Zwitterionic Conductive Hydrogel-Based Nerve Guidance Conduit Promotes Peripheral Nerve Regeneration in Rats.

In recent years, conductive biomaterials have been widely used to enhance peripheral nerve regeneration. However, most biomaterials use electronic conductors to increase the conductivity of materials. As information carriers, electronic conductors always transmit discontinuous electrical signals, while biological systems essentially transmit continuous signals through ions. Herein, an ion-based conductive hydrogel was fabricated by simple copolymerization of the zwitterionic monomer sulfobetin methacrylate and hydroxyethyl methacrylate. Benefiting from the excellent mechanical stability, suitable electrical conductivity, and good cytocompatibility of the zwitterionic hydrogel, the Schwann cells cultured on the hydrogel could grow and proliferate better, and dorsal root ganglian had an increased neurite length. The zwitterionic hydrogel-based nerve guidance conduits were then implanted into a 10 mm sciatic nerve defect model in rats. Morphological analysis and electrophysiological data showed that the grafts achieved a regeneration effect close to that of the autologous nerve. Overall, our developed zwitterionic hydrogel facilitates efficient and efficacious peripheral nerve regeneration by mimicking the electrical and mechanical properties of the extracellular matrix and creating a suitable regeneration microenvironment, providing a new material reserve for the repair of peripheral nerve injury.

Motor neuron-specific RhoA knockout delays degeneration and promotes regeneration of dendrites in spinal ventral horn after brachial plexus injury.

Dendrites play irreplaceable roles in the nerve conduction pathway and are vulnerable to various insults. Peripheral axotomy of motor neurons results in the retraction of dendritic arbors, and the dendritic arbor can be re-expanded when reinnervation is allowed. RhoA is a target that regulates the cytoskeleton and promotes neuronal survival and axon regeneration. However, the role of RhoA in dendrite degeneration and regeneration is unknown. In this study, we explored the potential role of RhoA in dendrites. A line of motor neuronal RhoA conditional knockout mice was developed by crossbreeding HB9Cre+ mice with RhoAflox/flox mice. We established two models for assaying dendrite degeneration and regeneration, in which the brachial plexus was transection or crush injured, respectively. We found that at 28 days after brachial plexus transection, the density, complexity, and structural integrity of dendrites in the ventral horn of the spinal cord of RhoA conditional knockout mice were slightly decreased compared with that in Cre mice. Dendrites underwent degeneration at 7 and 14 days after brachial plexus transection and recovered at 28-56 days. The density, complexity, and structural integrity of dendrites in the ventral horn of the spinal cord of RhoA conditional knockout mice recovered compared with results in Cre mice. These findings suggest that RhoA knockout in motor neurons attenuates dendrite degeneration and promotes dendrite regeneration after peripheral nerve injury.

Atractylenolide-III alleviates osteoarthritis and chondrocyte senescence by targeting NF-κB signaling.

Atractylenolide-III (AT-III) is well known as its role in antioxidant and anti-inflammatory. Present study was aimed to figure out its effects on osteoarthritis and potential mechanisms. Rat model, human osteoarthritis cartilage explants as well as rat/human chondrocyte cultures were prepared to test AT-III's effects on osteoarthritis progression and chondrocyte senescence. Potential targeted molecules of AT-III were predicted using network pharmacology and molecular docking, assessed by Western blotting and then verified with rescue experiments. AT-III treatment alleviated osteoarthritis severity (shown by OARSI grading score and micro-CT) and chondrocyte senescence (indexed by levels of SA-ß-gal, P16, P53, MMP13, ROS and ratio of healthy/collapsed mitochondrial membrane potentials). Network pharmacology and molecular docking suggested that AT-III might play role through NF-κB pathway. Further experiments revealed that AT-III reduced phosphorylation of IKKα/ß, IκBα and P65 in NF-κB pathway. As well as nuclear translocation of p65. Both in vivo and in vitro experiments indicated that AT-III's effects on osteoarthritis and anti-senescence were reversed by an NF-κB agonist. AT-III could alleviate osteoarthritis by inhibiting chondrocyte senescence through NF-κB pathway, which indicated that AT-III is a prospective drug for osteoarthritis treatment.

