Biological Effects of PMMA and Composite Resins on Human Gingival Fibroblasts: An In Vitro Comparative Study.

The use of temporary resin for provisional restorations is a fundamental step to maintain the position of prepared teeth, to protect the pulpal vitality and the periodontal health as well as the occlusion. The present study aimed at evaluating the biological effects of two resins used in dentistry for temporary restorations, Coldpac (Yates Motloid) and ProTemp 4™ (3M ESPE ™), and their eluates, in an in vitro model of human gingival fibroblasts (hGFs). The activation of the inflammatory pathway NFκB p65/NLRP3/IL-1ß induced by the self-curing resin disks was evaluated by real-time PCR, Western blotting and immunofluorescence analysis. The hGFs adhesion on resin disks was investigated by means of inverted light microscopy and scanning electron microscopy (SEM). Our results suggest that hGF cells cultured in adhesion and with eluate derived from ProTemp 4™ (3M ESPE ™) resin evidenced a downregulation in the expression of the inflammatory mediators such as NFκB p65, NLRP3 and IL-1ß compared to the cells cultured with Coldpac (Yates Motloid) after 24 h and 1 week of culture. Furthermore, the cells cultured with ProTemp 4™ (3M ESPE ™) after 24 h and 1 week of culture reported a higher cell viability compared to the cells cultured with Coldpac (Yates Motloid), established by MTS cell analysis. Similar results were obtained when hGFs were placed in culture with the eluate derived from ProTemp 4™ (3M ESPE ™) resin which showed a higher cell viability compared to the cells cultured with eluate derived from Coldpac (Yates Motloid). These results highlighted the lower pro-inflammatory action and improved cell biocompatibility of ProTemp 4™ (3M ESPE ™), suggesting a better performance in terms of cells-material interaction.

Emerging Role of Oral Mesenchymal Stem/Stromal Cells and Their Derivates.

Mesenchymal stem/stromal cells (MSCs) have fewer ethical, moral, and safety problems in comparison with embryonic stem cells [...].

Enriched Graphene Oxide-Polypropylene Suture Threads Buttons Modulate the Inflammatory Pathway Induced by Escherichia coli Lipopolysaccharide.

Graphene oxide (GO), derived from graphene, has remarkable chemical-physical properties such as stability, strength, and thermal or electric conductivity and additionally shows antibacterial and anti-inflammatory properties. The present study aimed to evaluate the anti-inflammatory effects of polypropylene suture threads buttons (PPSTBs), enriched with two different concentrations of GO, in the modulation of the inflammatory pathway TLR4/MyD 88/NFκB p65/NLRP3 induced by the Escherichia coli (E. coli) lipopolysaccharide (LPS-E). The gene and the protein expression of inflammatory markers were evaluated in an in vitro model of primary human gingival fibroblasts (hGFs) by real-time PCR, western blotting, and immunofluorescence analysis. Both GO concentrations used in the polypropylene suture threads buttons-GO constructs (PPSTBs-GO) decreased the expression of inflammatory markers in hGFs treated with LPS-E. The hGFs morphology and adhesion on the PPSTBs-GO constructs were also visualized by inverted light microscopy, scanning electron microscopy (SEM), and real-time PCR. Together, these results suggest that enriched PPSTBs-GO modulates the inflammatory process through TLR4/MyD 88/NFκB p65/NLRP3 pathway.

Improved osteogenic differentiation by extremely low electromagnetic field exposure: possible application for bone engineering.

