Engineered Enucleated Mesenchymal Stem Cells Regulating Immune Microenvironment and Promoting Wound Healing.

Persistent excessive inflammation caused by neutrophil and macrophage dysfunction in the wound bed leads to refractory response during wound healing. However, previous studies using cytokines or drugs often suffer from short half-lives and limited targeting, resulting in unsatisfactory therapeutic effects. Herein, the enucleated mesenchymal stem cell is engineered by aptamer bioorthogonal chemistry to modify the cell membrane and mRNA loading in the cell cytoplasm as a novel delivery vector (Cargocyte) with accurate targeting and sustained cytokine secretion. Cargocytes can successfully reduce NETosis by targeting the nuclear chromatin protein DEK protein with aptamers and sustaining interleukin (IL)-4 expression to overcome the challenges associated with the high cost and short half-life of IL-4 protein and significantly prevent the transition of macrophages into the M1 phenotype. Therapeutic effects have been demonstrated in murine and porcine wound models and have powerful potential to improve wound immune microenvironments effectively. Overall, the use of engineered enucleated mesenchymal stem cells as a delivery system may be a promising approach for wound healing.

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.

Melatonin promotes peripheral nerve repair through Parkin-mediated mitophagy.

Schwann cells (SCs) are the major glial cells in peripheral nervous system. They unsheathe and myelinate axons and play an essential role in peripheral nerve regeneration. Several studies report that Parkin-mediated mitophagy is associated with various diseases. Melatonin promotes proliferation of central glial cells. Little is known about the effect of melatonin and Parkin-mediated mitophagy on peripheral nerve repair. In this study, using a rat model of a peripheral nerve injury (PNI) and in vitro model established by RSC96 cells treated with tert-butyl hydroperoxide (TBHP), we found that Parkin-mediated mitophagy can effectively reduce the production of mitochondrial reactive oxygen species (ROS), maintain the balance of mitochondrial membrane potential, maintain autophagic flux, and inhibit mitochondrial apoptosis. At the same time, we found that the increase of Parkin under stress is a manifestation of the RSC96 cells' resistance to oxidative stress to maintain RSC96 cells' balance. In our experiment, melatonin is similar to a Parkin agonist, up-regulating the expression of Parkin, enhancing all the positive results of Parkin in a stress state, such as inhibiting active oxygen production, maintaining autophagic flux, and inhibiting mitochondrial apoptosis. In addition, we design in vivo experiments to verify in In vitro experiments. In in vivo, melatonin promotes the expression of Parkin, maintains autophagic flux, inhibits apoptosis, promotes myelin regeneration, reduces the regeneration of collagen fibers around damaged tissues, and promotes peripheral nerve repair. When adenovirus was used to down-regulate the expression of Parkin, we found that all the positive effects of melatonin were attenuated. Collectively, these findings indicate that melatonin upregulates Parkin-mediated mitophagy and promotes peripheral nerve repair. The results provide a basis for development of effective drugs for PNI treatment.

MFG-E8 alleviates intervertebral disc degeneration by suppressing pyroptosis and extracellular matrix degradation in nucleus pulposus cells via Nrf2/TXNIP/NLRP3 axis.

Intervertebral disc degeneration (IVDD) is a chronic age-related degenerative disease accompanied by complex pathophysiological mechanisms. Increasing evidence indicates that NLRP3 inflammasome mediated pyroptosis of nucleus pulposus (NP) cells displays an important role in the pathological progression of IVDD. Milk fat globule-EGF factor-8 (MFG-E8) is an endogenously secreted glycoprotein with beneficial effects of anti-inflammatory, antioxidant, and modulation of NLRP3 inflammasome. However, the effect of MFG-E8 on IVDD remains unclear. In this study, our purpose is to clarify the expression changes of MFG-E8 in the IVDD process and explore the role and mechanism of MFG-E8. We found that MFG-E8's expression was reduced in degraded nucleus pulposus tissues of humans and rats as well as hydrogen peroxide (H2O2)-treated NP cells. Exogenous supplementation of MFG-E8 could rescue H2O2-induced oxidative stress, mitochondrial dysfunction, and NLRP3 inflammasome activation and protect NP cells from pyroptosis and extracellular matrix (ECM) degradation. Mechanistically, Nrf2/TXNIP/NLRP3 axis plays a crucial role in MFG-E8-mediated suppression of the above-pathological events. In vivo, we established a rat intervertebral disc acupuncture model and found that MFG-E8 administration effectively alleviated IVDD development by imageological and histomorphological evaluation. Overall, our findings revealed the internal mechanisms underlying MFG-E8 regulation in NP cells and its intrinsic value for IVDD therapy.

Sparstolonin B suppresses free fatty acid palmitate-induced chondrocyte inflammation and mitigates post-traumatic arthritis in obese mice.

