Ang-1 Group Research Articles

C16 Peptide and Ang-1 Improve Functional Disability and Pathological Changes in an Alzheimer's Disease Model Associated with Vascular Dysfunction.

Alzheimer’s disease (AD) is a neurological disorder characterized by neuronal cell death, tau pathology, and excessive inflammatory responses. Several vascular risk factors contribute to damage of the blood–brain barrier (BBB), secondary leak-out of blood vessels, and infiltration of inflammatory cells, which aggravate the functional disability and pathological changes in AD. Growth factor angiopoietin-1 (Ang-1) can stabilize the endothelium and reduce endothelial permeability by binding to receptor tyrosine kinase 2 (Tie2). C16 peptide (KAFDITYVRLKF) selectively binds to integrin ανβ3 and competitively inhibits leukocyte transmigration into the central nervous system by interfering with leukocyte ligands. In the present study, 45 male Sprague-Dawley (SD) rats were randomly divided into three groups: vehicle group, C16 peptide + Ang1 (C + A) group, and sham control group. The vehicle and C + A groups were subjected to two-vessel occlusion (2-VO) with artery ligation followed by Aβ1-42 injection into the hippocampus. The sham control group underwent sham surgery and injection with an equal amount of phosphate-buffered saline (PBS) instead of Aβ1-42. The C + A group was administered 1 mL of drug containing 2 mg of C16 and 400 µg of Ang-1 daily for 2 weeks. The sham control and vehicle groups were administered 1 mL of PBS for 2 weeks. Our results showed that treatment with Ang-1 plus C16 improved functional disability and reduced neuronal death by inhibiting inflammatory cell infiltration, protecting vascular endothelial cells, and maintaining BBB permeability. The results suggest that these compounds may be potential therapeutic agents for AD and warrant further investigation.

Prolonged release of VEGF and Ang1 from intralesionally implanted hydrogel promotes perilesional vascularization and functional recovery after experimental ischemic stroke

Injectable hydrogels can generate and support pro-repair environments in injured tissue. Here we used a slow-releasing drug carrying in situ-forming hydrogel to promote post-stroke recovery in a rat model. Release kinetics were measured in vitro and in vivo with MRI, using gadolinium-labeled albumin (Galbumin), which demonstrated prolonged release over multiple weeks. Subsequently, this hydrogel was used for long-term delivery of vascular endothelial growth factor (VEGF) and angiopoietin-1 (Ang1) (Gel VEGF + Ang1, n = 14), in a photothrombotically induced cortical stroke lesion in rats. Control stroke animals were intralesionally injected with saline (Saline, n = 10), non-loaded gel (Gel, n = 10), or a single bolus of VEGF + Ang1 in saline (Saline VEGF + Ang1, n = 10). MRI was executed to guide hydrogel injection. Functional recovery was assessed with sensorimotor function tests, while tissue status and vascularization were monitored by serial in vivo MRI. Significant recovery from sensorimotor deficits from day 28 onwards was only measured in the Gel VEGF + Ang1 group. This was accompanied by significantly increased vascularization in the perilesional cortex. Histology confirmed (re)vascularization and neuronal sparing in perilesional areas. In conclusion, intralesional injection of in situ-forming hydrogel loaded with pro-angiogenic factors can support prolonged brain tissue regeneration and promote functional recovery in the chronic phase post-stroke.

Soluble Tei2 fusion protein inhibits retinopathy of prematurity occurrence via regulation of the Ang/Tie2 pathway.

