Dental stem cell therapy with calcium hydroxide in dental pulp capping

Calcium hydroxide has been extensively and steadily used for direct pulp capping in modern clinical dentistry. As it was known to have potential to induce hard tissue repair, this chemical has been applied to the exposed dental pulp and the hard tissue is expected to be regenerated above the pulp. During the reparative process of exposed pulp, primary odontoblasts that were lost as a result of extensive damage are replaced with newly differentiated odontoblast-like cells. This process is known to follow the sequential steps of proliferation, migration, and differentiation of progenitor cells. This research will examine the relationship between calcium hydroxide and the recruitment, proliferation, and mineralization of postnatal dental stem cells, obtained from an immature dental tissue of beagle dogs. Immunocytochemical staining and reverse transcriptase-polymerase chain reaction were used to identify the putative stem cell markers. Immunoblot analysis, wound healing assay, cell migration assay, and alizarin red staining were used to evaluate proliferation, migration, and mineralization capacity of the calcium hydroxide-treated stem cells. As an in vivo study, a combination of calcium hydroxide and autologous dental pulp stem cells (DPSCs) was applied for the treatment of intentionally created tooth defects on the premolars and the molars in beagle dogs to observe dentin regeneration. Ex vivo expanded DPSCs and periodontal ligament stem cells expressed STRO-1 and CD146, the mesenchymal stem cell markers. It was evident that calcium hydroxide increased recruitment, migration, proliferation, and mineralization of the DPSCs and periodontal ligament stem cells. Such results are valuable for future availability of DPSCs, which are recently focused as the stem cell reservoir for regeneration of dentin upon tooth injury, as well as for elucidation of the role of calcium hydroxide in pulp capping therapy.This work was supported by grants from the National Research Foundation of Korea (NRF) grant funded by the Korean government (MEST) (No. 2009-0066366) to Pill-Hoon Choung and by the Ministry of Health and Welfare, the Republic of Korea, through the Musculoskeletal Bioorgans Center Program (no. 0405-BO01-0204-0006)

Molecular Responses in Osteogenic Differentiation of Mesenchymal Stem Cells Induced by Physical Stimulation

Mesenchymal stem cells (MSCs) have been recognized as a great source of stem cells in the field of regenerative medicine and regulation of MSCs such as differentiation into specific cells. Particular interest is the use of physical stimulation for the expression of the osteoblast-specific genes from MSCs for bone tissue regeneration. The mechanical forces on MSCs, such as fluid flow, enhance the mineralized matrix and specific gene expressions. This process called mechanotransduction comprises of the steps of mechanoreception, biochemical coupling, transmission of signal and effector cell response. Physical stimuli effectively regulate extracellular and intracellular signaling pathways to enhance the expression of specific transcription factors, and the release of osteocytes, ultimately expedite the production of active osteoblasts. Thus understating, identification and functional characterization of the mechanotransduction underlying the physical stimulation of MSCs is a critical issue for devising new bone regenerative treatments for bone-related diseases. In this review, we focus on the molecular mechanism responsible for the mechanotransduction of osteogenic differentiation of MSCs induced by physical stimulation.

Establishment and Characterization of an In Vitro Model for Cholesteatoma

ObjectivesExperimental models are of importance to study the pathogenesis of middle ear cholesteatoma, however, they were not established until now. We aimed to develop in vitro model of middle ear cholesteatoma using primary keratinocytes and fibroblasts isolated from cholesteatoma tissue. HaCaT cell line was used as a "skin equivalent" and to compare the grade of homogeneity between cholesteatoma keratinocytes and HaCaT cells.MethodsPrimary keratinocytes were isolated from cholesteatoma tissue, co-cultured with preliminary prepared feeder layer from cholesteatoma fibroblasts and subsequently air-exposed. The protein profile of cholesteatoma keratinocytes and HaCaT cells was evaluated by means of immunoblot using monoclonal antibody against cytokeratin (CK) 13 and 16. Tissue localization of CK 13 and 16 was accomplished with immunohistochemistry.ResultsDifferent protein profile and stronger expression of CK 13 and 16 were demonstrated in cholesteatoma keratinocytes in comparison with HaCaT cells. Bigger stratification was observed in the 3D-in vitro systems when both cholesteatoma keratinocytes and HaCaT cells were respectively co-cultured with fibroblasts in comparison with the corresponding control groups without fibroblasts.Conclusion3D-model demonstrates the significance of intercellular interaction between components of cholesteatoma tissue

