Thread-structural microneedles loaded with engineered exosomes for annulus fibrosus repair by regulating mitophagy recovery and extracellular matrix homeostasis.

Low back pain is among the most grave public health concerns worldwide and the major clinical manifestation of intervertebral disc degeneration (IVDD). The destruction of annulus fibrosus (AF) is the primary cause of IVDD. A sustainable and stable treatment system for IVDD is lacking because of the special organizational structure and low nutrient supply of AF. We here found that IVDD results in the impaired mitochondrial function of AF tissue, and mitochondrial autophagy (mitophagy) plays a protective role in this process. We therefore reported a thread-structural microneedle (T-MN) matching the ring structure of AF. Based on the adsorption effect of laminin, our T-MN could load with bone marrow mesenchymal stem cell-derived exosomes to envelope the regulating mitophagy microRNA (miRNA 378), named as T-MN@EXO@miR-378. In general, we offered in situ locking in the defect site of AF to prevent nucleus pulposus leakage and promoted AF repair. The design of the thread structure was aimed at bionically matching the layered AF structure, thereby providing stronger adhesion. The T-MN@EXO@miR-378 effectively attached to AF and slowly released therapeutic engineered exosomes, and prevented IVDD progression by restoring mitophagy, promoting AF cell proliferation and migration, and inhibiting the pathological remodeling of the extracellular matrix. This functional system can be used as an excellent tool for sustained drug release and has a certain prospect in substituting the conventional treatment of IVDD.

Single-Cell Microgel Encapsulation Improves the Therapeutic Efficacy of Mesenchymal Stem Cells in Treating Intervertebral Disc Degeneration via Inhibiting Pyroptosis.

While mesenchymal stem cell (MSC) shows great potentials in treating intervertebral disc degeneration, most MSC die soon after intradiscal transplantation, resulting in inferior therapeutic efficacy. Currently, bulk hydrogels are the common solution to improve MSC survival in tissues, although hydrogel encapsulation impairs MSC migration and disrupts extracellular microenvironment. Cell hydrogel encapsulation has been proposed to overcome the limitation of traditional bulk hydrogels, yet this technique has not been used in treating disc degeneration. Using a layer-by-layer self-assembly technique, we fabricated alginate and gelatin microgel to encapsulate individual MSC for treating disc degeneration. The small size of microgel allowed intradiscal injection of coated MSC. We demonstrated that pyroptosis was involved in MSC death under oxidative stress stimulation, and microgel coating suppressed pyroptosis activation by maintaining mitochondria homeostasis. Microgel coating protected MSC in the harsh disc microenvironment, while retaining vital cellular functions such as migration, proliferation, and differentiation. In a rat model of disc degeneration, coated MSC exhibits prolonged retention in the disc and better efficacy of attenuating disc degeneration, as compared with bare MSC treatment alone. Further, microgel-coated MSC exhibited improved therapeutic effects in treating disc degeneration via suppressing the activation of pyroptosis in the disc. For the first time, microgel-encapsulated MSC was used to treat disc degeneration and obtain encouraging outcomes. The developed biocompatible single-cell hydrogel is an effective strategy to protect MSC and maintain cellular functions and may be an efficacious approach to improving the efficacy of MSC therapy in treating disc degeneration. The objective of this study is to improve the efficacy of cell therapy for treating disc degeneration using single-cell hydrogel encapsulation and further to understand related cytoprotective mechanisms.

Retinoic acid inhibits the pyroptosis of degenerated nucleus pulposus cells by activating Sirt1-SOD2 signaling.

