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First published online January 30, 2008

Increase in Circulating CD34-Positive Cells in Patients with Angiographic Evidence of Moyamoya-like Vessels

Tomoyuki Yoshihara, Akihiko Taguchi [email protected], Tomohiro Matsuyama, Yoko Shimizu, Akie Kikuchi-Taura, Toshihiro Soma, David M Stern, Hiroo Yoshikawa, Yukiko Kasahara, Hiroshi Moriwaki, Kazuyuki Nagatsuka, and Hiroaki NaritomiView all authors and affiliations
Volume 28, Issue 6
https://doi.org/10.1038/jcbfm.2008.1

Abstract

Increasing evidence points to a role for circulating endothelial progenitor cells, including populations of CD34-positive (CD34+) cells, in maintenance of cerebral blood flow. In this study, we investigated the link between the level of circulating CD34+ cells and neovascularization at ischemic brain. Compared with control subjects, a remarkable increase of circulating CD34+ cells was observed in patients with angiographic moyamoya vessels, although no significant change was observed in patients with major cerebral artery occlusion (or severe stenosis) but without moyamoya vessels. Our results suggest that the increased level of CD34+ cells associated with ischemic stress is correlated with neovascularization at human ischemic brain.

Introduction

Increasing evidence points to a role for bone marrow-derived immature cells, such as endothelial progenitor cells, in maintenance of vascular homeostasis and repair. CD34-positive (CD34+) cells comprise a population enriched for endothelial progenitor cells whose contribution to neovasculature includes both direct participation in forming the neovessel and regulatory roles as sources of growth/angiogenesis factors (Majka et al, 2001). Previously, we have shown accelerated neovascularization after administration of CD34+ cells in an experimental model of stroke (Taguchi et al, 2004b) and induced by autologous bone marrow mononuclear cells (rich cell fraction of CD34+ cells) transplanted locally into patients with limb ischemia (Taguchi et al, 2003). In addition, we have observed a positive correlation between the level of circulating CD34+ cells and regional blood flow (Taguchi et al, 2004a), and cognitive function (Taguchi et al, 2007) in patients with chronic cerebral ischemia.
In this study, we have evaluated the level of circulating CD34+ cells in patients with unusually accelerated neovascularization induced by progressive occlusion (or severe stenosis) of the supraclinoid portion of the internal carotid artery, the proximal region of the anterior, and/or middle cerebral artery characterized angiographically by the presence of moyamoya-like vessels (Natori et al, 1997) that supply ischemic brain as collaterals. We have investigated the hypothesis that circulating bone marrow-derived immature cells might be associated with neovascularization at ischemic sites in the human brain.

Patients and methods

The institutional review board of the National Cardiovascular Center approved this study. All subjects provided informed consent. A total of 50 individuals, including 24 patients with occlusion or severe stenosis (>90%) at the C1 portion of the internal carotid artery or the M1 portion of the middle cerebral artery, and 26 age-matched healthy volunteers with cardiovascular risk factors, but without history of vascular disease, were enrolled. The diagnosis of cerebral artery occlusion or stenosis was made angiographically and four patients were found to have classical angiographic evidence of moyamoya-like vessels, including one with right C1 occlusion, one with right M1 occlusion, and two with bilateral C1 severe stenosis. All patients with cerebral artery occlusion or stenosis had a history of cerebral infarction. Individuals excluded from the study included patients who experienced a vascular event within 30 days of measurements, premenopausal women, and those with evidence of infection and/or malignant disease. The number of circulating CD34+ cells was quantified as described (Taguchi et al, 2007). In brief, blood samples (200 μl) were incubated with phycoerythrin-labeled anti-CD34 antibody, fluorescein isothiocyanate-labeled anti-CD45 antibody, 7-aminoactinomycin-D (7-AAD), and internal control (all of these reagents are in the Stem-Kit; BeckmanCoulter, Marseille, France). After incubation, samples were centrifuged, and supernatant was removed to obtain concentrated cell suspensions. 7-Aminoactinomycin-D-positive dead cells and CD45-negative cells were excluded, and the number of cells forming clusters characteristic of CD34+ cells (i.e., low side scatter and low-to-intermediate CD45 staining) was counted. The absolute number of CD34+ cells was calculated using the internal control. Mean cell number of duplicate measurements was used for quantitative analysis. Statistical comparisons among groups were determined using analysis of variance or χ2 test. Individual comparisons were performed using a two-tailed unpaired Students’ t-test or Mann—Whitney's U-test. Mean±s.e. is shown.

