Development of a Wound Dressing Composed of Hyaluronic Acid and Collagen Sponge with Epidermal Growth Factor

This study was designed to investigate the effect of a wound dressing composed of hyaluronic acid (HA) and collagen (Col) sponge containing epidermal growth factor (EGF) on various parameters of wound healing in vitro and in vivo. High-molecular-weight (HMW) HA solution, hydrolyzed low-molecular-weight (LMW) HA solution and heat-denatured Col solution were mixed, followed by freeze-drying to obtain a spongy sheet. Cross-linkage between Col molecules was induced by UV irradiation to the spongy sheet (Type-I dressing). In a similar manner, a spongy sheet containing EGF was prepared (Type-II dressing). The efficacy of these products was firstly evaluated in vitro. Fibroblast proliferation was assessed in culture medium in the presence or absence of a piece of each wound dressing. EGF stimulated cell proliferation after UV irradiation and dry sterilization at 110°C for 1 h. In the second experiment, fibroblasts-embedded Col gels were elevated to the air–liquid interface to create a wound surface model, on which wound dressings were placed and cultured for 1 week. Cell proliferation and the production of vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) were investigated. With Type-II dressings, the amounts of VEGF and HGF released from fibroblasts in the Col gel were significantly increased compared with Type-I dressing. Next, the efficacy of these products was evaluated in vivo using Sprague–Dawley (SD) rats. Wound conditions after 1 and 2 weeks of treatment with the wound dressings were evaluated based on the gross and histological appearances. Type-II dressings promoted a decrease in wound size, re-epithelialization and granulation tissue formation associated with angiogenesis. These findings indicate that the combination of HA, Col and EGF promotes wound healing by stimulating fibroblast function.     

Development of a wound dressing composed of hyaluronic acid and collagen sponge with epidermal growth factor

This study was designed to investigate the effect of a wound dressing composed of hyaluronic acid (HA) and collagen (Col) sponge containing epidermal growth factor (EGF) on various parameters of wound healing in vitro and in vivo. High-molecular-weight (HMW) HA solution, hydrolyzed low-molecular-weight (LMW) HA solution and heat-denatured Col solution were mixed, followed by freeze-drying to obtain a spongy sheet. Cross-linkage between Col molecules was induced by UV irradiation to the spongy sheet (Type-I dressing). In a similar manner, a spongy sheet containing EGF was prepared (Type-II dressing). The efficacy of these products was firstly evaluated in vitro. Fibroblast proliferation was assessed in culture medium in the presence or absence of a piece of each wound dressing. EGF stimulated cell proliferation after UV irradiation and dry sterilization at 110°C for 1 h. In the second experiment, fibroblasts-embedded Col gels were elevated to the air-liquid interface to create a wound surface model, on which wound dressings were placed and cultured for 1 week. Cell proliferation and the production of vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) were investigated. With Type-II dressings, the amounts of VEGF and HGF released from fibroblasts in the Col gel were significantly increased compared with Type-I dressing. Next, the efficacy of these products was evaluated in vivo using Sprague-Dawley (SD) rats. Wound conditions after 1 and 2 weeks of treatment with the wound dressings were evaluated based on the gross and histological appearances. Type-II dressings promoted a decrease in wound size, re-epithelialization and granulation tissue formation associated with angiogenesis. These findings indicate that the combination of HA, Col and EGF promotes wound healing by stimulating fibroblast function.     

Development of novel wound dressing composed of hyaluronic acid and collagen sponge containing epidermal growth factor and vitamin C derivative

This study was designed to investigate the potential of a wound dressing composed of hyaluronic acid (HA) and collagen (Col) spongy sheet containing epidermal growth factor (EGF) and vitamin C derivative (VC). High-molecular-weight HA aqueous solution, hydrolyzed low-molecular-weight HA aqueous solution and heat-denatured Col aqueous solution were mixed, followed by freeze-drying to obtain a spongy sheet. Cross-linkage between Col molecules was induced by UV irradiation of the spongy sheet (C-wound dressing). In a similar manner, three types of spongy sheet containing EGF (EGF-wound dressing), containing VC (VC-wound dressing) or containing EGF and VC (EGF·VC-wound dressing) were prepared by freeze-drying the mixed solution containing the specified components. Cytokine production by fibroblasts was assessed in a wound surface model using a fibroblast-incorporating Col gel sheet (cultured dermal substitute; CDS). CDS was elevated to the air–medium interface, onto which each wound dressing was placed and cultured for 7 days. Fibroblasts in CDS covered with EGF-wound dressing released 3.6 times more VEGF and 3.0 times more HGF, as compared with the C-wound dressing. Fibroblasts in CDS covered with EGF·VC-wound dressing released 4.2 times more VEGF and 6.0 times more HGF, as compared with the C-wound dressing. The efficacy of these wound dressings was evaluated in animal tests using diabetic mice. Each wound dressing was applied to a full-thickness skin defect on the dorsal area measuring 1.5 × 2.0 cm. After 1 week of application, wound conditions were evaluated histologically. The EGF·VC-wound dressing more effectively promoted granulation tissue formation associated with angiogenesis, as compared with other wound dressings.     