The aim of the present study was to investigate the potential mechanism of retinopathy of prematurity (ROP) using an oxygen-induced retinopathy (OIR) mouse model. For experiments, mice were divided into either the OIR group or control group. Fluorescein isothiocyanate-dextran cardiac perfusion and stretched retina preparation were performed. The total retina area, area of instillation, density of microvascular network, area of new blood vessels, vein width and the tortuosity of arteries were measured. Next, mice were randomly assigned into the PBS, soluble TEK receptor tyrosine kinase (sTie2)-fusion protein (Fc), angiopoietin 1 (Ang1), ranibizumab, ranibizumab + sTie2-Fc and ranibizumab + Ang1 treatment groups. Following housing for 5 days, the body weight of each mouse was recorded. Mice in the OIR group presented smaller total retina area and larger area of instillation, larger area of new blood vessels, and higher microvascular network density compared with the control PBS group. Obvious retinal vein dilatation and arterial tortuosity were identified in the OIR group. The amount of endotheliocyte nuclei of new vessels beyond the inner limiting membrane was larger in the OIR group compared with the control group. Furthermore in the next set of experiments, a larger area of instillation, smaller area of new blood vessels and decreased amount of endotheliocyte nuclei of new vessels were observed in the sTie2-Fc group, Ang1 group, ranibizumab group, ranibizumab + sTie2-Fc group and ranibizumab + Ang1 group compared with the PBS group. Specifically, the ranibizumab + sTie2-Fc group and ranibizumab + Ang1 group demonstrated markedly reduced retina instillation area and microvascular network density in the instillation area. Total retina area and body weight following 10 days of the experiment for the ranibizumab group were significantly lower compared with other groups. In conclusion, the combined regulation of the Ang/Tie2 and the vascular endothelial growth factor (VEGF)/VEGF receptor pathways markedly increased the efficacy of treatment with retinal neovascularization (RNV). Regulation of these pathways has a potential for treating RNV, in particular ROP.

Synergistic Therapeutic Effect of Three-Dimensional Stem Cell Clusters and Angiopoietin-1 on Promoting Vascular Regeneration in Ischemic Region.

Peripheral artery disease (PAD) is an ischemic disease characterized by reduced blood flow to the legs, feet, and hands. Human mesenchymal stem cells are an attractive cell source to treat PAD in regenerative medicine. However, in clinical applications, the use of adult stem cells has several limitations, such as low cell viability and low therapeutic efficiency. In this study, we described an innovative method of culturing three-dimensional stem cell clusters (Angiocluster™ [AC]), demonstrated the potential for ACs to differentiate into vascular cells, and assessed the synergistic effects of ACs and angiopoietin-1 (Ang-1) on angiogenesis in ischemic animal models. ACs were formed by culturing human adipose-derived stem cells (hASCs) on a maltose-binding protein-linked basic fibroblast growth factor-immobilized polystyrene surface. ACs released various angiogenic factors, such as vascular endothelial growth factor and interleukin-8, and could differentiate into endothelial lineage cells. However, ACs did not secrete Ang-1, which is an essential component of vascular maturation and anti-inflammation. ACs were combined with Ang-1 and were transplanted into the ischemic lesions of mice for 28 days. Most of the mice receiving the AC + Ang-1 treatment exhibited limb salvage and exhibited similar blood perfusion ratio compared to normal limb. The combination therapy of AC and Ang-1 enhanced angiogenic efficacy by increasing blood vessel regeneration and facilitating the implantation of stem cells into host vessels. Importantly, fibrotic collagen was observed in most of the groups after 28 days of treatment, except for the AC + Ang-1 group. This indicates that the combination therapy is synergistic in minimizing ischemic fibrosis and muscle degeneration. Our results demonstrate that the combination therapy significantly enhanced tissue regeneration and angiogenic efficacy of hASCs and may have wide applications in regenerative medicine.

Dachengqi Decoction Attenuates Intestinal Vascular Endothelial Injury in Severe Acute Pancreatitis in Vitro and in Vivo

