Skip to main content
NCBI home page
ACS Omega logoLink to ACS Omega
. 2026 Sep 12;11(37):54886–54896. doi: 10.1021/acsomega.5c08053

Incorporation of Bone Marrow Stem Cell-Derived Exosomes into GelMA Hydrogel Effectively Attenuates Disc Degeneration in the Vertebrae of Rats

Yuwei Chen †, Yanjun Yang †, Yawei Chu †, Wanyi Wang †, Yu Liu †, Lifan Zhu †,‡,*
PMCID: PMC13613279  PMID: 42798950

Abstract

Chronic back pain and related disabilities are largely caused by intervertebral disc degeneration (IVDD). However, effective treatments remain limited. Existing treatment modalities, encompassing both conservative and surgical approaches, often fail to effectively arrest disease progression and may be associated with significant adverse effects. Recent studies have underscored the potential of tissue engineering and exosomes derived from bone marrow mesenchymal stem cells (BMSC-exos) as a promising strategy for the repair of intervertebral discs. This study aims to employ bioengineering techniques to enhance the retention and achieve sustained release of exosomes in the context of IVDD treatment. Our team developed a gelatin methacryloyl (GelMA) hydrogel system containing exosomes. Then Exosomes were mixed with GelMA to prepare a composite hydrogel (Exo-GelMA). This hydrogel-based system facilitates the targeted delivery of therapeutic agents to the degenerated disc, thereby modulating the proliferation, migration, and matrix synthesis of nucleus pulposus cells (NPCs). In a model of IVDD induced by puncture, this system notably slowed down the degeneration process and demonstrated considerable potential as a non-surgical treatment modality for IVDD, offering a promising strategy to attenuate the progression of IVDD and facilitate disc repair.

graphic file with name ao5c08053_0010.webp

graphic file with name ao5c08053_0008.webp

Introduction

Intervertebral disc degeneration (IVDD) is the leading factor in low back pain and is observed more often each year. At present, the primary therapeutic options for IVDD encompass surgical intervention and conservative management. However, surgical approaches are associated with a significant limitation: they may compromise the structural integrity of the spine. Conversely, conservative treatments encounter their own challenges, particularly regarding their suboptimal long-term prognoses. Consequently, stem cell-based tissue engineering technology has emerged as a promising alternative, offering innovative strategies for the restoration of the spine’s biological function. ,

Previous research has shown that exosomes derived from bone marrow mesenchymal stem cells (BMSC-exos) offer significant therapeutic advantages to nucleus pulposus cells (NPCs). These effects include promoting NPC proliferation, inhibiting inflammatory pathways, and enhancing the synthesis of the extracellular matrix (ECM). The homeostatic balance in the disc microenvironment is often disrupted by excessive catabolic enzyme expression, driving the progression of IVDD. Under inflammatory stress, NPCs increase the secretion of matrix-degrading enzymes like Matrix Metalloproteinase 3 (MMP3) and A Disintegrin and Metalloproteinase with Thrombospondin motifs (ADAMTS5). MMP3 breaks down collagen, while ADAMTS5 targets proteoglycans. Together, these enzymes erode the ECM, weakening the disc’s structure and function. In a rat model of IVDD, the intradiscal administration of BMSC-exos was found to effectively reduce the senescence of NPCs, preserve disc height, and attenuate the progression of degeneration. However, the therapeutic efficacy of free therapy in vivo is limited by restricted biological exposure and inadequate local retention. These factors hinder the achievement of sustained, therapeutically relevant concentrations, which are critical for ensuring long-term effectiveness.

Consequently, the development of engineered delivery systems capable of spatiotemporally regulating exosomes release is critically important. These systems have the potential to sustain localized concentrations of exosomes within degenerated intervertebral discs, which is essential for achieving enduring regenerative outcomes. Furthermore, such platforms should incorporate biomimetic design principles to replicate the natural paracrine signaling dynamics of MSCs, thereby addressing the pharmacokinetic limitations associated with free exosomes as well as the mechanical requirements of the disc microenvironment. This perspective highlights the pressing need for materials-driven innovations to facilitate the translation of exosomes-based therapies into clinically viable interventions for IVDD.

Gelatin, a natural macromolecular hydrophilic colloid, is a single-chain derivative obtained through the partial hydrolysis of collagen. , This hydrolysis preserves critical amino acid sequences, notably the Arg-Gly-Asp (RGD) motif. , Gelatin methacryloyl (GelMA) is synthesized by grafting methacryloyl groups onto the gelatin backbone. In the presence of a photoinitiator, GelMA undergoes photopolymerization to form a dynamic hydrogel network. GelMA is a top biomaterial in hydrogel research, offering a 3D environment that supports cell growth and differentiation, along with excellent biocompatibility and cellular responsiveness, making it a strong alternative to artificial basement membranes or natural collagen hydrogels. Its unique biological inertness preserves exosomes bioactivity, while the internal 3D porous architecture offers optimal spatial confinement for exosomes encapsulation. Under physiological conditions, GelMA undergoes enzyme-mediated degradation, enabling spatiotemporally controlled exosomes release. These attributes collectively position GelMA as an ideal platform for engineering exosomes delivery systems, balancing mechanical support, bioactivity preservation, and sustained cargo release for regenerative applications.

This study aims to counteract IVDD by engineering a GelMA hydrogel platform designed for the sustained release of BMSC-exos. Harnessing the regenerative potential of BMSC-exos for NP tissue, our team designed a hydrogel to deliver these bioactive substances in a controlled manner both spatially and temporally. When injected into a puncture-induced IVDD model, the Exo-GelMA hydrogel showed the ability to slow down degeneration and encourage NP regeneration. Our method targets mechanisms of disc degeneration, offering a promising biomimetic strategy to slow IVDD progression and aid natural disc repair.

Reagents

Rabbit polyclonal to Collagen II­(Col-II) , All-In-One RT Master Mix and Anti-CD63 antibody were purchased from Abcam (United Kingdom). Fetal Bovine Serum (Exosomes-Depleted) was purchased from Absin (China). 4′,6-Diamidino-2-phenylindole (DAPI), Crystal Violet Staining Solution, Western Blot Kit, Enhanced Bicinchoninic acid (BCA) Protein Assay Kit were purchased from Beyotime Biotechnology (China). FreeZol reagent kit, Hiscript II First Strand cDNA Synthesis kit, and SYBR qPCR Master Mix were purchased from Vazyme Biotech (China). HSP70 Antibody was purchased from CST (United States). Calcein/PI Cell Viability/Cytotoxicity Assay Kit was purchased from Dojindo (Japan). GelMA hydrogel was purchased from Suzhou Yongqinquan Intelligent Equipment Co., Ltd. DMEM/F12 and fetal bovine serum were purchased from Hyclone­(United States). Cell Plasma Membrane Red Fluorescence Staining Kit with PKH26 and Triton X-100 were purchased from Sigma-Aldrich (United States). Primers were purchased from Sangon Biotech­(United States). Exosomes Isolation Kit from Cell Culture Media by Precipitation Method was purchased from Thermo Fisher (United States). Type II Collagenase and TRITC Phalloidin were purchased from Yeasen (China).

