Apoptotic Bodies Derived from Fibroblast-Like Cells in Subcutaneous Connective Tissue Inhibit Ferroptosis in Ischaemic Flaps via the miR-339-5p/KEAP1/Nrf2 Axis.

Preventing and treating avascular necrosis at the distal end of the flaps are critical to surgery success, but current treatments are not ideal. A recent study shows that apoptotic bodies (ABs) generated near the site of apoptosis can be taken up and promote cell proliferation. The study reveals that ABs derived from fibroblast-like cells in the subcutaneous connective tissue (FSCT cells) of skin flaps promoted ischaemic flap survival. It is also found that ABs inhibited cell death and oxidative stress and promoted M1-to-M2 polarization in macrophages. Transcriptome sequencing and protein level testing demonstrated that ABs promoted ischaemic flap survival in endothelial cells and macrophages by inhibiting ferroptosis via the KEAP1-Nrf2 axis. Furthermore, microRNA (miR) sequencing data and in vitro and in vivo experiments demonstrated that ABs inhibited KEAP1 by delivering miR-339-5p to exert therapeutic effects. In conclusion, FSCT cell-derived ABs inhibited ferroptosis, promoted the macrophage M1-to-M2 transition via the miR-339-5p/KEAP1/Nrf2 axis and promoted ischaemic flap survival. These results provide a potential therapeutic strategy to promote ischaemic flap survival by administering ABs.

Dihydrocapsaicin suppresses the STING-mediated accumulation of ROS and NLRP3 inflammasome and alleviates apoptosis after ischemia-reperfusion injury of perforator skin flap.

Avascular necrosis frequently occurs as a complication following surgery involving the distal perforator flap. Dihydrocapsaicin (DHC) can protect tissue from ischemia-reperfusion (I/R) injury, but its specific role in multizone perforator flaps remains unclear. In this study, the prospective target of DHC in the context of I/R injury was predicted using network pharmacology analysis. Flap viability was determined through survival area analysis, laser Doppler blood flow, angiograms, and histological examination. The expressions of angiogenesis, apoptosis, NLR family pyrin domain containing 3 (NLRP3) inflammasome, oxidative stress, and molecules related to cyclic guanosine monophosphate (GMP)-adenosine monophosphate synthase (cGAS)-interferon gene stimulant (STING) pathway were assessed using western blotting, immunofluorescence, TUNEL staining, and dihydroethidium (DHE) staining. Our finding revealed that DHC promoted the perforator flap survival, which involves the cGAS-STING pathway, oxidative stress, NLRP3 inflammasome, apoptosis, and angiogenesis. DHC induced oxidative stress resistance and suppressed the NLRP3 inflammasome, preventing apoptosis in vascular endothelial cells. Through regulation of STING pathway, DHC controlled oxidative stress in endothelial cells and NLRP3 levels in ischemic flaps. However, activation of the cGAS-STING pathway led to the accumulation of reactive oxygen species (ROS) and NLRP3 inflammasome, thereby diminishing the protective role of DHC. DHC enhanced the survival of multidomain perforator flaps by suppressing the cGAS-STING pathway, oxidative stress, and the formation of NLRP3 inflammasome. These findings unveil a potentially novel mechanism with clinical significance for promoting the survival of multidomain perforator flaps.

A snake cathelicidin enhances transcription factor EB-mediated autophagy and alleviates ROS-induced pyroptosis after ischaemia-reperfusion injury of island skin flaps.

