A newly developed kit for dental apical root resorption detection: efficacy and acceptability.

OBJECTIVES: To determine the efficacy of a newly developed kit in dentine sialophosphoprotein (DSPP) detection and compare it with enzyme-linked immunosorbent assay (ELISA). User acceptance was also determined. MATERIALS AND METHODS: This cross-sectional study consisted of 45 subjects who were divided into 3 groups based on the severity of root resorption using radiographs: normal (RO), mild (RM), and severe (RS). DSPP in GCF samples was analyzed using both methods. Questionnaires were distributed to 30 orthodontists to evaluate future user acceptance. RESULTS: The sensitivity and specificity of the kit were 0.98 and 0.8 respectively. The DSPP concentrations measured using ELISA were the highest in the RS group (6.33 ± 0.85 ng/mL) followed by RM group (3.77 ± 0.36 ng/mL) and the RO group had the lowest concentration (2.23 ± 0.55 ng/mL). The new kit portrayed similar results as the ELISA, the optical density (OD) values were the highest in the RS group (0.62 ± 0.10) followed by RM group (0.33 ± 0.03) and the RO group (0.19 ± 0.06). The differences among all the groups were statistically significant (p < 0.05) for both methods. The Pearson correlation coefficient showed a statistically significant (p < 0.001) strong and positive correlation between DSPP concentrations and OD values. CONCLUSIONS: The new kit was validated to detect the colour intensities of different severity of root resorptions. Most of the responses to the survey were positive towards the new kit for being a safer and simpler method to detect apical root resorption.

PAX7, a Key for Myogenesis Modulation in Muscular Dystrophies through Multiple Signaling Pathways: A Systematic Review.

Muscular dystrophy is a heterogenous group of hereditary muscle disorders caused by mutations in the genes responsible for muscle development, and is generally defined by a disastrous progression of muscle wasting and massive loss in muscle regeneration. Pax7 is closely associated with myogenesis, which is governed by various signaling pathways throughout a lifetime and is frequently used as an indicator in muscle research. In this review, an extensive literature search adhering to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines was performed to identify research that examined signaling pathways in living models, while quantifying Pax7 expression in myogenesis. A total of 247 articles were retrieved from the Web of Science (WoS), PubMed and Scopus databases and were thoroughly examined and evaluated, resulting in 19 articles which met the inclusion criteria. Admittedly, we were only able to discuss the quantification of Pax7 carried out in research affecting various type of genes and signaling pathways, rather than the expression of Pax7 itself, due to the massive differences in approach, factor molecules and signaling pathways analyzed across the research. However, we highlighted the thorough evidence for the alteration of the muscle stem cell precursor Pax7 in multiple signaling pathways described in different living models, with an emphasis on the novel approach that could be taken in manipulating Pax7 expression itself in dystrophic muscle, towards the discovery of an effective treatment for muscular dystrophy. Therefore, we believe that this could be applied to the potential gap in muscle research that could be filled by tuning the well-established marker expression to improve dystrophic muscle.

A Comparative Analysis of Ascorbic Acid-induced Cytotoxicity and Differentiation between SHED and DPSC.

AIM: The aim of this study was to compare dental pulp tissue in human exfoliated deciduous teeth (SHEDs) and dental pulp stem cells (DPSCs) in response to ascorbic acid as the sole osteoblast inducer. BACKGROUND: A cocktail of ascorbic acid, ß-glycerophosphate, and dexamethasone has been widely used to induce osteoblast differentiation. However, under certain conditions, ß-glycerophosphate and dexamethasone can cause a decrease in cell viability in stem cells. OBJECTIVES: This study aims to determine the cytotoxic effect and potential of ascorbic acid as the sole inducer of osteoblast differentiation. METHODS: Cytotoxicity analyses in the presence of 10-500 µg/mL ascorbic acid were performed in both cell types using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The concentrations below the IC50 (i.e., 10-150 µg/mL) were used to determine osteoblast differentiation potential of ascorbic acid using the alkaline phosphatase (ALP) assay, von Kossa staining, and reverse transcription-polymerase chain reaction. RESULTS: SHEDs and DPSCs proliferated for 21 days, expressed a Mesenchymal Stem Cell (MSC) marker (CD73+), and did not express Hematopoietic Stem Cell (HSC) markers (CD34- and SLAMF1-). SHEDs had a higher range of IC50 values (215-240 µg/mL ascorbic acid), while the IC50 values for DPSCs were 177-211 µg/mL after 24-72 hours. SHEDs treated with 10-100 µg/mL ascorbic acid alone exhibited higher ALP-specific activity and a higher percentage of mineralisation than DPSCs. Both cell types expressed osteoblast markers on day 21, i.e., RUNX2+ and BSP+, in the presence of ascorbic acid. CONCLUSIONS: SHEDs survive at higher concentrations of ascorbic acid as compared to DPSC. The cytotoxic effect was only exhibited at ≥250 µg/mL ascorbic acid. In addition, SHED exhibited better ALP and mineralization activities, but lower osteoblast marker expression than DPSC in response to ascorbic acid as the sole inducer.