Human periodontal ligament mesenchymal stem cells (hPDLSCs) are a promising cell type model for regenerative medicine applications due to their anti-inflammatory, immunomodulatory and non-tumorigenic potentials. Extremely low-frequency electromagnetic fields (ELF-EMF) are reported to affect biological properties such as cell proliferation and differentiation and modulate gene expression profile. In this study, we investigated the effects of an intermittent ELF-EMF exposure (6 h/day) for the standard differentiation period (28 days) and for 10 days in hPDLSCs in the presence or not of osteogenic differentiation medium (OM). We evaluated cell proliferation, de novo calcium deposition and osteogenic differentiation marker expression in sham and ELF-EMF-exposed cells. After ELF-EMF exposure, compared with sham-exposed, an increase in cell proliferation rate (p < 0.001) and de novo calcium deposition (p < 0.001) was observed after 10 days of exposure. Real-time PCR and Western blot results showed that COL1A1 and RUNX-2 gene expression and COL1A1, RUNX-2 and OPN protein expression were upregulated respectively in the cells exposed to ELF-EMF exposure along with or without OM for 10 days. Altogether, these results suggested that the promotion of osteogenic differentiation is more efficient in ELF-EMF-exposed hPDLSCs. Moreover, our analyses indicated that there is an early induction of hPDLSC differentiation after ELF-EMF application.

Effects of growth hormone-releasing hormone receptor antagonist MIA-602 in mice with emotional disorders: a potential treatment for PTSD.

Anxiety and depression have been suggested to increase the risk for post-traumatic stress disorders (PTSD). A link between all these mental illnesses, inflammation and oxidative stress is also well established. Recent behavior studies by our group clearly demonstrate a powerful anxiolytic and antidepressant-like effects of a novel growth hormone releasing hormone (GHRH) antagonist of MIAMI class, MIA-690, probably related to modulatory effects on the inflammatory and oxidative status. In the present work we investigated the potential beneficial effects of MIA-602, another recently developed GHRH antagonist, in mood disorders, as anxiety and depression, and the possible brain pathways involved in its protective activity, in adult mice. MIA-602 exhibited antinflammatory and antioxidant effects in ex vivo and in vivo experimental models, inducing anxiolytic and antidepressant-like behavior in mice subcutaneously treated for 4 weeks. The beneficial effect of MIA-602 on inflammatory and oxidative status and synaptogenesis resulting in anxiolytic and antidepressant-like effects could be related by increases of nuclear factor erythroid 2-related factor 2 (Nrf2) and of brain-derived neurotrophic factor (BDNF) signaling pathways in the hippocampus and prefrontal cortex. These results strongly suggest that GHRH analogs should be tried clinically for the treatment of mood disorders including PTSD.

A Narrative Review: Gingival Stem Cells as a Limitless Reservoir for Regenerative Medicine.

The gingival tissue can be collected in an easy way and represent an accessible source to isolate gingival-derived mesenchymal stem cells (GMSCs). GMSCs are a subpopulation of dental-derived mesenchymal stem cells that show the mesenchymal stem cells (MSCs) features, such as differentiation abilities and immunomodulatory properties. Dental-derived stem cells are also expandable in vitro with genomic stability and the possibility to maintain the stemness properties over a prolonged period of passages. Moreover, several preclinical studies have documented that the extracellular vesicles (EVs) released from GMSCs possess similar biological functions and therapeutic effects. The EVs may represent a promising tool in the cell-free regenerative therapy approach. The present review paper summarized the GMSCs, their multi-lineage differentiation capacities, immunomodulatory features, and the potential use in the treatment of several diseases in order to stimulate tissue regeneration. GMSCs should be considered a good stem cell source for potential applications in tissue engineering and regenerative dentistry.

Role of ascorbic acid in the regulation of epigenetic processes induced by Porphyromonas gingivalis in endothelial-committed oral stem cells.