BACKGROUND AND PURPOSE: Ischaemia-reperfusion (I/R) injury is a major contributor to skin flap necrosis, which presents a challenge in achieving satisfactory therapeutic outcomes. Previous studies showed that cathelicidin-BF (BF-30) protects tissues from I/R injury. In this investigation, BF-30 was synthesized and its role and mechanism in promoting survival of I/R-injured skin flaps explored. EXPERIMENTAL APPROACH: Survival rate analysis and laser Doppler blood flow analysis were used to evaluate I/R-injured flap viability. Western blotting, immunofluorescence, TdT-mediated dUTP nick end labelling (TUNEL) and dihydroethidium were utilized to examine the levels of apoptosis, pyroptosis, oxidative stress, transcription factor EB (TFEB)-mediated autophagy and molecules related to the adenosine 5'-monophosphate-activated protein kinase (AMPK)-transient receptor potential mucolipin 1 (TRPML1)-calcineurin signalling pathway. KEY RESULTS: The outcomes revealed that BF-30 enhanced I/R-injured island skin flap viability. Autophagy, oxidative stress, pyroptosis and apoptosis were related to the BF-30 capability to enhance I/R-injured flap survival. Improved autophagy flux and tolerance to oxidative stress promoted the inhibition of apoptosis and pyroptosis in vascular endothelial cells. Activation of TFEB increased autophagy and inhibited endothelial cell oxidative stress in I/R-injured flaps. A reduction in TFEB level led to a loss of the protective effect of BF-30, by reducing autophagy flux and increasing the accumulation of reactive oxygen species (ROS) in endothelial cells. Additionally, BF-30 modulated TFEB activity via the AMPK-TRPML1-calcineurin signalling pathway. CONCLUSION AND IMPLICATIONS: BF-30 promotes I/R-injured skin flap survival by TFEB-mediated up-regulation of autophagy and inhibition of oxidative stress, which may have possible clinical applications.

Machine learning application for prediction of surgical site infection after posterior cervical surgery.

Surgical site infection (SSI) is one of the most common complications of posterior cervical surgery. It is difficult to diagnose in the early stage and may lead to severe consequences such as wound dehiscence and central nervous system infection. This retrospective study included patients who underwent posterior cervical surgery at The Second Affiliated Hospital and Yuying Childrens Hospital of Wenzhou Medical University from September 2018 to June 2022. We employed several machine learning methods, such as the gradient boosting (GB), random forests (RF), artificial neural network (ANN) and other popular machine learning models. To minimize the variability introduced by random splitting, the results underwent 10-fold cross-validation repeated 10 times. Five measurements were averaged across 10 repetitions with 10-fold cross-validation, the RF model achieved the highest AUROC (0.9916), specificity (0.9890) and precision (0.9759). The GB model achieved the highest sensitivity (0.9535) and the KNN achieved the highest sensitivity (0.9958). The application of machine learning techniques facilitated the development of a precise model for predicting SSI after posterior cervical surgery. This dynamic model can be served as a valuable tool for clinicians and patients to assess SSI risk and prevent it in clinical practice.

Tetrahydropalmatine: Orchestrating survival - Regulating autophagy and apoptosis via the PI3K/AKT/mTOR pathway in perforator flaps.

BACKGROUND: Introduced in clinical practice in 1989, perforator flaps are vital for tissue defect repair, but they are challenged by distal necrosis. Tetrahydropalmatine (THP) from celandine is renowned for its anti-inflammatory and analgesic effects. This study investigates THP's use in perforator flaps. METHODS: Thirty rats were divided into a control group and four THP concentration groups, while seventy-eight rats were categorized as control, THP, THP combined with rapamycin (RAP), and RAP alone. We created 11 cm by 2.5 cm multi-regional perforator flaps on rat backs, assessing survival blood flow and extracting skin flap tissue for autophagy, oxidative stress, apoptosis, and angiogenesis markers. RESULTS: The THP group exhibited significantly reduced distal necrosis, increased blood flow density, and survival area on the seventh day compared to controls. Immunohistochemistry and Western blot results demonstrated improved anti-oxidative stress and angiogenesis markers, along with decreased autophagy and apoptosis indicators. Combining THP with RAP diminished flap survival compared to THP alone. This was supported by protein expression changes in the PI3K-AKT-mTOR pathway. CONCLUSION: THP enhances flap survival by modulating autophagy, reducing tissue edema, promoting angiogenesis, and mitigating apoptosis and oxidative stress. THP offers a potential strategy for enhancing multi-regional perforator flap survival through the PI3K/AKT/mTOR pathway. These findings highlight THP's promise in combatting perforator flap necrosis, uncovering a novel mechanism for its impact on flap survival.

High-pressure infusion improves multi-territory perforator flap viability via choke artery dilation: A preliminary study in a rat model.

