DPSCs are MSCs collected from the pulpal tissue of permanent teeth, often impacted wisdom teeth or teeth taken for orthodontic procedures. They are thought to be readily obtainable sources of MSCs, but they are associated with the significant drawback of compromising the tooth`s vitality in order to collect the pulpal tissues. These cells exhibit the classic MSCs features, including multi-potency, rapid proliferation, and immune-modulating activities (85). Moreover, DPSCs can develop into endothelial cells, as well as their angiogenic potential (86). DPSCs demonstrated mineralization capacity and osteogenesis (87). Due to their limited ability to generate cementum, the therapeutic value of DPSCs for PDL reconstruction can be considered doubtful (88).

DPSCs can be effectively isolated from both periodontally healthy (hDPSCs) and periodontally compromised teeth (pDPSCs). Both of them have no morphological variations in early passage cells. Cryopreservation can alter the shape of pDSPCs. Early passage cells exhibited no substantial change in the favorable transcription of MSCs' biomarkers CD73, CD90, and CD105. Nevertheless, repeated passaging and cryopreservation influenced biomarker transcription in pDPSCs. Both cell types show modest transcription of the hematopoietic b, such as CD34, CD45, and the MHC class II antigen HLA-DR. PDPSCs express more HLA-DR than hDPSCs. pDPSCs exhibit much slower growth rates and wound healing characteristics than hDPSCs. The migration capacity of pDPSCs can be significantly boosted during late passage following cryopreservation. There is no discernible change in osteogenic capability between them. Yet, pDPSCs have much poorer chondrogenic capacity than hDPSCs. However, pDPSCs demonstrated increased osteogenesis and chondrogenesis at late passage and following cryopreservation (89).

SHEDs offer an exceptional, non-invasive source of MSCs. Because of their ease of separation, multipotential differentiating capability, and low antigenicity, they may be a viable choice for periodontal regeneration. SHEDs are conveniently accessible via noninvasive techniques since they are extracted from deciduous exfoliated teeth (90). SHEDs have an elevated level of multiplication and immune-modulating capabilities, comparable to BMMSCs, which are more challenging to get. SHEDs, like DPSCs, are derived from dental pulp (17, 91). Yet, SHEDs express greater quantities of stemness-related genes than DPSCs, preserving greater flexibility during in vitro passaging (92). SHEDS are robustly proliferating and can transform into several cellular types, notably osteoblasts and adipocytes (17). SHEDs can produce functional vessel-like constructs following transplantation (93).

SHEDs, when maintained in osteogenic settings, dramatically enhance the pro-angiogenic activity (94). Recently, Kato et al. (95), verified SHEDS's pro-angiogenic function, which secretes angiogenesis-promoting molecules for primary endothelium cells (95). SHEDs exosomes-shuttled miR-222 promote aggregation and angiogenesis of PDLSCs via upregulation of TGF-β/SMAD system (96).

Gao et al. (97), found that multiple dosing of SHEDs lowered gingival hemorrhage, promoted new PDL attachment, and suppressed osteoclast development. Micro-computed tomography research revealed that SHEDs delivery substantially enhanced periodontal regeneration and alveolar bone mass. Additionally, a spike in the levels of CD206+ M2 macrophages was detected after the application of SHEDs (97).

PDL comprises a range of cell types, notably MSCs called PDLSCs, that share in periodontal healing and reconstruction. Several investigations have been conducted to identify PDLSCs and examine their multipotency (98–101). PDLSCs are multipotent and can transform into osteogenic, neuronal, and adipogenic cell lines (102).

PDLSCs help maintain the physiologic homeostasis of periodontal tissues (103). PDLSCs have architectural and proliferative characteristics comparable to those of MSCs, as indicated by biomarkers (104). PDLSCs injections can improve periodontal regeneration, restore the decline in population diversity, and raise the number of colonies of Bifidobacterium and Lactobacillus. In vitro, PDLSCs prevent the development of periodontal pathogens, including Staphylococcus aureus and Fusobacterium nucleatum. The fundamental mechanism for activity is thought to entail the synthesis of LL-37 (105).

SCAPs are a distinct population of MSCs found in the apical papilla of young permanent teeth. These cells have critical MSCs behaviors such as particular biomarker activity, self-renewal, division, mobility, multipotency, and immunosuppression features (106). Furthermore, significant evidence suggests that SCAPs can develop into several types of cells, like osteoblasts and odontoblasts, which may be a promising option for periodontal engineering (106, 107). Clinical evaluations, CT scans, and histopathological data revealed that SCAPs might dramatically increase periodontal regeneration 12 weeks following injection into a periodontitis animal model (108). This work validates the idea of employing SCAPs as an appropriate substitute stem cell resource for PDL regeneration in the future. Moreover, under inflammatory settings, the human apical papilla was shown to be mildly inflamed, retaining SCAPs viability and stemness while increasing its osteogenic and angiogenic capabilities (109).