Periodontitis is a common inflammatory disease that affects the teeth-supporting tissue and causes bone and tooth loss. Moreover, in a worldwide population, periodontal disease is often associated with cardiovascular diseases. Emerging studies have reported that one of the major pathogens related to periodontitis is Porphyromonas gingivalis (P. gingivalis), which triggers the inflammatory intracellular cascade. Here, we hypothesized a possible protective effect of ascorbic acid (AA) in the restoration of the physiological molecular pathway after exposure to lipopolysaccharide derived from P. gingivalis (LPS-G). In particular, human gingiva-derived mesenchymal stem cells (hGMSCs) and endothelial-differentiated hGMSCs (e-hGMSCs) exposed to LPS-G showed upregulation of p300 and downregulation of DNA methyltransferase 1 (DNMT1), proteins associated with DNA methylation and histone acetylation. The co-treatment of AA and LPS-G showed a physiological expression of p300 and DNMT1 in hGMSCs and e-hGMSCs. Moreover, the inflammatory process triggered by LPS-G was demonstrated by evaluation of reactive oxygen species (ROS) and their intracellular localization. AA exposure re-established the physiological ROS levels. Despite the limitations of in vitro study, these findings collectively expand our knowledge regarding the molecular pathways involved in periodontal disease, and suggest the involvement of epigenetic modifications in the development of periodontitis.

The Effect of Liposomal Curcumin as an Anti-Inflammatory Strategy on Lipopolysaccharide e from Porphyromonas gingivalis Treated Endothelial Committed Neural Crest Derived Stem Cells: Morphological and Molecular Mechanisms.

Curcumin, a yellow polyphenol extracted from the turmeric root is used as a diet supplement. It exhibits anti-inflammatory, antioxidant, and antitumor properties by modulating different intracellular mechanisms. Due to their low solubility in water, the curcumin molecules must be encapsulated into liposomes to improve the bioavailability and biomedical potential. For the periodontal tissue and systemic health, it is essential to regulate the local inflammatory response. In this study, the possible beneficial effect of liposomes loaded with curcumin (CurLIP) in neural crest-derived human periodontal ligament stem cells (hPDLSCs) and in endothelial-differentiated hPDLSCs (e-hPDLSCs) induced with an inflammatory stimulus (lipopolysaccharide obtained from Porphyromonas gingivalis, LPS-G) was evaluated. The CurLIP formulation exhibited a significant anti-inflammatory effect by the downregulation of Toll-like receptor-4 (TLR4)/Myeloid differentiation primary response 88 (MyD88)/nuclear factor kappa light chain enhancer of activated B cells (NFkB)/NLR Family Pyrin Domain Containing 3 (NLRP3)/Caspase-1/Interleukin (IL)-1ß inflammation cascade and reactive oxygen species (ROS) formation. Moreover, the exposure to LPS-G caused significant alterations in the expression of epigenetic modifiers, such as DNA Methyltransferase 1 (DNMT1) and P300, while the CurLIP treatment showed physiological expression. Overall, our in vitro study provides novel mechanistic insights into the intracellular pathway exert by CurLIP in the regulation of inflammation and epigenetic modifications.

Oral Bone Tissue Regeneration: Mesenchymal Stem Cells, Secretome, and Biomaterials.

In the last few decades, tissue engineering has become one of the most studied medical fields. Even if bone shows self-remodeling properties, in some cases, due to injuries or anomalies, bone regeneration can be required. In particular, oral bone regeneration is needed in the dentistry field, where the functional restoration of tissues near the tooth represents a limit for many dental implants. In this context, the application of biomaterials and mesenchymal stem cells (MSCs) appears promising for bone regeneration. This review focused on in vivo studies that evaluated bone regeneration using biomaterials with MSCs. Different biocompatible biomaterials were enriched with MSCs from different sources. These constructs showed an enhanced bone regenerative power in in vivo models. However, we discussed also a future perspective in tissue engineering using the MSC secretome, namely the conditioned medium and extracellular vesicles. This new approach has already shown promising results for bone tissue regeneration in experimental models.

Stem Cells Secretome from Oral Tissue Could Represent a Promising Therapeutic Approach in COVID-19-Disease?

