| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
Department of Bioregulation, Yamaguchi University School of Medicine, Ube, Yamaguchi, Japan 755-8505
 |
ABSTRACT |
|---|
 TOP
 ABSTRACT
 INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
Therapeutic angiogenesis can be induced by local implantation of bone marrow cells. We tried to enhance the angiogenic potential of this treatment by ex vivo hypoxia stimulation of bone marrow cells before implantation. Bone marrow cells were collected and cultured at 33°C under 2% O2-5% CO2-90% N2 (hypoxia) or 95% air-5% CO2 (normoxia). Cells were also injected into the ischemic hindlimb of rats after 24 h of culture. Hypoxia culture increased the mRNA expression of vascular endothelial growth factor (VEGF), vascular endothelial (VE)-cadherin, and fetal liver kinase-1 (Flk-1) from 2.5- to fivefold in bone marrow cells. The levels of VEGF protein in the ischemic hindlimb were significantly higher 1 and 3 days after implantation with hypoxia-cultured cells than with normoxia-cultured or noncultured cells. The microvessel density and blood flow rate in the ischemic hindlimbs were also significantly (P < 0.001) higher 2 wk after implantation with hypoxia-cultured cells (89.7 ± 5.5%) than with normoxia-cultured cells (67.0 ± 9.6%) or noncultured cells (70.4 ± 7.7%). Ex vivo hypoxia stimulation increased the VEGF mRNA expression and endothelial differentiation of bone marrow cells, which together contributed to improved therapeutic angiogenesis in the ischemic hindlimb after implantation.
endothelial differentiation; vascular endothelial growth factor; vascular endothelial-cadherin; ischemic hindlimb.
| |
INTRODUCTION |
|---|
|
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
INDUCTION OF THERAPEUTIC ANGIOGENESIS by various methods has recently been developed to treat ischemic diseases (4, 12, 19, 22, 24). Both the increase of multiple angiogenic cytokines and the proliferation or migration of endothelial cells are critically important for improving angiogenesis (5, 6, 21). Bone marrow cell implantation is considered an ideal method for inducing therapeutic angiogenesis, because bone marrow cells secrete multiple angiogenic cytokines and differentiate into endothelial cells (2, 9).
We have shown previously that therapeutic angiogenesis can be induced by local implantation of bone marrow cells, and this finding was related to both the endothelial differentiation and the secretion of multiple angiogenic cytokines from the implanted bone marrow cells (10, 14, 17). We have also begun a clinical trial to induce therapeutic angiogenesis by local implantation of autologous bone marrow cells in patients with ischemic heart and limb diseases (11). Our preliminary clinical outcome indicates that the effectiveness of therapeutic angiogenesis induced by autologous bone marrow cell implantation was insufficient and required further study.
Theoretically, the angiogenic potency induced by bone marrow cell implantation could be enhanced by increasing the production of angiogenic cytokines or by improving the endothelial differentiation from bone marrow cells. mRNA expression of vascular endothelial growth factor (VEGF) can be increased in various types of cells when cultured under hypoxia (13, 20, 26). We have found that ex vivo hypoxic stimulation increases the production of VEGF from bone marrow cells (9). The effect of therapeutic angiogenesis induced by bone marrow cell implantation might be enhanced by ex vivo hypoxic stimulation of bone marrow cells before implantation.
In the present study, we investigated whether ex vivo hypoxic stimulation would influence the level of mRNA expression of VEGF and the endothelial differentiation of bone marrow cells. We also observed whether the angiogenic potency could be enhanced by the implantation of hypoxia-cultured bone marrow cells into the ischemic hindlimb of rats.
| |
MATERIALS AND METHODS |
|---|
|
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
Animals. Male Dark Agouti rats, 6-8 mo old, were used for all experiments, which were approved by the institutional Animal Care and Use Committee of Yamaguchi University School of Medicine. Animals were housed under clean conditions and allowed free access to food and water in a temperature-controlled environment with a 12:12-h light-dark cycle.
