Placenta mesenchymal stem cell accelerates wound healing by enhancing angiogenesis in diabetic Goto-Kakizaki (GK) rats

https://doi.org/10.1016/j.bbrc.2013.07.088Get rights and content

Highlights

  • PMSCs enhanced the extent of wound closure in diabetic GK rats.

  • Transplanted PMSCs localized to the wound tissue and were incorporated into recipient vasculature.

  • PMSCs secreted comparable amounts of proangiogenic molecules at bioactive levels.

Abstract

Multipotent mesenchymal stem cells have recently emerged as an attractive cell type for the treatment of diabetes-associated wounds. The purpose of this study was to examine the potential biological function of human placenta-derived mesenchymal stem cells (PMSCs) in wound healing in diabetic Goto-Kakizaki (GK) rats. PMSCs were isolated from human placenta tissue and characterized by flow cytometry. A full-thickness circular excisional wound was created on the dorsum of each rat. Red fluorescent CM-DiI-labeled PMSCs were injected intradermally around the wound in the treatment group. After complete wound healing, full-thickness skin samples were taken from the wound sites for histological evaluation of the volume and density of vessels. Our data showed that the extent of wound closure was significantly enhanced in the PMSCs group compared with the no-graft controls. Microvessel density in wound bed biopsy sites was significantly higher in the PMSCs group compared with the no-graft controls. Most surprisingly, immunohistochemical studies confirmed that transplanted PMSCs localized to the wound tissue and were incorporated into recipient vasculature with improved angiogenesis. Notably, PMSCs secreted comparable amounts of proangiogenic molecules, such as VEGF, HGF, bFGF, TGF-β and IGF-1 at bioactive levels. This study demonstrated that PMSCs improved the wound healing rate in diabetic rats. It is speculated that this effect can be attributed to the PMSCs engraftment resulting in vascular regeneration via direct de novo differentiation and paracrine mechanisms. Thus, placenta-derived mesenchymal stem cells are implicated as a potential angiogenesis cell therapy for repair-resistant chronic wounds in diabetic patients.

Introduction

Diabetes mellitus is increasing in incidence and represents a major health problem in the twenty-first century. Indeed, the total number of diabetic patients has been projected to increase from 171 million in 2000 to 366 million in 2030 [1]. Chronic skin ulcers are one of the most serious consequences of diabetes, representing a major contributing factor to amputation in diabetics [2]. The annual incidence of foot ulcers among diabetic patients has been variously estimated at between 1% and 4.1%, and the annual incidence of amputation is in the range 0.21% and 1.37% [3]. The economic and social costs associated with chronic wounds are enormous and include hospital costs, disability, decreased productivity and loss of independence. A number of factors have been implicated in the predisposition to non-healing wounds observed in diabetes, including microvascular disease, peripheral neuropathy and impaired angiogenesis of wounds [4]. Of these factors, angiogenesis is considered to play a pivotal pathophysiological role by virtue of its requirement in successful wound repair.

Human mesenchymal stem cells (MSCs)-based therapeutic angiogenesis has emerged as a promising strategy in regenerative medicine, including in the treatment of myocardial infarction [5], limb ischemia [6] and chronic skin ulcers [7], [8]. Particularly, bone marrow MSCs (BM-MSCs) [9] and endothelial progenitor cells (EPCs) [10] have been subjected to the most comprehensive translational and human studies of all stem cell approaches in wound healing. However, the use of BM-MSCs and EPCs is associated with some deficiencies. The survival and differentiation potential of BM-MSCs or progenitor cells obtained from aged patients or patients with age-related disorders are impaired, thus limiting their therapeutic efficiency [11], [12]. Furthermore, autologous delivery of BM-MSCs or EPCs is inevitably associated with a delay in treatment due to time required for cell collection, isolation and propagation of sufficient numbers for injection [13]. Therefore, there is emerging interest in the identification of alternative cell sources of MSCs.

In the present study, a population of multipotent stem cells was isolated from human term placenta, a temporary organ with fetal contributions that is discarded postpartum. These placenta mesenchymal stem cells (PMSCs) display typical mesenchymal characteristics such as great capacity for self-renewal while maintaining their multipotent differentiation potential and expression of common MSCs surface markers similar to those expressed by BM-MSCs. We hypothesized that PMSCs administration may restore wound healing in pathological conditions. Therefore, the effect of PMSCs transplantation on the wound healing rate in diabetic Goto-Kakizaki (GK) rats was evaluated. Furthermore, the mechanisms underlying the beneficial effects of PMSCs were assessed by analysis of their capacity of differentiation into endothelial cells and secretion of proangiogenic factors.

Section snippets

Patient selection and tissue processing

With consent, fresh placentas were collected from normal, full-term (38–40 weeks gestation), healthy donor mothers according to the regulations of the Independent Ethics Committee of the Shanghai Ninth People’s Hospital affiliated with Shanghai JiaoTong University School of Medicine. Written informed consent was signed prior to the study. Umbilical cord blood was allowed to drain from the placentas, which were then dissected carefully. All tissues were tested to exclude HIV-infection,

Characterization of PMSCs

We began our studies with the derivation and passaging of PMSCs as described under methods. To characterize the phenotypes of PMSCs, flow cytometry was performed to analyze surface markers of PMSCs. In this study, most of the PMSCs strongly expressed CD29, CD90, CD73, CD105 and CD49b but were negative for HLA-DR, CD45, and CD34. The immunophenotype of PMSCs remained unchanged for more than eight cell passages. PMSCs were also shown to give rise to adipogenic, chondrogenic and osteogenic

Discussion

The major goal of this study was to investigate the impact of transplanted PMSCs on the wound healing process in diabetic rats. To this end, our results demonstrate for the first time that PMSCs transplantation remarkably increased the wound healing rate and improved the condition of the scar by histological evaluation as well as by surface inspection. The greater potential of PMSCs may be related to the vascular differentiation capacity of these cells, as well as paracrine mechanisms. Local

Acknowledgments

This work was supported by the National Natural Science Foundation of China (Grant Nos. 81000345, 81200244) and by the Shanghai Natural Science Foundation (Grant Nos. 09ZR1417100 and 12ZR1428400). We thanks very much for the English editing by Medsci company.

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    Poren Kong (Chinese name: Boren Jiang) and Xiaoyun Xie contributed equally to this work.

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