Schwann cell-specific RhoA knockout accelerates peripheral nerve regeneration via promoting Schwann cell dedifferentiation.

Our previous studies indicated that RhoA knockdown or inhibition could alleviate the proliferation, migration, and differentiation of Schwann cells. However, the role of RhoA in Schwann cells during nerve injury and repair is still unknown. Herein, we developed two lines of Schwann cells conditional RhoA knockout (cKO) mice by breeding RhoAflox / flox mice with PlpCre -ERT2 or DhhCre mice. Our results indicate that RhoA cKO in Schwann cells accelerates axonal regrowth and remyelination after sciatic nerve injury, which enhances the recovery of nerve conduction and hindlimb gait, and alleviates the amyotrophy in gastrocnemius muscle. Mechanistic studies in both in vivo and in vitro models revealed that RhoA cKO could facilitate Schwann cell dedifferentiation via JNK pathway. Schwann cell dedifferentiation subsequently promotes Wallerian degeneration by enhancing phagocytosis and myelinophagy, as well as stimulating the production of neurotrophins (NT-3, NGF, BDNF, and GDNF). These findings shed light on the role of RhoA in Schwann cells during nerve injury and repair, indicating that cell type-specific RhoA targeting could serve as a promising molecular therapeutic strategy for peripheral nerve injury.

Perlecan Improves Blood Spinal Cord Barrier Repair Through the Integrin ß1/ROCK/MLC Pathway After Spinal Cord Injury.

Spinal cord injury (SCI) can lead to the destruction of the blood-spinal cord barrier (BSCB), causing various inflammatory cytokines, neutrophils, and macrophages to infiltrate the lesion area, resulting in secondary injury. Basement membranes (BMs) are maintained by all types of cells in the BSCB and contribute to BSCB maintenance. Perlecan is an important constituent of vascular BMs, maintaining vascular integrity and neuroprotection. However, it is not clear whether Perlecan is involved in BSCB repair after SCI. In this study, we found that Perlecan was specifically expressed in the BMs in the spinal cord and underwent degradation/remodeling after SCI. Subsequently, a CRISPR/Cas9-based SAM system was used to overexpress Perlecan in the injured spinal cord, resulting in significantly enhanced locomotor recovery and neural regeneration. Overexpression of Perlecan reduced BSCB permeability along with the neuroinflammatory response. Interestingly, Perlecan inhibited stress fiber formation by interacting with integrin ß1 and inhibiting downstream ROCK/MLC signaling, resulting in reduced tight junctions (TJs) disassembly and improved BSCB integrity. Furthermore, the integrin receptor antagonist GRGDSP abolished the effects of Perlecan overexpression on BSCB permeability and TJs integrity. Overall, our findings suggest that Perlecan reduces BSCB permeability and the neuroinflammatory response by interacting with integrin ß1 and inhibiting the downstream ROCK/MLC pathway to promote neurological recovery after SCI.

miR-301a Deficiency Attenuates the Macrophage Migration and Phagocytosis through YY1/CXCR4 Pathway.

(1) Background: the miR-301a is well known involving the proliferation and migration of tumor cells. However, the role of miR-301a in the migration and phagocytosis of macrophages is still unclear. (2) Methods: sciatic nerve injury, liver injury models, as well as primary macrophage cultures were prepared from the miR-301a knockout (KO) and wild type (WT) mice to assess the macrophage's migration and phagocytosis capabilities. Targetscan database analysis, Western blotting, siRNA transfection, and CXCR4 inhibition or activation were performed to reveal miR301a's potential mechanism. (3) Results: the macrophage's migration and phagocytosis were significantly attenuated by the miR-301a KO both in vivo and in vitro. MiR-301a can target Yin-Yang 1 (YY1), and miR-301a KO resulted in YY1 up-regulation and CXCR4 (YY1's down-stream molecule) down-regulation. siYY1 increased the expression of CXCR4 and enhanced migration and phagocytosis in KO macrophages. Meanwhile, a CXCR4 inhibitor or agonist could attenuate or accelerate, respectively, the macrophage migration and phagocytosis. (4) Conclusions: current findings indicated that miR-301a plays important roles in a macrophage's capabilities of migration and phagocytosis through the YY1/CXCR4 pathway. Hence, miR-301a might be a promising therapeutic candidate for inflammatory diseases by adjusting macrophage bio-functions.

