Using qPCR and ddPCR to study biodistribution of cell therapy products: a multi-site evaluation.

BACKGROUD AIMS: Regenerative therapies employing cell therapy products (CTPs) have attracted considerable attention. Biodistribution (BD) evaluation of CTPs is mainly performed to clarify the cell survival time, engraftment, and distribution site. This evaluation is crucial for predicting the efficacy and safety profiles of clinical studies based on non-clinical BD study outcomes. However, no internationally unified method has been established for assessing cell BD after administration. Here, we aimed to standardize the BD assay method used for CTPs, conducting the following evaluations using the same protocol across multiple study facilities: (1) in vitro validation of quantitative polymerase chain reaction (qPCR) and droplet digital PCR (ddPCR) analyses using the primate-specific Alu gene, and (2) in vivo BD studies after the intravenous administration of human mesenchymal stem cells (hMSCs) to immunodeficient mice, commonly used in non-clinical tumorigenicity studies. METHODS: Quality control samples were prepared and analyzed by adding a fixed number of human-derived cells to several mouse tissues. The respective quantitative performances of the qPCR and ddPCR methods were compared for accuracy and precision. hMSCs were intravenously administered to immunodeficient mice, and tissues were collected at 1, 4, and 24 h after administration. RESULTS: Both methods demonstrated an accuracy (relative error) generally within ±50% and a precision (coefficient of variation) generally less than 50%. While differences in calibration curve ranges were observed between qPCR and ddPCR, no significant differences in quantification were found among the assay facilities. The BD of hMSCs in mice was evaluated at seven facilities (qPCR at three facilities; ddPCR at four facilities), revealing similar tissue distribution profiles in all facilities, with the lungs showing the highest cell distribution among the tissues tested. CONCLUSIONS: Quantitative evaluation of qPCR and ddPCR using Alu sequences was conducted, demonstrating that the test method can be adapted for BD evaluation.

Mesial or distal to canine: Which is better for the position of closing loops? Analysis of tooth movements based on numerical simulation.

INTRODUCTION: Although many studies investigating the mechanical behavior of loop mechanics have focused on loop designs to produce a higher moment-to-force ratio, few studies have clarified the effect of loop position on the force system and resultant tooth movements. This study aimed to simulate orthodontic tooth movements during space closure and to compare the effects of loop position in association with different degrees of gable bend on tooth movements using the finite element method. METHODS: Two finite element models of the maxillary dentition were constructed, with the loop placed mesial or distal to the canine. Tooth movements during loop activation were simulated while varying the degree of gable bend. RESULTS: When the loop was placed distal to the canine, the incisor showed uncontrolled tipping even with the gable bend. Placement of the loop mesial to the canine produced controlled tipping or root movement of the incisor, depending on the degree of gable bend. CONCLUSIONS: Placement of the closing loop mesial to the canine in combination with the incorporation of a gable bend into the archwire distal to the canine could provide better control of incisor movements, such as controlled tipping or root movement, as compared with placement of a gable bend into the loop located distal to the canine.

Biomechanical features of tooth movement from a lingual appliance in comparison with a labial appliance during space closure in sliding mechanics.

INTRODUCTION: The objectives of this study were to simulate long-term orthodontic tooth movement in en-masse retraction using the finite element method and investigate the effects of power arms on tooth movements when using a lingual appliance in comparison with a labial appliance. METHODS: A 3-dimensional finite element model of the maxillary dentition was constructed with 0.018-in brackets and 0.016 × 0.022-in stainless steel archwire. An en-masse retraction was performed by applying retraction force at various lengths of the power arm (4, 6, 8, and 10 mm) to the second molar tube, and long-term tooth movements with the lingual and labial appliances were analyzed using the finite element method. RESULTS: Although lingual crown tipping of the incisor was more marked with the lingual appliance than with the labial appliance in the early phase of space closure, only a slight difference was evident after space closure. Although the power arm was effective for achieving better-controlled tooth movement and reducing vertical and transverse bowing effects, bodily movement of the incisor could not be achieved, and bowing effects could not be eliminated. CONCLUSIONS: To provide better torque control of the incisor or prevent a vertical bowing effect, the incorporation of extra torque into brackets of incisors was recommended, and the use of power arms for the lingual appliance. To prevent a transverse bowing effect, incorporation of the antibowing bend or application of retraction force from both buccal and lingual sides or temporary skeletal anchorage devices was recommended.