Congenital Stapes Anomalies with Normal Eardrum

ObjectivesA non-progressive and conductive hearing loss with normal eardrum, but no history of trauma and infection, is highly suggestive of a congenital ossicular malformation. Among ossicular anomalies, stapes anomaly is the most common. The purpose of this study is to describe patterns of stapes anomaly and to analyze its surgical outcome with special reference to its patterns.MethodsWe conducted a retrospective case review. The subjects comprised 66 patients (76 ears) who were decisively confirmed by the exploratory tympanotomy as congenital stapes anomalies without any anomalies of the tympanic membrane and external auditory canal. The preoperative and postoperative audiological findings, temporal bone computed tomography scan, and operative findings were analyzed.ResultsThere were 16 anomalous patterns of stapes among which footplate fixation was the most common anomaly. These 16 patterns were classified into 4 types according to the status of stapes footplate. Successful hearing gain was achieved in 51 out of 76 ears (67.1%) after surgical treatment.ConclusionFootplate fixation was usually bilateral, whereas stapes anomalies associated with other ossicular anomaly were usually unilateral. The success of the surgical treatment of stapes anomaly might depend on its developmental status of the footplate. Stapes anomalies were detected without any fixed patterns, therefore, it is quite possible to detect a large variety of patterns in future

Management for the Children with Otitis Media with Effusion in the Tertiary Hospital

ObjectivesRecently, new evidence-based recommendations have been introduced for diagnosing and managing otitis media with effusion (OME) in children. However, there are some difficulties to follow the general guidelines in the tertiary hospitals. The purpose is to evaluate the efficiency of antibiotics or antihistamines for treatment of children with OME in the tertiary hospital with a randomized prospective clinical study.MethodsEighty-four children with OME who had been diagnosed in the tertiary hospital were randomized to receive 5 different medications for 2 weeks. We prescribed antibiotics (amoxicillin-clavulanate syrup) in Group I (n=16), antibiotics/steroids (prednisolone) in Group II (n=18), antibiotics/antihistamines (ebastine) in Group III (n=15), antibiotics/steroids/antihistamines in Group IV (n=17), and mucolytics (ivy leaf extract) in Group V (n=17) for control. We followed-up children every 2 weeks and evaluated the state of OME at 3 months.ResultsThirty six (42.9%) of 84 children were resolved within average 6.9 weeks after the treatments. Thirty-six (42.9%) were treated with ventilation tube insertion and 12 patients (14.3%) were observed. There was no difference in the resolution rates of OME among the five different protocols (P>0.05). There was no difference in the resolution rates among groups who used steroids, antihistamines, steroids and antihistamines, or other medications to manage 42 children with allergies (P>0.05).ConclusionIn the tertiary hospital, the cure rate of children with OME was not as high as well-known, and antibiotics or anti-allergic medications were not more effective than control. We may, therefore, need any other guidelines which are different from the previous evidence-based recommendations, including early operation in the tertiary hospitals

CC Chemokine Ligand 3 Overcomes the Bacteriocidal and Phagocytic Defect of Macrophages and Hastens Recovery from Experimental Otitis Media in TNF-/- Mice

Innate immune mechanisms are crucial in defense against bacterial illnesses in humans, as evidenced by abnormal antibacterial responses due to defects in TLR signaling, seen in children with MyD88 or IL-1R–associated kinase 4 deficiency. Otitis media (OM) is the most common disease of childhood, and the role of innate immune molecules in this disorder remains unclear. In a murine model of OM, we show that, in the absence of TNF, a key effector of innate immunity, this disease is prolonged after middle ear infection with nontypeable Haemophilus influenzae (NTHi). In the absence of TNF, mice fail to upregulate both TLRs and downstream genes and proteins, such as CCL3, resulting in defects in both inflammatory cell recruitment and macrophage function. Peritoneal macrophages of mice lacking TNF have a diminished ability to phagocytose and kill NTHi, and this defect is partially corrected in vitro by exogenous rTNF. Addition of rCCL3 alone or in combination with rTNF restores phagocytosis and killing by TNF-deficient macrophages to that of unstimulated wild-type macrophages. In vivo administration of rCCL3 to animals deficient in TNF fully restores the ability to control OM due to NTHi, whereas a CCL3-blocking Ab impaired the ability of wild-type mice to recover from OM. Thus, CCL3 is a potent downstream effector of TNF-mediated inflammation in vitro and in vivo. Manipulation of CCL3 and/or TNF may prove to be effective therapeutic approaches in OM or other conditions associated with defective TNF generation

