Human Periodontal Ligament Fibroblast Cells Research Articles

Gene Expression Profile in Immortalized Human Periodontal Ligament Fibroblasts Through hTERT Ectopic Expression: Transcriptome and Bioinformatic Analysis.

Human periodontal ligament fibroblast (hPLF) cells play an important role in maintaining oral cavity homeostasis with special function in tissue regeneration and maintenance of dental alveoli. Although their primary cell cultures are considered a good experimental model with no genetic changes, the finite life span may limit some experimental designs. The immortalization process increases cell life span but may cause genetic changes and chromosomal instability, resulting in direct effects on physiological cell responses. In this way, we aimed to investigate the global gene expression of hPLFs after the immortalization process by the ectopic expression of the catalytic subunit of the enzyme telomerase reverse transcriptase (hTERT) through transcriptome analysis. The embryonic origin of the primary culture of hPLF cells and immortalized hPLF-hTERT was also tested by vimentin staining, hTERT synthesis evaluated by indirect immunocytochemistry, analysis of cell proliferation, and morphology. The results indicated that hPLFs and hPLF-hTERT were positive for vimentin. On the 20th cell passage, hPLFs were in senescence, while hPLF-hTERT maintained their proliferation and morphology characteristics. At the same passage, hPLF-hTERT presented a significant increase in hTERT synthesis, but transcriptome did not reveal overexpression of the hTERT gene. Fifty-eight genes had their expression altered (11 upregulated and 47 downregulated) with the absence of changes in the key genes related to these cell types and in the main cancer-associated genes. In addition, the increase in hTERT protein expression without the overexpression of its gene indicates posttranscriptional level regulation. Successful immortalization of hPLFs through the ectopic expression of hTERT encourages further studies to design experimental protocols to investigate clinical questions from a translational perspective.

Physicochemical properties and cytocompatibility of newly developed calcium silicate-based sealers.

The purpose of this study was to compare the physical properties and cytocompatibility of contemporary calcium silicate-based sealers. Four calcium silicate-based sealers (BrightEndo MTA sealer, CeraSeal, EndoSeal TCS and One-Fil) were compared to an epoxy resin-based sealer (AH Plus). Flow, setting time, radiopacity and dimensional change were evaluated according to ISO 6876 standards. Cytotoxicity on human periodontal ligament fibroblast (hPDLF) cells was compared for biological properties using MTT assay. The surface of the sealer was analysed using scanning electron microscopy to evaluate cell attachment. Flow and radiopacity of all sealers met ISO standards, while setting time and dimensional stability did not meet the ISO standards. Calcium silicate-based sealers showed favourable cytocompatibility, and hPDLF cells were well attached to the calcium silicate-based sealers. Calcium silicate-based sealers have clinically acceptable flow and radiopacity, and cytocompatibility. However, these sealers had longer setting time and higher dimensional change than those required by ISO 6876.

Ultrafine-Grained Ti-31Mo-Type Composites with HA and Ag, Ta2O5 or CeO2 Addition for Implant Applications.

Ultrafine-grained Ti31Mo alloy and Ti31Mo5HA, Ti31Mo5HA-Ag (or Ta2O5, CeO2) composites with a grain size of approximately 2 μm were produced by the application of mechanical alloying and powder metallurgy. Additionally, the surface of the Ti31Mo alloy was modified. In the first stage, the specimens were immersed in 5M NaOH for 24 h at 60 °C. In the second stage, hydroxyapatite (HA) was deposited on the sample surface. The cathodic deposition at −5 V vs. open circuit potential (OCP) in the electrolyte containing 0.25M CaNa2-EDTA (di-calcium ethylenediaminetetraacetic acid), 0.25M K2HPO4 in 1M NaOH at 120 °C for 2 h was applied. The bulk Ti31Mo alloy is a single β-type phase. In the alkali-modified surface titanium oxide, Ti3O is formed. After hydrothermal treatment, the surface layer mostly consists of the Ca10(PO4)6(OH)2 (81.23%) with about 19% content of CaHPO4·2H2O. Using optical profiler, roughness 2D surface topography parameters were estimated. The in vitro cytocompatibility of synthesized materials was studied. The cell lines of normal human osteoblasts (NHost) and human periodontal ligament fibroblasts (HPdLF) was conducted in the presence of tested biomaterials. Ultrafine-grained Ti-based composites altered with HA and Ag, Ta2O5 or CeO2 have superior biocompatibility than the microcrystalline Ti metal. NHost and HPdLF cells in the contact with the synthesized biomaterial showed stable proliferation activity. Biocompatibility tests carried out indicate that the ultrafine-grained Ti31Mo5HA composites with Ag, Ta2O5, or CeO2 could be a good candidate for implant applications.

