A comprehensive search was conducted across three English-language databases: PubMed, Scopus, and Web of Science. The search focused on identifying studies related to nano-hydrogel systems for osteochondral repair. Search terms included combinations of MeSH and free-text keywords: (“nanohydrogel” OR “nanogel” OR “nano-hydrogel scaffold” OR “nanoscale hydrogel” OR “nano-sized hydrogel” OR “nanocomposite hydrogel”) AND (“osteochondral repair” OR “cartilage regeneration” OR “cartilage repair” OR “osteochondral defect”). Filters were applied to include only English-language publications. A detailed list of search terms and strategies for each database is provided in Supplementary Table S1.

Additionally, reference lists of retrieved articles were manually reviewed to identify any further relevant studies. Two authors (AFA and LQ) independently screened titles and abstracts to assess eligibility based on the inclusion criteria. Full-text articles were further reviewed to exclude any duplicates or studies that did not meet the criteria (Figure 1). Discrepancies were resolved through discussion with a third reviewer (JH). The last update search was conducted on 29 September 2024.

This systematic review was performed to address the following focused question: “What is the current status of nano-hydrogel preparations in promoting osteochondral repair, specifically regarding their composition, preparation methods, mechanical properties, biocompatibility, and therapeutic efficacy?”

To ensure the inclusion of high-quality and relevant studies, specific eligibility criteria were established prior to the screening process. Studies were included if they were original research articles published in peer-reviewed journals, written in English, and focused on the preparation and application of nano-hydrogel systems specifically for osteochondral or cartilage repair. Eligible studies were required to provide sufficient detail on the hydrogel’s composition, crosslinking or functionalization strategies, and report at least one form of biological or functional evaluation, whether in vitro, ex vivo, or in vivo.

Studies were excluded if they were review articles, conference abstracts, dissertations, clinical case reports, editorials, or other forms of grey literature. Additionally, publications that did not focus on osteochondral repair, or those that lacked essential data on hydrogel characterization or biological performance, were omitted. There were no restrictions on publication year; however, only articles published in English were considered. These criteria were designed to ensure methodological rigor and relevance to the focused research question.

Titles and abstracts were screened by two independent reviewers (AFA and LQ), followed by full-text assessments for studies that met the inclusion criteria. Disagreements on study eligibility were resolved through consultation with a third reviewer (JH). The extracting data were following PICO (P: sources, I: interventions, C: control study, O: outcomes) standards.

The data extraction process focused on gathering information about general study characteristics, including nano-hydrogel composition, types of nanoparticles, preparation methods, crosslinking strategies, and controlled release mechanisms. It also covered mechanical and bioactivity properties, such as mechanical strength, degradation rates, biocompatibility, swelling ratios, and functionalization aspects. For in vitro studies, details on cell types, culture conditions, cell viability, and proliferation were collected. In vivo studies were evaluated based on animal models, group allocation, implantation techniques, histological assessments, and outcomes related to subchondral bone and cartilage regeneration, including immunohistochemical findings, inflammation, infection, and hydrogel degradation. Lastly, the extraction included identification of research limitations and recommendations for future studies, ensuring a comprehensive overview of each study’s approach and findings.

The methodological quality of the included studies was evaluated using a customized CAMARADES checklist, which I adapted to better assess the relevance of each study (Macleod et al., 2004). The adapted checklist incorporated 11 key criteria to assess study relevance: (1) publication in a peer-reviewed journal, (2) random allocation to treatment or control groups, (3) blinded outcome assessment, (4) Control of the temperature in the animal facilities, (5) use of appropriate controls, (6) adequate sample size, (7) clear description of the animal model, (8) adherence to animal welfare guidelines, (9) reproducibility and replication of findings, (10) thorough outcome reporting, and (11) disclosure of any potential conflicts of interest. Given the nature of the data, analysis was conducted descriptively, as the variability across studies precluded meta-analysis.

