Autologous Bone Marrow–Derived Cultured Mesenchymal Stem Cells Delivered in a Fibrin Spray Accelerate Healing in Murine and Human Cutaneous Wounds

Abstract

The nonhematopoietic component of bone marrow includes multipotent mesenchymal stem cells (MSC) capable of differentiating into fat, bone, muscle, cartilage, and endothelium. In this report, we describe the cell culture and characterization, delivery system, and successful use of topically applied autologous MSC to accelerate the healing of human and experimental murine wounds. A single bone marrow aspirate of 35-50 mL was obtained from patients with acute wounds (n = 5) from skin cancer surgery and from patients with chronic, long-standing, nonhealing lower extremity wounds (n = 8). Cells were grown in vitro under conditions favoring the propagation of MSC, and flow cytometry and immunostaining showed a profile (CD29+, CD44+, CD105+, CD166+, CD34-, CD45-) highly consistent with published reports of human MSC. Functional induction studies confirmed that the MSC could differentiate into bone, cartilage, and adipose tissue. The cultured autologous MSC were applied up to four times to the wounds using a fibrin polymer spray system with a double-barreled syringe. Both fibrinogen (containing the MSC) and thrombin were diluted to optimally deliver a polymerized gel that immediately adhered to the wound, without run-off, and yet allowing the MSC to remain viable and migrate from the gel. Sequential adjacent sections from biopsy specimens of the wound bed after MSC application showed elongated spindle cells, similar to their in vitro counterparts, which immunostained for MSC markers. Generation of new elastic fibers was evident by both special stains and antibodies to human elastin. The application of cultured cells was safe, without treatment-related adverse events. A strong direct correlation was found between the number of cells applied (greater than 1 x 10(6) cells per cm2 of wound area) and the subsequent decrease in chronic wound size (p = 0.0058). Topical application of autologous MSC also stimulated closure of full-thickness wounds in diabetic mice (db/db). Tracking of green fluorescent protein (GFP)+ MSC in mouse wounds showed GFP+ blood vessels, suggesting that the applied cells may persist as well as act to stimulate the wound repair process. These findings indicate that autologous bone marrow-derived MSC can be safely and effectively delivered to wounds using a fibrin spray system.

Activity of mesenchymal stem cells in therapies for chronic skin wound healing

“…As MSCs play a normal role in the wound healing process, they are an obvious candidate for study in this context as opposed to embryonic stem cells or other regenerative sources. Recent studies have outlined some successful approaches to promoting wound healing with MSCs, including autologous bone marrow-derived MSCs in fibrin matrix 52 or, more recently, intramuscular injection of autologous MSCs to improve diabetic wound closure. 54 While some trials have been aimed primarily at safety of MSC use for wound healing, 50 several clinical trials have shown the potential benefit of MSCs for inclusion in wound healing devices, including improved average rate of wound healing and general limb perfusion after treatment 53 and also improved acute wound healing correlating to the number of injected cells.…”

67: Longterm in-Vivo Tumorigenic Assessment of Human Culture-Expanded Adipose Stromal/Stem Cells

“…Previous attempts to determine the migration and persistence of stem cells in vivo include GFP labeling methods and reporter gene analysis [30][31][32][33]. These studies either noted a lack of stem cell persistence, or experienced time-dependent limits of assay efficacy [33][34][35].…”

Do Mesenchymal Stem Cells Have a Role to Play in Cutaneous Wound Healing?

“…56 MSCs utilized from more easily accessible sites like adipose tissue found to have a similar healing potential. For instance, in a mouse wound healing model, adipose tissue MSCs loaded on collagen gel resulted in acceleration of the healing, enhancement of the secretion of type I collagen and the extracellular matrix proteins.…”

“…Falanga et al implanted autologous BM-MSCs loaded in a fibrin spray in five patients with acute wounds from skin cancer surgery and eight patients with chronic, long-standing, non-healing lower extremity wounds. 56 The application of MSCs has stimulated the wound healing process leading to a decrease in size or healing within 20 weeks. It is of note that a strong correlation between the number of cells applied and the subsequent decrease in chronic wound size was noted.…”