Hybrid Nanocomposite Mini-Tablet to Be Applied into the Post-Extraction Socket: Matching the Potentialities of Resveratrol-Loaded Lipid Nanoparticles and Hydroxyapatite to Promote Alveolar Wound Healing

Abstract

Background/Objectives: Following tooth extraction, resveratrol (RSV) can support healing by reducing inflammation and microbial risks, though its poor solubility limits its effectiveness. This study aims to develop a solid nanocomposite by embedding RSV in lipid nanoparticles (mLNP) within a hydrophilic matrix, to the scope of improving local delivery and enhancing healing. Hydroxyapatite (HXA), often used as a bone substitute, was added to prevent post-extraction alveolus volume reduction.

Methods: The mLNP-RSV dispersion was mixed with seven different polymers in various mLNP/polymer ratios. Following freeze-drying, the powders were redispersed, and the resulting dispersions were tested by DLS experiments. Then, the best two nanocomposites underwent extensive characterization by SEM, XRD, FTIR, Raman spectroscopy, and thermal analysis as well as in vitro partitioning studies aimed at verifying their ability to yield the mLNP-RSV from the hydrophilic matrix to a lipophilic tissue. The characterizations led to identify the best nanocomposite, which was further combined with HXA to obtain hybrid nanocomposites, further evaluated as pharmaceutical powders or in form of mini-tablets.

Results: PEG-based nanocomposites emerged as optimal and, following HXA insertion, the resulting powders revealed adequate bulk properties, making them useful as a pharmaceutical intermediate to produce ≈59 mm³ mini-tablets, compliant with the post-extraction socket. Moreover, they were proven ex vivo to be able to promote RSV and GA accumulation into the buccal tissue over time.

Conclusions: The here-proposed mini-tablet offers an innovative therapeutic approach for alveolar wound healing promotion as they led to a standardized dose administration, while being handy and stable in terms of physical solid identity as long as it takes to suture the wound.

Introduction

Tooth extraction is one of the most common surgical procedures in dentistry [1]. The healing process of the socket involves both soft (e.g., periodontal ligament, gum) and hard tissues (e.g., alveolar bone) [2]. The healing of the socket after tooth extraction occurs in four distinct phases. The first one is governed by hemostasis and coagulation; bleeding from the socket allows platelets to interact with endothelial cells and the extracellular matrix, starting the coagulation cascade that forms an initial blood clot, establishing hemostasis and creating a structure for further cell adhesion [3]. In the second phase, cytokines and growth factors play their role in the inflammatory stage, leading to the organization of the fibrin clot, which is replaced by granulation tissue. In the third phase there is rapid deposition of a temporary matrix (fibroplasia) which displaces granulation tissue and periodontal ligament remnants, followed by the formation of woven bone around newly developed blood vessels. Finally, the modeling and remodeling stages reshape the bone structure involving the deposition of lamellar and bone marrow, replacing woven bone without affecting overall bone shape or architecture. Osteoblasts and osteoclasts are crucial in this phase, during which both bone and surrounding soft tissue loss may be observed due to increased osteoclastic activity [1,4,5,6].

The exacerbation and prolongation of even just one of these phases leads to an incomplete healing process. Since the four phases are all interconnected, and considering the ability of inflammatory processes to self-sustain, as well as the importance of reactive oxygen species (ROS) in the various signaling pathways involved in the wound repair process, a strategy to ensure prompt healing includes the use of natural substances with scavenging, anti-inflammatory and antimicrobial activities. Nowadays, innovative dental materials, intended as mouthwashes, endodontics, orthodontics and filling materials, containing natural phyto-components are an emerging trend due to their efficacy, low cost, and non-toxicity against humans. Particularly, polyphenols belong to an emerging class of useful natural bioactives in dentistry [7].

Among them, resveratrol (RSV) currently stands out. It is a polyphenolic phytoalexin, belonging to the stilbene subfamily, capable of exerting antioxidant, anti-inflammatory, and cardioprotective actions [8,9,10]. RSV exerts protective effects on bone tissue and, also, bone formation, since it is an active against osteoporosis and bone resorption due to aging [11,12]; as well as it can also stimulate the proliferation and differentiation of osteoblasts [13]. Furthermore, it possesses antimicrobial activity which could be helpful in protecting the damaged tissue from microbial colonization that otherwise could slow down or prevent the healing process [14]. Unfortunately, RSV has an unfavorable pharmacokinetic profile due to its low solubility in aqueous media and high susceptibility to the hepatic first-pass metabolism, resulting in a bioavailability of less than 1% when taken orally [15]. Consequently, considering that the post-extraction socket healing process is limited to a certain area of the oral cavity, it should be more effective to act locally.