Results

Enrolled individuals were divided into three groups: control subjects, patients with cerebral occlusion or severe stenosis, but without the presence of vessels with angiographic characteristics of moyamoya disease, and patients with angiographic evidence of moyamoya-like vessels. Baseline characteristics of the groups are shown in Table 1. The modified Rankin scale evaluation of patients with and without moyamoya-like vessels was 0.5±0.5 and 1.3±0.2, respectively (P=0.15). Comparing these groups, there was a significant difference in the ratio of gender and treatment with aspirin between groups. However, no significant difference was observed in the number of circulating CD34+ cells in control group between genders (male, n=13, CD34+ cells=0.93±0.10/μL; female, n=13, CD34+ cells=0.85±0.11/μL: P=0.59) and treatment with aspirin (aspirin (+), n=6, CD34+ cells=0.76±0.12/μL; aspirin (−), n=20, CD34+ cells=0.93±0.09/μL: P=0.26), indicating mild and nonsignificant effects of gender and treatment with aspirin on the level of circulating CD34+ cells. In univariate analysis of control subjects, each cerebrovascular risk factor and treatment with statins showed no significant difference in the number of circulating CD34+ cells (data not shown).
Table 1 Baseline characteristics
 TotalControlMajor artery occlusion/stenosisP-value for trend
   Moyamoya (−)Moyamoya (+) 
N5026204 
Age, years60.8±1.160.5±1.961.5±1.059.3±5.90.85
Male, n (%)33 (66)13 (50)18 (90)2 (50)0.01
      
Risk factor, n (%)
 Hypertension35 (70)16 (62)15 (75)4 (100)0.24
 Hyperlipidemia26 (52)14 (54)10 (50)2 (50)0.96
 Diabetes mellitus11 (22)7 (27)4 (20)0 (0)0.46
 Smoking15 (30)7 (27)8 (40)0 (0)0.25
      
Treatment, n (%)
 Ca channel blockers20 (40)10 (38)8 (40)2 (50)0.91
 β-Blockers5 (10)3 (11)1 (5)1 (25)0.44
 ACE inhibitor7 (14)4 (15)2 (10)1 (25)0.70
 ARB12 (24)5 (19)5 (25)2 (50)0.40
 Diuretics4 (8)2 (7)1 (5)1 (25)0.40
 Statin therapy14 (28)9 (34)4 (20)1 (25)0.54
 Aspirin19 (38)6 (23)10 (50)3 (75)0.05
 Ticlopidine12 (24)3 (11)8 (40)1 (25)0.08
Abbreviations: ACE, angiotensin-converting enzyme; ARB, angiotensin 2 receptor blocker.
A representative angiogram showing characteristics of moyamoya-like vessels is shown in Figures 1A and 1B. Angiographic moyamoya-like vessels were observed around the M1 portion of an occluded middle cerebral artery. Compared with a normal subject (Figure 1C) and patients without angiographic evidence of moyamoya-like vessels (Figure 1D), a remarkable increase in levels of peripheral CD34+ cells was observed in patients with moyamoya-like vessels (Figure 1E) based on fluorescence-activated cell sorting. To confirm this impression, levels of circulating CD34+ cells were quantified (control, CD34+ cells=0.89±0.07/μL; moyamoya (−), CD34+ cells=0.98±0.13/μL; moyamoya (+), CD34+ cells=2.28±0.53/μL) and found to be significantly increased in patients with moyamoya-like vessels more than two-fold higher than in controls (Figure 1F, P<0.001).
Figure 1 Increased levels of circulating CD34+ cells in patients with angiographic evidence of moyamoya-like vessels. (A, B) Representative angiogram from a patient with moyamoya-like vessels. Unusually accelerated neovascularization (based on angiographic features of moyamoya-like vessels, arrowheads) was observed around an occlusive M1 lesion (arrow). Anterior—posterior view (A) and lateral view (B) of the right internal carotid artery showed angiographically. (C–E) After exclusion of 7-aminoactinomycin-D (7-AAD)-positive dead cells and CD45-negative cells (nonleukocytes), CD34+ cells cluster at low side scatter. Representative fluorescence-activated cell sorting analyses from a control subject (C), a patient without moyamoya-like vessels (D), and a patient with moyamoya-like vessels (E) are shown. (F) A more than two-fold increase in circulating CD34+ cells was observed in patients with moyamoya-like vessels, compared with control subjects and patients without moyamoya-like vessels (∗P<0.001). SS Lin: side-scatter linear scale.