Development of an artificial dermis composed of hyaluronic acid and collagen

This study aimed to investigate the efficacy of an artificial dermis composed of hyaluronic acid (HA) and collagen (Col) with or without epidermal growth factor (EGF), both in in vitro and in vivo. The cross-linked high molecular weight HA spongy sheet was prepared by freeze-drying. The spongy sheet was immersed in a mixed solution of high molecular weight HA, low molecular weight HA, and heat-denatured Col, and then lyophilized to obtain a two-layered spongy sheet. Cross-linking among Col molecules was induced by ultraviolet irradiation to prepare the artificial dermis (Type I). In a similar manner, a two-layered artificial dermis containing EGF (Type II) was prepared using a similar mixed solution containing EGF. The in vitro experiments demonstrated that EGF released from the Type II artificial dermis stimulates fibroblasts to produce increased amounts of vascular endothelial growth factor and hepatocyte growth factor. The therapeutic efficacy of artificial dermis was evaluated in animal tests using Sprague Dawley (SD) rats. The dorsal skin of the SD rat was shaved and then exposed to boiling water for 3?s to induce a deep dermal burn. The necrotic tissue was then excised 3?days later. Each artificial dermis was applied to the skin defect for 7?days and assessed for its ability to generate a wound bed. The in vivo experiments demonstrated that the Type II artificial dermis promotes angiogenesis to a greater extent at an early stage (within 3?days), and also suppresses the inflammatory reaction more successfully compared with the Type I artificial dermis. In further animal tests, an autologous skin graft was performed by excising a piece of skin from the abdominal region and then grafting it onto the wound bed prepared using each artificial dermis for 7?days. Although the Type II artificial dermis had the highest potential to promote angiogenesis, in this animal study, each artificial dermis induced excellent wound bed formation acceptable for autologous skin grafting.     

Development of a Wound Dressing Composed of Hyaluronic Acid Sponge Containing Arginine and Epidermal Growth Factor

Hyaluronic acid (HA) has the ability to promote wound healing. Epidermal growth factor (EGF) is able to promote the proliferation of various cell types, in addition to epidermal cells. A novel wound dressing was designed using high-molecular-weight hyaluronic acid (HMW-HA) and low-molecular-weight hyaluronic acid (LMW-HA). Spongy sheets composed of cross-linked high-molecular-weight hyaluronic acid (c-HMW-HA) were prepared by freeze-drying an aqueous solution of HMW-HA containing a crosslinking agent. Each spongy sheet was immersed into an aqueous solution of LMW-HA containing arginine (Arg) alone or both Arg and epidermal growth factor (EGF), and were then freeze-dried to prepare two types of product. One was a wound dressing composed of c-HMW-HA sponge containing LMW-HA and Arg (c-HMW-HA/LMW-HA + Arg; Group I). The other was a wound dressing composed of c-HMW-HA sponge containing LMW-HA, Arg and EGF (c-HMW-HA/LMW-HA + Arg + EGF; Group II). The efficacy of these products was evaluated in animal tests using rats. In the first experiment, each wound dressing was applied to a full-thickness skin defect with a diameter of 35 mm in the abdominal region of Sprague–Dawley (SD) rats, leaving an intact skin island measuring 15 mm in diameter in the central area of this skin defect. Commercially available polyurethane film dressing was then applied to each wound dressing as a covering material. In the control group, the wound surface was covered with polyurethane film dressing alone. Both wound dressings (Group I and Group II) potently decreased the size of the full-thickness skin defect and increased the size of the intact skin island, when compared with the control group. The wound dressing in Group II showed particularly potent activity in increasing the distance of epithelization from the intact skin island. This suggests that EGF release from the spongy sheet serves to promote epithelization. The wound dressing in Group II enhanced early-stage inflammation after 1 week, as compared with the other two groups. In the second experiment, each wound dressing was applied to a full-thickness skin defect measuring 35 mm in diameter in the abdominal region of SD rats, after removing necrotic skin caused by dermal burns. Polyurethane film dressing was applied to each wound dressing as a covering material. In the control group, the wound surface was covered with polyurethane film dressing alone. Both wound dressings (Group I and Group II) potently decreased the size of the full-thickness skin defect and increased epithelization from the wound margin, as compared with the control group. The wound dressing in Group II was found to enhance early-stage inflammation after 1 week, as compared with the other two groups. The findings in both experiments indicate that the wound dressing composed of HA-based spongy sheets containing Arg and EGF potently promotes wound healing by inducing moderate inflammation. The release of EGF in the early stages of wound healing induces moderate inflammation. This suggests that wound healing is facilitated directly by topical application of EGF, and indirectly by cytokines derived from inflammatory cells stimulated by EGF.     

