Dedifferentiation-like reprogramming of degenerative nucleus pulposus cells into notochordal-like cells by defined factors.

The extensive degeneration of functional somatic cells and the depletion of endogenous stem/progenitor populations present significant challenges to tissue regeneration in degenerative diseases. Currently, a cellular reprogramming approach enabling directly generating corresponding progenitor populations from degenerative somatic cells remains elusive. The present study focused on intervertebral disc degeneration (IVDD) and identified a three-factor combination (OCT4, FOXA2, TBXT [OFT]) that could induce the dedifferentiation-like reprogramming of degenerative nucleus pulposus cells (dNPCs) toward induced notochordal-like cells (iNCs). Single-cell transcriptomics dissected the transitions of cell identity during reprogramming. Further, OCT4 was found to directly interact with bromodomain PHD-finger transcription factor to remodel the chromatin during the early phases, which was crucial for initiating this dedifferentiation-like reprogramming. In rat models, intradiscal injection of adeno-associated virus carrying OFT generated iNCs from in situ dNPCs and reversed IVDD. These results collectively present a proof-of-concept for dedifferentiation-like reprogramming of degenerated somatic cells into corresponding progenitors through the development of a factor-based strategy, providing a promising approach for regeneration in degenerative disc diseases.

Development of Uveal Melanoma-Specific Aptamer for Potential Biomarker Discovery and Targeted Drug Delivery.

Uveal melanoma (UM) is the most common primary intraocular malignancy in adults. However, challenges in early diagnosis, high risk of liver metastasis, and lack of effective targeted therapy lead to poor prognosis and high mortality of UM. Therefore, generating an effective molecular tool for UM diagnosis and targeted treatment is of great significance. In this study, a UM-specific DNA aptamer, PZ-1, was successfully developed, which could specifically distinguish molecular differences between UM cells and noncancerous cells with nanomolar-range affinity and presented excellent recognition ability for UM in vivo and clinical UM tissues. Subsequently, the binding target of PZ-1 on UM cells was identified as JUP (junction plakoglobin) protein, which held great potential as a biomarker and therapeutic target for UM. Meanwhile, the strong stability and internalization capacity of PZ-1 were also determined, and a UM-specific aptamer-guided "nanoship" was engineered to load and selectively release doxorubicin (Dox) to targeted UM cells, with lower toxicity to nontumor cells. Taken together, the UM-specific aptamer PZ-1 could serve as a molecular tool to discover the potential biomarker for UM and to achieve the targeted therapy of UM.

Swelling-Mediated Mechanical Stimulation Regulates Differentiation of Adipose-Derived Mesenchymal Stem Cells for Intervertebral Disc Repair Using Injectable UCST Microgels.

Mechanical stimulation is an effective approach for controlling stem cell differentiation in tissue engineering. However, its realization in in vivo tissue repair remains challenging since this type of stimulation can hardly be applied to injectable seeding systems. Here, it is presented that swelling of injectable microgels can be transformed to in situ mechanical stimulation via stretching the cells adhered on their surface. Poly(acrylamide-co-acrylic acid) microgels with the upper critical solution temperature property are fabricated using inverse emulsion polymerization and further coated with polydopamine to increase cell adhesion. Adipose-derived mesenchymal stem cells (ADSCs) adhered on the microgels can be omnidirectionally stretched along with the responsive swelling of the microgels, which upregulate TRPV4 and Piezo1 channel proteins and enhance nucleus pulposus (NP)-like differentiation of ADSCs. In vivo experiments reveal that the disc height and extracellular matrix content of NP are promoted after the implantation with the microgels. The findings indicate that swelling-induced mechanical stimulation has great potential for regulating stem cell differentiation during intervertebral disc repair.

A conductive supramolecular hydrogel creates ideal endogenous niches to promote spinal cord injury repair.

The current effective method for treatment of spinal cord injury (SCI) is to reconstruct the biological microenvironment by filling the injured cavity area and increasing neuronal differentiation of neural stem cells (NSCs) to repair SCI. However, the method is characterized by several challenges including irregular wounds, and mechanical and electrical mismatch of the material-tissue interface. In the current study, a unique and facile agarose/gelatin/polypyrrole (Aga/Gel/PPy, AGP3) hydrogel with similar conductivity and modulus as the spinal cord was developed by altering the concentration of Aga and PPy. The gelation occurred through non-covalent interactions, and the physically crosslinked features made the AGP3 hydrogels injectable. In vitro cultures showed that AGP3 hydrogel exhibited excellent biocompatibility, and promoted differentiation of NSCs toward neurons whereas it inhibited over-proliferation of astrocytes. The in vivo implanted AGP3 hydrogel completely covered the tissue defects and reduced injured cavity areas. In vivo studies further showed that the AGP3 hydrogel provided a biocompatible microenvironment for promoting endogenous neurogenesis rather than glial fibrosis formation, resulting in significant functional recovery. RNA sequencing analysis further indicated that AGP3 hydrogel significantly modulated expression of neurogenesis-related genes through intracellular Ca2+ signaling cascades. Overall, this supramolecular strategy produces AGP3 hydrogel that can be used as favorable biomaterials for SCI repair by filling the cavity and imitating the physiological properties of the spinal cord.

