**11. Characterization of the DPSCs**

impact of vitamin K on mir-760 on SP1 and mir-149 with reciprocal regulatory loops), and the mir-760 "junction" between RUNX1, SP1, SP3, and PPARG versus DIO2, ADIPOQ, and RUNX2, which are all part of a mutually interacting network regulated by vitamin K2

Finally, it should be emphasized that vitamin K2 (MK-7) upregulates Amelx and DKK1 in tooth germs, the former is instrumental in the building and maintenance of tooth enamel, and thus their resilience toward enamel erosion; the latter, DKK1 (dickopf1 WNT signaling pathway inhibitor 1), takes part in the modulation of osteoclast-related bone degradation, and in this context, the healthy transition between osteoclast-induced resorption and renewal of

**10. Pertinent question: is the dental filling material toxic to the living** 

Monomers from methacrylate based dental materials both prior to and post polymerization have demonstrated adverse effects both *in vitro* and *in vivo* in terms of cytotoxicity [68], mutagenicity/genotoxicity [69–72], negative effects on fertility [73], xenoestrogenicity [74–76], and allergy induction [77]. The degree of cytoxicity will vary from the type assay used, materials

It is most pertinent to perform *in vitro* cytotoxicity testing on cells from cell types and tissues relevant to the area of *in vivo* placement of dental materials [79]. Recent studies on elution of monomers available in both dental composites and methacrylate- and epoxy-based root canal sealers looked at reactions in submandibular salivary gland acinar cells for the evaluation of cytotoxicity, cell proliferation, and apoptosis [69, 80]. The findings in such studies are of great interest and importance, but the authors of one of these studies [80] stated that their model would have been more realistic had they utilized human primary cells from direct target tissue. Tissues that often share the closest proximity to dental fillings are certainly not

The pulp is a loose connective tissue within a nonresilient capsule of dentin and enamel. Pulpal inflammation is considered a protective mechanism and can either be of an acute or chronic nature. Acute and chronic responses are related to the "magnitude and duration of the insult [81]." Inflammation will inevitably cause vasodilation, increased vessel permeability which in turn will result in relatively large changes in tissue pressure [82]. Bacterial infection is the most common reason for pulpal inflammation, but any insult or stimuli will most probably result in a response. It is an established fact that many of the constituents in dental adhesive resin are cytotoxic [81], and the difference in cytoxicity varies among commercial materials commonly used by public dentists in Norway [unpublished results in a report to the

This project aims at elucidating the cellular effects of "leachables" (residual monomers) from dental filling materials exerted on dental pulp stem cells (DPSCs) *ex vivo*. It is important to

**tooth? Contemplations on the making of live and artificial teeth**

salivary glands but rather gingiva, mucosa, and, in particular, pulp tissues [79].

tested, time intervals for testing, and cell types tested [78].

Norwegian National Directorate of Health].

odontoblasts/osteoblasts.

142 Vitamin K2 - Vital for Health and Wellbeing

bone tissue with microcracks [67].

Dental specimens were obtained from extraction after signed informed consent, and the pulp was exposed by cutting the tooth, while maintaining sterile conditions: The enamel of the tooth crown is partially cut, following the sagittal plane, applying a diamond bur. Thereafter, the cut is completed using a piezoelectric ultrasound scalpel to avoid overheating of the tissue. The pulp is then treated collagenase and dispase for 1 hour at 37°C, and then incubated in a bioSpherix chamber under normoxic conditions [85].

Phenotyping of the DPSCs yielded a CD-profile very much like the one seen for bone marrow mesenchymal stem cells (BM-MSCs) with an approximately identical percentage of cells expressing CD10 (CALLA), CD 13 (Aminopeptidase N), CD29 (β1-integrin), CD44 (H-CAM, Pgp-1), CD49acd (VLA-1,3,4 = α1,3,4-integrin), CD54 (I-CAM-1), CDw90 (THY-1), CD105 (Endoglin, TGFβ-R), CD140b (PDGF-Rb), CD146 (M-CAM), CD147 (Neurothelin/basigin), CD166 (Alcam, CD6-ligand), and also comparable amounts of GD2 (Neural ganglioside).
