Flap Survival Area Research Articles

Dietary nitrate maintains homeostasis of oxidative stress and gut microbiota to promote flap survival in type 2 diabetes mellitus rats

BackgroundRandom-pattern skin flaps are commonly used to repair skin tissue defects in surgical tissue reconstruction. However, flap necrosis in the distal area due to ischemia injury is still challenging for its applications in plastic surgery. The complications of diabetes will further increase the risk of infection and necrosis.MethodsThis study induced type 2 diabetes mellitus (T2DM) rats with a high-fat diet and STZ. The survival rate of the skin flap was observed by adding inorganic sodium nitrate to drinking water. Histology and immunohistochemistry were used to detect the damage to the skin flap. The nitrate content was measured by total nitric oxide and nitrate/nitrite parameter assay. Dihydroethidium and malondialdehyde (MDA) assays were used to value oxidative stress. Rat colon feces were collected for 16s rRNA gene sequence.ResultsOur studies showed that nitrate administration leads to anti-obesity and anti-diabetic effects. Nitrate directly increased the survival area of skin flaps in diabetic rats and mean blood vessel density by enhancing angiogenesis, inhibiting apoptosis, and reducing oxidative stress. The 16s rRNA sequence revealed that nitrate may regulate the homeostasis of the gut microbiota and re-store energy metabolism.ConclusionDietary nitrate has been shown to maintain the homeostasis of oxidative stress and gut microbiota to promote flap survival in rats with T2DM.

Quercetin activates autophagy in the distal ischemic area of random skin flaps through Beclin1 to enhance the adaptability to energy deficiency

Random flaps are frequently employed in treating substantial skin abnormalities and in surgical tissue-rebuilding interventions. The random flap technique provides flaps of specific dimensions and contours to fit the surgical incision. However, blood supply deficiency and subsequent ischemia-reperfusion injury can cause severe oxidative stress and apoptosis, eventually leading to distal necrosis, which limits the clinical application of the flap. Quercetin (QUE) is primarily found in the glycoside form in many plant parts, such as stem bark, flowers, leaves, buds, seeds, and fruits. Cellular, animal, and clinical studies have demonstrated the antioxidant, anti-apoptosis, anti-inflammatory, and activation of autophagy properties of QUE. In previous studies, high doses of QUE effectively suppressed the survival of human umbilical vein endothelial cells (HUVECs) stimulated by hydrogen peroxide. However, different concentration gradients of QUE on HUVECs revealed a significant protective effect at a concentration of 10 mM. The protective impact of QUE on HUVECs was evaluated using scratch tests, CCK-8 assays, and EDU assays. Simultaneously, a mouse model of random skin flap was created, and the impact of QUE on skin flap survival was examined by intragastric injection. The QUE group showed a significantly larger survival area of the random flap and higher blood flow intensity compared to the control group. Furthermore, the beneficial effects of QUE were reversed by the autophagy inhibitor 3-MA. Therefore, autophagy plays a significant role in the therapeutic benefits of QUE on flap survival.

Erastin promotes random-pattern skin flaps survival by inducing mTORC1-TFEB mediated autophagy

Random-pattern skin flaps are important method for skin reconstruction after defect; however, the distal end of flaps is not easily viable due to inadequate nutrient supply. Erastin is a well-established ferroptosis inducer, but our study found that low-dose of erastin (2 μM) may reduce nutrient deficiency induced cell death in human umbilical vein endothelial cells (HUVECs). RNA-seq analysis suggested that its role was related to autophagy regulation. Follow-up studies have shown that the use of autophagy inhibitors or the knockdown of TFEB in HUVECs can both reduce the anti-apoptotic effect of erastin in HUVECs. Mechanism study demonstrated that erastin can suppress mTORC1 and promote TFEB activity in HUVECs, suggesting that the effect of erastin on the survival of HUVECs under nutrient deprivation conditions is regulated by mTORC1/TFEB. Subsequently, we evaluated the effect of erastin on the survival of random-pattern skin flaps in mice in vivo. On the postoperative day 7, we observed a significant increase in flap survival area, blood perfusion, and microvascular density after erastin treatment; also, erastin treatment showed enhanced autophagy within the ischemic region. In summary, our study demonstrates that low-dose of erastin may suppress cell death in endothelial cells under nutrient deficiency condition, and its effects may relate to the mTORC1-TFEB medicated autophagy regulation, erastin treatment may be a potential therapy for random-pattern skin flaps.