AIM: Intervertebral disc (IVD) degeneration is a common disease initiated by the degeneration of the nucleus pulposus (NP). The pyroptosis of degenerated NP cells (dNPCs) plays an important role in NP degeneration. The purpose of this study is to identify a feasible solution that can inhibit NP cell pyroptosis to therapy the degeneration of the intervertebral disc. METHODS: Cell viability and proliferation were quantified by Cell Counting Kit-8 assay. The measurement of cellular reactive oxygen species (ROS) was detected by 2,7-Dichlorodi-hydrofluorescein diacetate. The death of cells was analyzed by the Terminal Deoxynucleotidyl Transferase-mediated dUTP Nick-End Labeling (TUNEL) method of fluorescence analysis. The pyroptosis of cells was assessed by flow cytometry analyses. The contents of sulfate glycosaminoglycans were detected by a blyscan assay kit. RESULT: In this study, we determined the effects of retinoic acid (RA) on dNPCs and investigated the underlying mechanism of RA-mediated pyroptosis in dNPCs. We also verified the effects of RA on IVD degeneration in vivo. Our results demonstrated that RA significantly increased the proliferation and the protein expression of sox9, aggrecan, and collagen II of dNPCs. Pyroptosis-related proteins and the pyroptosis rate of dNPCs were significantly decreased by RA. We found that Sirt1-SOD2 signaling was activated, while ROS generation and TXNIP/NLRP3 signaling in dNPCs were inhibited after the addition of RA. Furthermore, RA also recovered the structure of NP and increased the contents of sulfated glycosaminoglycans and collagen in vivo. CONCLUSION: Our study demonstrated that RA could inhibit the pyroptosis and increase the extracellular matrix synthesis function of dNPCs and verified that RA has a protective effect on IVD degeneration.

Causal Relationships Between Total Physical Activity and Ankylosing Spondylitis: A Mendelian Randomization Study.

Background: Currently, there is little literature about the association between physical activity (PA) and the risk of ankylosing spondylitis (AS). The present study aimed to understand the causal relationships between PA and AS. Methods: We performed two-sample Mendelian randomization (MR) using publicly released genome-wide association studies summary statistics to estimate the causal associations of PA with AS risk. The inverse variance weighted (IVW) method was utilized as primary MR analysis. Furthermore, sensitivity, pleiotropy, and heterogeneity analyses were then conducted to assess the robustness of the findings of the present study. Results: Results of the IVW analysis suggested a protective relationship between accelerometer-based PA and AS (average acceleration, odds ratio [OR] = 0.9995, 95% CI, 0.9988-0.9999, P = 0.014). On the contrary, there was no causal relationship between accelerometer-based PA (acceleration fraction >425 mg; OR = 0.9981, 95% CI = 0.9936-1.0026, P = 0.402) and AS. Furthermore, there was no significant relationship between self-reported vigorous PA and AS (OR = 1.0005, 95% CI = 0.9875-1.0136, P = 0.943), or even between self-reported moderate-to-vigorous PA and AS (OR = 1.0000, 95% CI, 0.9947-1.0052; P = 0.990). Conclusions: The use of genetic approach in the present study revealed that total physical activity (TPA) has a protective relationship with AS risk. Furthermore, it was evident that vigorous PA or moderate-to-vigorous physical levels are not causally associated with AS. Therefore, the present study evidently supports the hypothesis that enhancing TPA rather than PA intensity is an effective prevention strategy for AS.

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.

Mesenchymal Stem Cell-Derived Extracellular Vesicles: Immunomodulatory Effects and Potential Applications in Intervertebral Disc Degeneration.

Intervertebral disc (IVD) degenerative disease is a common health problem worldwide. Administration of mesenchymal stem cells (MSCs) in intervertebral disc degeneration (IVDD) has been widely explored in recent years. However, transplantation of MSCs is restricted by several factors. Currently, paracrine signaling is one of the main mechanisms by which MSCs play a therapeutic role in disc regeneration. Extracellular vehicles (EVs) are the main paracrine products of MSCs. They show great potential as an effective alternative to MSCs and play immunomodulation roles such as anti-inflammatory effects, antioxidative stress, antiapoptosis, and antiextracellular matrix (ECM) degradation during treatment of IVDD. This review focuses on the immunomodulatory effect of MSC EVs and their potential applications.

Injectable exosome-functionalized extracellular matrix hydrogel for metabolism balance and pyroptosis regulation in intervertebral disc degeneration.

