Pornprom Muangman

Nerve growth factor accelerates wound healing in diabetic mice

Patients with diabetic neuropathy have reduced numbers of cutaneous nerves, which may contribute to an increased incidence of nonhealing wounds. Nerve growth factor (NGF) has been reported to augment wound closure. We hypothesized that topical 2.5S NGF, a biologically active subunit of the NGF polymer, would accelerate wound repair, augment nerve regeneration, and increase inflammation in excisional wounds in diabetic mice. A full‐thickness 6‐mm punch biopsy wound was created on the dorsum of C57BL/6 J‐m + Leprdb mice (db/db) and heterozygous (db/–) littermates and treated daily with normal saline or 2.5S NGF (1 µg/day or 10 µg/day) on post‐injury days 0–6. Time to closure, wound epithelialization, and degree of inflammation were compared using a Students t‐test. Color subtractive‐computer‐assisted image analysis was used to quantify immunolocalized nerves in wounds. Non‐overlapping (20×) digital images of the wound were analyzed for nerve profile counts, area density (number of protein gene product 9.5 positive profiles per unit dermal area) and area fraction (protein gene product 9.5 positive area per unit dermal area). Healing times in db/db mice decreased from 30 days in normal saline‐treated mice to 26 days in mice treated with 1 µg/day NGF (p < 0.05) and 24 days in mice treated with 10 µg/day NGF (p < 0.02). A similar trend in db/– mice was not significant. NGF treatment augmented epithelialization in the db/db mice (p < 0.05). Histological evaluation of inflammation in healed wounds showed no statistical difference between treatment groups. Total nerve number, area density, and area fraction were increased in NGF‐treated wounds at 14, 21, and 35 days (p < 0.05). The 2.5 NGF subunit may improve wound closure kinetics by promoting epithelialization and nerve regeneration. Further studies to determine the role of nerves in wound repair are warranted.

Neutral endopeptidase inhibition in diabetic wound repair

In response to cutaneous injury, sensory nerves release substance P, a proinflammatory neuropeptide. Substance P stimulates mitogenesis and migration of keratinocytes, fibroblasts, and endothelial cells. Neutral endopeptidase (NEP), a cell surface metallopeptidase, degrades substance P. Chronic nonhealing wounds and skin from patients with diabetes mellitus show increased NEP localization and activity. We hypothesized that increased NEP may retard wound healing and that NEP inhibition would improve closure kinetics in an excisional murine wound model. NEP enzyme activity was measured in skin samples from mutant diabetic mice (db/db) and nondiabetic (db/–) littermates by degradation of glutaryl‐ala‐ala‐phe‐4‐methoxy‐2‐naphthylamine. Full‐thickness 6‐mm dorsal excisional wounds treated with normal saline or the NEP inhibitor thiorphan (10 µM or 25 µM) for 7 days were followed until closure. Histological examination and NEP activity were evaluated in a subset of wounds. NEP activity in unwounded db/db skin (20.6 pmol MNA/hr/µg) significantly exceeded activity in db/–skin (7.9 pmol MNA/hr/µg; p = 0.02). In db/db mice, 25 µM thiorphan shortened time to closure (18.0 days; p < 0.05) compared to normal saline (23.5 days). NEP inhibition did not alter closure kinetics in db/–mice. While the inflammatory response appeared enhanced in early wounds treated with thiorphan, blinded histological scoring of healed wounds using a semiquantitative scale showed no difference in inflammation. Unwounded skin from diabetic mice shows increased NEP activity and NEP inhibition improved wound closure kinetics without affecting contraction, suggesting that its principal effect was to augment epithelialization. (WOUND REP REG 2002;10:295–301)