Hydrogels containing KYNA promote angiogenesis and inhibit inflammation to improve the survival rate of multi-territory perforator flaps

BackgroundA new spray adhesive (KYNA-PF127) was established through the combination of thermosensitive hydrogel (Pluronic F127) and KYNA, aimed to investigate the effect of KYNA-PF127 on multi-territory perforator flaps and its possible molecular mechanism. Materials and methods36 SD male rats with 250–300 g were randomly divided into 3 groups (n = 12): control group, blank glue group and KYNA-PF127 group. KYNA-PF127 hydrogel was prepared and characterized for its morphology and properties using scanning electron microscopy. CCK-8 assay, scratch wound assay, transwell assay, tube formation assay and Ki67 staining were used to study the effect of KYNA-PF127 on the proliferation, migration, and tube formation of HUVECs. VEGF and FGF2 were measured by qPCR to evaluate the angiogenesis capacity of HUVECs in vitro. In vivo, the effect of each group on the survival area of the cross-zone perforator flap was evaluated, and angiogenesis was evaluated by HE and immunofluorescence (CD31 and MMP-9). The effect of inflammation on skin collagen fibers was assessed by Masson. Immunohistochemistry (SOD1, IL-1β, TNF-α) was used to evaluate the effects of oxidative stress and inflammatory factors on multi-territory flaps. ResultsKYNA-PF127 has good sustained release and biocompatibility at 25% concentration. KYNA-PF127 promoted the proliferation, migration, and angiogenesis of HUVECs in vitro. In vivo, the survival area of multi-territory perforator flaps and angiogenic capability have increased after KYNA-PF127 intervention. KYNA-PF127 could effectively reduce the oxidative stress and inflammation of multi-territory perforator flaps. ConclusionKYNA-PF127 promotes angiogenesis through its antioxidant stress and anti-inflammatory effects, and shows potential clinical value in promoting the survival viability and drug delivery of multi-territory perforator flaps.

Protective effects of the bioactive peptide from maggots against skin flap ischemia‒reperfusion injury in rats

Ischemia‒reperfusion (I/R) injury is a frequently observed complication after flap surgery, and it affects skin flap survival and patient prognosis. Currently, there are no proven safe and effective treatment options to treat skin flap I/R injury. Herein, the potential efficacies of the bioactive peptide from maggots (BPM), as well as its underlying mechanisms, were explored in a rat model of skin flap I/R injury and LPS- or H2O2-elicited RAW 264.7 cells. We demonstrated that BPM significantly ameliorated the area of flap survival, and histological changes in skin tissue in vivo. Furthermore, BPM could markedly restore or enhance Nrf2 and HO-1 levels, and suppress the expression of pro-inflammatory cytokines, including TLR4, p-IκB, NFκB p65, p-p65, IL-6, and TNF-α in I/R-injured skin flaps. In addition, BPM treatment exhibited excellent biocompatibility with an adequate safety profile, while it exhibited superior ROS-scavenging ability and the upregulation of antioxidant enzymes in vitro. Mechanistically, the above benefits related to BPM involved the activation of Nrf2/HO-1 and suppression of TLR4/NF-κB pathway. Taken together, this study may provide a scientific basis for the potential therapeutic effect of BPM in the prevention of skin flap I/R injury and other related diseases.

Cutting scheme and clinical application effects of ultrathin thoracodorsal artery perforator flap assisted by color Doppler ultrasound