Discussion

In this study, we have found that a feature of unusually accelerated neovascularization, evidence of moyamoya-like vessels in the immediate locale of an occluded major cerebral artery, can be correlated with a robust increase in the level of circulating CD34+ cells. The latter was determined using a newly developed method that enables quantification of few CD34+ cells in peripheral blood in a highly reproducible manner.
After acute cerebral ischemia, mobilization of CD34+ cells from bone marrow has been shown in stroke patients (Taguchi et al, 2004a). Furthermore, transplantation of CD34+ cells (Taguchi et al, 2004b) and bone marrow cells (Borlongan et al, 2004a, 2004b) has been shown to restore cerebral blood flow in experimental models of stroke. In chronic ischemia, transplantation of CD34+ cells has also been shown to accelerate neovascularization, including formation of collateral vessels, in patients with chronic ischemic heart disease (Boyle et al, 2006) and limb ischemia (Kudo et al, 2003). In addition, there is a report regarding the correlation between inadequate coronary collateral development and reduced numbers of circulating endothelial progenitor cells in patients with myocardial ischemia (Lambiase et al, 2004). In this study, we show, for the first time, a correlation between neovascularization of the cerebral arterial circulation and increased levels of circulating CD34+ cells. Our results support the hypothesis that circulating CD34+ cells potentially contribute to neovascularization at sites of ischemic brain injury.

Acknowledgements

We thank K Obata and Y Okinaka for technical assistance.

Footnote

The authors state no conflict of interest.

