Platinum Nanoparticles Regulated V2 C MXene Nanoplatforms with NIR-II Enhanced Nanozyme Effect for Photothermal and Chemodynamic Anti-Infective Therapy.

Given the challenge of multidrug resistance in antibiotics, non-antibiotic-dependent antibacterial strategies show promise for anti-infective therapy. V2 C MXene-based nanomaterials have demonstrated strong biocompatibility and photothermal conversion efficiency (PCE) for photothermal therapy (PTT). However, the limitation of V2 C MXene's laser irradiation to the near-infrared region I (NIR-I) restricts tissue penetration, making it difficult to achieve complete bacterial eradication with single-effect therapeutic strategies. To address this, Pt nanoparticles (Pt NPs) are attached to V2 C, forming artificial nanoplatforms (Pt@V2 C). Pt@V2 C exhibits enhanced PCE (59.6%) and a longer irradiation laser (NIR-II) due to the surface plasmon resonance effect of Pt NPs and V2 C. Notably, Pt@V2 C displays dual enzyme-like activity with chemodynamic therapy (CDT) and NIR-II enhanced dual enzyme-like activity. The biocatalytic mechanism of Pt@V2 C is elucidated using density functional theory. In an in vivo animal model, Pt@V2 C effectively eliminates methicillin-resistant Staphylococcus aureus from deep-seated tissues in subcutaneous abscesses and bacterial keratitis environments, accelerating abscess resolution and promoting wound and cornea healing through the synergistic effects of PTT/CDT. Transcriptomic analysis reveals that Pt@V2 C targets inflammatory pathways, providing insight into its therapeutic mechanism. This study presents a promising therapeutic approach involving hyperthermia-amplified biocatalysis with Pt NPs and MXene nanocomposites.

Ursolic acid alleviates steroid-induced avascular necrosis of the femoral head in mouse by inhibiting apoptosis and rescuing osteogenic differentiation.

Steroid-induced avascular necrosis of femoral head (SANFH) is a common disorder worldwide with high disability. Overdose of glucocorticoid (GC) is the most common non-traumatic cause of SANFH. Up until now, there are limited therapeutic strategies for curing SANFH, and the mechanisms underlying SANFH progression remain unclear. Nevertheless, Osteogenic dysfunction is considered to be one of the crucial pathobiological mechanisms in the development of SANFH, which involves mouse bone marrow mesenchymal stem cells (BMSCs) apoptosis and osteogenic differentiation disorder. Ursolic acid (UA), an important component of the Chinese medicine formula Yougui Yin, has a wide range of pharmacological properties such as anti-tumor, anti-inflammatory and bone remodeling. Due to the positive effect of Yougui Yin on bone remodeling, the purpose of this study was to investigate the effects of UA on dexamethasone (DEX)-induced SANFH in vitro and vivo. In vitro, we demonstrated that UA can promote mouse BMSCs proliferation and resist DEX-induced apoptosis by CCK8, Western blotting, TUNEL and so on. In addition, vitro experiments such as ALP and Alizarin red staining assay showed that UA had a beneficial effect on the osteogenic differentiation of mouse BMSCs. In vivo, the results of H&E staining, immunohistochemistry staining, Elisa and micro-CT analysis showed that UA had a bone repair-promoting effect in SANFH model. Moreover, the results of Western blot and TUNEL experiments showed that UA could delay the disease progression of SANFH in mice by inhibiting apoptosis. Overall, our study suggests that UA is a potential compound for the treatment of SANFH.

3D-printed porous functional composite scaffolds with polydopamine decoration for bone regeneration.

Large size bone defects affect human health and remain a worldwide health problem that needs to be solved immediately. 3D printing technology has attracted substantial attention for preparing penetrable multifunctional scaffolds to promote bone reconditioning and regeneration. Inspired by the spongy structure of natural bone, novel porous degradable scaffolds have been printed using polymerization of lactide and caprolactone (PLCL) and bioactive glass 45S5 (BG), and polydopamine (PDA) was used to decorate the PLCL/BG scaffolds. The physicochemical properties of the PLCL/BG and PLCL/BG/PDA scaffolds were measured, and their osteogenic and angiogenic effects were characterized through a series of experiments both in vitro and in vivo. The results show that the PLCL/BG2/PDA scaffold possessed a good compression modulus and brilliant hydrophilicity. The proliferation, adhesion and osteogenesis of hBMSCs were improved in the PDA coating groups, which exhibited the best performance. The results of the SD rat cranium defect model indicate that PLCL/BG2/PDA obviously promoted osteointegration, which was further confirmed through immunohistochemical staining. Therefore, PDA decoration and the sustained release of bioactive ions (Ca, Si, P) from BG in the 3D-printed PLCL/BG2/PDA scaffold could improve surface bioactivity and promote better osteogenesis and angiogenesis, which may provide a valuable basis for customized implants in extensive bone defect repair applications.

