Cutaneous Ischemia-reperfusion Injury Research Articles

Exposure to volatile ferroptosis inhibitor, TEMPO, reduced cutaneous ischemia-reperfusion injury progression to pressure ulcer formation in a mouse model

BackgroundIschemia– reperfusion (I/R) injury-induced oxidative stress is a key factor in the pathogenesis of pressure ulcer formation. Ferroptosis is an iron-dependent programmed cell death that connects oxidative stress and inflammation in various diseases. Recent studies revealed the protective effect of inhibition of ferroptosis in I/R injury. However, the role of ferroptosis in cutaneous I/R injury remains elusive. ObjectiveTo assess the role of ferroptosis in the progression of cutaneous I/R injury. MethodsCutaneous I/R injury experiments and histopathological studies were performed in wild-type mice with or without exposure to volatile ferroptosis inhibitor, TEMPO (2,2,6,6-Tetramethylpiperidine-1-oxyl). The suppressive effects of TEMPO on ferroptosis inducing cell death and oxidative stress were examined in vitro. ResultsInhibition of ferroptosis with TEMPO significantly reduced ulcer formation after cutaneous I/R injury. Fluctuated ferroptosis markers, such as GPX4, ACSL4, and 4-HNE expression in the I/R skin site, were reversed by TEMPO treatment. Inhibition of ferroptosis reduced apoptosis, CD3+ infiltrating lymphocytes, and improved vascularity in the I/R skin site. Inhibition of ferroptosis also suppressed the enhancement of Nrf2 activation. In vitro, ferroptosis and the activation of ferroptosis-related gene expression by RSL3 stimulation were markedly ameliorated by TEMPO treatment in mouse fibroblasts. Inhibiting ferroptosis also suppressed the elevation of the mRNA levels of NOX2 and HO-1 caused by ferroptosis. ConclusionCutaneous I/R injury-induced ferroptosis likely promotes cell death, vascular loss, infiltration of inflammatory cells, and oxidative stress. The inhibition of ferroptosis with TEMPO might have potential clinical application as novel therapeutic agent for cutaneous I/R injury.

Keratinocyte-Specific SOX2 Overexpression Suppressed Pressure Ulcer Formation after Cutaneous Ischemia–Reperfusion Injury via Enhancement of Amphiregulin Production

Ischemia-reperfusion (I/R) injury is a key player in the pathogeneses of pressure ulcer formation. Our previous work demonstrated that inducing the transcription factor SOX2 promotes cutaneous wound healing through EGFR signaling pathway enhancement. However, its protective effect on cutaneous I/R injury was not well-characterized. We aimed to assess the role of SOX2 in cutaneous I/R injury and the tissue-protective effect of SOX2 induction in keratinocytes (KCs) in cutaneous I/R injury. SOX2 was transiently expressed in KCs after cutaneous I/R injury. Ulcer formation was significantly suppressed in KC-specific SOX2-overexpressing mice. SOX2 in skin KCs significantly suppressed the infiltrating inflammatory cells, apoptotic cells, vascular damage, and hypoxic areas in cutaneous I/R injury. Oxidative stress-induced mRNA levels of inflammatory cytokine expression were suppressed, and antioxidant stress factors and amphiregulin were elevated by SOX2 induction in skin KCs. Recombinant amphiregulin administration suppressed pressure ulcer development after cutaneous I/R injury in mice and suppressed oxidative stress-induced ROS production and apoptosis invitro. These findings support that SOX2 in KCs might regulate cutaneous I/R injury through amphiregulin production, resulting in oxidative stress suppression. Recombinant amphiregulin can be a potential therapeutic agent for cutaneous I/R injury.

NKG2D Ligand Expression Induced by Oxidative Stress Mitigates Cutaneous Ischemia-Reperfusion Injury.