25th annual computational neuroscience meeting: CNS-2016

The same neuron may play different functional roles in the neural circuits to which it belongs. For example, neurons in the Tritonia pedal ganglia may participate in variable phases of the swim motor rhythms [1]. While such neuronal functional variability is likely to play a major role the delivery of the functionality of neural systems, it is difficult to study it in most nervous systems. We work on the pyloric rhythm network of the crustacean stomatogastric ganglion (STG) [2]. Typically network models of the STG treat neurons of the same functional type as a single model neuron (e.g. PD neurons), assuming the same conductance parameters for these neurons and implying their synchronous firing [3, 4]. However, simultaneous recording of PD neurons shows differences between the timings of spikes of these neurons. This may indicate functional variability of these neurons. Here we modelled separately the two PD neurons of the STG in a multi-neuron model of the pyloric network. Our neuron models comply with known correlations between conductance parameters of ionic currents. Our results reproduce the experimental finding of increasing spike time distance between spikes originating from the two model PD neurons during their synchronised burst phase. The PD neuron with the larger calcium conductance generates its spikes before the other PD neuron. Larger potassium conductance values in the follower neuron imply longer delays between spikes, see Fig. 17.Neuromodulators change the conductance parameters of neurons and maintain the ratios of these parameters [5]. Our results show that such changes may shift the individual contribution of two PD neurons to the PD-phase of the pyloric rhythm altering their functionality within this rhythm. Our work paves the way towards an accessible experimental and computational framework for the analysis of the mechanisms and impact of functional variability of neurons within the neural circuits to which they belong

Vascularized bone flap for access on the maxillary sinus

This article describes a vascularized bony window for access to the maxillary sinus and reports the clinical results. A bony U-shaped window in the anterior sinus wall was pedicled on the surrounding soft tissue and periosteum. After the described sinus was cleared of disease, the window was repositioned in its original site either using resorbable sutures or not. The method was used in 47 maxillary sinus operations in 45 patients. Twenty-four patients were followed-up for more than 48 months. The vascularized bony window technique showed uneventful healing in all patients and none of the 24 patients reported any problems. The vascularized bony window technique provides a large antrostomy, which gives good access and visibility and results in satisfactory postoperative healing

Regulating the role of bone morphogenetic protein 4 in tooth bioengineering

Purpose Culture of the whole organ and regulation of its development using biologic and engineering principles can be used to produce structures and organs for reconstructing defects. The application of these bioengineering approaches in artificial tooth development may be the alternative way to replace missing dentition. Materials and Methods For the artificial bioengineering of a mouse tooth, tooth buds were dissected and transplanted into the diastema of the developing mandible. The mandiblular primordia containing transplanted tooth buds were culture in vitro and in vivo using a bioengineering method. In addition, to regulate the development of tooth germs, bone morphogenetic protein 4 (BMP4) or its antagonist, Noggin was administered. Results After the period of in vitro and in vivo culture, the transplanted tooth germ in the diastema showed tooth development with supportive structure formation. In the BMP-treated group, the bioengineered tooth was observed with increased maturation of cusp and enamel matrix. However, in the Noggin-treated tooth germs, the developing molar had a crater-like appearance with the immature development of the cusp and suppressed formation of the enamel matrix. Conclusions This study confirmed that tooth germ transplantation in the diastema and culture with administration of BMP4 could lead to the mature development of the dental structures. In addition, these results suggest the possibility of bioengineering the tooth in morphogenesis and differentiation even in the toothless area

Dental Stem Cell Therapy with Calcium Hydroxide in Dental Pulp Capping

Calcium hydroxide has been extensively and steadily used for direct pulp capping in modern clinical dentistry. As it was known to have potential to induce hard tissue repair, this chemical has been applied to the exposed dental pulp and the hard tissue is expected to be regenerated above the pulp. During the reparative process of exposed pulp, primary odontoblasts that were lost as a result of extensive damage are replaced with newly differentiated odontoblast-like cells. This process is known to follow the sequential steps of proliferation, migration, and differentiation of progenitor cells. This research will examine the relationship between calcium hydroxide and the recruitment, proliferation, and mineralization of postnatal dental stem cells, obtained from an immature dental tissue of beagle dogs. Immunocytochemical staining and reverse transcriptase-polymerase chain reaction were used to identify the putative stem cell markers. Immunoblot analysis, wound healing assay, cell migration assay, and alizarin red staining were used to evaluate proliferation, migration, and mineralization capacity of the calcium hydroxide-treated stem cells. As an in vivo study, a combination of calcium hydroxide and autologous dental pulp stem cells (DPSCs) was applied for the treatment of intentionally created tooth defects on the premolars and the molars in beagle dogs to observe dentin regeneration. Ex vivo expanded DPSCs and periodontal ligament stem cells expressed STRO-1 and CD146, the mesenchymal stem cell markers. It was evident that calcium hydroxide increased recruitment, migration, proliferation, and mineralization of the DPSCs and periodontal ligament stem cells. Such results are valuable for future availability of DPSCs, which are recently focused as the stem cell reservoir for regeneration of dentin upon tooth injury, as well as for elucidation of the role of calcium hydroxide in pulp capping therapy.This work was supported by grants from the National Research Foundation of Korea (NRF) grant funded by the Korean government (MEST) (No. 2009-0066366) to Pill-Hoon Choung and by the Ministry of Health and Welfare, the Republic of Korea, through the Musculoskeletal Bioorgans Center Program (no. 0405-BO01-0204-0006).