Ischemia-reperfusion (IR) injury is a leading cause of flap compromise and organ dysfunction during free-tissue transfer, and remains a great challenge for plastic surgeons. Thioredoxin-1 (Trx-1) was proved to protect the IR flap by mitigating the oxidative stress, and inhibiting the activation of apoptosis signal-regulating kinase-1 (ASK-1) and mitogen-activated protein kinase (MAPK) pathway. The aim of this study is to investigate the distinction of Trx-1 expression, apoptosis indices in different layers of IR flaps, and the feasibility of tissue-layer-specific administration of Trx-1.

Ten patients′ specimens of IR flaps for DIEP breast reconstruction were collected and assessed for apoptosis and Trx-1 expression. Twenty mice were used to establish the IR flap model. The mice were sacrificed twenty-four hours after reperfusion. The flap tissues were harvested and tested by immunohistochemistry staining and TUNEL assay. The tissue-layer-specific dermoprotective effect of Trx-1 and the molecular mechanisms were assessed by an in vitro epithelial skin cell hypoxia-reoxygenation model. The statistics were conducted by t test and ANOVA using SPSS 20.0.

Trx-1 expression and apoptotic cells were observed mainly located in the basal layer of epidermis and the papillary layer of dermis in human IR flaps and mice models. Trx-1 depletion was 24.19 %± 2.23% in the basal layer of epidermis and the papillary layer of dermis of patient IR flaps, decreasing significantly compared with 70.71% ± 6.38% in control group (t = 27.54, P< 0.001). Similar tissue-layer-specific down regulation of Trx-1 also displayed in mice IR flap models (19.83% ± 2.34% vs. 76.59% ± 4.88%; t = 34.71, P<0.001). The apoptotic index in human samples significantly increased from 1.32% ± 1.52% in control group to 43.71 %± 3.17% in IR group (t =38.23, P<0.001); while it was proved to be dramatically raised in mice models from 0.86% ± 1.15% in control group to 41.14 %± 4.21% in IR group (t= 36.96, P < 0.001). Western Blot analysis revealed Trx-1 down regulation and a significant increase in ASK-1, p-p38, and c-PARP abundance in the hypoxia-reoxygenation-treated HaCaT cells (P < 0.01). Supplementation of recombinant human Trx-1 significantly reduced the apoptosis-related protein expression.

The basal layer of epidermis and the papillary layer of dermis are the main damaged tissue layers in the early stage of skin flap ischemia-reperfusion injury. The IR flap can be protected by precisely replenishing the vulnerable layers with Trx-1.

Surgical flap; Free Flap; Ischemia-reperfusion injury; Thioredoxin; Cell Apoptosis; Stratum basale; Dermal papilla of dermis