Enhancement of nucleus pulposus repair by glycoengineered adipose-derived mesenchymal cells.

Adipose-derived mesenchymal stem cells (ADSCs) are promising candidates for repairing degenerated intervertebral discs through multiple means, including: i. Secretion of bioactive factors to regulate inflammation and, ii. The potential to differentiate into nucleus pulposus (NP)-like cells, which can integrate into host tissues. However, the differentiation ability of ADSCs to NP-like cells is limited, which emphasizes on the need for alternative approaches to regulate cell differentiations. Given that cell functions are influenced by interactions between the extracellular matrix (ECM) and cells, we hypothesize that cell surface modification promotes ADSCs adhesion and differentiation towards NP-like cells. In this study, cell surfaces of ADSCs were functionalized with unnatural sialic acid via metabolic glycoengineering. Subsequently, adhesion abilities of modified cells to three main ECM (laminin, collagen and fibronectin) were compared. The adhesion assay revealed that glycoengineered ADSCs had the highest affinity for collagen, compared to laminin and fibronectin. Moreover, cultures with collagen coated plates enhanced the differentiation of glycoengineered ADSCs to NP-like cells. Metabolic glycoengineering prolonged ADSCs viability. The glycoengineered ADSCs increased the height and elasticity of intervertebral discs, as well as the water content and ECM volumes of nucleus pulposus. In conclusion, metabolic glycoengineering of cell surfaces has a significant role in modulating cell biological functions and promoting NP tissue repair.

Partial reprogramming strategy for intervertebral disc rejuvenation by activating energy switch.

Rejuvenation of nucleus pulposus cells (NPCs) in degenerative discs can reverse intervertebral disc degeneration (IDD). Partial reprogramming is used to rejuvenate aging cells and ameliorate progression of aging tissue to avoiding formation of tumors by classical reprogramming. Understanding the effects and potential mechanisms of partial reprogramming in degenerative discs provides insights for development of new therapies for IDD treatment. The findings of the present study show that partial reprogramming through short-term cyclic expression of Oct-3/4, Sox2, Klf4, and c-Myc (OSKM) inhibits progression of IDD, and significantly reduces senescence related phenotypes in aging NPCs. Mechanistically, short-term induction of OSKM in aging NPCs activates energy metabolism as a "energy switch" by upregulating expression of Hexokinase 2 (HK2) ultimately promoting redistribution of cytoskeleton and restoring the aging state in aging NPCs. These findings indicate that partial reprogramming through short-term induction of OSKM has high therapeutic potential in the treatment of IDD.

A bioactive injectable self-healing anti-inflammatory hydrogel with ultralong extracellular vesicles release synergistically enhances motor functional recovery of spinal cord injury.

The repair and motor functional recovery after spinal cord injury (SCI) remains a worldwide challenge. The inflammatory microenvironment is one of main obstacles on inhibiting the recovery of SCI. Using mesenchymal stem cells (MSCs) derived extracellular vesicles to replace MSCs transplantation and mimic cell paracrine secretions provides a potential strategy for microenvironment regulation. However, the effective preservation and controlled release of extracellular vesicles in the injured spinal cord tissue are still not satisfied. Herein, we fabricated an injectable adhesive anti-inflammatory F127-polycitrate-polyethyleneimine hydrogel (FE) with sustainable and long term extracellular vesicle release (FE@EVs) for improving motor functional recovery after SCI. The orthotopic injection of FE@EVs hydrogel could encapsulate extracellular vesicles on the injured spinal cord, thereby synergistically induce efficient integrated regulation through suppressing fibrotic scar formation, reducing inflammatory reaction, promoting remyelination and axonal regeneration. This study showed that combining extracellular vesicles into bioactive multifunctional hydrogel should have great potential in achieving satisfactory locomotor recovery of central nervous system diseases.

Interpreting the Mechanisms by which Integrins Promote the Differentiation of Mesenchymal Stem Cells and Integrin Application Prospects.

