Dale S. Feldman

Experimental and clinical applications of fibrin glue.

A 2-year experience with laboratory and clinical applications of fibrin glue is presented. An autologous technique, which eliminates the danger of multidonor preparations, has been developed in our blood bank. While one can obtain different fibrinogen concentrations from the same amount of a patients blood, in vitro mechanical testing demonstrated that at higher fibrinogen concentrations there is an increase in shear adhesive strength. Evaluation of skin-graft take in 16 Sprague-Dawley rats did not demonstrate significant differences in healing when adhesive use was compared with suture technique. In a clinical study, four different groups of patients (facial burns, hand burns, difficult graft sites, and miscellaneous surgical applications) benefited from autologous or single-donor fibrin glue for a total of 82 cases. There are several distinct advantages to the use of fibrin adhesive: The autologous technique eliminates the risk of transmissible viral diseases (AIDS, hepatitis); it can be used as a sealant in the treatment of seromas, dural leaks, and lymphoceles; and it improves hemostasis and early graft adherence. Face and hands are resurfaced with sheet grafts in a single procedure, obtaining a better aesthetic result with complete graft take and immediate start of physical therapy. Neither sutures nor pressure dressings are required. The minimal postoperative care associated with early return to normal activities seems to increase the satisfaction of patients and nurse personnel.

Promotion of incisional wound repair by human mesenchymal stem cell transplantation.

Abstract:  The purpose of this study was to determine the effect of transplanted human mesenchymal stem cells (hMSCs) on wound healing. In this model, full‐thickness cutaneous wounds were created by incision in the skin of adult New Zealand white rabbits and treated by transplanted hMSCs into the wounds. Wound healing was evaluated by histological analysis and tensiometry over time. A total of 15 New Zealand white rabbits with 10 wounds per animal were examined in this study. Animals were treated with hMSCs and euthanised at 3, 7, 14, 21 and 80 days after manipulation. The hMSCs were labelled with a fluorescent dye (CM‐DiI), suspended in phosphate‐buffered saline and used to treat full‐thickness incisional wounds in rabbit skin. Tensiometry and histology were used to characterise the wound‐healing rate of the incisional wounds. These results showed that transplanted hMSCs significantly inhibited scar formation and increased the tensile strength of the wounds. Importantly, MSCs from genetically unrelated donors did not appear to induce an immunologic response. In conclusion, human mesenchymal stem cell therapy is a viable approach to significantly affect the course of normal cutaneous wound healing and significantly increase the tensile strength.

The effect of TGF-beta delivered through a collagen scaffold on wound healing.

In this preliminary study, the wound healing response of full-thickness skin defects to transforming growth factor beta (TGF-beta) incorporated in a collagen scaffold was evaluated. The objectives of this study were (1) to compare the effects of TGF-beta on the cellular and tissue response and mechanical properties and (2) to determine the effects of a collagen scaffold on wound healing when compared to control. Three 3 x 3 cm, full-thickness defects were created on the dorsi of 15 New Zealand White rabbits. Each rabbit had a control (no treatment), collagen scaffold, and collagen scaffold with TGF-beta (2 microg/cm2). All the wounds were covered with a transparent polyurethane dressing. There were three periods of study (1, 2, and 3 weeks) with five rabbits in each period. The volume fraction of the tissue and cells was histomorphometrically determined for each wound. A greater inflammatory response was found in the collagen scaffold-treated group, but the fastest epithelialization and contraction rates were associated with TGF-beta and collagen. No significant differences in the mechanical strength between the different treatment groups were seen. Overall, TGF-beta delivered through a collagen scaffold enhanced the healing process and showed promise for future clinical applications.