BML-111 inhibits osteoclast differentiation by suppressing the MAPK and NF-κB pathways, alleviating deterioration of the knee joints in a CIA rat model.

Irreversible destruction of joints is the hallmark of rheumatoid arthritis (RA). Osteoclasts are the only bone-resorbing cells and play an important role in joint rebuilding. BML-111 (5(S),6(R),7-trihydroxyheptanoic acid methyl ester, C8 H16 O5 ) is a synthetic lipoxin A4 agonist with antioxidant and anti-inflammatory properties. The present study aimed to investigate the effect of BML-111 on osteoclasts in vivo and in vitro, to investigate its therapeutic effect on joint destruction in RA. Cell Counting Kit-8 assay and flow cytometry were used to exclude cytotoxic effects of BML-111 to bone marrow-derived macrophages (BMMs). Then, osteoclasts were differentiated in vitro from BMMs by used macrophage colony-stimulating factor and receptor activator of nuclear factor-κB ligand, and osteoclasts were observed following tartrate-resistant acid phosphatase staining with or without BML-111 treatment. Meanwhile, absorption pit assay and immunofluorescence staining of the fibrous actin ring were used to observe osteoclast function. Moreover, we examined mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB) activation. We established collagen-induced arthritis in a rat model and, after treatment with BML-111, joint swelling was measured and the knee joints were processed for histology. We also examined serum and tissue for osteoclastogenesis-related markers. BML-111 inhibited osteoclast formation and differentiation in a time- and concentration-dependent manner, and downregulated the expression levels of MAPK and NF-κB in vitro. Meanwhile, BML-111 effectively alleviated joint structural damage and inhibited osteoclast formation in vivo. BML-111 inhibited osteoclast formation and differentiation in vitro and in vivo, and delayed the progression of joint destruction.

Freeze-Casted Keratin Matrix as an Organic Binder to Integrate Hydroxyapatite and BMP2 for Enhanced Cranial Bone Regeneration.

An ideal bone regenerative scaffold is expected to possess architectural characteristics that mimic the bone tissue, osteoconductive properties, and osteoinductive functionality. Key challenges to creating a scaffold with these ideal characteristics simultaneously are the selection of appropriate processing methods and biocompatible materials. Herein, human hair keratin is proposed as an organic binder for the simultaneous incorporation of bone's major inorganic component, hydroxyapatite and bone's growth factor, recombinant human bone morphogenetic protein 2 (rhBMP2) to enable both osteoconductive and osteoinductive characteristics in the creation of bone scaffolds. Furthermore, a freeze-casting method is selected to fabricate this rhBMP2-incorporated keratin/hydroxyapatite (KHA) scaffold with aligned lamellar pores to guide and promote bone regeneration. The aligned KHA scaffolds display better mechanical properties, sustained rhBMP2 release, good cell compliance, and 3D cellular infiltration. Implantation of KHA scaffolds in vivo reveals that scaffolds with aligned pores effectively accelerate the healing process of bone defects compared to scaffolds with random pores. This work indicates the distinctive potential of freeze-casted rhBMP-2 incorporated KHA scaffolds for bone regeneration.

Corrigendum: Whole-exome sequencing identifies a novel CPT2 mutation in a pedigree with gout.

[This corrects the article DOI: 10.3389/fcell.2022.802635.].

Whole-Exome Sequencing Identifies a Novel CPT2 Mutation in a Pedigree With Gout.