Mesenchymal stem cells: A comprehensive methods for odontoblastic induction.

BACKGROUND: In the area of oral and maxillofacial surgery, regenerative endodontics aims to present alternative options to conventional treatment strategies. With continuous advances in regenerative medicine, the source of cells used for pulp tissue regeneration is not only limited to mesenchymal stem cells as the non-mesenchymal stem cells have shown capabilities too. In this review, we are systematically assessing the recent findings on odontoblastic differentiation induction with scaffold and non-scaffold approaches. METHODS: A comprehensive search was conducted in Pubmed, and Scopus, and relevant studies published between 2015 and 2020 were selected following the PRISMA guideline. The main inclusion criteria were that articles must be revolving on method for osteoblast differentiation in vitro study. Therefore, in vivo and human or animal clinical studies were excluded. The search outcomes identified all articles containing the word "odontoblast", "differentiation", and "mesenchymal stem cell". RESULTS: The literature search identified 99 related studies, but only 11 articles met the inclusion criteria. These include 5 odontoblastic differentiation induction with scaffold, 6 inductions without scaffolds. The data collected were characterised into two main categories: type of cells undergo odontoblastic differentiation, and odontoblastic differentiation techniques using scaffolds or non-scaffold. CONCLUSION: Based on the data analysis, the scaffold-based odontoblastic induction method seems to be a better option compared to the non-scaffold method. In addition of that, the combination of growth factors in scaffold-based methods could possibly enhance the differentiation. Thus, further detailed studies are still required to understand the mechanism and the way to enhance odontoblastic differentiation.

Comparative evaluation of osteogenic differentiation potential of stem cells derived from dental pulp and exfoliated deciduous teeth cultured over granular hydroxyapatite based scaffold.

BACKGROUND: Mesenchymal stem cells isolated from the dental pulp of primary and permanent teeth can be differentiated into different cell types including osteoblasts. This study was conducted to compare the morphology and osteogenic potential of stem cells from exfoliated deciduous teeth (SHED) and dental pulp stem cells (DPSC) in granular hydroxyapatite scaffold (gHA). Preosteoblast cells (MC3T3-E1) were used as a control group. METHODOLOGY: The expression of stemness markers for DPSC and SHED was evaluated using reverse transcriptase-polymerase chain reaction (RT-PCR). Alkaline phosphatase assay was used to compare the osteoblastic differentiation of these cells (2D culture). Then, cells were seeded on the scaffold and incubated for 21 days. Morphology assessment using field emission scanning electron microscopy (FESEM) was done while osteogenic differentiation was detected using ALP assay (3D culture). RESULTS: The morphology of cells was mononucleated, fibroblast-like shaped cells with extended cytoplasmic projection. In RT-PCR study, DPSC and SHED expressed GAPDH, CD73, CD105, and CD146 while negatively expressed CD11b, CD34 and CD45. FESEM results showed that by day 21, dental stem cells have a round like morphology which is the morphology of osteoblast as compared to day 7. The osteogenic potential using ALP assay was significantly increased (p < 0.01) in SHED as compared to DPSC and MC3T3-E1 in 2D and 3D cultures. CONCLUSION: gHA scaffold is an optimal scaffold as it induced osteogenesis in vitro. Besides, SHED had the highest osteogenic potential making them a preferred candidate for tissue engineering in comparison with DPSC.

Insights into the Effects of the Dental Stem Cell Secretome on Nerve Regeneration: Towards Cell-Free Treatment.