Pressure ulcers represent a crucial clinical problem, especially in hospitalized patients. Ischemia-reperfusion (I-R) is an important cause of these lesions. Natural killer (NK), invariant NK T (iNKT), and dendritic epidermal T-cells, which express the natural killer group 2, member D (NKG2D) receptor, have been reported to have physiological roles in skin tissue repair and wound healing. However, a role for NKG2D-NKG2D ligand interactions in I-R-induced skin injury has not been determined. Using a murine pressure ulcer model, we demonstrated that I-R-induced ulcers in NKG2D-deficient mice were larger than those in wild-type or T-cell receptor δ knockout mice. Histopathological evaluation revealed that accumulation of macrophages and neutrophils at the peripheral deep dermis and subcutaneous tissue of the ulcers was enhanced in NKG2D-deficient mice. Rae-1 mRNA, which encodes an NKG2D ligand, was induced, and RAE-1 protein was detected immunohistochemically in fibroblasts and inflammatory cells in the dermis after reperfusion. RAE-1 expression was also increased in primary mouse fibroblasts treated with sodium arsenite. These results suggested that NKG2D ligand expression was induced by oxidative stress after I-R injury and support a putative role for this ligand in wound repair. Furthermore, the influx of NKG2D-positive cells at I-R sites may mitigate pressure ulcers via NKG2D-NKG2D ligand interactions.

Cutaneous ischemia-reperfusion injury is exacerbated by IL-36 receptor antagonist deficiency.

Loss-of-function homozygous or compound heterozygous mutations in IL36RN, which encodes interleukin-36 receptor antagonist (IL-36Ra), has been implicated in the pathogenesis of skin disorders. However, the pathogenic role of IL-36Ra in cutaneous ischemia-reperfusion (I/R) injury remains unclear. We investigated the role of IL36Ra in cutaneous I/R injury. We examined I/R injury in Il36rn-/- mice. The area of wounds, numbers of infiltrated cells, apoptotic cells and neutrophil extracellular trap (NET) formation were assessed. The expression levels of various genes were analysed using real-time RT-PCR. The expression of high mobility group box 1 (HMGB1), an endogenous toll-like receptor (TLR) 4 ligand, was confirmed using immunohistology, and serum HMGB1 levels were measured by ELISA. Cytokine production by stimulated cultured J774A.1 and HaCaT cells was examined. IL-36Ra deficiency resulted in significantly delayed wound healing and increased neutrophil and macrophage infiltration into the wound tissues. Il36rn-/- mice had increased mRNA expression levels of CXCL1, CXCL2, CCL4, TNF-α, TGF-β, IL-1β, IL-6 and IL-36γ relative to wild-type mice. Apoptosis was identified in keratinocytes by TUNEL assay. HMGB1 expression in the I/R site was decreased in both keratinocytes and adnexal cells, while serum HMGB1 levels were significantly elevated after reperfusion. The mRNA levels of various cytokines, including IL-1β, were elevated in J774A.1 cells through TLR4 signalling by HMGB1 stimulation. In addition, HaCaT cells stimulated with IL-1β showed significantly increased CXCL1, TNF-α, IL-6, IL-36β and IL-36γ mRNA expression. Furthermore, NET formation was increased by IL-36Ra deficiency. Finally, either the blockade of TLR4 signalling by TAK-242 or inhibition of NET formation by Cl-amidine normalized exacerbated I/R injury in Il36rn-/- mice. This study indicated that IL-36Ra deficiency exacerbates cutaneous I/R injury due to excessive inflammatory cell recruitment, NET formation, and excessive cytokine and chemokine production via the TLR4 pathway by HMGB1 released from epidermal apoptotic cells.

Inhibition of skin fibrosis in systemic sclerosis by botulinum toxin B via the suppression of oxidative stress.

Oxidative stress has been reported to play an important role in the pathogenesis of skin fibrosis in systemic sclerosis (SSc). We previously identified that botulinum toxin (BTX) injection suppresses pressure ulcer formation in a cutaneous ischemia-reperfusion injury mouse model by regulation of oxidative stress. However, the therapeutic possibility of BTX administration for preventing skin fibrosis in SSc is unclear. The objective of this study was to investigate the effect of BTX-B on skin fibrosis in a murine model of SSc and determine the underlying mechanism. We found that BTX-B injection significantly reduced dermal thickness and inflammatory cell infiltration in bleomycin-induced skin fibrosis lesion in mice. We also identified that the oxidative stress signal detected through bioluminescence in OKD48 mice after bleomycin injection in the skin was significantly decreased by BTX-B. Additionally, mRNA levels of oxidative stress associated factors (NOX2, HO-1, Trx2) were significantly decreased by BTX-B. Apoptotic cells in the lesional skin of bleomycin-treated mice were significantly reduced by BTX-B. Oxidant-induced intracellular accumulation of reactive oxygen species in SSc fibroblasts was also inhibited by BTX-B. In conclusion, BTX-B might improve bleomycin-induced skin fibrosis via the suppression of oxidative stress and inflammatory cells in the skin. BTX-B injection may have a therapeutic effect on skin fibrosis in SSc.