Application of collagen-chondroitin sulfate scaffolds with different pore sizes combined with acidic fibroblast growth factor in repairing full thickness skin defects in nude mice.

Wound healing of skin defects is complex. For the treatment of large and deep wounds, it is a good alternative to accept artificial dermis grafting at the first stage surgery, and autologous split-thickness skin grafting 2-3 weeks later at the second stage surgery. In addition, the effectiveness of numerous cytokines such as fibroblast growth factor (FGF) on wounds healing has been widely researched. The traditional view is that direct external application orin vivoinjection of exogenous FGFs may not achieve the desired therapeutic effect as the effective concentration cannot be maintained for a long time. Therefore, some researchers have tried to integrate various cytokines into skin substitutes for combined application. However, we believe that considering the current situation, it is still difficult to achieve mass production of these types of artificial dermis. Here, we manufactured a collagen-chondroitin sulfate scaffold material by imitating the marketed artificial dermis materials. Then, we combined it with recombinant human acidic FGF in a single full dose during the first-stage artificial dermis transplantation, which is simple and completely feasible but always controversial in the current clinical work, to explore whether this combinatorial therapy could serve as an efficient way wound healing in the Balb/c-nu mice full-thickness skin defect model.

Nsun4 and Mettl3 mediated translational reprogramming of Sox9 promotes BMSC chondrogenic differentiation.

The chondrogenic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs) has been used in the treatment and repair of cartilage defects; however, the in-depth regulatory mechanisms by which RNA modifications are involved in this process are still poorly understood. Here, we found that Sox9, a critical transcription factor that mediates chondrogenic differentiation, exhibited enhanced translation by ribosome sequencing in chondrogenic pellets, which was accompanied by increased 5-methylcytosine (m5C) and N6-methyladenosine (m6A) levels. Nsun4-mediated m5C and Mettl3-mediated m6A modifications were required for Sox9-regulated chondrogenic differentiation. Interestingly, we showed that in the 3'UTR of Sox9 mRNA, Nsun4 catalyzed the m5C modification and Mettl3 catalyzed the m6A modification. Furthermore, we found that Nsun4 and Mettl3 co-regulated the translational reprogramming of Sox9 via the formation of a complex. Surface plasmon resonance (SPR) assays showed that this complex was assembled along with the recruitment of Ythdf2 and eEF1α-1. Moreover, BMSCs overexpressing Mettl3 and Nsun4 can promote the repair of cartilage defects in vivo. Taken together, our study demonstrates that m5C and m6A co-regulate the translation of Sox9 during the chondrogenic differentiation of BMSCs, which provides a therapeutic target for clinical implications.

3D Printed Platelet-Rich Plasma-Loaded Scaffold with Sustained Cytokine Release for Bone Defect Repair.