Background: Gout is a common inflammatory arthritis, and its exact pathogenesis remains unclear. Multiple studies have demonstrated that genetic factors play important roles in the development of gout. This study aims to investigate the genetic basis of gout in a three-generation pedigree of affected individuals. Methods: Whole-exome sequencing (WES), comprehensive variant analyses, and co-segregation testing were performed. The effects of candidate variants on protein localization and cellular expression were analyzed, as were interactions with gout-related genes. Results: After comprehensive bioinformatic analysis, Sanger sequencing validation, and pedigree co-segregation analysis, we identified a rare heterozygous missense variant (c.1891C > T, p.R631C) in CPT2. Although no associated changes in localization were observed, the fluorescence intensity of p.R631C mutants was obviously reduced in comparison to the wild-type protein, suggesting that protein degradation is induced by the mutant. Furthermore, our results also indicate that the c.1891C > T variant influences the ability of CPT2 to bind UCP2. Conclusion: This study identified a rare CPT2 mutation in a large Chinese pedigree with gout. Functional studies were used to define the effect of this mutant. This study provides novel insight into the genetic etiology of gout.

Light manipulation for fabrication of hydrogels and their biological applications.

The development of biocompatible materials with desired functions is essential for tissue engineering and biomedical applications. Hydrogels prepared from these materials represent an important class of soft matter for mimicking extracellular environments. In particular, dynamic hydrogels with responsiveness to environments are quite appealing because they can match the dynamics of biological processes. Among the external stimuli that can trigger responsive hydrogels, light is considered as a clean stimulus with high spatiotemporal resolution, complete bioorthogonality, and fine tunability regarding its wavelength and intensity. Therefore, photoresponsiveness has been broadly encoded in hydrogels for biological applications. Moreover, light can be used to initiate gelation during the fabrication of biocompatible hydrogels. Here, we present a critical review of light manipulation tools for the fabrication of hydrogels and for the regulation of physicochemical properties and functions of photoresponsive hydrogels. The materials, photo-initiated chemical reactions, and new prospects for light-induced gelation are introduced in the former part, while mechanisms to render hydrogels photoresponsive and their biological applications are discussed in the latter part. Subsequently, the challenges and potential research directions in this area are discussed, followed by a brief conclusion. STATEMENT OF SIGNIFICANCE: Hydrogels play a vital role in the field of biomaterials owing to their water retention ability and biocompatibility. However, static hydrogels cannot meet the dynamic requirements of the biomedical field. As a stimulus with high spatiotemporal resolution, light is an ideal tool for both the fabrication and operation of hydrogels. In this review, light-induced hydrogelation and photoresponsive hydrogels are discussed in detail, and new prospects and emerging biological applications are described. To inspire more research studies in this promising area, the challenges and possible solutions are also presented.

The potential utility of hybrid photo-crosslinked hydrogels with non-immunogenic component for cartilage repair.

Finding a suitable biomaterial for scaffolding in cartilage tissue engineering has proved to be far from trivial. Nonetheless, it is clear that biomimetic approaches based on gelatin (Gel) and hyaluronic acid (HA) have particular promise. Herein, a set of formulations consisting of photo-polymerizable Gel; photo-polymerizable HA, and allogenic decellularized cartilage matrix (DCM), is synthesized and characterized. The novelty of this study lies particularly in the choice of DCM, which was harvested from an abnormal porcine with α-1,3-galactose gene knockout. The hybrid hydrogels were prepared and studied extensively, by spectroscopic methods, for their capacity to imbibe water, for their behavior under compression, and to characterize microstructure. Subsequently, the effects of the hydrogels on contacting cells (in vitro) were studied, i.e., cytotoxicity, morphology, and differentiation through monitoring the specific markers ACAN, Sox9, Coll2, and Col2α1, hypertrophy through monitoring the specific markers alkaline phosphatase (ALP) and Col 10A1. In vivo performance of the hydrogels was assessed in a rat knee cartilage defect model. The new data expand our understanding of hydrogels built of Gel and HA, since they reveal that a significant augmenting role can be played by DCM. The data strongly suggest that further experimentation in larger cartilage-defect animal models is worthwhile and has potential utility for tissue engineering and regenerative medicine.

Hydrogel composite scaffolds with an attenuated immunogenicity component for bone tissue engineering applications.

Xenogeneic bones are potential templates for bone regeneration. In this study, decellularized porcine bone powder with attenuated immunogenicity was incorporated into a photocurable hydrogel, gelatin methacryloyl (GelMA), to obtain scaffolds with good mechanical properties for bone tissue engineering. The decellularized bone powder (DCB)-GelMA hybrid scaffolds had higher compressive strength and stiffness values when the DCB content was increased. In vitro evaluations revealed the biocompatibility of these scaffolds. The scaffolds could induce human bone marrow mesenchymal stem cells (hMSCs) to undergo osteogenic differentiation even in the absence of an induction medium. The efficiency of the scaffolds for bone regeneration applications was further evaluated using an in vivo cranial bone defect model in rats. Micro-CT images showed that the hybrid scaffolds with 20% DCB content had the best effect in promoting new bone regeneration. Thus, it was concluded that the DCB-GelMA hybrid scaffolds have high potential in bone tissue engineering applications.