Cytotoxic evaluation of two orthodontic silver solder materials on human periodontal ligament fibroblast cells and the effects of antioxidant and antiapoptotic reagents.

To evaluate the cytotoxicity effects of two different solder materials used for orthodontic appliances on human periodontal ligament fibroblast (HPLF) cells, and to determine whether the mechanism of toxicity may involve oxidative stress and apoptosis. The silver solder samples (Leone and Summit) were soldered to orthodontic stainless steel bands and exposed to HPLF cells via cell culture inserts for 48 hours. Cytotoxicity effect of the soldered materials on HPLF cells was measured via tetrazolium salt 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) colorimetric assay (n = 10/sample) and morphological observation. In addition, the mechanism of cytotoxicity of the most toxic silver solder was investigated using both a caspase inhibitor Z-VAL-Ala-Asp-flu-oromethylketone (ZVAD-fmk) and the free radical scavenger Trolox (n = 8/sample). Statistical analysis was performed using one-way analysis of variance with a Bonferroni test. P < .05 was considered statistically significant. Compared to the control (no treatment, cells only), both silver solders were cytotoxic (P < .001). The bands alone were significantly cytotoxic compared to the control. There was a significant difference in cytotoxicity between the stainless steel bands alone and the Summit silver solder (P < .001), but not the Leone silver solder. The Summit silver solder was more cytotoxic than the Leone silver solder (P < .05). MTT results were supported by the microscopic morphological changes of the HPLF cells. Neither ZVAD-fmk nor Trolox provided significant protection. The two silver solder materials demonstrated different levels of cytotoxicity, and neither oxidative stress nor apoptosis is involved in the mechanism of cytotoxicity.

An in-vitro evaluation of cytotoxicity of fungal derived nanosilver particle endodontic irrigant on human periodontal ligament fibroblast cells.

Introduction: Recently, silver nanoparticles were indicated for root canal irrigation. The cytotoxicity evaluation of these agents helps to ascertain confident clinical use. This article aims at evaluating the cytotoxic effect of fungal-derived nanosilver particle irrigant on human periodontal ligament fibroblast (hPDLF) cells using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Materials and Methods: The test “nanoparticle” irrigant was produced using the fungi “Fusarium semitectum.” The cells were cultured in Dulbecco's Modified Eagle's Medium solution and adjusted to 5 × 103 cells/ml. About 100 μl of cells were seeded into a 96 well microplate and 100 μl test irrigant ranging from 5 to 640 μg/ml concentrations was added to a microplate and incubated at 37°C, 5% CO2 humidified conditions for 24 h. Untreated cells were used as a control group. About 5 mg/ml MTT was added to the plates and incubated at 37°C in 5% CO2 conditions for 4 h. The viability of cells and the percentage inhibition of cell were determined. Results: The 50% inhibition of cells was found to be around 320 μg/ml concentration. Cytotoxicity was found to be dose dependent and increased with higher concentrations of the nanosolution. Conclusion: Fungal-derived nanosilver irrigant is safe to hPDLF cells “in vitro” at a concentration of <320 μg/ml.

A Human Periodontal Ligament Fibroblast Cell Line as a New Model to Study Periodontal Stress.

The periodontal ligament (PDL) is exposed to different kinds of mechanical stresses such as bite force or orthodontic tooth movement. A simple and efficient model to study molecular responses to mechanical stress is the application of compressive force onto primary human periodontal ligament fibroblasts via glass disks. Yet, this model suffers from the need for primary cells from human donors which have a limited proliferative capacity. Here we show that an immortalized cell line, PDL-hTERT, derived from primary human periodontal ligament fibroblasts exhibits characteristic responses to glass disk-mediated compressive force resembling those of primary cells. These responses include induction and secretion of pro-inflammatory markers, changes in expression of extracellular matrix-reorganizing genes and induction of genes related to angiogenesis, osteoblastogenesis and osteoclastogenesis. The fact that PDL-hTERT cells can easily be transfected broadens their usefulness, as molecular gain- and loss-of-function studies become feasible.