Ex vivo hypoxic stimulation of bone marrow cells. Rats were killed under deep general anesthesia induced by intraperitoneal pentobarbital injection. Bone marrow was collected from the femur and tibia and placed in PBS. Simple bone marrow cell suspensions were prepared by gently pressing bone marrow segments through fine wire mesh. Bone marrow cells were separated from red blood cells by 1,800 rpm centrifugation for 20 min in Histopaque 1077 (Sigma, St. Louis, MO). Collected cells were washed two times, and the survival rate of cells was higher than 98%, as evaluated by staining the cells with 0.4% trypan blue solution (Sigma). Purified bone marrow cells were then resuspended at a density of 1 × 107/ml in RPMI 1640 (GIBCO, Grand Island, NY) supplemented with 10% fetal bovine serum (FBS), 100 U/ml penicillin, and 100 µg/ml streptomycin (all from GIBCO). Cells were cultured at 33°C (7), under 2% O2-5% CO2-90% N2 (hypoxia) or under 95% air-5% CO2 (normoxia) condition, in a gas-tight humidified incubator (ASTEC, Fukuoka, Japan).
RT-PCR Analysis. Cells were collected after 0 (baseline), 4, 8, 12, 24, and 48 h of culture, and five separate cultures were performed for each time point. Total RNA was extracted from cells with the use of an EZNA total RNA kit (Omega Bio-Tek; Doraville, GA) according to the manufacturer's instructions. cDNA was synthesized and amplified by PCR with the BcaBEST RNA PCR Kit Version 2.1 (Takara, Osaka, Japan) according to manufacturer's instructions. We used the following primers: VEGF (353 bp), sense 5'-CCCTGGTGGACATCTTCCAGG-3' and reverse 5'-CGCCTTGCAACGCGAGTCTG-3'; VE-cadherin (336 bp), sense 5'-AACCATGACAACACCGCC-3' and reverse 5'-CTCGTGAATCTCCAGCGC-3'; fetal liver kinase-1 (Flk-1) (434 bp), sense 5'-TGGCTCACAGGCAACATC-3' and reverse 5'-CTTCTTTCCTCACCCTTCG-3'. GAPDH (726 bp), sense 5'-CTTCATTGACCTCAACTACATGGT-3' reverse 5'-CTCAGTGTAGCCCAGGATGCCCTT-3', was used as a positive standard control for mRNA quantitation and for determining stability between specimens.
Amplifications of these cDNAs were performed under the following conditions: denaturing for 1 min at 94°C, annealing for 1 min at 60°C, and elongation for 1 min at 72°C. Optimal numbers of cycles were determined to be 28 for VEGF, 30 for VE-cadherin, 27 for Flk-1, and 25 for GAPDH. The PCR products were labeled with [
-32P]dCTP and separated on 5% polyacrylamide gels.
Relative radioactivity of the bands was estimated with an imaging
analyzer (model BAS-2000; Fuji Photo Film, Tokyo, Japan). Expressions
of mRNA were normalized to constant GAPDH expression, and for
statistical analysis, the expression level for each factor measured was
calculated as the percentage of baseline (i.e., 0 h) level for
that factor.
Bone marrow cell implantation in the ischemic hindlimb of rats. Bone marrow cells were collected after 24 h of culture under hypoxia or normoxia. Survival rate of these collected cells was higher than 98% according to the results of trypan blue staining as described in Ex vivo hypoxic stimulation of bone marrow cells. Cells were suspended in PBS at the density of 1 × 109 /ml for injection. The induction of angiogenesis by bone marrow cell implantation was investigated in a rat ischemic hindlimb model described previously (10). Briefly, rats were anesthetized with 50 mg/kg of pentobarbital given intraperitoneally. A vertical longitudinal incision was made in the left limb. The femoral artery and its branches were dissected, ligated, and completely excised. The bone marrow cells (1 × 107 in 10 µl of PBS) with 24 h of culture under hypoxia (hypoxia group, n = 23) or under normoxia (normoxia group, n = 23) were then injected with a 26-gauge needle into the ischemic muscles in the thigh of rats at six points. For control, uncultured (control group, n = 23) bone marrow cells were also injected into the ischemic hindlimb of rats. Finally, the incision was closed by suture.
