Tissue Biocompatibility and Antimicrobial Properties of Sympathomimetic Nasal Solutions for Potential Use in Dental Pulpal Management

BackgroundThis study evaluated the antibacterial properties of a potential pulpal medicament, a nasal solution with oxymetazoline (NS-OXY, 0.05%), against a dentinal caries pathogen.MethodsUsing a disc diffusion susceptibility test (n = 6), Rothia dentocariosa was grown on brain–heart infusion (BHI) agar plates and exposed to OXY (0.05%), benzalkonium chloride (BKC-0.025%), OXY-NS (with OXY-0.05% and BKC), ferric sulfate (20%; ViscoStat), and distilled water (DI). This was followed by exposure of an artificial dental caries model with sheep blood to simulate the clinical pulpotomy procedure. An antibacterial broth inhibition test was conducted by adding the test samples in BHI broth at 37 ± 0.5 °C in an aerobic chamber.ResultsIn the disc diffusion test, NS-OXY and BKC had the largest zone of inhibition (ZOI) measuring 14.42 mm (± 1.62) and 18.92 mm (± 4.14) respectively, indicating antibacterial activity. Ferric sulfate demonstrated a smaller ZOI, while OXY alone had no ZOI. The antibacterial broth test showed antibacterial effects with stable OD and pH levels for test samples containing BKC (0.025%) and diluted NS-OXY (0.01%) for up to 20 h. DI- and OXY-treated samples showed an increase in OD, indicating an increase in bacterial count and a concurrent drop in pH. BKC treatment statistically (P < 0.05) reduced polyP extracts, which may contribute to blood clot formation. NS-OXY demonstrated antibacterial properties, likely due to the addition of BKC to Rothia dentocariosa. NS-OXY showed concentration dependent biocompatibility with dental pulp stem cells while FS was cytotoxic at the same dilution.ConclusionThese antimicrobial properties, together with OXY’s hemostatic effects, suggest the potential off-label use of NS-OXY during a pulpotomy procedure in primary and permanent teeth. This study provides support for potential future clinical trials of repurposing FDA-approved drugs consisting of oxymetazoline and benzalkonium chloride for dental and other similar applications.Featured applicationAn over-the-counter nasal solution containing oxymetazoline has the potential to be used off-label to manage surgical bleeding from dental pulp exposures and provide antimicrobial properties against Rothia dentocariosa, a model gram-positive bacteria associated with deep dentinal caries.

Objective: The objective of our study was to successfully isolate progenitor cells from dental pulp and umbilical tissue and perform a comparative investigation of their potential transdifferentiation into osteo- and neuronal-like cells. Methods: Progenitor cells were harvested from dental pulp tissue as well as cord tissue and cultured through explant culture method over the span of 4 weeks. Image-based cytometric analysis was done to determine the cell viability along with phenotypic analysis to validate the occurrence of stem cell surface markers such as CD13, CD29, CD31, CD34, CD45, CD73, CD90, CD105, HLA-DR, and HLA-ABC. After culturing mesenchymal progenitor cells, osteogenic and neurogenic differentiation potential of both tissue sources was studied. The cells were seeded in two different surfaces tissue culture treated dishes and titanium sheets and cultured along with osteogenic differentiation medium (for 28 days) and neurogenic differentiation medium (for 5 days). The osteogenic potential of the progenitor cells were checked with the detection of calcium deposits by Vonn Kossa staining and PCR studies were done to confirm the presence of osteogenic genes like BMP2, HDAC1, HNF1A. The neurogenic potential of the progenitors were phenotypically determined by the observation of neuronal cells in the culture medium. Post differentiation PCR studies were done to confirm the presence of neuronal genes like NESTIN, AGRIN, MAG, DAPDH, NF-M. Findings: Progenitor cells extracted from cord tissue and dental pulp were positive for markers such as CD13, CD29, CD73, CD90, and CD105 and were found negative for markers such CD31, CD34, CD45, and HLA-DR. Progenitors obtained from dental pulp tissue showed a higher expression of cell surface markers indicating a stronger mesenchymal lineage. After culturing progenitor cells in osteogenic differentiation specific medium, these cells were successfully differentiated into cells of osteogenic lineage. Within 28 days of culture calcium deposits were detected by Von kossa staining. The differentiated cells were also found positive for osteogenic markers such as BMP2, HDAC1, HNF1A.In neural differentiation, post day 5 of culture neurospheres of varying sizes were observed floating in the culture medium. The fraction of the cells differentiated into osteogenic and neurogenic lineages were higher in progenitor cells derived from dental pulp in comparison with umbilical cord tissue. The higher potentialityof progenitor cells derived from dental pulp for neurogenic trans-differentiation could be explained by the fact that human adult dental pulp stem cells residing within the perivascular niche are thought to originate from the migrating cranial neural crest cells. There was no difference observed in the osteogenic and neurogenic differentiation capabilities of mesenchymal cells when plated in a plastic dish or titanium surface. Improvement: In-depth studies needs to be carried out on progenitor cells from dental pulp tissues in order to enhance the clinical efficacy of stem cells based therapies.

