Injection of a PMMA-doped MSC spheroid gel for the treatment of painful osteoporotic vertebral compression fractures.

We aimed to develop a biocompatible treatment to overcome the limitations of polymethyl methacrylate (PMMA) vertebroplasty for osteoporotic compression fracture patients. We synthesized an injectable hydrogel containing PMMA. Mesenchymal stem cell (MSC) spheroids were included in the injectable PMMA-doped gel (= PMMA-doped spheroid gel). In vitro, the osteogenic/anti-inflammatory effects of the embedded spheroids were investigated by the quantitative real-time polymerase chain reaction method. In vivo, we used ovariectomy (OVX)-induced osteoporotic rats with injured femurs to investigate the pain-relief effects. The OVX rats were divided into four groups according to the materials injected (non, PMMA, PMMA gel, and PMMA-spheroid gel) into the lesion. The immunofluorescence (IF) intensity levels of painful markers in dorsal root ganglia (DRG) were measured. In vitro, a volumetric ratio of the gel of 8 (gel):2 (PMMA) was non-cytotoxic for MSCs and promoted the expression of osteogenic/anti-inflammatory markers. In vivo, the values of several bone parameters in the PMMA-doped spheroid gel group showed remarkable increases compared to those in the PMMA group. In addition, the IF intensity levels of the painful markers were noticeably decreased in the PMMA-spheroid gel group. We, therefore, suggest that this treatment can be useful for osteoporotic vertebral compression fracture patients.

Axon guidance gene-targeted siRNA delivery system improves neural stem cell transplantation therapy after spinal cord injury.

BACKGROUND: Neural stem cells (NSCs) derived from the embryonic spinal cord are excellent candidates for the cellular regeneration of lost neural cells after spinal cord injury (SCI). Semaphorin 3 A (Sema3A) is well known as being implicated in the major axon guidance of the growth cone as a repulsive function during the development of the central nervous system, yet its function in NSC transplantation therapy for SCI has not been investigated. Here, we report for the first time that embryonic spinal cord-derived NSCs significantly express Sema3A in the SCI environment, potentially facilitating inhibition of cell proliferation after transplantation. METHODS: siRNA-Sema3A was conjugated with poly-l-lysin-coated gold nanoparticles (AuNPs) through a charge interaction process. NSCs were isolated from embryonic spinal cords of rats. Then, the cells were embedded into a dual-degradable hydrogel with the siRNA- Sema3A loaded-AuNPs and transplanted after complete SCI in rats. RESULTS: The knockdown of Sema3A by delivering siRNA nanoparticles via dual-degradable hydrogels led to a significant increase in cell survival and neuronal differentiation of the transplanted NSCs after SCI. Of note, the knockdown of Sema3A increased the synaptic connectivity of transplanted NSC in the injured spinal cord. Moreover, extracellular matrix molecule and functional recovery were significantly improved in Sema3A-inhibited rats compared to those in rats with only NSCs transplanted. CONCLUSIONS: These findings demonstrate the important role of Sema3A in NSC transplantation therapy, which may be considered as a future cell transplantation therapy for SCI cases.

Anti-Inflammatory Effects of Tegoprazan in Lipopolysaccharide-Stimulated Bone-Marrow-Derived Macrophages.

The purpose of this study was to investigate the anti-inflammatory effect of tegoprazan (TEGO) in lipopolysaccharide (LPS)-stimulated bone-marrow-derived macrophages (BMMs). To this end, compared to methylprednisolone (MP; positive control), we evaluated whether TEGO effectively differentiates LPS-stimulated BMMs into M2-phenotype macrophages. Moreover, the expression of pro- and anti-inflammatory cytokines genes influenced by TEGO was measured using quantitative real-time polymerase chain reaction (qRT-PCR) analysis. TEGO was found to reduce nitric oxide (NO) production in BMMs significantly. In addition, TEGO significantly decreased and increased the gene expression levels of pro-inflammatory and anti-inflammatory cytokines, respectively. In addition, we evaluated the phosphorylated values of the extracellular signal-regulatory kinase (ERK) and p38 in the mitogen-activated protein (MAP) kinase signaling pathway through Western blotting. TEGO significantly reduced the phosphorylated values of the ERK and p38. In other words, TEGO suppressed the various pro-inflammatory responses in LPS-induced BMMs. These results show that TEGO has the potential to be used as an anti-inflammatory agent.