Measurement of VEGF level in ischemic hindlimb by ELISA.
On days 1, 3, and 7 after the
implantation of bone marrow cells, five rats per group per day were
killed, samples of the adductor muscles were harvested, and wet weights
were measured. All samples were minced and homogenated with 2.5 ml of
PBS on ice. These samples were centrifuged at 10,000 g for 5 min at 4°C, and the resulting supernatants were stored at 
80°C.
VEGF in the supernatant was measured with VEGF ELISA kits (R&D Systems,
Minneapolis, MN) according to the manufacturer's instructions. The
detection limit was 5 pg/ml. Levels of VEGF per gram wet tissue was
used for statistical analysis.
Histological analysis of microvessel density. Five rats from each group were killed 2 wk after treatment (n = 7 or n = 5 in each group), and the quadriceps and adductor muscles were harvested. Samples of harvested muscle were embedded in optimum cutting temperature compound, and snap-frozen in liquid nitrogen. To detect the development of microvessels, 5-µm-thick frozen sections were stained for alkaline phosphatase with an indoxyl tetrazolium method and then were counterstained with eosin. The number of microvessels and muscle fibers were counted under a microscope using ×200 magnification by a single observer blind to the treatment regimen, and a total of 20 different fields were randomly selected. Density of microvessels was estimated by the microvessel/muscle fiber ratio.
Evaluation of blood flow recovery in the ischemic hindlimb. Eight rats from each group were reanesthetized 2 wk after the implantation of bone marrow cells. Rats were placed on a heating plate set at 37°C, and then 6 × 105/200 µl of eosin Dye-Trak microspheres (15 µm in diameter; Triton Technology, Loughborough, UK) were injected into the abdominal aorta. Rats were killed by cutting the abdominal aorta ~30 s after the injection of the microspheres. Tissue specimens were collected from the hindlimb, weighed, and then digested in 16N KOH for 48 h at 60°C. The microspheres in the tissues were reclaimed with a vacuum filter, and dye from the microspheres was extracted with dimethyl formamide. The optical density (OD) of these dye samples was measured with a spectrophotometer. The recovery of perfusion in the ischemic hindlimb was evaluated by determining the percentage of limb blood flow in relation to of that of a normal right hindlimb, which was calculated as (ischemic limb OD/normal limb OD) × (normal limb tissue wt /ischemic limb tissue wt) × 100 (23).
Statistical analysis. All data are expressed as means ± SD. Statistical significance was evaluated with unpaired Student's t-test for comparisons between two means, with ANOVA, followed by Scheffé's procedure for more than two means, and with repeated-measures ANOVA to test for interaction. Data were considered significant when the P value was <0.05.
| |
RESULTS |
|---|
|
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
Expression of VEGF, VE-cadherin, and Flk-1 mRNA in bone marrow
cells.
The level of expression of VEGF mRNA in bone marrow cells increased
rapidly after culture under hypoxia and showed a peak at ~24 h of
culture. However, the level of VEGF mRNA expression in bone marrow
cells did not change significantly within the first 12 h of
culture under normoxia and only increased slightly after 24 h of
culture under normoxia. Quantitative analysis showed that the level of
VEGF mRNA expression in bone marrow cells was increased about fivefold
after 24 h of culture under hypoxia but only ~1.2-fold after
48 h of culture under normoxia (Fig.
1).
|
|
VEGF levels in ischemic hindlimb after bone marrow cell
implantation.
Levels of VEGF protein in the ischemic hindlimb of rats
decreased with time after treatment in all groups (Fig.
3). The VEGF level in ischemic
hindlimb of rats was significantly higher in the hypoxia group than in
the normoxia or control groups at 1 and 3 days after treatment
(245.7 ± 46.6 vs. 119.0 ± 25.3 and 127.4 ± 24.6 pg/g
tissue on day 1, P < 0.001; 173.2 ± 30.4 vs. 88.1 ± 24.9 and 89.9 ± 15.4 pg/g tissue on
day 3, P < 0.01). However, no significant
difference was found among the three groups 7 days after treatment.
