axis direction. We investigated the effects of WBPA on blood flow recovery in a mouse model of hindlimb ischemia and in patients with peripheral arterial disease. After unilateral femoral artery excision, mice were assigned to either the WBPA (n=15) or the control (n=13) group. WBPA was applied at 150 cpm for 45 minutes under anesthesia once a day. WBPA significantly increased blood flow recovery after ischemic surgery, as determined by laser Doppler perfusion imaging. Sections of ischemic adductor muscle stained with anti-CD31 antibody showed a significant increase in capillary density in WBPA mice compared with control mice. WBPA increased the phosphorylation of endothelial nitric oxide synthase (eNOS) in skeletal muscle. The proangiogenic effect of WBPA on ischemic limb was blunted in eNOS-deficient mice, suggesting that the stimulatory effects of WBPA on revascularization are eNOS dependent. Quantitative real-time polymerase chain reaction analysis showed significant increases in angiogenic growth factor expression in ischemic hindlimb by WBPA. Facilitated blood flow recovery was observed in a mouse model of diabetes despite there being no changes in glucose tolerance and insulin sensitivity. Furthermore, both a single session and 7-day repeated sessions of WBPA significantly improved blood flow in the lower extremity of patients with peripheral arterial disease. WBPA increased blood supply to ischemic lower extremities through activation of eNOS signaling and upregulation of proangiogenic growth factor in ischemic skeletal muscle. WBPA is a potentially suitable noninvasive intervention to facilitate therapeutic angiogenesis.

A Novel Approach for Studying Microcirculation in Bone Defects by Intravital Fluorescence Microscopy

Angiogenic and inflammatory responses to biodegradable scaffolds were previously studied using the dorsal skinfold chamber for testing different scaffold materials. In this model, the angiogenic response originates from the soft tissue of the skin. Herein, we introduce a new model that allows the study of developing microcirculation of bone defects for testing tissue-engineered constructs. A bone defect was prepared in the femur of Balb/c mice by inserting a pin for intramedullary fixation, and a custom-made observation window fixed over the defect allowed constant observation. This study included three different groups: empty defect (control), defect filled with porous poly(L-lactide-co-glycolide), and beta-tricalcium-phosphate scaffolds. Starting from 6 days after surgery, angiogenesis, neovascularization, leukocyte-endothelial cell interaction, and microvascular permeability were analyzed over 22 days by using intravital fluorescence microscopy. The empty defects showed no signs of angiogenesis during the observation period, but a distinct increase of capillary density was detected in the scaffold-containing defects. Surprisingly, the histological sections of the scaffold-treated defects showed new bone formation 22 days after implantation. We present a new bone chamber model for intravital long-term study of scaffold materials suitable for bone reconstruction in mice by using fluorescence microscopy.

A nanobiohybrid complex of recombinant baculovirus and Tat/DNA nanoparticles for delivery of Ang-1 transgene in myocardial infarction therapy

The study aims to design a new gene delivery method utilizing the complementary strengths of baculovirus, such as relatively high transduction efficiency and easy scale-up, and non-viral nanodelivery systems, such as low immunogenicity. This formulation was developed by generating a self assembled binary complex of negatively charged baculovirus (Bac) and positively charged endosomolytic histidine rich Tat peptide/DNA nanoparticles (NP). The synergistic effect of this hybrid (Bac-NP) system to induce myocardial angiogenesis in acute myocardial infarction (AMI) model has been explored in this study, using Angiopoietin-1 (Ang-1) as the transgene carried by both vector components. Under optimal transduction conditions, Bac-NP Ang1 showed 1.75 times higher and sustained Ang-1 expression in cardiomyocytes than Bac Ang1, with significantly high angiogenic potential as confirmed by functional assays. For in vivo analysis, we intramyocardially delivered Bac-NP Ang1 to AMI rat model. 3 weeks post AMI, data showed increase in capillary density ( p < 0.01) and reduction in infarct sizes ( p < 0.05) in Bac-NP Ang1 compared to Bac Ang1, NP Ang1 and control groups due to enhanced myocardial Ang-1 expression at peri-infarct regions (1.65 times higher than Bac Ang1). Furthermore, the Bac-NP Ang1 group showed significantly higher cardiac performance in echocardiography than Bac Ang1 (44.2 ± 4.77% vs 37.46 ± 5.2%, p < 0.01), NP Ang1 and the control group (32.26 ± 2.49% and 31.58 ± 2.26%). Collectively, this data demonstrates hybrid Bac-NP as a new and improved gene delivery system for therapeutic applications.