Following the removal of duplicates, a total of 1,126 unique publications were identified through database screening. Title and abstract screening narrowed these to 56 articles for full-text evaluation. After applying the inclusion criteria, 11 studies were excluded. Consequently, 41 studies were included in this systematic review (Figure 1). Of these, 34 studies employed both in vitro and in vivo methodologies, while seven were limited to in vitro experiments (Adedoyin et al., 2015; Castro et al., 2015; Kosik-Kozioł et al., 2019; Qin et al., 2020; Fan et al., 2021; Banihashemian et al., 2024; Brown et al., 2024). The assessment of bias showed a spectrum from low to high risk, and detailed findings on methodological quality are illustrated in Figures 2,3, .

The studies summarized in Table 1 highlight the structural and compositional diversity of nano-hydrogel systems used for osteochondral repair. These range from simple, single-phase injectable formulations to more complex preformed multilayered scaffolds—each engineered to address distinct mechanical and biological requirements. Scaffold configurations were stratified into single-phase, bilayered, trilayered, and gradient systems. Many bilayered and trilayered constructs were designed to emulate the zonal architecture of osteochondral tissue, allowing site-specific modulation of chondrogenesis and osteogenesis.

Integration of nanoparticles such as hydroxyapatite (HA), chitosan montmorillonite, silica, and polydopamine (PDA) has been shown to enhance the mechanical integrity, osteoconductivity, and cellular interactions of hydrogels (Shalumon et al., 2016; Gong et al., 2020; Korpayev et al., 2020; Sheng et al., 2022; Hu et al., 2024; Jin et al., 2024). For instance, a study by Cao et al. (2023) utilized Cu-based nanoparticles embedded in a silk fibroin (SF) matrix via enzymatic crosslinking to create a single-phase injectable hydrogel with antioxidative and immunomodulatory properties Similarly, preformed bilayer hydrogels composed of polyvinyl alcohol (PVA), biphasic calcium phosphate (BCP), and carbon nanotubes (CNTs) were fabricated through a freeze-thawing process to generate a gradient interface, mimicking native cartilage–bone transition zones (Lan et al., 2021). These examples illustrate how both formulation type and nanoparticle selection directly influence the functional performance of nano-hydrogels.

The choice of crosslinking strategy is another determinant of scaffold performance, affecting mechanical stability, degradation behavior, and cellular response. Studies included a wide array of crosslinking approaches, enzymatic, photo-initiated, thermal, chemical, ionic, and dual-crosslinking methods, each tailored to the specific polymer systems and application needs (Adedoyin et al., 2015; Xu et al., 2019; Zhang et al., 2022b; Cao et al., 2023; Wu et al., 2023; Chen et al., 2024). For instance, photo-crosslinking has been employed to allow spatially controlled gelation, ideal for constructing gradient or multi-layered hydrogels (Zhang et al., 2024). However as highlighted in multiple reports, optimization is needed to reduce cytotoxicity from residual initiators, which may impact cell viability and tissue integration (Berry et al., 2019; Hu et al., 2019; Tomal and Ortyl, 2020). In terms of polymer origin, systems were broadly classified as natural, synthetic, or hybrid. Natural polymers like chitosan, gelatin (GelMA), alginate, and hyaluronic acid offer favorable biocompatibility and degradation profiles. Synthetic polymers such as PEGDA, PVA, and PLGA provide enhanced mechanical tunability and process control. Hybrid systems, which combine the strengths of both natural and synthetic components, emerged as especially promising in balancing bioactivity with structural integrity, several trilayered and bilayered scaffolds utilized such combinations to achieve distinct zone-specific functions.

Moreover, the application of advanced fabrication methods such as 3D printing, electrospinning, microsphere sintering, and solvent casting enabled precise spatial organization of materials. These techniques facilitated the development of functionally graded scaffolds, often incorporating nano-hydroxyapatite (nHA) or exosome-loaded layers, to mimic the mechanical and biochemical gradients of native osteochondral tissue (Zhang et al., 2022b; Brown et al., 2024). Several preformed multilayered systems were constructed with dual or triple layers, each designed with distinct pore architectures, ion release kinetics, and biofunctional molecules to modulate regeneration in a zone-specific manner.