DFPCs are a kind of dental MSCs that live in the dental follicle and is essential for tooth formation and function. DFPCs are neural crest-derived cells with multipotential transformation capabilities. More significantly, they offer advantages over other stem cells, such as ease of isolation and abundance, dynamic self-regenerating capacity, and absence of ethical concerns, which makes them an appealing choice in biotechnology. DFSCs are MSCs located in the tooth follicle and so share biological similarities with PDLSCs (24). In the inflammatory periodontal milieu, DFSCs have the potential to stimulate the division, osteogenic, and adipogenic transformation of both PDLSCs and inflammation-prone PDLSCs to various extents. Furthermore, when cultured together with DFSCs, the cell layering and ECM of PDLSCs/inflamed PDLSCs sheets expanded in vitro, whereas periodontal regeneration accelerated in vivo (110).

DFSCs, which are more undeveloped and demonstrate more DSPP than PDLSCs, can produce periodontal ligament (PDL) like constructions in vitro (111). A range of pluripotency biomarkers, notably octamer-binding transcription factor 4 (OCT-4), and NANOG, were demonstrated to be produced by DFPCs, confirming their multipotency and self-renewing capabilities (112). In contrast to other oral MSCs, DFSCs have a greater proliferation ability and osteogenic characteristics (113). Following in vivo implantation, DFSCs can replace the root by generating cementum and PDL (114). DFSCs had greater concentrations of osteogenic biomarkers, such as RUNX2 and ALP, than DPSCs and SHEDs (115).

GMSCs are a separate homogeneous group of MSCs that arise from neural ectomesenchymal tissues (116). GMSCs distinguish themselves from other oral MSCs due to their simplified accessibility and availability, as well as their exceptionally lengthy cultivation sustainability, lack of tumorigenicity, and persistent telomerase activity (117, 118).

Mitrano et al. (119), identified and analyzed GMSCs, which meet the basic specifications for MSCs, including multilineage differentiation, expression of MSCs markers, and increased adhesion (119). GMSCs demonstrated immune-modulating properties similar to those of other oral MSCs, inducing anti-inflammatory macrophage polarization and inhibiting osteoclasts, thereby lowering periodontal bone resorption in vivo (120). GMSCs' osteogenic ability has been established, and when transplanted into rats' gingival lesions, they restored healthy tissue (121). In rats, GMSCs-derived CM had a similar potential to stimulate PDL regeneration as PDLSCa-derived CM (122). In vivo transplantation of GMSCs can successfully regenerate bones (123).

The mouth houses a unique fatty tissue known as the buccal pad of fat or Bichat's pads (124).

Several investigations have employed BFPs as an autogenous transplant to reconstruct small- to medium-sized maxillofacial lesions (125–127). Furthermore, they are currently being utilized to generate MSCs called BFPSCs, which share characteristics and behavior with the more well-known dermal MSCs (128). This innovative procedure for producing BFPSCs has significant advantages because BFP collection is simple, involves just a small incision with local anesthetic, and generates low donor-area complications (129).

Farre-Guasch and colleagues (130) were among the initial researchers to identify BFPSCs (130). Utilizing rhBMP-2, Hiraishi et al. (131), verified the osteogenic capacity of BFPSCs. Additionally, only in cells containing recombinant bone morphogenic protein-2 (rhBMP-2) and osteoinductive reagents (OSR) were adipogenic genes readily visible. Yet, transplanting BFPSCs grown in this setting resulted in the most significant in vivo bone production. Therefore, when subjected to rhBMP-2 to induce mature osteoblastic development, BFPSCs consistently produced manufactured bone (131).

In surgically produced defects in rabbits' jaws, BFPSCs and cellular matrix (CM) both promote bone regeneration, indicating that BFPSCs primarily enhance bone regeneration via releasing paracrine substances. Regenerative dentistry is significantly influenced by the findings of MSCs' paracrine action on bone regeneration, and utilizing their CM can help tackle several problems and issues associated with cell transplantation. Specifically, CM provides greater convenience for medical professionals during clinical applications and is portable and easy to store (132). It is predicted that buccal fat pad tissue could offer valuable transplant material because it is readily accessible and has a rich vascularized area; however, further research is necessary to confirm this.

TGSCs have grown in popularity as a cell origin with great promise for transformation into many lineages. MSCs with endothelium and epithelial cells are essential for tooth formation, rendering them an ideal cellular reservoir for dental regeneration. TGSCs were discovered by a pedodontist, Dr. Songtao Shi, while working on his six-year-old daughter's deciduous teeth in 2003 (133). The tooth germ is a cluster of primitive cells that participate in the formation of teeth and related structures (134). MSCs-like properties can be observed in cells generated from the third molar tooth germ. MSCs-specific surface antigens are expressed by human dental germ cells (135). TGSCs' multipotency enables them to develop into osteoblasts, odontoblasts, adipocytes, and brain cells. Human TGSCs exhibit immune-regulatory characteristics (136). Guzman et al. (137), demonstrated that the application of human TGSCs has immune-suppressing actions in mice (137).

Extracellular matrix (ECM) components, including proteins and glycans, are secreted by living cells in tissues to form complex networks that regulate cell behavior and enable cells to fulfill specific roles, providing crucial cues and substances for cell migration and proliferation (149). Nevertheless, since 2D well plates are unable to facilitate biomacromolecule agglomeration or the spatial space necessary for cell adhesion, they are absent from conventional 2D cell culture techniques (150). Complex photophilic polymeric chains, protostructures, and elevated water levels make hydrogel matrices the ideal substrates for simulating in vivo cell culture conditions (151). As a result, hydrogels are frequently employed as synthetic substrates or frameworks in biohybrid networks, offering the benefits of long-term survivability, self-healing, and bottom-to-top construction (152).