The combination of three-dimensional (3D) printed scaffold materials and various cytokines can achieve the purpose of tissue reconstruction more efficiently. In this study, we prepared platelet-rich plasma (PRP)/gelatin microspheres combined with 3D printed polycaprolactone/ß-tricalcium phosphate scaffolds to solve the key problem that PRP cannot be released under control and the release time is too short, and thus better promote bone repair. Consequently, the composite scaffold displayed a good mechanical property and sustained cytokine release for ∼3 weeks. Increased survival, proliferation, migration, and osteogenic and angiogenic differentiation of bone marrow mesenchymal stem cells were observed compared with the control groups. The in vivo study demonstrated that the composite scaffold with PRP/gelatin microspheres led to greater positive effects in promoting large bone defect repair. In conclusion, in this study, a new type of PRP long-term sustained-release composite scaffold material was constructed that effectively improved the survival, proliferation, and differentiation of cells in the transplanted area, thereby better promoting the repair of large bone defects. Impact statement Reconstruction of bone tissue and blood vessels at bone defects takes time. Platelet-rich plasma (PRP) has been widely used in bone defect repair because it contains a variety of cytokine that can promote local osteogenesis and angiogenesis. In this study, we constructed a new type of polycaprolactone/ß-tricalcium phosphate/PRP/gelatin scaffold to solve the predicament of short cytokine release time in PRP-related materials. We proved that this scaffold can not only achieve long-term PRP-related cytokine release (more than 3 weeks) but also promote osteogenesis and bone defect repair. We believe that this is a novel concept of developing the sustained PRP-related cytokine releasing bioscaffold for treating large bone defect.

Role of microtubule dynamics in Wallerian degeneration and nerve regeneration after peripheral nerve injury.

Wallerian degeneration, the progressive disintegration of distal axons and myelin that occurs after peripheral nerve injury, is essential for creating a permissive microenvironment for nerve regeneration, and involves cytoskeletal reconstruction. However, it is unclear whether microtubule dynamics play a role in this process. To address this, we treated cultured sciatic nerve explants, an in vitro model of Wallerian degeneration, with the microtubule-targeting agents paclitaxel and nocodazole. We found that paclitaxel-induced microtubule stabilization promoted axon and myelin degeneration and Schwann cell dedifferentiation, whereas nocodazole-induced microtubule destabilization inhibited these processes. Evaluation of an in vivo model of peripheral nerve injury showed that treatment with paclitaxel or nocodazole accelerated or attenuated axonal regeneration, as well as functional recovery of nerve conduction and target muscle and motor behavior, respectively. These results suggest that microtubule dynamics participate in peripheral nerve regeneration after injury by affecting Wallerian degeneration. This study was approved by the Animal Care and Use Committee of Southern Medical University, China (approval No. SMU-L2015081) on October 15, 2015.

Stem Cell-Seeded 3D-Printed Scaffolds Combined with Self-Assembling Peptides for Bone Defect Repair.

Bone defects caused by infection, tumor, trauma, and so on remain difficult to treat clinically. Bone tissue engineering (BTE) has great application prospect in promoting bone defect repair. Polycaprolactone (PCL) is a commonly used material for creating BTE scaffolds. In addition, self-assembling peptides (SAPs) can function as the extracellular matrix and promote osteogenesis and angiogenesis. In the work, a PCL scaffold was constructed by 3D printing, then integrated with bone marrow mesenchymal stem cells (BMSCs) and SAPs. The research aimed to assess the bone repair ability of PCL/BMSC/SAP implants. BMSC proliferation in PCL/SAP scaffolds was assessed via Cell Counting Kit-8. In vitro osteogenesis of BMSCs cultured in PCL/SAP scaffolds was assessed by alkaline phosphatase staining and activity assays. Enzyme-linked immunosorbent assays were also performed to detect the levels of osteogenic factors. The effects of BMSC-conditioned medium from 3D culture systems on the migration and angiogenesis of human umbilical vein endothelial cells (HUVECs) were assessed by scratch, transwell, and tube formation assays. After 8 weeks of in vivo transplantation, radiography and histology were used to evaluate bone regeneration, and immunohistochemistry staining was utilized to detect neovascularization. In vitro results demonstrated that PCL/SAP scaffolds promoted BMSC proliferation and osteogenesis compared to PCL scaffolds, and the PCL/BMSC/SAP conditional medium (CM) enhanced HUVEC migration and angiogenesis compared to the PCL/BMSC CM. In vivo results showed that, compared to the blank control, PCL, and PCL/BMSC groups, the PCL/BMSC/SAP group had significantly increased bone and blood vessel formation. Thus, the combination of BMSC-seeded 3D-printed PCL and SAPs can be an effective approach for treating bone defects. Impact statement Both polycaprolactone (PCL) and self-assembling peptides (SAPs) have been broadly applied in bone defect repair. However, the poor osteoinductivity of PCL and weak mechanical strength of SAPs have limited their clinical application. Here, a 3D-printed PCL scaffold was fabricated for seeding bone marrow mesenchymal stem cells (BMSCs), then combined with SAPs to construct a composite PCL/BMSC/SAP implant for treating the calvarial defect. We showed that transplantation of PCL/BMSC/SAP composite implants clearly promoted bone regeneration and neovascularization. To our knowledge, this is the first study to treat bone defects by combination of BMSC-seeded 3D-printed PCL and SAPs.