Protective effect of dimethyl fumarate for the development of pressure ulcers after cutaneous ischemia-reperfusion injury.

Ischemia-reperfusion (I/R) is associated with various pathogenic conditions, and there has been increasing evidence that cutaneous I/R injury is associated with the pathogenesis of pressure ulcers (PUs), especially at the early stage presenting as non-blanchable erythema. Several studies demonstrated that oxidative stress is a key player in I/R injury, and the inhibition of oxidative stress may be capable of protecting tissue damage after I/R injury in various organs including skin. Dimethyl fumarate (DMF) approved by the Food and Drug Administration is Nrf2 activator, and recent studies revealed the antioxidative and anti-inflammatory effects of DMF on I/R injury in animal models. Our objective was to assess the effects of oral administration of DMF on the development of PUs after cutaneous I/R injury in mice. We found that DMF administration significantly decreased the size of PUs after cutaneous I/R. Cutaneous I/R-induced oxidative stress was also significantly inhibited by DMF in OKD48 mice, in which oxidative stress can be visually assessed. In addition, DMF treatment decreased hypoxic area, the numbers of apoptotic cells, and vascular loss in I/R area. DMF treatment suppressed the infiltration of MPO+ neutrophils and the production of proinflammatory cytokines in I/R site after cutaneous I/R injury. in vitro experiments, DMF treatment suppressed the production of reactive oxygen species in pericyte-like cells. These results suggest that DMF treatment might prevent the formation of PUs induced by cutaneous I/R injury via suppressing oxidative stress and subsequent inflammation. DMF treatment during the early phase of decubitus ulcers might protect against further progression.

The expression of NKG2D ligand is induced in the cutaneous ischemia-reperfusion injury and mitigates pressure ulcer via NKG2D-NKG2D ligand interaction

Abstract NKG2D is an activating receptor on NK cells, NKT cells, and several types of T-cells. The ligands for NKG2D are expressed on distressed cells, and NKG2D-NKG2D ligand interaction plays a major role in the surveillance against tumors and viral infection. Several studies have reported the physiological role of NKG2D-positive cells, such as NK cells, iNKT cells, and dendritic epidermal T-cells (DETCs) in skin tissue repair and wound healing. It has been proposed that a major etiology of pressure ulcers is reperfusion injury following ischemia, though the role of NKG2D and its ligands in the process remains to be clarified. Here we report the involvement of NKG2D-NKG2D ligand interaction in the cutaneous ischemia-reperfusion (I-R) injury. The wound size of cutaneous I-R injury was expanded in NKG2D-deficient mice, compared with wild-type and TCRδ knock-out mice. Histopathological evaluation revealed that the accumulation of macrophages and neutrophils was prominent at the periphery of the I-R injury site in NKG2D-deficient mice. The transcripts and the protein expression of murine NKG2D ligands, retinoic acid early inducible gene 1 (RAE-1), were detected in fibrocytes at the I-R injury site. The expression of RAE-1 transcripts was accelerated in primary fibroblasts cultures with sodium arsenite treatment. These results suggest that NKG2D ligand expression is induced by the oxidative stress at the I-R injury site. The pressure ulcer formation by the I-R injury is mitigated via NKG2D-NKG2D interaction. This process may be mediated by the influx of NKG2D-positive cells, which regulate the degree of acute inflammatory cells accumulation at the I-R injury site.

Apelin/APJ signaling suppresses the pressure ulcer formation in cutaneous ischemia-reperfusion injury mouse model

Several studies have demonstrated potential roles for apelin/APJ signaling in the regulation of oxidative stress associated with ischemia-reperfusion (I/R) injury in several organs. Objective was to assess the role of apelin/APJ signaling in the development of pressure ulcers (PUs) formation after cutaneous I/R injury in mice. We identified that cutaneous I/R injury increased the expression of apelin in the skin at I/R site. Administration of apelin significantly inhibited the formation of PUs. The reductions of blood vessels, hypoxic area and apoptosis in I/R site were inhibited by apelin injection. Oxidative stress signals in OKD48 mice and the expressions of oxidative stress related genes in the skin were suppressed by apelin injection. H2O2-induced intracellular ROS and apoptosis in endothelial cells and fibroblasts were suppressed by apelin in vitro. Furthermore, MM07, biased agonist of APJ, also significantly suppressed the development of PUs after cutaneous I/R, and the inhibitory effect of MM07 on PUs formation was higher than that in apelin. We conclude that apelin/APJ signaling may inhibit cutaneous I/R injury-induced PUs formation by protecting the reduction of vascularity and tissue damage via suppression of oxidative stress. Exogenous application of apelin or MM07 might have therapeutic potentials against the development of PUs.