References

Borlongan CV, Lind JG, Dillon-Carter O, Yu G, Hadman M, Cheng C, Carroll J, Hess DC (2004a) Bone marrow grafts restore cerebral blood flow and blood brain barrier in stroke rats. Brain Res 1010:108–16.
Crossref
PubMed
ISI
Google Scholar
Borlongan CV, Lind JG, Dillon-Carter O, Yu G, Hadman M, Cheng C, Carroll J, Hess DC (2004b) Intracerebral xenografts of mouse bone marrow cells in adult rats facilitate restoration of cerebral blood flow and blood–brain barrier. Brain Res 1009:26–33.
Crossref
PubMed
ISI
Google Scholar
Boyle AJ, Whitbourn R, Schlicht S, Krum H, Kocher A, Nandurkar H, Bergmann S, Daniell M, O'Day J, Skerrett D, Haylock D, Gilbert RE, Itescu S (2006) Intra-coronary high-dose CD34+ stem cells in patients with chronic ischemic heart disease: a 12-month follow-up. Int J Cardiol 109:21–7.
Crossref
PubMed
ISI
Google Scholar
Kudo FA, Nishibe T, Nishibe M, Yasuda K (2003) Autologous transplantation of peripheral blood endothelial progenitor cells (CD34+) for therapeutic angiogenesis in patients with critical limb ischemia. Int Angiol 22:344–8.
PubMed
ISI
Google Scholar
Lambiase PD, Edwards RJ, Anthopoulos P, Rahman S, Meng YG, Bucknall CA, Redwood SR, Pearson JD, Marber MS (2004) Circulating humoral factors and endothelial progenitor cells in patients with differing coronary collateral support. Circulation 109:2986–92.
Crossref
PubMed
ISI
Google Scholar
Majka M, Janowska-Wieczorek A, Ratajczak J, Ehrenman K, Pietrzkowski Z, Kowalska MA, Gewirtz AM, Emerson SG, Ratajczak MZ (2001) Numerous growth factors, cytokines, and chemokines are secreted by human CD34(+) cells, myeloblasts, erythroblasts, and mega-karyoblasts and regulate normal hematopoiesis in an autocrine/paracrine manner. Blood 97:3075–85.
Crossref
PubMed
ISI
Google Scholar
Natori Y, Ikezaki K, Matsushima T, Fukui M (1997) ‘Angiographic moyamoya’ its definition, classification, and therapy. Clin Neurol Neurosurg 99(Suppl 2): S168–72.
Crossref
PubMed
ISI
Google Scholar
Taguchi A, Matsuyama T, Moriwaki H, Hayashi T, Hayashida K, Nagatsuka K, Todo K, Mori K, Stern DM, Soma T, Naritomi H (2004a) Circulating CD34-positive cells provide an index of cerebrovascular function. Circulation 109:2972–5.
Crossref
PubMed
ISI
Google Scholar
Taguchi A, Matsuyama T, Nakagomi T, Shimizu Y, Fukunaga R, Tatsumi Y, Yoshikawa H, Kikuchi-Taura A, Soma T, Moriwaki H, Nagatsuka K, Stern DM, Naritomi H (2007) Circulating CD34-positive cells provide a marker of vascular risk associated with cognitive impairment. J Cereb Blood Flow Metab; e-pub ahead of print 8 August 2007.
Google Scholar
Taguchi A, Ohtani M, Soma T, Watanabe M, Kinosita N (2003) Therapeutic angiogenesis by autologous bone-marrow transplantation in a general hospital setting. Eur J Vase Endovasc Surg 25:276–8.
Crossref
Google Scholar
Taguchi A, Soma T, Tanaka H, Kanda T, Nishimura H, Yoshikawa H, Tsukamoto Y, Iso H, Fujimori Y, Stern DM, Naritomi H, Matsuyama T (2004b) Administration of CD34+ cells after stroke enhances neurogenesis via angiogenesis in a mouse model. J Clin Invest 114:330–8.
Crossref
PubMed
ISI
Google Scholar

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Published In

Volume 28, Issue 6
Pages: 1086 - 1089
Article first published online: January 30, 2008
Issue published: June 2008

Keywords

  1. antigens
  2. CD34
  3. moyamoya vessel
  4. neovascularization

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© 2008 ISCBFM.
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PubMed: 18231114

Authors

Affiliations

Tomoyuki Yoshihara
Department of Cerebrovascular Disease, National Cardiovascular Center, Osaka, Japan
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Akihiko Taguchi
Department of Cerebrovascular Disease, National Cardiovascular Center, Osaka, Japan
[email protected]
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Tomohiro Matsuyama
Institute for Advanced Medical Sciences, Hyogo College of Medicine, Hyogo, Japan
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Yoko Shimizu
Department of Cerebrovascular Disease, National Cardiovascular Center, Osaka, Japan
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Akie Kikuchi-Taura
Department of Hematology, Osaka Minami National Medical Center, Osaka, Japan
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Toshihiro Soma
Department of Hematology, Osaka Minami National Medical Center, Osaka, Japan
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David M Stern
Dean's Office, College of Medicine, University of Cincinnati, Cincinnati, Ohio, USA
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Hiroo Yoshikawa
Department of Internal Medicine, Hyogo College of Medicine, Hyogo, Japan
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Yukiko Kasahara
Department of Cerebrovascular Disease, National Cardiovascular Center, Osaka, Japan
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Hiroshi Moriwaki
Department of Cerebrovascular Disease, National Cardiovascular Center, Osaka, Japan
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Kazuyuki Nagatsuka
Department of Cerebrovascular Disease, National Cardiovascular Center, Osaka, Japan
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Hiroaki Naritomi
Department of Cerebrovascular Disease, National Cardiovascular Center, Osaka, Japan
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Notes

This work was supported by a Grant-in-Aid for Scientific Research from the Ministry of Health, Labour, and Welfare (H19-Choujyu-029).
Correspondence: Dr A Taguchi, Department of Cerebrovascular Disease, National Cardiovascular Center, 5-7-1 Fujishiro-dai, Suita, Osaka 565-8565, Japan. E-mail: [email protected]

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