Moringa oleifera Use in Maintaining Oral Health and Its Potential Use in Regenerative Dentistry

Phytomedicine refers to the use of naturally derived products to cure and mitigate human conditions. Natural products have the advantages of causing minimum side effects, being biocompatible, available, and economical, with a wide array of biological activities. Reports have described the use of natural products with antimicrobial and anti-inflammatory properties to treat oral conditions and promote wound healing. Moringa oleifera, known as the “drumstick” or “horseradish” tree, is believed to have medicinal properties regarding a range of medical conditions, though there is limited information on its use in oral medicine. This narrative review focuses on the use of Moringa extracts in the management of oral conditions, including oral infections, inflammatory conditions, the remineralization of hard tissues, oral wound healing, and tissue regeneration, drawing from both in vitro and in vivo studies which indicate that the potential of Moringa extracts in supporting dentin-pulp regeneration after caries or trauma is worthy of more careful consideration.


Introduction
Phytomedicine refers to a body of therapeutic knowledge based on natural extracts of plants, animals, fungi, and minerals [1].Reports are available describing the use of "natural" products in the treatment of gingivitis and periodontitis, promoting periodontal wound healing after traumatic injury or pathologic disease [2], the healing of oral ulcers, pulp and periapical treatment [3], and therapeutic use for oral cancer both in vitro and in vivo [4][5][6].
Moringa oleifera (M.oleifera) is a mineral and vitamin rich, nutritious, and medicinally important tree species, belonging to the family Moringaceae.Common names include drumstick, ben oil, or horseradish tree [7].Its cultivation is widespread in the Himalayan foothills, with subsequent introduction to south east and west Asia, Arabian lands, east and west Africa, some states of the USA, and South America [8].M. oleifera is considered to be a "super" plant by virtue of its exceptional properties to combat various illnesses in the human body [9].
Te phytochemistry of M. oleifera reveals diferent classes of compounds with the potential to confer health benefts.Te presence and amount of these compounds vary according to the geographical location of cultivation, soil type, climatic conditions, and sun exposure [10].Derivatives of this plant may also vary according to extraction methods, especially the use of diferent solvents including methanol, ethanol, and water [11,12].Numerous bioactive compounds can be extracted from this plant: favonoids, phenolic acids, glucosinolates, saponins, tannins, steroids, alkaloids, and terpenes [13].Te favonoids present include rutin, quercetin, rhamnetin, kaempferol, apigenin, and myricetin [14], and these compounds have numerous therapeutic efects including anti-infammatory, antioxidant, antibacterial, and hypoglycemic [15] and help in wound healing and tissue regeneration [16].In addition, numerous phenolic acids are present in leaves, including cafeic acid, chlorogenic acid, gallic acid, and others [13,17].Furthermore, all parts of the M. oleifera plant contain glucosinolates, giving the plant its potential cancer chemopreventive [18,19], hypotensive and antibacterial [20,21], and potential activities against neurodegenerative diseases [22].Tannins are proposed to contribute to anticancer, antimicrobial, and antihepatoxic activities [23].Moringa also contains alkaloids with calciumchannel blocking activity used in antihypertensive therapy [24].Moreover, the plant leaf and seed extracts were reported to be potent natural coagulants in the water purifcation process [25].
Diferent traditional and nontraditional methods have been developed for extraction of the active components from M. oleifera leaves and seeds [26].Traditional solvent extraction using ethanol and methanol is efective for production of M. oleifera leaf extract with high yield and the highest contents of phenolic compounds and highly antioxidant favonoids [11,12].Other modern nontraditional methods, such as ultrasound [27] and microwave-assisted [28] extraction, have also been employed.Tese diferent methods have been reported in the fabrication of various formulations of M. oleifera extracts to be used for therapeutic purposes.Examples of such products include the nanomicelle of seed oil to treat cancer [29], herbal nasal gels for the treatment of allergic rhinitis [30], alginate-pectin flm dressing containing extracts of M. oleifera for wound healing application [31], seed oil cream with anti-infammatory properties [32], and granules of Moringa extract as antiarthritic therapy [33].
Recently, active molecules have been incorporated into dental materials with the objective of simulation of the dentin-pulp complex behavior to promote repair and regeneration and manage the infammatory process and the deposition of mineralized tissue [34].Terefore, the purpose of this narrative review is to present a comprehensive summary of the properties of M. oleifera extracts which may be benefcial in the regeneration of tissues and specifcally in the damaged dental pulp.