SIRT6 Negatively Regulates Schwann Cells Dedifferentiation via Targeting c-Jun During Wallerian Degeneration After Peripheral Nerve Injury.

Silent information regulator 6 (SIRT6) is a mammalian homolog of the nicotinamide adenine dinucleotide (NAD)-dependent deacetylase sirtuin family. Previous studies have been reported a pro-regenerative role of SIRT6 in central nervous system injury. However, the role of SIRT6 in peripheral nerve injury is still unknown. Given the importance and necessity of Schwann cell dedifferentiation response to peripheral nerve injury, we aim to investigate the molecular mechanism of SIRT6 steering Schwann cell dedifferentiation during Wallerian degeneration in injured peripheral nerve. Herein, we first examined the expression pattern of SIRT6 after peripheral nerve injury. Using the explants of sciatic nerve, an ex vivo model of nerve degeneration, we provided evidences indicating that SIRT6 inhibitor accelerates Schwann cell dedifferentiation as well as axonal and myelin degeneration, while SIRT6 activator attenuates this process. Moreover, in an in vitro Schwann cell dedifferentiation model, we found SIRT6 inhibitor promotes Schwann cell dedifferentiation through upregulating the expression of c-Jun. In addition, downregulation of c-Jun reverse the effects of SIRT6 inhibition on the Schwann cells dedifferentiation and axonal and myelin degeneration. In summary, we first described SIRT6 acts as a negative regulator for Schwann cells dedifferentiation during Wallerian degeneration and c-Jun worked as a direct downstream partner of SIRT6 in injured peripheral nerve.

SIRT6 inhibition delays peripheral nerve recovery by suppressing migration, phagocytosis and M2-polarization of macrophages.

BACKGROUND: Silent information regulator 6 (SIRT6) is a mammalian homolog of the nicotinamide adenine dinucleotide (NAD)-dependent deacetylase sirtuin family. Prior evidences suggested that the anti-inflammatory function of SIRT6 after spinal cord and brain injury, and it plays a crucial role in macrophages polarization of adipose tissue and skin. However, the role of SIRT6 in macrophages involved peripheral nerve injury is still unknown. Given the prominent role of macrophages in peripheral nerve recovery, we aim to investigate the role of SIRT6 in the regulation of phenotypes shift and functions in macrophages after peripheral nerve injury. RESULTS: In the present study, we first identified a significant increase of SIRT6 expression during nerve degeneration and macrophages phagocytosis. Next, we found nerve recovery was delayed after SIRT6 silencing by injected shRNA lentivirus into the crushed sciatic nerve, which exhibited a reduced expression of myelin-related proteins (e.g., MAG and MBP), severer myoatrophy of target muscles, and inferior nerve conduction compared to the shRNA control injected mice. In vitro, we found that SIRT6 inhibition by being treated with a selective inhibitor OSS_128167 or lentivirus transfection impairs migration and phagocytosis capacity of bone marrow-derived macrophages (BMDM). In addition, SIRT6 expression was discovered to be reduced after M1 polarization, but SIRT6 was enhanced after M2 polarization in the monocyte-macrophage cell line RAW264.7 and BMDM. Moreover, SIRT6 inhibition increased M1 macrophage polarization with a concomitant decrease in M2 polarization both in RAW264.7 and BMDM via activating NF-κB and TNF-α expression, and SIRT6 activation by UBCS039 treatment could shift the macrophages from M1 to M2 phenotype. CONCLUSION: Our findings indicate that SIRT6 inhibition impairs peripheral nerve repair through suppressing the migration, phagocytosis, and M2 polarization of macrophages. Therefore, SIRT6 may become a favorable therapeutic target for peripheral nerve injury.