Multifunctional Composite Inverse Opal Film with Multiactives for Wound Healing.

A film with an elaborate microstructure and multifunctions is urgently needed in wound healing. Here, we present a multiactive encapsulated inverse opal film with a monitorable delivery system for chronic wound healing. The inverse opal film is prepared by using poly(lactic-co-glycolic acid) to negatively replicate a colloidal crystal template, which presents a high specific surface area and interconnected nanopores. It could be imparted with a potent antibacterial effect and promote angiogenesis by loading the vascular endothelial growth factor into the nanopores and encapsulating by chitosan. In addition, it is demonstrated that the structure color change of the film could intuitively reflect the drug release progress from the nanopores, which made the film a real-time drug monitoring system. In the affected wound model, the properties of the multifunctional film in promoting wound healing are certified by the faster healing speed, more granulation tissue, less inflammation, and even a distribution of new blood vessels and collagen. These results indicate that the resultant multifunctional film has a practical application value in clinical wound care.

Smart materials for point-of-care testing: From sample extraction to analyte sensing and readout signal generator.

The last decade has seen a surge of technical developments in the field on point-of-care testing (POCT). While these developments are extremely diverse, the common aim is to implement improved methods for quick, reliable and inexpensive diagnosis of patients within the clinical setting. While examples of successful introduction and use of POCT techniques are growing, further developments are still necessary to create POCT devices with better portability, usability and performance. Advances in smart materials emerge as potentially valuable know-hows to provide a competitive edge to the development of next generation POCT devices. This review describes the key advantages of adopting smart material-based technologies at different analytical stages of a POCT platform. Under these analytical stages which involves sample pre-treatment, analyte sensing and readout signal generator, several concepts and approaches from contemporary research work in using smart material-based technologies will be the major focus in this review. Lastly, challenges and potential outlook in implementing materials technologies from the application point of view for POCT will be discussed.

Stabilization of HIF-1α alleviates osteoarthritis via enhancing mitophagy.

Mitochondrial dysfunction leads to osteoarthritis (OA) and disc degeneration. Hypoxia inducible factor-1α (HIF-1α) mediated mitophagy has a protective role in several diseases. However, the underlying mechanism of HIF-1α mediated mitophagy in OA remains largely unknown. This current study was performed to determine the effect of HIF-1α mediated mitophagy on OA. Therefore, X-ray and tissue staining including HE staining, safranin O-fast green (S-O) and Alcian Blue were used to assess imageology and histomorphology differences of mouse knee joint. Transcriptional analysis was used to find the possible targets in osteoarthritis. Western blot analysis, RT-qPCR and immunofluorescence staining were used to detect the changes in gene and protein levels in the vitro experiment. The expression of HIF-1α was increased in human and mouse OA cartilage. HIF-1α knockdown by siRNA further impair the hypoxia-induced mitochondrial dysfunction; In contrast, HIF-1α mediated protective role was reinforced by prolylhydroxylase (PHD) inhibitor dimethyloxalylglycine (DMOG). In addition, HIF-1α stabilization could alleviate apoptosis and senescence via mitophagy in chondrocytes under hypoxia condition, which could also ameliorate surgery-induced cartilage degradation in mice OA model. In conclusion, HIF-1α mediated mitophagy could alleviate OA, which may serve as a promising strategy for OA treatment.

Unraveling the genetic causes in large pedigrees with gout by whole­exome sequencing.