At present, severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection has quickly become a health emergency because no specifics vaccines or drugs, at this moment, are available. Recent studies have shown that the transplantation of mesenchymal stem cells (MSCs) into Coronavirus Disease 2019 (COVID-19) patients could represent a promising strategy for the development of new therapeutic methods. We speculate and suggest that the secretome of human Oral Tissue Stem Cells (hOTSCs), for their immunomodulatory and anti-inflammatory specific properties, could exert beneficial effects on the COVID-19 patients through an innovative aerosolisation technique. This non-invasive technique can offer multiple advantages in prophylaxis, as well as the prevention and treatment of severe epidemic respiratory syndrome with minimum risk and optimal therapeutic effects. This has the potential to create a novel pathway towards immunomodulatory therapy for the treatment of COVID-19 positive patients.

Short ELF-EMF Exposure Targets SIRT1/Nrf2/HO-1 Signaling in THP-1 Cells.

Extremely low frequency electromagnetic fields (ELF-EMFs) have been known to modulate inflammatory responses by targeting signal transduction pathways and influencing cellular redox balance through the generation of oxidants and antioxidants. Here, we studied the molecular mechanism underlying the anti-oxidative effect of ELF-EMF in THP-1 cells, particularly with respect to antioxidant enzymes, such as heme oxygenase-1 (HO-1), regulated transcriptionally through nuclear factor E2-related factor 2 (Nrf2) activation. Cells treated with lipopolysaccharides (LPS) were exposed to a 50 Hz, 1 mT extremely low frequency electromagnetic fields for 1 h, 6 h and, 24 h. Our results indicate that ELF-EMF induced HO-1 mRNA and protein expression in LPS-treated THP-1 cells, with peak expression at 6 h, accompanied with a concomitant migration to the nucleus of a truncated HO-1 protein form. The immunostaining analysis further verified a nuclear enrichment of HO-1. Moreover, ELF-EMF inhibited the protein expressions of the sirtuin1 (SIRT1) and nuclear factor kappa B (NF-kB) pathways, confirming their anti-inflammatory/antioxidative role. Pretreatment with LY294002 (Akt inhibitor) and PD980559 (ERK inhibitor) inhibited LPS-induced Nrf2 nuclear translocation and HO-1 protein expression in ELF-EMF-exposed cells. Taken together, our results suggest that short ELF-EMF exposure exerts a protective role in THP-1 cells treated with an inflammatory/oxidative insult such as LPS, via the regulation of Nrf-2/HO-1 and SIRT1 /NF-kB pathways associated with intracellular glutathione (GSH) accumulation.

Functional Relationship between Osteogenesis and Angiogenesis in Tissue Regeneration.

Bone tissue renewal can be outlined as a complicated mechanism centered on the interaction between osteogenic and angiogenic events capable of leading to bone formation and tissue renovation. The achievement or debacle of bone regeneration is focused on the primary role of vascularization occurrence; in particular, the turning point is the opportunity to vascularize the bulk scaffolds, in order to deliver enough nutrients, growth factors, minerals and oxygen for tissue restoration. The optimal scaffolds should ensure the development of vascular networks to warrant a positive suitable microenvironment for tissue engineering and renewal. Vascular Endothelial Growth Factor (VEGF), a main player in angiogenesis, is capable of provoking the migration and proliferation of endothelial cells and indirectly stimulating osteogenesis, through the regulation of the osteogenic growth factors released and through paracrine signaling. For this reason, we concentrated our attention on two principal groups involved in the renewal of bone tissue defects: the cells and the scaffold that should guarantee an effective vascularization process. The application of Mesenchymal Stem Cells (MSCs), an excellent cell source for tissue restoration, evidences a crucial role in tissue engineering and bone development strategies. This review aims to provide an overview of the intimate connection between blood vessels and bone formation that appear during bone regeneration when MSCs, their secretome-Extracellular Vesicles (EVs) and microRNAs (miRNAs) -and bone substitutes are used in combination.

Diameters and Fluorescence Calibration for Extracellular Vesicle Analyses by Flow Cytometry.