Collectively, the reviewed studies demonstrate how scaffold architecture (e.g., single-phase, bilayered, trilayered), formulation type (injectable vs. preformed), polymer composition (natural, synthetic, hybrid), nanoparticle inclusion, crosslinking strategy, and fabrication technique can be tailored in concert to engineer next-generation nano-hydrogels for osteochondral repair. This multi-dimensional classification, as summarized in Table 1, provides a comparative framework to inform rational scaffold design and translational scaffold development.

Mechanical properties are essential for nano-hydrogel systems, particularly for osteochondral repair, where the scaffold must withstand the mechanical stresses of both cartilage and subchondral bone environments. As observed in Table 2, studies report varied mechanical strengths, with compressive moduli ranging from 0.4 MPa (Mpa) to over 73 MPa depending on the hydrogel composition (Gong et al., 2020; Zhang et al., 2022b; Brown et al., 2024; Hu et al., 2024; Kang et al., 2024). For instance, polycaprolactone-hydroxyapatite (PCL-HA) scaffolds have demonstrated compressive moduli as high as 73 ± 1 MPa, while IL-4-loaded GelMA-PCL-HA composites exhibit lower values around 4.7 ± 0.6 MPa (Gong et al., 2020). These scaffold values are within the range of trabecular (cancellous) bone, which exhibits compressive moduli typically between 10 and 200 MPa, depending on site and density. In contrast, the modulus of natural cortical bone is substantially higher, with a longitudinal elastic modulus ranging from 17.2 to 23.2 GPa and a transverse modulus ranging from 10.8 to 13.9 GPa, as demonstrated through multiscale modeling validated by nanoindentation and ultrasound measurements (Hamed et al., 2010). These comparisons highlight the potential of HA-containing scaffolds to approximate native bone behavior in osteochondral repair applications, particularly when enhanced with structural reinforcements like hydroxyapatite.

Biomimetic designs incorporating GelMA and HA have shown promise in enhancing mechanical stability and bioactivity for bone regeneration applications. GelMA hydrogels, while beneficial for tissue engineering, lack sufficient mechanical strength and osteogenic factors (Wang et al., 2022a). Incorporating HA into GelMA hydrogels improves their mechanical properties, biocompatibility, and osteogenic potential (Suvarnapathaki et al., 2020). Mineralized HA nanofibers further enhance the mechanical and bone regenerative performances of GelMA composites (Wang et al., 2022a). GelMA-based biomaterials can be tailored to overcome challenges in bone tissue engineering, such as insufficient mechanical properties and uncontrolled degradation (Dong et al., 2019). Advanced designs combining GelMA with other materials, like methacrylated HA nanoparticles and l-arginine-based unsaturated poly (ester amide), can create periosteum-mimicking scaffolds with improved mechanical strength, tissue adhesion, and osteogenic-angiogenic coupling effects (Yang et al., 2021). Double-crosslinking and freeze-drying methods have also been widely applied, producing physically and chemically reinforced structures that retain mechanical properties under physiological conditions (Yan et al., 2015; Filová et al., 2020; Zheng et al., 2023).

Balancing degradation rates with tissue regeneration remains another core challenge. An ideal scaffold degrades gradually, transferring mechanical load to newly forming tissue to aid integration (Hu et al., 2022; Li et al., 2022; Banihashemian et al., 2024; Chen et al., 2024). Studies have shown that adjusting crosslinking density and introducing bioactive molecules can customize degradation profiles for specific applications (Radhakrishnan et al., 2018; Zhang et al., 2023a; Zhang et al., 2023b; Deng et al., 2024). For example, Chen et al. developed a trilayered hydrogel with varied degradation rates across layers to replicate the native tissue gradient from cartilage to bone, facilitating sustained cell infiltration and extracellular matrix formation (Chen et al., 2024). Recent research has focused on developing multilayered hydrogel scaffolds to mimic the zonal organization of native cartilage tissue. These scaffolds feature gradients in mechanical properties, extracellular matrix composition, and bioactive factors across layers to guide cell differentiation and tissue formation (Brady et al., 2017; Qiao et al., 2021). Furthermore, a study demonstrated that layer-specific biomaterial compositions could direct a single stem cell population into zone-specific chondrocytes, resulting in native-like cartilage with varying mechanical and biochemical properties (Nguyen et al., 2011). In addition, a study further showed that stiffness gradient hydrogels could induce zone-specific responses in both chondrocytes and mesenchymal stem cells, mimicking cartilage zonal organization (Zhu et al., 2018). These approaches offer promising strategies for engineering complex osteochondral tissues with spatially-varying properties that more closely resemble native tissue structure and function.