Macrophage-specific RhoA knockout delays Wallerian degeneration after peripheral nerve injury in mice.

BACKGROUND: Plenty of macrophages are recruited to the injured nerve to play key roles in the immunoreaction and engulf the debris of degenerated axons and myelin during Wallerian degeneration, thus creating a conducive microenvironment for nerve regeneration. Recently, drugs targeting the RhoA pathway have been widely used to promote peripheral axonal regeneration. However, the role of RhoA in macrophage during Wallerian degeneration and nerve regeneration after peripheral nerve injury is still unknown. Herein, we come up with the hypothesis that RhoA might influence Wallerian degeneration and nerve regeneration by affecting the migration and phagocytosis of macrophages after peripheral nerve injury. METHODS: Immunohistochemistry, Western blotting, H&E staining, and electrophysiology were performed to access the Wallerian degeneration and axonal regeneration after sciatic nerve transection and crush injury in the LyzCre+/-; RhoAflox/flox (cKO) mice or Lyz2Cre+/- (Cre) mice, regardless of sex. Macrophages' migration and phagocytosis were detected in the injured nerves and the cultured macrophages. Moreover, the expression and potential roles of ROCK and MLCK were also evaluated in the cultured macrophages. RESULTS: 1. RhoA was specifically knocked out in macrophages of the cKO mice; 2. The segmentation of axons and myelin, the axonal regeneration, and nerve conduction in the injured nerve were significantly impeded while the myoatrophy was more severe in the cKO mice compared with those in Cre mice; 3. RhoA knockout attenuated the migration and phagocytosis of macrophages in vivo and in vitro; 4. ROCK and MLCK were downregulated in the cKO macrophages while inhibition of ROCK and MLCK could weaken the migration and phagocytosis of macrophages. CONCLUSIONS: Our findings suggest that RhoA depletion in macrophages exerts a detrimental effect on Wallerian degeneration and nerve regeneration, which is most likely due to the impaired migration and phagocytosis of macrophages resulted from disrupted RhoA/ROCK/MLCK pathway. Since previous research has proved RhoA inhibition in neurons was favoring for axonal regeneration, the present study reminds us of that the cellular specificity of RhoA-targeted drugs is needed to be considered in the future application for treating peripheral nerve injury.

3D printed polycaprolactone/beta-tricalcium phosphate/magnesium peroxide oxygen releasing scaffold enhances osteogenesis and implanted BMSCs survival in repairing the large bone defect.

Ischemia and hypoxia in the bone defect area remain an intractable problem when treating large bone defects. Thus, oxygen-releasing biomaterials have been widely researched in recent years. Magnesium peroxide (MgO2) can release oxygen (O2), and magnesium ions (Mg2+), simultaneously, which is seen to have significant potential in bone substitutes. In this study, we used 3D printing technology to fabricate a MgO2-contained composite scaffold, which was composed of polycaprolactone (PCL), beta-tricalcium phosphate (ß-TCP) and magnesium peroxide (MgO2). Physical properties and O2/Mg2+ releasing behavior of the scaffold were studied. Then, we evaluated the effects of the scaffold on cell survival, proliferation, migration, adhesion and osteogenic differentiation by the co-culture of bone marrow mesenchymal stem cells (BMSCs) and scaffold under normoxia and hypoxia in vitro. Finally, the osteogenic properties of the scaffold in vivo were evaluated via the rat femoral condylar bone defect model. The PCL/ß-TCP/MgO2 scaffold showed good mechanical properties and sustained O2 and Mg2+ release for about three weeks. Meanwhile, the scaffold showed appreciable promotion on the survival, proliferation, migration and osteogenic differentiation of BMSCs under hypoxia compared with control groups. The results of imaging studies and histological analysis showed that implantation of PCL/ß-TCP/MgO2 scaffold could promote seed cell survival and significantly increased new bone formation. In sum, the PCL/ß-TCP/MgO2 scaffold is promising with great potential for treating large bone defects.