Extracellular vesicles (EVs) play a crucial role in the intercellular crosstalk. Mesenchymal stem cell-derived EVs (MSC-EVs), displaying promising therapeutic roles, contribute to the strong rationale for developing EVs as an alternative therapeutic option. EV analysis still represents one of the major issues to be solved in order to translate the use of MSC-EV detection in clinical settings. Even if flow cytometry (FC) has been largely applied for EV studies, the lack of consensus on protocols for FC detection of EVs generated controversy. Standard FC procedures, based on scatter measurements, only allows the detection of the "tip of the iceberg" of all EVs. We applied an alternative FC approach based on the use of a trigger threshold on a fluorescence channel. The EV numbers obtained by the application of the fluorescence triggering resulted significantly higher in respect to them obtained from the same samples acquired by placing the threshold on the side scatter (SSC) channel. The analysis of EV concentrations carried out by three different standardized flow cytometers allowed us to achieve a high level of reproducibility (CV < 20%). By applying the here-reported method highly reproducible results in terms of EV analysis and concentration measurements were obtained.

AEDG Peptide (Epitalon) Stimulates Gene Expression and Protein Synthesis during Neurogenesis: Possible Epigenetic Mechanism.

It was shown that AEDG peptide (Ala-Glu-Asp-Gly, Epitalon) regulates the function of the pineal gland, the retina, and the brain. AEDG peptide increases longevity in animals and decreases experimental cancerogenesis. AEDG peptide induces neuronal cell differentiation in retinal and human periodontal ligament stem cells. The aim of the study was to investigate the influence of AEDG peptide on neurogenic differentiation gene expression and protein synthesis in human gingival mesenchymal stem cells, and to suggest the basis for the epigenetic mechanism of this process. AEDG peptide increased the synthesis of neurogenic differentiation markers: Nestin, GAP43, ß Tubulin III, Doublecortin in hGMSCs. AEDG peptide increased Nestin, GAP43, ß Tubulin III and Doublecortin mRNA expression by 1.6-1.8 times in hGMSCs. Molecular modelling method showed, that AEDG peptide preferably binds with H1/6 and H1/3 histones in His-Pro-Ser-Tyr-Met-Ala-His-Pro-Ala-Arg-Lys and Tyr-Arg-Lys-Thr-Gln sites, which interact with DNA. These results correspond to previous experimental data. AEDG peptide and histones H1/3, H1/6 binding may be one of the mechanisms which provides an increase of Nestin, GAP43, ß Tubulin III, and Doublecortin neuronal differentiation gene transcription. AEDG peptide can epigenetically regulate neuronal differentiation gene expression and protein synthesis in human stem cells.

In vivo and in vitro results of an automated preloaded delivery system for IOL implantation in cataract surgery.

PURPOSE: To compare the corneal tissue trauma after the use of an automated preloaded injector and a manual injector and assess scanning electron microscope (SEM) and atomic force microscope (AFM) features of both injector cartridges. SETTING: Ophthalmology Clinic and Laboratory of Stem Cells and Regenerative Medicine University "G. d'Annunzio" of Chieti-Pescara, Chieti, Italy; DESIGN: Prospective randomized clinical study METHODS: Forty eyes of 40 patients for phacoemulsification were divided into two groups: implantation of intraocular lens was performed with AutonoMe automated delivery system (AutonoMe group: 20 eyes) and Monarch III injector system (Monarch group: 20 eyes). In vivo confocal microscopy (IVCM) and anterior segment optical coherence tomography (AS-OCT) were performed before surgery, at 1 h, 1 day and 1 month post-operatively. In addition, SEM and AFM were performed on cartridges of both injector systems after injection of the IOL. RESULTS: A greater increase in central corneal thickness and corneal thickness at the incision site were observed in Monarch group versus AutonoMe group 1 h and 1 day post-operatively (p < 0.05). Endothelial cell count loss was significantly higher in Monarch group compared with AutonoMe group (p < 0.05) at 1 and 24 h. AS-OCT showed less endothelial misalignment at 30 days (p < 0.05), and IVCM showed less tunnel inflammation at all time points (p < 0.05) in AutonoMe group compared with Monarch group; roughness analysis at AFM of the AutonoMe cartridge was significantly lower compared to Monarch D cartridge (p < 0.05). CONCLUSIONS: The AutonoMe injector provided less corneal tissue trauma compared with Monarch III injector. The AutonoMe cartridge showed lower roughness at AFM compared to the Monarch D cartridge.