In Vitro Experiments

Harvesting and Cultivation of Rat BMSCs

Rats aged 4 weeks, obtained from the Experimental Animal Center in Hangzhou Ziyuan, were anesthetized using an intraperitoneal injection of 3.5% chloral hydrate. The tibiae and femora from both back legs were isolated in a sterile manner and washed with phosphate-buffered saline (PBS). The medullary cavities were accessed by taking off the epiphyses, and bone marrow was collected by flushing the cavities with 5 mL of low-glucose DMEM containing 10% Fetal Bovine Serum (FBS). Once centrifuged at 1000 rpm for 15 min, the supernatant was poured off. After washing the cell pellet three times with PBS, it was resuspended in low-glucose DMEM with 10% FBS. The cells were then seeded into 25 cm2 culture flasks at a density of 1 × 104 cells/cm2 and incubated at 37 °C in a humidified atmosphere with 5% CO2 until they reached a confluence of ≥90%. Subsequently, BMSCs were co-cultured with NPCs in transwell plates for a period of 72 h.

Culturing and Isolating NPCs

Surgical removal of the rats’ caudal vertebrae was performed, followed by aseptic extraction of the NP tissues from the intervertebral discs (IVDs). The NP tissues were subsequently minced into 1 mm3 fragments and subjected to enzymatic digestion at 37 °C for 4 h using a solution containing 0.2% Dispase II and 0.2% type II collagenase. Following digestion, the tissue suspension was gently pipetted 20 times and filtered through a 200 μm cell strainer. The resulting filtrate was collected and centrifuged at 300 × g for 5 min, after which the pelleted cells were cultured in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum and 1% penicillin–streptomycin. Following the seeding of the cells into culture flasks, incubation was conducted at 37 °C within a 5% CO2 atm. The culture medium was replenished every 2 to 3 days, and P1 NPCs were utilized for subsequent experiments.

RNA Isolation and Real-Time Polymerase Chain Reaction

Total RNA was extracted utilizing the TRIzol reagent, followed by reverse transcription of cDNA employing the Revert Aid First Strand cDNA Synthesis Kit. Quantitative reverse-transcription polymerase chain reaction (RT-qPCR) was conducted under the following thermal cycling conditions: an initial incubation at 50 °C for 2 min, followed by 2 min at 95 °C, and 40 amplification cycles consisting of 3 s at 95 °C and 32 s at 60 °C. The relative expression levels of the genes were calculated using the 2̂–ΔΔCt method. Primer sequences used are provided in Table S1.

Immunofluorescence

NPCs were seeded at a density of 1000 cells/cm2 in the lower chamber of Transwell inserts, while the hydrogel was positioned in the upper chamber. After a three-day culture period, the samples were fixed using 4% paraformaldehyde for 20 min. Following fixation, samples were incubated in a blocking and permeabilization solution containing 5% bovine serum albumin (BSA) and 0.1% Triton X-100 for 1 h. Subsequently, the anti-Collagen II primary antibody was introduced, and the samples were incubated overnight at 4 °C. Afterward, a Cy3-conjugated fluorescent secondary antibody was applied and incubated for 1 h in the dark. The samples were then stained with DAPI for 10 min, ensuring protection from light exposure. Finally, images were captured and analyzed using an inverted fluorescence microscope.

Cytotoxicity Assay

NPCs were seeded onto the hydrogel at a density of 3000 cells/cm2. To evaluate the effects of the composite hydrogel on NPC proliferation, CCK-8 assays were performed on days 1, 3, and 5 of the culture. Specifically, the CCK-8 solution was diluted with the culture medium at a ratio of 1:9, transferred to a 96-well plate, and incubated for 2 h in a cell culture incubator at 37 °C. Finally, the optical density (OD) values were measured to assess cell viability and proliferation.

Exosomes Isolation and Characterization

BMSCs at the second passage were cultured for 48 h in an exosomes-depleted medium supplemented with 10% fetal bovine serum. Briefly, two centrifugation steps (300 × g, 5 min, 2–8 °C; 2000 × g, 30 min, 2–8 °C) were performed on conditioned medium before exosomes

Pre-enrichment by precipitation. The Invitrogen Total Exosomes Isolation Kit was used for precipitation of exosomes. Enriched exosomes were resuspended in PBS. For protein analysis, exosomesomal proteins were extracted using a commercial protein extraction kit, separated via SDS-PAGE, and subsequently transferred onto PVDF membranes. The membranes were blocked for 1 h at room temperature and then incubated overnight at 4 °C with primary antibodies targeting CD63, TSG101, HSP70, and Calnexin. This was followed by a 1 h incubation with appropriate HRP-conjugated secondary antibodies. After three washes with TBST, protein bands were visualized using chemiluminescent detection methods.

Fabrication of Exo-GelMA Hydrogel

A commercial GelMA hydrogel was utilized as a carrier for exosomes to develop an Exo-loaded composite hydrogel. Initially, a 5% (w/v) GelMA hydrogel solution was prepared and subsequently filtered through a 0.22 μm sterile filter to yield a sterile GelMA precursor solution. Following this, 200 μg of purified exosomes were combined with 50 μL of the sterile solution containing 5% (w/v) GelMA and 0.05% (w/v) lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP). The resulting mixture underwent photo-crosslinking for 30 s under 405 nm light, thereby forming the Exo–GelMA hydrogel system.

Assessment of In Vitro Exosomes Release Kinetics

The exosomes release profile from the Exo-GelMA hydrogel was evaluated under simulated physiological conditions. Briefly, the prepared Exo-GelMA hydrogel samples were immersed in phosphate-buffered saline (PBS, pH 7.4) and incubated at 37 °C in a thermostatic shaking incubator with a continuous rotation speed of 75 rpm. At predetermined intervals (days 1, 3, 7, and 14), the supernatant was collected. The cumulative release of exosomes was quantified using a BCA protein assay kit. All experiments were performed in triplicate (n = 3) to ensure statistical reliability.

Viewing through Scanning Electron Microscopy

The morphology of the composite hydrogel was examined using scanning electron microscopy (SEM) following the freeze-drying of the prepared Exo-GelMA hydrogel in a lyophilizer. Subsequent to embrittlement by liquid nitrogen, the composite hydrogel was affixed to a metal sample stage using conductive adhesive. It was then subjected to an ion sputtering apparatus for a 30 s gold coating. Ultimately, the surface structure of the sample was analyzed via SEM.

Crystal Violet Staining

The hydrogel was positioned in the lower chamber of a Transwell plate, while 5000 NPCs were seeded in the upper chamber. The cells were maintained in a serum-free medium for a duration of 6 h, after which crystal violet staining was performed. The detailed protocol is as follows: the culture medium was removed, and the cells were rinsed with PBS. Subsequently, the cells were treated with 4% paraformaldehyde (PFA) and stained with crystal violet at room temperature, avoiding light exposure, for 30 min. After the staining solution was removed, images were captured using microscopy for additional analysis.