Cell-free treatment is emerging as an alternative to cell delivery to promote endogenous regeneration using cell-derived factors. The purpose of this article was to systematically review studies of the effects of the dental stem cell secretome on nerve regeneration. PubMed and Scopus databases were used where searched and related studies were selected. The primary search identified 36 articles with the utilized keywords; however, only 13 articles met the defined inclusion criteria. Eight out of thirteen articles included in vivo and in vitro studies. We classified the dental stem cell-derived secretome with its nerve regeneration potential. All studies demonstrated that dental stem cell-derived factors promote neurotrophic effects that can mechanistically stimulate nerve regeneration in neurodegenerative diseases and nerve injury. This data collection will enable researchers to gather information to create a precise formulation for future prescribed treatments.

A Perspective on Stem Cells as Biological Systems that Produce Differentiated Osteoblasts and Odontoblasts.

Stem cells (SCs) are capable of self-renewal and multilineage differentiation. Human mesenchymal stem cells (MSCs) and haematopoietic stem cells (HSCs) which can be obtained from multiple sources, are suitable for application in regenerative medicine and transplant therapy. The aim of this review is to evaluate the potential of genomic and proteomic profiling analysis to identify the differentiation of MSCs and HSCs towards osteoblast and odontoblast lineages. In vitro differentiation towards both of these lineages can be induced using similar differentiation factors. Gene profiling cannot be utilised to confirm the lineages of these two types of differentiated cells. Differentiated cells of both lineages express most of the same markers. Most researchers have detected the expression of genes such as ALP, OCN, OPN, BMP2 and RUNX2 in osteoblasts and the expression of the DSPP gene in odontoblasts. Based on their cell-type specific protein profiles, various proteins are differentially expressed by osteoblasts and odontoblasts, except for vimentin and heterogeneous nuclear ribonucleoprotein C, which are expressed in both cell types, and LOXL2 protein, which is expressed only in odontoblasts.

Cytotoxicity effect of degraded and undegraded kappa and iota carrageenan in human intestine and liver cell lines.

BACKGROUND: Carrageenan is a linear sulphated polysaccharide extracted from red seaweed of the Rhodophyceae family. It has broad spectrum of applications in biomedical and biopharmaceutical field. In this study, we determined the cytotoxicity of degraded and undegraded carrageenan in human intestine (Caco-2; cancer and FHs 74 Int; normal) and liver (HepG2; cancer and Fa2N-4; normal) cell lines. METHODS: Food grade k-carrageenan (FGKC), dried sheet k-carrageenan (DKC), commercial grade k-carrageenan (CGKC), food grade i-carrageenan (FGIC) and commercial grade i-carrageenan (CGIC) were dissolved in hydrochloric acid and water to prepare degraded and undegraded carrageenan, respectively. Carrageenan at the concentration range of 62.5 - 2000.0 µg mL(-1) was used in the study. MTT assay was used to determine the cell viability while the mode of cell death was determined by May-Grunwald Giemsa (MGG) staining, acridine orange-ethidium bromide (AO/EtBr) staining, agarose gel electrophoresis and gene expression analysis. RESULTS: Degraded FGKC, DKC and CGKC showed IC50 in 24, 48 and 72 hours treated Caco-2, FHs 74 Int, HepG2 and Fa2N-4 cell lines as tested by MTT assay. Degraded FGIC and CGIC only showed its toxicity in Fa2N-4 cells. The characteristics of apoptosis were demonstrated in degraded k-carrageenan treated Caco-2, FHs 74 Int, HepG2 and Fa2N-4 cells after MGG staining. When Caco-2 and HepG2 cells were undergone AO/EtBr staining, chromatin condensation and nuclear fragmentation were clearly seen under the microscope. However, DNA ladder was only found in HepG2 cells after gel electrophoresis analysis. Degraded k-carrageenan also inactivated PCNA, Ki-67 and survivin gene in HepG2. On the other hand, undegraded FGKC, DKC, CGKC, FGIC and CGIC treated cells showed no cytotoxic effect after analyzed by the same analyses as in degraded carrageenan. CONCLUSION: Degraded k-carrageenan inhibited cell proliferation in Caco-2, FHs 74 Int, HepG2 and Fa2N-4 cell lines and the anti-proliferative effect was related to apoptosis together with inactivation of cell proliferating genes as determined by morphological observation and molecular analysis. However, no cytotoxic effect was found in undegraded carrageenan towards normal and cancer intestine and liver cell lines.