Future advancements will likely focus on refining crosslinking techniques, such as enzyme-catalyzed, thermal, and photo-crosslinking, to develop materials that meet both mechanical and degradation needs for effective tissue engineering.

Nano-hydrogel systems have consistently demonstrated excellent biocompatibility and functional characteristics, making them highly suitable for applications in tissue engineering, particularly in osteochondral regeneration. Studies have reported cell viability rates exceeding 90% and enhanced cell proliferation, supporting the potential of these materials to promote tissue growth and regeneration (Table 3). For example, a study showed that LiMn₂O₄ nanozyme-functionalized hydrogels effectively supported the proliferation of rat chondrocytes and bone marrow-derived mesenchymal stem cells (BMSCs), promoting cell adhesion and growth (Hu et al., 2024). In addition, in vitro studies have highlighted that nano-hydrogels, such as GH@LM + GA@HLM and Zn-AlgMA, significantly enhance the proliferation of both chondrocytes and BMSCs, while maintaining high levels of cell viability (Hu et al., 2024; Zhang et al., 2024). Similarly, functionalized scaffolds, including those with CK2.1/LL37 and SF-MMT, further promote the regenerative processes of BMSCs and chondrocytes, reinforcing the critical role of scaffold composition in optimizing cellular responses (Liu et al., 2021b; Sheng et al., 2022).

Nano-hydrogels mimicking the extracellular matrix (ECM) have emerged as promising scaffolds for tissue engineering and regenerative medicine. These biomimetic materials create a three-dimensional (3D) environment that closely resembles the native ECM’s nanoscale architecture (Geckil et al., 2010; Gough et al., 2012; Brown et al., 2024). By incorporating nanostructured components, such as nanofibers or nanosilicates, these hydrogels can actively modulate cellular responses, including attachment, proliferation, and differentiation (Wei and Ma, 2008). For instance, nanoengineered collagen-based hydrogels reinforced with disk-shaped nanosilicates have been shown to enhance osteogenic differentiation of human mesenchymal stem cells without the need for exogenous growth factors (Paul et al., 2016). These ECM-mimicking hydrogels not only provide structural support but also create a regulatory milieu that guides tissue formation and organization (Geckil et al., 2010). Furthermore, their biocompatibility and ability to induce regenerative processes make them promising candidates for various biomedical applications, including bone tissue engineering and in vitro disease modeling (Wei and Ma, 2008; Paul et al., 2016).

Furthermore, functionalization techniques are crucial for enhancing the bioactivity of hydrogels in osteochondral tissue engineering. By incorporating growth factors, bioactive molecules, and nanoparticles, these hydrogels can promote both osteogenesis and chondrogenesis. For example, research has shown that embedding polydopamine-encapsulated kartogenin (KGN) and calcium phosphate-encapsulated miRNA-26a within hydrogels effectively promotes regeneration in both cartilage and bone layers (Kang et al., 2024). Additionally, KGN has been grafted onto ultrasmall superparamagnetic iron-oxide nanoparticles, which are then integrated into hydrogels for cartilage repair while enhancing MRI contrast (Yang et al., 2019). Another study developed microscaffold-hydrogel composites containing KGN and peptides to accelerate osteochondral repair through endochondral ossification (Zhang et al., 2022a). Moreover, a versatile hydrogel system using click chemistry has been created to provide tissue-specific cues for either chondrogenesis or osteogenesis (You et al., 2018; Guo et al., 2020; Liu et al., 2021a; Li et al., 2023a). These approaches highlight the potential of functionalized hydrogels in addressing the complex requirements of osteochondral tissue regeneration.