Uptake of Exosomes by NPCs

Exosomes were labeled with PKH26 for 5 min at room temperature. Following the labeling process, the exosomes were washed with PBS and subjected to centrifugation. The exosomes were then resuspended in a basal medium and incubated with NPCs for 24 h at 37 °C. Subsequently, the cells were then fixed with 4% PFA for 20 min and counterstained with DAPI. Fluorescence microscopy was employed to capture images of the cells.

Wound-Healing Assay

NPCs were planted at a concentration of 5000 cells per square centimeter in the bottom chamber of a Transwell plate. After the cells adhered, a straight scratch was made with a sterile 1 mL pipette tip. Cells that had detached were then washed away using PBS, and the hydrogel was placed in the upper chamber. The culture medium was then replaced with a 1% FBS medium and incubated for 12 h, after which microscopic observation and image capture were conducted.

In Vivo Experiments

Construction of the Model for Degeneration of Intervertebral Discs

A total of 40 6 week-old female Sprague-Dawley rats were randomly allocated into four experimental groups (n = 10 per group): sham operation (Sham), simple injury (SI), GelMA, and GelMA-Exo. Using 2% pentobarbital sodium for intraperitoneal anesthesia, the skin, fascia, and muscle layers were cut in sequence to reveal the Co7-8 intervertebral disc. Degeneration was induced using a 21G needle, which was inserted into the center of the disc, rotated for 5 s, and maintained in position for an additional 30 s to ensure adequate trauma. Following this, 10 μL of GelMA, free exosomes, or a GelMA-Exo solution was injected into the respective discs and crosslinked with ultraviolet light for 1 min. After the surgical procedure, the incisions were sutured and disinfected, and the rats were housed individually with ad libitum access to food and water. All rats were euthanized at 4 and 8 weeks post-surgery, and any rats that died during the modeling process were replaced to ensure that each group maintained a total of 10 rats.

Radiological Evaluation

At 4 and 8 weeks post-operation, five rats were randomly selected from each group for magnetic resonance imaging (MRI). Following administration of intraperitoneal anesthesia, the rats were positioned supine on a custom-designed restraint board, with their tails secured in linear alignment. MRI scanning was conducted, focusing on the Co7-8 intervertebral level. Subsequent to imaging, the rats were euthanized, and their spinal columns were harvested. The specimens were then subjected to fixation and decalcification, sectioned sagittally along the midline of the intervertebral discs, and photographed for gross examination.

Histological Examination

Subsequent to gross morphological photography, the intervertebral discs were rehydrated through a series of graded ethanol washes. This was followed by histological evaluation utilizing hematoxylin and eosin (H & E) staining, safranin O-fast green (S.O.) staining, and immunohistofluorescence staining to assess the extent of disc repair.

Statistical Analysis

The data are shown as the mean ± standard deviation from at least three experiments. GraphPad Prism 7.0 was used for statistical analysis. For comparing two groups, a Student’s t-test was applied, while one-way ANOVA was used for multiple group comparisons. Statistical significance was indicated by a p-value under 0.05.

Results

Evaluation of Exosomes Produced by BMSCs

Primary nucleus pulposus cells (NPCs) were identified via morphology and phenotypic markers. Specifically, NPCs were characterized using Hematoxylin–Eosin (HE) staining, Toluidine blue staining for GAG secretion, and immunofluorescence staining for Col-II (Supporting Information Figure S1). According to SEM images, the exosomes appeared spherical with a teardrop-like configuration (Figure A). Nanoparticle tracking analysis (NTA) indicated that the diameters of the exosomes were predominantly distributed within the range of 50–100 nm (Figure B). Western blot assays demonstrated that the exosomesomal markers CD63, TSG101 and HSP70 were expressed, while the endoplasmic reticulum protein Calnexin was not detected (Figure C). To assess cellular uptake, exosomes were co-cultured with NPCs. The exosomes were marked with PKH26, and the co-culture that followed displayed PKH26 fluorescence in the cytoplasm of the surrounding NPCs (Figure D). The results suggest that NPCs can internalize exosomes originating from bone marrow stem cells.

1.

1

Open in a new tab

Characterization of Exosomes Derived from Bone Marrow Stem Cells. (A) Representative SEM images of exosomes; (n = 3, Scale bar = 500 nm). (B) Size distribution analysis of exosomes as determined by NTA; (n = 3). (C) WB illustrating the expression of exosomesomal biomarkers CD63, TSG101, HSP70, and Calnexin; (n = 3). (D) Uptake of PKH26-labeled exosomes by NPCs. (n = 3, Scale bar = 10 μm).

Characterization of Exo-GelMA Hydrogels

GelMA and Exo-GelMA hydrogels were synthesized through UV-induced crosslinking in the presence of a photoinitiator. Scanning electron microscopy (SEM) analysis demonstrated that both Exo-GelMA and control-GelMA hydrogels possessed a highly porous, interconnected network structure, with no significant modifications in the three-dimensional microarchitecture following exosomes incorporation (Figure A,B). To evaluate the release kinetics, exosomes were encapsulated within the GelMA hydrogel, and their sustained release was monitored over a period of 14 days. The composite hydrogel exhibited controlled exosomes release, achieving approximately 60% cumulative release by day 14 (Figure C), suggesting its potential for long-term therapeutic efficacy in both in vitro and in vivo applications.

2.

2

Open in a new tab

Characterization of Exo-GelMA hydrogels. (A,B) Representative SEM images of GelMA and Exo-GelMA hydrogels, respectively; (n = 3, Scale bar = 5 μm). (C) Cumulative release of Exo from the Exo-GelMA composite hydrogel over 14 days. (n = 3).

Cellular Activity on Exo-Infused 3D Hydrogel

NPCs were cultured on Exo-GelMA composite hydrogels to assess their biocompatibility and cellular responses. Immunofluorescence staining indicated that NPCs adhered and spread effectively on both GelMA and Exo-GelMA hydrogels, displaying characteristic morphological features (Figure A). Live/dead staining analysis revealed a high cell viability rate of over 90% (indicated by green staining for live cells) with minimal cell death (indicated by red staining) (Figure B,C). Additionally, CCK-8 assays confirmed robust NPCs proliferation, with a significant increase in metabolic activity over time (Figure D). Notably, NPCs cultured on Exo-GelMA hydrogels exhibited enhanced viability compared to those cultured on GelMA alone (p < 0.05), suggesting that the release of exosomes may play a beneficial role in maintaining cell function. Furthermore, Transwell migration and scratch wound assays demonstrated that Exo-GelMA hydrogels significantly enhanced NPC migration compared to control groups (Figure E,F), likely due to the bioactive cues released from the encapsulated exosomes.

3.