Effect of adenoviral mediated overexpression of fibromodulin on human dermal fibroblasts and scar formation in full-thickness incisional wounds

Fibromodulin, a member of the small leucine-rich proteoglycan family, has been recently suggested as a biologically significant mediator of fetal scarless repair. To assess the role of fibromodulin in the tissue remodeling, we constructed an adenoviral vector expressing human fibromodulin cDNA. We evaluated the effect of adenovirus-mediated overexpression of fibromodulin in vitro on transforming growth factors and metalloproteinases in fibroblasts and in vivo on full-thickness incisional wounds in a rabbit model. In vitro, we found that Ad-Fibromodulin induced a decrease of expression of TGF-β1 and TGF-β2 precursor proteins, but an increase in expression of TGF-β3 precursor protein and TGF-β type II receptor. In addition, fibromodulin overexpression resulted in decreased MMP-1 and MMP-3 protein secretion but increased MMP-2, TIMP-1, and TIMP-2 secretion, whereas MMP-9 and MMP-13 were not influenced by fibromodulin overexpression. In vivo evaluation by histopathology and tensile strength demonstrated that Ad-Fibromodulin administration could ameliorate wound healing in incisional wounds. In conclusion, although the mechanism of scar formation in adult wounds remains incompletely understood, we found that fibromodulin overexpression improves wound healing in vivo, suggesting that fibromodulin may be a key mediator in reduced scarring.

Fibrin Scaffold as an Effective Vehicle for the Delivery of Acidic Fibroblast Growth Factor (FGF-1):

The effect of wound healing by fibrin and acidic fibroblast growth factor (FGF-1) in an in vivo model was evaluated in this study. Four full-thickness wounds were made on the dorsum of each rabbit (n = 5). Each of these wounds had different treatment groups: control, topical FGF-1 (100 μg/9 cm2), fibrin (2.0 mL at 60 mg/mL fibrinogen), and FGF-1 (100 gg/9 cm2)/fibrin. The animals were sacrificed at the end of 2 weeks. Histomorphometric analysis and mechanical testing were conducted to assess the healing response. FGF-1/fibrin treatment improved the mechanical properties of the healed tissue. Fibrin scaffold exhibited the desired tissue response, as demonstrated by the lack of inflammation, and was deemed an effective carrier for FGF-1.

Analysis of a biodegradable composite for bone healing

A degradable L-PLA/calcium carbonate composite made of interconnecting phases was examined. This structure was used both to slow the degradation rate and to reduce the brittleness of the ceramic. Both in vitro and in vivo degradation studies were performed. Samples were incubated in buffered saline or placed in the dorsum of rats for 0, 1, or 4 weeks. Mechanical testing was performed on both groups, volume fraction of each component was determined for in vitro samples, and histology was performed on in vivo samples. Failure load, tensile strength, and elastic modulus significantly decreased during the 1st week for both groups. Continued decreases were seen at 4 weeks for in vitro samples but not for in vivo. Failure strain and tensile strength decreased only for in vitro specimens. PLA fraction significantly decreased during the 1st week and then stabilized. Histology showed that tissue ingrowth occurred at 4 weeks. The decrease in mechanical properties was probably a result of the decreased PLA fraction. The stabilization and even a slight increase in tensile strength and failure strain in the in vivo samples was probably due to the tissue ingrowth forming an implant-tissue composite.

Investigation of acidic fibroblast growth factor delivered through a collagen scaffold for the treatment of full-thickness skin defects in a rabbit model.

&NA; In this study the wound healing response of full‐thickness skin defects was enhanced through the use of acidic fibroblast growth factor using a collagen scaffold. The objectives of this study were (1) to compare the facilitatory effects of acidic fibroblast growth factor on the cellular response (fibroblast, neutrophil, macrophage, epithelial), tissue response (angiogenesis, collagen) and mechanical properties in a healing wound; and (2) to determine the effect of a collagen scaffold on wound healing when compared with the control. Three 3 × 3 cm full‐thickness defects were created on the dorsi of 15 New Zealand White rabbits. Each rabbit had a control (no treatment), collagen scaffold, and collagen scaffold with FGF‐1 (100 &mgr;g/ cm2). All the wounds were covered with a transparent polyurethane dressing. There were three periods of study (1, 2, and 3 weeks) with five rabbits in each period. The volume fraction of the tissue and cells was histomorphometrically determined for each wound. The acidic fibroblast growth factor/collagen system showed promise in enhancing the healing process. Acidic fibroblast growth factor/collagen treatment increased angiogenesis, enhanced epithelialization, and reduced contraction rate over the control. A higher inflammatory response was indicated in the collagen scaffold treated group. Hence acidic fibroblast growth factor delivered through a collagen scaffold shows promise in future clinical applications.