Transplantation of neuron-inducing grafts embedding positively charged gold nanoparticles for the treatment of spinal cord injury.

In this study, we aimed to investigate the recovery after traumatic spinal cord injury (SCI) by inducing cellular differentiation of transplanted neural stem cells (NSCs) into neurons. We dissociated NSCs from the spinal cords of Fisher 344 rat embryos. An injectable gel crosslinked with glycol chitosan and oxidized hyaluronate was used as a vehicle for NSC transplantation. The gel graft containing the NSC and positively charged gold nanoparticles (pGNP) was implanted into spinal cord lesions in Sprague-Dawley rats (NSC-pGNP gel group). Cellular differentiation of grafted NSCs into neurons (stained with ß-tubulin III [also called Tuj1]) was significantly increased in the NSC-pGNP gel group (***p < 0.001) compared to those of two control groups (NSC and NSC gel groups) in the SCI conditions. The NSC-pGNP gel group showed the lowest differentiation into astrocytes (stained with glial fibrillary acidic protein). Regeneration of damaged axons (stained with biotinylated dextran amines) within the lesion was two-fold higher in the NSC-pGNP gel group than that in the NSC gel group. The highest locomotor scores were also found in the NSC-pGNP gel group. These outcomes suggest that neuron-inducing pGNP gel graft embedding embryonic spinal cord-derived NSCs can be a useful type of stem cell therapy after SCI.

Lobeglitazone Exerts Anti-Inflammatory Effect in Lipopolysaccharide-Induced Bone-Marrow Derived Macrophages.

The purpose of this study is to elucidate the anti-inflammatory effect of lobeglitazone (LOBE) in lipopolysaccharide (LPS)-induced bone-marrow derived macrophages (BMDMs). We induced nitric oxide (NO) production and pro-inflammatory gene expression through LPS treatment in BMDMs. The changes of NO release and expression of pro-inflammatory mediators by LOBE were assessed via NO quantification assay and a real-time quantitative polymerase chain reaction (RT-qPCR), respectively. In addition, the regulatory effect of LOBE on activation of mitogen-activated protein kinase (MAPK) signaling pathway was investigated by measuring the phosphorylation state of extracellular regulatory protein (ERK) and c-Jun N-terminal kinase (JNK) proteins by Western blot. Our results show that LOBE significantly reduced LPS-induced NO production and pro-inflammatory gene expression of interleukin-1ß (IL-1ß), IL-6, inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and monocyte chemoattractant protein-1 (MCP-1). Moreover, LOBE reduced phosphorylation levels of ERK and JNK of MAPK signaling pathway. In conclusion, LOBE exerts an anti-inflammatory effect in LPS-induced BMDMs by suppression of NO production and pro-inflammatory gene expression, and this effect is potentially through inhibition of the MARK signaling pathway.

Chirality-Dependent Anti-Inflammatory Effect of Glutathione after Spinal Cord Injury in an Animal Model.

Neuroinflammation forms a glial scar following a spinal cord injury (SCI). The injured axon cannot regenerate across the scar, suggesting permanent paraplegia. Molecular chirality can show an entirely different bio-function by means of chiral-specific interaction. In this study, we report that d-chiral glutathione (D-GSH) suppresses the inflammatory response after SCI and leads to axon regeneration of the injured spinal cord to a greater extent than l-chiral glutathione (L-GSH). After SCI, axon regrowth in D-GSH-treated rats was significantly increased compared with that in L-GSH-treated rats (*** p < 0.001). Secondary damage and motor function were significantly improved in D-GSH-treated rats compared with those outcomes in L-GSH-treated rats (** p < 0.01). Moreover, D-GSH significantly decreased pro-inflammatory cytokines and glial fibrillary acidic protein (GFAP) via inhibition of the mitogen-activated protein kinase (MAPK) signaling pathway compared with L-GSH (*** p < 0.001). In primary cultured macrophages, we found that D-GSH undergoes more intracellular interaction with activated macrophages than L-GSH (*** p < 0.001). These findings reveal a potential new regenerative function of chiral GSH in SCI and suggest that chiral GSH has therapeutic potential as a treatment of other diseases.

Direct Injection of Hydrogels Embedding Gold Nanoparticles for Local Therapy after Spinal Cord Injury.