BACKGROUND: Multi-territory perforator flaps have become the preferred option for the repair and reconstruction of large soft tissue defects. Although methods (e.g., pharmacological agents, mechanical stimulation, and thermal stimulation) were developed to open choke vessels to improve flap survival, the flap necrosis rate is still as high as 28.8%. The authors hypothesized that high-pressure infusion might enhance flap viability by dilating choke arteries intraoperatively in a rat model of multi-territory perforator flap. METHODS: Two-month-old male Sprague-Dawley rats were randomized into two groups (n = 32 each). During the multi-territory perforator flap elevation based on the right superficial epigastric angiosome, one group received continuous high-pressure infusion (mean pressure, 250 mmHg; duration, 1 min) of an isotonic heparin sodium solution (12,500 U/L) via the artery in the pedicle, whereas the other group received no infusion. At 7 days postoperatively, arteriography was performed; endothelial nitric oxide synthase (eNOS) and vascular endothelial growth factor (VEGF) expression and microvascular density were evaluated by western blot and histology, respectively; and flap survival was compared. Moreover, intraluminal diameters were examined at 1 day and 7 days postoperatively using hematoxylin and eosin staining, and coagulation function was assessed immediately postoperatively. RESULTS: High-pressure infusion significantly promoted the dilation of choke arteries at 1 day and 7 days postoperatively. It also increased eNOS and VEGF expression, flap survival, and microvascular density. The coagulation function remained unaffected. CONCLUSIONS: High-pressure infusion allowed intraoperative and postoperative dilation of the choke arteries that enhanced the viability of multi-territory perforator flaps in rats.

Efficacy of a fingertip reconstruction technique using the tongue-shaped flap and advancement flap for repairing congenital syndactyly with osseous fusion of the distal phalanges.

BACKGROUND: To explore the clinical efficacy of using tongue-shaped flaps and advancement flaps to reconstruct the fingertips in congenital syndactyly patients with osseous fusion of the distal phalanges. METHODS: From January 2016 to January 2019, 12 patients with congenital syndactyly, involving 30 digits in total, presented to our hospital and were surgically treated with tongue-shaped flaps, as well as with advancement flaps to reconstruct the fingertips. The flap infection rate, necrosis rate and any other early complications were recorded. Fingertip aesthetics were reported according to the modified Bulic scale. A questionnaire was used to assess the satisfaction of the patients' family members. RESULTS: All cases were thoroughly reviewed. The postoperative period for inclusion in this study ranged from 36 to 60 months, with an average follow-up time of 45 months. During this period, no complications such as infection and/or necrosis of any flap were observed. Significant improvements in finger aesthetics and functioning compared to preoperative values were observed in most cases. Based on the modified Bulic scale, of 30 fingertips, an excellent result was obtained for 3, a very good result for 13, a good result for 13 and a poor result for just 1. Family members were satisfied with the treatment outcome. CONCLUSIONS: This technique employing tongue-shaped flaps and advancement flaps to reconstruct fingertips is effective, which enables the attainment of favourable aesthetic and functional outcomes in congenital syndactyly patients with osseous fusion of the distal phalanges.

Management of wounds with exposed bone structures using an induced-membrane followed by polymethyl methacrylate and second-stage skin grafting in the elderly with a 3-year follow-up.

The treatment of traumatic wounds with exposed bone or tendons is often challenging. An induced membrane (IM) is used to reconstruct bone defects, as it provides an effective and sufficient blood supply for bone and soft-tissue reconstruction. This study explored a novel two-stage strategy for wound management, consisting of initial wound coverage with polymethyl methacrylate (PMMA) and an autologous split-thickness skin graft under the IM. Fifty inpatients were enrolled from December 2016 to December 2019. Each patient underwent reconstruction according to a two-stage process. In the first stage, the defect area was thoroughly debrided, and the freshly treated wound was then covered using PMMA cement. After 4-6 weeks, during the second stage, the PMMA cement was removed to reveal an IM covering the exposed bone and tendon. An autologous split-thickness skin graft was then performed. Haematoxylin and eosin (H&E) staining and immunohistochemical analysis of vascular endothelial growth factor (VEGF), CD31 and CD34 were used to evaluate the IM and compare it with the normal periosteal membrane (PM). The psychological status and the Lower Extremity Function Scale (LEFS) as well as any complications were recorded at follow-up. We found that all skin grafts survived and evidenced no necrosis or infection. H&E staining revealed vascularised tissue in the IM, and immunohistochemistry showed a larger number of VEGF-, CD31- and CD34-positive cells in the IM than in the normal PM. The duration of healing in the group was 5.40 ± 1.32 months with a mean number of debridement procedures of 1.92 ± 0.60. There were two patients with reulceration in the group. The self-rating anxiety scale scores ranged from 35 to 60 (mean 48.02 ± 8.12). Postoperatively, the LEFS score was 50.10 ± 9.77. Finally, our strategy for the management of a non-healing wound in the lower extremities, consisting of an IM in combination with skin grafting, was effective, especially in cases in which bony structures were exposed in the elderly. The morbidity rate was low.