Recent studies demonstrate the effectiveness of functionalized biomaterials in advancing osteochondral repair, primarily by supporting both osteogenic and chondrogenic differentiation. Composite hydrogels with anti-CD44-labeled microparticles have shown to significantly improve osteogenic regeneration in animal models of osteochondral defects (Filová et al., 2020). Likewise, bilayer scaffolds that guide stem cell differentiation spatially have been effective in directing cells into osteogenic and chondrogenic lineages, enhancing repair outcome (Kang et al., 2024; Lowen et al., 2024). Furthermore, microscaffold-hydrogel composites, incorporating bioactive modifications like RGD peptides, have demonstrated accelerated osteochondral repair through endochondral ossification, achieved by controlled delivery of bioactive molecules within the scaffold layers (Zhang et al., 2022a; Brown et al., 2024; Deng et al., 2024). Other studies reinforce these findings, with functionalized hydrogels designed for dual osteogenic and chondrogenic applications showing sustained, layer-specific release of growth factors and bioactive ions, thus promoting cell proliferation and tissue integration (Cao et al., 2023; Wu et al., 2023).

These findings underscore the potential of multi-functionalized nano-hydrogels in tissue engineering, with customizable layers enabling the spatially controlled release of bioactive agents that foster site-specific tissue regeneration. Such approaches pave the way for advanced therapies for osteochondral defects and other complex tissue engineering applications (Wu et al., 2023; Brown et al., 2024).

These findings suggest that nano-hydrogels are capable of providing a supportive 3D microenvironment that mimics the native ECM. However, achieving consistent differentiation and integration remains challenging, particularly when translating in vitro success to in vivo conditions. Variability in cell behavior across studies suggests that more standardized protocols are needed to optimize cell-scaffold interactions, ensuring predictable outcomes in clinical settings.

The in vivo studies summarized in Table 4 illustrate the promising efficacy of nano-hydrogels in promoting osteochondral repair, using diverse animal models such as rabbits, rats, and mice to assess the regenerative potential of these systems. Significant cartilage regeneration and subchondral bone repair were observed in a rabbit model using a bi-layered GelMA-PCL-HA scaffold, where histological analyses confirmed the formation of a smooth cartilage surface and well-integrated bone layer (Gong et al., 2020). Similarly, a bilayer hydrogel containing GH@LM + GA@HLM demonstrated notable regeneration, with micro-CT and histological assessments indicating smooth hyaline cartilage formation and robust subchondral bone repair (Hu et al., 2024) (Table 4). These advanced hydrogel systems have demonstrated improvements in defect filling, cartilage thickness, and bone regeneration compared to control groups (Gan et al., 2019; Guo et al., 2021). However, a critical review of in vivo cartilage repair studies highlights the need for standardized experimental designs and careful interpretation of results (Vilela et al., 2015).

Histological assessments across various studies frequently highlighted improved tissue integration. A GTU-Fe/KGN@PDA/miRNA@CaP scaffold led to enhanced chondrogenic and osteogenic marker expression, indicating successful differentiation and maturation of regenerated tissue, with elevated glycosaminoglycans (GAG) and collagen deposition contributing to effective cartilage and bone regeneration (Table 4) (Kang et al., 2024). Further corroborating these findings, a Zn-AlgMA@Mg scaffold achieved significant osteochondral integration, facilitating seamless cartilage repair and trabecular bone formation within femoral condyle defects in rabbits (Zhang et al., 2024) (Table 4). Despite these advancements, scaffold-cartilage integration remains a significant challenge in tissue engineering. Recent strategies to address this issue include manipulating cellular, material, and biomolecular composition of engineered tissue (Jelodari et al., 2022). These findings highlight the potential for improved cartilage repair and integration using advanced scaffolds and tissue engineering techniques.