Weissella cibaria EIR/P2-derived exopolysaccharide: A novel alternative to conventional biomaterials targeting periodontal regeneration

Healing and regeneration of periodontium are considered as a complex physiological process. Therefore, treatments need to be addressed with highly effective components modulating the multiple pathways. In this study, exopolysaccharide (EPS) produced by Weissella cibaria EIR/P2, was partially purified from the culture supernatant and subjected to characterization within the aim of evaluating its potential for periodontal regeneration. High-Performance Liquid Chromatography analysis revealed a single-peak corresponding to the glucose which identified the EPS as dextran. Fourier transform-infrared spectra were also displayed characteristic peaks for polysaccharides. According to the results of gel permeation/size exclusion-chromatography, the molecular mass was determined to be 8 × 106 Da. To clarify its anti-bacterial activity on Streptococcus mutans, effects on viability and biofilm formation was evaluated. At 50 mg/mL, dextran exhibited a bactericidal effect with 70% inhibition on biofilm formation. Besides, dose-dependent antioxidant effects were also detected. The efficacy of dextran in enhancing the viability of human periodontal ligament fibroblast cells (hPDLFCs) was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium-bromide (MTT) assay, and an increase was observed in the viability of hPDLFCs. In conclusion, dextran derived from W. cibaria can be potentially used as a multi-functional bioactive polymer in the design of new therapeutic strategies to promote healing and regeneration of periodontium.

Resveratrol and Celastrol Loaded Collagen Dental Implants Regulate Periodontal Ligament Fibroblast Growth and Osteoclastogenesis of Bone Marrow Macrophages.

Collagen is widely used for dental therapy in several ways such as films, 3D matrix, and composites, besides traditional Chinese medicine (TCM), has been used in tissue regeneration and wound healing application for centuries. Hence, the present study was targeted for the first time to fabricate collagen film with TCM such as resveratrol and celastrol in order to investigate the human periodontal ligament fibroblasts (HPLF) growth and bone marrow macrophages (BMM) derived osteoclastogenesis. Further, the physicochemical, mechanical and biological activities of collagen-TCM films crosslinked by glycerol and EDC-NHS (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide-N-hydroxysulfosuccinimide) were investigated. Collagen film characterization was significantly regulated by the nature of plasticizers like hydrophobic and degree of polarity. Interestingly, the collagen film's denaturation temperature was increased by EDC-NHS than glycerol. FT-IR data confirmed the functional group changes due to chemical interaction of collagen with TCM. Morphological changes of HPLF cells cultured in control and collagen films were observed by SEM. Importantly, the addition of resveratrol upregulated the proliferation of HPLF cells, while osteoclastogenesis of BMM cells treated with mCSF-RANKL was significantly downregulated by celastrol. Accordingly, the collagen-TCM film could be an interesting material for dental regeneration, and especially it is a therapeutic target to restrain the elevated bone resorption during osteoporosis.

Survival of human periodontal ligament fibroblast cells in Cornisol and HBSS for transportation of avulsed teeth: a comparative ex vivo study

Objective Viability of periodontal ligament fibroblast cells (PDFC) is one of the key factors in determining the success of replantation of avulsed teeth. Extra-oral time and transport media are closely related to the same. The present study aims to evaluate and compare the efficiency of Cornisol, Hank’s balanced salt solution (HBSS) and normal saline in preserving the viability of PDFC. Materials and methods The human PDFC were isolated from primary culture from freshly extracted human premolars. Effect of Cornisol, HBSS and normal saline on viability of isolated PDFC was assessed using standard MTT assay. The cells were exposed to the experimental solutions (Cornisol/HBSS/normal saline) for varying time points (30 min, 1 h, 24 h, 48 h and 96 h) and viability was determined by colorimetric MTT method by quantifying the amount of formazan crystal formed (optical density). Experiment was performed in triplicates and the data were subjected to statistical analysis. Results Statistical analysis was performed using the Kruskal–Wallis ANOVA with post hoc Bonferroni’s test with a significance level of p value ≤.05. Cornisol ≥ HBSS > saline. Conclusion Cornisol can be used as a storage media for avulsed teeth and is significantly more effective than HBSS in maintaining the periodontal ligament cell viability at tested time intervals.