Transmembrane integrin receptors represent a major component of cell-extracellular matrix (ECM) communications that mediate cellular biological activities, including proliferation and differentiation. Stem cells, especially mesenchymal stem cells (MSC), have rapidly emerged as promising therapies for various diseases. Dynamic links exist between extracellular and intracellular environments that profoundly influence the cellular activities via integrin receptors, such as cell morphology transformation and differentiation. Interpreting the roles of integrin receptors in the regulation of MSC differentiation may potentially lead to an amplified therapeutic effect. In this review, we summarize, for the first time, the potential mechanisms by which integrins promote MSC multilineage differentiation, including integrin downstream signaling cascades and the interactions between integrin and ion channels, the cytoskeleton, and nuclear mechanoresponses. Furthermore, we focus on the current state and future prospects of the application of integrins to promote cell differentiation.

Strategies and prospects of effective neural circuits reconstruction after spinal cord injury.

Due to the disconnection of surviving neural elements after spinal cord injury (SCI), such patients had to suffer irreversible loss of motor or sensory function, and thereafter enormous economic and emotional burdens were brought to society and family. Despite many strategies being dealing with SCI, there is still no effective regenerative therapy. To date, significant progress has been made in studies of SCI repair strategies, including gene regulation of neural regeneration, cell or cell-derived exosomes and growth factors transplantation, repair of biomaterials, and neural signal stimulation. The pathophysiology of SCI is complex and multifaceted, and its mechanisms and processes are incompletely understood. Thus, combinatorial therapies have been demonstrated to be more effective, and lead to better neural circuits reconstruction and functional recovery. Combinations of biomaterials, stem cells, growth factors, drugs, and exosomes have been widely developed. However, simply achieving axon regeneration will not spontaneously lead to meaningful functional recovery. Therefore, the formation and remodeling of functional neural circuits also depend on rehabilitation exercises, such as exercise training, electrical stimulation (ES) and Brain-Computer Interfaces (BCIs). In this review, we summarize the recent progress in biological and engineering strategies for reconstructing neural circuits and promoting functional recovery after SCI, and emphasize current challenges and future directions.

Transplantation Strategies for Spinal Cord Injury Based on Microenvironment Modulation.

Spinal cord injury (SCI) is different from peripheral nerve injury; it results in devastating and permanent damage to the spine, leading to severe motor, sensory and autonomic dysfunction. SCI produces a complex microenvironment that can result in hemorrhage, inflammation and scar formation. Not only does it significantly limit regeneration, but it also challenges a multitude of transplantation strategies. In order to promote regeneration, researchers have recently begun to focus their attention on strategies that manipulate the complicated microenvironment produced by SCI. And some have achieved great therapeutic effects. Hence, reconstructing an appropriate microenvironment after transplantation could be a potential therapeutic solution for SCI. In this review, first, we aim to summarize the influential compositions of the microenvironment and their different effects on regeneration. Second, we highlight recent research that used various transplantation strategies to modulate different microenvironments produced by SCI in order to improve regeneration. Finally, we discuss future transplantation strategies regarding SCI.

An injectable recombinant human milk fat globule-epidermal growth factor 8-loaded copolymer system for spinal cord injury reduces inflammation through NF-κB and neuronal cell death.

Spinal cord injury (SCI) is a common disease and a major cause of paralysis, carrying much burden around the world. Despite the progress made with growth factors therapy, the response rate of acute SCI treatment still remains unsatisfactory, due largely to complex and severe inflammatory reactions. Herein, we prepare a MFG-E8-loaded copolymer system-based anti-inflammation therapy for SCI treatment. It is shown that the MFG-E8-loaded copolymer system can decrease pro-inflammatory cytokine expression and neuron death. In a rat model of crush-caused SCI, the copolymer system shows significant therapeutic efficacy by ameliorating inflammation, decreasing fibrotic scar, promoting myelin regeneration and suppressing overall SCI severity.

Effects of Rhizoma Drynariae Cataplasm on Fracture Healing in a Rat Model of Osteoporosis.

BACKGROUND Osteoporosis is an increasingly prevalent disease characterized by decreased bone mass and deterioration of the bone microstructure, which contribute to increased fragility and subsequent fragility fractures, especially in elderly individuals. Rhizoma Drynariae (DRE) is among the most frequently used herbal medicines for the treatment of osteoporosis. Transdermal delivery is a proven novel pathway for drug treatment and has several advantages over traditional drug delivery routes. MATERIAL AND METHODS Female Sprague-Dawley osteoporotic fracture model rats were divided into 3 groups: the control group, the DRE (90 mg/kg/day) group and the DRE cataplasm (containing 30 mg DRE, administered at right femur site daily) group. At 3 and 6 weeks after operation, we performed x-ray, histological, and biomechanical analyses, and evaluated bone marrow density of the femur. RESULTS Treatment with DRE increased callus formation and bone union compared with the control group. Moreover, DRE enhanced bone strength at the femoral diaphysis in the osteoporotic fractures in rats by increasing the ultimate load and stiffness compared with the control group. Furthermore, DRE restored the trabecular bone mineral density in the femur compared with the control group. DRE cataplasm application further enhanced the therapeutic effects against osteoporotic fracture in this rat model. CONCLUSIONS DRE cataplasm application might be useful against osteoporotic fracture.