Wound dressing composed of hyaluronic acid and collagen containing EGF or bFGF: comparative culture study

We developed a novel wound dressing composed of a hyaluronic acid (HA) and collagen (Col) spongy sheet containing epidermal growth factor (EGF) or basic fibrolast growth factor (bFGF) by freeze-drying method (EGF-wound dressing or bFGF-wound dressing, respectively). A wound dressing without any growth factor was prepared as a control in a similar manner as above (C-wound dressing). Intermolecular cross-linkage between Col molecules was induced by UV irradiation. The release behavior of free HA from the wound dressing was investigated using a C-wound dressing. The weight of C-wound dressing after 1 day, 3, 5, and 7?days of incubation on top of a Col gel sheet at the air–water interface (wound surface model) was 55, 36, 30, and 19% of the original weight, respectively. Most free HA and a part of Col was released from the cross-linked Col network in the wound dressing during incubation, as the original Col content in the wound dressing was 33%. Next, fibroblast proliferation was assessed in conventional culture medium preconditioned by immersion of a piece of C-, EGF-, or bFGF-wound dressing, i.e. C-conditioned medium, EGF-conditioned medium, or bFGF-conditioned medium. Cell proliferation in C-conditioned medium increased to approximately the same level as that in conventional medium. Cell proliferation in EGF- and bFGF-conditioned medium was 1.9 times and 2.6 times greater than that in conventional medium after 7?days of cultivation, respectively. Finally, cytokine production of fibroblasts was assessed in a wound surface model using a fibroblast-incorporating Col gel sheet (cultured dermal substitute [CDS]). CDS was elevated to the air–medium interface, on which each wound dressing was placed and cultured for 7?days. Fibroblasts in CDS covered with EGF-wound dressing released 3.6 times more vascular endothelial growth factor (VEGF) and 4.6 times more hepatocyte growth factor (HGF) when compared with the C-wound dressing. Fibroblasts in CDS covered with bFGF-wound dressing released 10.2 times more VEGF and 6.3 times more HGF when compared with the C-wound dressing. This finding indicates that bFGF-wound dressing can facilitate more effectively the VEGF and FGF production compared with EGF-wound dressing.     

Development of a Wound Dressing Composed of a Hyaluronic Acid Sponge Containing Arginine

Spongy sheets composed of cross-linked high-molecular-weight (HMW) hyaluronic acid (HA) were prepared by freeze-drying an aqueous HMW-HA solution containing cross-linking agent (Group I). The Group I sheet was immersed into an aqueous low-molecular-weight (LMW) HA solution with or without L-arginine (Arg) and was then freeze-dried to prepare several types of spongy sheets (Groups II–V). The amount of Arg was 1.0 g, 0.5 g, 0.2 g and 0 g in Groups III, IV, V and II, respectively. In the first experiment, each spongy sheet was applied to a full-thickness skin defect with a diameter of 35 mm in the abdominal region of SD rats, with intact skin in the central area measuring 15 mm in diameter. Commercially available polyurethane film dressing was applied over each spongy sheet as a covering material. The control group was covered with polyurethane film dressing alone. All spongy sheets promoted epithelization, as well as angiogenesis, as compared with controls. These findings indicate that HA and Arg are essential for wound healing. Re-epithelizaion was particularly active in Groups IV and V. In the second experiment, each spongy sheet was applied to a full-thickness burn injury measuring 35 mm in diameter in the abdominal region of SD rats, after necrotic skin was surgically removed. Groups II–V showed decreased wound size when compared with Group I and controls. The present findings indicate that the release of LMW-HA and Arg from a cross-linked HMW-HA spongy sheet effectively stimulates wound healing.     