CaO2/gelatin oxygen slow-releasing microspheres facilitate tissue engineering efficiency for the osteonecrosis of femoral head by enhancing the angiogenesis and survival of grafted bone marrow mesenchymal stem cells.

The osteonecrosis of femoral head (ONFH), a common refractory disease, is still not fully understood today. Hypoxia caused by ischemia is not only an important pathogenic factor but also a critical challenge for the survival of seed cells in the tissue engineering therapy of ONFH. To explore an efficient strategy to treat ONFH by targeting hypoxia, newly designed CaO2/gelatin microspheres were composited with 3D printed polycaprolactone/nano-hydroxyapatite (PCL/nHA) porous scaffold, sodium alginate/gelatin hydrogel, and bone marrow mesenchymal stem cells (BMSCs) to develop a novel tissue engineering scaffold and then transplanted into the core depression area of the ONFH rabbit model. The current data demonstrated that CaO2/gelatin microspheres can constantly release oxygen for 19 days. In vitro assays with BMSCs illustrated that scaffolds have high biocompatibility and are favorable for cell proliferation in extreme hypoxia (1% O2). The in vivo study demonstrated that the transplanted scaffold with oxygen-generating microspheres significantly enhanced the osteogenic and angiogenic effects compared to the scaffold without microspheres. Further assessments revealed that microspheres in the scaffold can reduce the local cell apoptosis and enhance the survival of grafted cells in the host. Collectively, the present study developed a novel oxygen slow-releasing composite scaffold, which can facilitate tissue engineering efficiency for treating the osteonecrosis of the femoral head by enhancing the angiogenesis and survival of grafted stem cells.

Cotransplantation of mesenchymal stem cells and endothelial progenitor cells for treating steroid-induced osteonecrosis of the femoral head.

Steroid-induced osteonecrosis of the femoral head (ONFH) is characterized by decreased osteogenesis, angiogenesis, and increased adipogenesis. While bone tissue engineering has been widely investigated to treat ONFH, its therapeutic effects remain unsatisfactory. Therefore, further studies are required to determine optimal osteogenesis, angiogenesis and adipogenesis in the necrotic area of the femoral head. In our study, we developed a carboxymethyl chitosan/alginate/bone marrow mesenchymal stem cell/endothelial progenitor cell (CMC/ALG/BMSC/EPC) composite implant, and evaluated its ability to repair steroid-induced ONFH. Our in vitro studies showed that BMSC and EPC coculture displayed enhanced osteogenic and angiogenic differentiation. When compared with single BMSC cultures, adipogenic differentiation in coculture systems was reduced. We also fabricated a three-dimensional (3D) CMC/ALG scaffold for loading cells, using a lyophilization approach, and confirmed its good cell compatibility characteristics, that is, high porosity, low cytotoxicity and favorable cell adhesion. 3D coculture of BMSCs and EPCs also promoted secretion of osteogenic and angiogenic factors. Then, we established an rabbit model of steroid-induced ONFH. The CMC/ALG/BMSC/EPC composite implant was transplanted into the bone tunnel of the rabbit femoral head after core decompression (CD) surgery. Twelve weeks later, radiographical and histological analyses revealed CMC/ALG/BMSC/EPC composite implants had facilitated the repair of steroid-induced ONFH, by promoting osteogenesis and angiogenesis, and reducing adipogenesis when compared with CD, CMC/ALG, CMC/ALG/BMSC and CMC/ALG/EPC groups. Thus, our data show that cotransplantation of BMSCs and EPCs in 3D scaffolds is beneficial in treating steroid-induced ONFH.

Ascorbic acid accelerates Wallerian degeneration after peripheral nerve injury.