Objective: To explore the cutting scheme and clinical application effects of ultrathin thoracodorsal artery perforator flap assisted by color Doppler ultrasound. Methods: This study was a retrospective historical control study. From February 2017 to October 2019, 20 patients who were admitted to the Third Department of Orthopedics of Xingtai General Hospital of North China Medical and Health Group (hereinafter referred to as our department), met the inclusion criteria, and underwent repair of skin and soft tissue defects of extremities with ultrathin thoracodorsal artery perforator flap designed and harvested based on the surgeon's clinical experience were selected as control group, including 16 males and 4 females, aged (37±5) years. From November 2019 to December 2022, 21 patients who were admitted to our department, met the inclusion criteria, and underwent repair of skin and soft tissue defects of extremities with ultrathin thoracodorsal artery perforator flap designed and harvested under the assistance of color Doppler ultrasound were selected as ultrasound-assisted group, including 15 males and 6 females, aged (38±6) years. After debridement, the area of skin and soft tissue defects of extremities ranged 5.0 cm×4.0 cm to 19.0 cm×8.0 cm, and the area of thoracodorsal artery perforator flaps ranged 6.0 cm×5.0 cm to 20.0 cm×9.0 cm. The wounds in flap donor sites were closed directly. For patients in ultrasound-assisted group, the time and cost required for color Doppler ultrasound examination were recorded, and the number, type, and location of thoracodorsal artery perforator vessels detected by preoperative color Doppler ultrasound were compared with those of intraoperative actual detection. The time required for complete flap harvest of patients in 2 groups was recorded. On postoperative day (POD) 1, 3, 5, 7, and 14, the blood perfusion of flaps in the 2 groups of patients was assessed using a flap perfusion assessment scale. On POD 14, flap survival of patients in 2 groups was observed, and the percentage of flap survival area was calculated. In postoperative 6 months, satisfaction of patients with the treatment outcome in the 2 groups was assessed using 5-grade Likert scale, and the satisfaction rate was calculated. Results: For patients in ultrasound-assisted group, the time required for preoperative color Doppler ultrasound examination was (10.5±2.3) min, and the cost was 120 yuan; 21 thoracodorsal artery perforator vessels were detected and marked using preoperative color Doppler ultrasound, including 8 (38.10%) type 1 perforator vessels, 10 (47.62%) type 2 perforator vessels, and 3 (14.29%) type 3 perforator vessels; the number, type, and location of thoracodorsal artery perforator vessels detected preoperatively were consistent with those detected intraoperatively. The time required for complete flap harvest of patients in ultrasound-assisted group was (41±10) min, which was significantly shorter than (63±12) min in control group (t=6.32, P<0.05). On POD 1, 3, 5, 7, and 14, the blood perfusion scores of flaps of patients in ultrasound-assisted group were significantly better than those in control group (with t values of 6.67, 7.48, 8.03, 8.75, and 7.99, respectively P<0.05). On POD 14, only one patient in ultrasound-assisted group had partial flap necrosis and 6 patients in control group had complete or partial necrosis of the flap; the percentage of flap survival area of patients in ultrasound-assisted group was (99±8)%, which was significantly higher than (87±8)% in control group (t=4.57, P<0.05). In postoperative 6 months, there was no significant difference in the satisfaction rate of patients with the treatment outcome between the two groups (P>0.05). Conclusions: Preoperative color Doppler ultrasound is highly accurate in detecting the number, type, and location of perforator vessels. The cutting scheme of ultrathin thoracodorsal artery perforator flaps can be designed according to the different types of perforator vessels, with shorted flap cutting time and improved flap survival rate.

Propionyl-l-carnitine mitigates ischemia-reperfusion injury in rat epigastric island flaps

BackgroundIschemia-reperfusion injury presents a substantial concern in various medical scenarios, notably in reconstructive surgery involving tissue flaps. Despite reports on the protective benefits of Propionyl-l-carnitine against ischemia-reperfusion injury, a thorough assessment of its efficacy in epigastric island flap models is currently lacking. MethodsSixteen male Sprague-Dawley rats underwent epigastric island flap surgery and were divided into two groups: a Propionyl-l-carnitine group that received intraperitoneal Propionyl-l-carnitine prior to ischemia induction and a sham group that received saline treatment. A comprehensive evaluation was performed including macroscopic, biochemical and histological assessments encompassing measurements of flap survival areas, lipid peroxidation (malondialdehyde), glutathione, myeloperoxidase, nitric oxide and peripheral neutrophil counts. ResultsThe Propionyl-l-carnitine group demonstrated significantly increased flap survival areas when compared to the sham group. Administration of Propionyl-l-carnitine led to reduced malondialdehyde levels and elevated glutathione levels indicating a reduction in oxidative stress. Furthermore, the Propionyl-l-carnitine group exhibited lower myeloperoxidase levels, higher nitric oxide levels and reduced peripheral neutrophil counts, suggesting a decrease in the inflammatory response. Histopathological analysis revealed decreased levels of inflammation, necrosis, polymorphonuclear leukocyte infiltration and edema in the Propionyl-l-carnitine group. Additionally, vascularity was enhanced in the Propionyl-l-carnitine group. ConclusionThis study provides compelling evidence that Propionyl-l-carnitine administration effectively mitigates the deleterious effects of ischemia-reperfusion injury in epigastric island flaps. This is substantiated by the improved flap survival, diminished oxidative stress and inflammation, as well as the enhanced vascularity observed. Propionyl-l-carnitine emerges as a promising therapeutic intervention to enhance tissue flap survival in reconstructive surgery, warranting further exploration through larger-scale investigations.