Visualization of mandibular movement relative to the maxilla during mastication in mice: integration of kinematic analysis and reconstruction of a three-dimensional model of the maxillofacial structure.

BACKGROUND: Mastication is one of the most fundamental functions for the conservation of human life. To clarify the pathogenetic mechanism of various oral dysfunctions, the demand for devices for evaluating stomatognathic function has been increasing. The aim of the present study was to develop a system to reconstruct and visualize 3-dimensional (3D) mandibular movements relative to the maxilla, including dynamic transition of occlusal contacts between the upper and lower dentitions during mastication in mice. METHODS: First, mandibular movements with six degrees of freedom were measured using a motion capture system comprising two high-speed cameras and four reflective markers. Second, 3D models of maxillofacial structure were reconstructed from micro-computed tomography images. Movement trajectories of anatomical landmark points on the mandible were then reproduced by integrating the kinematic data of mandibular movements with the anatomical data of maxillofacial structures. Lastly, 3D surface images of the upper dentition with the surrounding maxillofacial structures were transferred to each of the motion capture images to reproduce mandibular movements relative to the maxilla. We also performed electromyography (EMG) of masticatory muscles associated with mandibular movements. RESULTS: The developed system could reproduce the 3D movement trajectories of arbitrary points on the mandible, such as incisor, molars and condylar points with high accuracy and could visualize dynamic transitions of occlusal contacts between upper and lower teeth associated with mandibular movements. CONCLUSIONS: The proposed system has potential to elucidate the mechanisms underlying motor coordination of masticatory muscles and to clarify their roles during mastication by taking advantage of the capability to record EMG data synchronously with mandibular movements. Such insights will enhance our understanding of the pathogenesis and diagnosis of oral motor disorders by allowing comparisons between normal mice and genetically modified mice with oral behavioral dysfunctions.

Biodistribution studies for cell therapy products: Current status and issues.

Information on the biodistribution (BD) of cell therapy products (CTPs) is essential for prediction and assessment of their efficacy and toxicity profiles in non-clinical and clinical studies. To conduct BD studies, it is necessary to understand regulatory requirements, implementation status, and analytical methods. This review aimed at surveying international and Japanese trends concerning the BD study for CTPs and the following subjects were investigated, which were considered particularly important: 1) comparison of guidelines to understand the regulatory status of BD studies in a global setting; 2) case studies of the BD study using databases to understand its current status in cell therapy; 3) case studies on quantitative polymerase chain reaction (qPCR) used primarily in non-clinical BD studies for CTPs; and 4) survey of imaging methods used for non-clinical and clinical BD studies. The results in this review will be a useful resource for implementing BD studies.

Simulation of orthodontic tooth movement during activation of an innovative design of closing loop using the finite element method.

INTRODUCTION: Although many attempts have been made to study the mechanical behavior of closing loops, most have been limited to analyses of the magnitude of forces and moments acting on the end of the closing loop. The objectives of this study were to simulate orthodontic tooth movement during the activation of a newly designed closing loop combined with a gable bend and to investigate the optimal loop activation condition to achieve the desired tooth movement. METHODS: We constructed a 3-dimensional model of maxillary dentition reproducing the state wherein a looped archwire combined with a gable bend was engaged in brackets and tubes. Orthodontic tooth movements were simulated for both anterior and posterior teeth while varying the degree of gable bend using the finite element method. RESULTS: The incorporation of a 5° gable bend into the newly designed closing loop produced lingual crown tipping for the central incisor and bodily movement for the first molar. The incorporation of 10° and 15° gable bends produced bodily movement and root movement, respectively, for the central incisor and distal tipping for the first molar. CONCLUSIONS: Torque control of the anterior teeth and anchorage control of the posterior teeth can be carried out effectively and simply by reducing by half the thickness of a teardrop loop with a height of 10 mm and a 0.019 × 0.025-in cross-section, to a distance of 3 mm from its apex, and by incorporating various degrees of gable bend into the loop corresponding to the treatment plan.

Biomechanical considerations for uprighting impacted mandibular molars.