Gout is a common type of inflammatory arthritis that is clinically and genetically heterogeneous. The genetic aetiology remains unclear, and mainly relies on previous genome­wide association studies focused on sporadic cases. The present study aimed to identify the genetic basis of gout in three families using whole­exome sequencing (WES). WES was performed in the probands, and family members were involved in the co­segregation analysis. In total, three deleterious rare or novel missense mutations were identified in ATP­binding cassette super­family G member 2 (ABCG2), protein kinase CGMP­dependent 2 (PRKG2) and adrenoceptor ß3 (ADRB3) genes in three different families. In addition, certain gout­associated candidate genes were revealed to be shared among the co­expression and protein­protein interaction (PPI) networks of ABCG2, PRKG2 and ADRB3. Furthermore, the disease ontology analysis of the genes present in the co­expression network exhibited significant (P<0.05) enrichment in hyperuricemia, gout, cardiovascular system disease and metabolic disease. In addition, genes involved in the PPI network were significantly enriched in the purine nucleoside monophosphate biosynthetic process, urate transport and biological processes associated with glycose metabolism. Collectively, to the best of our knowledge, the present study was the first to use WES to identify three candidate rare or novel deleterious mutations in three families with gout. The present results provided novel insights that may improve the current understanding of the molecular genetic basis underlying gout. Importantly, the present results may facilitate the improvement of clinical diagnosis and the development of novel personalized therapies.

Whole-Exome Sequencing Reveals a Rare Missense Variant in SLC16A9 in a Pedigree with Early-Onset Gout.

Gout is a common inflammatory arthritis triggered by monosodium urate deposition after longstanding hyperuricemia. In the general community, the disease is largely polygenic in genetic architecture, with many polymorphisms having been identified in gout or urate-associated traits. In a small proportion of cases, rare high penetrant mutations associated with monogenic segregation of the disease in families have been demonstrated to be disease causative. In this study, we recruited a two-generation pedigree with early-onset gout. To elucidate the genetic predisposition, whole-exome sequencing (WES) was performed. After comprehensive variant analyses and cosegregation testing, we identified a missense variant (c.277C>A, p.L93M) in SLC16A9, an extremely rare variant in genetic databases. Moreover, in silico assessments showed strong pathogenicity. This variant cosegregated with the disease phenotype perfectly in the family and is located in a highly conserved functional domain. A few studies supported our results of the association between SLC16A9 and gout and serum urate levels. In conclusion, we provide the first evidence for the association of rare missense in SLC16A9 with early-onset gout. These findings not only expand our current understanding of gout but also may have further implications for the treatment and prevention of gout.

Angiogenesis and bone regeneration by mesenchymal stem cell transplantation with danshen in a rabbit model of avascular necrotic femoral head.

The present study aimed to explore the potential of combined treatment with mesenchymal stem cells (MSCs) and danshen for angiogenesis and bone regeneration in a rabbit model of avascular necrosis of femoral head (ANFH). A rabbit model of ANFH was established using the Shwartzman reaction with methylprednisolone and Escherichia coli endotoxin injection. Magnetic resonance imaging (MRI) and histopathological examination were used to evaluate the rabbit model of ANFH. The rabbits were randomly divided into the danshen group, the MSCs group, the danshen combined with MSCs group and the model group (treated with physiological saline). The expression level of monocyte chemoattractant protein-1 (MCP-1) and stromal cell-derived factor-1 (SDF-1) were determined by reverse transcription polymerase chain reaction (RT-PCR). The expression level of bone morphogenetic protein-2 (BMP-2) and vascular endothelial growth factor (VEGF) were detected by immunofluorescence and the mRNA expression of BMP-2 and VEGF were detected by RT-PCR. Typical osteonecrosis occurred in the rabbit model of ANFH, which indicated that the model was successfully established. MCP-1 and SDF-1 were significantly increased in the model group compared with the normal group (P<0.05). Following the administration of MSCs and Salvia miltiorrhiza (danshen), MSCs labeled with 5-bromo-2-deoxyuridine were observed to be gathered in the necrotic area. The increased migration of MSCs to the necrotic area may be due to the upregulated expression of the chemokines MCP-1 and SDF-1. ANFH treated with danshen combined with MSCs may promote revascularization by increasing the expression of VEGF and BMP-2 in the femoral head, promoting re-ossification and revascularization. Danshen combined with the transplantation of MSCs may be regarded as a novel therapy for the treatment of ANFH in a clinical setting.

Adenosine A2A Receptor Mediates Inhibition of Synovitis and Osteoclastogenesis after Electroacupuncture in Rats with Collagen-Induced Arthritis.