Cells can be directly embedded in the hydrogel matrix, seeded on films or fibers, or seeded on a decellularized matrix to create common hydrogel-based biohybrid systems (153). The development of tissue engineering has led to notable breakthroughs in sophisticated manufacturing techniques, with 3D bioprinting being the most promising of these (154–157). Hierarchical designs can be formed from bottom to top thanks to this technological capacity to build cells at the microscale in customizable 3D areas (158). 3D printing is more reliable than traditional manufacturing techniques when it comes to creating various biological networks and systems, and accurately specifying the architecture of cells. These benefits over conventional manufacturing techniques have been demonstrated in domains such as clinical healthcare, biological science, and organ regeneration (159). Inkjet, extrusion, and photosensitive approaches are among the various types of 3D printing techniques that are primarily accomplished by sequential layering of sensitive inks (160, 161). These techniques are also well-suited for cell-laden, sensitive hydrogel engineering, which enables a variety of biohybrid functions. It is now simpler to load cells into scaffolds to assist cell functionalization, thanks to the 3D printing process that creates hydrogel scaffolds (162). There are several advantages to using hydrogels as platforms for cell seeding, particularly in regenerative applications, such as bone regeneration and repair. As cells are suspended directly in the hydrogel solution, cell-containing hydrogel bioinks enable the direct fabrication of 3D designs (163).

Because of its intricacy, reconstruction of the periodontium requires the synchronized repair of several structures. Numerous 3D printing processes, such as the freeform reversible embedding of suspended hydrogels (FRESH), employed by Lin et al. (164) are utilized to carry out this operation (164) with a bioink incorporating type I collagen, thereby building collagen microfibers. The findings demonstrated that the growth, attachment, and vitality of the cytoskeleton were satisfactory, and the PDLSCs were effectively implanted (164).

However, Tian et al. (165) utilized a hydrogel, hydroxyapatite nanoparticles, and PDLSCs to combine synthetic and natural materials, thereby generating a bioink (165). This bioscaffold enhanced the mechanical characteristics and swelling capacity while also effectively stimulating cellular viability, division, and differentiation. Zhu et al. (166),, also employed PDLSCs in GelMA hydrogel at various levels (3%, 5%, and 10%). The addition of PDLSCs helped create new cells, but the 10% GelMa demonstrated lower cell longevity (166). One benefit of hydrogels is that they may be administered directly via injection into the targeted region using less invasive techniques (Figure 3). Kandalam et al. (167) effectively enclosed GMSCs in PuraMatrix™, a self-assembling hydrogel. The GMSCs immobilized with 0.5% PuraMatrix showed exceptional attachment and multiplication ratios.

Hydrogels must be both economical and simple to manufacture, in addition to having a therapeutic impact, to be commercially successful and widely used. Enhancing the ability to attract native cells might encourage the therapeutic application of biomaterials by eliminating the challenges and costs related to the development, preservation and delivery of cellular materials, besides safety and ethical issues (168).

Hydrogels can be manufactured from a broad spectrum of materials, including both artificial and organic sources, and can be utilized in various formulations (169, 170). Natural hydrogels exhibit good biological compatibility and minimal immunological response (171, 172), yet this is often accompanied by inadequate scalability and limited control over mechanical properties (170, 173–175).

Manufactured hydrogels possess an identifiable framework, consistent material sources, and an extended shelf life, and can be constructed in several batches with reproducibility. The fundamental challenges are their low bioactivity, antigenic byproducts, and static makeup, which provides no biological data to cells. Thus, they may be employed in combination with bioactive molecules to mitigate these shortcomings (176–178).

Isaac et al. (179), used immersed electrosprayed polyethylene glycol (PEG) hydrogel microparticles, which were subsequently used to make tetrazine click chemistry to prepare microporous annealed particle-based hydrogels (TzMAP) by attaching norbornene-loaded PEG hydrogel microspheres. Incorporating PDLSCs during annealing resulted in cellular viability proportions of 87% ± 5% at 24 h. The inclusion of PDLSCs and platelet-derived growth factor (PDGF)-BB into TzMAP led to enhanced cellular division and movement, which are crucial for periodontal tissue regeneration (179).

Hydrogels can be carefully manufactured for optimized cell dispersion and tissue formation through self-assembling or 3D printing processes (180–182). For encapsulation of PDLSCs, a bioprinting technique used injectable composite hydrogels made of GelMA and PEG dimethacrylate (PEGDA). The use of PEG improved droplet control. In vivo tests demonstrated that PDLSCs-containing hydrogels stimulated bone formation in rat periodontal defects when compared to hydrogels lacking cells (183).

Natural polymeric hydrogels derived from microbes, plants, or animals have garnered considerable interest recently, owing to their exceptional biodegradability and biocompatibility (184). However, their practical uses are limited by their weak mechanical characteristics, unpredictable rates of breakdown, and the likelihood of immunological responses. To meet the potential needs for dental applications, both artificially produced and organic composite hydrogels have been proposed. The most often used injectable matrix for cellular transportation is composite hydrogels (185).