Biomimetic collagen-sodium alginate-titanium oxide (TiO2) 3D matrix supports differentiated periodontal ligament fibroblasts growth for periodontal tissue regeneration

Fabrication of biomaterial that mimics a suitable biological microenvironment is still a major challenge in the field of periodontitis treatment. Hence, in this report, we presented for the first time the fabrication of a novel biomaterial 3D matrix using collagen combined with sodium alginate and titanium oxide (TiO2) to recreate the in-vivo microenvironment and to act as a platform for the culture of human periodontal ligament fibroblasts (HPLF) towards osteogenic differentiation. Further, we explored the changes of differentiated and undifferentiated HPLF cells in morphological and cellular level comparing 2D (standard culture plates) and 3D cell culture systems. The physicochemical parameters such as stiffness, water binding capacity, swelling, shrinkage factor, porosity and in-vitro biodegradation show the suitability of this 3D matrix to act as a scaffold for in-vitro periodontal regeneration. The differentiated HPLF cells in the 3D matrix secrete high levels of collagen, osteocalcin, alkaline phosphatase compared to the conventional 2D cell culture. Morphological analysis revealed the structural changes of HPLF cells before and after differentiation in 2D and 3D cell culture. In this study, we find that the level of osteocalcin secretion towards osteogenic differentiation was enhanced in HPLF cells by 3D matrix as compared with 2D cell culture, which demonstrates the osteogenic stimulatory potential of 3D matrix. Overall, the fabricated 3D matrix supports the differentiation of the HPLF cells into osteoblastogenic lineage cells in-vitro and is a promising approach for further investigations in in-vivo treatment of periodontal tissue impairment.

Cytotoxicity of Different Nano Composite Resins on Human Gingival and Periodontal Ligament Fibroblast Cell Lines: An In Vitro Study.

The aim of this study is to determine the cytotoxicity of three different nano composite resins (CRs) on human gingival fibroblast (hGF) and periodontal ligament fibroblast (hPDLF) cell lines. These CRs selected were nanohybrid organic monomer-based Admira Fusion (AF), nanohybrid Bis-(acryloyloxymethyl) tricyclo [5.2.1.0.sup.2,6] decane-based Charisma Topaz (CT), and supra nano filled resin-based Estelite Quick Sigma (EQS). MTT assay was performed to assess the cytotoxicity of CRs at 24 h and one week. AF and EQS applied on hGF cells at 24 h and one week demonstrated similar cytotoxic outcomes. Cytotoxicity of CT on hGF cells at one week was higher than 24 h (p = 0.04). Cytotoxicity of CT on hGF cells was higher at 24 h (p = 0.002) and one week (p = 0.009) compared to control. All composites showed higher cytotoxicity on hPDLF cells at one week than the 24 h (AF; p = 0.02, CT; p = 0.02, EQS; p = 0.04). AF and EQS demonstrated lower cytotoxicity on hPDLF cells than the control group at 24 h (AF; p = 0.01, EQS; p = 0.001). CT was found more cytotoxic on hPDLF cells than the control (p = 0.01) and EQS group (p = 0.008) at one week. The cytotoxicity of CRs on hGF and hPDLF cells vary, according to the type of composites, cell types, and exposure time.

Oxidative Damage in Human Periodontal Ligament Fibroblast (hPLF) after Methylmercury Exposure.

Human exposure to mercury (Hg) is primary associated with its organic form, methylmercury (MeHg), through the ingestion of contaminated seafood. However, Hg contamination is also positively correlated with the number of dental restorations, total surface of amalgam, and organic mercury concentration in the saliva. Among the cells existing in the oral cavity, human periodontal ligament fibroblast (hPLF) cells are important cells responsible for the production of matrix and extracellular collagen, besides sustentation, renewal, repair, and tissue regeneration. In this way, the present study is aimed at investigating the potential oxidative effects caused by MeHg on hPLF. Firstly, we analyzed the cytotoxic effects of MeHg (general metabolism status, cell viability, and mercury accumulation) followed by the parameters related to oxidative stress (total antioxidant capacity, GSH levels, and DNA damage). Our results demonstrated that MeHg toxicity increased in accordance with the rise of MeHg concentration in the exposure solutions (1-7 μM) causing 100% of cell death at 7 μM MeHg exposure. The general metabolism status was firstly affected by 2 μM MeHg exposure (43.8 ± 1.7%), while a significant decrease of cell viability has arisen significantly only at 3 μM MeHg exposure (68.7 ± 1.4%). The ratio among these two analyses (named fold change) demonstrated viable hPLF with compromised cellular machinery along with the range of MeHg exposure. Subsequently, two distinct MeHg concentrations (0.3 and 3 μM) were chosen based on LC50 value (4.2 μM). hPLF exposed to these two MeHg concentrations showed an intracellular Hg accumulation as a linear-type saturation curve indicating that metal accumulated diffusively in the cells, typical for metal organic forms such as methyl. The levels of total GSH decreased 50% at exposure to 3 μM MeHg when compared to control. Finally, no alteration in the DNA integrity was observed at 0.3 μM MeHg exposure, but 3 μM MeHg caused significant damage. In conclusion, it was observed that MeHg exposure affected the general metabolism status of hPLF with no necessary decrease on the cell death. Additionally, although the oxidative imbalance in the hPLF was confirmed only at 3 μM MeHg through the increase of total GSH level and DNA damage, the lower concentration of MeHg used (0.3 μM) requires attention since the intracellular mercury accumulation may be toxic at chronic exposures.