Cardioprotection of Insulin-Like Growth Factor-1 During Reperfusion Therapy

In the setting of acute myocardial infarction, reperfusion of ischemic myocardium carried out early after coronary occlusion can salvage reversibly injured, viable myocardium. Although still controversial, however, reperfusion itself may cause a population of reversibly injured cells to die, a phenomenon termed lethal reperfusion injury .1 A variety of pharmacological agents have been studied to limit ischemia/reperfusion injury as an adjunct to current reperfusion, and some beneficial effects have been demonstrated in experimental animal research, but their clinical applications have not been achieved.2–4 Article see p 327 Among the various agents claimed to have cardioprotective effects is insulin-like growth factor-1 (IGF-1), which is a hormone produced primarily by the liver. Its molecular structure is similar to that of insulin. IGF-1 plays an important role in cellular survival and growth by binding to its specific receptor (IGF1R), which is present on many cell types, including cardiac cells. Activation of IGF1R, a receptor tyrosine kinase, has been demonstrated to attenuate both apoptotic and necrotic cell death induced by ischemia/reperfusion injury through stimulating both the intracellular phosphoinositide 3-kinase/protein kinase B signaling pathway and extracellular signal-regulated kinase 1/2 signaling cascades.5 Buerke at al6 administered IGF-I 1 hour before ischemia in a murine model of 20 minutes of myocardial ischemia followed by 24 hours of reperfusion. IGF-I preserved ischemic/reperfused myocardium through inhibition of polymorphonuclear leukocyte-induced cardiac necrosis and inhibition of reperfusion-induced apoptosis of cardiac myocytes. Davani et al7 subjected isolated murine hearts to 20 minutes of global ischemia followed by 2 hours of reperfusion with either modified Kreb's solution alone or …

Placental Growth Factor Regulates Cardiac Adaptation and Hypertrophy Through a Paracrine Mechanism

Paracrine growth factor-mediated crosstalk between cardiac myocytes and nonmyocytes in the heart is critical for programming adaptive cardiac hypertrophy in which myocyte size, capillary density, and the extracellular matrix function coordinately. To examine the role that placental growth factor (PGF) plays in the heart as a paracrine regulator of cardiac adaptation to stress stimulation. PGF is induced in the heart after pressure-overload stimulation, where it is expressed in both myocytes and nonmyocytes. We generated cardiac-specific and adult inducible PGF-overexpressing transgenic mice and analyzed Pgf(-/-) mice to examine the role that this factor plays in cardiac disease and paracrine signaling. Although PGF transgenic mice did not have a baseline phenotype or a change in capillary density, they did exhibit a greater cardiac hypertrophic response, a greater increase in capillary density, and increased fibroblast content in the heart in response to pressure-overload stimulation. PGF transgenic mice showed a more adaptive type of cardiac growth that was protective against signs of failure with pressure overload and neuroendocrine stimulation. Antithetically, Pgf(-/-) mice rapidly died of heart failure within 1 week of pressure overload, they showed an inability to upregulate angiogenesis, and they showed significantly less fibroblast activity in the heart. Mechanistically, we show that PGF does not have a direct effect on cardiomyocytes but works through endothelial cells and fibroblasts by inducing capillary growth and fibroblast proliferation, which secondarily support greater cardiac hypertrophy through intermediate paracrine growth factors such as interleukin-6. PGF is a secreted factor that supports hypertrophy and cardiac function during pressure overload by affecting endothelial cells and fibroblasts that in turn stimulate and support the myocytes through additional paracrine factors.

Vascular Gene Transfer of SDF-1 Promotes Endothelial Progenitor Cell Engraftment and Enhances Angiogenesis in Ischemic Muscle

Gene therapy approaches to enhance endothelial progenitor cell (EPC) homing may augment cell engraftment to ischemic tissue and lead to a greater therapeutic response. Therefore, we assessed the effects of ultrasound-mediated (UM) transfection of the chemokine stromal cell-derived factor-1 (SDF-1) on homing and engraftment of intravenously administered EPCs and the subsequent angiogenic response in chronically ischemic skeletal muscle. Bone marrow-derived EPCs were isolated from donor Fisher 344 rats, cultured and labeled in preparation for injection into recipient animals via a jugular vein. Using a model of chronic hindlimb ischemia in rats, we demonstrated that UM destruction of intravenous carrier microbubbles loaded with SDF-1 plasmid DNA resulted in targeted transfection of the vascular endothelium within ischemic muscle and greater local engraftment of EPCs. The combination of SDF-1gene therapy and EPCs lead to the greatest increase in tissue perfusion and microvascular density within ischemic muscle, compared to no treatment or either monotherapy alone. Our results demonstrate that UM transfection of SDF-1 improves EPC targeting to chronically ischemic tissue, enhancing vascular engraftment and leading to a more robust neovascularization response.