Tissue engineering is based on hydrogels, which provide a framework for cell adhesion, proliferation, and transformation. By modifying their functional components, cross-linking techniques, and synthetic materials, the physical-chemical characteristics of hydrogels can be tuned to satisfy the unique biomechanical requirements of various tissues. Because hydrogels can be customized for specific medical applications, they have been utilized in medication delivery, regenerative medicine, and other fields of medical research. Many novel hydrogels have been manufactured to enhance the interaction between targeted cells and hydrogels, thereby promoting tissue regeneration (186).

Rapidly cured microporous hydrogels based on gelatin and GelMA were shown in a prior work (187). Via photopolymerization and enzymatic cross-linking, these hydrogels set after 2.5 min of injection, enabling consistent cell dispersion and substantial cellular dissemination and division within a week. Furthermore, these hydrogels may carry hMSCs primed with interferon-gamma, increasing the production of anti-inflammatory substances, including interleukin-6 and prostaglandin E2. Therefore, these hydrogels offer a potential method of delivering cells. Another safe method for cell transplantation is the use of hydrogel particles. Hydrogel particles, which are formed of microscopic 3D network structures built of biopolymers, shield cells from shear pressures during transplanting, maintaining cell viability (187).

According to recent research, the physical characteristics of hydrogels (like the stiffness, durability, surface charge, and other physical factors of ECM can be altered to control cell expansion, division, and transformation; therefore, they impact the regenerative processes. The majority of research has produced hydrogels with varying stiffness by altering their composition or applying external stimuli. The investigation of variations in cell activity has also made extensive use of these hydrogels with varying stiffness. For instance, the stiffer the hydrogel, the more likely the oral stem cells can develop into bone, making it a potential option for regenerative dentistry (188, 189).

He et al. (190) developed and evaluated stiff transglutaminase-crosslinked gelatins (TG-GELs) with SDF-1α and/or IL-4 as a potential platform in periodontal therapy. According to their findings, IL-4 could encourage the transformation of Mφs to the M2 phenotype, which may promote BMSCs' osteogenesis in vitro. Furthermore, the inclusion of SDF-1α might continuously encourage the functionality of BMSCs. Periodontal regeneration was considerably enhanced after implantation of these bioactive gels into periodontal defects as opposed to TG-GELs carrying either IL-4 or SDF-1α alone. These findings suggest that improving the ability of high-stiffness hydrogels to modulate macrophages (Mφ) and attract cells is a feasible and successful method for performing targeted periodontal regeneration. Therefore, if the synchronized crosstalk between stem cells and Mφs is appropriately guided, very simple, designed biomaterials can give significant functional advantages and good regeneration results with the help of carefully chosen signaling molecules (190).

Biodegradability is a key design factor that influences the pharmacokinetics and pharmacodynamics of hydrogels. Hydrolysis of ester, carbonate, or anhydride chains; metabolic breakdown of peptide, polysaccharide, or proteolysis by enzymes like matrix metalloproteinases; or redox- and pH-triggered disintegration of dynamic covalent bonds. To optimize decomposition dynamics, polymer molecular weight, crosslinking capacity, hydrophilic properties, and the introduction of labile functional chains must all be balanced out. Customized breakdown allows synchronicity with tissue healing timeframes, ranging from fast disintegration for bursting delivery to delayed degradability for extended reinforcement. Furthermore, breakdown metabolites must be non-toxic, immune-compatible, and quickly eliminated to avoid long-term inflammation (191).

Biological compatibility is determined by the hydrogels' capacity to keep encapsulated or invading cells viable and functioning. Polymeric purity, minimal endotoxin concentration, the absence of harmful crosslinking by-products, and the preservation of homeostasis during gelation are all important variables. Moderate gel formation settings are required, whether by light triggering and enzymatic processes. Mechanical qualities also impact cellular activity; hydrogels with insufficient durability can impede cell dissemination, relocation, lineage determination, and ECM deposition. To avoid hypoxic or apoptotic areas inside cell-laden structures, nutrition, oxygen, and waste byproducts flow must be optimal (192).

Biocompatibility refers to the capacity of hydrogels to interact with host tissues without causing unfavorable immunologic or inflammatory responses. After administration, hydrogels interact with immune cell populations, notably macrophages. Biomaterial chemical composition, charge concentration, crosslinking methods, architectural characteristics, and decomposition products all impact the behavior of macrophages toward pro-inflammatory or pro-regenerative phenotypes (190).

Stimuli-sensitive hydrogels provide dynamic features that allow for sensing and responding to surrounding signals. They can be activated by heat, light, pH, and chemicals, thereby imparting them with environment-sensitive properties (193–195). Along with these qualities, hydrogels also exhibit other common traits, such as softness, swelling capacity, and water retention capacity. Additionally, the polymer chains in hydrogels can be purposefully altered or tailored to enhance particular qualities (196). For instance, hydrogels can acquire stimuli-sensing capabilities by linking polymers to biological molecules, which adds living things, such as proteins, to polymer chains (197). Temperature-sensitive platforms encounter sol-gel transformations around physiological temperatures when the hydrophobic-hydrophilic balance shifts within the polymeric matrix. pH-sensitive hydrogels with ionized motifs exhibit swelling or deswelling activity depending on protonation status. Enzyme-sensitive hydrogels include peptide chains or degradable components that can be preferentially degraded by illness or tissue-related enzymes. Glucose and redox-reactive hydrogels use reactive boronate ester bonds or oxidative breakdown of highly reactive links to provide tailored medication release under metabolic or inflammatory conditions. Light-sensitive hydrogels contain photosensitive molecules, allowing for precise time- and space-based manipulation of gelation and breakdown. These behavioral modifications improve accuracy, decrease undesirable outcomes, and increase therapeutic variety (198).