Vesicular nucleotide transporter mediates adenosine triphosphate release in compressed human periodontal ligament fibroblast cells and participates in tooth movement-induced nociception in rats

ObjectivePain control is imperative in orthodontic treatment. Adenosine triphosphate (ATP) is a key mediator released from periodontal ligament cells that excites nociceptive nerve endings. Vesicular nucleotide transporter (VNUT), encoded by the Solute carrier family 17 member 9 (SLC17A9) gene, participates in ATP uptake into secretory vesicles; thus, it may mediate tooth movement-induced pain. In the present study, we examined whether VNUT in periodontal ligament cells participates in tooth movement-induced nociception. DesignExpression levels of SLC17A9, connexin 43, and pannexin 1 in human periodontal ligament fibroblasts (HPDLFs) were examined by quantitative reverse transcription-polymerase chain reaction. Mechanical force via centrifugation-induced ATP release was measured using an ATP bioluminescence assay. Inhibitors were used to evaluate the role of ATP transporters. Face-grooming behaviors were assessed as indicators of nociceptive responses after experimental tooth movement in rats, as well as the effects of drugs for the pain-like behavior. ResultsAfter HPDLFs underwent mechanical stimulation by centrifugation, SLC17A9 mRNA expression in the cells was significantly upregulated. Increased ATP release from HPDLFs after mechanical stimulation was suppressed by treatment with clodronic acid, a VNUT inhibitor, at concentrations of 0.1 and 1.0 μM. In rats, face-grooming behaviors (indicators of nociception) were significantly increased on day 1 after experimental tooth movement. Increased face-grooming behaviors were suppressed by systemic administration of clodronic acid (0.1 mg/kg). ConclusionsThese results indicate that release of ATP from periodontal ligament cells via VNUT is important for nociceptive transduction during orthodontic treatment. Thus, VNUT may provide a novel drug target for tooth movement-induced pain.

Effect of Sequestosome-1/p62 Expression on Autophagy of Human Periodontal Ligament Fibroblasts Induced by Porphyromonas gingivalis

Porphyromonas gingivalis is the main pathogen of periodontitis. The purpose of this study was to analyze the effect of p62 protein on P. gingivalis-induced autophagy in human periodontal ligament fibroblast (HPDLF). HPDLF cells were first infected with P. gingivalis; autophagy and p62 expression levels were observed at different time points (0, 12, 18, and 24 h) to detect the relationship between p62 and autophagy over time. Autophagy and apoptosis of HPDLF cells infected with P. gingivalis were detected using siRNA specific interference with p62 expression. Fluorescence microscopy and electron microscopy showed autophagic vacuoles in HPDLF cells infected with P. gingivalis. Western blotting showed that p62 expression peaked at 12 h and began to decrease at 18 h. The expression of p62 was silenced by siRNA compared with the control group; the ratio of LC3-II/LC3-I decreased significantly, while the expression level of caspase-3 increased significantly. The the CCK-8 cell viability assay showed that the silencing of p62 reduced HPDLF cell viability, and the plate coating method showed that the silencing of p62 reduced the survival rate of P. gingivalis in the HPDLF cells. Therefore, in vitro studies showed that the presence of p62 is beneficial for maintaining the level of autophagy in HPDLF cells induced by P. gingivalis and inhibiting apoptosis. p62 facilitates the removal of P. gingivalis that are invading cells during this process.

N-acetyl cysteine inhibits lipopolysaccharide-mediated synthesis of interleukin-1β and tumor necrosis factor-α in human periodontal ligament fibroblast cells through nuclear factor-kappa B signaling.