Objective: The objective of our study was to successfully isolate progenitor cells from dental pulp and umbilical tissue and perform a comparative investigation of their potential transdifferentiation into osteo- and neuronal-like cells. Methods: Progenitor cells were harvested from dental pulp tissue as well as cord tissue and cultured through explant culture method over the span of 4 weeks. Image-based cytometric analysis was done to determine the cell viability along with phenotypic analysis to validate the occurrence of stem cell surface markers such as CD13, CD29, CD31, CD34, CD45, CD73, CD90, CD105, HLA-DR, and HLA-ABC. After culturing mesenchymal progenitor cells, osteogenic and neurogenic differentiation potential of both tissue sources was studied. The cells were seeded in two different surfaces tissue culture treated dishes and titanium sheets and cultured along with osteogenic differentiation medium (for 28 days) and neurogenic differentiation medium (for 5 days). The osteogenic potential of the progenitor cells were checked with the detection of calcium deposits by Vonn Kossa staining and PCR studies were done to confirm the presence of osteogenic genes like BMP2, HDAC1, HNF1A. The neurogenic potential of the progenitors were phenotypically determined by the observation of neuronal cells in the culture medium. Post differentiation PCR studies were done to confirm the presence of neuronal genes like NESTIN, AGRIN, MAG, DAPDH, NF-M. Findings: Progenitor cells extracted from cord tissue and dental pulp were positive for markers such as CD13, CD29, CD73, CD90, and CD105 and were found negative for markers such CD31, CD34, CD45, and HLA-DR. Progenitors obtained from dental pulp tissue showed a higher expression of cell surface markers indicating a stronger mesenchymal lineage. After culturing progenitor cells in osteogenic differentiation specific medium, these cells were successfully differentiated into cells of osteogenic lineage. Within 28 days of culture calcium deposits were detected by Von kossa staining. The differentiated cells were also found positive for osteogenic markers such as BMP2, HDAC1, HNF1A.In neural differentiation, post day 5 of culture neurospheres of varying sizes were observed floating in the culture medium. The fraction of the cells differentiated into osteogenic and neurogenic lineages were higher in progenitor cells derived from dental pulp in comparison with umbilical cord tissue. The higher potentialityof progenitor cells derived from dental pulp for neurogenic trans-differentiation could be explained by the fact that human adult dental pulp stem cells residing within the perivascular niche are thought to originate from the migrating cranial neural crest cells. There was no difference observed in the osteogenic and neurogenic differentiation capabilities of mesenchymal cells when plated in a plastic dish or titanium surface. Improvement: In-depth studies needs to be carried out on progenitor cells from dental pulp tissues in order to enhance the clinical efficacy of stem cells based therapies.

Successful isolation of human dental pulp stem cells (hDPSCs) has been documented at least 120h after tooth extraction. Viable hDPSCs have been isolated chiefly from cryopreserved healthy molar teeth and their undigested dental pulp tissue. Isolation of hDPSCs from diseased but vital teeth after cryopreservation has not been reported. This study aimed to isolate hDPSCs from cryopreserved diseased but vital teeth of various tooth types. Fifty tooth samples were divided into group A (n = 20) - freshly derived dental pulp tissues, group B (n = 20) - liquid nitrogen (liq N(2) )-stored dental pulp tissues and group C (n = 10) - liq N(2) -stored intact teeth. The success rate for hDPSCs isolation was 100% for groups A and B and only 20% for group C. hDPSCs from all groups demonstrated self-renewal properties and similar multipotent potential characteristics of adipogenic, chondrogenic and osteogenic differentiation. In addition, hDPSCs showed high expression of bone-marrow mesenchymal stem-cell markers (CD29, CD90 and CD105) and very low expression of specific hematopoietic cells markers (CD14, CD34 and CD45). Our results indicate that hDPSCs isolated from diseased but vital teeth of various tooth types can be stored in liq N(2) for future usage.