Hydrogels intended for periodontal regeneration should reduce acute inflammation, prevent persistent fibrotic encapsulation, and optimally establish an immunological microenvironment favorable to tissue regeneration. The inclusion of immune-modulation elements, bioactive substances, or anti-inflammatory drugs might help modulate immunological reactions at the location of administration. Additionally, by influencing redox reactions, hydrogels can be utilized for tissue restoration (199). Elevated ROS synthesis can control the associated biological reactions during tissue regeneration and raise the amount of MSCs-related protein secretome (200). Additionally, some composite hydrogels can alter the behavior of macrophages by making the hydrogel more rigid, changing them to an anti-inflammatory M2 type, and encouraging tissue reconstruction by controlling associated immunological and inflammatory responses (201). By altering the associated biophysical characteristics of hydrogels and influencing the presentation of relevant factors, it can generally impact the phenotypic or proliferative maturation of cells.

Despite the use of modern treatments, chronic oral inflammatory illnesses such as pulpitis, periodontitis, and peri-implantitis present serious clinical problems and frequently cause irreparable tissue loss. By actively encouraging the suppression of inflammation and tissue regeneration while maintaining host defense, specialized pro-resolving mediators (SPMs) provide a revolutionary treatment paradigm. Nevertheless, SPMs' low affinity to inflammatory tissues, limited bioavailability, and quick disintegration impede their practical application. Promising platforms to overcome these obstacles include smart biomaterial-based delivery systems, particularly stimuli-responsive hydrogels. These devices enhance the stability and therapeutic efficacy of SPMs by enabling their regulated, localized, and environmentally induced release (202).

Hydrogel-mediated SPM administration not only reduces inflammation but also maintains tissue integrity and encourages regeneration, according to preclinical research in oral inflammation models. To facilitate clinical acceptance, future strategies will focus on enhancing dosage procedures, ensuring long-term bioactivity, and addressing manufacturing and regulatory challenges. Biomaterial-based approaches have an opportunity for transforming the medical management of oral inflammatory illnesses and promoting regenerative dental treatments by improving the delivery and prolonged bioactivity of SPMs (203).

Numerous inflammatory conditions, such as periodontitis, are characterized by excessive ECM degradation by MMP-1. Under optimal circumstances, MMP activity is carefully regulated—for example, by tissue inhibitors of metalloproteinases (TIMPs)—to maintain homeostasis. MMPs hydrolyze peptide bonds with a high degree of amino acid affinity. Nevertheless, MMP activity persists under pathophysiological conditions, leading to unfavorable alterations in tissue structure and function, and accelerating the course of the disease. Over the past 25 years, numerous investigations have been conducted on the construction and creation of molecules that block MMP activity in an effort to alleviate this (204–206).

Huang et al. (207) constructed an injectable dual-crosslinked protein hydrogel by integrating gelatin and bovine serum albumin (BSA) using a rapid, straightforward, and cross-linker-free manufacturing technique. Its cross-linked, interconnected design offered extra support to optimize thermal stability and mechanical characteristics. CD/BSA/GEL hydrogel was produced by adding the potent oxidant ClO₂. To administer targeted high-concentration medication effectively, this hydrogel is easily injectable into periodontal pockets. It reacted with and alleviated elevated protease components in the periodontal inflammatory milieu, allowing for the prolonged release of ClO₂ and bioactive cargo. This system downregulates the expression of inflammatory genes and maintains cell motility, expansion, division, and osteogenic potential while exhibiting substantial antimicrobial characteristics. The CD/BSA/GEL hydrogel successfully minimized inflammatory reactions and encouraged periodontal bone regeneration (207).

The prevalence and progression of periodontitis are clearly positively correlated with H2S, a common metabolite of periodontal bacteria. H2S can control numerous biological processes at physiological quantities. Nonetheless, excessive H2S in the periodontal pocket can exacerbate the progression of periodontitis by inducing oxidative stress, releasing proinflammatory cytokines, causing mitochondrial damage, and promoting apoptosis in human gingival fibroblasts. Even worse, by preserving bacterial redox balance and boosting antibiotic resistance, H2S promotes bacterial survival and growth. However, eliminating H2S is often overlooked when treating periodontitis. To improve the management of periodontitis, Xie et al. (208) developed a type of hyaluronic acid methacryloyl/ZnO (HMZ) hydrogel with the ability to scavenge H2S. By reacting with ZnO, the HMZ hydrogel was able to eliminate H2S and had high injectability and cytocompatibility. Consequently, the HMZ hydrogel restored mitochondrial homeostasis, reduced inflammation mediated by the cGAS-STING signaling pathway, and increased cell survival from 13% to 120% for human gingival fibroblasts and from 22% to 94% for human periodontal ligament fibroblasts after 48 h. Additionally, the HMZ hydrogel demonstrated effective plaque biofilm removal and adequate antibacterial properties in vitro and in vivo. In summary, a potential approach based on H2S elimination was created to increase the efficacy of periodontitis treatment (208).