Background/Aims: Dachengqi decoction (DCQD) is a well-known traditional Chinese herbal drug with strong anti-inflammatory effects. Angiopoietin-1 (Ang-1) plays a vital role in maintaining the stability and integrity of the vascular wall and prevents vascular leakage due to inflammatory mediators. Our previous work found that DCQD protects against pancreatic injury in rats with severe acute pancreatitis (SAP). This study aims to investigate the effects of DCQD on intestinal endothelial damage in both damaged human umbilical vein endothelial cells (HUVECs) and SAP rats. Methods: HUVECs were randomly divided into four groups: control group, TNF-α group, TNF-α plus Ang-1 group (Ang-1 group), and TNF-α plus DCQD group (DCQD group). Cells were incubated for 6 h, 12 h, and 24 h, before collection. The treatment concentration of DCQD was decided based on a Cell Counting Kit-8 (CCK-8) assay. The monolayer permeability of the HUVECs was assessed by measuring the transendothelial electrical resistance (TEER). Apoptosis was analyzed by flow cytometry. mRNA and protein expression of aquaporin 1 (AQP-1), matrix metalloproteinase 9 (MMP9), and junctional adhesion molecule-C (JAM-C) was evaluated by RT-PCR, immunocytofluorescence, and western blot. Forty male Sprague-Dawley rats were randomized into a control group, SAP group, SAP plus Ang-1 group (Ang-1 group), and SAP plus DCQD group (DCQD group). SAP was induced by intraperitoneal injection of cerulein and lipopolysaccharide (LPS), while the control group received 0.9% saline solution. Evans blue was injected through the penile vein and the rats were then sacrificed 12 h after modeling. Levels of serum amylase, TNF-α, IL-1β, IL-2, and IL-6 were determined by using ELISA. Intestinal tissue was analysed by histology, and capillary permeability in the tissues was evaluated by Evans blue extravasation assay. Protein and mRNA expression of AQP-1, MMP9, and JAM-C were assessed by immunohistofluorescence, western blot, and RT-PCR. Results: DCQD reduced the permeability of HUVEC induced by TNF-α in vitro. Furthermore, DCQD altered the mRNA and protein levels of JAM-C, MMP9, and AQP-1 in HUVECs after TNF-α induction. SAP intestinal injury induced by cerulein combined with lipopolysaccharides was concomitant with increased expression of JAM-C and MMP9, and reduced expression of AQP-1 in intestinal tissue. Pretreatment with DCQD attenuated SAP intestinal injury and lowered the levels of serum amylase, TNF–α, IL-1β, IL-2, and IL-6 effectively. Our study demonstrated that DCQD decreased the expression of JAM-C and MMP9 and increased the expression of AQP-1 both in vitro and in vivo. Conclusion: DCQD can reduce capillary endothelial damage in acute pancreatitis-associated intestinal injury and the mechanism may be associated with the regulation of endothelial barrier function-associated proteins AQP-1, MMP9, and JAM-C.

Cartilage Oligomeric Matrix Protein-Angiopoietin-1 Has a Protective Effect of Vascular Endothelial Barrier in Rat With Acute Necrotizing Pancreatitis

To investigate the protective effect of angiopoietin-1 (Ang-1) from capillary endothelial damage in rats with acute necrotizing pancreatitis (ANP). 96 male Sprague-Dawley rats were randomly averaged and divided into control group, ANP group, Si-Ang-1 group, and COMP (cartilage oligomeric matrix protein)-Ang-1 group. Animals were killed at 6, 12, and 24 hours after molding. Levels of serum amylase, porcine endothelin 1, C-reactive protein, and Ang-1 were detected; histopathological changes in the pancreas were observed; capillary permeability and Ang-1 expression of the pancreatic tissue were detected by Evans Blue extravasation assay, immunohistochemistry, Western blot, and quantitative polymerase chain reaction. (1) Levels of serum amylase, C-reactive protein, and porcine endothelin-1 increased and level of Ang-1 decrease in the ANP group and Si-Ang-1 group compared with the control group, whereas COMP-Ang-1 group could improve the changes. (2) The order of pancreas pathological changes (mild to severe) is: control group, COMP-Ang-1 group, ANP group, and Si-Ang-1 group. (3) Capillary permeability of the pancreatic tissue in the COMP-Ang-1 group was lower than that in the ANP group. (4) Ang-1 mRNA and protein expression in the COMP-Ang-1 group was significantly higher than in the ANP group. COMP-Ang-1 can upregulate the expression of Ang-1 protein to promote angiogenesis and improve early inflammatory and pathological damage in ANP group.