Activating Parkin-dependent mitophagy alleviates oxidative stress, apoptosis, and promotes random-pattern skin flaps survival.

The random-pattern skin flap is a crucial technique in reconstructive surgery and flap necrosis caused by ischemia/reperfusion injury is a major postoperative complication. Herein, we investigated the mechanism of mitophagy induced by Melatonin (ML) and its effect on the survival of skin flaps. Our results demonstrated that ML could activate mitophagy, ameliorate oxidative stress and alleviate apoptosis in Tert-Butyl hydroperoxide solution (TBHP)-stimulated human umbilical vein endothelial cells in vitro. Inhibiting ML-induced mitophagy considerably abolished its protective effects. Moreover, knockdown of Parkin by siRNA inhibited ML-induced mitophagy, and subsequently exacerbated oxidative stress and apoptosis. Further study demonstrated that inhibition of AMPK reversed these protective effects of ML and downregulated the expression of TFEB. In the vivo study, ML effectively promoted flap survival by activating mitophagy and subsequently ameliorating oxidative stress and mitigating apoptosis. These results established that ML is a potent agent capable for increasing random-pattern skin flap survival by activating Parkin-dependent mitophagy through the AMPK-TFEB signaling pathway.

Which one is better for multi-territory perforator flap survival, central perforator artery or central perforator vein?

BACKGROUND: The survival of multi-territory perforator flap is associated with the position of the perforators. This study aimed to explore whether use of the central perforator artery or vein was better for flap survival. METHODS: 75 male Sprague-Dawley rats were randomly divided into three groups (n=25 per group). The flap contained the right and left iliolumbar, left posterior intercostal, and left thoracodorsal angiosomes, termed angiosomes Ⅰ to Ⅳ, respectively. The anastomosis between angiosomes Ⅱ and Ⅲ was termed choke 2. In experimental group 2, only the right iliolumbar vein and the left iliolumbar artery were preserved; in experimental group 1, only the right iliolumbar artery and the left iliolumbar vein were preserved; and in the control group, only the right iliolumbar artery and vein were preserved. On day-7 after the operation, the flap arteriography, intraluminal diameter, average microvascular density, vascular endothelial growth factor (VEGF) expression and flap survival were compared among groups. Moreover, the percentages of the angiosomes were measured. RESULTS: The dilation of the choke 2 artery was most pronounced in experimental group 2, followed by experimental group 1, and, finally, the control group (p<0.05). Similar results regarding average microvascular density, VEGF expression, and survival rate were found among the three groups. The percentages of angiosomes Ⅰ to Ⅳ were 23.1%, 23.0±3.1%, 23.0±1.9%, and 31.0±3.1%, respectively. CONCLUSIONS: Compared with the central perforator vein, the central perforator artery was more beneficial in enhancing flap survival. A multi-territory perforator flap with the central perforator artery could capture 3 angiosomes safely.

Primary closure using a perforator flap-based technique for the treatment of Dupuytren's contracture.