Development of a functional wound dressing composed of hyaluronic acid spongy sheet containing bioactive components: evaluation of wound healing potential in animal tests

This study aimed to develop a novel wound dressing composed of hyaluronic acid (HA) spongy sheet containing bioactive components. The wound dressing prepared by the freeze-drying method has a two-layered structure: an upper layer composed of cross-linked high-molecular-weight HA (HMW-HA) and a lower layer composed of low-molecular-weight HA (LMW-HA) containing arginine (Arg), magnesium ascorbyl phosphate (vitamin C derivative: VC), and epidermal growth factor (EGF) (referred to as EGF-dressing). A wound dressing containing only Arg and VC was prepared in a similar manner (referred to as EGF-free-dressing). The potential of each wound dressing was evaluated in animal tests using Sprague Dawley (SD) rats and diabetic mice. In the first experiment, each wound dressing was applied to a full-thickness skin defect in the abdominal region of SD rats. Wound conditions after 1?week and 2?weeks of treatment were evaluated based on macroscopic and histological appearance. A commercially available non-woven alginate wound dressing (Alg-dressing) was used in a control group. Both EGF-free-dressing and EGF-dressing decreased wound size and promoted granulation tissue formation associated with angiogenesis more effectively when compared with Alg-dressing. In particular, EGF-dressing promoted re-epithelialization. In the second experiment, each wound dressing was applied to a full-thickness skin defect in the dorsal region of diabetic mice. Wound conditions after 1?week and 2?weeks of treatment were evaluated based on macroscopic and histological appearance. A commercially available Alg-dressing was used in a control group. Both EGF-free-dressing and EGF-dressing decreased wound size and promoted granulation tissue formation associated with angiogenesis more effectively when compared with Alg-dressing. These findings indicate that EGF-free-dressing and EGF-dressing have the potential for more effective wound healing when compared with Alg-dressing. In particular, EGF-dressing has a higher potential for wound healing when compared with EGF-free-dressing.     

Potential of wound dressing composed of hyaluronic acid containing epidermal growth factor to enhance cytokine production by fibroblasts

This study aimed to investigate the potential of a wound dressing composed of hyaluronic acid (HA) containing epidermal growth factor (EGF) to enhance cytokine production by fibroblasts. The present wound dressing has a two-layered spongy structure: an upper layer composed of crosslinked high-molecular-weight HA, and a lower layer composed of low-molecular-weight HA containing arginine (Arg) and vitamin C derivative (VC) with or without EGF. Human fibroblast-embedded collagen gel sheet (cultured dermal substitute: CDS) was elevated to the interface between the air and culture medium to create a wound surface model onto which each wound dressing was placed, which was followed by culture for 7 days. The EGF dressing (with EGF, Arg, VC) significantly enhanced the production of vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) by CDS as compared to the EGF-free dressing (with Arg, VC). To evaluate if this enhanced production of VEGF and HGF achieved with the EGF dressing is sustained, a second experiment was conducted using a wound surface model. Each wound dressing was placed on the CDS in the wound surface model. Culture was then performed for 3 days (first period), after which each dressing was placed on another CDS for a further 3-day culture period (second period). The EGF dressing enhanced the production of VEGF and HGF by CDS during the first and second periods as compared to the corresponding production when using the EGF-free dressing. These results suggest that EGF can be maintained in the hydrated layer of a wound dressing composed of crosslinked high-molecular-weight HA.     

