Infrared diode laser enhances human periodontal ligament stem cells behaviour on titanium dental implants.

Low level laser treatment (LLLT) is known for its photobiostimulatory and photobiomodulatory characteristics, which stimulate cell proliferation, increase cellular metabolism, and improve cellular regeneration. The objective of the present research was to assess the possible influence of infrared diode laser irradiation on the behaviour, attachment, and osteogenic differentiation of human periodontal ligament stem cells (hPDLSCs) seeded on different types of dental implants. Two distinct types of implants, one subjected to laser surface treatment and the other treated with acid etching, were longitudinally divided into two halves and submerged in six wells culture plates. Both implants were subjected to infrared diode laser treatment, and subsequently, the morphology and attachment of cells were examined using scanning electron microscopy (SEM) after 14 and 21 days. The behaviour of (hPDLSCs) towards two types of implants, when exposed to osteogenic medium and low-level laser therapy (LLLT), was assessed using quantitative real-time polymerase chain reaction to measure the expression of stemness markers and osteogenic markers. The scanning electron microscopy (SEM) demonstrated that the application of infrared diode laser irradiation substantially improved the attachment of cells to both types of implants. The stemness gene markers were significantly down regulated in cells seeded on both surfaces when challenged with osteogenic media in relation to control. At 14 days, early osteogenic markers, were upregulated, while late osteogenic markers, were downregulated in both challenged groups. At the 21-day mark, hPDLSCs seeded on an acid-etched implant exhibited increased expression of all osteogenic markers in response to stimulation with osteogenic media and infra-red diode laser, in contrast to hPDLSCs seeded on a laser surface treated implant under the same conditions. Finally, the findings of our research revealed that when subjected to infrared diode laser, human periodontal ligament stem cells cultured on both types of implants demonstrated improved cellular attachment and differentiation. This suggested that infrared diode laser enhanced the activity of the cells surrounding the implants. Hence, the use of infrared diode laser could be pivotal in improving and expediting the clinical osseointegration process around dental implants.

The regenerative potential of Tideglusib and CHIR99021 small molecules as potent odontogenic differentiation enhancers of human dental pulp stem cells.

OBJECTIVES: To assess the effect of Tideglusib and CHIR99021 small molecules on the odontogenic differentiation potential of human dental pulp stem cells (hDPSCs) via Wnt/ß-catenin pathway activation. METHODOLOGY: hDPSCs were isolated from impacted third molars indicated for extraction and were characterized by flow cytometry. hDPSCs were then induced to differentiate into odontogenic lineage in the presence of Tideglusib and CHIR99021. Odontogenic differentiation was evaluated using Alizarin Red stain and RT-PCR for expression of odontogenic specific differentiation markers: DSPP, DMP1, ALP, OPN, and RUNX2 in relation to undifferentiated cells. RT-PCR was also conducted to assess the expression of Wnt/ß-catenin pathway activation marker (AXIN2). One-way ANOVA Kruskal-Wallis test was used for statistical analysis. RESULTS: Wnt/ß-catenin pathway was successfully activated by Tideglusib and CHIR99021 in hDPSCs where AXIN2 was significantly upregulated. Successful odontogenic differentiation was confirmed by Alizarin Red staining of calcified nodules. RT-PCR for odontogenic differentiation markers DSPP, DMP1, and RUNX expression by hDPSCs induced by CHIR99021 was higher than that expressed by hDPSCs induced by Tideglusib, whereas expression of OPN and ALP was higher in Tideglusib-induced cells than in CHIR99021-induced cells. CONCLUSIONS: Both small molecules successfully induced odontogenic differentiation of hDPSCs through Wnt/ß-catenin pathway activation. CLINICAL RELEVANCE: These findings suggest that Tideglusib and CHIR99021 can be applied clinically in pulp regeneration to improve strategies for vital pulp regeneration and to promote dentine repair.

Small Molecule GSK-3 Inhibitors Safely Promote the Proliferation and Viability of Human Dental Pulp Stem Cells-In Vitro.