LB1612 Botulinum toxin B suppresses the pressure ulcer formation in cutaneous ischemia-reperfusion injury mouse model: Possible regulation of oxidative and endoplasmic reticulum stress

There is increasing evidence that cutaneous ischemia-reperfusion (I/R) injury is associated with the development of pressure ulcers (PUs). Botulinum toxin (BTX) suppress the release of neurotransmitters such as acetylcholine in the neuromuscular junction. Several studies have demonstrated that BTX enhanced the blood flow and survival of ischemic skin flaps in animal cutaneous flap models. The objective of this study is to assess the effects of BTX-B on the formation of PUs in the cutaneous I/R injury. In the cutaneous I/R injury mouse model, I/R injury-induced vascular damage, hypoxic area, and the number of oxidative stress-associated DNA-damaged cells and apoptotic cells were suppressed by BTX-B injection. BTX-B significantly inhibited cutaneous I/R injury-induced oxidative stress in OKD48 mice by bioluminescence. BTX-B reduced the I/R-induced mRNA levels of oxidative stress-associated factors, including HO-1, Nox2, and Nrf2. In an in vitro assay, BTX-B significantly inhibited the oxidant-induced ROS and apoptotic endothelial cells and fibroblasts. In addition, BTX-B significantly inhibited cutaneous I/R injury-induced endoplasmic reticulum (ER) stress in ERAI mice by bioluminescence. Cutaneous I/R injury-induced ER stress-response factors and GRP78/BiP- and CHOP-positive cells in the I/R area were significantly decreased by BTX-B injection. These results suggest that BTX-B injection might have protective effects against PU formation after cutaneous I/R injury by reducing vascular damage, inhibiting hypoxia-induced oxidative cellular damage, decreasing oxidative and ER stress, and preventing apoptosis.

Botulinum toxin B suppresses the pressure ulcer formation in cutaneous ischemia-reperfusion injury mouse model: Possible regulation of oxidative and endoplasmic reticulum stress

BackgroundWe previously identified that botulinum toxin A (BTX-A) suppressed pressure ulcer (PU) formation after cutaneous ischemia-reperfusion (I/R) injury; however, regulation of cutaneous I/R-induced oxidative and endoplasmic reticulum (ER) stress by BTX-B was not investigated. Additionally, the efficacy of BTX-B injection has never been examined. ObjectiveObjective was to assess the effects of BTX-B on the formation of PU by cutaneous I/R injury, and the regulation of oxidative and ER stress in I/R injury by BTX-B. MethodsBTX-B was subcutaneously injected into I/R area, and wound size, vascular damage, hypoxic area, and apoptotic cells in I/R area were analyzed. We evaluated the extent of oxidative and ER stress in I/R area by using OKD48 mice and ERAI mice, respectively, which enabled evaluating oxidative and ER stress through bioluminescence detection. ResultsBTX-B injection significantly suppressed the formation of PU by cutaneous I/R injury. Cutaneous I/R-induced vascular damage, hypoxic area, and number of oxidative-damaged cells and apoptotic cells were suppressed by BTX-B injection. BTX-B administration significantly inhibited I/R-induced oxidative stress signal in OKD48 mice. BTX-B reduced the I/R-induced oxidative stress-associated factors. BTX-B significantly inhibited the oxidant-induced reactive oxygen species and apoptosis of endothelial cells and fibroblasts. BTX-B significantly inhibited I/R-induced ER stress signal in ERAI mice. Cutaneous I/R injury-induced ER stress-response factors and GRP78/BiP and CHOP-positive cells in I/R area were significantly decreased by BTX-B injection. ConclusionBTX-B injection might have protective effects against PU formation after cutaneous I/R injury by reducing vascular damage, hypoxia-induced oxidative and ER stress, and apoptosis.

Protective effect of MFG-E8 after cutaneous ischemia-reperfusion injury

Protective effect of MFG-E8 after cutaneous ischemia-reperfusion injury

Topical betamethasone butyrate propionate exacerbates pressure ulcers after cutaneous ischemia-reperfusion injury.