Development of anti-adhesive spongy sheet composed of hyaluronic acid and collagen containing epidermal growth factor

Anti-adhesive products need to be designed while considering the concept of wound healing. Two main events must proceed simultaneously: facilitating wound healing in surgically excised tissue, as well as preventing injured tissue from adhering to the surrounding tissue. The present study aimed to develop an anti-adhesive spongy sheet composed of hyaluronic acid and collagen (Col) containing epidermal growth factor, and to investigate the potential of this spongy sheet using an in vitro wound surface model (placing a spongy sheet on a fibroblast-incorporating Col gel sheet) and an in vitro inter-tissue model (placing a spongy sheet between two fibroblast-incorporating Col gel sheets). These in vitro experiments demonstrated that this spongy sheet effectively stimulates fibroblasts to release an increased amount of vascular endothelial growth factor and hepatocyte growth factor, which are essential for wound healing to proceed succesfully. In addition, anti-adhesive performance of this spongy sheet was evaluated in animal experiments using Sprague Dawley rats. Under anesthesia, a 1?cm?×?2?cm segment of peritoneum was superficially excised from walls, and the cecum was then abraded by scraping with a scalpel blade over a 1?cm?×?2?cm area. A piece of spongy sheet was placed on the peritoneal defect. Both defects were placed in contact, and the incision was closed by suturing. Peritoneal condition was evaluated after one week. This spongy sheet was capable of facilitating the wound healing of surgically excised tissue and preventing surgically excised tissue from adhering to surrounding tissues.     

Spongy matrix of hyaluronic acid and collagen as a cultured dermal substitute: evaluation in an animal test

 The authors have developed a two-layered spongy matrix composed of hyaluronic acid (HA) and atelo-collagen (Col) as a cultured dermal substitute (CDS). This spongy matrix was applied to a full-thickness skin defect on the dorsum of Sprague-Dawley rats, and the wound conditions were observed over a period of 2 weeks. A piece of two-layered spongy matrix was applied to the wound, over which a commercially available polyurethane film dressing was applied. Both the macroscopic and the histological evaluations indicate that the two-layered spongy matrix is able to prepare a highly vascularized granulation tissue at an early stage. These findings suggest that this type of spongy matrix would be useful as a CDS. Received: October 4, 2002 / Accepted: January 23, 2003 Correspondence to:Y. Kuroyanagi     

Development of cultured dermal substitute composed of hyaluronic acid and collagen spongy sheet containing fibroblasts and epidermal growth factor

The present study aimed to develop a two-layered cultured dermal substitute (CDS). The upper layer is a hyaluronic acid (HA) and collagen (Col) spongy sheet with or without epidermal growth factor (EGF). The lower layer is a HA spongy sheet and Col gel containing fibroblasts. The CDS is prepared in serum-free medium, followed by placing on the wound surface. Corresponding to clinical application, CDS was incubated in serum-free medium for a period of 1, 3 or 5?days, followed by placing onto the air and culture medium interface (wound surface model), and culture for 6?days using conventional culture medium supplemented with serum. Metabolic activity and cytokine production were considerably higher in EGF-incorporating CDS, as compared with EGF-free CDS. Metabolic activity of EGF-incorporating CDS was maintained for a period of 3?days, but decreased slightly after 5?days. EGF-incorporating CDS is able to effectively stimulate fibroblasts within CDS to release increased amounts of vascular endothelial growth factor and hepatocyte growth factor, which are essential for wound healing. CDS is promising for wound therapy, because there is no risk of cellular damage caused by cryopreservation, thawing and rinsing processes. The critical issue is how to reduce the cellular damage during a prolonged period of incubation in serum-free medium. EGF-incorporating CDS can be used after a period of 3–5?days incubation in serum-free medium. This period is sufficient for transport of CDS from manufacturing facilities to hospitals.     

Effect of EGF and bFGF on Fibroblast Proliferation and Angiogenic Cytokine Production from Cultured Dermal Substitutes

Abstract

Growth factors accelerate wound healing but the underlying mechanisms remain poorly understood. The aim of this study was to investigate the effect of epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) on fibroblast proliferation and production of angiogenic factors from cultured dermal substitutes (CDS). In the first experiment, fibroblasts were seeded into a flask at a density of 1 × 10⁴ cells/cm².Cell proliferation was assessed after culturing in media containing EGF or bFGF at concentrations ranging from 2 to 50 μg. The number of fibroblasts increased significantly in the presence of EGF or bFGF, but fibroblasts detached from the flasks in the presence of 50 μg bFGF. In the second experiment, CDS were prepared by incorporating fibroblasts into collagen gels. To make a wound surface model, the CDS was elevated to the air–liquid interface, on which a spongy sheet of hyaluronic acid (HA) containing EGF or bFGF was placed. The amount of vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) released from the CDS after 1 week of cultivation was measured by ELISA. When the CDS was covered with a HA sponge containing EGF (Group 1), fibroblasts released 3.5-times more VEGF compared with a HA-alone sponge (control group). When covered with a HA sponge containing bFGF (Group 2), 8.7-times more VEGF was released compared with the control group. Fibroblasts in Groups 1 and 2 released 9.6- and 9.3-times more HGF, respectively, compared with the control group. Thus, EGF stimulates fibroblasts to produce VEGF and HGF, in addition to its ability to enhance epidermal cell proliferation.     