Small molecules have demonstrated promising results as successful alternatives to growth factors. In this study, focus was drawn to CHIR99021 and tideglusib as GSK-3 inhibitors known for their anti-inflammatory and regenerative potential. The effect of both tideglusib and CHIR99021 on the proliferation, viability, and stemness of human dental pulp stem cells (hDPSCs) was investigated to assess their possible role in regenerative dentistry. Briefly, hDPSCs were isolated from sound premolars extracted for orthodontic purposes. Cytotoxicity and proliferation assessment were performed via cell counting kit-8 followed by flow cytometric analysis of apoptotic marker ANNEXIN V. The effect of both small molecules on the stemness of hDPSCs was analyzed by qRT-PCR. Both tideglusib and CHIR99021 were proven to be safe on hDPSCs. The tideglusib concentration that resulted in higher viable cells was 100 nM, while the concentration for CHIR99021 was 5 nM. Both small molecules successfully induced cellular proliferation and demonstrated minimal expression of ANNEXIN V, indicative of the absence of cellular apoptosis and further confirming their positive effect on proliferation. Finally, both small molecules enhanced stemness markers expression as evidenced by qRT-PCR, which, again, highlighted the positive effect of both tideglusib and CHIR99021 on safely promoting the proliferation of hDPSCs while maintaining their stemness.

Optimization of differentiation protocols of dental tissues stem cells to pancreatic ß-cells.

BACKGROUND: Despite the recent progress in the differentiation strategies of stem cells into pancreatic beta cell lineage, current protocols are not optimized for different cell types. The purpose of this study is to investigate and compare the ability of stem cells derived from dental pulp (DPSCs) and periodontal ligament (PDLSCs) as two anatomically different dental tissues to differentiate into pancreatic beta cells while assessing the most suitable protocol for each cell type. METHODS: DPSCs & PDLSCs were isolated and characterized morphologically and phenotypically and then differentiated into pancreatic beta cells using two protocols. Differentiated cells were assessed by qRT-PCR for the expression of pancreatic related markers Foxa-2, Sox-17, PDX-1, Ngn-3, INS and Gcg. Functional assessment of differentiation was performed by quantification of Insulin release via ELISA. RESULTS: Protocol 2 implementing Geltrex significantly enhanced the expression levels of all tested genes both in DPSCs & PDLSCs. Both DPSCs & PDLSCs illustrated improved response to increased glucose concentration in comparison to undifferentiated cells. Moreover, DPSCs demonstrated an advanced potency towards pancreatic lineage differentiation over PDLSCs under both protocols. CONCLUSION: In conclusion, the current study reports the promising potential of dental derived stem cells in differentiating into pancreatic lineage through selection of the right protocol.

Efficient generation of functional pancreatic ß cells from dental-derived stem cells via laminin-induced differentiation.

BACKGROUND: This study was designed to generate functional insulin-producing cells (IPCs) from dental-derived mesenchymal stem cells (MSCs) and further explore their therapeutic potential against diabetes mellitus in vivo. MSCs were isolated from human dental pulp and periodontal ligament and were induced to differentiate into insulin-producing cells (IPCs) using laminin-based differentiation protocol for 14 days. Confirmation of IPCs was performed through real-time PCR analysis and insulin release assay. Then, the generated IPCs were labeled with PKH26 dye prior to transplantation in experimental animals. Twenty-eight days later, blood glucose, serum insulin (INS), c-peptide (CP), and visfatin (VF) levels and pancreatic glucagon (GC) level were estimated. Pancreatic forkhead box protein A2 (Foxa2) and SRY-box transcription factor 17 (Sox17), insulin-like growth factor I (IGF-1), and fibroblast growth factor10 (FGF 10) gene expression levels were analyzed. RESULTS: Dental stem cells were successfully differentiated into IPCs that demonstrated increased expression of pancreatic endocrine genes. IPCs released insulin after being subjected to high levels of glucose. In vivo findings uncovered that the implanted IPCs triggered significant decrease in blood glucose, serum VF, and pancreatic GC levels with significant increase in serum INS and CP levels. Furthermore, the implanted IPCs provoked significant upregulation in the expression level of pancreatic genes. Histopathological description of the pancreas tissues revealed that transplantation of IPCs ameliorated the destabilization of pancreas tissue architecture. CONCLUSION: This study demonstrates the significant role of the implantation of IPCs generated from dental-derived stem cells in treatment of diabetes mellitus.

Preconditioned human dental pulp stem cells with cerium and yttrium oxide nanoparticles effectively ameliorate diabetic hyperglycemia while combatting hypoxia.