Making sense of hypertrophic scar: a role for nerves

Healed partial thickness wounds including burns and donor sites cause hypertrophic scar formation and patient discomfort. For many patients with hypertrophic scars, pruritis is the most distressing symptom, which leads to wound excoriation and chronic wound formation. In spite of the clinical significance of abnormal innervation in scars, the nervous system has been largely ignored in the pathophysiology of hypertrophic scars. Evidence that neuropeptides contribute to inflammatory responses to injury include inflammatory cell chemotaxis, cytokine and growth factor production. The neuropeptide substance P, which is released from nerve endings after injury, induces inflammation and mediates angiogenesis, keratinocyte proliferation, and fibrogenesis. Substance P activity is tightly regulated by neutral endopeptidase (NEP), a membrane bound metallopeptidase that degrades substance P at the cell membrane. Altered substance P levels may contribute to impaired cutaneous healing responses associated with diabetes mellitus or hypertrophic scar formation. Topical application of exogenous substance P or an NEP inhibitor enhances wound closure kinetics in diabetic murine wounds suggesting that diabetic wounds have insufficient substance P levels to promote a neuroinflammatory response necessary for normal wound repair. Conversely, increased nerve numbers and neuropeptide levels with reduced NEP levels in human and porcine hypertrophic scar samples suggest that excessive neuropeptide activity induces exuberant inflammation in hypertrophic scars. Given these observations about the role of neuropeptides in cutaneous repair, neuronal modulation of repair processes at two extremes of abnormal wound healing, chronic non‐healing ulcers in type II diabetes mellitus and hypertrophic scars in deep partial thickness wounds, may provide therapeutic targets.

Complex wound management utilizing an artificial dermal matrix.

The benefits of the Integra Dermal Regeneration Template in the management of extensive burn injuries have been well documented. Integra can reduce donor- and graft-site scarring and has been reported to be capable of vascularizing over small areas of exposed bone and tendon. Given these potential advantages, we have used Integra for a variety of other reconstruction applications. We performed a retrospective review of patients with complex wounds treated with Integra at our burn center. Integra was used in the management of a variety of wounds, including necrotizing fasciitis, extremity degloving injury, meningococcemia, Marjolin ulcer, postburn lip reconstruction, and fourth-degree burns with exposed bone or tendon. Engraftment rates of Integra and autograft were 98% ± 4% and 97% ± 4%, respectively. All areas of graft loss healed without need for regrafting. The benefits of Integra in the management of acute burn wounds can be extended to other traumatic and complex wounds.

A prospective, randomized trial of silver containing hydrofiber dressing versus 1% silver sulfadiazine for the treatment of partial thickness burns

Silver sulfadiazine has been used as a topical burn wound treatment for many years. Pain associated with dressing changes is a common problem in burn wounds. Aquacel Ag, a hydrofiber dressing coated with ionic silver has been reported to reduce burn wound infection and promote antimicrobial activity. The purpose of this study was to show the benefits of Aquacel Ag for the treatment of partial thickness burns. This prospective randomized study was conducted in 70 patients who had partial thickness burns less than 15% of total body surface area and were treated at Siriraj outpatient burn clinic during December 2006–February 2008. Patients were divided into two groups: Aquacel Ag‐treated group with dressing changes every 3 days (35 patients) and 1% silver sulfadiazine‐treated group, with daily dressing changes (35 patients). There was no difference in demographic data including age, gender, burn percentage between groups. Time‐to‐wound healing pain score during dressing change and cost of treatment were compared between both groups. Time‐to‐wound closure was significantly shorter in the Aquacel Ag‐treated group (10 ± 3 versus 13.7 ± 4 days, P < 0·02) as well as pain scores at days 1, 3 and 7 (4·1 ± 2·1, 2·1 ± 1·8, 0·9 ± 1·4 versus 6·1 ± 2·3, 5·2 ± 2·1, 3·3 ± 1·9, respectively, P < 0·02). Total cost of treatment was 52 ± 29 US dollars for the Aquacel Ag‐treated group versus 93 ± 36 US dollars for the silver sulfadiazine‐treated group. This study showed that Aquacel Ag increased time to healing, decreased pain symptoms and increased patient convenience because of limiting the frequency of replacement of the dressing at lower total cost. This study confirms the efficacy of Aquacel Ag for the treatment of partial thickness burns at an outpatient clinic.