3

Open in a new tab

Biocompatibility of the Exo-GelMA composite hydrogel. (A) Cytoskeleton staining showing the morphology of NPCs cultured on the hydrogels (red: cytoskeleton; blue: nuclei); (n = 3, Scale bar = 50 μm). (B) Live/dead staining of NPCs on the hydrogels (red: dead cells; green: live cells). (n = 3, Scale bar = 10 μm). (C) Quantitative analysis of live cells from the live/dead staining. (n = 3). (D) Cell viability and proliferation of NPCs cultured on the hydrogels on days 1, 3, and 5, as determined by CCK-8 assay. (n = 3). *p < 0.05. (E,F) Transwell migration assay and scratch wound healing assay, respectively. (n = 3).

Impact of Exo-Loaded 3D Hydrogel on Matrix Production in NPCs

The RT-qPCR analysis of cellular matrix-related gene expression, including Sox-9, Aggrecan (ACAN), and Col-II, revealed that the exo-GelMA composite significantly enhanced the expression levels of Sox-9, ACAN, and Col-II in NPCs compared to the GelMA group (Figure A). Additionally, cellular immunofluorescence staining demonstrated an increased secretion of Col-II by NPCs within the exo-GelMA composite hydrogel group (Figure B,C), which may be attributed to the release of exosomes.

4.

4

Open in a new tab

Effect of the Exo-GelMA composite hydrogel on extracellular matrix (ECM) synthesis in NPCs. (A) Expression of ECM synthesis-related genes in NPCs cultured on the hydrogels, as detected by RT-qPCR. (n = 3). *p < 0.05. (B) Immunofluorescence staining showing Col-II expression levels in NPCs cultured on the hydrogels; (n = 3, Scale bar = 20 μm). (C) Quantitative analysis of Col-II expression in NPCs. (n = 3).*p < 0.05.

Visual and Magnetic Resonance Imaging Assessment of Rat Tail Intervertebral Discs

Gross morphological examination of the tissue samples indicated that the nucleus pulposus region of the intervertebral disc in the GelMA group exhibited a progressive decrease and eventual disappearance, being supplanted by fibroblastic-like tissue at both 4 and 8 weeks post-surgery. In the exosomes group, the nucleus pulposus region showed an increase; however, this was significantly lower than that observed in the exo-GelMA group. At 4 weeks, the size and morphology of the medullary region in the exo-GelMA group closely resembled those in the Sham group, with a clear demarcation between the medullary tissue and the fibrous ring tissue. By 8 weeks, although the nucleus pulposus area in the GelMA-Exo group had decreased, it remained the largest among the three groups (Figure A). Furthermore, MRI assessment of the disc nucleus pulposus region using T2-weighted imaging revealed a significant reduction in the nucleus pulposus signal in the GelMA group compared to the Sham group. Conversely, the intervertebral disc nucleus pulposus signal in the exosomes group exhibited a slight decrease at 4 weeks, followed by a significant reduction in the nucleus pulposus signal area by 8 weeks. Notably, the MRI T2-weighted signal intensity of the exo-GelMA group at 4 weeks was comparable to that of the Sham group, exhibiting elevated nucleus pulposus signals, which persisted until the 8 week mark (Figure B–D). This finding suggests that the sustained release of exosomes from GelMA-exosomes may significantly mitigate IVDD following acupuncture treatment.

5.

5

Open in a new tab

Evaluation of rat coccygeal intervertebral discs by gross appearance and magnetic resonance imaging (MRI). (A)­Gross appearance images of the experimental groups. (n = 5). (B) MRI images of the experimental groups. (C,D) Pfirrmann grading scores for the groups after 4 and 8 weeks, respectively. (n = 5).*p < 0.05.

Histological Evaluation

Histological analyses utilizing Hematoxylin and Eosin (H & E) and Safranin O (S.O.) staining revealed distinct morphological characteristics among the experimental groups. In the Sham group, the nucleus pulposus exhibited a central, oval morphology, while the annulus fibrosus was organized in well-defined concentric layers, with clearly discernible collagen fibers and distinct boundaries separating the annulus fibrosus from the nucleus pulposus. In contrast, the intervertebral disc tissue of the GelMA group at 4 and 8 weeks demonstrated a progressive reduction in the area of the nucleus pulposus, which was increasingly replaced by fibrous tissue exhibiting a disordered arrangement. The exosomes group retained a greater amount of nucleus pulposus tissue, with a more distinct demarcation between the nucleus pulposus and the annulus fibrosus. Notably, the exo-GelMA group exhibited the highest preservation of nucleus pulposus tissue, characterized by a clear delineation from the annulus fibrosus (Figure ).

6.

6

Open in a new tab

Histological assessment of intervertebral disc tissues. (A) H & E staining showing disc morphology in the experimental groups at 4 and 8 weeks postoperatively; Scale bar = 1 mm. (n = 5, Scale bar = 1 mm). (B) Safranin O (S.O.) staining showing disc morphology in the experimental groups at 4 and 8 weeks postoperatively; (n = 5, Scale bar = 1 mm).

Matrix Expression in Intervertebral Disc Tissue

The immunofluorescence staining results for detecting Col-II expression in disc tissues across different groups revealed that Col-II expression was high in normal discs but reduced in the GelMA group. Notably, discs treated with exosomes and GelMA-Exo exhibited increased Col-II expression, with the GelMA-Exo treatment demonstrating a significantly enhanced effect compared to exosomes alone. At the 4 week mark, Col-II levels in the exosomes group surpassed those in the GelMA group; however, by 8 weeks, this difference was no longer statistically significant (Figure ). These findings suggest that the GelMA hydrogel effectively sustained matrix synthesis by modulating the release of exosomes.

7.

7

Open in a new tab

Expression of Col-II in the ECM of intervertebral disc tissues. (A) Representative images of Col-II expression in the experimental groups at 4 weeks postoperatively. (n = 5, Scale bar = 50 μm). (B) Quantitative analysis of Col-II expression at 4 weeks. (n = 5). *p < 0.05. (C) Representative images of Col-II expression in the experimental groups at 8 weeks postoperatively. (n = 5, Scale bar = 50 μm). (D) Quantitative analysis of Col-II expression at 8 weeks. (n = 5). *p < 0.05.

Discussion

IVDD poses a major socioeconomic issue due to its commonness and the chronic pain and disability it causes. Current treatments, including surgery and conservative methods, often fail to address the root causes of disc degeneration, resulting in limited success and recurring symptoms. Full disc regeneration necessitates restoring both the disc’s biomechanical properties and the biological functions of its cells.

BMSCs are pivotal in regenerative therapies for IVDD due to their capacity to downregulate inflammatory mediators and matrix-degrading enzymes. , Current research suggests that these therapeutic effects are predominantly facilitated by exosomes, which are bilayer lipid vesicles with diameters ranging from 50 to 200 nm. − Through the delivery of diverse molecular cargo, exosomes precisely regulate critical biological processes in NPCs, including proliferation, autophagy, and apoptosis.