Human Oral Stem Cells, Biomaterials and Extracellular Vesicles: A Promising Tool in Bone Tissue Repair.

Tissue engineering and/or regenerative medicine are fields of life science exploiting both engineering and biological fundamentals to originate new tissues and organs and to induce the regeneration of damaged or diseased tissues and organs. In particular, de novo bone tissue regeneration requires a mechanically competent osteo-conductive/inductive 3D biomaterial scaffold that guarantees the cell adhesion, proliferation, angiogenesis and differentiation into osteogenic lineage. Cellular components represent a key factor in tissue engineering and bone growth strategies take advantage from employment of mesenchymal stem cells (MSCs), an ideal cell source for tissue repair. Recently, the application of extracellular vesicles (EVs), isolated from stem cells, as cell-free therapy has emerged as a promising therapeutic strategy. This review aims at summarizing the recent and representative research on the bone tissue engineering field using a 3D scaffold enriched with human oral stem cells and their derivatives, EVs, as a promising therapeutic potential in the reconstructing of bone tissue defects.

Transcriptomic Analysis of Stem Cells Treated with Moringin or Cannabidiol: Analogies and Differences in Inflammation Pathways.

Inflammation is a common feature of many neurodegenerative diseases. The treatment of stem cells as a therapeutic approach to repair damage in the central nervous system represents a valid alternative. In this study, using Next-Generation Sequencing (NGS) technology, we analyzed the transcriptomic profile of human Gingival Mesenchymal Stem Cells (hGMSCs) treated with Moringin [4-(α-l-ramanosyloxy)-benzyl isothiocyanate] (hGMSCs-MOR) or with Cannabidiol (hGMSCs-CBD) at dose of 0.5 or 5 µM, respectively. Moreover, we compared their transcriptomic profiles in order to evaluate analogies and differences in pro- and anti-inflammatory pathways. The hGMSCs-MOR selectively downregulate TNF-α signaling from the beginning, reducing the expression of TNF-α receptor while hGMSCs-CBD limit its activity after the process started. The treatment with CBD downregulates the pro-inflammatory pathway mediated by the IL-1 family, including its receptor while MOR is less efficient. Furthermore, both the treatments are efficient in the IL-6 signaling. In particular, CBD reduces the effect of the pro-inflammatory JAK/STAT pathway while MOR enhances the pro-survival PI3K/AKT/mTOR. In addition, both hGMSCs-MOR and hGMSCs-CBD improve the anti-inflammatory activity enhancing the TGF-ß pathway.

3D Printing PLA/Gingival Stem Cells/ EVs Upregulate miR-2861 and -210 during Osteoangiogenesis Commitment.

Bone tissue regeneration strategies require approaches that provide an osteogenic and angiogenic microenvironment able to drive the bone growth. Recently, the development of 3D printing biomaterials, including poly(lactide) (3D-PLA), enriched with mesenchymal stem cells (MSCs) and/or their derivatives, such as extracellular vesicles (EVs) has been achieving promising results. In this study, in vitro results showed an increased expression of osteogenic and angiogenic markers, as RUNX2, VEGFA, OPN and COL1A1 in the living construct 3D-PLA/human Gingival MSCs (hGMSCs)/EVs. Considering that EVs carry and transfer proteins, mRNA and microRNA into target cells, we evaluated miR-2861 and miR-210 expression related to osteoangiogenesis commitment. Histological examination of rats implanted with 3D-PLA/hGMSCs/EVs evidenced the activation of bone regeneration and of the vascularization process, confirmed also by MicroCT. In synthesis, an upregulation of miR-2861 and -210 other than RUNX2, VEGFA, OPN and COL1A1 was evident in cells cultured in the presence of the biomaterial and EVs. Then, these results evidenced that EVs may enhance bone regeneration in calvaria defects, in association with an enhanced vascularization offering a novel regulatory system in the osteoangiogenesis evolution. The application of new strategies to improve biomaterial engraftment is of great interest in the regenerative medicine and can represent a way to promote bone regeneration.