Since hydrogel stiffness affects stem cell activity, including development, attachment, and relocation, it is an essential component of stem cell administration (209). To influence the destinies of stem cells, investigators can modify the stiffness of hydrogels. For instance, stiffer hydrogels could stimulate osteogenic development, whereas softer hydrogels may support adipogenic transformation or preserve stem cell properties. Due to this adaptability, specific niches may be created that can enhance the effectiveness of stem cell treatments (210, 211).

Osteogenic differentiation results from a greater nuclear-cytoplasmic ratio of Yes-associated protein (YAP)/transcriptional coactivator with PDZ-binding motif (TAZ) on the surface of the stiffer hydrogel. The nuclear-cytoplasmic proportion of YAP/TAZ is lower on the scaffold's less rigid surface, causing the cells to develop into adipocytes (212, 213).

Since they may be utilized to carry medications and cells and replicate the chemical makeup of the extracellular matrix, hydrogels are regarded as promising biomaterials. Properly controlled synthetic procedures can produce artificial polymeric hydrogels with well-specified, well-defined chemical compositions, precise molecular weights, enhanced stiffness, and customizable microstructures. Nevertheless, synthetic polymer hydrogels are often either biodegradable or biocompatible, which makes them inappropriate for cell reinforcement (21).

Although hydrogels have demonstrated potential as transporters for stem cell transplants, there are still obstacles (19). Cell mortality after delivery, limited cell attachment after transplantation, challenges to extended cell longevity, and insufficient reinforcement are all significant factors influencing the effectiveness of periodontal regenerative therapies (204, 214). Currently, there is no ideal hydrogel cellular vehicle to address these challenges (215, 216).

DFSCs sheets exhibit high extracellular secretion capacity and efficiency in periodontal regeneration. DFSCs sheets produced from passage 4 cells outperformed DFSCs suspensions in terms of vitality and osteogenic transformation potential. Following 10 days of culturing, DFSCs sheets showed upregulation of osteogenic biomarkers. In comparison to the cell suspension, the cell sheet exhibited superior regenerative and paracrine capabilities in vivo. The modified DFSCs sheets revealed considerably greater concentrations of VEGF and angiopoietin-1 in comparison with DFSCs suspensions, as well as improved osteogenic initiation effects (223).

Tri-layered hydrogel formed of chitin-poly(lactic-co-glycolic acid) (PLGA) composite hydrogel. The first layer contained nanoinspired bioactive glass (nBG). The second layer contained fibroblast growth factor 2, and the third layer was loaded with nBG/platelet-rich plasma. It was biocompatible and promoted cementogenesis, fibrogenesis, and osteogenesis of human DFSCs both in vitro and in vivo (224). A 3D bioprinted biomimetic cell-laden bioink containing gelatin methacrylate/decellularized extracellular matrix (GelMA/dECM). The dECM exhibited strong immune-modulating properties while minimizing local inflammation in vivo. This hydrogel substantially promoted harmonious PDL fiber regeneration and enhanced bone mineralization in vivo (225).

Although deferoxamine (DFO) promotes angiogenesis during bone regeneration and wound healing, its effect on DPSCs driven angiogenesis remains uncertain. Here, DFO was loaded into gelatin-based microspheres (GMSs), and thermally responsive injectable hydrogels containing chitosan and collagen were created to allow for regulated DFO release. Due to its advantageous physical characteristics and biocompatibility, the DFO-GMS-laden hydrogel composite enabled prolonged DFO administration for up to 15 days. In vitro, DFO successfully induced tube formation, increased the release of molecules linked to angiogenesis, and encouraged DPSCs' movement (226).

In addition, Divband et al. (227), created new injectable hydrogels in situ using chitosan biguanidine and carboxymethylcellulose that were loaded with rhBMP-2 and VEGF. They investigated the impact of the fabricated hydrogels on the osteoblastic development of DPSCs. These hydrogels considerably boosted the growth of DPSCs and were non-toxic. Additionally, they displayed noticeably increased ALP, COL1α1, and OCN genetic and protein transcriptions (227).

Wang et al. (228), investigated hyaluronic acid (HA) and polyethylene glycol loaded with DPSCs' extracellular vesicles (EVs), which successfully relieved inflammation, expedited revascularization, and encouraged tissue mineralization in vivo (228).

GelMA-based biomaterials are frequently employed as scaffolds due to their biological compatibility, adjustable characteristics, and functional capacities. Nevertheless, the biological effects of its photo-crosslinking mechanism on MSCs, as well as stress-reduction methods, have yet to be fully investigated. Integrating DPSCs-CM into the GelMA hydrogel demonstrated increased cellular survival, growth, movement, and osteogenic transformation (229).