The aim of this study was to investigate the role of n-acetyl cysteine (NAC) in the lipopolysaccharide (LPS)-mediated induction of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) synthesis by human periodontal ligament fibroblast cells (hPDLFs). In addition, we aimed to determine the involvement of the nuclear factor-kappa B (NF-κB) pathway in any changes in IL-1β and TNF-α expression observed in response to LPS and NAC. HPDLFs were obtained by primary culture. The culture medium used in this experiment was Dulbecco's Modified Eagle Medium (DMEM low-glucose). Cells were stimulated with various concentrations of NAC or LPS. Cell proliferation was measured at various time-points with the cell Counting Kit 8 (CCK-8) assay. mRNA levels of IL-1β and TNF-α were determined by real-time quantitative polymerase chain reaction (RT-qPCR) analysis. Protein levels of IL-1β and TNF-α were measured by enzyme-linked immunosorbent assay (ELISA). Protein and mRNA expression levels of NF-κB were measured by western blot and RT-qPCR. The results showed that LPS treatment in hPDLFs induced mRNA and protein expression of IL-1β, TNF-α, and NF-κB. However, these effects were eliminated by pretreatment with NAC. Pretreatment with both NAC (1 mmol/L) and BAY11-7082 (10 μmol/L) significantly inhibited the NF-κB activity induced by LPS. NAC inhibits the LPS-mediated synthesis of tumor TNF-α and IL-1β in hPDLFs, through the NF-κB pathway.

The effects of different 650 nm laser diode irradiation times on the viability and proliferation of human periodontal ligament fibroblast cells

Background: Endo-perio lesions are clinical manifestations of inflammation in the periodontal and pulp tissue. Damage to the periodontal ligament can inhibit its ability to regenerate. Therefore, laser therapy use is expected to improve the prognosis with regard to healing lesions. Unfortunately, the duration of irradiation during laser diode therapy can influence the viability and proliferation of human periodontal ligament fibroblast (hPDLF) cells. Purpose: This study aims to determine the effects of different irradiation exposure times of the 650 nm laser diode of the pulsed mode type on the viability and proliferation of human periodontal ligament fibroblast cells. Methods: This study constituted a laboratory experiment on hPDLF cells using 650 nm laser diode irradiation. Six groups formed the research subjects in this study, namely; two control groups, two radiation groups respectively subjected to irradiation exposure of 15 seconds and 35 seconds duration followed by 24-hour incubation, and two radiation groups exposed to irradiation for 15 and 35 seconds respectively followed by 72-hour incubation period. The viability and proliferation of those cells were subsequently calculated by ELISA reader, while the data was analyzed by means of one-way ANOVA and Tukey tests. Results: The significance value of the viability scores between the 15-second irradiation group and the 35-second irradiation group was less than 0.05, indicating that there was a significant difference between these treatment groups. Similarly, the significance value of proliferation scores between the 15-second irradiation group and the 35-second irradiation group was less than 0.05, again indicating a significant difference between these treatment groups. Conclusion: Irradiation using a 650 nm laser diode 15 seconds and 35 seconds in duration can induce an increase in the viability and proliferation of hPDLF cells.

Tumor-derived exosomes educate fibroblasts to promote salivary adenoid cystic carcinoma metastasis via NGF-NTRK1 pathway.

Salivary adenoid cystic carcinoma (SACC) is a rare head and neck malignancy characterized by unpredictable expansion, considerable perineural invasion and high risk of metastasis; however, the underlying mechanism of SACC progression remains unclear. Cancer-associated fibroblasts localized within the tumor microenvironment may promote cancer malignant transformation by enhancing tumor growth, blood vessel formation, inflammation development and metastasis occurrence. Small extracellular vesicles, including exosomes, are mediators of intercellular communication and can influence major tumor-associated pathways. The present study aimed to explore the exosome-mediated communication between SACC cells and fibroblasts. The results from confocal microscopy demonstrated that exosomes derived from the human cell line SACC-83 were internalized by the human periodontal ligament fibroblast (HPLF) cells. Following exosome internalization, HPLF cells appeared to enhance SACC-83 cell metastasis and were educated toward a protumorigenesis phenotype according to transcriptome RNA sequencing and reverse transcription-quantitative polymerase chain reaction analysis. This phenomenon included exosome-mediated stimulation of proinflammatory cytokines and nerve growth factor (NGF) secretion. Furthermore, NGF blockage reduced the enhanced SACC-83 cell invasion stimulated by the supernatant isolated from exosome-educated HPLF cells. In addition, the results reported that neurotrophic receptor tyrosine kinase 1 (NTRK1), which is the high-affinity NGF receptor, was significantly upregulated in human SACC-83 cells. These results demonstrated that SACC-83 cell-derived exosomes educated HPLF cells toward the protumorigenic phenotype via the NGF-NTRK1 pathway, which suggested that this type of exosomes may be used as a potential therapeutic target for SACC.