Search Strategy
An electronic search of PubMed and Science Direct databases was undertaken using keywords including combinations of "Moringa oleifera" with "dental," "dentistry," and "regeneration."Te search of articles published in English was conducted from January 2010 to July 2022.Te identifed articles were fltered according to their relevance to dental aspects and regeneration, and the bibliographies of relevant articles were manually searched for additional articles of interest.Te search was limited to original articles, research papers, case reports, and reviews of the literature.Materials published as letters to the editor, website entries, and social media posts were excluded.Te search resulted in collection of 113 initial potential articles, and after excluding duplicates (n � 17), the remaining articles were initially fltered by reading their titles, abstracts, and full texts to identify 29 articles that ftted the scope of the review.

Antimicrobial Activity of Moringa oleifera against Oral Pathogens
Te antibacterial activity of the methanolic extract of M. oleifera inhibited the growth of Streptococcus mutans, Streptococcus salivarius, Streptococcus mitis, Lactobacillus fermentum, Streptococcus anginosus, Streptococcus gordonii, Lactobacillus acidophilus, and Staphylococcus aureus using the in vitro well difusion method [35].Elgamily et al. [36] reported that extracts from diferent parts of Moringa (leaves, roots, seeds, and their mixture) showed antibacterial activity against Staphylococcus aureus and Streptococcus mutans growth, although there was no inhibition of oral fungus Candida albicans.In the same study, the incorporation of ethanolic extract from leaves with a passive tooth paste and mouth wash signifcantly increased their antimicrobial efectiveness.An in vitro study performed by Arévalo-Híjar et al. [37] found that methanolic extracts of M. oleifera leaves suppressed the growth of Enterococcus faecalis with a greater antibacterial efect in comparison to 2% chlorhexidine.Tese extracts showed minimal cytotoxic efects on cultured Madin-Darby canine kidney (MDCK) cells.Tis study suggested the possible use of these extracts as antimicrobial agents in root canal therapy.Aqueous and ethyl alcohol extracts from M. oleifera leaves were tested in vitro for their antimicrobial activity on bioflms from volunteer specimens against Streptococcus mutans in comparison to 70% ethanol.Both extracts displayed powerful antimicrobial efects and restrained cariogenic bioflm formation, suggesting that they could be employed as preventive measures for dental caries [38].Another study by Noushad et al. [39] demonstrated a signifcant antibacterial efect of aqueous extract of M. oleifera seeds against Enterococcus faecalis bacterial growth in comparison to 5.25% sodium hypochlorite by using an agar difusion test.
Tese reports demonstrated in vitro antibacterial activity of M. oleifera against caries-related bacteria and inhibition of artifcially grown cariogenic bioflms.It is important to note that some of these studies had no conventional controls for comparison and that the methods of evaluation may not ideally represent in vivo conditions.Further work is clearly required to optimize the extraction of active ingredients, identify and purify potentially useful compounds, and conduct more rigorous evaluation using test models that refect in vivo conditions.Tis should be combined with careful investigation of the safety and toxicity of these agents prior to their application.

Effects of Moringa in Oral Inflammatory Conditions
Dos Santos et al. [40] studied natural isothiocyanate from M. oleifera and its seven analogue molecules (semisynthetic of derivatives MC-D1, MC-D6, MC-D7, MC-D8, and MC-D9) in a TMJ infammatory hypernociception model in rats.Tey found that MC-D7 and MC-D9 were efective in reducing nociception and formalin-induced infammation, while MC-H was more efective against serotonin-induced hypernociception in comparison to indomethacin as a positive control.In another in vivo study of temporomandibular joint pain using male Wistar rats, the semisynthetic derivative MC-H exhibited antinociceptive and antiinfammatory efects when administered orally.Tis potential analgesic efect could be peripherally mediated by 2 Te Scientifc World Journal action of the heme oxygenase-1 (HO-1) pathway, as well as through inhibition of intercellular adhesion molecule levels while centrally by the activation of opioid receptors (µ and δ) [41].
In a study employing network pharmacology and molecular docking, phenolic compounds derived from M. oleifera leaf were investigated for antiperiodontitis efects both in an in vitro RAW 264.7 macrophage cell culture and in an in vivo ligature-induced periodontitis rat model.Moringa leaf extract achieved antiperiodontitis activity by regulating the p38α/MAPK14-OPG/RANKL pathway.Te extract also decreased serum proinfammatory cytokines and increased anti-infammatory cytokines and reduced alveolar bone resorption within the in vivo model [42].Herbal lozenges composed of M. oleifera leaf and Cyanthillium cinereum (Less.)H. Rob extracts showed an antioxidant activity with powerful efects in reducing gingivitis and oral infammation in a double-blinded, randomized, controlled clinical trial of smoker volunteers [43].Mouthwashes containing ethanolic leaf extracts of Citrus hystrix, M. oleifera, and Azadirachta indica have been shown to decrease the gingival index and plaque index with a reduction in both Staphylococcus and Candida species in gingivitis subjects after 14 days in comparison with chlorhexidine gluconate [44].Tus, these new mouthwashes could reduce gingival infammation and be a complementary treatment in microbial-induced gingivitis.A randomized clinical crossover study by Duarte et al. [45] found that commercially available Moringa-based dentifrice was associated with a signifcant reduction in the gingivitis and plaque index compared with miswak dentifrice in subjects with mild-to-moderate gingivitis.However, this study has several limitations, such as short experimental duration, small sample size (only 20 subjects), and absence of a control group.
It can be concluded that diferent parts of M. oleifera demonstrate anti-infammatory properties with the potential to reduce proinfammatory mediators and control gingivitis.Tese studies were again limited by the absence of conventional controls.However, derivatives of medicinal plants such as M. oleifera are worthy of further exploration as alternative anti-infammatory agents in the management of a range of mucosal, periodontal, and possibly pulpal infammatory conditions.