We aim to present a perforator flap-based technique that is useful in the resection and reconstruction of the palmar fascia for the treatment of Dupuytren's contracture with primary closure. A curve incision through the palmar skin radial to the hypothenar area was made. The ulnar side of the palmar skin and the subcutaneous fat was raised to the ulnar side, exposing the palmar fascia. The ulnar palmar digital artery extending from the superficial palmar arch curves distally towards the little finger at a point perpendicular to the fourth interdigital space with parting branches into the subcutaneous fat radial of the flap was carefully identified and preserved, and the Dupuytren's cords were excised. Depending on contracture involvement, additional incision extending from the arc of the palmar incision to the proximal interphalangeal joint is made to raise the digital flap similar to that of the palmar incision. The perforator flap was raised along the hypothenar region in 53 hands of 48 patients, nine women and 39 men, and their age at the time of surgery averaged 56 years. Two patients complained of paresthesia in the ring and little fingers after surgery in 2015, and the symptom had disappeared without further intervention before the latest follow-up in 2017. There was no incidence of skin necrosis, delayed healing, and no recurrence within the follow-up period. This perforator flap-based technique is technically reliable and straightforward with better exposure and easier removal of all the diseased fascia, making it possible for primary healing without skin necrosis and acceptable for the treatment of patients at all stages of the disease.

A hitchhiking approach to reconstruct the finger pulp and the subsequent 1st toe hemi-pulp donor site defect.

BACKGROUND: The free 1st toe hemi-pulp transfer for finger pulp reconstruction was acknowledged as the optimal one. However, the treatment of the 1st toe donor defect, owning to the impossibility of primary closure, was frequently oversimplified. This study presented a "hitchhiking" approach to resurface finger pulp and the subsequent 1st toe donor site defect in a one-stage procedure. METHODS: From 2014 to 2019, finger pulp amputations (13 digits in 12 patients) were reconstructed with free 1st toe pulp flaps, and the donor site was resurfaced by the 2nd toe pedicled flap with the 2nd toe's primary closure. Therapeutic evaluation of repaired fingers and toes was based on cold intolerance, two-point discrimination (2PD), and gait disturbance. RESULTS: All finger and toe pulp flaps survived uneventfully. The average size of free 1st toe and pedicled 2nd toe flap was 3.1 cm × 2.0 cm (3.5 cm × 1.4 cm to 4.2 cm × 2.5 cm) and 3.0 cm × 1.1 cm (2.0 cm × 0.9 cm to 3.8 cm × 1.5 cm), respectively. The regained average static 2PD on the finger and 1st toe pulps was 6 mm (ranged 5-10 mm) and 4 mm (ranged 2-6 mm), respectively. All reconstructed 1st toe pulps were qualified for normal gait. One patient complained the mild cold intolerance, and hammer-toe deformities were involved in two cases. CONCLUSION: To fulfill donor site care and cost-effective rule, the toe-to-finger pulp reconstruction can't underestimate the morbidity on 1st toe donor site due to inappropriate intervention. Equally importantly, the hitchhiking pedicled 2nd toe flap should be recruited in the reconstructive scheme.

Inhibition of PLA2G4E/cPLA2 promotes survival of random skin flaps by alleviating Lysosomal membrane permeabilization-Induced necroptosis.