Exosome therapy is a promising therapeutic approach for intervertebral disc degeneration (IVDD) and achieves its therapeutic effects by regulating metabolic disorders, the microenvironment and cell homeostasis with the sustained release of microRNAs, proteins, and transcription factors. However, the rapid clearance and disruption of exosomes are the two major challenges for the application of exosome therapy in IVDD. Herein, a thermosensitive acellular extracellular matrix (ECM) hydrogel coupled with adipose-derived mesenchymal stem cell (ADSC) exosomes (dECM@exo) that inherits the superior properties of nucleus pulposus tissue and ADSCs was fabricated to ameliorate IVDD. This thermosensitive dECM@exo hydrogel system can provide not only in situ gelation to replenish ECM leakage in nucleus pulposus cells (NPCs) but also an environment for the growth of NPCs. In addition, sustained release of ADSC-derived exosomes from this system regulates matrix synthesis and degradation by regulating matrix metalloproteinases (MMPs) and inhibits pyroptosis by mitigating the inflammatory response in vitro. Animal results demonstrated that the dECM@exo hydrogel system maintained early IVD microenvironment homeostasis and ameliorated IVDD. This functional system can serve as a powerful platform for IVD drug delivery and biotherapy and an alternative therapy for IVDD.

The preconditioning of lithium promotes mesenchymal stem cell-based therapy for the degenerated intervertebral disc via upregulating cellular ROS.

Adipose-derived stem cell (ADSC) is one of the most widely used candidate cell for intervertebral disc (IVD) degeneration-related disease. However, the poor survival and low differentiation efficacy in stressed host microenvironment limit the therapeutic effects of ADSC-based therapy. The preconditioning has been found effective to boost the proliferation and the functioning of stem cells in varying pathological condition. Lithium is a common anti-depression drug and has been proved effective to enhance stem cell functioning. In this study, the effects of preconditioning using LiCl on the cellular behavior of ADSC was investigated, and specially in a degenerative IVD-like condition. METHOD: The cellular toxicity on rat ADSC was assessed by detecting lactate dehydrogenase (LDH) production after treatment with a varying concentration of lithium chloride (LiCl). The proliferative capacity of ADSC was determined by detecting Ki67 expression and the relative cell number of ADSC. Then, the preconditioned ADSC was challenged by a degenerative IVD-like condition. And the cell viability as well as the nucleus pulpous (NP) cell differentiation efficacy of preconditioned ADSC was evaluated by detecting the major marker expression and extracellular matrix (ECM) deposit. The therapeutic effects of preconditioned ADSC were evaluated using an IVD degeneration rat model, and the NP morphology and ECM content were assessed. RESULTS: A concentration range of 1-10 mmol/L of LiCl was applied in the following study, since a higher concentration of LiCl causes a major cell death (about 40%). The relative cell number was similar between preconditioned groups and the control group after preconditioning. The Ki67 expression was elevated after preconditioning. Consistently, the preconditioned ADSC showed stronger proliferation capacity. Besides, the preconditioned groups exhibit higher expression of NP markers than the control group after NP cell induction. Moreover, the preconditioning of LiCl reduced the cell death and promoted ECM deposits, when challenged with a degenerative IVD-like culture. Mechanically, the preconditioning of LiCl induced an increased cellular reactive oxidative species (ROS) level and activation of ERK1/2, which was found closely related to the enhanced cell survival and ECM deposits after preconditioning. The treatment with preconditioned ADSC showed better therapeutic effects than control ADSC transplantation, with better NP preservation and ECM deposits. CONCLUSION: These results suggest that the preconditioning with a medium level of LiCl boosts the cell proliferation and differentiation efficacy under a normal or hostile culture condition via the activation of cellular ROS/ERK axis. It is a promising pre-treatment of ADSC to promote the cell functioning and the following regenerative capacity, with superior therapeutic effects than untreated ADSC transplantation.

miRNA-141 Induced Pyroptosis in Intervertebral Disk Degeneration by Targeting ROS Generation and Activating TXNIP/NLRP3 Signaling in Nucleus Pulpous Cells.