Effect of Moringa Extracts in Enamel Remineralization
Only one related study was found, which investigated the remineralization efect of lyophilized extract from M. oleifera leaves incorporated within plain dental varnish on in vitro-induced enamel lesions [46].Tis study found that the formulated varnish reinforced mineralization of the lesions with complete reestablishment of the enamel surface in comparison to plain and fuoride varnishes.Tis could be due to the high mineral contents (Ca and P) and large range of amino acids within the M. oleifera extract which may play a role in peptide-guided remineralization of enamel surface lesions.

Regeneration and Healing Potential of Moringa oleifera
Extracts from M. oleifera have been investigated for their capacity to induce bony growth and enhance the integration of intraosseous implants.Tese studies are based on the assertion that extracts of this plant leaf can act as osteoconductive and osteoinductive agents when combined with demineralized freeze-dried bovine bone xenograft.Tis formula was evaluated in a Cavia cobaya animal model which revealed superior bone regeneration and socket preservation after tooth extraction associated with signifcant expression of transforming growth factor-beta 1 (TGF-β1) and osteocalcin [47].Extracts of acemannan and M. oleifera, in the form of hydrophilic gel, were used to coat titanium dental implants before implanting within the tibia and femur of rabbits.Tese coats were able to produce hydrophilic implant surfaces which improved bone to implant contact, stimulated new bone formation, and reduced infammation, fbrosis, and degenerative and necrotic changes within the newly formed bone [48].Tis bioactive behavior of the M. oleifera extract inducing bone regeneration still needs further investigations in medically compromised models such as diabetes mellitus, postradiation therapy, and osteoporosis where new bone formation may be hindered by the systemic condition of patients.
Another study was conducted to identify the healing efect of the M. oleifera leaf extract in gel form with two concentrations (2% and 4%) on oral excisional wounds in the palate of Sprague-Dawley rats.Povidone iodine gel (10%) was used as a control [49].Histopathological sections from Moringa-treated samples revealed a faster rate of fbroblast and collagen fber deposition in the course of the initial phase of palatal wound healing, whereas a slower rate was seen in the control group.Tis suggested that the M. oleifera leaf extract could accelerate wound healing in the oral cavity by virtue of its antibacterial and anti-infammatory capacities.However, actual mechanisms of such actions are currently unclear and require further clarifcation.Also, the optimal concentration and physical stability of the formulation to be used need to be explored before its clinical application.
An isothiocyanate extracted from M. oleifera seeds named moringin was reported in an in vitro study to stimulate human periodontal ligament stem cells (hPDLSCs) into neural progenitor, osteogenic, and adipogenic activation, with no sign of tumorgenic activation [50].Another study explored gene deregulation in the mitophagy of hPDLSCs pretreated with moringin.Most of the genes showed signifcant downregulation in comparison to untreated cells.Accordingly, this advantageous efect seemed to improve hPDLSCs for potential application in stem cell therapies, specifcally for disorders that involve oxidative stress as pathological process mechanisms, such as neurodegenerative diseases [51].
Aqueous extracts of M. oleifera were found capable of enhancing proliferation, viability, and migration of human dermal fbroblast cells in vitro [16].M. oleifera aqueous Te Scientifc World Journal Table 1: Green synthesis of nanoparticles from M. oleifera extracts.
Aqueous extracts of M. oleifera fower showed a noticeable ability in promoting rat fbroblast proliferation, bone marrow-derived mesenchymal stem cell diferentiation, and angiogenesis, with no observed efect on abnormal cancer cell formation, whereas aqueous leaf extract showed lesser ability to stimulate cell proliferation but had prominent cytotoxicity on Mcf7 breast cancer cell lines [53].In another study, the latter extract was demonstrated to decrease skin wound areas in diabetic rats and increase macrophage and fbroblast population, leading to a high level of wound contraction in comparison to a control group treated with normal saline [54].n-Hexane extract and hydrogels of M. oleifera seeds showed wound healing capacity and accelerated regeneration of tissue with decreased infammatory cells and increased vascularity of the newly formed skin in excision