Our study demonstrates that BMSC-exos enhance the viability of rat NPCs and promote their proliferation, migration, and ECM synthesis. Furthermore, intradiscal administration of these exosomes partially preserved disc structure and hydration in a rat model of IVDD, thereby mitigating disease progression. However, this therapeutic effect diminished significantly over time. This attenuation is likely attributable to limitations inherent in bolus exosomes injection, specifically rapid clearance and a short half-life, which prevent the maintenance of sustained, therapeutically effective local concentrations. As a result, ensuring that delivery methods keep exosomes levels high in the degenerated disc is key to maximizing the therapeutic advantages of exosomes based IVDD treatments.

Tissue engineering utilizing hydrogel-encapsulated BMSC-exos is a potent strategy for IVDD, as it prevents rapid exosomes clearance while leveraging matrix-exosomes synergy. This platform has demonstrated therapeutic efficacy across various tissues, including skin repair and cartilage regeneration via photo-induced imine hydrogels. Furthermore, injectable alginate systems have been developed for bone repair, while advanced innovations such as functionalized ECM scaffolds and self-healing chitosan-based hydrogels effectively delay IVDD progression by modulating disc metabolism, inhibiting pyroptosis, and suppressing NPC senescence. ,

GelMA has gained prominence as a versatile biomimetic scaffold in various tissue engineering applications, including the regeneration of skin, bone, and cartilage, owing to its tunable mechanical properties and compatibility with advanced microfabrication techniques. − Although GelMA concentrations generally range from 1% to 30% (w/v), we have deliberately chosen a 5% (w/v) formulation to optimize injectability for the deep-seated intervertebral disc environment, where maintaining fluidity during delivery is crucial for effective therapeutic outcomes. − This biomimetic matrix effectively mimics the native ECM through its inherent cell-attachment sites and matrix metalloproteinase (MMP)-responsive motifs, thereby facilitating critical NPC functions such as adhesion, proliferation, and migration. By incorporating BMSC-exos into this photo-cross-linkable matrix, our platform modulates the inflammatory microenvironment and prevents NPC degeneration through sustained exosomesomal release, thereby balancing mechanical protection with targeted biological signals. −

Material characterization validated the successful synthesis of GelMA hydrogel, which exhibits the properties of a stable viscoelastic solid with an elastic modulus that aligns with the optimal range for in vivo applications. The hydrogel effectively accommodates the loading of exosomes onto its matrix. Notably, GelMA hydrogel closely resembles the native ECM in several critical attributes, thereby demonstrating exceptional biocompatibility and positioning it as an ideal carrier for exosomes. Additionally, its porous structure and hydrophilic characteristics create a conducive microenvironment for cell adhesion.

In terms of drug release kinetics, exosomes encapsulated within GelMA hydrogel demonstrated a sustained release profile over a 14 day period, achieving a cumulative release rate exceeding 60%. This extended release characteristic facilitates optimal bioactivity delivery, thereby enhancing therapeutic efficacy. In contrast to bolus administration, the sustained-release kinetics of GelMA hydrogel prolong the bioactivity of exosomes within the target microenvironment. Co-culture experiments involving NPCs and the test samples indicated a significant enhancement in cell viability on the hydrogels, with progressive increases observed throughout the culture duration, suggesting a robust proliferative capacity. Importantly, NPCs cultured on the Exo-GelMA composite hydrogel exhibited significantly higher viability compared to those cultured on GelMA hydrogel alone. Both Transwell migration and scratch wound assays corroborated the ability of the Exo-GelMA composite to promote NPC migration. These findings provide compelling evidence that GelMA-encapsulated exosomes retain their intrinsic bioactivity and facilitate sustained delivery into the microenvironment, thereby exerting long-term therapeutic effects on target cells.

A significant finding of our study is the elucidation of the molecular pathways through which the Exo-GelMA system mitigates IVDD. Our results indicate that the upregulation of Sox-9, Aggrecan (ACAN), and Col-II is not merely a descriptive occurrence but is likely facilitated by the suppression of the pro-inflammatory NF-κB signaling pathway. Recent literature corroborates that BMSC-exos cargo can specifically inhibit the nuclear translocation of the p65 subunit, thereby reducing the expression of catabolic enzymes such as MMP3 and ADAMTS5a , Through sustained inhibition of this pathway, our hydrogel system establishes a localized anti-inflammatory “niche” that stabilizes the ECM against the adverse disc microenvironment.

To assess the in vivo function of Exo-GelMA hydrogel, we developed a rat model of IVDD induced through caudal acupuncture. Previous research has demonstrated that puncturing with needles of 21-gauge or larger results in substantial mechanical damage, which accelerates disc degeneration within a 4 week period, with the progression of degeneration stabilizing between 4 and 8 weeks. As a result, these two time points were chosen for observation. In vivo studies indicated that the intradiscal administration of Exo-GelMA promoted histological restoration of the nucleus pulposus by the fourth week. By the eighth week, the hydrogel-based delivery system exhibited significantly greater efficacy compared to free exosomes alone, as demonstrated by improved maintenance of disc integrity and consistently higher proteoglycan content.

The enhanced long-term therapeutic outcomes observed in the Exo-GelMA group can be attributed to the intricate “mechano-biological” synergy inherent in our delivery system. Building on our previous work with biomimetic scaffolds and controlled-release kinetics for disc repair, , we employed the 5% GelMA platform as an advanced bio-niche to facilitate the targeted and sustained release of BMSC-exos. This localized delivery effectively activates the Nrf2/HO-1 antioxidant pathway within NPCs. Given that oxidative stress contributes to both cellular apoptosis and matrix degradation, the continuous release of exosomesomal miR-142-3p-a key regulator identified in our prior molecular axis studies-plays a crucial role in reestablishing intracellular redox balance. Specifically, this exosomesomal component enhances the scavenging of reactive oxygen species and stabilizes mitochondrial membrane potential, thereby preventing the mitochondria-mediated apoptotic cascade. By concurrently mitigating inflammation and oxidative stress, this system suppresses the senescence-associated secretory phenotype (SASP), establishing a robust, multi-faceted defense that significantly postpones degenerative progression, as validated by our longitudinal in vivo MRI and histological assessments.

Our study developed a GelMA hydrogel-based system for sustained release of BMSC-exos to treat IVDD. The system mimics the native ECM, supports NPCs, and allows controlled exosomes release, enhancing cell proliferation, migration, and ECM synthesis, thereby reducing IVDD progression in vitro and in vivo. The GelMA hydrogel enabled sustained exosomes release for over 14 days, explaining the superior long-term efficacy at week 8 in the Exo-GelMA group compared to free exosomes. This improved outcome is likely due to the 5% GelMA formulation, which maintained injectability and formed a porous structure that balanced mechanical protection with efficient nutrient and exosomes delivery. The primary importance of this study is the development of an exosomes-encapsulated GelMA hydrogel aimed at improving exosomes retention and facilitating sustained release for the treatment of IVDD. This approach signifies a strategic transition from the mere supplementation of active substances to the reconstruction of a regenerative microenvironment, balancing mechanical protection with the efficient delivery of bioactive cargo. Despite limitations like model translatability and long-term safety, this study offers strong proof-of-concept for next-generation exosomes-based therapies for IVDD. Future research should optimize hydrogel parameters and clarify how exosomesomal cargo, such as specific miRNAs, influences disc metabolism to advance clinical application.