VEGF concentration in the ischemic hindlimb of rats was not
significantly different between the normoxia and control groups at any
of the time points after treatment.
|
Microvessel density in ischemic hindlimb.
The microvessel-to-muscle fiber ratio was significantly higher in the
hypoxia group (1.78 ± 0.09) than in the control (1.41 ± 0.18) and normoxia (1.34 ± 0.22) groups (P < 0.01, Fig. 4). No significant difference
was seen between the control and normoxia groups.
|
Blood flow recovery of ischemic hindlimb after bone marrow
cell implantation.
Blood flow recovery in the ischemic hindlimb was evaluated and
expressed as the percentage of normal blood flow in the right (nonischemic) hindlimb. At 2 wk postimplantation with bone
marrow cells, recovery of blood flow in the ischemic hindlimb
was significantly greater in the hypoxia group (89.7% ± 5.5%) than
in either the normoxia group (67.0% ± 9.6%) or the control groups
(70.4% ± 7.7%) (P < 0.001; Fig.
5).
|
| |
DISCUSSION |
|---|
|
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
We have safely performed local implantation of autologous bone marrow cells to induce therapeutic angiogenesis for patients with ischemic heart or limb disease (11). To find a safe, easy, and simple method for improving the angiogenic effect of this treatment, we undertook the present investigation. We cultured bone marrow cells under 33°C (7), which might depress the metabolism of cells and prevent the rapid maturation or differentiation of these stem cells during culture. Cells were subjected to mild hypoxic stimulation under 2% O2 rather than full hypoxia under 0% O2, because we wanted to maintain the cells in good bioactivity for implantation in vivo after culturing. Under such culture conditions, the survival rate of bone marrow cells was higher than 98% even after 24 h of hypoxia, which was equivalent to the rate for freshly collected bone marrow cells. This result indicates that it might also be safe and reasonable to implant the bone marrow cells even after 24 h of culture under 2% O2 at 33°C.
The main aim of the present study was to develop a method for increasing the angiogenic effect of bone marrow cell implantation. Therapeutic angiogenesis can be induced by direct administration or gene transfer of VEGF or basic fibroblast growth factor (bFGF) (4, 19, 21, 24). One potential method for increasing the angiogenic potency of bone marrow cell implantation is to increase the production of angiogenic cytokines, such as VEGF and bFGF, from the implanted bone marrow cells. In the present study, we tried to enhance VEGF mRNA expression of bone marrow cells ex vivo before implantation, because the enhanced VEGF mRNA expression of bone marrow cells might result in higher VEGF production and more effective angiogenesis in vivo after implantation. As with other cells cultured under hypoxia (13, 20, 26), 2% O2 hypoxic stimulation rapidly increased the expression of VEGF mRNA in bone marrow cells with the expression level increased fivefold after 24 h of hypoxic culture.
Bone marrow as an origin of endothelial progenitor cells has been shown to contribute to postnatal vasculogenesis in physiological and pathological neovascularization (1, 3). Implantation of ex vivo expanded endothelial progenitor cells has also been shown to induce therapeutic angiogenesis effectively for ischemic tissues in experimental models (15, 16). For increasing the angiogenic effect of bone marrow cell implantation, we believed that increasing the endothelial differentiation of bone marrow cells might provide more endothelial cells and thus improve angiogenesis. Ex vivo hypoxic stimulation is known to upregulate the expression of VE-cadherin and Flk-1 in some cells (8, 25). We investigated whether mild hypoxia stimulation would also influence the endothelial differentiation of bone marrow cells, which should be a key factor for improving angiogenesis. Expression of VE-cadherin, a special surface antigen of endothelial cells, increased rapidly to ~3.3-fold of baseline after 48 h of culture under 2% O2 hypoxia. Flk-1, a receptor of the most important angiogenic growth factor of VEGF, is a marker used to evaluate the endothelial differentiation of bone marrow cells (2, 18). In the present study, we observed that the expression of Flk-1 mRNA in bone marrow cells increased significantly after culture under 2% O2 hypoxia, showing the level increased ~2.5 of baseline after 48 h of hypoxic culture. mRNA expression of VE-cadherin and Flk-1 increased to only ~1.3-fold or less of baseline in the bone marrow cells after culture under normoxia. Significant increase of VE-cadherin and Flk-1 gene expression in bone marrow cells after culture under hypoxia indicates the possibility that ex vivo hypoxia stimulation could improve the endothelial differentiation of bone marrow cells.