Necrosis that appears at the ischemic distal end of random-pattern skin flaps increases the pain and economic burden of patients. Necroptosis is thought to contribute to flap necrosis. Lysosomal membrane permeabilization (LMP) plays an indispensable role in the regulation of necroptosis. Nonetheless, the mechanisms by which lysosomal membranes become leaky and the relationship between necroptosis and lysosomes are still unclear in ischemic flaps. Based on Western blotting, immunofluorescence, enzyme-linked immunosorbent assay, and liquid chromatography-mass spectrometry (LC-MS) analysis results, we found that LMP was presented in the ischemic distal portion of random-pattern skin flaps, which leads to disruption of lysosomal function and macroautophagic/autophagic flux, increased necroptosis, and aggravated necrosis of the ischemic flaps. Moreover, bioinformatics analysis of the LC-MS results enabled us to focus on the role of PLA2G4E/cPLA2 (phospholipase A2, group IVE) in LMP of the ischemic flaps. In vivo inhibition of PLA2G4E with an adeno-associated virus vector attenuated LMP and necroptosis, and promoted flap survival. In addition, microRNA-seq helped us determine that Mir504-5p was differentially expressed in ischemic flaps. A string of in vitro and in vivo tests was employed to verify the inhibitory effect of Mir504-5p on PLA2G4E, LMP and necroptosis. Finally, we concluded that the inhibition of PLA2G4E by Mir504-5p reduced LMP-induced necroptosis, thereby promoting the survival of random-pattern skin flaps.Abbreviations: AAV: adeno-associated virus; ACTA2/α;-SMA: actin alpha 2, smooth muscle, aorta; ALOX15/12/15-LOX: arachidonate 15- lipoxygenase; c-CASP8: cleaved caspase; c-CASP3: cleaved caspase 3; CTSD: cathepsin D; CTSB: cathepsin B; CTSL: cathepsin L; DMECs: primary mouse dermal microvascular endothelial cells; ELISA: enzyme-linked immunosorbent assay; F-CHP: 5-FAM-conjugated collagen hybridizing peptide; FISH: fluorescence in situ hybridization; HUVECs: human umbilical vein endothelial cells; LAMP1: lysosomal-associated membrane protein 1; LAMP2: lysosomal-associated membrane protein 2; LC-MS: liquid chromatography-mass spectrometry; LDBF: laser doppler blood flow; LMP: lysosomal membrane permeabilization; LPE: lysophosphatidylethanolamine; LPC: lysophosphatidylcholine; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MLKL: mixed lineage kinase domain-like; NDI: N-dodecylimidazole; PECAM1/CD31: platelet/endothelial cell adhesion molecule 1; PLA2G4A/cPLA2: phospholipase A2, group IVA (cytosolic, calcium-dependent); PLA2G4E/cPLA2: phospholipase A2, group IVE; qPCR: quantitative real-time polymerase chain reaction; RIPK1: receptor (TNFRSF)-interacting serine-threonine kinase 1; RIPK3: receptor-interacting serine-threonine kinase 3; RISC: RNA-induced silencing complex; ROS: reactive oxygen species; shRNA: short hairpin RNA; SQSTM1: sequestosome 1; TBHP: tert-butyl hydroperoxide; TUNEL: terminal deoxynucleotidyl transferase dUTP nick end labelling.

Paeoniflorin Suppresses TBHP-Induced Oxidative Stress and Apoptosis in Human Umbilical Vein Endothelial Cells via the Nrf2/HO-1 Signaling Pathway and Improves Skin Flap Survival.

Random-pattern skin flap is a vital technique frequently applied in reconstruction surgeries for its convenience and effectiveness in solving skin defects. However, ischemic necrosis, especially in the distal areas of the flap, still needs extra attention after surgery. Earlier evidence has suggested that paeoniflorin (PF) could stimulate angiogenesis and suppress ischemic cardiovascular disease. However, few studies have focused on the role of PF in flap survival. In this study, we have demonstrated that the human umbilical vein endothelial cells (HUVECs) treated with PF can alleviate tert-butyl hydroperoxide (TBHP)-stimulated cellular dysfunction and apoptosis. To better evaluate, HUVECs' physiology, cell tube formation, migration, and adhesion were assessed. Mechanistically, PF protects HUVECs against apoptosis via stimulating the nuclear factor (erythroid-derived 2)-like 2 (Nrf2)/heme oxygenase 1 (HO-1) pathway. PF also downregulates mitochondrial ROS production to reduce excessive intracellular ROS production induced by TBHP and restore TBHP-induced mitochondrial depolarization. As a result, silencing Nrf2 partially abolishes the protective effect of PF exposure on HUVECs. In in vivo experiments, the oral administration of PF was shown to have enhanced the vascularization of regenerated tissues and promote flap survival. However, the PF-mediated protection was partially lost after co-treatment with ML385, a selective Nrf2 inhibitor, suggesting that PF is a crucial modulator regulating the Nrf2/HO-1 signaling pathway. In summary, our data have provided a new insight into PF as a potential therapy for enhancing random-pattern flap viability.

Betulin Alleviates the Inflammatory Response in Mouse Chondrocytes and Ameliorates Osteoarthritis via AKT/Nrf2/HO-1/NF-κB Axis.