and incision wound-healing mouse models [55].Topical contact of the wound surface with a gel layer of M. oleifera leaf extract-loaded chitosan microparticles formed a protective coat above the wound with sustained release of M. oleifera active constituents.Tis was reported to stimulate keratinocyte growth in an ex vivo pig skin model, which suggested the potential of this combination in exuding wound treatment [56].Dressing containing 0.5% M. oleifera leaf extract-loaded flm showed satisfactory drug release properties to facilitate the healing process of wounds in streptozotocin-induced diabetic type II rat models [57].Furthermore, M. oleifera gum polysaccharide carbopol-based hydrogel dressing formulated by [58] was able to maintain a moist environment over the wound bed to enhance re-epithelialization and wound healing.
Alginate-PEG methyl ether methacrylate-Moringa oleifera-aloe vera scafolds showed notable increases in the number of viable human skin fbroblast cells, compared to control scafolds used in cell-free and cell-based assays [59].
A phenolic glycoside in M. oleifera seeds named 1-O-(4hydroxymethylphenyl)-α-L-rhamnopyranoside MPG was tested for its hepatoprotection against carbon tetrachloride CCl4-induced hepatotoxicity in L02 cell lines (immortalized human hepatocyte cell line L02) and mice.Both in vitro and in vivo studies showed that MPG can notably protect the liver against acute hepatotoxicity induced by CCl4 through its antioxidant ability, regulating infammatory mediators and antiapoptosis [60].
M. oleifera part extracts were used as a stabilizing and reducing/oxidizing agent in the biosynthesis of metal and metal oxide nanoparticles [61], with the beneft of this plant's phytochemicals, such as tannins, favonoids, saponins, and alkaloids along with vitamins [62].Some of the metals and metal oxides used in green synthesis of nanoparticles utilizing M. oleifera extracts are listed with their publications in Table 1.
Green gold nanoparticle synthesized using methanolic leaf extracts of M. oleifera as stabilizing agents was proved to be biologically active, less cytotoxic against blood cells in vitro, and helps in regeneration of neuronal cells in a Swiss albino mouse model based on the observation of their efect on brain architecture [75].According to the authors, biosynthesis of these nanoparticles was achieved using gold chloride HAuCl 4 .3H 2 O, with the addition of methanolic extract of M. oleifera leaf as the stabilizing agent.Te mixture was kept under continuous stirring at 75 °C and then fltered and centrifuged to remove unwanted heavy materials [75].In another study, M. oleifera seed polysaccharide was used to produce nanoparticles.Te extract was mixed with AgNO 3 solution and distilled water, and then, the mixture was centrifuged to completely remove unbounded compounds.A pellet was obtained that was dried in an oven at 80 °C to produce nanoparticles.Tese nanoparticles exhibited no cytotoxic efect on mouse fbroblast L929 cells and promoted their migration in vitro, with antibacterial efects against wound infection pathogens.Furthermore, these nanoparticles facilitated infected wound healing with excellent biocompatibility, eliminating pathogenic bacteria and reducing infammation and maximum epithelization after 9 days of administration in an in vivo male rat wound-healing model [76].
Tere is increasing evidence to demonstrate that M. oleifera extracts from diferent parts of the plant have the ability to promote the proliferation of various normal stem cells and other cells [77], promote wound healing and tissue regeneration, and even promote angiogenesis.Tese efects, combined with their antimicrobial properties, suggest that extracts of M. oleifera may be worthy of further investigation as therapeutic agents in the regeneration of the dentin-pulp complex after dental injury.

Conclusion
Te present review summarized published information on the potential of Moringa oleifera extracts to control infections and support tissue healing in a range of in vitro and in vivo models.At this stage, the evidence is limited as many published laboratory studies do not exactly refect clinical conditions.Diverse mechanisms have been suggested for these biological actions, including antioxidant, neuroprotective, antiinfammatory, and antimicrobial activities.Tese may in turn have the potential to be induced by a range of biologically active derivatives including polyphenols, favonoids, glycosides, glucosinolates, and many others.Although direct evidence is still limited, the potential of such plant derivatives in supporting wound and tissue regeneration after dental injury is worthy of further investigation.