Stimulation of Angiogenesis by FGF-1 Delivered Through a Modified Fibrin Scaffold

A few studies have indicated that repeated dosing of acidic fibroblast growth factor (FGF-1) is essential to be effective in modulating the wound-healing response. However, little investigation has been done to determine the effective dosing regimen of FGF-1 or the appropriate carrier vehicle for this growth factor. The main objective of this study was to determine the effective angiogenic stimulatatory dose of FGF-1 delivered through a modified fibrin matrix, using a rabbit ear ulcer model. Specifically, the aim was to test the effects of FGF-1 on the angiogenic, fibroblastic, and epithelial responses in a wound model. Five 6-mm diameter ulcers to the depth of bare cartilage were created on each rabbit ear. Four different combinations (0.8, 8, 80, and 800 micrograms/ml) of the growth factor were examined across two periods of study. Pooled modified fibrin was used to deliver the growth factor. Histomorphometrical quantification was conducted after routine histological processing of the ulcers sites. Data analysis indicated a strong correlation between concentration and the histomorphometric response. In general, the growth factor treatments affected the healing response and exhibited a dose-dependent behavior. The addition of FGF-1 led to an increase in the angiogenic and fibroblastic responses, as well as an increase in the epithelialization rate. The preferred dose of 8 micrograms initiated a high epithelialization rate, fibroblastic, and angiogenic responses, and was the lowest dose required to initiate these responses.

In Vivo Wound Healing Response to a Modified Degradable Fibrin Scaffold

Pooled donor fibrin with an ultimate fibrinogen concentration of 60 mg/ml was used to study its effect on wound healing of surgically created ulcers in a rabbit ear. Water soluble polymer (PEG Mw = 20 KD) beads of 100-150 μm were added (12% by volume) to the fibrinogen to obtain a porous and rough structure. Five 6 mm-diameter ulcers to the depth of bare cartilage were created on each rabbit ear. There were two periods of study (4 and 8 days), with 15 ulcers in each time period, 5 of which were treated with a modified fibrin scaffold, 5 with a non-modified fibrin scaffold, and 5 served as control ulcers. The ulcer sites were subjected to routine histological processing and histomorphometrical quantification. Data analysis revealed significant increases in volume fraction of fibroblast and number of blood vessels in the modified fibrin scaffold treated ulcers over control and non-modified fibrin scaffold treated groups.

Modification of fibrinogen with poly(ethylene glycol) and its effects on fibrin clot characteristics

The suitability of existing topical fibrin glue preparations for tissue sealing or local drug delivery applications is greatly limited by their poor mechanical properties and the limited capacity of fibrinogen (Fgn) to actively bind growth factors or other therapeutic agents. Poly(ethylene glycol) (PEG) offers potential solutions to these problems by providing a mechanism for increasing the number of crosslinks between adjacent fibrin monomer molecules or for covalently crosslinking Fgn to therapeutic agents. The feasibility of this approach requires the full biological activity, or clottability, of PE glycolated Fgn. This study characterizes the clot characteristics of Fgn modified to varying degrees with monofunctional succinimidyl propionate PEG (5000 Da). The data indicate that, although thrombin clotting times are significantly altered, Fgn maintains 90% of its capacity to clot upon the addition of up to 5 PEG/Fgn. Further derivatization significantly decreases the Fgn clottability. The addition of up to 5 PEG/Fgn has little, if any, effect on the kinetics of degradation by plasmin. The results suggest that limited modification of Fgn with lysine-reactive PEG allows therapeutic enhancement of fibrin glues.