VEGF and Other Gene Therapies Improve Flap Survival-A Systematic Review and Meta-Analysis of Preclinical Studies.

Surgical flaps are basic tools in reconstructive surgery. Their use may be limited by ischemia and necrosis. Few therapies address or prevent them. Genetic therapy could improve flap outcomes, but primary studies in this field present conflicting results. This systematic review and meta-analysis aimed to appraise the efficacy of external gene delivery to the flap for its survival in preclinical models. This review was registered with PROSPERO (CRD42022359982). PubMed, Embase, Web of Science, and Scopus were searched to identify studies using animal models reporting flap survival outcomes following any genetic modifications. Random-effects meta-analysis was used to calculate mean differences in flap survival with accompanying 95% CI. The risk of bias was assessed using the SYRCLE tool. Subgroup and sensitivity analyses were performed to ascertain the robustness of primary analyses, and the evidence was assessed using the GRADE approach. The initial search yielded 690 articles; 51 were eventually included, 36 of which with 1576 rats were meta-analyzed. VEGF gene delivery to different flap types significantly improved flap survival area by 15.66% (95% CI 11.80-19.52). Other interventions had smaller or less precise effects: PDGF-13.44% (95% CI 3.53-23.35); VEGF + FGF-8.64% (95% CI 6.94-10.34); HGF-5.61% (95% CI 0.43-10.78); FGF 3.84% (95% CI 1.13-6.55). Despite considerable heterogeneity, moderate risk of bias, and low quality of evidence, the efficacy of VEGF gene therapy remained significant in all sensitivity analyses. Preclinical data indicate that gene therapy is effective for increasing flap survival, but further animal studies are required for successful clinical translation.

Electrical stimulation promoting the angiogenesis in diabetic rat perforator flap through attenuating oxidative stress-mediated inflammation and apoptosis.

Skin flap transplantation is one of the effective methods to treat the diabetes-related foot ulceration, but the intrinsic damage to vessels in diabetes mellitus (DM) leads to the necrosis of skin flaps. Therefore, the discovery of a non-invasive and effective approach for promoting the survival of flaps is of the utmost importance. Electrical stimulation (ES) promotes angiogenesis and increases the proliferation, migration, and elongation of endothelial cells, thus being a potential effective method to improve flap survival. The purpose of this study was to elucidate the mechanism used by ES to effectively restore the impaired function of endothelial cells caused by diabetes. A total of 79 adult male Sprague-Dawley rats were used in this study. Gene and protein expression was assessed by PCR and western blotting, respectively. Immunohistochemistry and hematoxylin-eosin staining were performed to evaluate the morphology and density of the microvessels in the flap. The optimal duration for preconditioning the flap with ES was 7 days. The flap survival area percentage and microvessels density in the DMES group were markedly increased compared to the DM group. VEGF, MMP2, and MMP9 protein expression was significantly upregulated. ROS intensity was significantly decreased and GSH concentration was increased. The expression of IL-1β, MCP‑1, cleaved caspase-3, and Bax were downregulated in the DMES group, while TGF-β expression was upregulated. ES improves the angiogenesis in diabetic ischemic skin flaps by attenuating oxidative stress-mediated inflammation and apoptosis, eventually increasing their viability.

Evaluation of the effects of ectopic replantation on amputate survival in the management of crush traumatic amputations in rats: An experimental study.

This controlled experimental study aimed to compare ectopic replantation with other replantation techniques in a rat model of crush amputations. It also assessed the impact of different replantation methods on the viability of amputates. Forty male Wistar albino rats were divided into four groups. Groin flaps served as the amputation model. Group 1 un-derwent guillotine-style amputation followed by orthotopic replantation, Group 2 experienced crush-type amputation and orthotopic replantation, Group 3 had crush-type amputation and orthotopic replantation with a vein graft, and Group 4 underwent crush-type amputation followed by ectopic replantation. Flap viability and perfusion rates were assessed on day 3 using an infrared perfusion as-sessment system. The ratio of viable area to total flap area and thrombus formation in the pedicle vessels were evaluated on day 7. Infrared evaluations on day 3 post-replantation revealed flap perfusion percentages of 73.5% in Group 1, 11.1% in Group 2, 65% in Group 3, and 64.1% in Group 4. Statistical analysis indicated that Group 1 exhibited the highest perfusion rates, while Group 2 showed the lowest. No differences were observed between Groups 3 and 4. On the seventh day, the average surviving flap areas were found to be 74.6% in Group 1, 2.5% in Group 2, 64.5% in Group 3, and 64% in Group 4. Statistically, Group 1 exhibited the best outcomes, while Group 2 had the poorest, with no differences between Groups 3 and 4. Additionally, thrombus formation was observed in the vessels of two animals in Group 1, nine in Group 2, and three each in Groups 3 and 4. Significant statistical differences were noted among the groups. The results indicate that ectopic replantation and replantation with a vein graft are equally effective. The preferred method for crush-type replantations may depend on the patient's and the amputated limb's conditions. In crush-type amputations, we recommend vein graft repair if the patient's overall condition supports replantation and if crushed segments can be debrided without excessive shortening of the amputated part. If these conditions are not met, temporary ectopic replantation is advised to preserve the amputated limb.