Reviewing the literature, hepatic differentiation of human dental pulp stem cells (hDPSCs) from cryopreserved dental pulp tissues of vital extracted teeth with disease has not been studied. This study is aimed to evaluate the hypothesis that hDPSCs from cryopreserved dental pulp tissues of vital extracted teeth with disease could possess potential hepatic differentiation. Forty vital extracted teeth with disease recruited for hDPSCs isolation, stem cell characterization and hepatic differentiation were randomly and equally divided into group A (liquid nitrogen-stored dental pulp tissues) and group B (freshly derived dental pulp tissues). Samples of hDPSCs isolated from groups A and B but without hepatic growth factors formed negative controls. A well-differentiated hepatocellular carcinoma cell line was employed as a positive control. All the isolated hDPSCs from groups A and B showed hepatic-like differentiation with morphological change from a spindle-shaped to a polygonal shape and normal karyotype. Differentiated hDPSCs and the positive control expressed hepatic metabolic function genes and liver-specific genes. Glycogen storage of differentiated hDPSCs was noted from day 7 of differentiation-medium culture. Positive immunofluorescence staining of low-density lipoprotein and albumin was observed from day 14 of differentiation-medium culture; urea production in the medium was noted from week 6. No hepatic differentiation was observed for any of the samples of the negative controls. We not only demonstrated the feasibility of hepatic-like differentiation of hDPSCs from cryopreserved dental pulp tissues of vital extracted teeth with disease but also indicated that the differentiated cells possessed normal karyotype and were functionally close to normal hepatic-like cells. Copyright © 2013 John Wiley & Sons, Ltd.

Guanylate-binding protein 5 (GBP5) is an interferon (IFN)-inducible GTPase that plays a crucial role in the cell-autonomous immune response against microbial infections. In this study, we investigated the immunoregulatory role of GBP5 in the pathogenesis of dental pulpitis. Gene-set enrichment analysis (GSEA) was utilized to evaluate the IFN-γ signalling pathway, and the differential expression of GBP mRNA in normal versus inflamed dental pulp tissues was screened, based on Gene Expression Omnibus (GEO) datasets associated with pulpitis. Both normal pulp tissues and inflamed pulp tissues were used for experiments. The expression of IFNs and GBPs was determined by qRT-PCR. Immunoblotting and double immunofluorescence were performed to examine the cellular localization of GBP5 in dental pulp tissues. For the functional studies, IFN-γ priming or lentivirus vector-delivered shRNA was used to, respectively, overexpress or knock down endogenous GBP5 expression in human dental pulp stem cells (HDPSCs). Subsequently, LPS was used to stimulate HDPSCs (overexpressing or with knocked-down GBP5) to establish an invitro model of inflammation. qRT-PCR and ELISA were employed to examine the expression of proinflammatory cytokines (IL-6, IL-8 and IL-1β) and cyclooxygenase 2 (COX2). Every experiment has three times of biological replicates and three technical replicates, respectively. Statistical analysis was performed using the Student's t-test and one-way ANOVA, and a p-value of <.05 was considered statistically significant. GSEA analysis based on the GEO dataset revealed a significant activation of the IFN-γ signalling pathway in the human pulpitis group. Among the human GBPs evaluated, GBP5 was selectively upregulated in inflamed dental pulp tissues and predominantly expressed in dental pulp cells. Invitro experiments demonstrated that IFN-γ robustly induced the expression of GBP5 in HDPSCs. Knockdown of GBP5 expression in HDPSCs significantly amplified the LPS-induced upregulation of inflammatory mediators (IL-6, IL-8, IL-1β and COX2) both with and without IFN-γ priming. Our findings demonstrated that GBP5 partook in the pathogenesis of dental pulpitis. The involvement of GBP5 in pulpitis appeared to coordinate the regulation of inflammatory cytokines. Knockdown of GBP5 contributed to the exacerbation of LPS-mediated inflammation.