Salvadora persica Extract-laden Jellyfish Collagen Hybrid Constructs for Periodontal Tissue Regeneration

Considerable effort in the field of periodontal tissue engineering has been expended in the construction of advanced biomatrix for the treatment of periodontal diseases caused by poor oral hygiene, malnutrition, genetic factors, and systemic disorders. With this in mind, the ultimate goal of this investigation is to fabricate sophisticated scaffolds using jellyfish collagen (JC) and aqueous Salvadora persica (Miswak) extracts. Rhizostoma pulmo species JC was isolated and characterized in depth. Miswak was extracted using two different methods. The extraction yield was calculated to be 14.2 ± 0.9 % and 17.1 ± 0.4 % for the Method I and II, respectively. gas chromatography-mass spectroscopy (GC-MS) results revealed the extract to be composed of 1,8-cineole (49.3 %), benzyl nitrile (36.2 %), benzyl isothiocyanate (5.9 %), limonene (2.4 %), eugenol (0.8 %) and palmitic acid (0.3 %). Total phenolic content and antioxidant capacities of the extracts were also determined by spectrophotometrically. Human periodontal ligament fibroblast cells were isolated and expanded. Cell viability on JC and miswak extract-laden JC scaffolds was determined by 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) assay. Microarchitectures of the JC, 0.05 and 0.1% miswak extract-laden JC scaffolds and also cellular behaviors on these surfaces were evaluated by scanning electron microscopy (SEM) analysis. This study suggests that miswak extract-laden JC scaffolds would present new opportunities for periodontal tissue engineering.

Thermal annealed silk fibroin membranes for periodontal guided tissue regeneration.

Guided tissue regeneration (GTR) is a surgical procedure applied in the reconstruction of periodontal defects, where an occlusive membrane is used to prevent the fast-growing connective tissue from migrating into the defect. In this work, silk fibroin (SF) membranes were developed for periodontal guided tissue regeneration. Solutions of SF with glycerol (GLY) or polyvinyl alcohol (PVA) where prepared at several weight ratios up to 30%, followed by solvent casting and thermal annealing at 85 °C for periods of 6 and 12 h to produce high flexible and stable membranes. These were characterized in terms of their morphology, physical integrity, chemical structure, mechanical and thermal properties, swelling capability and in vitro degradation behavior. The developed blended membranes exhibited high ductility, which is particular relevant considering the need for physical handling and adaptability to the defect. Moreover, the membranes were cultured with human periodontal ligament fibroblast cells (hPDLs) up to 7 days. Also, the higher hydrophilicity and consequent in vitro proteolytic degradability of these blends was superior to pure silk fibroin membranes. In particular SF/GLY blends demonstrated to support high cell adhesion and viability with an adequate hPDLs' morphology, make them excellent candidates for applications in periodontal regeneration.

Cytotoxicity assay of sodium hypochlorite and QMix on cultured human periodontal ligament fibroblast cells

Aims and Objectives: Sodium hypochlorite (NaOCl) and QMix are solutions used for root canal irrigation. During the endodontic procedure, irrigation solutions are often leaked beyond the apical foramen and as a result cause periapical tissue complication because of their toxicity. This study was aimed at proving the cytotoxicity effect of NaOCl and QMiX on cultured human periodontal ligament fibroblast cells (HPDLFc). Materials and Methods: HPDLFc were exposed to NaOCl and QMix at various concentrations. Cell viability was assessed with Mosmann’s tetrazolium toxicity assay and the results were measured and statistically analyzed by probit analysis to determine the median lethal concentration (LC50), the concentration that is lethal to 50% of the cells and therefore considered cytotoxic. Results: NaOCl and QMix are cytotoxic to HPDLFc at 0.254 and 0.363 µL/mL, respectively. Conclusion: Both solutions are cytotoxic to HPDLFc at certain concentrations.