The development of efficient insulin producing cells (IPC) induction system is fundamental for the regenerative clinical applications targeting Diabetes Mellitus. This study was set to generate IPC from human dental pulp stem cells (hDPSCs) capable of surviving under hypoxic conditions in vitro and in vivo. METHODS: hDPSCs were cultured in IPCs induction media augmented with Cerium or Yttrium oxide nanoparticles along with selected growth factors & cytokines. The generated IPC were subjected to hypoxic stress in vitro to evaluate the ability of the nanoparticles to combat hypoxia. Next, they were labelled and implanted into diabetic rats. Twenty eight days later, blood glucose and serum insulin levels, hepatic hexokinase and glucose-6-phosphate dehydrogenase activities were measured. Pancreatic vascular endothelial growth factor (VEGF), pancreatic duodenal homeobox1 (Pdx-1), hypoxia inducible factor 1 alpha (HIF-1α) and Caspase-3 genes expression level were evaluated. RESULTS: hDPSCs were successfully differentiated into IPCs after incubation with the inductive media enriched with nanoparticles. The generated IPCs released significant amounts of insulin in response to increasing glucose concentration both in vitro & in vivo. The generated IPCs showed up-regulation in the expression levels of anti-apoptotic genes in concomitant with down-regulation in the expression levels of hypoxic, and apoptotic genes. The in vivo study confirmed the homing of PKH-26-labeled cells in pancreas of treated groups. A significant up-regulation in the expression of pancreatic VEGF and PDX-1 genes associated with significant down-regulation in the expression of pancreatic HIF-1α and caspase-3 was evident. CONCLUSION: The achieved results highlight the promising role of the Cerium & Yttrium oxide nanoparticles in promoting the generation of IPCs that have the ability to combat hypoxia and govern diabetes mellitus.

Current state of stem cell-based therapies: an overview.

Recent research reporting successful translation of stem cell therapies to patients have enriched the hope that such regenerative strategies may one day become a treatment for a wide range of vexing diseases. In fact, the past few years witnessed, a rather exponential advancement in clinical trials revolving around stem cell-based therapies. Some of these trials resulted in remarkable impact on various diseases. In this review, the advances and challenges for the development of stem-cell-based therapies are described, with focus on the use of stem cells in dentistry in addition to the advances reached in regenerative treatment modalities in several diseases. The limitations of these treatments and ongoing challenges in the field are also discussed while shedding light on the ethical and regulatory challenges in translating autologous stem cell-based interventions, into safe and effective therapies.

Nanotechnological Applications Hold a Pivotal Position in Boosting Stem Cells Osteogenic Activity: In Vitro and In Vivo Studies.

This approach was constructed to appraise the therapeutic effectiveness of a single i.v. dose of osteoblasts generated from co-culturing BM-MSCs with nano-HA, Pt-NPs, or Pt-HA-nanocomposite in osteoporotic rats. MSCs were grown, propagated in culture, and characterized. The effect of the suggested nanoplatforms on the survival, osteogenic differentiation, and mineralization of BM-MSCs was assessed by MTT assay, real-time PCR analysis, and Alizarin red S staining, respectively. Thereafter, the generated osteoblasts were employed for the treatment of ovariectomized rats. Our results revealed that the selected nanoplatforms upregulate the expression of osteogenic differentiation related genes (Runx-2 and BMP-2) significantly and enhance calcium deposition in BM-MSCs after 7 and 21 days, respectively, whereas the in vivo study validated that the infusion of the generated osteoblasts considerably downturn serum BALP, BSP, and SOST levels; upswing OSX level; and regain femur bone mineralization and histoarchitecture. Conclusively, the outcomes of this work provide scientific evidence that transplanting osteoblasts derived from differentiation of BM-MSCs in the presence of nanoplatforms in ovariectomized rats restores bone remodeling balance which constitutes a new hope for the treatment of osteoporosis.

Optimizing a serum-free/xeno-free culture medium for culturing and promoting the proliferation of human dental pulp stem cells.