Protective effect of bone marrow mesenchymal stem cells on the survival zone of the perforator flaps in rats.

Perforator flaps have recently been used in the field of plastic surgery. Skin defects can be reconstructed to reach functional and cosmetic goals. With the development of reconstructive approaches, utilizing stem cells is a hopeful approach to enhance wound healing and tissue recovery. In this study, we assessed the effect of bone marrow mesenchymal stem cells on the perforator flap's survival in rats. Perforator flaps (2.5 × 11 cm) were transplanted into rats and focused on the thoracodorsal, intercostal, and deep circumflex arteries, which were randomly divided into three groups: control, saline, and bone marrow mesenchymal stem cells (7 × 106 ml). Seven days after the surgery, tissue edema, inflammation, and discharge were observed and photographed. Histological analyses were performed to determine flap survival. Hematoxylin and eosin staining was performed to assess levels of microvascular density determined in skin flaps. Rats in the bone marrow mesenchymal stem cells group exhibited higher average flap survival area, and higher microvascular density levels at the dynamical regions of the flaps compared with the other two groups. Subdermal injection of bone marrow mesenchymal stem cells significantly increased ischemic perforator flap survival due to stimulated neovascularization in rats. Our findings suggest the potential usefulness of bone marrow mesenchymal stem cells in preventing skin flap perforator tissue necrosis.

Neuregulin-1, a member of the epidermal growth factor family, mitigates STING-mediated pyroptosis and necroptosis in ischaemic flaps

Abstract Background Ensuring the survival of the distal end of a random flap during hypoperfusion (ischaemia) is difficult in clinical practice. Effective prevention of programmed cell death is a potential strategy for inhibiting ischaemic flap necrosis. The activation of stimulator of interferon genes (STING) pathway promotes inflammation and leads to cell death. The epidermal growth factor family member neuregulin-1 (NRG1) reduces cell death by activating the protein kinase B (AKT) signalling pathway. Moreover, AKT signalling negatively regulates STING activity. We aimed to verify the efficacy of NRG1 injection in protecting against flap necrosis. Additionally, we investigated whether NRG1 effectively enhances ischemic flap survival by inhibiting pyroptosis and necroptosis through STING suppression. Methods A random-pattern skin flap model was generated on the backs of C57BL/6 mice. The skin flap survival area was determined. The blood supply and vascular network of the flap was assessed by laser Doppler blood flow analysis. Cluster of differentiation 34 immunohistochemistry (IHC) and haematoxylin and eosin (H&amp;E) staining of the flap sections revealed microvessels. Transcriptome sequencing analysis revealed the mechanism by which NRG1 promotes the survival of ischaemic flaps. The levels of angiogenesis, oxidative stress, necroptosis, pyroptosis and indicators associated with signalling pathways in flaps were examined by IHC, immunofluorescence and Western blotting. Packaging adeno-associated virus (AAV) was used to activate STING in flaps. Results NRG1 promoted the survival of ischaemic flaps. An increased subcutaneous vascular network and neovascularization were found in ischaemic flaps after the application of NRG1. Transcriptomic gene ontology enrichment analysis and protein level detection indicated that necroptosis, pyroptosis and STING activity were reduced in the NRG1 group. The phosphorylation of AKT and forkhead box O3a (FOXO3a) were increased after NRG1 treatment. The increased expression of STING in flaps induced by AAV reversed the therapeutic effect of NRG1. The ability of NRG1 to phosphorylate AKT-FOXO3a, inhibit STING and promote flap survival was abolished after the application of the AKT inhibitor MK2206. Conclusions NRG1 inhibits pyroptosis and necroptosis by activating the AKT-FOXO3a signalling pathway to suppress STING activation and promote ischaemic flap survival.