Osteoarthritis (OA) is a common degenerative joint disease featuring the degeneration, destruction, and ossification of cartilage. Inflammation which may facilitate OA occurrence and development is considered as the main pathological factor. Betulin, a natural product extracted from birch bark, has been commonly used for inflammation treatment; however, its role in OA remains unclear. This study is aimed to explore whether betulin can suppress IL-1ß-induced inflammation in chondrocytes and alleviate OA in vitro and in vivo. In in vitro studies, the generation of pro-inflammatory factors, such as interleukin-6 (IL-6), tumor necrosis factor alpha (TNF-α), prostaglandin E2 (PGE2), and nitric oxide (NO), was assessed using the enzyme-linked immunosorbent assay (ELISA) and Griess reaction. As revealed by results, betulin inhibited the expression of pro-inflammatory mediators. In addition, the protein expressions of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), matrix metalloproteinase (MMP-13), thrombospondin motifs 5 (ADAMTS5), Collagen II, and Aggrecan were quantified using Western blot analysis. We found that betulin could inhibit the generation of COX-2 and iNOS induced by IL-1ß, indicating that betulin has anti-inflammatory effects in chondrocytes. Furthermore, betulin downregulates the expression of MMP-13 and ADAMTS-5 and upregulates the expression of Collagen II and Aggrecan, indicating that it can inhibit the degradation of the extracellular matrix. In mechanism, betulin activated the AKT/Nrf2 pathway and inhibited the phosphorylation of p65. In in vivo studies, administration of betulin in vivo could inhibit cartilage destruction and inflammatory progression. Therefore, these findings suggest that betulin may alleviate IL-1ß-induced OA via the AKT/Nrf2/HO-1/NF-κB signal axis, and betulin may be a potential drug for the treatment of OA.

Distal Arterialized Venous Supercharging Improves Perfusion and Survival in an Extended Dorsal Three-Perforasome Perforator Flap Rat Model.

BACKGROUND: Perforator flaps are commonly applied for a variety of skin defects. Many strategies (e.g., hyperbaric oxygen and preconditioning) have been investigated to improve flap survival, but a postoperative 2.03 to 18.2 percent flap necrosis frequency remains a major complication. The authors hypothesized that a distal arterialized venous supercharged (DAVS) flap procedure might improve perfusion and survival in an extended three-perforasome perforator flap rat model and rescue flap ischemia intraoperatively. METHODS: One hundred twenty male Sprague-Dawley rats (200 to 300 g) were divided into the thoracodorsal artery (TDA) flap group and the DAVS flap group (n = 60 per group). An approximately 11 × 2.5-cm2 flap based on the TDA perforasome was designed in the TDA flap. A DAVS flap was designed based on the TDA flap and supercharged by anastomosing the rat caudal artery with the deep circumflex iliac vein. At postoperative times 1, 3, 6, and 12 hours and 1, 3, 5, and 7 days, perfusion and angiography were compared. On day 7, flap viability and angiogenesis were assessed using histology and Western blotting. RESULTS: The DAVS flap showed a higher survival rate compared with the TDA flap (100 percent versus 81.93 ± 5.38 percent; p < 0.001). All blood flow ratios of deep circumflex iliac artery to TDA perforasome and of choke zone II to choke zone I were higher in the DAVS flap (all p < 0.05). Angiography qualitatively revealed that choke vessels in choke zone II dilated earlier and extensively in the DAVS flap group. CD34+ vessels (68.66 ± 12.53/mm2 versus 36.82 ± 8.99/mm2; p < 0.001) and vascular endothelial growth factor protein level (0.22 ± 0.03 versus 0.11 ± 0.03; p < 0.001) were significantly increased in the DAVS flap group. CONCLUSIONS: The DAVS procedure improves three-perforasome perforator flap survival and can be used for rescuing flap ischemia intraoperatively. Further study is needed before possible clinical adoption for reconstructive operations.

Liraglutide, a TFEB-Mediated Autophagy Agonist, Promotes the Viability of Random-Pattern Skin Flaps.