An injectable thermally responsive Cs/β-glycerophosphate/hydroxyapatite hydrogel can preserve typical adherent DPSCs cell shape, encourage fast multiplication, promote high cell viability, and enhance osteogenic activity (230).

It is found that alginate is a potential non-toxic platform for GMSCs encapsulation (231). Moreover, alginate hydrogel could promote osseointegration of GMSCs in vitro (231). Fawzy El-Sayed et al. (232) explored the regenerative capacity of GMSCs to reconstruct periodontal tissue when combined with HA-sECM that releases IL-1ra. GMSCs exhibited stem and MSCs properties. Interleukin (IL)-1ra-loaded and unloaded GMSCs/HA-sECM demonstrated increased clinical attachment level (CAL), decreased junctional epithelium (JE), and enhanced bleeding on probing (BOP) compared to negative controls (232). Wang et al. (76) developed a CS/ oxidized chondroitin sulfate (OCS) hydrogel by freeze-casting process to carry PDLSCs and GMSCs with the goal of inducing periodontal tissue regeneration. The PDLSCs and GMSCs loaded hydrogels demonstrated outstanding biocompatibility, more significant bone tissue healing, and generated more organized PDLs in vivo (76).

Furthermore, Ansari et al. (233) created an alginate/GelMA hydrogel formulation that encapsulates GMSCs. The GMSC-hydrogel could speed up wound healing by increasing collagen production, encouraging angiogenesis, and blocking local proinflammatory cytokines (233). Balaban et al. (120) verified that the local treatment of GMSCs in fibroin/chitosan oligosaccharide lactate hydrogel (F/COS) resulted in significant new bone growth and less lengthy junctional epithelium creation with well-organized PDLs and connective tissues (120).

Moshaverinia et al. (234) demonstrated that microencapsulation of PDLSCs and GMSCs in RGD-alginate hydrogel improved MSCs survival and osteogenic transformation in vitro. Moreover, PDLSCs loaded hydrogels could restore the damaged bone by stimulating the creation of mineralized tissue, while GMSCs had much inferior osteogenic differentiation competence (234). Pouraghaei Sevari et al. (235) developed alginate-whitlockite (WHMP) hydrogels, which showed enhanced elasticity without impacting the viability of GMSCs. Additionally, alginate-WHMP hydrogels stimulate the mitogen-activated protein kinase (MAPK) system, which regulates various osteogenic biomarkers in encapsulated GMSCs, particularly RUNX2 and osteocalcin (OCN). They could inhibit osteoclastic activity, likely thanks to the liberation of WHMP Mg²⁺ ions and GMSCs osteoprotegerin. GMSCs encapsulated in an osteogenic niche may improve bone repair in vivo (235). Kandalam et al. (167) employed a self-assembled hydrogel scaffold, PuraMatrix™ (PM), and/or BMP2 and loaded them with GMSCs. GMSCs-laden hydrogels could enhance bone regeneration upon in vivo application (167).

The good physical characteristics and biodegradability of a temperature-sensitive hydrogel have been demonstrated, making it an effective scaffold. Thermosensitive hydrogel in conjunction with lentiviral PDGF-BB loaded with SCAPS dramatically increased new bone growth and mineralization (236). Dutta et al. (237) demonstrated that 3D bioprinted temperature-sensitive poloxamer-407 (P407) hydrogels are non-toxic for encapsulating SCAPs, leading to high cellular survival and increased cell migratory potential. After 14 days of in vitro growth, the 3D hydrogels containing PAI-1 showed significantly higher levels of osteogenic biomarkers (237).

Chen et al. (238), created an injectable antimicrobial peptide-/(GelMA-AMP) hydrogel that contains hypoxia-inducible factor (HIF-1α), which promoted osteogenic transformation in SHEDs (238). Qu et al. develop SHEDs laden with metformin (MF)-loaded mesoporous silica nanospheres (MSNs)-/ (GelMA) photo-cured hydrogels. This bioactive hydrogel did not affect cell survival and showed significant activation of osteogenic-related genes (excluding OCN) (238).

Ivanov et al. (239), demonstrated that PDLSCs grown in a 3D collagen I hydrogel with dECM for 14 days exhibited phenotypic traits comparable to those of osteoblast-like cells. The discharge of bone-forming biomarkers (OC, OPN, and ALP) demonstrates this capacity. In 3D culture, the inclusion of fibronectin to the dECM promotes the most efficient conversion of PDLSCs into osteoblasts (240). PDLSC-laden GelMA and PEG dimethacrylate hydrogel can promote PDLSC survival and spreading (239). HydroMatrix™ (HydM) is a synthesized self-assembly peptide that can produce hydrogels with thermal or ion concentration variations. Nagy et al. (183) discovered that PDLSCs can attach, live, relocate, and multiply on HydM, and the gel promotes osteogenic transformation, encourages the viability of PDLSCs and their osteogenic transformation in vitro, as well as hastens osteogenesis in vivo (183).