Comparison of self- and conventional-ligating brackets in the alignment stage.

This prospective study investigated the difference in clinical efficiency between Damon™ 3 self-ligating brackets (SLB) compared with Mini Diamond conventional ligating brackets (CLBs) during tooth alignment in straightwire fixed appliance therapy. Twenty-nine patients (10 males and 19 females), aged between 14 and 30 years, were randomly divided into two groups: 14 patients received the SLB and 15 received the CLB. Upper arch impressions were taken for pre-treatment records (T(0)). A transpalatal arch was soldered to both maxillary first molar bands prior to extraction of the maxillary first premolars, followed by straightwire fixed appliances (0.022 × 0.028 inch). A 0.014 inch nickel titanium (NiTi) wire was used as the levelling and aligning archwire. Four monthly reviews were undertaken and impressions of the upper arch were taken at each appointment (T(1), T(2), T(3), and T(4)). Displacements of the teeth were determined using Little's irregularity index (LII). Data were analysed using the Mann-Whitney U-test. In the aligning stage, the CLB group showed significantly faster alignment of the teeth compared with the SLB group at the T(1)-T(2) interval (P < 0.05). However, there were no differences at T(2)-T(3), and T(3)-T(4) for either group (P > 0.05). The CLB group showed 98 per cent crowding alleviation compared with 67 per cent for the SLB after 4 months of alignment and levelling. Mini Diamond brackets aligned the teeth faster than Damon™ 3 but only during the first month. There was no difference in efficacy between the two groups in the later 3 weeks. Alleviation of crowding was faster with CLB than with SLB.

Differentiation analyses of adult suspension mononucleated peripheral blood cells of Mus musculus.

BACKGROUND: The purpose of this study is to determine whether isolated suspension mouse peripheral mononucleated blood cells have the potential to differentiate into two distinct types of cells, i.e., osteoblasts and osteoclasts. RESULTS: Differentiation into osteoblast cells was concomitant with the activation of the Opn gene, increment of alkaline phosphatase (ALP) activity and the existence of bone nodules, whereas osteoclast cells activated the Catk gene, increment of tartrate resistant acid phosphatase (TRAP) activity and showed resorption activities via resorption pits. Morphology analyses showed the morphology of osteoblast and osteoclast cells after von Kossa and May-Grunwald-Giemsa staining respectively. CONCLUSIONS: In conclusion, suspension mononucleated cells have the potentiality to differentiate into mature osteoblasts and osteoclasts, and hence can be categorized as multipotent stem cells.

Determination of the differentiation capacities of murines' primary mononucleated cells and MC3T3-E1 cells.

BACKGROUND: The main morphological features of primitive cells, such as stem and progenitor cells, are that these cells consists only one nucleus. The main purpose of this study was to determine the differentiation capacities of stem and progenitor cells. This study was performed using mononucleated cells originated from murine peripheral blood and MC3T3-E1 cells. Three approaches were used to determine their differentiation capacities: 1) Biochemical assays, 2) Gene expression analysis, and 3) Morphological observations. RESULTS: We found that both cells were able to differentiate into mature osteoblasts, as assayed by ALP activity. RT-PCR analysis showed the activation of the Opn gene after osteoblast differentiation. Morphological observations of both cells revealed the formation of black or dark-brown nodules after von Kossa staining. Nevertheless, only mononucleated cells showed the significant increase in TRAP activity characteristic of mature osteoclasts. The osteoclast-specific CatK gene was only upregulated in mononucleated cells. Morphological observations indicated the existence of multinucleated osteoclasts. Sca-1 was activated only in undifferentiated mononucleated cells, indicating that the cells were hematopoietic stem cells. In both cell lines, the housekeeping Gapdh gene was activated before and after differentiation. CONCLUSION: The isolated mononucleated cells were able to differentiate into both osteoblasts and osteoclasts; indicating that they are stem cells. On the other hand, MC3T3-E1 cells can only differentiate into osteoblasts; a characteristic of progenitor cells.