Random skin flaps are commonly used in reconstruction surgery. However, distal necrosis of the skin flap remains a difficult problem in plastic surgery. Many studies have shown that activation of autophagy is an important means of maintaining cell homeostasis and can improve the survival rate of flaps. In the current study, we investigated whether liraglutide can promote the survival of random flaps by stimulating autophagy. Our results show that liraglutide can significantly improve flap viability, increase blood flow, and reduce tissue oedema. In addition, we demonstrated that liraglutide can stimulate angiogenesis and reduce pyroptosis and oxidative stress. Through immunohistochemistry analysis and Western blotting, we verified that liraglutide can enhance autophagy, while the 3-methylladenine- (3MA-) mediated inhibition of autophagy enhancement can significantly reduce the benefits of liraglutide described above. Mechanistically, we showed that the ability of liraglutide to enhance autophagy is mediated by the activation of transcription factor EB (TFEB) and its subsequent entry into the nucleus to activate autophagy genes, a phenomenon that may result from AMPK-MCOLN1-calcineurin signalling pathway activation. Taken together, our results show that liraglutide is an effective drug that can significantly improve the survival rate of random flaps by enhancing autophagy, inhibiting oxidative stress in tissues, reducing pyroptosis, and promoting angiogenesis, which may be due to the activation of TFEB via the AMPK-MCOLN1-calcineurin signalling pathway.

Cardamonin protects nucleus pulposus cells against IL-1ß-induced inflammation and catabolism via Nrf2/NF-κB axis.

Intervertebral disc degeneration (IVDD) is one of the major causes of low back pain, but effective therapies are still lacking because of its complicated pathology. It has been demonstrated that increased levels of interleukin-1ß (IL-1ß) may promote the development of IVDD. Cardamonin (CAR) is a chalcone extracted from Alpinia katsumadai and other plants. It exhibits an anti-inflammatory effect in multiple diseases. In the present study, we investigated the protective effects of CAR on rat nucleus pulposus (NP) cells under IL-1ß stimulation in vitro and in a puncture-induced rat IVDD model in vivo. We explored the CAR treatment's inhibition of the expression of inflammatory factors such as cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), prostaglandin E2 (PGE2), nitric oxide (NO), tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) in rat NP cells. Moreover, the up-regulation of matrix metalloproteinase-13 (MMP-13) and thrombospondin motifs 5 (ADAMTS-5) and the degradation of aggrecan and collagen II induced by IL-1ß were reversed by CAR. Mechanistically, we demonstrated that CAR inhibited nuclear factor kappa B (NF-κB) signaling by activating the nuclear factor erythroid-derived 2-like 2 (Nrf2) in IL-1ß-induced rat NP cells. Furthermore, the protective effect of CAR was shown in the IVDD model through persistent intragastric administration. Taken together, our results revealed that CAR could activate the Nrf2/HO-1 signaling axis and be a novel agent for IVDD therapy.

Apelin/APJ-Manipulated CaMKK/AMPK/GSK3ß Signaling Works as an Endogenous Counterinjury Mechanism in Promoting the Vitality of Random-Pattern Skin Flaps.

A random-pattern skin flap plays an important role in the field of wound repair; the mechanisms that influence the survival of random-pattern skin flaps have been extensively studied but little attention has been paid to endogenous counterinjury substances and mechanism. Previous reports reveal that the apelin-APJ axis is an endogenous counterinjury mechanism that has considerable function in protecting against infection, inflammation, oxidative stress, necrosis, and apoptosis in various organs. As an in vivo study, our study proved that the apelin/APJ axis protected the skin flap by alleviating vascular oxidative stress and the apelin/APJ axis works as an antioxidant stress factor dependent on CaMKK/AMPK/GSK3ß signaling. In addition, the apelin/APJ-manipulated CaMKK/AMPK/GSK3ß-dependent mechanism improves HUVECs' resistance to oxygen and glucose deprivation/reperfusion (OGD/R), reduces ROS production and accumulation, maintained the normal mitochondrial membrane potential, and suppresses oxidative stress in vitro. Besides, activation of the apelin/APJ axis promotes vascular migration and angiogenesis under relative hypoxia condition through CaMKK/AMPK/GSK3ß signaling. In a word, we provide new evidence that the apelin/APJ axis is an effective antioxidant and can significantly improve the vitality of random flaps, so it has potential be a promising clinical treatment.