This case report demonstrates two different uprighting mechanics separately applied to mesially tipped mandibular first and second molars. The biomechanical considerations for application of these mechanisms are also discussed. For repositioning of the first molar, which was severely tipped and deeply impacted, a novel cantilever mechanics was used. The molar tube was bonded in the buccolingual direction to facilitate insertion of a cantilever from the buccal side. By twisting the distal end of the cantilever, sufficient uprighting moment was generated. The mesial end of the cantilever was hooked over the miniscrew placed between the canine and first premolar, which could prevent exertion of an intrusive force to the anterior portion of the dentition as a side effect. For repositioning of the second molar, an uprighting mechanics using a compression force with two step bends incorporated into a nickel-titanium archwire was employed. This generated an uprighting moment as well as a distal force acting on the tipped second molar to regain the lost space for the first molar and bring it into its normal position. This epoch-making uprighting mechanics could also minimize the extrusion of the molar, thereby preventing occlusal interference by increasing interocclusal clearance between the inferiorly placed two step bends and the antagonist tooth. Consequently, the two step bends could help prevent occlusal interference. After 2 years and 11 months of active treatment, a desirable Class I occlusion was successfully achieved without permanent tooth extraction.

Development and evaluation of a jaw-tracking system for mice: reconstruction of three-dimensional movement trajectories on an arbitrary point on the mandible.

BACKGROUND: Mastication is one of the most fundamental functions for the conservation of life. The demand for devices for evaluating stomatognathic function, for instance, recording mandibular movements or masticatory muscle activities using animal models, has been increasing in recent years to elucidate neuromuscular control mechanisms of mastication and to investigate the etiology of oral motor disorders. To identify the fundamental characteristics of the jaw movements of mice, we developed a new device that reconstructs the three-dimensional (3D) movement trajectories on an arbitrary point on the mandible during mastication. METHODS: First, jaw movements with six degrees of freedom were measured using a motion capture system comprising two high-speed cameras and four reflective markers. Second, a 3D model of the mandible including the markers was created from micro-computed tomography images. Then, the jaw movement trajectory on the certain anatomical point was reproduced by integrating the kinematic data of the jaw movements with the geometric data of the mandible. RESULTS: The 3D movements at any points on the mandible, such as the condyle, molar, and incisor during mastication, could be calculated and visualized with an accuracy > 0.041 mm in 3D space. The masticatory cycle was found to be clearly divided into three phases, namely, the opening, closing, and occlusal phases in mice. CONCLUSIONS: The proposed system can reproduce and visualize the movements of internal anatomical points such as condylar points precisely by combining kinematic data with geometric data. The findings obtained from this system could facilitate our understanding of the pathogenesis of eating disorders or other oral motor disorders when we could compare the parameters of stomatognathic function of normal mice and those of genetically modified mice with oral behavioral dysfunctions.

Numeric simulation model for long-term orthodontic tooth movement with contact boundary conditions using the finite element method.

INTRODUCTION: Although many attempts have been made to simulate orthodontic tooth movement using the finite element method, most were limited to analyses of the initial displacement in the periodontal ligament and were insufficient to evaluate the effect of orthodontic appliances on long-term tooth movement. Numeric simulation of long-term tooth movement was performed in some studies; however, neither the play between the brackets and archwire nor the interproximal contact forces were considered. The objectives of this study were to simulate long-term orthodontic tooth movement with the edgewise appliance by incorporating those contact conditions into the finite element model and to determine the force system when the space is closed with sliding mechanics. METHODS: We constructed a 3-dimensional model of maxillary dentition with 0.022-in brackets and 0.019 × 0.025-in archwire. Forces of 100 cN simulating sliding mechanics were applied. The simulation was accomplished on the assumption that bone remodeling correlates with the initial tooth displacement. RESULTS: This method could successfully represent the changes in the moment-to-force ratio: the tooth movement pattern during space closure. CONCLUSIONS: We developed a novel method that could simulate the long-term orthodontic tooth movement and accurately determine the force system in the course of time by incorporating contact boundary conditions into finite element analysis. It was also suggested that friction is progressively increased during space closure in sliding mechanics.

Innovative design of closing loops producing an optimal force system applicable in the 0.022-in bracket slot system.