Potential of a Cryopreserved Cultured Dermal Substitute Composed of Hyaluronic Acid and Collagen to Release Angiogenic Cytokine

Abstract

An allogeneic cultured dermal substitute (CDS) was prepared by culturing fibroblasts on a spongy matrix of hyaluronic acid (HA) and collagen (Col), which was then cryopreserved. This cryopreserved allogeneic CDS (CDS-1; cryopreserved for 1 month, CDS-6; cryopreserved for 6 months) was thawed and re-cultured for a period of 7 days to investigate the potential of the CDS for wound treatment. The cell metabolic activity in the CDS and their cytokine production were measured using an MTT assay and ELISA. Fibroblast metabolic activity in each CDS-1 and CDS-6 immediately after thawing and following 3 and 7 days of re- cultivation was 56, 67 and 93%, and 49, 64 and 86%, respectively, of that before cryopreservation. The amount of vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) released from the CDS-1 on days 1, 3 and 7 of re-cultivation after thawing was 8, 44 and 92% (VEGF) and 3, 7 and 28% (HGF), respectively, of that before cryopreservation. The amount of VEGF and HGF released from the CDS-6 on days 1, 3 and 7 of re-cultivation after thawing was 9, 32 and 45% (VEGF) and 6, 10 and 27% (HGF), respectively, of that before cryopreservation. These findings showed that the potential of the CDS was restored to some extent over the first 3 days of re-cultivation after thawing. The potential of the CDS for wound treatment was then evaluated using a wound surface model, in which the each CDS-1 and CDS-6 that was re-cultured for 3 days after thawing was elevated at the air/culture medium interface, and a wound dressing was placed on top, and then cultured for 5 days. Two different types of wound dressing were tested. Fibroblasts in the CDS in Group II (placing a wound dressing with EGF) released increased amount of VEGF and HGF compared with that in Group I (placing a wound dressing without EGF). These findings suggest that re-culture of the CDS for 3 days following thawing results in a CDS with improved wound healing potential and that an EGF-incorporating wound dressing is useful as a top dressing for the CDS.     

EGF containing gelatin-based wound dressings

In case of bulk loss of tissue or non-healing wounds such as burns, trauma, diabetic, decubitus and venous stasis ulcers, a proper wound dressing is needed to cover the wound area, protect the damaged tissue, and if possible to activate the cell proliferation and stimulate the healing process. In this study, synthesis of a novel polymeric bilayer wound dressing containing epidermal growth-factor (EGF) -loaded microspheres was aimed. For this purpose, a natural, nontoxic and biocompatible material, gelatin, was chosen as the underlying layer and various porous matrices in sponge form were prepared from gelatin by freeze-drying technique. As the external layer, elastomeric polyurethane membranes were used. Two different doses of EGF was added into the prepared gelatin sponges (1 and 15 microg/cm2) to activate cell proliferation. EGF addition was carried out either in free form or within microspheres to achieve prolonged release of EGF for higher efficiency. The prepared systems were tested in in vivo experiments on full-thickness skin defects created on rabbits. At certain intervals, wound areas were measured and tissues from wound areas were biopsied and processed for histological examinations. The wound areas decreased upon low-dose EGF application but the difference between the affects of free EGF and microsphere loaded EGF was not so distinct. Upon increasing the dose of EGF by a factor of 15, it was observed that controlled release of EGF from microspheres provided a higher degree of reduction in the wound areas. Histological investigations showed that the prepared dressings were biocompatible and did not cause any mononuclear cell infiltration or foreign body reaction. The structure of the newly formed dermis was almost the same as that of the normal skin.     