Dental pulp regeneration is significantly aided by human dental pulp stem cells (hDPSCs). An increasing number of studies have demonstrated that circular RNAs (circRNAs) are crucial in the multidirectional differentiation of many mesenchymal stem cells, but their specific functions and mechanisms remain unknown. This work aimed at elucidating the molecular mechanism by which hsa_circ_0001599 works in hDPSCs during odontogenic differentiation. The expression of hsa_circ_0001599 in hDPSCs and dental pulp tissue was determined by using quantitative real-time PCR (qRT‒PCR). The role of hsa_circ_0001599 in the odontogenic differentiation of hDPSCs and its mechanism were studied using a variety of in vivo and in vitro assessments. The odontogenic differentiation of hDPSCs was facilitated by the overexpression of hsa_circ_0001599, which activated the PI3K/AKT signalling pathway in vitro. In vivo, hsa_circ_0001599 can promote the formation of new dentin-like structures. Mechanistically, hsa_circ_0001599 enhanced ITGA2 expression by sponging miR-889-3p. Furthermore, hsa_circ_0001599 interacts with the methylation reader hnRNPA2B1, promoting hnRNPA2B1 translocation from the nucleus to the cytoplasm and increasing ITGA2 mRNA stability. This research revealed the important role of hsa_circ_0001599 in odontogenic differentiation. Thus, hDPSCs engineered with hsa_circ_0001599 have the potential to be effective therapeutic targets for dental pulp repair and regeneration.

As the most important organs of occlusion, teeth are subjected to a variety of mechanical stresses. These stresses are transmitted into the dental pulp tissue and affect the dental pulp stem cells. In this study, human dental pulp stem cells were isolated from human impacted third molars and their multilineage differentiation abilities were tested. Human dental pulp stem cells were then exposed to cyclic tensile stretch. The results showed that the cyclic tensile stretch inhibited the expression of osteogenic marker genes and proteins such as BMP-2, OCN and ALP. Simultaneously, odontogenic marker genes and proteins such as DSPP, DSP and BSP were also inhibited by the mechanical stress. It was concluded that cyclic tensile stretch inhibits the osteogenic and odontogenic differentiation of dental pulp stem cells.

Multiple side effects related to bleaching were found to occur in the dental pulp tissue, including decreased cell metabolism and viability. In this work we evaluated the in vitro diffusion capacity, cytotoxicity and biocompatibility of four commercial bleaching products on stem cells from human dental pulp (hDPSCs). Two commercial bleaching gels hydrogen peroxide-based (HP), Norblanc Office 37.5% (Nor-HP) and Opalescence Boost 40% (Opal-HP) were applied for 30 min to enamel/dentine discs. Another two gels from the same manufacturers, 16% carbamide peroxide-based (CP), Norblanc Home (Nor-CP) and Opalescence CP 16% (Opal-CP), were applied for 90 min. The diffusion of HP was analysed by fluorometry. Cytotoxicity was determined using the MTT assays, the determination of apoptosis, immunofluorescence assays and intracellular reactive oxygen species (ROS) level. Tissue inflammatory reactions were evaluated histopathologically in rats. Statistical differences were performed by one-way ANOVA and Bonferroni post-test (α < 0.05). Normon products showed lower cytotoxicity and diffusion capacity than the Ultradent products. A high intracellular ROS level was measured in hDPSCs after exposure to Opal-HP. Finally, a severe necrosis of both coronal and radicular pulp was observed with Opal-HP. Similar concentrations of hydrogen peroxide and carbamide peroxide in a variety of bleaching products exhibited different responses in cells and dental pulp tissue, suggesting that bleaching products contain unknown agents that could influence their toxicity.

Nell-1 promotes the neural-like differentiation of dental pulp cells

The dental pulp is a highly vascularized and innervated connective tissue composed of various cell types, including fibroblasts, odontoblasts, mesenchymal stem cells, neuronal, and endothelial cells. The interplay between these diverse cell populations is pivotal for dental pulp tissue homeostasis and regeneration after carious infections and traumatic tooth lesions. Despite the great clinical need, comprehensive in vitro models that accurately recapitulate the complexity of the dental pulp are still missing, hampering the development of novel, faster, and more effective therapies. In this study, an innovative “tooth‐on‐chip” microfluidic device is presented to emulate the composition and three‐dimensional structure of the dental pulp tissue in vitro. Co‐culture of human dental pulp stem cells, odontoblast‐like cells, endothelial cells, and trigeminal neurones in this miniaturized system successfully reproduced the structural organization and physiology of the dental pulp. The microfluidic device integrated various compartments that allowed the generation of complex vascular and neuronal networks, the formation of stem cell perivascular niches, and the formation of an odontoblast/dentine interface. The “tooth‐on‐chip” device represents a conceptual leap in replicating dental pulp physiology in vitro, offering a state‐of‐the‐art platform to study dental pulp physiology and pathology and serving as a benchmark to create more advanced tooth simulation systems.