INTRODUCTION: Most closing loops designed for producing higher moment-to-force (M/F) ratios require complex wire bending and are likely to cause hygiene problems and discomfort because of their complicated configurations. We aimed to develop a simple loop design that can produce optimal force and M/F ratio. METHODS: A loop design that can generate a high M/F ratio and the ideal force level was investigated by varying the portion and length of the cross-sectional reduction of a teardrop loop and the loop position. The forces and moments acting on closing loops were calculated using structural analysis based on the tangent stiffness method. RESULTS: An M/F ratio of 9.3 (high enough to achieve controlled movement of the anterior teeth) and an optimal force level of approximately 250 g of force can be generated by activation of a 10-mm-high teardrop loop whose cross-section of 0.019 × 0.025 or 0.021 × 0.025 in was reduced in thickness by 50% for a distance of 3 mm from the apex, located between a quarter and a third of the interbracket distance from the canine bracket. CONCLUSIONS: The simple loop design that we developed delivers an optimal force and an M/F ratio for the retraction of anterior teeth, and is applicable in a 0.022-in slot system.

Skeletal stability after sagittal split ramus osteotomy with physiological positioning in patients with skeletal mandibular prognathism and facial asymmetry.

The correction of deformities of the jaw in patients with facial asymmetry is challenging because of the high rate of relapse, which may partly be caused by skeletal interference and inappropriate seating of the condylar head. We evaluated outcomes in 30 patients who were treated by short lingual osteotomy with physiological positioning. Nine had facial symmetry (absolute displacement of the menton<2mm), 14 had minor asymmetry (displacement of >2 to <4mm), and 7 severe asymmetry (displacement of >4mm). The postoperative position of the menton (Me) was stable in each group, but deviated by 3.56mm in those with severe asymmetry. This deviation remained immediately after operation and after more than one year in this group, which implies that the Me was not on the mandibular midline. The lateral swing of both sides of the proximal segment did not change immediately after operation in any group. Although short lingual osteotomy with physiological positioning can result in skeletal stability, it is important to assess the association between the dental arch and the mandible using computed tomography to ensure a good outcome in patients with a skeletal class III deformity and facial asymmetry.

Biomechanical aspects of segmented arch mechanics combined with power arm for controlled anterior tooth movement: A three-dimensional finite element study.

The porpose of this study was to determine the optimal length of power arms for achieving controlled anterior tooth movement in segmented arch mechanics combined with power arm. A three-dimensional finite element method was applied for the simulation of en masse anterior tooth retraction in segmented power arm mechanics. The type of tooth movement, namely, the location of center of rotation of the maxillary central incisor in association with power arm length, was calculated after the retraction force was applied. When a 0.017 × 0.022-in archwire was inserted into the 0.018-in slot bracket, bodily movement was obtained at 9.1 mm length of power arm, namely, at the level of 1.8 mm above the center of resistance. In case a 0.018 × 0.025-in full-size archwire was used, bodily movement of the tooth was produced at the power arm length of 7.0 mm, namely, at the level of 0.3 mm below the center of resistance. Segmented arch mechanics required shorter length of power arms for achieving any type of controlled anterior tooth movement as compared to sliding mechanics. Therefore, this space closing mechanics could be widely applied even for the patients whose gingivobuccal fold is shallow. The segmented arch mechanics combined with power arm could provide higher amount of moment-to-force ratio sufficient for controlled anterior tooth movement without generating friction, and vertical forces when applying retraction force parallel to the occlusal plane. It is, therefore, considered that the segmented power arm mechanics has a simple appliance design and allows more efficient and controllable tooth movement.

Effect of bracket slot and archwire dimensions on anterior tooth movement during space closure in sliding mechanics: a 3-dimensional finite element study.

INTRODUCTION: It has been found that controlled movement of the anterior teeth can be obtained by attaching a certain length of power arm onto an archwire in sliding mechanics. However, the impact of the archwire/bracket play on anterior tooth movement has not been clarified. The purpose of this study was to compare the effect of the power arm on anterior tooth movements with different dimensions of bracket slots and archwires. METHODS: A 3-dimensional finite element method was used to simulate en-masse anterior tooth retraction in sliding mechanics. Displacements of the maxillary central incisor and the archwire deformation were calculated when applying retraction forces from different lengths of power arms. RESULTS: When a 0.017 × 0.022-in archwire was engaged into the 0.018-in slot bracket, bodily movement of the incisor was obtained with 9.1-mm length of the power arm. When a 0.022-in slot system was coupled with a 0.019 × 0.025-in archwire, bodily movement was observed with a power arm length of 11.6 mm. CONCLUSIONS: Archwire/bracket play has a remarkable impact on anterior tooth movement. An effective torque application to the anterior teeth becomes clinically difficult in sliding mechanics combined with power arms when the archwire/bracket play is large.