This study possesses several limitations that merit discussion. Firstly, the interspecies differences in disc microenvironments between rodents and humans require validation using translationally relevant animal models, such as non-human primates or large mammals, to enhance clinical applicability. Secondly, the limited in vivo observation period restricts a comprehensive assessment of long-term safety and therapeutic durability. Extending the follow-up period to 6–12 months would provide robust evidence for evaluating sustained treatment efficacy and the potential mitigation of degeneration recurrence.

Supplementary Material

ao5c08053_si_001.pdf (128.8KB, pdf)

The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acsomega.5c08053.

  • qPCR primer sequences (Table S1) (PDF)

The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript. Yuwei Chen and Yanjun Yang contributed equally.

This study was supported by the people’s livelihood Science and Technology Plan of Suzhou Municipal Government (SS202090) and the grants from the Jiangsu Provincial Health Commission Research Project (Z2022082).

The authors declare no competing financial interest.

References

  1. Hoy D., March L., Brooks P., Blyth F., Woolf A., Bain C., Williams G., Smith E., Vos T., Barendregt J.. et al. The global burden of low back pain: estimates from the Global Burden of Disease 2010 study. Ann. Rheum Dis. 2014;73(6):968–974. doi: 10.1136/annrheumdis-2013-204428. [DOI] [PubMed] [Google Scholar]
  2. Kirnaz S., Capadona C., Wong T., Goldberg J. L., Medary B., Sommer F., McGrath L. B. Jr., Härtl R.. Fundamentals of Intervertebral Disc Degeneration. World Neurosurg. 2022;157:264–273. doi: 10.1016/j.wneu.2021.09.066. [DOI] [PubMed] [Google Scholar]
  3. Hussain I., Sloan S. R., Wipplinger C., Navarro-Ramirez R., Zubkov M., Kim E., Kirnaz S., Bonassar L. J., Härtl R.. Mesenchymal Stem Cell-Seeded High-Density Collagen Gel for Annular Repair: 6-Week Results From In Vivo Sheep Models. Neurosurgery. 2019;85(2):E350–E359. doi: 10.1093/neuros/nyy523. [DOI] [PubMed] [Google Scholar]
  4. Sloan S. R. Jr., Wipplinger C., Kirnaz S., Navarro-Ramirez R., Schmidt F., McCloskey D., Pannellini T., Schiavinato A., Härtl R., Bonassar L. J.. Combined nucleus pulposus augmentation and annulus fibrosus repair prevents acute intervertebral disc degeneration after discectomy. Sci. Transl Med. 2020;12(534):eaay2380. doi: 10.1126/scitranslmed.aay2380. [DOI] [PubMed] [Google Scholar]
  5. Ukeba D., Yamada K., Suyama T., Lebl D. R., Tsujimoto T., Nonoyama T., Sugino H., Iwasaki N., Watanabe M., Matsuzaki Y., Sudo H.. Combination of ultrapurified stem cells with an in situ-forming bioresorbable gel enhances intervertebral disc regeneration. EBioMedicine. 2022;76:103845. doi: 10.1016/j.ebiom.2022.103845. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bhat S., Kannan S., Kolkundkar U. K., Seetharam R. N.. Exosomes Therapy: A Promising Avenue for Treating Intervertebral Disc Degeneration. Tissue Eng. Regen Med. 2025;22(7):895–909. doi: 10.1007/s13770-025-00746-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Miyamoto T., Muneta T., Tabuchi T., Matsumoto K., Saito H., Tsuji K., Sekiya I.. Intradiscal transplantation of synovial mesenchymal stem cells prevents intervertebral disc degeneration through suppression of matrix metalloproteinase-related genes in nucleus pulposus cells in rabbits. Arthritis Res. Ther. 2010;12(6):R206. doi: 10.1186/ar3182. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Zhu G., Yang X., Peng C., Yu L., Hao Y.. Exosomesomal miR-532-5p from bone marrow mesenchymal stem cells reduce intervertebral disc degeneration by targeting RASSF5. Exp Cell Res. 2020;393(2):112109. doi: 10.1016/j.yexcr.2020.112109. [DOI] [PubMed] [Google Scholar]
  9. Wang F., Zhou H., Du Z., Chen X., Zhu F., Wang Z., Zhang Y., Lin L., Qian M., Zhang X.. et al. Cytoprotective effect of melatonin against hypoxia/serum deprivation-induced cell death of bone marrow mesenchymal stem cells in vitro. Eur. J. Pharmacol. 2015;748:157–165. doi: 10.1016/j.ejphar.2014.09.033. [DOI] [PubMed] [Google Scholar]
  10. Vadalà G., Sowa G., Hubert M., Gilbertson L. G., Denaro V., Kang J. D.. Mesenchymal stem cells injection in degenerated intervertebral disc: cell leakage may induce osteophyte formation. J. Tissue Eng. Regen Med. 2012;6(5):348–355. doi: 10.1002/term.433. [DOI] [PubMed] [Google Scholar]
  11. Zhang Z., Qin F., Feng Y., Zhang S., Xie C., Huang H., Sang C., Hu S., Jiao F., Jiang J., Qin Y.. Icariin regulates stem cell migration for endogenous repair of intervertebral disc degeneration by increasing the expression of chemotactic cytokines. BMC Complement Med. Ther. 2022;22(1):63. doi: 10.1186/s12906-022-03544-x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Colella F., Garcia J. P., Sorbona M., Lolli A., Antunes B., D’Atri D., Barré F. P. Y., Oieni J., Vainieri M. L., Zerrillo L.. et al. Drug delivery in intervertebral disc degeneration and osteoarthritis: Selecting the optimal platform for the delivery of disease-modifying agents. J. Control Release. 2020;328:985–999. doi: 10.1016/j.jconrel.2020.08.041. [DOI] [PubMed] [Google Scholar]
  13. Tam V., Rogers I., Chan D., Leung V. Y., Cheung K. M.. A comparison of intravenous and intradiscal delivery of multipotential stem cells on the healing of injured intervertebral disk. J. Orthop Res. 2014;32(6):819–825. doi: 10.1002/jor.22605. [DOI] [PubMed] [Google Scholar]
  14. Oehme D., Goldschlager T., Ghosh P., Rosenfeld J. V., Jenkin G.. Cell-Based Therapies Used to Treat Lumbar Degenerative Disc Disease: A Systematic Review of Animal Studies and Human Clinical Trials. Stem Cells Int. 2015;2015:946031. doi: 10.1155/2015/946031. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Schol J., Sakai D.. Cell therapy for intervertebral disc herniation and degenerative disc disease: clinical trials. Int. Orthop. 2019;43(4):1011–1025. doi: 10.1007/s00264-018-4223-1. [DOI] [PubMed] [Google Scholar]