Sequential transplantation of exosomes and BMSCs pretreated with hypoxia efficiently facilitates perforator skin flap survival area in rats

Bone marrow mesenchymal stem cells (BMSC) are promising candidates for the treatment of trans-territory perforator flap necrosis. However, the low retention and survival rate of engrafted BMSCs limit their therapeutic efficacy. Strategies either modifying BMSCs or alleviating the inflammatory environment may solve this problem. Thus, we aimed to explore the therapeutic efficacy of sequential transplantation of exosomes and hypoxia pretreated BMSCs on flap necrosis. After the perforator flap model was created, the exosomes derived from BMSCs were injected immediately into choke zone II followed by transplantation of hypoxia pretreated BMSCs on Day 2. Gross view was performed to assess the flap survival, enzyme-linked immunosorbent assay was performed to evaluate the inflammatory factor level, microvessel number was assessed and quantitative polymerase chain reaction (qPCR) was performed to assess angiogenesis. We found that exosome delivery significantly reduced inflammatory cytokines levels on Day 1 and Day 3 and promoted the engrafted BMSCs’ survival on Day 7. After combining with transplantation of hypoxia pretreated BMSCs, the flap survival rate and the angiogenesis-related gene expression were significantly higher than in the other three groups; the von Willebrand factor (vWF) vascular diameter and vWF vascular count were significantly higher than in the phosphate buffered saline (PBS) group. Thus, we concluded that sequential transplantation of exosomes and BMSCs combinatorially pretreated with hypoxia further facilitated flap survival. This sequential transplantation approach provides novel insights into the clinical treatment of flap necrosis.

Effects of Intramuscular Epinephrine Injection and Systemic Sildenafil Citrate on Rat Transverse Rectus Abdominis Musculocutaneous Flap Survival

Background and Objectives Flaps are commonly employed for reconstruction, and many studies have sought to improve flap survival. In this study, we determined whether sildenafil and epinephrine synergistically improve the survival rate of transverse rectus abdominis musculocutaneous (TRAM) flaps.Materials and Method Twenty-four Sprague-Dawley rats were divided into four groups of six rats each: a control group, an epinephrine group, a sildenafil group, and an epinephrineplus- sildenafil group. TRAM flaps were elevated on the upper abdomen.Results The average flap survival area and number of blood vessels differed significantly between the control and experimental groups; however, the various experimental groups did not differ significantly. The level of vascular epithelial growth factor was higher in the experimental groups than in the control group.Conclusion Both epinephrine and sildenafil citrate significantly improve TRAM flap survival in rats. However, whether effects are better when the drugs are combined rather than when used alone remains unclear. Further research is required.

M2 macrophage-derived exosomes induce angiogenesis and increase skin flap survival through HIF1AN/HIF-1α/VEGFA control

BackgroundSkin flap transplantation is a routine strategy in plastic and reconstructive surgery for skin-soft tissue defects. Recent research has shown that M2 macrophages have the potential for pro-angiogenesis during tissue healing. MethodsIn our research, we extracted the exosomes from M2 macrophages(M2-exo) and applied the exosomes in the model of skin flap transplantation. The flap survival area was measured, and the choke vessels were assessed by morphological observation. Hematoxylin and eosin (H&E) staining and Immunohistochemistry were applied to assess the neovascularization. The effect of M2-exo on the function of Human umbilical vein endothelial cells (HUVECs) was also investigated. We also administrated 2-methoxyestradiol (2-ME2, an inhibitor of HIF-1α) to explore the underlying mechanism. We tested the effects of M2-Exo on the proliferation of HUVECs through CCK8 assay and EdU staining assay. ResultsThe survival area and number of micro-vessels in the skin flaps were increased in the M2-exo group. Besides, the dilation rate of choke vessels was also enhanced in the M2-exo group. Additionally, compared with the control group, M2-exo could accelerate the proliferation, migration and tube formation of HUVECs in vitro. Furthermore, the expression of the pro-angiogenesis factors, HIF-1α and VEGFA, were overexpressed with the treatment of the M2-exo. The expression of HIF1AN protein level was decreased in the M2-exo group. Finally, treatment with HIF-1α inhibitor reverses the pro-survival effect of M2-exo on skin flaps by interfering with the HIF1AN/HIF-1α/VEGFA signaling pathway. ConclusionThis study showed that M2-exosomes promote skin flap survival by enhancing angiogenesis, with HIF1AN/HIF-1α/VEGFA playing a crucial role in this process.