Many studies achieved substantial subchondral bone regeneration, suggesting that functionalization strategies including the incorporation of miRNAs, bioactive molecules, and structurally adaptive hydrogels play a crucial role in promoting dual regeneration for osteochondral repair. For example, bi-layer hydrogels and trilayered scaffolds demonstrated enhanced bone volume and trabecular thickness, ultimately supporting comprehensive osteochondral regeneration (Lan et al., 2021; Chen et al., 2024). Moreover, these studies predominantly used femoral condyle defect models, effectively showing that nano-hydrogels, when tailored to recreate the native extracellular environment, support robust tissue regeneration over extended periods. Functionalization strategies, such as incorporating tissue-specific peptides or drugs, have shown enhanced chondrogenesis and osteogenesis both in vitro and in vivo (Guo et al., 2021; Chen et al., 2024). These advanced scaffolds have demonstrated improved bone volume, trabecular thickness, and overall defect filling in femoral condyle defect models, supporting comprehensive osteochondral regeneration (Cao et al., 2024; Chen et al., 2024; Guo et al., 2024).

The variability in regenerative outcomes observed across studies, characterized by differing degrees of bone density and cartilage smoothness, highlights the necessity for a standardized approach to evaluating scaffold performance. Future research should focus on adopting consistent animal models, such as femoral defect models, and harmonized assessment criteria, such as specific histological markers and imaging techniques, to enable comparative evaluations across various hydrogel systems. Such standardization could accelerate the translation of nano-hydrogel-based technologies into clinical settings, supporting more predictable outcomes and broader applicability.

Recent advances in osteochondral tissue engineering have focused on developing scaffolds that support cell growth and tissue regeneration. Scaffold degradation plays a crucial role in the repair process, with different degradation modalities and speeds influencing outcomes (Tortorici et al., 2022). Despite considerable advances in osteochondral repair, several critical limitations remain across studies, as outlined in Table 5. One major challenge involves inconsistent degradation rates in scaffold materials. Achieving a uniform degradation timeline has proven difficult, with some hydrogel systems degrading faster than intended, reducing structural support for newly forming tissue, while others degrade too slowly, limiting cell infiltration and impeding tissue remodeling. For instance, study conducted by Adedoyin et al. noted this inconsistency in their dual-gelation scaffold, where uneven degradation impacted overall regenerative outcomes (Adedoyin et al., 2015). To address this, further research should investigate advanced crosslinking techniques to fine-tune degradation kinetics, ensuring scaffold resorption aligns more closely with native tissue growth.

Another prevalent issue is the variability in scaffold mechanical strength, particularly when scaling up for larger defects. Achieving a mechanical resilience that closely mimics native tissue properties remains challenging. Li et al. reported that preserving compressive strength in bilayer scaffolds was difficult over long-term in vivo applications, highlighting a critical need for more durable biomaterials (Li et al., 2023a). Novel scaffold compositions and innovative crosslinked structures could offer the increased load-bearing capacities necessary to provide robust support in osteochondral applications, particularly those involving weight-bearing joints.

Additionally, there is limited long-term in vivo data on the efficacy and safety of these scaffolds. While short-term successes are frequently observed, the potential for chronic inflammation or complications related to scaffold degradation requires longer follow-up. Studies highlight the necessity for prolonged trials to thoroughly assess scaffold stability, biocompatibility, and integration with native tissue structures, all critical for achieving successful clinical translation (Brown et al., 2024; Hu et al., 2024).

To overcome these challenges, future research could focus on innovative materials and scaffold designs. The use of in situ forming hydrogels, which adapt to irregular defect sites during implantation, may enhance scaffold integration (Zheng et al., 2014; Park and Park, 2018; Kang et al., 2024). Smart, stimuli-responsive hydrogels capable of controlled therapeutic release could also support sustained regeneration and more effective clinical outcomes. Additionally, combining nano-hydrogels with synergistic regenerative approaches such as gene therapy, bioelectronics, or cell-based treatments may lead to multifunctional scaffolds that facilitate not only osteogenesis and chondrogenesis but also angiogenesis (Kumar et al., 2022; Chen et al., 2023). Together, these integrated approaches have the potential to advance osteochondral repair, bringing the field closer to scalable, reliable therapeutic solutions.