Ischaemia-reperfusion (I/R) is involved in the development of various organ diseases. There has been increasing evidence that cutaneous I/R injury is associated with the pathogenesis of pressure ulcers (PUs), especially at the early stage presenting as non-blanchable erythema. However, there is no evidence-based treatment for early-stage PUs. Our objective was to assess the effects of topical steroid on the development of PUs after cutaneous I/R injury in mice. Cutaneous I/R was performed by trapping the dorsal skin between two magnetic plates for 12 h, followed by plate removal. Topical application of betamethasone butyrate propionate (BBP) in I/R areas significantly increased the size of PUs after I/R. The number of thromboses was increased, and CD31(+) vessels were decreased in the I/R area treated with topical BBP. The number of oxidative stress-associated DNA-damaged cells and apoptotic cells in the I/R area was increased by topical BBP treatment. In addition, the mRNA level of NADPH oxidase 4 (Nox4), the essential enzyme that produces reactive oxygen species, was significantly increased and that of NF-E2-related factor 2 (Nrf2), a transcription factor that regulates the expression of antioxidant proteins, was inhibited in the I/R area treated by BBP. The number of CD68(+) macrophages and the level of transforming growth factor-beta in lesional skin were also decreased by BBP. These results suggest that a topical steroid might accelerate the formation of PUs induced by cutaneous I/R injury by aggravating oxidative stress-induced tissue damage. Topical steroids might not be recommended for the treatment of acute-phase decubitus ulcers.

Protective Effect of MFG-E8 after Cutaneous Ischemia–Reperfusion Injury

We recently demonstrated that the secreted glycoprotein and integrin-ligand MFG-E8 promotes cutaneous wound healing by enhancing angiogenesis. Several studies have identified potential roles for MFG-E8 in regulation of ischemia-reperfusion (I/R) injury in the brain, kidney, and liver. Our objective was to assess the role of MFG-E8 in the formation of skin ulcers using a murine model of cutaneous I/R injury-cutaneous pressure ulcers. Cutaneous I/R was performed by trapping the dorsal skin between two magnetic plates for 12 hours, followed by plate removal. Expression of MFG-E8 increased in the dermis during ischemia, and then decreased after reperfusion. Administration of recombinant (r)MFG-E8 in I/R areas at the beginning of reperfusion significantly inhibited the formation of cutaneous pressure ulcers, and the number of CD31(+) vessel and NG2(+) pericytes in wounds were increased in I/R mice treated with rMFG-E8. The number of M1 macrophages and the amount of proinflammatory mediators monocyte chemotactic protein-1,induced nitric oxide synthase, IL-6, tumor necrosis factor-α, and IL-1β in the wound area were reduced by the administration of rMFG-E8. We conclude that MFG-E8 may inhibit the formation of pressure ulcers induced by cutaneous I/R injury by regulating angiogenesis and inflammation. Exogenous application of MFG-E8 might have therapeutic potential for cutaneous I/R injuries, including decubitus ulcers and Raynaud's phenomenon-induced digital ulcers.

Protective effect of botulinum toxin A after cutaneous ischemia-reperfusion injury.

Botulinum toxin A (BTX-A) blocks the release of acetylcholine vesicles into the synaptic space, and has been clinically used for aesthetic indications, neuromuscular disorders and hyperhidrosis. Several studies have demonstrated that BTX-A enhanced the blood flow and improved ischemia in animal models. Our objective was to assess the effects of BTX-A on cutaneous ischemia-reperfusion (I/R) injuries, mimicking decubitus ulcers. The administration of BTX-A in I/R areas significantly inhibited the formation of decubitus-like ulcer in cutaneous I/R injury mouse model. The number of CD31+ vessels and αSMA+ pericytes or myofibroblasts in wounds were significantly increased in the I/R mice treated with BTX-A. The hypoxic area and the number of oxidative stress-associated DNA-damaged cells and apoptotic cells in the I/R sites were reduced by BTX-A administration. In an in vitro assay, BTX-A significantly prevented the oxidant-induced intracellular accumulation of reactive oxygen species (ROS) in vascular endothelial cells. Furthermore, the administration of BTX-A completely suppressed the ulcer formation in an intermittent short-time cutaneous I/R injury model. These results suggest that BTX-A might have protective effects against ulcer formation after cutaneous I/R injury by enhancing angiogenesis and inhibiting hypoxia-induced cellular damage. Exogenous application of BTX-A might have therapeutic potential for cutaneous I/R injuries.