Characterization of a cultured dermal substitute composed of a spongy matrix of hyaluronic acid and collagen combined with fibroblasts

 The authors have developed an allogeneic cultured dermal substitute (CDS) through cultivation of fibroblasts on a two-layered spongy matrix of hyaluronic acid (HA) and atelocollagen (Col). The Col spongy layer is essential for attachment and proliferation of fibroblasts on the two-layered spongy matrix. The HA spongy layer is necessary for maintaining the moisture environment on the wound surface. The optimal weight ratio of HA/Col is determined by considering the following characteristics: mechanical properties for handling, cell viability after thawing, potency of vascular endothelial growth factor (VEGF) release after thawing, efficacy of wound healing, and manufacturing cost. This study is designed to investigate the physical properties for handling, the growth behavior of fibroblasts on the spongy matrix, and the quantitative analysis of VEGF released from fibroblasts in the fresh or cryopreserved CDS. The results of this study suggest that a CDS composed of Col spongy matrix alone has the highest potency in regard to the release of VEGF. However, taking into account the manufacturing cost, coupled with the potency of VEGF release, a two-layered sponge of HA and Col with a weight ratio of 5/2 is very promising for commercial application. Received: October 4, 2002 / Accepted: March 20, 2003     

Defects in Innate Immunity Predispose C57BL/6J-Leprdb/Leprdb Mice to Infection by Staphylococcus aureus

Foot and ankle infections are the most common cause of hospitalization among diabetic patients, and Staphylococcus aureus is a major pathogen implicated in these infections. Patients with insulin-resistant (type 2) diabetes are more susceptible to bacterial infections than nondiabetic subjects, but the pathogenesis of these infections is poorly understood. C57BL/6J-Lepr^db/Lepr^db (hereafter, db/db) mice develop type 2 diabetes due to a recessive, autosomal mutation in the leptin receptor. We established a S. aureus hind paw infection in diabetic db/db and nondiabetic Lepr^+/+ (+/+) mice to investigate host factors that predispose diabetic mice to infection. Nondiabetic +/+ mice resolved the S. aureus hind paw infection within 10 days, whereas db/db mice with persistent hyperglycemia developed a chronic infection associated with a high bacterial burden. Diabetic db/db mice showed a more robust neutrophil infiltration to the infection site and higher levels of chemokines in the infected tissue than +/+ mice. Blood from +/+ mice killed S. aureus in vitro, whereas db/db blood was defective in bacterial killing. Compared with peripheral blood neutrophils from +/+ mice, db/db neutrophils demonstrated a diminished respiratory burst when stimulated with S. aureus. However, bone marrow-derived neutrophils from +/+ and db/db mice showed comparable phagocytosis and bactericidal activity. Our results indicate that diabetic db/db mice are more susceptible to staphylococcal infection than their nondiabetic littermates and that persistent hyperglycemia modulates innate immunity in the diabetic host.Approximately 2 million of the estimated 16 million individuals with diabetes in the United States will develop chronic foot ulcers or infections during the course of their disease (38). An infection is initiated when the skin barrier is breached and bacteria, mostly skin commensals, gain access to the underlying tissues. Although limb-threatening infections are usually polymicrobial, Staphylococcus aureus is a major determinant of these infections (21). S. aureus is the predominant pathogen in non-limb-threatening infections, particularly in patients who have not received antimicrobial therapy (5, 26, 27). The emergence of S. aureus strains resistant to multiple antibiotics has made treatment of staphylococcal infections especially problematic. Methicillin-resistant S. aureus strains have become increasingly prevalent among both nosocomial and community-acquired infections within the United States (22, 36, 43). The prevalence of methicillin-resistant S. aureus is higher among diabetic patients than in the general population (11, 44, 45). Seven S. aureus strains resistant to vancomycin have been isolated in the United States, and four of these strains were isolated from patients with diabetes (42, 50). Complications of type 2 diabetes such as peripheral neuropathy and vasculopathy contribute to delayed wound healing. Although the increased susceptibility of the diabetic host to bacterial infections is well established, the chronicity of these infections is poorly understood. A consistent defect in the humoral or cell-mediated host immune system of diabetic patients has not been demonstrated. However, deficiencies in the host innate immune response are apparent since clinical investigations have indicated that phagocytes from type 2 diabetic patients are in a heightened state of oxidative stress and have impaired bactericidal activity and chemotaxis (6, 15, 41, 48). Clearly, numerous pathophysiologic perturbations contribute to the recurrence of soft tissue and bone infections in the lower extremity of patients with diabetes.C57BL/6J-Lepr^db/Lepr^db (hereafter, db/db) mice are a valuable model of type 2 diabetes since they are hyperglycemic and resistant to insulin, and they experience peripheral neuropathy, delayed wound healing, and myocardial disease. In this study we inoculated the hind paws of wild-type Lepr^+/+ (+/+) and diabetic db/db mice with S. aureus and evaluated the course of the ensuing infection in each host type, as well as the resultant host innate immune response to infection. Diabetic mice that were ≥4 months of age were more susceptible to staphylococcal infection than age-matched nondiabetic control animals. The db/db mice showed a heightened inflammatory response that was characterized by defects in phagocyte function.     