To determine whether the over expression of VEGF mRNA in hypoxia-cultured bone marrow cells would also contribute to higher VEGF production in vivo after implantation, we injected bone marrow cells into the ischemic hindlimb of rats and then measured the local level of VEGF protein in the ischemic hindlimb. We found a significantly higher VEGF concentration in the ischemic hindlimb after implantation with hypoxia-cultured bone marrow cells than with normoxia-cultured or noncultured bone marrow cells. This result indicates clearly that the overexpression of VEGF mRNA in bone marrow cells contributed to higher VEGF production in vivo after implantation into ischemic tissue.
The final purpose of our investigation was to develop a method for inducing an improved therapeutic angiogenesis in ischemic tissues by bone marrow cell implantation. Using an ischemic hindlimb model, we found that the microvessel density was significantly higher in the hypoxia group than in the control and normoxia groups. Furthermore, blood flow in the ischemic hindlimb recovered to ~90% of normal level after implantation with hypoxia-cultured bone marrow cells, which was much higher than the recovering level after implantation with normoxia-cultured (67%) or noncultured (70%) bone marrow cells. The significant enhancement of angiogenic potency after implantation of hypoxia-cultured bone marrow cells is apparently associated with both the upregulation of VEGF mRNA and the improvement of endothelial differentiation of bone marrow cells by ex vivo hypoxia stimulation before implantation.
In conclusion, we found that ex vivo mild hypoxia stimulation of bone marrow cells increased their level of VEGF mRNA expression and of endothelial differentiation. Furthermore, blood flow recovery in the ischemic hindlimb was improved more in rats implanted with ex vivo hypoxia-stimulated bone marrow cells than in those implanted with noncultured bone marrow cells. Our results indicate that improved effectiveness of therapeutic angiogenesis might be induced by ex vivo hypoxia prestimulation of bone marrow cells before implantation.
| |
ACKNOWLEDGEMENTS |
|---|
This work was supported by Ministry of Education, Science, and Culture Grant-in-Aid 13671391 for scientific research and by the Research Fund for the Development of New Medical Treatments of the Ministry of Education.
| |
FOOTNOTES |
|---|
Address for reprint requests and other correspondence: K. Hamano, Division of Cardiovascular Surgery, Dept. of Bioregulation, Yamaguchi University School of Medicine, 1-1-1 Minami-Kogushi, Ube, Yamaguchi, Japan 755-8505 (E-mail: kimikazu{at}yamaguchi-u.ac.jp).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
10.1152/ajpheart.00261.2002
Received 25 March 2002; accepted in final form 17 April 2002.
| |
REFERENCES |
|---|
|
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES
|
|---|
1.
Asahara, T,
Masuda H,
Takahashi T,
Kalka C,
Pastore C,
Silver M,
Kearne M,
Magner M,
and
Isner JM.
Bone marrow origin of endothelial progenitor cells responsible for postnatal vasculogenesis in physiological and pathological neovascularization.
Circ Res
85:
221-228,
1999
2.
Asahara, T,
Murohara T,
Sullivan A,
Silver M,
van der Zee R,
Li T,
Witzenbichler B,
Schatteman G,
and
Isner JM.
Isolation of putative progenitor endothelial cells for angiogenesis.
Science
275:
964-967,
1997
3.
