Genetic Comparison of Stemness of Human Umbilical Cord and Dental Pulp

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The Bio-Molecular Dynamics of Dental Pulp in Different Clinical Scenarios

Dental pulp (DP) is a very dynamic tissue both in health and in disease. When exposed to stressors and pathological conditions. It undergoes a complex series of biological reactions whereby alterations affect the pulp tissue at tissue cellular and molecular levels. The aim of this review is to update the reader on the various bio-molecular alterations in the dental pulp under different clinical conditions: orthodontic treatment (OT), caries, pulpitis and others. The morphological changes in the composition of the DP rang from the reversible remodeling to apoptosis and sometimes necrosis. Many apoptotic factors are involved like Bcl2, Bax and the significant increase in Caspases 9 and 3, as well as, Hsp60, its possible role and its mitochodrial localization. The inflammatory responses in dental pulp and the role of diffusible and cellular factors as well as DP stem cells were highlighted, in particular, where caries was involved in the pulpitis.Recent data report changes in tissue metabolism and homeostasis inside the DP caused by OT leading to increased levels of iNOS reactivity in the nerve fibers of the pulp. Moreover, remodeling of the extra cellular matrix(ECM) is an important feature in clinical scenarios like OT and caries whereby alterations in MMP-2 and MMP-9 expression patterns are reported leading to degradation of type IV and V collagens in the ECM. Furthermore, neurogenic factors are also modified after injuries and OT. Neuropeptides play a significant role not onlyin pain perception but also in vascular responses. Substance P increases in DP and enhances pain perception and so is the increase in CGRP which is correlated with concomitant gain in bone morphogenetic protein expression resulting in more dentin formation. The role of stem cells and the possible molecular mechanisms of dentin genesis are presented in this review. They focus on important signaling proteins and the possible role of various scaffolds in this regeneration process. In conclusion, most alterations inpulpal structure are reversible unless the pulp has a history of caries, restorations, trauma or prolonged heavy orthodontic forces. Pulpal symptoms arising from these clinical conditions should be treated appropriately and swiftly.Otherwise, exacerpation of pulpitis and the interplay of the various bio-molecular factors will lead to inhibition of repair and regeneration

Defining properties of neural crest-derived progenitor cells from the apex of human developing tooth

The connective tissue of the human tooth arises from cells that are derived from the cranial neural crest and, thus, are termed as "ectomesenchymal cells." Here, cells being located in a pad-like tissue adjacent to the apex of the developing tooth, which we designated the third molar pad, were separated by the microexplant technique. When outgrowing from the explant, dental neural crest-derived progenitor cells (dNC-PCs) adhered to plastic, proliferated steadily, and displayed a fibroblast-like morphology. At the mRNA level, dNC-PCs expressed neural crest marker genes like Sox9, Snail1, Snail2, Twist1, Msx2, and Dlx6. Cytofluorometric analysis indicated that cells were positive for CD49d (alpha4 integrin), CD56 (NCAM), and PDGFRalpha, while negative for CD31, CD34, CD45, and STRO-1. dNC-PCs could be differentiated into neurogenic, chondrogenic, and osteogenic lineages and were shown to produce bone matrix in athymic mice. These results demonstrate that human third molar pad possesses neural crest-derived cells that represent multipotent stem/progenitor cells. As a rather large amount of dNC-PCs could be obtained from each single third molar, cells may be used to regenerate a wide range of tissues within the craniofacial region of humans