Tetrandrine promotes the survival of the random skin flap via the PI3K/AKT signaling pathway.

Flaps are mainly used for wound repair. However, postoperative ischemic necrosis of the distal flap is a major problem, which needs to be addressed urgently. We evaluated whether tetrandrine, a compound found in traditional Chinese medicine, can prolong the survival rate of random skin flaps. Thirty-six rats were randomly divided into control, low-dose tetrandrine (25 mg/kg/day), and high-dose tetrandrine (60 mg/kg/day) groups. On postoperative Day 7, the flap survival and average survival area were determined. After the rats were sacrificed, the levels of angiogenesis, apoptosis, and inflammation in the flap tissue were detected with immunology and molecular biology analyses. Tetrandrine increased vascular endothelial growth factor and Bcl-2 expression, in turn promoting angiogenesis and anti-apoptotic processes, respectively. Additionally, tetrandrine decreased the expression of Bax, which is associated with the induction of apoptosis, and also decreased inflammation in the flap tissue. Tetrandrine improved the survival rate of random flaps by promoting angiogenesis, inhibiting apoptosis, and reducing inflammation in the flap tissue through the modulation of the PI3K/AKT signaling pathway.

Nesfatin-1 regulates the HMGB1-TLR4-NF-κB signaling pathway to inhibit inflammation and its effects on the random skin flap survival in rats

ObjectiveRandom skin flaps are often placed by plastic surgeons to treat limb deformities and dysfunction. Nesfatin-1 (NES) is a peptide that exerts angiogenic, anti-inflammatory, and anti-oxidant effects. We assessed the impact of NES on flap survival and the underlying mechanism. MethodsWe modified the McFarlane random skin flap rat model. Thirty-six male Sprague-Dawley rats were randomly divided into a control group (corn oil solution with DMSO), low-dose group (NES-L at 10 µg/kg/day), and high-dose group (NES-H at 20 µg/kg/day). On day 7 after surgery, average flap survival areas were calculated. Laser Doppler blood flow monitoring and lead oxide/gelatin angiography were used to evaluate blood perfusion and neovascularization, respectively. Flap histopathological status was evaluated by hematoxylin and eosin (H&E) staining. The levels of superoxide dismutase (SOD) and malondialdehyde (MDA) were determined. Immunohistochemical techniques were used to evaluate the expression of angiogenetic and inflammatory factors. ResultsIn the experimental groups, the mean skin flap survival areas and blood perfusion increased considerably. The SOD activities in the experimental groups increased and the MDA contents decreased. Immunohistochemically, VEGF expression was upregulated in the experimental groups and the expression levels of inflammatory factors decreased markedly. ConclusionNES inhibited ischemic skin flap necrosis, promoted angiogenesis, and reduced ischemia-reperfusion injury and inflammation. Inhibition of the inflammatory HMGB1-TLR4-NF-κB signal pathway, which reduced flap inflammation and oxidative stress, may explain the enhanced flap survival.

Effects of pharmacological delay with roxadustat on multi-territory perforator flap survival in rats.

Roxadustat (FG-4592) is a specific hypoxia-inducible factor (HIF) prolyl hydroxylase inhibitor. We investigated the effects of FG-4592 pretreatment on survival and second choke vessels of multi-territory perforator flaps in rats. In total, 72 rats were divided into two groups (n = 36 each): the experimental (FG-4592) group and the control group. FG-4592 was administered orally as a single dose of 60 mg/kg every other day; the first drug solution was administered to the animals 7 days before the surgical procedure. On postoperative day 7, the surviving flap area was calculated. At 12 h post-surgery, in the second choke zone in the flaps, macrovascular hinges were compared by angiography and imaging, and microvascular changes were assessed by histology. Laser Doppler imaging was used to evaluate flap perfusion at the second choke zone at 12 h and 7 days after surgery. At 7 days after surgery, the flap survival area and perfusion were significantly greater in rats given FG-4592 compared with controls. At 12 h after surgery, the diameter of macrovascular and microvascular vessels, nitric oxide content, perfusion, and the protein levels of HIF-1α and inducible nitric oxide synthase were also significantly greater in FG-4592-treated rats than controls. In conclusion, pretreatment with roxadustat may improve initial flap survival and dilate the second choke zone vessels in a multi-territory perforator flap.