Influence of malocclusion on the development of masticatory function and mandibular growth.

OBJECTIVE: To verify the hypothesis that appropriate acquisition of masticatory function and normal growth of the mandible are modified by malocclusion. MATERIALS AND METHODS: Eighteen Jcl:ICR mice were divided into two groups. In one group we shifted the mandible laterally using an occlusal guidance appliance, creating a posterior crossbite at 5 weeks of age. The other group served as control. After 10 weeks, three-dimensional jaw movements and muscle activities were recorded simultaneously during mastication. Microcomputed tomography scans were obtained in vivo to evaluate morphometric changes in the mandible. RESULTS: (1) The jaw movement pattern in the sagittal plane showed significantly less anteroposterior excursion in the malocclusion group during the late-closing phase (power phase). (2) Electromyography showed significantly less masseter activity in the malocclusion group. (3) The condylar width and mandibular bone mineral density (BMD) were significantly reduced in the malocclusion mice compared to the normal mice. CONCLUSIONS: These findings suggest that optimization of the chewing pattern and acquisition of appropriate masticatory function is impeded by malocclusion. Altered mechanical loading to the mandible may cause significant reduction of condylar width and mandibular BMD.

Maxillary canine substitution for the severely resorbed root of central incisor: 12-year follow-up.

Ectopically erupting maxillary canines can cause problems that necessitate surgical, orthodontic, and restorative treatment. When a canine eruption disturbance causes resorption and requires subsequent extraction of the affected teeth, the resulting spaces are candidates for orthodontic repositioning and recontouring of the remaining teeth. To achieve successful results, the clinician must have a proper knowledge of tooth anatomy, root angulation, gingival margin position, restorative techniques, and occlusion. A collaborative effort from the pediatric dentist, orthodontist, and surgeon is required to produce an esthetic and functional result. This case report describes the substitution of maxillary canines for both the left central and right lateral incisors and substitution of the maxillary right lateral incisor for the maxillary right central incisor.

Effect of play between bracket and archwire on anterior tooth movement in sliding mechanics: A three-dimensional finite element study.

OBJECTIVES: The aim of this study was to clarify the effect of the play between the bracket and the archwire on anterior tooth movement subjected to the retraction force from various lengths of power arms in sliding mechanics. MATERIALS AND METHODS: A three-dimensional finite element method was used to simulate en masse anterior tooth retraction in sliding mechanics. The displacements of the maxillary incisor and the archwire deformation were calculated when the retraction force was applied. RESULTS: When a play did not exist, bodily movement was obtained at 5.0 mm length of power arm. In case a play existed, bodily movement was observed at the power arm length of 11.0 mm. CONCLUSIONS: In the actual clinical situation, a bracket/archwire play and the torsion of the archwire within the bracket slot should be taken into consideration to prescribe an optimal power arm length and to achieve effective anterior tooth movement.

Optimal loading conditions for controlled movement of anterior teeth in sliding mechanics.

OBJECTIVE: To determine optimal loading conditions such as height of retraction force on the power arm and its position on the archwire in sliding mechanics. MATERIALS AND METHODS: A 3D finite element method (FEM) was used to simulate en masse anterior teeth retraction in sliding mechanics. The degree of labiolingual tipping of the maxillary central incisor was calculated when the retraction force was applied to different heights of a power arm set mesial or distal to the canine. RESULTS: When the power arm was placed mesial to the canine, at the level of 0 mm (bracket slot level), uncontrolled lingual crown tipping of the incisor was observed and the anterior segment of the archwire was deformed downward. At a power arm height of 5.5 mm, bodily movement was produced and the archwire was less deformed. When the power arm height exceeded 5.5 mm, the anterior segment of the archwire was raised upward and lingual root tipping occurred. When the power arm was placed distal to the canine, lingual crown tipping was observed up to a level of 11.2 mm. CONCLUSIONS: Placement of the power arm of an archwire between the lateral incisor and canine enables orthodontists to maintain better control of the anterior teeth in sliding mechanics. Both the biomechanical principles associated with the tooth's center of resistance and the deformation of the archwire should be taken into consideration for predicting and planning orthodontic tooth movement.