Glucagon-like peptide-1 receptor regulates endoplasmic reticulum stress-induced apoptosis and the associated inflammatory response in chondrocytes and the progression of osteoarthritis in rat.

Treatments for osteoarthritis (OA) are designed to restore chondrocyte function and inhibit cell apoptosis. Previous studies have shown that activation of the glucagon-like peptide-1 receptor (GLP-1R) leads to anti-inflammatory and anti-apoptotic effects. However, the role of GLP-1R in the pathological process of OA is unclear. In present work, we aimed to demonstrate the potential effect of GLP-1R on chondrocytes and elucidate its underlying mechanisms. We found that activation of GLP-1R with liraglutide could protect chondrocytes against endoplasmic reticulum stress and apoptosis induced by interleukin (IL)-1ß or triglycerides (TGs). These effects were partially attenuated by GLP-1R small interfering RNA treatment. Moreover, inhibiting PI3K/Akt signaling abolished the protective effects of GLP-1R by increase the apoptosis activity and ER stress. Activating GLP-1R suppressed the nuclear factor kappa-B pathway, decreased the release of inflammatory mediators (IL-6, tumor necrosis factor α), and reduced matrix catabolism in TG-treated chondrocytes; these effects were abolished by GLP-1R knockdown. In the end, liraglutide attenuated rat cartilage degeneration in an OA model of knee joints in vivo. Our results indicate that GLP-1R is a therapeutic target for the treatment of OA, and that liraglutide could be a therapeutic candidate for this clinical application.

Neuron and microglia/macrophage-derived FGF10 activate neuronal FGFR2/PI3K/Akt signaling and inhibit microglia/macrophages TLR4/NF-κB-dependent neuroinflammation to improve functional recovery after spinal cord injury.

Therapeutics used to treat central nervous system (CNS) injury were designed to repair neurites and inhibit cell apoptosis. Previous studies have shown that neuron-derived FGF10 exerts potential neuroprotective effects after cerebral ischemia injury. However, little is known about the role of endogenous FGF10 in the recovery process after spinal cord injury (SCI). In this study, we found that FGF10 is mainly produced by neuron and microglia/macrophages, and its expression is increased after SCI. Exogenous treatment of FGF10 improved functional recovery after injury by reducing apoptosis, as well as repairing neurites via FGFR2/PI3K/Akt pathway. On another hand, inhibiting the PI3K/Akt pathway with LY294002 partially reversed the therapeutic effects of FGF10. In addition, small interfering RNA knockdown of FGFR2 suppressed PI3K/Akt pathway activation by FGF10 and abolished its anti-apoptotic and neurite repair effects in vitro. Furthermore, FGF10 treatment inhibited the activation and proliferation of microglia/macrophages through regulation of TLR4/NF-κB pathway, and attenuated the release of pro-inflammatory cytokines after SCI. Thus, the increased expression of FGF10 after acute SCI is an endogenous self-protective response, suggesting that FGF10 could be a potential treatment for CNS injury.

Celastrol reduces IL-1ß induced matrix catabolism, oxidative stress and inflammation in human nucleus pulposus cells and attenuates rat intervertebral disc degeneration in vivo.

Celastrol has been reported to exert therapeutic potential on pro-inflammatory diseases including asthma, Crohn's disease, arthritis and neurodegenerative disorders via inhibiting NF-κB pathway. While the effect of celastrol on intervertebral disc degeneration (IDD), which is also a pro-inflammatory disease, remains unknown. In this study, we evaluated the effect of celastrol on IDD in IL-1ß treated human nucleus pulposus cells in vitro as well as in puncture induced rat IDD model in vivo. Our results showed that celastrol reduced the expression of catabolic genes (MMP-3, 9, 13, ADAMTS-4, 5), oxidative stress factors (COX-2, iNOS) and pro-inflammatory factors (IL-6, TNF-a) induced by IL-1ß in nucleus pulposus cells, also phosphorylation of IκBα and p65 were attenuated by celastrol, indicating NF-κB pathway was inhibited by celastrol in nucleus pulposus cells. In vivo study showed that celastrol treated rats had stronger T2-weighted signal than vehicle-treated rats at 2 weeks and 6 weeks' time point, suggesting celastrol could attenuate intervertebral disc degeneration in vivo. Together, our study demonstrates that celastrol could reduce IL-1ß induced matrix catabolism, oxidative stress and inflammation in human nucleus pulposus cells and attenuates rat intervertebral disc degeneration in vivo, which shows its potential to be a therapeutic drug for IDD.