Human adult dental pulp stem cells (DPSCs) reside within the perivascular niche of dental pulp and are thought to originate from migrating cranial neural crest (CNC) cells. During embryonic development, CNC cells differentiate into a wide variety of cell types, including neurons of the peripheral nervous system. Previously, we have demonstrated that DPSCs derived from adult human third molar teeth differentiate into cell types reminiscent of CNC embryonic ontology. We hypothesized that DPSCs exposed to the appropriate environmental cues would differentiate into functionally active neurons. The data demonstrated that ex vivo-expanded human adult DPSCs responded to neuronal inductive conditions both in vitro and in vivo. Human adult DPSCs, but not human foreskin fibroblasts (HFFs), acquired a neuronal morphology, and expressed neuronal-specific markers at both the gene and protein levels. Culture-expanded DPSCs also exhibited the capacity to produce a sodium current consistent with functional neuronal cells when exposed to neuronal inductive media. Furthermore, the response of human DPSCs and HFFs to endogenous neuronal environmental cues was determined in vivo using an avian xenotransplantation assay. DPSCs expressed neuronal markers and acquired a neuronal morphology following transplantation into the mesencephalon of embryonic day-2 chicken embryo, whereas HFFs maintained a thin spindle fibroblastic morphology. We propose that adult human DPSCs provide a readily accessible source of exogenous stem/precursor cells that have the potential for use in cell-therapeutic paradigms to treat neurological disease.

Success of Maxillary Alveolar Defect Repair in Rats Using Osteoblast-Differentiated Human Deciduous Dental Pulp Stem Cells

“Mirror, mirror on the wall, who’s the fairest of them all?“ This is certainly the most cited expression of the 21 century. We do not want to get old, and in this context a lot of hopeful are set on stem cell therapy for tissue regeneration: regeneration of skin, dermis, cartilage, bone, nervous sistem and so on. Most of all on our smile that it is our business card. Rapid and good regeneration of alveolar bone to support new dental implant is up to now ensured by the use adult mesenchymal stem cells. But are they able to conserve their stemness ability for all the age of the donors? In light of such consideration aim of the present work has been the study of the biological properties such as proliferation and stemness ability of human adult dental pulp stem cells (DPSC) in relation to the age of the donors and to the in vitro aging. Human dental pulps derived from adult subjects aged 16–over 66 years have been isolated and cultured in presence of differentiative medium. Results obtained confirmed a correlation between age and conservation of stemness during in vitro aging.

In general medicine, melatonin is known to enhance wound healing and promote stem cell differentiation. Its potential relevance in endodontics, however, remains underexplored. This scoping review aimed to systematically assess the available evidence on the effects of melatonin (a) on dental pulp tissue and (b) on human dental pulp stem cells (hDPSCs), particularly regarding cell proliferation and differentiation with regard to endodontics. A comprehensive literature search was conducted in PubMed, Clarivate Analytics' Web of Science and Scopus from inception to July 1, 2025, using Medical Subject Headings (MeSH terms) and supplemented by hand searching and screening of major subject journals. The initial search yielded 252 records, with one additional record identified through citation mining and relevant journal screening. A total of 22 studies met the inclusion criteria: 11 investigated melatonin's effect on dental pulp tissue regarding anti-inflammatory properties, treatment of pulpitis, wound healing and pulp capping and 11 examined its impact on hDPSCs in terms of cell proliferation and differentiation. The limited evidence obtained from laboratory and animal studies suggests a dose- and time-dependent influence of melatonin, though evidence is insufficient to establish optimal concentrations. (a) Melatonin demonstrates anti-inflammatory, antioxidant and anti-fibrinolytic effects on dental pulp tissue. (b) Melatonin has potential as a stem cell modulator by promoting odontogenic differentiation and may improve migration and proliferation of hDPSCs.