Pretreatment with docosahexaenoic acid enhances the protective effect of angiopoietin-1 on rat brain microvascular endothelial cells during oxygen glucose deprivation

Objective To evaluate the role of pretreatment with docosahexaenoic acid (DHA) in the protective effect of angiopoietin-1 (Ang-1) on rat brain microvascular endothelial cells (BMVECs) during oxygen glucose deprivation (OGD). Methods BMVECs were sub-cultured in vitro and divided with a random number table into 7 groups: normal control group, normal control+ Ang-1 group, OGD group, OGD+ Ang-1 group, OGD+ DHA group, OGD+ DHA+ Ang-1 group, and OGD+ DHA+ GW9662+ Ang-1 group. The normal control and normal control+ Ang-1 groups were cultured in DMEM containing serum, 5 mmol/L glucose, and 1.25 mmol/L pyruvate; OGD groups were cultured in glucose- and serum-free DMEM. DHA (40 μmol/L) was added to OGD+ DHA, OGD+ DHA+ Ang-1, and OGD+ DHA+ GW9662+ Ang-1 groups, and 5 μmol/L GW9662 [inhibitor of peroxisome proliferator-activated receptor gamma (PPAR-γ)] to OGD+ DHA+ GW9662+ Ang-1 group, before pretreatment for 1 hour in 5% CO2 and 95% air. After the pretreatment, Ang-1 (250 ng/ml) was added to normal control+ Ang-1, OGD+ Ang-1, OGD+ DHA+ Ang-1, and OGD+ DHA+ GW9662+ Ang-1 groups. The cells were cultured in 94% N2∶5% CO2∶1%O2 for 24 hours, except for normal control and normal control+ Ang-1 groups, which were cultured in 5% CO2 and 95% air instead. The cell apoptosis rate was detected with flow cytometry, expressions of Bax, Bcl-2, caspase-3, Tie-1, Tie-2, pTie-2, pAkt, ZO-1 proteins with Western blot. Results Compared with normal control group, the cell apoptosis rate in OGD, OGD+ Ang-1, OGD+ DHA, OGD+ DHA+ Ang-1, and OGD+ DHA+ GW9662+ Ang-1 groups were significantly increased (P=0.000, 0.000, 0.000, 0.004, 0.000); the expression levels of Bax, caspase-3, and Tie-1 were significantly enhanced (Bax: 0.62±0.03, 0.38±0.03, 0.45±0.03, 0.26±0.02, 0.33±0.02 vs. 0.16±0.01; caspase-3: 0.76±0.05, 0.42±0.04, 0.52±0.02, 0.32±0.02, 0.40±0.02 vs. 0.15±0.01; Tie-1: 0.51±0.03, 0.25±0.01, 0.33±0.02, 0.16±0.01, 0.22±0.02 vs. 0.12±0.01; all P=0.000); the expression levels of Bcl-2, Bcl-2/Bax, Tie-2, and pTie-2 were significantly decreased [Bcl-2: 0.09±0.01, 0.20±0.01, 0.16±0.02, 0.31±0.01, 0.22±0.01 vs. 0.34±0.01; Bcl-2/Bax: (14.93±1.86)%, (68.03±5.56)%, (36.93±2.22)%, (119.1±13.3)%, (64.23±6.07)% vs. (208.33±7.37)%; Tie-2: 0.07±0.01, 0.16±0.02, 0.11±0.01, 0.21±0.01, 0.18±0.01 vs. 0.26±0.01; pTie-2: 0.05±0.01, 0.15±0.01, 0.07±0.01, 0.22±0.02, 0.16±0.01 vs. 0.27±0.01; all P=0.000], in addition, pAkt and ZO-1 in OGD, OGD+ Ang-1, OGD+ DHA, and OGD+ DHA+ GW9662+ Ang-1 groups were also significantly reduced (pAkt: 0.13±0.01, 0.26±0.01, 0.14±0.01, 0.28±0.02 vs. 0.39±0.02; ZO-1: 0.08±0.01, 0.18±0.01, 0.10±0.01, 0.19±0.01 vs. 0.23±0.02; all P=0.000). Compared with the OGD group, OGD+ Ang-1 and OGD+ DHA+ Ang-1 groups demonstrated significantly reduced cell apoptosis (both P=0.000). Compared with normal control+ Ang-1 group, the expression levels of pTie-2, pAkt, and ZO-1 were significantly decreased in OGD+ Ang-1 and OGD+ DHA+ Ang-1 groups (pTie-2: 0.15±0.01, 0.22±0.02 vs. 0.52±0.02; pAkt: 0.26±0.01, 0.37±0.04 vs. 0.67±0.05; ZO-1: 0.18±0.01, 0.24±0.02 vs. 0.39±0.05; all P=0.000). Compared with OGD+ Ang-1 group, OGD+ DHA+ Ang-1 group had significantly decreased cell apoptosis and expression levels of Bax, caspase-3 and Tie-1, and significantly increased expressions of Bcl-2, Bcl-2/Bax, Tie-2, pTie-2, pAkt, and ZO-1 (all P=0.000), while OGD+ DHA+ GW9662+ Ang-1 group showed no significant differences in these indexes (P=0.202, 0.770, 0.382, 0.448, 0.233, 0.736, 0.143, 0.526, 0.495, 0.670). Conclusion Pretreatment with DHA may enhance the protective effect of Ang-1 on rat BMVECs under the condition of OGD, possibly via activating PPAR-γ, suppressing Tie-1 expression, and hence amplifying the activation of Tie-2. Key words: Docosahexaenoid acid; Oxygen glucose deprivation; Cell apoptosis; Angiopoietin-1; Brain microvascular endothelial cells