Asahara, T,
Takahashi T,
Masuda H,
Kalka C,
Chen D,
Iwaguro H,
Inai Y,
Silver M,
and
Isner JM.
VEGF contributes to postnatal neovascularization by mobilizing bone marrow-derived endothelial progenitor cells.
EMBO J
18:
3964-3972,
1999[Web of Science][Medline].
4.
Baumgartner, I,
Pieczek A,
Manor O,
Blair R,
Kearney M,
Walsh K,
and
Isner JM.
Constitutive expression of phVEGF165 after intramuscular gene transfer promotes collateral vessel development in patients with critical limb ischemia.
Circulation
97:
1114-1123,
1998
5.
Bouck, N,
Stellmach V,
and
Hsu SC.
How tumors become angiogenic.
Adv Cancer Res
69:
135-174,
1996[Web of Science][Medline].
6.
Carmeliet, P.
Mechanisms of angiogenesis and arteriogenesis.
Nat Med
6:
389-395,
2000[Web of Science][Medline].
7.
Dexter, TM,
Allen TD,
and
Lajtha LG.
Conditions controlling the proliferation of haemopoietic stem cells in vitro.
J Cell Physiol
91:
335-344,
1977[Web of Science][Medline].
8.
Gerber, HP,
Condorelli F,
Park J,
and
Ferrara N.
Differential transcriptional regulation of the two vascular endothelial growth factor receptor genes. Flt-1, but not Flk-1/KDR, is up-regulated by hypoxia.
J Biol Chem
272:
23659-23667,
1997
9.
Hamano, K,
Li TS,
Kobayashi T,
Kobayashi S,
Matsuzaki M,
and
Esato K.
Angiogenesis induced by the implantation of self-bone marrow cells: a new material for therapeutic angiogenesis.
Cell Transplant
9:
439-443,
2000[Web of Science][Medline].
10.
Hamano, K,
Li TS,
Kobayashi T,
Tanaka N,
Kobayashi S,
Matsuzaki M,
and
Esato K.
The induction of angiogenesis by the implantation of autologous bone marrow cells: a novel and simple therapeutic method.
Surgery
130:
44-54,
2001[Web of Science][Medline].
11.
Hamano, K,
Nishida M,
Hirata K,
Mikamo A,
Li TS,
Harada M,
Miura T,
Matsuzaki M,
and
Esato K.
Local implantation of autologous bone marrow cells for therapeutic angiogenesis in patients with ischemic heart disease: clinical trial and preliminary results.
Jpn Circ J
65:
845-847,
2001[Medline].
12.
Horvath, KA,
Cohn LH,
Cooley DA,
Crew JR,
Frazier OH,
Griffith BP,
Kadipasaoglu K,
Lansing A,
Mannting F,
March R,
Mirhoseini MR,
and
Smith C.
Transmyocardial laser revascularization: results of a multicenter trial with transmyocardial laser revascularization used as sole therapy for end-stage coronary artery disease.
J Thorac Cardiovasc Surg
113:
645-653,
1997
13.
Iizuka, M,
Yamauchi M,
Ando K,
Hori N,
Furusawa Y,
Itsukaichi H,
Fukutsu K,
and
Moriya H.
Quantitative RT-PCR assay detecting the transcriptional induction of vascular endothelial growth factor under hypoxia.
Biochem Biophys Res Commun
205:
1474-1480,
1994[Web of Science][Medline].
14.
Ikenaga, S,
Hamano K,
Nishida M,
Kobayashi T,
Li TS,
Kobayashi S,
Matsuzaki M,
Zempo N,
and
Esato K.
Autologous bone marrow implantation induced angiogenesis and improved deteriorated exercise capacity in a rat ischemic hindlimb model.
J Surg Res
96:
277-283,
2001[Web of Science][Medline].
15.
Kalka, C,
Masuda H,
Takahashi T,
Kalka-Moll WM,
Silver M,
Kearney M,
Li T,
Isner JM,
and
Asahara T.
Transplantation of ex vivo expanded endothelial progenitor cells for therapeutic neovascularization.