The role and mechanism of pyroptosis in intervertebral disk (IVD) degeneration are unclear. MicroRNAs (miRNAs) regulate the viability and function of nucleus pulposus cells (NPCs) in IVDs and are related to pyroptosis. We performed microarray analyses of normal and degenerated nucleus pulposus (NP) to assess the role of pyroptosis and identify key miRNAs in IVD degeneration. We also evaluated the underlying mechanism of miRNA-mediated pyroptosis in NPCs. In addition, we demonstrated the preventative effects of miRNAs on IVD degeneration in a rat model. The levels of the pyroptosis-related proteins cleaved caspase-1, N-terminal gasdermin D (GSDMD), interleukin (IL)-1ß, and IL-18 in the degenerative NP were significantly higher than those in the normal NP. miRNA-141 was significantly upregulated in the degenerated NP. miR-141 mimic suppressed the matrix synthesis function of NPCs. By contrast, reactive oxygen species (ROS) generation, and the expression of TXNIP and NLRP3 were significantly downregulated by an miR-141 inhibitor. Furthermore, the miRNA-141 inhibitor prevented the degeneration of IVDs in vivo. Our findings suggest that miRNA-141 induces pyroptosis and extracellular matrix (ECM) catabolism in NPCs by increasing ROS generation and activating TXNIP/NLRP3 signaling. miRNA-141-regulated pyroptosis may be a novel therapeutic target for IVD degeneration.

The multiple function of long noncoding RNAs in osteosarcoma progression, drug resistance and prognosis.

Osteosarcoma is a bone tumor prevalent in children and young adults. LncRNAs are a family of non-protein-coding transcripts longer than 200 nucleotides. The tumor-related pathological functions of lncRNAs include proliferation, migration, and chemotherapy resistance, all of which have been widely acknowledged in research on osteosarcoma. In addition, compelling evidence suggests that lncRNAs could serve as diagnostic indicators, prognostic biomarkers, and targets for disease treatment. In this review, we systematically summarize how lncRNAs regulate tumorigenesis, invasion and therapeutic resistance. By deepening our knowledge of the relationship between lncRNAs and osteosarcoma, we hope to translate research findings into clinical applications as soon as possible.

[The anti-leukemic effects of imatinib, daunorubicin and bortezomib on two leukemia cell lines with Ph(+)].

OBJECTIVE: To study the proliferative inhibition effects of imatinib, daunorubicin and bortezomib on two leukemia cell lines with Ph(+), chronic myelogenous leukemia cell line K562 expressing P210 protein and acute lymphoblastic leukemia cell line SUP-B15 expressing P190 protein. METHODS: (1) The cells of the two cell lines treated with imatinib, daunorubicin and bortezomib for 72 hours were analyzed by MTT assay for proliferation. The proliferative activity was displayed by growth curve and IC50 value. (2) The bcr-abl transcriptant in the cells treated with imatinib (final concentration at 0, 0.35, 1 micromol/L) for 48 hours was detected by reverse transcription polymerase chain reaction (RT-PCR). RESULTS: (1) The IC50 values of K562 and SUP-B15 cell lines treated with imatinib, daunorubicin and bortezomib for 72 hours were respectively (0.286 +/- 0.060) micromol/L, (0.303 +/- 0.009) micromol/L, (22.127 +/- 3.592) nmol/L and (1.387 +/- 0.180) micromol/L, (0.117 +/- 0.017) micromol/L, (12.350 +/- 0.740) nmol/L. (2) There was no change of bcr-abl expression level in both cell lines after the treatment of imatinib. CONCLUSION: Imatinib, daunorubicin and bortezomib showed anti-cancer effects on Ph(+) leukemia cells in vitro. K562 cells were more sensitive to imatinib than the other two drugs, whereas SUP-B15 cells are more sensitive to daunorubicin and bortezomib. The short time intervention of imatinib has no effect on the expression of bcr-abl in Ph (+) leukemia cell lines.