BACKGROUND: Dental pulp stem cells (DPSCs) hold great promise for utilization in tissue repair and regenerative medicine. Routinely, culture media used for culturing stem cells are supplemented with animal serum for promoting growth and successful maintenance of stem cells. However, there is a growing demand for optimizing a well-defined culture media that could safely increase the efficacy and reproducibility of the cultured cells. In this study, we aimed at optimizing a serum-free/xeno-free culture medium. METHODS: A cocktail of various supplements intended to enrich DPSCs proliferation in defined concentrations was designed. It consisted of recombinant human basic fibroblast growth factor (hbFGF), insulin transferrin selenium (ITS), ascorbic acid (vitamin C), Beta mercaptoethanol and cholesterol. The effect of this optimized media on the proliferation of DPSCs was assessed by MTT assay and flow cytometric analysis (FACS) of early apoptotic marker annexin V. Expression of stemness-related genes (OCT4, SOX and NANOG) was assessed by qRT-PCR. RESULTS: Proliferation results by MTT illustrated a significant increase in the proliferation rate of DPSCs cultured in the proposed media. FACS analysis of annexin V expression was nil. Expression of OCT4, SOX and NANOG genes was also up-regulated. CONCLUSIONS: The proposed combination of supplements utilized in the proposed culture media successfully increased the proliferation potential of DPSCs in addition to enhancing the stemness properties. Thus, it can be considered a promising and safe substitute to traditional animal derived supplements like fetal bovine serum (FBS).

Nano Hydroxyapatite & Mineral Trioxide Aggregate Efficiently Promote Odontogenic Differentiation of Dental Pulp Stem Cells.

BACKGROUND: There has been an urge to shift from conventional therapies to the more promising regenerative strategy since conventional treatment relies on synthetic materials to fill defects and replace missing tissues, lacking the ability to restore the tissues' physiological architecture and function. AIM: The present study focused on the assessment of the role of two commonly used biomaterials namely; mineral trioxide aggregate (MTA) and nano hydroxy-apatite as promoters of odontogenic differentiation of dental pulp stem cells (DPSCs). METHODS: DPSCs were isolated, cultured in odontogenic media and divided into three groups; control group, MTA group and nanohydroxyapatite group. Odontogenic differentiation was assessed by tracing genes characteristic of different stages of odontoblasts via qRT-PCR. Calcific nodules formation was evaluated by Alizarin red staining. RESULTS: Results demonstrated that both MTA and nanohydroxyapatite were capable of enhancing odontogenic differentiation of DPSCs. CONCLUSION: Nano hydroxyapatite was found to have a higher promoting effect. However, in the absence of an odontogenic medium, MTA and nanohydroxyapatite could not enhance the odontogenic differentiation of DPSCs.

Preclinical Assessment of the Proliferation Capacity of Gingival and Periodontal Ligament Stem Cells from Diabetic Patients.

BACKGROUND: Stem cells have recently received great interest as potential therapeutics alternative for a variety of diseases. The oral and maxillofacial region, in particular, encompasses a variety of distinctive mesenchymal (MSC) populations and is characterized by a potent multilineage differentiation capacity. AIM: In this report, we aimed to investigate the effect of diabetes on the proliferation potential of stem cells isolated from controlled diabetic patients (type 2) and healthy individuals. SUBJECTS & METHODS: The proliferation rate of gingival and periodontal derived stem cells isolated from diabetic & healthy individuals were compared using MTT Assay. Expression levels of Survivin in isolated stem cells from all groups were measured by qRt - PCR. RESULTS: There was a significantly positive correlation between proliferation rate and expression of Survivin in all groups which sheds light on the importance of Survivin as a reliable indicator of proliferation. The expression of Survivin further confirmed the proliferation results from MTT Assay where the expression of stem cells from non - diabetic individuals was higher than diabetic patients. CONCLUSION: Taking together all the results, it could be concluded that PDLSC and GSC are promising candidates for autologous regenerative therapy due to their ease of accessibility in addition to their high proliferative rates.

Relationship of deciduous teeth emergence with physical growth.

INTRODUCTION: Tooth eruption, recognized as an aspect of human growth and development, could possibly be influenced by a number of factors. It may reflect the general body development. AIM: The aim of the present research is to investigate the relationship of deciduous teeth emergence with physical growth (weight/height indices). MATERIALS AND METHODS: A study was conducted among a sample of 1132 Egyptian infants whose ages range from 4 to 36 months. The sample was collected from some randomly selected health centers affiliated to the ministry of health where various socioeconomic strata from different geographic localities were recruited. The children visit there regularly for vaccination at definite ages. Weight and height were evaluated as factors that might have influence on the time of deciduous teeth emergence. RESULTS: The results showed a certain degree of correlation between the number of deciduous teeth emerged and the studied anthropometric measurements which differed by sex and age. CONCLUSION: All anthropometric parameters showed relationship with the number of teeth at different levels. Although weight showed influence on the number of teeth emerged, it was less significant than height.