Abnormal lipid metabolism, such as systemic increased free fatty acid, results in overproduction of pro-inflammatory enzymes and cytokines, which is crucial in the development of obesity-related osteoarthritis (OA). However, there are only a few drugs that target the lipotoxicity of OA. Recent researches have documented that the traditional Chinese medicine, Sparstolonin B (Ssn B), exerted anti-inflammatory effects in various diseases, but not yet in OA. On the basis of this evidence, our works purposed to evaluate the effect of Ssn B on free fatty acid (FFA) palmitate (PA)-stimulated human osteoarthritic chondrocytes and obesity-associated mouse OA model. We found that Ssn B suppressed PA-triggered inflammatory response and extracellular matrix catabolism in a concentration-dependent approach. In vivo, Ssn B treatment inhibited cartilage degeneration and subchondral bone calcification caused by joint mechanical imbalance and alleviated metabolic inflammation in obesity. Mechanistically, co-immunoprecipitine and molecular docking analysis showed that the formation of toll-like receptor 4 (TLR4)/myeloid differentiation protein-2 (MD-2) complex caused by PA was blocked by Ssn B. Subsequently, it leads to inactivation of PA-caused myeloid differentiation factor 88 (MyD88)-dependent nuclear factor-kappaB (NF-κB) cascade. Together, these findings demonstrated that Ssn B is a potential treatment agent for joint degenerative diseases in obese individuals.

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.

Inhibition of PI3K/Akt/NF-κB signaling by Aloin for ameliorating the progression of osteoarthritis: In vitro and in vivo studies.

Osteoarthritis (OA) is a progressive and degenerative joint disease. Aloin is a bitter and yellow-brown-coloured compound from the Aloe plant and is allowed for use in foods as a "natural flavour". In our study, we examined the protective effects of Aloin on the inhibition of OA development as well as its underlying mechanism in both in vitro and vivo experiments. In in-vitro experiments, the protective effect of aloin on the anabolism and catabolism of the extracellular matrix (ECM) induced by IL-1 ß in chondrocytes by inhibiting the expression of pro-inflammatory factors, including TNF-α (p = 0.016), IL-6 (p = 0.006), iNOS (p = 0.001) and COX-2 (p = 0.006). Mechanistically, Aloin suppressed the IL-1ß-induced activation of the PI3K/Akt/NF-κB signalling pathway cascades. Moreover, molecular docking studies demonstrated that Aloin bound strongly to PI3K. In vivo, Aloin ameliorated the OA process in the destabilization of the medial meniscus (DMM) model. In summary, our findings demonstrate that Aloin ameliorates the progression of OA via the PI3K/Akt/NF-κB signalling pathways, which supports Aloin as a promising therapeutic agent for the treatment of OA.

Detrimental Effect of Sitagliptin Induced Autophagy on Multiterritory Perforator Flap Survival.

Multiterritory perforator flap survival is commonly applied in surgical tissue reconstructions and covering of large skin defects. However, multiple risk factors such as ischemia, reperfusion injury, and apoptosis after reconstructive surgeries cause necrosis in distal parts with outcomes ranging from poor aesthetic appearance to reconstructive failure. A few studies have reported that sitagliptin (Sit) promotes angiogenesis and inhibits apoptosis. However, little is known about Sit-induced autophagy especially on the flap model. Therefore, our study investigated the effect of Sit and its induced autophagy on the perforator flap survival. Ninety male Sprague-Dawley rats were randomly separated into control, Sit, and Sit+3-methyladenine group. Results revealed that Sit significantly promoted flap survival by enhancing angiogenesis, reducing oxidative stress, and attenuating apoptosis. In addition, flap survival was further improved after co-administration with 3-methyladenine to inhibit autophagy. Overall, our results established that Sit has positive effects in promoting survival of multiterritory perforator flap. Sit-induced autophagy was detrimental for flap survival and its inhibition may further improve flap survival.

Butylphthalide Inhibits Autophagy and Promotes Multiterritory Perforator Flap Survival.

Multiterritory perforator flap is an important plastic surgery technique, yet its efficacy can be limited by partial necrosis at the choke Ⅱ zone. Butylphthalide (NBP) has been used for many diseases but has not been studied in the multiterritory perforator flap. With the effect of NBP, we observed increasing in capillary density, inhibition of autophagy and oxidative stress, and a reduction in apoptosis of cells, all consistent with increased flap survival. However, the protective effect of NBP on multiterritory perforator flap was lost following administration of the autophagy agonist rapamycin (Rap). Through the above results, we assumed that NBP promotes flap survival by inhibiting autophagy. Thus, this study has found a new pharmacological effect of NBP on the multiterritory perforator by inhibiting autophagy to prevent distal postoperative necrosis and exert effects on angiogenesis, oxidative stress, and apoptosis within the flap.