A Role of Macrophage and Cytokine in a Cutaneous Ischemia-reperfusion Injury Model of Mice

A Role of Macrophage and Cytokine in a Cutaneous Ischemia-reperfusion Injury Model of Mice

Anti-HMGB1 antibody attenuates vascular hyperpermeability and promotes wound healing during ischemia-reperfusion injury model in mouse skin

High mobility group box-1 (HMGB1), a highly conserved non-histone nuclear architectural protein, shows an inflammatory cytokine-like activity in the skin. Furthermore, HMGB1 is capable of activating macrophages in injured tissues during ischemia reperfusion (IR) injury. IR, one of the major causes of pressure ulcer, is mediated through tissue ischemia, leading to excessive inflammation and enhanced skin damage. However, it remains unclear whether HMGB1 is induced in cutaneous IR injury, and plays a role in pressure ulcer formation.

Investigation of Mechanism Concerning Pressure Ulcer Formation Using A Mouse Model of Cutaneous Ischemia-Reperfusion Injury

Investigation of Mechanism Concerning Pressure Ulcer Formation Using A Mouse Model of Cutaneous Ischemia-Reperfusion Injury

The Loss of MCP-1 Attenuates Cutaneous Ischemia–Reperfusion Injury in a Mouse Model of Pressure Ulcer

The formation of pressure ulcers is dependent on multiple factors including ischemia-reperfusion (IR). This study assessed the mechanism of a previously reported murine model of cutaneous IR injury. Three cycles of IR (days 1-3) by external application of two magnetic plates were performed to induce pressure ulcer formation. Increased infiltration of neutrophils and macrophages, and augmented expression of proinflammatory cytokines and inducible nitric oxide synthase (iNOS), were observed during IR cycles. In this model, monocyte chemoattractant protein-1 (MCP-1) was remarkably increased at day 1 in the skin followed by inflammatory cell infiltration. Therefore, IR cycles were performed in MCP-1-deficient (MCP-1(-/-)) mice to evaluate the role of this chemokine in pressure ulcer development. MCP-1(-/-) mice showed reduced macrophage infiltration and expression of tumor-necrosis factor-alpha (TNF)-alpha and iNOS during IR cycles leading to attenuated apoptosis and skin injury. Importantly, MCP-1 played a role in apoptosis and injury via inducing iNOS during the reperfusion rather than the ischemic period. These findings indicate that MCP-1 may be a critical factor for macrophage recruitment and subsequent skin inflammation and injury during IR cycles. We propose that this is a useful model for investigating the mechanism of pressure ulcer formation using various transgenic mice.

Attenuated stress- and hypoxia-driven gene expression in a rat model of repetitive cutaneous ischemia-reperfusion injury

Attenuated stress- and hypoxia-driven gene expression in a rat model of repetitive cutaneous ischemia-reperfusion injury

A novel murine model of cyclical cutaneous ischemia-reperfusion injury

Background Increasing evidence points to a principal role of ischemia-reperfusion in the pathogenesis of chronic skin ulceration, including pressure sores, diabetic ulcers, and venous ulcers. An incomplete understanding of this process and the limitations of current animal models of chronic wounds mandate a reproducible model in mice, in which transgenic and knockout technology are continually evolving. Materials and methods A murine model of chronic skin ulceration based on cyclical magnetic compression is presented. Forty-three C57BL/6J mice underwent varying degrees of cyclical compression with defined periods of reperfusion. Injury was measured grossly as regional necrosis, and tissue was harvested for histology, DNA electrophoresis, and reverse transcription polymerase chain reaction. Results Skin necrosis became apparent only 12 h post cycling, and was cycle-responsive and quantitative in cycled subjects. Histopathologic analysis revealed a statistically significant doubling of the leukocyte count in sections from compressed skin versus sham controls. Moreover, apoptotic DNA laddering was evident in post ischemic skin and absent in controls. Real-time PCR analysis revealed a 300-fold higher expression in iNOS mRNA from cyclically compressed skin compared with normal skin: such expression was temporal in nature. Conclusions A murine model of pressure necrosis, which bears all of the gross, histological, and molecular features of ischemia-reperfusion injury, has been established. Application of this model to the vast number of transgenic mice available will further our understanding of the mechanism of pressure sore development.