Moringin Pretreatment Inhibits the Expression of Genes Involved in Mitophagy in the Stem Cell of the Human Periodontal Ligament.

Moringin [4-(α-L-rhamnosyloxy) benzyl isothiocyanate] is an isothiocyanate extracted from Moringa oleifera seeds. It is an antioxidant known for several biological properties useful in the treatment of neurodegenerative diseases. Several neurodegenerative disorders such as Parkinson's and Alzheimer's diseases are linked to dysfunctional mitochondria due to the resulting increase of Reactive Oxygen Species (ROS). Stem cell-based therapeutic treatments in neurodegenerative diseases provide an alternative strategy aimed to replace the impaired tissue. In this study were investigated the deregulated genes involved in mitophagy in the human periodontal ligament stem cells pretreated with moringin. The RNA-seq study reveals the downregulation of PINK1, with a fold change (FC) of -0.56, such as the genes involved in the phagophore formation (MAP1LC3B FC: -0.73, GABARAP FC: -0.52, GABARAPL1 FC: -0.70, GABARAPL2 FC: -0.39). The moringin pretreatment downregulates the pro-apoptotic gene BAX (-0.66) and upregulates the anti-apoptotic genes BCL2L12 (FC: 1.35) and MCL1 (FC: 0.36). The downregulation of the most of the caspases (CASP1 FC: -1.43, CASP4 FC: -0.18, CASP6 FC: -1.34, CASP7 FC: -0.46, CASP8 FC: -0.65) implies the inactivation of the apoptotic process. Our results suggest that mitochondrial dysfunctions induced by oxidative stress can be inhibited by moringin pretreatment in human periodontal ligament stem cells (hPDLSCs).

Endothelial committed oral stem cells as modelling in the relationship between periodontal and cardiovascular disease.

In the present study we have mimicked, in vitro, an inflammatory process using Lipopolysaccharide derived from Porphyromonas Gingivalis (LPS-G) and human Periodontal Ligament Stem Cells induced to endothelial differentiation (e-hPDLSCs). The research project has been organized into the three following steps: i) induction of hPDLSCs toward endothelial differentiation; ii) evaluation of the molecular signaling pathway involved in the response to the LPS-G, and iii) functional response evaluation of the living construct constituted by porcine decellularized valve/e-hPDLSCs treated with LPS-G. Obtained results showed that 5 µg/ml LPS-G stimulus provokes: a slowdown of cell growth starting from 24 hr and the release of IL6, IL8, and MCP1 molecules. Signaling network analyzed showed the activation of TLR4/ NFkB/ERK1/2/p-ERK1/2 signaling mediated by MyD88 in LPS-G stimulated e-hPDLSCs, moreover a time course put in evidence a nuclear traslocation of ERK1/2 and p-ERK1/2 in differentiated samples. Following, the ability of e-hPDLSCs to expand and colonize the decellularized porcine heart valves was appraised at ultrastructural level. Considering that, the Reactive Oxygen Species (ROS) play an important role in the progression and development of cardiovascular disease (CVD), in LPS-G living construct model e-hPDLSCs/decellularized porcine heart valves (dPHV), ROS production was assessed. Time lapse experiments evidenced that LPS-G provokes in e-hPDLSCs a rapid and sustained increase in ROS generation, negligible on undifferentiated cells. From obtained data, by multiparametric analyses, a reasonable conclusion may be that the inflammation process activated by LPS-G can affect endothelial cells and could represent in vivo a possible pathological and predictor state of CVD.