Proc Natl Acad Sci USA
97:
3422-3427,
2000
16.
Kawamoto, A,
Gwon HC,
Iwaguro H,
Yamaguchi JI,
Uchida S,
Masuda H,
Silver M,
Ma H,
Kearney M,
Isner JM,
and
Asahara T.
Therapeutic potential of ex vivo expanded endothelial progenitor cells for myocardial ischemia.
Circulation
103:
634-637,
2001
17.
Kobayashi, T,
Hamano K,
Li TS,
Katoh T,
Kobayashi S,
Matsuzaki M,
and
Esato K.
Enhancement of angiogenesis by the implantation of self bone marrow cells in a rat ischemic heart model.
J Surg Res
89:
189-195,
2000[Web of Science][Medline].
18.
Kocher, AA,
Schuster MD,
Szabolcs MJ,
Takuma S,
Burkhoff D,
Wang J,
Homma S,
Edwards NM,
and
Itescu S.
Neovascularization of ischemic myocardium by human bone-marrow-derived angioblasts prevents cardiomyocyte apoptosis, reduces remodeling and improves cardiac function.
Nat Med
7:
430-436,
2001[Web of Science][Medline].
19.
Losordo, DW,
Vale PR,
Symes JF,
Dunnington CH,
Esakof DD,
Maysky M,
Ashare AB,
Lathi K,
and
Isner JM.
Gene therapy for myocardial angiogenesis: initial clinical results with direct myocardial injection of phVEGF165 as sole therapy for myocardial ischemia.
Circulation
98:
2800-2804,
1998
20.
Minchenko, A,
Bauer T,
Salceda S,
and
Caro J.
Hypoxic stimulation of vascular endothelial growth factor expression in vitro and in vivo.
Lab Invest
71:
374-379,
1994[Web of Science][Medline].
21.
Risau, W.
Mechanisms of angiogenesis.
Nature
386:
671-674,
1997[Medline].
22.
Rosengart, TK,
Lee LY,
Patel SR,
Sanborn TA,
Parikh M,
Bergman GW,
Hachamovitch R,
Szulc M,
Kligfield PD,
Okin PM,
Hahn RT,
Devereux RB,
Post MR,
Hackett NR,
Foster T,
Grasso TM,
Lesser ML,
Isom OW,
and
Crystal RG.
Angiogenesis gene therapy: phase I assessment of direct intramyocardial administration of an adenovirus vector expressing VEGF121 cDNA to individuals with clinically significant severe coronary artery disease.
Circulation
100:
468-474,
1999
23.
Takeshita, S,
Isshiki T,
Ochiai M,
Eto K,
Mori H,
Tanaka E,
Umetani K,
and
Sato T.
Endothelium-dependent relaxation of collateral microvessels after intramuscular gene transfer of vascular endothelial growth factor in a rat model of hindlimb ischemia.
Circulation
98:
1261-1263,
1998
24.
Unger, EF,
Goncalves L,
Epstein SE,
Chew EY,
Trapnell CB,
Cannon RO, III,
and
Quyyumi AA.
Effects of a single intracoronary injection of basic fibroblast growth factor in stable angina pectoris.
Am J Cardiol
85:
1414-1419,
2000[Web of Science][Medline].
25.
Vasir, B,
Jonas JC,
Steil GM,
Hollister-Lock J,
Hasenkamp W,
Sharma A,
Bonner-Weir S,
and
Weir GC.
Gene expression of VEGF and its receptors Flk-1/KDR and Flt-1 in cultured and transplanted rat islets.
Transplantation
71:
924-935,
2001[Web of Science][Medline].
26.
Xiong, M,
Elson G,
Legarda D,
and
Leibovich SJ.
Production of vascular endothelial growth factor by murine macrophages: regulation by hypoxia, lactate, and the inducible nitric oxide synthase pathway.