Clinical effectiveness of alginate silver dressing in outpatient management of partial-thickness burns

Askina Calgitrol Ag® (B. Braun Hospicare Ltd, Collooney Co. Sligo, Ireland), alginate silver wound dressing, is an advanced wound dressing which combines the potent broad‐spectrum antimicrobial action of silver with enhanced exudate management properties of calcium alginate and polyurethane foam. The purpose of this study was to compare the efficacy of Askina Calgitrol Ag® and 1% silver sulfadiazine (1% AgSD) in the outpatient management of partial‐thickness burn wounds at Burn Unit, Siriraj Hospital. A prospective descriptive study was conducted between January 2008 and January 2009 in Burn Unit, Division of Trauma Surgery, Siriraj Hospital, Mahidol University, Thailand. The 65 patients with partial‐thickness burn wounds, less than 24 hours post‐burn injury, had a total body surface area (TBSA%) less than 15% were treated at Siriraj Outpatient Burn Clinic. All patients were divided into Askina Calgitrol Ag® treated group (30 patients) and 1% AgSD treated group (35 patients). The data were compared by the demographics including age, gender, % TBSA burn, pain score, number of wound dressing change, nursing time and time of wound healing. Patients included in both groups were comparable with no significant differences in demographic data of age, gender, location of burn and type of burn injury (P > 0·05 evaluated by paired Students t‐test) between both group. The present results showed that average pain scores in the Askina Calgitrol Ag® treated group were significantly lower than the 1% AgSD treated group (2·23 ± 1·87 versus 6·08 ± 2·33, respectively) between both groups (P < 0·02). Patients treated with Askina Calgitrol Ag® had significantly lower number of wound dressing change (P < 0·02) and nursing time (P < 0·02) compared with 1% AgSD treated group. The Askina Calgitrol Ag® group needed less frequent wound dressing. Healing time was 7 ± 3·51 days after the application of Askina Calgitrol Ag®. This was significantly shorter than that of control wounds (14 ± 4·18 days). Application of Askina Calgitrol Ag® leads to a good burn wound outcome. The present study confirms the effectiveness of Askina Calgitrol Ag® in the outpatient management of partial‐thickness burn wounds.

Fatty acids and glucose increase neutral endopeptidase activity in human microvascular endothelial cells

Neutral endopeptidase (NEP), a membrane-bound metallopeptidase enzyme that degrades neuropeptides, bradykinin, atrial natriuretic factor, enkephalins, and endothelin may regulate response to injury. We have previously demonstrated increased NEP localization and enzyme activity in diabetic wounds and skin compared with normal controls. We hypothesized that hyperlipidemia and hyperglycemia associated with type 2 diabetes mellitus may induce excessive NEP activity and thereby diminish normal response to injury. Human microvascular endothelial cells were treated with five different fatty acids (40 &mgr;M) with varying degrees of saturation, including oleic acid, linoleic acid, palmitic acid, stearic acid, and linolenic acid and/or glucose (40 mM) for 48 h. The effect of the antioxidative agents vitamin E and C on NEP enzyme activation was determined by treating the cultured cells with alpha-tocopherol succinate and/or L-ascorbic acid. Cell membrane preparations were assayed for NEP activity by incubation with glutaryl-Ala-Ala-Phe-4-methoxy-&bgr; naphthylamide to generate a fluorescent degradation product methoxy 2 naphthylamine. High glucose or fatty acid concentration upregulated NEP activity. The highest NEP activity was observed with combined elevated glucose, linoleic acid, and oleic acid (P < 0.05). Antioxidant vitamin E and C treatment significantly reduced NEP enzyme activity after fatty acid exposure (P < 0.05). Thus, hyperglycemia and hyperlipidemia associated with type 2 diabetes mellitus may increase endothelial cell NEP activity and thereby decrease early pro-inflammatory responses. The modulator effect of vitamin E and C on NEP membrane enzyme activity after exposure to fatty acid stimulation suggests that lipid oxidation may activate NEP.