  16. Jin R., Cui Y., Chen H., Zhang Z., Weng T., Xia S., Yu M., Zhang W., Shao J., Yang M.. et al. Three-dimensional bioprinting of a full-thickness functional skin model using acellular dermal matrix and gelatin methacrylamide bioink. Acta Biomater. 2021;131:248–261. doi: 10.1016/j.actbio.2021.07.012. [DOI] [PubMed] [Google Scholar]
  17. Kurian A. G., Singh R. K., Patel K. D., Lee J. H., Kim H. W.. Multifunctional GelMA platforms with nanomaterials for advanced tissue therapeutics. Bioact Mater. 2022;8:267–295. doi: 10.1016/j.bioactmat.2021.06.027. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Lv B., Lu L., Hu L., Cheng P., Hu Y., Xie X., Dai G., Mi B., Liu X., Liu G.. Recent advances in GelMA hydrogel transplantation for musculoskeletal disorders and related disease treatment. Theranostics. 2023;13(6):2015–2039. doi: 10.7150/thno.80615. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Sun Y., Leung V. Y., Cheung K. M.. Clinical trials of intervertebral disc regeneration: current status and future developments. Int. Orthop. 2019;43(4):1003–1010. doi: 10.1007/s00264-018-4245-8. [DOI] [PubMed] [Google Scholar]
  20. Serigano K., Sakai D., Hiyama A., Tamura F., Tanaka M., Mochida J.. Effect of cell number on mesenchymal stem cell transplantation in a canine disc degeneration model. J. Orthop Res. 2010;28(10):1267–1275. doi: 10.1002/jor.21147. [DOI] [PubMed] [Google Scholar]
  21. Rani S., Ryan A. E., Griffin M. D., Ritter T.. Mesenchymal Stem Cell-derived Extracellular Vesicles: Toward Cell-free Therapeutic Applications. Mol. Ther. 2015;23(5):812–823. doi: 10.1038/mt.2015.44. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Willms E., Cabañas C., Mäger I., Wood M. J. A., Vader P.. Extracellular Vesicle Heterogeneity: Subpopulations, Isolation Techniques, and Diverse Functions in Cancer Progression. Front Immunol. 2018;9:738. doi: 10.3389/fimmu.2018.00738. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Vaiasicca S., Melone G., James D. W., Quintela M., Xiao J., Yao S., Finnell R. H., Conlan R. S., Francis L. W., Corradetti B.. Transcriptomic analysis reveals the anti-cancer effect of gestational mesenchymal stem cell secretome. Stem Cells Transl Med. 2024;13(7):693–710. doi: 10.1093/stcltm/szae024. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Chang Y. H., Wu K. C., Harn H. J., Lin S. Z., Ding D. C.. Exosomes and Stem Cells in Degenerative Disease Diagnosis and Therapy. Cell Transplant. 2018;27(3):349–363. doi: 10.1177/0963689717723636. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Wangler S., Peroglio M., Menzel U., Benneker L. M., Haglund L., Sakai D., Alini M., Grad S.. Mesenchymal Stem Cell Homing Into Intervertebral Discs Enhances the Tie2-positive Progenitor Cell Population, Prevents Cell Death, and Induces a Proliferative Response. Spine. 2019;44(23):1613–1622. doi: 10.1097/BRS.0000000000003150. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Khayambashi P., Iyer J., Pillai S., Upadhyay A., Zhang Y., Tran S. D.. Hydrogel Encapsulation of Mesenchymal Stem Cells and Their Derived Exosomes for Tissue Engineering. Int. J. Mol. Sci. 2021;22(2):684. doi: 10.3390/ijms22020684. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Liu X., Yang Y., Li Y., Niu X., Zhao B., Wang Y., Bao C., Xie Z., Lin Q., Zhu L.. Integration of stem cell-derived exosomes with in situ hydrogel glue as a promising tissue patch for articular cartilage regeneration. Nanoscale. 2017;9(13):4430–4438. doi: 10.1039/C7NR00352H. [DOI] [PubMed] [Google Scholar]
  28. Yang S., Zhu B., Yin P., Zhao L., Wang Y., Fu Z., Dang R., Xu J., Zhang J., Wen N.. Integration of Human Umbilical Cord Mesenchymal Stem Cells-Derived Exosomes with Hydroxyapatite-Embedded Hyaluronic Acid-Alginate Hydrogel for Bone Regeneration. ACS Biomater. Sci. Eng. 2020;6(3):1590–1602. doi: 10.1021/acsbiomaterials.9b01363. [DOI] [PubMed] [Google Scholar]
  29. Zhang K., Zhao X., Chen X., Wei Y., Du W., Wang Y., Liu L., Zhao W., Han Z., Kong D.. et al. Enhanced Therapeutic Effects of Mesenchymal Stem Cell-Derived Exosomes with an Injectable Hydrogel for Hindlimb Ischemia Treatment. ACS Appl. Mater. Interfaces. 2018;10(36):30081–30091. doi: 10.1021/acsami.8b08449. [DOI] [PubMed] [Google Scholar]
  30. Guan M., Liu C., Zheng Q., Chu G., Wang H., Jin J., Wu H., Chen J., Huang Q., Deng Z., Wang Y.. Exosomes-laden injectable self-healing hydrogel based on quaternized chitosan and oxidized starch attenuates disc degeneration by suppressing nucleus pulposus senescence. Int. J. Biol. Macromol. 2023;232:123479. doi: 10.1016/j.ijbiomac.2023.123479. [DOI] [PubMed] [Google Scholar]
  31. Eke G., Mangir N., Hasirci N., MacNeil S., Hasirci V.. Development of a UV crosslinked biodegradable hydrogel containing adipose derived stem cells to promote vascularization for skin wounds and tissue engineering. Biomaterials. 2017;129:188–198. doi: 10.1016/j.biomaterials.2017.03.021. [DOI] [PubMed] [Google Scholar]
  32. Goto R., Nishida E., Kobayashi S., Aino M., Ohno T., Iwamura Y., Kikuchi T., Hayashi J. I., Yamamoto G., Asakura M., Mitani A.. Gelatin Methacryloyl-Riboflavin (GelMA-RF) Hydrogels for Bone Regeneration. Int. J. Mol. Sci. 2021;22(4):1635. doi: 10.3390/ijms22041635. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Jiang G., Li S., Yu K., He B., Hong J., Xu T., Meng J., Ye C., Chen Y., Shi Z.. et al. A 3D-printed PRP-GelMA hydrogel promotes osteochondral regeneration through M2 macrophage polarization in a rabbit model. Acta Biomater. 2021;128:150–162. doi: 10.1016/j.actbio.2021.04.010. [DOI] [PubMed] [Google Scholar]
  34. Zhao S., Fang J., Wang Y., Zhang Y., Zhou Y., Zhuo S.. Construction of three-dimensional mesoporous carbon nitride with high surface area for efficient visible-light-driven hydrogen evolution. J. Colloid Interface Sci. 2020;561:601–608. doi: 10.1016/j.jcis.2019.11.035. [DOI] [PubMed] [Google Scholar]