BACKGROUND: This study was to investigate the role of adenosine A2A receptors (A2AR) in inhibiting the effect of electroacupuncture (EA) on osteoclastogenesis in collagen-induced arthritis (CIA). METHODS: Wistar rats were divided into four groups: sham-control group, CIA-control group, CIA-EA group, and CIA-EA-SCH58261 (A2AR antagonist) group. We detected tumor necrosis factor-α (TNF-α), nuclear transcription factor-κB (NF-κB), receptor activator of NF-κB ligand (RANKL), protein kinase A (PKA), and extracellular regulatory protein kinase 1/2 (ERK1/2) in peripheral blood by ELISA. PKA, ERK1/2, and NF-κB in ankle joints were determined by western blotting. We evaluated the arthritis damage by histological examination and determined the number of osteoclasts by tartrate-resistant acid phosphatase (TRAP) staining. RESULTS: EA treatment downregulated the expression of TNF-α, RANKL, PKA, ERK1/2, and NF-κB in peripheral blood but increased the levels of PKA and ERK1/2 in ankle joints. Importantly, EA treatment reduced bone erosion as evidenced by the histological findings and inhibited osteoclastogenesis as revealed by TRAP staining. All these effects of the EA treatment were reversed by combining EA treatment with the A2AR antagonist SCH58261. CONCLUSION: Our data suggest that EA treatment activated A2AR. The effects of the A2AR antagonist SCH58261 suggest that the inhibition of osteoclast formation, the inhibition of TNF-α, RANKL, and NF-κB expression, and the increase of ERK1/2 are all dependent on this EA-induced A2AR activation. It is therefore likely that these pathways with clearly defined roles in inflammation and bone erosion are at least partially involved in the mediation of the inhibition of synovitis and osteoclast formation induced by EA.

Luteolin inhibits angiogenesis of the M2­like TAMs via the downregulation of hypoxia inducible factor­1α and the STAT3 signalling pathway under hypoxia.

The imbalance between angiogenic inducers and inhibitors appears to be a critical factor in tumour pathogenesis. Angiogenesis serves a key role in the occurrence, invasion and metastasis of tumours. Macrophages are a major cellular component of human and rodent tumours, where they are usually termed tumour­associated macrophages (TAMs). In malignant tumours, TAMs tend to resemble alternatively activated macrophages (M2­like), promote TA angiogenesis, strengthen tumour migration and invasive abilities, and simultaneously inhibit antitumor immune responses. In our previous study, luteolin, commonly found in a wide variety of plants, had a strong antitumor effect under normoxia; however, it is unknown whether luteolin serves a similar role under hypoxia. In the present study, cobalt chloride (CoCl2) was used to simulate hypoxia. Hypoxia­inducible factor­1α (HIF­1α), which is difficult to detect under normoxic conditions, was significantly increased. Additionally, vascular endothelial growth factor (VEGF) was also significantly increased in response to CoCl2 treatment. Subsequently, luteolin was applied with CoCl2 to examine the effects of luteolin. Luteolin decreased the expression of VEGF and matrix metalloproteinase­9, which promote angiogenesis. In addition, luteolin also suppressed the activation of HIF­1 and phosphorylated­signal transducer and activator of transcription 3 (STAT3) signalling, particularly within the M2­like TAMs. The results of the present study provide novel evidence that luteolin, under hypoxic conditions, has a strong anticancer effect via the HIF­1α and STAT3 signalling pathways.

AMP-activated protein kinase-dependent induction of autophagy by erythropoietin protects against spinal cord injury in rats.

AIMS: Autophagy has been regarded as a promising therapeutic target for spinal cord injury (SCI). Erythropoietin (EPO) has been demonstrated to exhibit neuroprotective effects in the central nervous system (CNS); however, the molecular mechanisms of its protection against SCI remain unknown. This study aims to investigate whether the neuroprotective effects of EPO on SCI are mediated by autophagy via AMP-activated protein kinase (AMPK) signaling pathways. METHODS: Functional assessment and Nissl staining were used to investigate the effects of EPO on SCI. Expressions of proteins were detected by Western blot and immunohistochemistry. RESULTS: Treatment with EPO significantly reduced the loss of motor neurons and improved the functional recovery following SCI. Erythropoietin significantly enhanced the SCI-induced autophagy through activating AMPK and inactivating mTOR signaling. The inhibitor of AMPK, compound C, could block the EPO-induced autophagy and beneficial action on SCI, whereas the activator of AMPK, metformin, could mimic the effects of EPO. In the in vitro studies, EPO enhanced the hypoxia-induced autophagy in an AMPK-dependent manner. CONCLUSIONS: The AMPK-dependent induction of autophagy contributes to the neuroprotection of EPO on SCI.