The protective role of angiopoietin-1 in blood brain barrier permeability after cerebral ischemia reperfusion in rats

Objective To investigate the protective role of angiopoietin-1 (Ang-1) in blood brain barrier permeability after cerebral ischemia reperfusion in rats.Methods Rat cerebral ischemia reperfusion was established by middle cerebral artery occlusion (MCAO) and randomly divided into 3 groups:sham operated group,saline group and angiopoietin-1 treatment group (n =10).The cerebral water content,brain barrier permeability,infarct volume and the neurological deficits scores were measured 48 h after reperfusion.Results 48 h after reperfusion,the cerebral water content of Ang-1 group was (68.69 ±4.46)％,significantly lower than that of saline group [(79.08 ± 4.12) ％,t =4.33,P ＜ 0.05],and similar to the sham group [(64.37 ± 3.28) ％,t =1.387,P ＞ 0.05].The evans blue (EB) content of three groups were (87.40 ±9.42) μg/g tissue,(381.40 ±22.30) μg/g tissue and (130.90 ± 16.80) μg/g tissue,respectively.The EB content of Ang-1 group was significantly lower than that of saline group (t =15.12,P ＜ 0.01) and similar to the sham group (t =0.934,P ＞ 0.05).The infarct volume of Ang-1 group was (0.119 ± 0.022) cm3,significantly lower than that of saline group [(0.407 ± 0.036) cm3,t =6.327,P ＜ 0.01] and similar to the sham group [(0.104 ± 0.011) cm3,t =1.099,P ＞ 0.05].The neurological deficits scores of three groups were (0.79 ± 0.11),(2.47 ± 0.34) and (1.83 ± 0.29),respectively.The neurological deficits scores of Ang-1 group was significantly lower than that of saline group (t =4.37,P＜0.05) and similar to the sham group (t=2.31,P＜0.05).Conclusion Ang-1 has a protective effect on the permeability of blood brain barrier after cerebral ischemia-reperfusion. Key words: Angiopoietin-1 ; Cerebral ischemia; Reperfusion injury; Blood brain barrier

COMP-Angiopoietin-1 Promotes Cavernous Angiogenesis in a Type 2 Diabetic Rat Model