Effect of celecoxib on emotional stress and pain-related behaviors evoked by experimental tooth movement in the rat.

OBJECTIVE: To test the efficacy of an animal model of pain and stress and evaluate the effects of celecoxib administered when orthodontic force is applied. MATERIALS AND METHODS: A 20-g reciprocal force was applied via an orthodontic appliance to the maxillary left first and second molars of 7-week-old male Sprague-Dawley rats. Rat behavior was evaluated at 5, 24, and 48 hours after the appliance was set. Behavior was assessed in a test field by the number of lines crossed in the first 30 seconds and 5 minutes following force application; number of lines crossed to the center; rearing time; and facial grooming time. Experimental group 1 received intraperitoneal administration of 30 mg/kg celecoxib before every behavioral test. Experimental group 2 received 90 mg/kg before the first behavioral test, and physiologic saline was administered before the remaining behavioral tests. Control groups received saline before every behavioral test and were given passive (passive control group) and active (active control group) appliances, respectively. RESULTS: Parameters related to pain increased in the active controls, whereas the parameters in the experimental groups decreased to the level seen in the passive controls. Statistically significant differences in pain-related behavior between control and experimental groups were found at 5 and 24 hours after placing the appliance. Stress-related behavior was significantly less in the experimental groups compared to the active control group during experimental periods. CONCLUSIONS: The administration of celecoxib relieves pain- and stress-related behavior evoked by orthodontic tooth movement in the rat. This model might be a useful tool for the evaluation of pain and stress.

A novel method for the assessment of three-dimensional tooth movement during orthodontic treatment.

OBJECTIVE: To (1) evaluate the stability of palatal rugae as landmarks for superimposition of dental casts and (2) establish a three-dimensional superimposition method of maxillary dental casts for analyzing orthodontic tooth movement. MATERIALS AND METHODS: The sample consisted of dental casts obtained from 10 patients treated with extraction of bilateral maxillary first premolars and placement of three palatal miniscrews as anchorage for retraction of the anterior teeth. Dental casts were measured by means of laser surface scanning system, and three-dimensional images were reconstructed. Serial dental casts were superimposed on the three miniscrews as registration landmarks (miniscrew-superimposition method), and the displacement of each palatal ruga point during the closure of extraction spaces was measured. Displacement of the central incisors was measured by the miniscrew-superimposition method and the proposed superimposition technique (ruga-palate-superimposition method). Correlation analysis and paired t-tests were performed to determine whether a significant difference existed between the measurements of the two superimposition methods. RESULTS: The medial points of the third palatal rugae and the shape of the palatal vault were stable throughout the treatment. The displacement of the central incisors measured using the ruga-palate-superimposition method showed no significant difference with that measured using the miniscrew-superimposition method. CONCLUSION: The maxillary dental casts can be reliably superimposed on the medial points of the third palatal rugae and the palatal vault as reference landmarks.

Experimental determination of optimal force system required for control of anterior tooth movement in sliding mechanics.

INTRODUCTION: This study was designed to determine the optimum vertical height of the retraction force on the power arm that is required for efficient anterior tooth retraction during space closure with sliding mechanics. METHODS: Three adults (1 man, 2 women) with Angle Class II Division 1 malocclusions were selected for this study. In each subject, the maxillary right central incisor was the target tooth. Initial tooth displacements of that tooth with sliding mechanics with various heights of retraction forces were measured in vivo by a 2-point 3-dimensional displacement magnetic sensor device. The tooth's motion trajectories on the midsagittal plane were studied. RESULTS: The location of the center of rotation of the target tooth varied according to the different heights of the retraction forces. Controlled anterior tooth movement (ie, lingual-crown tipping, lingual-root movement) can be predicted, simulated, or even manipulated by different heights of retraction forces on the power arm in the sliding mechanics force system. A power arm length of 3 to 5 mm is estimated to produce controlled lingual-crown tipping (with the apex as the center of rotation) for efficient anterior tooth retraction during sliding space closure in adults with Angle Class II Division 1 malocclusion. CONCLUSIONS: Knowing and applying the correct height of retraction force on the power arm is the key to efficient anterior tooth retraction.