Occlusive Dressings and the Healing of Standardized Abrasions

Context:

Acute skin trauma during sport participation, resulting in partial-thickness abrasions, is common. The limited investigations focusing on the acute wound environment and dressing techniques and the subsequent lack of evidence-based standards complicate clinical wound care decisions.

Objective:

To examine the effects of occlusive dressings on healing of standardized, partial-thickness abrasions.

Design:

Controlled, counterbalanced, repeated-measures design.

Setting:

University laboratory.

Patients or Other Participants:

Sixteen healthy women (n  =  10) and men (n  =  6).

Intervention(s):

Four standardized, partial-thickness abrasions were inflicted. Film, hydrogel, and hydrocolloid occlusive dressings and no dressing (control) were applied. Participants returned on postwound days 1, 3, 5, 7, 10, and 14 for digital imaging. Wound healing time was measured by change in wound contraction (cm²) and change in wound color (chromatic red) and luminance in red, green, and blue color values.

Main Outcome Measure(s):

Wound contraction, color (chromatic red), and luminance.

Results:

A day-by-dressing interaction was found for wound contraction, color, and luminance. Post hoc testing indicated that the film and hydrocolloid dressings produced greater wound contraction than the hydrogel and no dressing on days 7 and 10. Film, hydrogel, and hydrocolloid dressings also resulted in greater wound contraction than the control on day 14. Hydrocolloid dressings produced smaller measures of color and greater measures of luminance than no dressing on day 7. Film, hydrogel, and hydrocolloid dressings also resulted in smaller measures of color and greater measures of luminance compared with no dressing on days 10 and 14.

Conclusions:

When compared with the control (no dressing), the film, hydrogel, and hydrocolloid occlusive dressings were associated with a faster healing rate of partial-thickness abrasions across time measured by wound contraction, color, and luminance. Overall, these data indicate that occlusive dressings were more effective in healing than no dressing was.     

Hyaluronic acid nanofiber wound dressing--production, characterization, and in vivo behavior

Hyaluronic acid (HA) was electrospun. The effects of flow rate, spin length, and the applied voltage on the diameter of the HA nanofibers were analyzed. The average thickness of the webs was 0.041 cm. The air permeability of sterilized HA nanofiber wound dressing was much higher than that of gauze with Vaseline. The degree of crystallinity of HA nanofibers was characterized using powder X-ray technique and was found to be 20.6%. A preclinical study was conducted to compare healing of wounds covered by an adhesive bandage, a sterilized solid HA, gauze with Vaseline, an antibiotic dressing, and a sterilized HA nanofiber wound dressing. Results of the two methods of appraisal showed that the sterilized HA nanofiber wound dressing was the best type of dressing out of the five types of dressings compared.     

不同敷料在糖尿病足溃疡伤口治疗中的研究与应用

BACKGROUND: To date, various types of dressings have been proved to promote healing of diabetic foot ulcers. OBJECTIVE: To introduce the principle and application of various kinds of wound dressing as well as other adjuvant therapies for diabetic foot ulcers. METHODS: CNKI, PubMed, Medline and Foreign Medical Journal Full-Text Service were retrieved by computer for relevant literatures published from 2005 to 2015. The search terms were “diabetic foot ulcer, wound healing, dressing, treatment” in Chinese and English, respectively. RESULTS AND CONCLUSION: The most commonly used dressings for diabetic foot ulcers include gels, hydrogels, sponges, foam dressings, film dressings. Polymer materials with different components exhibit their unique chemical, physical and biological properties, such as natural polymer dressings (chitin, chitosan and its derivatives, fibroin, dextran), synthetic polymer dressings (polyvinyl alcohol, polyethylene glycol/polyethylene oxide, poly(α-ester)s, all of which can promote the healing of diabetic foot ulcers. Other therapies for diabetic foot ulcers include herbal therapy, becaplermin, and maggot therapy.