Cartilage oligomeric matrix protein-angiopoietin-1 (COMP-Ang1) is an angiogenic factor for vascular angiogenesis. The aim was to investigate the effect of an intracavernosal injection of COMP-Ang1 on cavernosal angiogenesis in a diabetic rat model. Male Otsuka Long-Evans Tokushima Fatty (OLETF) rats made up the experimental group (1 yr old) and Long-Evans Tokushima Otsuka (LETO) rats made up the control group. The experimental group was divided into vehicle only, 10 µg COMP-Ang1, and 20 µg COMP-Ang1. COMP-Ang1 was injected into the corpus cavernosum of the penis. After 4 weeks, the penile tissues of the rats were obtained for immunohistochemistry and Western blot analysis. The immunoreactivity of PECAM-1 and VEGF was increased in the COMP-Ang1 group compared with the vehicle only group. Moreover, the expression of PECAM-1 and VEGF was notably augmented in the 20 µg Comp Ang-1 group. In the immunoblotting study, the expression of PECAM-1 and VEGF protein was significantly less in the OLEFT rats than in the control LETO rats. However, this expression was restored to control level after intracavernosal injection of COMP-Ang1. These results show that an intracavernosal injection of COMP-Ang1 enhances cavernous angiogenesis by structurally reinforcing the cavernosal endothelium.

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.

E0044 The role of Ang1 and eNOS in the proangiogenic effect of simvastatin after myocardial infarction in rats

ObjectiveTo investigate the roles of angiopoietin-1 (Ang-1) and endothelial nitric oxide synthase (eNOS) in pro-angiogenic effect of simvastatin after experimental myocardial infarction (MI).Methods60 healthy adult SD rats were randomly divided...

Transient transfection of angiopoietin-1 into vascular endothelial cells by ultrasound combined with microbubbles

Objective To investigate the effects of ultrasound and microbubbles on transient transfection of angiopoietin-1(Ang-1) into vascular endothelial CRL1730 cells and the effects of cell migration. Methods CRL1730 cells were divided into four groups:a control group (without any treatment), an Ang-1 group (with Ang-1 added only), a microbubble group (with only microbubbles added) and an ultrasound with microbubbles group (Ang-1, microbubbles and 1 MHz ultrasound irradiation for 30 s). All were cultivated for 24 h. The expression of Ang-1 mRNA and protein were measured using RT-PCR and Western blotting. The cells' migration ability was detected using the Transwell method. Results There was no difference in the expression of Ang-1 mRNA or protein among the control group, the Ang-1 group and the microbubbles group, but the expression of Ang-1 mRNA and protein was increased significantly in the ultrasound plus microbubbles group. Cell migration ability was also enhanced significantly in the ultrasound and microbubbles group compared with the other groups. Conclusion A combination of ultrasound and microbubbles might successfully transfect Ang-1 genes into vascular endothelial cells, and the migration ability of the transfected cells is also enhanced. Key words: Microbubbles; Ultrasound; Angiopoietin-1; Migration

COMP–angiopoietin-1 accelerates bone formation during distraction osteogenesis

IntroductionDuring distraction osteogenesis, new and highly vascularized bone is formed, with angiogenesis preceding osteogenesis. We investigated the possibility that COMP–Ang1, an angiogenic factor, may facilitate bone formation. MethodsRats were divided into three groups. Control rats underwent tibial distraction without treatment. In the two remaining groups, BSA (100μg) or COMP–Ang1 (100μg) were injected transcutaneously into the center of the distraction zone. Using radiographic and histologic analyses, we assessed total bone volume, vascular density, and bone mineral density. Total RNA was prepared from regenerated bone and analyzed for osteogenic marker protein expression using real-time RT–PCR analysis. ResultsBone formation in the distraction gap progressed more quickly in the COMP–Ang1-treated group than in the BSA-treated group. Histological findings and immunostaining of endothelial cells for factor VIII revealed that Comp–Ang1 group animals exhibited higher levels of vascularity. NanoCT and dual-energy X-ray absorptiometry analyses revealed increased new bone formation along capillaries in the COMP–Ang1 group compared with the BSA group. Runt-related transcription factor 2 and its target genes, including bone sialoprotein, type 1 collagen, osteopontin, and osterix, were significantly upregulated in the COMP–Ang1 group. ConclusionsOur results are consistent with previous descriptions of the positive relationship between angiogenesis and osteogenesis. In addition, our results suggest the potential use of COMP–Ang1 as a therapeutic agent for treatment of distracted limbs by enhancing angiogenesis.

