J Korean Med Sci.  2014 Nov;29(Suppl 3):S237-S248. 10.3346/jkms.2014.29.S3.S237.

Effect of Endogenous Bone Marrow Derived Stem Cells Induced by AMD-3100 on Expanded Ischemic Flap

• Jeong HS
• Lee HK
• Tark KC
• Lew DH
• Koh YW
• Kim CH
• Seo IS

• KMID: 2151419
• DOI: http://doi.org/10.3346/jkms.2014.29.S3.S237

Abstract

The purpose of this study was to devise an expanded ischemic flap model and to investigate the role of AMD-3100 (Plerixafor, chemokine receptor 4 inhibitor) in this model by confirming its effect on mobilization of stem cells from the bone marrow. Male Sprague-Dawley rats were used as an animal research model. The mobilization of stem cells from the bone marrow was confirmed in the AMD-3100-treated group. The fractions of endothelial progenitor cells (EPC) and the vascular endothelial growth factor receptor (VEGFR) 2+ cells in the peripheral blood were increased in groups treated with AMD-3100. The expression of vascular endothelial growth factor (VEGF) was increased in response to expansion or AMD injection. The expression of stromal cell derived factor (SDF)-1 and VEGFR2 were increased only in unexpanded flap treated with AMD-3100. Treatment with AMD-3100 increased both the number and area of blood vessels. However, there were no statistically significant differences in the survival area or physiologic microcirculation in rats from the other groups. This endogenous neovascularization induced by AMD-3100 may be a result of the increase in both the area and number of vessels, as well as paracrine augmentation of the expression of VEGF and EPCs. However, the presence of a tissue expander under the flap could block the neovascularization between the flap and the recipient regardless of AMD-3100 treatment and expansion.

Keyword

Endothelial Progenitor Cell; Hematopoietic Stem Cell; Tissue Expansion; Ischemic Flap; AMD-3100; CXCR4 Inhibitor; Plerixafor

MeSH Terms

Animals
Anti-HIV Agents/pharmacology
Bone Marrow Cells/cytology
Chemokine CXCL12/biosynthesis
Endothelial Progenitor Cells/*cytology
Hematopoietic Stem Cells/*cytology
Heterocyclic Compounds/*pharmacology
Hypoxia-Inducible Factor 1, alpha Subunit/metabolism
Male
Neovascularization, Physiologic
Nitric Oxide Synthase Type III/metabolism
Rats
Rats, Sprague-Dawley
Receptors, CXCR4/antagonists & inhibitors
Surgical Flaps/*blood supply/surgery
Tissue Expansion/*methods
Vascular Endothelial Growth Factor A/biosynthesis
Vascular Endothelial Growth Factor Receptor-2/biosynthesis/metabolism
Anti-HIV Agents
Chemokine CXCL12
Heterocyclic Compounds
Hypoxia-Inducible Factor 1, alpha Subunit
Receptors, CXCR4
Vascular Endothelial Growth Factor A
Nitric Oxide Synthase Type III
Vascular Endothelial Growth Factor Receptor-2

Figure

• 

  Fig. 1 Expanded skin flap animal model. (A) Design of expanded ischemic random pattern skin flap. (B) Edge of the distal flap removed for skin expansion. Flap elevation above the submuscular connective tissue. (C) Silicone placement on the floor. (D) Final suture state.

• 

  Fig. 2 Flap survival in the preliminary study of unexpanded flaps. (A) Gross findings of the skin flaps. GA, Group A, control without silicone sheet; GB, Group B, control with silicone sheet; GC, Group C, AMD-3100 injection without silicone sheet; GD, Group D, AMD-3100 injection with silicone sheet. (B) Treatment with AMD-3100 significantly increased survival rates in classical unexpanded flaps, with or without insertion of a silicone sheet. (C) Insertion of a silicone sheet under the flap blocked the blood supply from the bed. In a classical unexpanded flap, the mean survival rate of flaps without insertion of a silicone sheet was greater than those with a silicone sheet, regardless of treatment with AMD-3100 (*P<0.05, Mann-Whitney test).

• 

  Fig. 3 Immunohistochemical staining for CD31 (A, 200×). Arrows indicate vessels. Vessel areas (B) and counts (C) by Group III and IV (received AMD-3100 injection) showed statistically significant increases in vessel number compared with group I (control) (*P<0.05, Mann-Whitney test).

• 

  Fig. 4 Immunohistochemical staining (A) for VEGF (200×) and density of VEGF in flap tissue (B) (*P<0.05, Kruskal-Wallis test).

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  Fig. 5 Real time quantitative RT-PCR measurement of VEGFA, VEGFR2, NOS3, SDF-1 and HIF-1 (*P<0.05, significant difference compared between control levels of group I by Mann-Whitney test).

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