Saputra et al. (241) constructed a nanohydroxyapatite-chitosan (nHPA-CS) hydrogel injectable scaffold loaded with PRF and BFPSCs. The NHPA-CS scaffold develops a native microenvironment that stimulates BFPSCs' driven regenerative capacity, whereas PRF promotes osteogenic differentiation and multiplication of BFPSCs. The integration of nHPA-CS, platelet-rich fibrin (PRF), and BFPSCs exhibits osteoinductive capacity in patients with aggressive periodontitis (241). Bastami et al. (242) developed a 3D β-tricalcium phosphate (β-TCP)/gelatin/BMP2/chitosan (CS)/collagen composite and explored its impact on hBFPSCs. This hydrogel might serve not only as a structurally and physiologically suitable scaffolding, but also as an osteoinductive graft, supplying rhBMP2 during a therapeutic window for hBFPSC development into the osteoblast lineage (242).

Limitations with employing stem cells include decreased survival of fewer cells, restricted division, and the stem cells' capability to differentiate. Although oral stem cells have demonstrated enhanced periodontal regeneration, there is still a lack of pre-clinical and clinical evidence, and more carefully thought-out studies with a bigger sample size are necessary to determine whether employing stem cells to boost periodontal healing is feasible (252). The effectiveness of stem cells in promoting periodontal regeneration is reduced when their sustainability is reduced as a result of late glycosylation byproducts in a hyperglycemic condition (253). DPSCs have shown the ability to mineralize. Indeed, it has been established that human DPSCs encourage osteogenesis both in vitro and in vivo. DPSCs' low ability to generate cementum raises doubts about their efficacy for periodontal regeneration. Despite these experiments showing oral stem cells' capacity for regeneration, the majority of them lack comprehensive quantitative methods to assess the cells' capacity for self-renewal, proliferation, and differentiation, particularly in vivo (87, 254, 255).

Furthermore, preceding their clinical use, the laboratory tests must address the subsequent concerns: 1) Significant apoptosis in cells at the transplantation site, thus the longevity and functionality of oral stem cells in vivo must be enhanced. 2) The relationship between implanted cells and the native cells must be investigated; 3) In vivo cell lineage mapping of implanted oral stem cells is essential to comprehend their destiny and activities. 4) Because some oral stem cells are frequently engaged in carcinoma, the biochemical pathways that enable oral stem cells to select between self-renewal, malignancy, and transformation should be thoroughly investigated (22, 256).

In addition, while these experiments are extremely helpful in discovering the characteristics of oral stem cells, they do not completely mirror the biological and pathological state of the injured tissues in the human body. Double-blind, randomized controlled studies are required to prove their genuine regeneration ability. Because clinical studies require a high quantity of clinical-grade cells within a short period, preservation and handling of oral stem cells are viable options for clinical use (256). After Hiroshima University established an institutional tooth bank in Japan in 2004, numerous biotechnology companies specializing in teeth banking have begun operations. With improved preservation technologies, regenerative dentistry could potentially act as a portal to an extensive spectrum of rejuvenating therapies by successfully tapping these beneficial stem cell reserves (257).

While the technical details of the banking procedure should worry an interested person, there is typically little information accessible from the public side of dental banks that enables people to distinguish between solutions. Their selection will most certainly be heavily affected by both regional accessibility and industry-specific, or more specifically, financial variations across banking centers (258). Additionally, prolonged stem cell preservation is not currently covered by medical insurance policies in the United States and is not a regular service in the majority of nations. A straight cost analysis may be useless because charges are liable to fluctuation, and there are many other types of plans available. However, as a rule of thumb, the initial processing may cost between $500 and $2,000 (US dollars), and the yearly maintenance can cost between $99 and $264. For a 20-year contract with no annual maintenance fees, several businesses offer a fixed fee of between $2,000 and $3,000 (259). In fact, this is Oothy's sole product, with the notion of preserving stem cells from deciduous teeth as a sustainable therapeutic investment. Therefore, choosing a service based solely on expense is undoubtedly a challenging and intimidating task, and it still fails to account for the specific services provided by each tooth bank. In this regard, NDPL, Oothy, and Stem-Save can collect a large number of teeth simultaneously. Still, BioEden and Tooth Bank do not charge for handling teeth until a satisfactory stem cell extraction is achieved. Some businesses provide full reimbursement if the entire procedure fails, while others offer credits for future teeth. Licensing and reliability are two additional unpredictable criteria. While every center guarantees security for cryopreservation operations, liquid nitrogen storage, and patient privacy, it is challenging for individual customers to compare centers based on these features. Eventually, it is realistic to expect dentists or other healthcare professionals to attract potential patients to the notion of oral stem cell banking and to suggest personalized banking services, which will have the most significant impact on any subsequent preference (260).

Despite the promising potentials of stem cells, numerous investigations are necessary to establish the safety and efficacy of these cells in periodontal applications. However, the standardized design and improvement of oral stem cell cryopreservation techniques must overcome significant hurdles, such as culture-driven variances, patient-associated variations, and the effects of culture medium additions. Only in this manner can we enhance and strengthen oral stem cells as potential therapeutic options for patients who have few or no other treatment options (261). These stem cells are becoming increasingly essential not just in dental care, where they play a crucial role in tissue regeneration and preservation, but also in other medical sectors, where they are gaining popularity. In summary, further examinations are required to confirm the seemingly regenerative ability of these stem cells. Still, it appears extremely promising to examine their regeneration capacities in a wide range of disorders, given that they are easily accessible, exist throughout life, and possess remarkable multipotency (262).