Mesenchymal stem cells modified with angiopoietin-1 improve remodeling in a rat model of acute myocardial infarction

We used human angiopoietin-1 (hAng1)-modified mesenchymal stem cells (MSCs) to treat acute myocardial infarction (AMI) in rats. The hAng1 gene was transfected into cultured rat MSCs using an adenoviral vector. Five million hAng-transfected MSCs (MSC Ang1) or green fluorescent protein transfected MSCs (MSC GFP) or PBS only (PBS group) were injected intramyocardially into the inbred Lewis rat hearts immediately after myocardial infarction. MSC Ang1 survived in the infarcted myocardium, and expressed hAng1 at both mRNA and protein levels. The vascular density was higher in the MSC Ang1 and MSC GFP groups than in the PBS group. The measurements of infarcted ventricular wall thickness, infarction area, and left ventricular diameter indicated that heart remodeling was inhibited and heart function was improved in both the MSC Ang1 and MSC GFP groups. However, in contrast to the MSC GFP group, the MSC Ang1 group showed enhanced angiogenesis and arteriogenesis (by 11–35%), infarction area was reduced by 30% and the left ventricular wall was 46% thicker ( P < 0.05). The results indicated that hAng1-modified MSCs improved heart function, followed by angiogenic effects in salvaging ischemic myocardium and reduced cardiac remodeling.

Combination of In Vivo Angiopoietin-1 Gene Transfer and Autologous Bone Marrow Cell Implantation for Functional Therapeutic Angiogenesis

Autologous bone marrow mononuclear cell (BM-MNC) implantation into ischemic tissues promotes angiogenesis, but a large amount of marrow aspiration is required, which is a major clinical limitation. Angiopoietin-1 (Ang-1) is requisite for vascular maturation during angiogenesis. We examined the impacts of combinatorial Ang-1 gene transfer and low-dose autologous BM-MNC implantation on therapeutic angiogenesis in a rabbit model of hind limb ischemia. Rabbits were divided into 4 groups: phosphate-buffered saline (control), 500 microg Ang-1 plasmid (Ang-1), 1 x 10(6) autologous BM-MNCs (BMC), and Ang-1 plasmid plus BM-MNCs (combination). The Ang-1 group had a greater angiographic score and capillary density compared with the control (P<0.05), but the Ang-1 gene therapy alone did not improve transcutaneous oxygen pressure (TcO2) and skin ulcer score. However, the combination group showed a significant improvement in not only angiographic score and capillary density (P<0.05) but also TcO2 (P<0.05) and skin ulcer score. These efficacies were greater in the combination group compared with the BMC group. This Ang-1 gene and BM-MNC combination therapy enhances not only quantitative but also qualitative angiogenesis in ischemic tissues. Moreover, the combination therapy will enable a reduction in the amount of BM aspiration required for significant therapeutic angiogenesis.

Adenoviral-delivered angiopoietin-1 reduces the infarction and attenuates the progression of cardiac dysfunction in the rat model of acute myocardial infarction

In acute myocardial infarction (AMI), prognosis and mortality rate are closely related to the infarct size and the progression of postinfarction cardiac failure. Angiogenic gene therapy has presented a new approach for the treatment of AMI. Angiopoietin-1 (Ang1) is a critical angiogenic factor for vascular maturation and enhances vascular endothelial growth factor (VEGF)-induced angiogenesis in a complementary manner. We hypothesized that gene therapy using Ang1 for AMI might promote angiogenesis cooperatively with intrinsic VEGF, since high concentrations of circulating VEGF have been reported in AMI. To evaluate our hypothesis, we employed a rat AMI model and adenoviral Ang1 (HGMW-approved gene symbol ANGPT1) gene transfer to the heart. A significant increase in capillary density and reduction in infarct sizes were noted in the infarcted hearts with adenoviral Ang1 gene treatment compared with control infarcted hearts treated with saline or adenoviral vector containing the β-galactosidase gene. Furthermore, the Ang1 group showed significantly higher cardiac performance in echocardiography (55.0% of ejection fraction, P < 0.05 vs control) than the saline or adenoviral controls (36.0 or 40.5%, respectively) 4 weeks after myocardial infarction. The adenoviral delivery of Ang1 during the acute phase of myocardial infarction would be feasible to attenuate the progression of cardiac dysfunction in the rat model.