Activation of aldehyde dehydrogenase-2 improves ischemic random skin flap survival in rats.

Random skin flaps have many applications in plastic and reconstructive surgeries. However, distal flap necrosis restricts wider clinical utility. Mitophagy, a vital form of autophagy for damaged mitochondria, is excessively activated in flap ischemia/reperfusion (I/R) injury, thus inducing cell death. Aldehyde dehydrogenase-2 (ALDH2), an allosteric tetrameric enzyme, plays an important role in regulating mitophagy. We explored whether ALDH2 activated by N-(1,3-benzodioxol-5-ylmethyl)-2,6-dichlorobenzamide (Alda-1) could reduce the risk of ischemic random skin flap necrosis, and the possible mechanism of action. Modified McFarlane flap models were established in 36 male Sprague-Dawley rats assigned randomly to three groups: a low-dose Alda-1 group (10 mg/kg/day), a high-dose Alda-1 group (20 mg/kg/day) and a control group. The percentage surviving skin flap area, neutrophil density and microvessel density (MVD) were evaluated on day 7. Oxidative stress was quantitated by measuring the superoxide dismutase (SOD) and malondialdehyde (MDA) levels. Blood perfusion and skin flap angiogenesis were assessed via laser Doppler flow imaging and lead oxide-gelatin angiography, respectively. The expression levels of inflammatory cytokines (IL-1β, IL-6, and TNF-α), vascular endothelial growth factor (VEGF), ALDH2, PTEN-induced kinase 1 (PINK1), and E3 ubiquitin ligase (Parkin) were immunohistochemically detected. Indicators of mitophagy such as Beclin-1, p62, and microtubule-associated protein light chain 3 (LC3) were evaluated by immunofluorescence. Alda-1 significantly enhanced the survival area of random skin flaps. The SOD activity increased and the MDA level decreased, suggesting that Alda-1 reduced oxidative stress. ALDH2 was upregulated, and mitophagy-related proteins (PINK1, Parkin, Beclin-1, p62, and LC3) were downregulated, indicating that ALDH2 inhibited mitophagy through the PINK1/Parkin signaling pathway. Treatment with Alda-1 reduced neutrophil infiltration and expressions of inflammatory cytokines. Alda-1 significantly upregulated VEGF expression, increased the MVD, promoted angiogenesis, and enhanced blood perfusion. ALDH2 activation can effectively enhance random skin flap viability via inhibiting PINK1/Parkin-dependent mitophagy. Moreover, enhancement of ALDH2 activity also exerts anti-inflammatory and angiogenic properties.

Extrinsic Vascular Pathway Preservation Improves Survival in a Rat Three-Territory Flap Model Based on the Deep Circumflex Iliac Artery.

Extended flaps are commonly applied for large defects. However, a postoperative flap necrosis incidence of 11% to 44% remains a major complication. Previous clinical studies have shown that maintaining the extrinsic vascular pathway (EVP) can increase the survival area of extended flaps. The authors hypothesized that preserving the EVP would improve flap survival by reducing blood resistance within the vascular territory. Twenty-four adult male Sprague-Dawley rats were used. Tissue samples were obtained from eight untreated rats as a baseline control. Three-territory flaps were elevated in the remaining 16 rats. The EVP was preserved or ligated. Flap perfusion was assessed immediately using indocyanine green angiography. Rats were euthanized on day 7. The flap survival area was measured using Adobe Photoshop. Hematoxylin and eosin staining, CD31 immunostaining, and Western blot analysis of vascular endothelial growth factor protein expression were used to quantitatively assess vasodilation and angiogenesis in choke zones. Indocyanine green angiography revealed that blood could flow through the preserved EVP and perfuse the third vascular territory of the flap. EVP preservation significantly increased flap survival area (86.3%, 19.3% difference; P < 0.001), promoted vasodilation (5.0/choke zone, 3.0/choke zone difference; P = 0.013) and angiogenesis (29.3/mm 2 , 14.3/mm 2 difference; P = 0.002), and increased vascular endothelial growth factor expression (0.6, 0.2 difference; P = 0.067) in the second choke zone. EVP preservation improves flap survival in this rat three-territory flap model. Further investigation in large-animal models is required for clinical translation. Although further validation in large animal models and prospective clinical trials are necessary to verify the efficacy of the authors' hypothesis, their findings suggest that the EVP preservation procedure could provide an alternative for surgeons to create an extended flap in defect reconstruction.