In this study, we created a hydrogel composed of glycol chitosan (gC) and oxidized hyaluronate (oHA). Gold nanoparticles (GNPs) were conjugated with ursodeoxycholic acid (UDCA). The GNP-UDCA complex was embedded into gC-oHA (CHA) hydrogels to form a CHA-GNP-UDCA gel. This CHA-GNP-UDCA gel was injected once into an epicenter of an injured region in SCI rats. Near-infrared (NIR) irradiation was then applied to the lesion as a means of local therapy. To optimize the viscosity for injection into a lesion, several volume ratios of gC and oHA were investigated using scanning electron microscopy and a rotating rheometer. The optimally synthesized CHA-GNP-UDCA gel under NIR irradiation suppressed the production of inflammatory cytokines in vitro. In addition, the optimized CHA-GNP-UDCA gel under NIR irradiation inhibited the cystic cavity of the lesion and significantly improved the hindlimb function. The production of inflammatory cytokines following SCI was significantly inhibited in the CHA-GNP-UDCA gel + NIR group. CHA-GNP-UDCA gels with NIR irradiation can therefore have therapeutic effects for those with spinal cord injuries.

Transplantation of tauroursodeoxycholic acid-inducing M2-phenotype macrophages promotes an anti-neuroinflammatory effect and functional recovery after spinal cord injury in rats.

OBJECTIVES: In this study, we study the transplantation of tauroursodeoxycholic acid (TUDCA)-induced M2-phenotype (M2) macrophages and their ability to promote anti-neuroinflammatory effects and functional recovery in a spinal cord injury (SCI) model. METHODS: To this end, compared to the granulocyte-macrophage colony-stimulating factor (GM-CSF), we evaluated whether TUDCA effectively differentiates bone marrow-derived macrophages (BMDMs) into M2 macrophages. RESULTS: The M2 expression markers in the TUDCA-treated BMDM group were increased more than those in the GM-CSF-treated BMDM group. After the SCI and transplantation steps, pro-inflammatory cytokine levels and the mitogen-activated protein kinase (MAPK) pathway were significantly decreased in the TUDCA-induced M2 group more than they were in the GM-CSF-induced M1 group and in the TUDCA group. Moreover, the TUDCA-induced M2 group showed significantly enhanced tissue volumes and improved motor functions compared to the GM-CSF-induced M1 group and the TUDCA group. In addition, biotinylated dextran amine (BDA)-labelled corticospinal tract (CST) axons and neuronal nuclei marker (NeuN) levels were increased in the TUDCA-induced M2 group more than those in the GM-CSF-induced M1 group and the TUDCA group. CONCLUSIONS: This study demonstrates that the transplantation of TUDCA-induced M2 macrophages promotes an anti-neuroinflammatory effect and motor function recovery in SCI. Therefore, we suggest that the transplantation of TUDCA-induced M2 macrophages represents a possible alternative cell therapy for SCI.

Injectable Hydrogel Containing Tauroursodeoxycholic Acid for Anti-neuroinflammatory Therapy After Spinal Cord Injury in Rats.

We investigate the anti-inflammatory effects of injectable hydrogel containing tauroursodeoxycholic acid (TUDCA) in a spinal cord injury (SCI) model. To this end, TUDCA-hydrogel (TC gel) is created by immersing the synthesized hydrogel in a TUDCA solution for 1 h. A mechanical SCI was imposed on rats, after which we injected the TC gel. After the SCI and injections, motor functions and lesions were significantly improved in the TC gel group compared with those in the saline group. The TC gel significantly decreased pro-inflammatory cytokine levels compared with the saline; TUDCA and glycol chitosan-oxidized hyaluronate were mixed at a ratio of 9:1 (CHA) gel independently. In addition, the TC gel significantly suppressed the phosphorylation of extracellular signal-regulated kinase (p-ERK) and c-Jun N-terminal kinase (p-JNK) in the mitogen-activated protein kinase (MAPK) pathway compared with the saline, TUDCA, and CHA gel independently. It also decreased tumor necrosis factor-α (TNF-α) and glial fibrillary acidic protein (GFAP), inflammatory marker, at the injured sites more than those in the saline, TUDCA, and CHA gel groups. In conclusion, the results of this study demonstrate the neuroinflammatory inhibition effects of TC gel in SCI and suggest that TC gel can be an alternative drug system for SCI cases.