Wallerian degeneration occurs after peripheral nerve injury and provides a beneficial microenvironment for nerve regeneration. Our previous study demonstrated that ascorbic acid promotes peripheral nerve regeneration, possibly through promoting Schwann cell proliferation and phagocytosis and enhancing macrophage proliferation, migration, and phagocytosis. Because Schwann cells and macrophages are the main cells involved in Wallerian degeneration, we speculated that ascorbic acid may accelerate this degenerative process. To test this hypothesis, 400 mg/kg ascorbic acid was administered intragastrically immediately after sciatic nerve transection, and 200 mg/kg ascorbic acid was then administered intragastrically every day. In addition, rat sciatic nerve explants were treated with 200 µM ascorbic acid. Ascorbic acid significantly accelerated the degradation of myelin basic protein-positive myelin and neurofilament 200-positive axons in both the transected nerves and nerve explants. Furthermore, ascorbic acid inhibited myelin-associated glycoprotein expression, increased c-Jun expression in Schwann cells, and increased both the number of macrophages and the amount of myelin fragments in the macrophages. These findings suggest that ascorbic acid accelerates Wallerian degeneration by accelerating the degeneration of axons and myelin in the injured nerve, promoting the dedifferentiation of Schwann cells, and enhancing macrophage recruitment and phagocytosis. The study was approved by the Southern Medical University Animal Care and Use Committee (approval No. SMU-L2015081) on October 15, 2015.

Lithium Loaded Octa-Poly(Ethylene Glycol) Based Adhesive Facilitates Axon Regeneration and Reconnection of Transected Peripheral Nerves.

At present, reconnecting the transected nerve in clinic is still mainly reliant on surgery suture. This is a procedure that requires thorough training and is also time consuming. Here, an octa-poly(ethylene glycol) (PEG)-based adhesive for fast reconnecting of the transected peripheral nerve is reported. To enhance the therapeutic efficacy, a succinyl unit is applied to endow the controllably dissolvable property of the adhesive, and lithium is loaded in the adhesive to improve the axonal regeneration. Present data reveal that this adhesive possesses good cytocompatibility and can significantly shorten the reconnecting time of the transected nerve ends compared to that required for suture surgery. Histology, electrophysiological, and behavior assessments indicate that the adhesive reconnected nerves exhibit a low grade of fibrosis, inflammation response, and myoatrophy as well as robust axonal regeneration and functional recovery. Together, these results indicate that this octa-PEG adhesive can act as an alternative to traditional nerve suture in peripheral nerve injury.

Electric Conductivity on Aligned Nanofibers Facilitates the Transdifferentiation of Mesenchymal Stem Cells into Schwann Cells and Regeneration of Injured Peripheral Nerve.

Schwann cells (SCs) are the most promising seed cells for peripheral nerve tissue engineering, but clinical applications are limited by the lack of cell sources. Existing data demonstrate that bone marrow mesenchymal stem cells (BMSCs) can be induced to differentiate into Schwann-like cells and aligned nanofibers can enhance the differentiation. Considering that SCs are living along with the electrical conductive axons, it is hypothesized that conductivity properties may play roles in SCs differentiation and then facilitate nerve regeneration. To verify this hypothesis, amine functionalized multi-walled carbon nanotubes (MWCNTs) are incorporated with polycaprolactone and gelatin to fabricate aligned or random conductive nanofibers by electrospinning. Current data demonstrate that MWCNTs can dramatically increase the electrical conductive properties but do not alter the biocompatibility of the nanofibers. It is found that endowing conductive properties into the aligned nanofibers can significantly enhance their capability to promote the SCs differentiation. Furthermore, the aligned and conductive nanofibers with induced BMSCs can dramatically promote peripheral axonal regeneration. Collectively, the present study demonstrates that the conductive properties in the aligned nanofiber plays significant roles in SCs differentiation and the aligned and conductive nanofibers can be used as a promising scaffold for SCs differentiation and peripheral nerve tissue engineering.