Am J Pathol
153:
587-598,
1998
This article has been cited by other articles:
|
|
 |
|
  J. Moriya, T. Minamino, K. Tateno, N. Shimizu, Y. Kuwabara, Y. Sato, Y. Saito, and I. Komuro Long-Term Outcome of Therapeutic Neovascularization Using Peripheral Blood Mononuclear Cells for Limb Ischemia Circ Cardiovasc Interv, June 1, 2009; 2(3): 245 - 254. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 X. Hu, S. P. Yu, J. L. Fraser, Z. Lu, M. E. Ogle, J.-A. Wang, and L. Wei Transplantation of hypoxia-preconditioned mesenchymal stem cells improves infarcted heart function via enhanced survival of implanted cells and angiogenesis. J. Thorac. Cardiovasc. Surg., April 1, 2008; 135(4): 799 - 808. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Kubo, T.-S. Li, R. Suzuki, B. Shirasawa, N. Morikage, M. Ohshima, S.-L. Qin, and K. Hamano Hypoxic preconditioning increases survival and angiogenic potency of peripheral blood mononuclear cells via oxidative stress resistance Am J Physiol Heart Circ Physiol, February 1, 2008; 294(2): H590 - H595. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Kubo, T.-S. Li, R. Suzuki, M. Ohshima, S.-L. Qin, and K. Hamano Short-term pretreatment with low-dose hydrogen peroxide enhances the efficacy of bone marrow cells for therapeutic angiogenesis Am J Physiol Heart Circ Physiol, June 1, 2007; 292(6): H2582 - H2588. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. Uemura, M. Xu, N. Ahmad, and M. Ashraf Bone Marrow Stem Cells Prevent Left Ventricular Remodeling of Ischemic Heart Through Paracrine Signaling Circ. Res., June 9, 2006; 98(11): 1414 - 1421. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Tateno, T. Minamino, H. Toko, H. Akazawa, N. Shimizu, S. Takeda, T. Kunieda, H. Miyauchi, T. Oyama, K. Matsuura, et al. Critical Roles of Muscle-Secreted Angiogenic Factors in Therapeutic Neovascularization Circ. Res., May 12, 2006; 98(9): 1194 - 1202. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T.-S. Li, A. Furutani, M. Takahashi, M. Ohshima, S.-L. Qin, T. Kobayashi, H. Ito, and K. Hamano Impaired potency of bone marrow mononuclear cells for inducing therapeutic angiogenesis in obese diabetic rats Am J Physiol Heart Circ Physiol, April 1, 2006; 290(4): H1362 - H1369. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. J. O'Neill IV, B. R. Wamhoff, G. K. Owens, and T. C. Skalak Mobilization of Bone Marrow-Derived Cells Enhances the Angiogenic Response to Hypoxia Without Transdifferentiation Into Endothelial Cells Circ. Res., November 11, 2005; 97(10): 1027 - 1035. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. K. Haider and M. Ashraf Bone marrow stem cell transplantation for cardiac repair Am J Physiol Heart Circ Physiol, June 1, 2005; 288(6): H2557 - H2567. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T.-S. Li, H. Ito, M. Hayashi, A. Furutani, M. Matsuzaki, and K. Hamano Cellular expression of integrin-{beta}1 is of critical importance for inducing therapeutic angiogenesis by cell implantation Cardiovasc Res, January 1, 2005; 65(1): 64 - 72. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Pesce, A. Orlandi, M. G. Iachininoto, S. Straino, A. R. Torella, V. Rizzuti, G. Pompilio, G. Bonanno, G. Scambia, and M. C. Capogrossi Myoendothelial Differentiation of Human Umbilical Cord Blood-Derived Stem Cells in Ischemic Limb Tissues Circ. Res., September 5, 2003; 93 (5): e51 - e62. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T.-S. Li, K. Hamano, M. Nishida, M. Hayashi, H. Ito, A. Mikamo, and M. Matsuzaki CD117+ stem cells play a key role in therapeutic angiogenesis induced by bone marrow cell implantation Am J Physiol Heart Circ Physiol, August 7, 2003; 285(3): H931 - H937. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| Visit Other APS Journals Online |
P < 0.05, 