Peripheral nerve glia as multipotent progenitors in craniofacial development

Craniofacial development is complex. Numerous populations of progenitor cells coordinate activities to produce an array of highly integrated tissues inside the developing head. However, it is not clear how some key multipotent progenitors continue to exist in late developing head compartments. The general hypothesis of this thesis is focused around the idea of an embryonic infrastructure represented by peripheral nerves that serves as a niche for glial multipotent neural crest-like cells. The nerve-adjacent glial cells can change their fate and be recruited in a targeted way to produce tissues at remote destinations during fast growth, development and regeneration. Results presented in this thesis explain how the nerves contribute pulp cells and matrix-producing cells of odontoblast lineage to the developing and growing tooth. Glial cells as an unexpected progenitor source give rise to almost half of all pulp cells and odontoblasts in the growing incisor. Furthermore, lineage tracing with colour-coding of individual recombination events allowed us to discover new aspects of tooth development and coordination between pulp cell lineage and odontoblasts. Another important component of the craniofacial compartment, the parasympathetic nervous system that targets glands in the head, is crucial for "rest-and-digest" or "feed and breed" activities especially during eating, salivation and lacrimation. Importantly, neurons of the autonomic parasympathetic nervous system are located very close to or inside the tissues they innervate and appear late in embryonic development. The discrepancy in developmental timing raised new questions: how do early neural crest-derived progenitors of parasympathetic neurons reach their destinations, and how do they acquire neuronal properties in situ? Furthermore, what is the nature of those progenitor cells? Our results clearly demonstrate that cells of glial origin located in the peripheral nervous system possess multipotency and gives rise to parasympathetic neurons during later developmental stages. Peripheral glial cells arrive to late-developing tissues on the pioneer presynaptic nerve fibres. Subsequently, some glial cells change fate, navigate for short distances and then convert into neurons and satellite cells of parasympathetic ganglia. Our conclusions redraw a fundamental principle on how the peripheral nervous system develops and provide a new type of logic, where both the cellular elements, as well as, the wiring are solved by a simple deposition of the postsynaptic elements from the presynaptic. During our work we used a wide spectrum of approaches including advanced genetic tracing with multicolor reporters, analysis of numerous mouse mutants, in vitro cell cultures and 3D imaging of developing embryos. We have applied both genetic and surgical ablation techniques to the peripheral nerves and investigated targeted recruitment of glia from the nerves in each case. Peripheral glia represents a novel amenable source of multipotent progenitor cells with putative regenerative potential that in the future might be applied for the treatment of congenital craniofacial pathologies, trauma cases or used for aesthetic body treatments

Assessment of the phenotypic effects of Platelet Rich Fibrin on Mesenchymal Stem Cells derived from Minced Pulp

Our aim is to investigate the effects of autologous platelet-rich fibrin (PRF) on Mesenchymal Stem cell derived from Minced Pulp (MP-MSCs). We first developed a mouse model of PRF to study the phenotypic effects of PRF in cultured cells. We obtained PRF from the blood and prepared PRF-enriched culture media. The phenotypic effects of PRF on MP-MSCs were determined by assessing the changes in cell proliferation, differentiation and immunophenotypic profiling. The mRNA levels of ALP, OCN, DMP1 and DSPP were determined by qRT-PCR. It was found that PRF increased the proliferation capacity of MP-MSCs and reduced the cell doubling time. With PRF exposure, the MP-MSCs were able to retain their immunophenotypic characteristics defining them as MSCs, as the cells expressing surface markers CD105, CD146 and CD73 were higher. MP-MSCs were able to undergo osteogenic differentiation in the presence of PRF and the mRNA levels of OCN was significantly increased in the presence of PRF. To assess the odontogenic differentiation of cells in response to PRF, we prepared dentin-slice model in which we cultured MP-MSCs embedded in PRF. Histological sections of the dentin slice model revealed that there was increase in the cellularity of the pulp tissue along the edges of the pulp tissue. Based on our findings, PRF can act as a source of growth factors cell proliferation, migration and differentiation

Glial cell line-derived neurotrophic factor influences proliferation of osteoblastic cells

Little is known about the role of neurotrophic growth factors in bone metabolism. This study investigated the short-term effects of glial cell line-derived neurotrophic factor (GDNF) on calvarial-derived MC3T3-E1 osteoblasts. MC3T3-E1 expressed GDNF as well as its canonical receptors, GFRα1 and RET. Addition of recombinant GDNF to cultures in serum-containing medium modestly inhibited cell growth at high concentrations; however, under serum-free culture conditions GDNF dose-dependently increased cell proliferation. GDNF effects on cell growth were inversely correlated with its effect on alkaline phosphatase (ALP) activity showing a significant dose-dependent inhibition of relative ALP activity with increasing concentrations of GDNF in serum-free culture medium. Live/dead and lactate dehydrogenase assays demonstrated GDNF did not significantly affect cell death or survival under serum-containing and serum-free conditions. The effect of GDNF on cell growth was abolished in the presence of inhibitors to GFR α 1 and RET indicating that GDNF stimulated calvarial osteoblasts via its canonical receptors. Finally, this study found that GDNF synergistically increased tumor necrosis factor-α (TNF-α)-stimulated MC3T3-E1 cell growth suggesting that GDNF interacted with TNF-α-induced signaling in osteoblastic cells. In conclusion, this study provides evidence for a direct, receptor-mediated effect of GDNF on osteoblasts highlighting a novel role for GDNF in bone physiology. \ud \u