Nevertheless, RSV is unwieldy due to the already mentioned poor water solubility together with instability due to light exposure, alkaline pH, and high temperatures [16]. Therefore, considering its lipophilic nature, its insertion into lipid-based nano-platforms could represent a valuable solution to protect RSV and promote its interaction with soft tissues which also have hydrophobic nature. Lipid-based nano-platforms have evolved over the years: (i) the Solid Lipid Nanoparticles (SLN) are composed of physiologically tolerated solid lipids, finely dispersed in an aqueous phase containing at least one surfactant [17]; (ii) the Nanostructured Lipid Carriers (NLC) consist of a mixture of at least two lipids, one solid and one liquid at room temperature and pressure, allowing higher stability and drug encapsulation capacity than the SLN [18]; (iii) the Multicomponent Lipid Nanoparticles (mLNP) represent a further progression of the NLC as they are still based on a mixture of at least two lipids (one solid and one liquid) but also present some functional components capable of assisting and synergizing the therapeutic effect of the encapsulated active ingredient, e.g., for wound healing purposes, functional excipients characterized by proper scavenging and antimicrobial properties should be chosen. Specifically, we have already developed and characterized RSV-loaded mLNPs based on a mixture of Labrasol® (liquid lipid), Glyceryl Monostearate (solid lipid), Menthol, and 18-β-Glycyrrhetinic Acid (functional lipids) [19].

These functional excipients were generally chosen for their wound healing potential. Specifically, glycyrrhetinic acid (GA) is a natural triterpene glycoconjugate present in licorice roots and it is a well-known HMGB1 inhibitor. GA has been proven to possess immunomodulatory, antioxidant, antibacterial, and anti-inflammatory activities, and has been widely used to clinically treat such chronic inflammatory diseases (e.g., chronic hepatitis, atopic dermatitis and other skin diseases) [20,21]. Furthermore, a recent study published in 2018 underlined its osteoprotective ability by inhibiting osteoclasts activity [22]. On the other hand, menthol is a natural terpene that has been widely investigated as a penetration enhancer as well as for its antimicrobial potential, enabling both the maintenance of wound asepsis and the promotion of RSV entry into the target injured tissue [23,24]. The accurate design of the proposed mLNP-RSV led to a promising nanosystem which displayed powerful antioxidant activity due to the synergistic effect of RSV and GA, as highlighted by the DPPH assay, as well as interesting wound healing properties, as proven by a scratch assay on fibroblasts [19]

Considering that together with persisting inflammation and microbial colonization, a further issue to be addressed in post-extraction socket healing is related to the reduction in alveolus volume, which physiologically occurs following extraction; the objective of this work was to propose a therapeutic system capable of supporting alveolar healing in a comprehensive manner. Generally, to minimize the phenomenon of alveolar volume reduction, it is possible to use a bone substitute such as hydroxyapatite (HXA; Ca10(PO4)6(OH)2). At present, HXA is the mainly used bone substitute as it is able to create a close bond with the bone tissue, while also promoting osteoconduction. Moreover, it is easily bioabsorbable and has no negative effects on the organism [25].

Therefore, the final aim of this work is to create a hybrid solid nanocomposite, containing HXA and the already characterized mLNP loaded with RSV, both being dispersed into a hydrophilic matrix, which could be directly inserted inside the post-extraction socket to promote the wound healing process.

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Materials

Trans-Resveratrol (RSV) and 18-β-Glycyrrhetinic Acid (GA) were purchased from A.C.E.F. spa (Fiorenzuola D’Arda, Piacenza, Italy). Labrasol® (LB) was kindly supplied by Gattefossé (Lyon, France). Glyceryl Monostearate 55–60 (GMS) was obtained from Farmalabor (Canosa di Puglia, Italy). Menthol (ME), Polyethylene glycol 6000 (PEG6K), Polyethylene glycol 10000 (PEG10K), Hyaluronic acid sodium salt (HA), Carboxymethylcellulose (CMC), Polyvinylalcohol (PVA), Polyvinylpyrrolidone K90 (PVP K90), and Polyvinylpyrrolidone K30 (PVP K30) were purchased from Carlo Erba Reagents (Milan, Italy). The Hydroxyapatite (HXA) was purchased as SpherHA granules (dense granules, 0.5–1 mm, SHA-D1006) from Tiss’You srl (Domagnano, Republic of San Marino). β-cyclodextrin (Kleptose)(β-CD) was obtained from Roquette Italia S.P.A (Cassano Spinola, AL, Italy). Pluronic F-127 was supplied by Sigma Aldrich (Milan, Italy). Trifluoroacetic acid (TFA) was obtained from Merck (Darmstadt, Germany). Sodium Citrate Dihydrate and Citric Acid Monohydrate were supplied by VWR International (Leuven, Belgium). The citrate buffer pH 5.5 was prepared by dissolving 3.024 g of sodium citrate dihydrate and 0.636 g of Citric Acid Monohydrate in 1 L of ultrapure water.

All chemicals and solvents (analytical grade) were purchased from Carlo Erba Reagents (Milan, Italy) and were used without any further purification.

De Caro, V.; Tranchida, G.; La Mantia, C.; Megna, B.; Angellotti, G.; Di Prima, G. Hybrid Nanocomposite Mini-Tablet to Be Applied into the Post-Extraction Socket: Matching the Potentialities of Resveratrol-Loaded Lipid Nanoparticles and Hydroxyapatite to Promote Alveolar Wound Healing. Pharmaceutics 2025, 17, 112. https://doi.org/10.3390/pharmaceutics17010112