In Vitro Evaluation of the Antimicrobial Effectiveness and Moisture Binding Properties of Wound Dressings

A variety of silver-coated dressings and some impregnated with other chemicals are now available in the market; however, there have been few studies analyzing their comparative efficacies as antimicrobial agents. Moreover, their properties for retaining an appropriate level of moisture that is critical for effective wound healing have never been reported. Five commercially available silver-containing and chlorhexidine dressings, Urgotul SSD®, Bactigras®, Acticoat®, Askina Calgitrol Ag® and Aquacel Ag®, were tested to determine their comparative antimicrobial effectiveness in vitro against five common wound pathogens, namely methicillin-sensitive and -resistant Staphylococcus aureus, Bacillus subtilis, Escherichia coli and Pseudomonas aeruginosa. Mepitel®, a flexible polyamide net coated with soft silicone, was used as a control. The zones of inhibition and both the rapidity and the extent of killing of these pathogens were evaluated. All five antimicrobial dressings investigated exerted some bactericidal activity, particularly against E. coli. The spectrum and rapidity of action ranged widely for the different dressings. Acticoat® had a broad spectrum of action against both Gram-positive and -negative bacteria. Other dressings demonstrated a narrower range of bactericidal activities. Regarding the absorption and release of moisture, Askina Calgitrol Ag® absorbed and released the most moisture from the environment. Aquacel Ag® also exhibited good moisture absorption and moisture release, but to a lower degree. The other tested dressings absorbed or released very little moisture. Askina Calgitrol Ag® and Aquacel Ag® are good alternative dressings for treating wounds with high exudates and pus. An understanding of the characteristics of these dressings will be useful for utilizing them for specific requirements under specified conditions.

Efficiency of Microbial Cellulose Dressing in Partial-Thickness Burn Wounds

Microbial cellulose is a natural polymer that can hold a quantity of water without any disconformities. Therefore, it is proposed for use as wound dressing material. We report a case of 28% total body surface area partial-thickness flame burn: approximately 4.5% superficial partial-thickness burns on anterior face and 23.5% combined superficial and deep partial-thickness burns on both upper arms and anterior trunk. A microbial cellulose dressing, Nanocell (Thai Nano Cellulose Co Ltd, Bangkok, Thailand), was applied to the face wound only once, without any further dressing change. Progress of healing, until full epithelialization on the face, was observed for 2 weeks. During the treatment period, the patient did not show any irritation or allergic reaction to this new dressing, and wound swab culture showed no evidence of bacteria presence. This innovative material can be an alternative dressing for superficial partial-thickness burn wounds.

SUBSTANCE P ENHANCES WOUND CLOSURE IN NITRIC OXIDE SYNTHASE KNOCKOUT MICE

INTRODUCTION The neuropeptide, substance P (SP), up-regulates nitric oxide production (NO). The purpose of this study was to determine whether SP enhances response to cutaneous injury in nitric oxide synthase knockout (NOS null) mice. METHODS We studied mice with targeted deletions of the 3 NOS genes, neuronal NOS, inducible NOS, or endothelial NOS. Full thickness dorsal wounds were treated daily (d 0-6) with topical SP or normal saline (NaCl). Wounds were analyzed by flow cytometry for macrophage, leukocyte, endothelial, and dendritic cells. Healing time and wound epithelialization were compared using analysis of variance. RESULTS Wound closure in the 3 NOS null mice was slower than the control mice (P < 0.05). SP treatment enhanced wound closure in NOS null mice (P < 0.02). NOS null wounds exhibited reduced inflammation. SP increased macrophage, leukocyte, and dendritic cell densities at d 3 and d 7 (P < 0.05) in all NOS null mice. SP increased endothelial cell number in neuronal NOS and inducible NOS null mice, but not in endothelial NOS null mice (P > 0.05). CONCLUSIONS SP ameliorated the impaired wound healing response observed in NOS null mice by enhancing wound closure kinetics and epithelialization. SP increased inflammatory cell density in the wounds supporting the essential role of inflammatory cells, especially macrophages, in wound repair.