  35. Zhu S., Yu C., Liu N., Zhao M., Chen Z., Liu J., Li G., Huang H., Guo H., Sun T.. et al. Injectable conductive gelatin methacrylate / oxidized dextran hydrogel encapsulating umbilical cord mesenchymal stem cells for myocardial infarction treatment. Bioact. Mater. 2022;13:119–134. doi: 10.1016/j.bioactmat.2021.11.011. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Jia W., Gungor-Ozkerim P. S., Zhang Y. S., Yue K., Zhu K., Liu W., Pi Q., Byambaa B., Dokmeci M. R., Shin S. R., Khademhosseini A.. Direct 3D bioprinting of perfusable vascular constructs using a blend bioink. Biomaterials. 2016;106:58–68. doi: 10.1016/j.biomaterials.2016.07.038. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Xu W., Molino B. Z., Cheng F., Molino P. J., Yue Z., Su D., Wang X., Willför S., Xu C., Wallace G. G.. On Low-Concentration Inks Formulated by Nanocellulose Assisted with Gelatin Methacrylate (GelMA) for 3D Printing toward Wound Healing Application. ACS Appl. Mater. Interfaces. 2019;11(9):8838–8848. doi: 10.1021/acsami.8b21268. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Yin J., Yan M., Wang Y., Fu J., Suo H.. 3D Bioprinting of Low-Concentration Cell-Laden Gelatin Methacrylate (GelMA) Bioinks with a Two-Step Cross-linking Strategy. ACS Appl. Mater. Interfaces. 2018;10(8):6849–6857. doi: 10.1021/acsami.7b16059. [DOI] [PubMed] [Google Scholar]
  39. Boularaoui S., Shanti A., Lanotte M., Luo S., Bawazir S., Lee S., Christoforou N., Khan K. A., Stefanini C.. Nanocomposite Conductive Bioinks Based on Low-Concentration GelMA and MXene Nanosheets/Gold Nanoparticles Providing Enhanced Printability of Functional Skeletal Muscle Tissues. ACS Biomater Sci. Eng. 2021;7(12):5810–5822. doi: 10.1021/acsbiomaterials.1c01193. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Wang W., Zhu Y., Liu Y., Chen B., Li M., Yuan C., Wang P.. 3D bioprinting of DPSCs with GelMA hydrogel of various concentrations for bone regeneration. Tissue Cell. 2024;88:102418. doi: 10.1016/j.tice.2024.102418. [DOI] [PubMed] [Google Scholar]
  41. Leu Alexa R., Cucuruz A., Ghiţulică C. D., Voicu G., Stamat Balahura L. R., Dinescu S., Vlasceanu G. M., Stavarache C., Ianchis R., Iovu H., Costache M.. 3D Printable Composite Biomaterials Based on GelMA and Hydroxyapatite Powders Doped with Cerium Ions for Bone Tissue Regeneration. Int. J. Mol. Sci. 2022;23(3):1841. doi: 10.3390/ijms23031841. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Van den Steen P. E., Dubois B., Nelissen I., Rudd P. M., Dwek R. A., Opdenakker G.. Biochemistry and molecular biology of gelatinase B or matrix metalloproteinase-9 (MMP-9) Crit Rev. Biochem Mol. Biol. 2002;37(6):375–536. doi: 10.1080/10409230290771546. [DOI] [PubMed] [Google Scholar]
  43. Pramanik S., Alhomrani M., Alamri A. S., Alsanie W. F., Nainwal P., Kimothi V., Deepak A., Sargsyan A. S.. Unveiling the versatility of gelatin methacryloyl hydrogels: a comprehensive journey into biomedical applications. Biomed Mater. 2024;19(4):042008. doi: 10.1088/1748-605X/ad4df7. [DOI] [PubMed] [Google Scholar]
  44. Yue K., Trujillo-de Santiago G., Alvarez M. M., Tamayol A., Annabi N., Khademhosseini A.. Synthesis, properties, and biomedical applications of gelatin methacryloyl (GelMA) hydrogels. Biomaterials. 2015;73:254–271. doi: 10.1016/j.biomaterials.2015.08.045. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Mushtaq F., Raza Z. A., Batool S. R., Zahid M., Onder O. C., Rafique A., Nazeer M. A.. Preparation, properties, and applications of gelatin-based hydrogels (GHs) in the environmental, technological, and biomedical sectors. Int. J. Biol. Macromol. 2022;218:601–633. doi: 10.1016/j.ijbiomac.2022.07.168. [DOI] [PubMed] [Google Scholar]
  46. Li X., Li X., Hu J., Zhu Y., Pang Z., Chen M., Liu C., Zhao C., Luo L., Gao Y.. et al. Spheroid-Derived BMSC Exosomes Enhance Proliferation and Matrix Remodeling of Nucleus Pulposus Cell Clusters in Degenerative Disc Repair. Small. 2025;21(45):e07073. doi: 10.1002/smll.202507073. [DOI] [PubMed] [Google Scholar]
  47. Su K. K., Yu D. C., Cao X. F., Li P., Chang L., Yu X. L., Li Z. Q., Li M.. Bone Marrow Mesenchymal Stem Cell-Derived Exosomes Alleviate Nuclear Pulposus Cells Degeneration Through the miR-145a-5p/USP31/HIF-1α Signaling Pathway. Stem Cell Rev. Rep. 2024;20(8):2268–2282. doi: 10.1007/s12015-024-10781-9. [DOI] [PubMed] [Google Scholar]
  48. Hu M. H., Yang K. C., Chen Y. J., Sun Y. H., Lin F. H., Yang S. H.. Optimization of puncture injury to rat caudal disc for mimicking early degeneration of intervertebral disc. J Orthop Res. 2018;36(1):202–211. doi: 10.1002/jor.23628. [DOI] [PubMed] [Google Scholar]
  49. Zhu L., Yang Y., Yan Z., Zeng J., Weng F., Shi Y., Shen P., Liu L., Yang H.. Controlled Release of TGF-β3 for Effective Local Endogenous Repair in IDD Using Rat Model. Int. J. Nanomedicine. 2022;17:2079–2096. doi: 10.2147/IJN.S358396. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Zhu L., Shi Y., Liu L., Wang H., Shen P., Yang H.. Mesenchymal stem cells-derived exosomes ameliorate nucleus pulposus cells apoptosis via delivering miR-142-3p: therapeutic potential for intervertebral disc degenerative diseases. Cell Cycle. 2020;19(14):1727–1739. doi: 10.1080/15384101.2020.1769301. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Xu G., Zhu K., Chen Z., Li M., Zhang Z.. Exosomesomal miR-221-3p derived from bone marrow mesenchymal stem cells alleviates endoplasmic reticulum stress-mediated apoptosis in nucleus pulposus cells via the Nrf2/HO-1 signalling pathway. Int. J. Exp Pathol. 2025;106(6):e70008. doi: 10.1111/iep.70008. [DOI] [PMC free article] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

ao5c08053_si_001.pdf (128.8KB, pdf)

Articles from ACS Omega are provided here courtesy of American Chemical Society

RESOURCES