The authors would like to thank Dr. David Rowe for kindly providing the transgenic mice and Li Chen for the histological assistance. 
The research reported in this publication was supported by the National Institute of Dental &amp; Craniofacial Research of the National Institutes of Health under Award Number K08DE025914 and by the American Association of Orthodontic Foundation to Sumit Yadav.

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	<li>LeResche, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epidemiology+of+Temporomandibular+Disorders:+Implications+for+the+Investigation+of+Etiologic+Factors.">Epidemiology of Temporomandibular Disorders: Implications for the Investigation of Etiologic Factors.</a> Crit Rev Oral Biol Med. 8 (3), 291-305 (1997).</li><li>Chen, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Altered+functional+loading+causes+differential+effects+in+the+subchondral+bone+and+condylar+cartilage+in+the+temporomandibular+joint+from+young+mice.">Altered functional loading causes differential effects in the subchondral bone and condylar cartilage in the temporomandibular joint from young mice.</a> Osteoarthr Cartil. 17 (3), 354-361 (2009).</li><li>Pirttiniemi, P., Kantomaa, T., Sorsa, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+decreased+loading+on+the+metabolic+activity+of+the+mandibular+condylar+cartilage+in+the+rat.">Effect of decreased loading on the metabolic activity of the mandibular condylar cartilage in the rat.</a> Eur J Orthod. 26 (1), 1-5 (2004).</li><li>Chavan, S. J., Bhad, W. A., Doshi, U. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+temporomandibular+joint+changes+in+Twin+Block+and+Bionator+appliance+therapy:+a+magnetic+resonance+imaging+study.">Comparison of temporomandibular joint changes in Twin Block and Bionator appliance therapy: a magnetic resonance imaging study.</a> Prog Orthod. 15 (57), (2014).</li><li>Dutra, E. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cellular+and+Matrix+Response+of+the+Mandibular+Condylar+Cartilage+to+Botulinum+Toxin.">Cellular and Matrix Response of the Mandibular Condylar Cartilage to Botulinum Toxin.</a> PLoS ONE. 11 (10), 0164599 (2016).</li><li>Benjamin, M., Ralphs, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biology+of+fibrocartilage+cells.">Biology of fibrocartilage cells.</a> Int Rev Cytol. 233, 1-45 (2004).</li><li>Shen, G., Darendeliler, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Adaptive+Remodeling+of+Condylar+Cartilage-+A+Transition+from+Chondrogenesis+to+Osteogenesis.">The Adaptive Remodeling of Condylar Cartilage- A Transition from Chondrogenesis to Osteogenesis.</a> J Dent Res. 84 (8), 691-699 (2005).</li><li>Utreja, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cell+and+matrix+response+of+temporomandibular+cartilage+to+mechanical+loading.">Cell and matrix response of temporomandibular cartilage to mechanical loading.</a> Osteoarthr Cartil. 24 (2), 335-344 (2016).</li><li>Kaul, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Effect+of+Altered+Loading+on+Mandibular+Condylar+Cartilage.">The Effect of Altered Loading on Mandibular Condylar Cartilage.</a> PLoS ONE. 11 (7), 0160121 (2016).</li><li>Dyment, N. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-Throughput,+Multi-Image+Cryohistology+of+Mineralized+Tissues.">High-Throughput, Multi-Image Cryohistology of Mineralized Tissues.</a> J Vis Exp. , e54468 (2016).</li><li>Kawamoto, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Use+of+a+new+adhesive+film+for+the+preparation+of+multi-purpose+fresh-frozen+sections+from+hard+tissues,+whole-animals,+insects+and+plants.">Use of a new adhesive film for the preparation of multi-purpose fresh-frozen sections from hard tissues, whole-animals, insects and plants.</a> Arch Histol Cytol. 66 (2), 123-143 (2003).</li><li>Hayman, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tartrate-resistant+acid+phosphatase+(TRAP)+and+the+osteoclast/immune+cell+dichotomy.">Tartrate-resistant acid phosphatase (TRAP) and the osteoclast/immune cell dichotomy.</a> Autoimmunity. 41 (3), 218-223 (2008).</li></ol>

The authors have no competing financial interests.

This manuscript described methods for the morphometric measurement and cellular analysis of murine mandibular condyles and mandibles. The radiographic morphometric measurements can also be used to analyze other bones from small experimental animals. In addition, the cellular analysis (cell quantification and cartilage distance mapping) are not limited to the rodent mandibular condyle, but can be used to quantify histological sections of numerous tissues. 
Transgenic mouse models expressing flu...

The TMJ is a unique load-bearing joint located in the craniofacial region and is formed of fibrocartilage. The MCC of the TMJ is essential for joint function, including unhindered jaw movement while speaking and masticating, but it is commonly affected by degenerative diseases, including osteoarthritis1. The TMJ has the capacity to adapt to external stimuli and loading alterations, leading to structural and cellular changes to the components of the MCC2,3,4,5. The load-bearing properties of the MCC can be explained by the interactions between its constituents, including water, the collagen network, and densely packed proteoglycans. The MCC has four distinct cellular zones that express different types of collagen and non-collagen proteins: 1) the superficial or articular zone; 2) the proliferative zone, composed of undifferentiated mesenchymal cells and that responds to loading demands; 3) the prehypertrophic zone, composed of mature chondrocytes expressing collagen type 2; and 4) the hypertrophic zone, the region where the hypertrophic chondrocytes expressing collagen type 10 die and undergo calcification. The non-mineralized region is rich in proteoglycans which provide resistance to compressive forces6.
There is continuous mineralization at the hypertrophic zone of the MCC, where the transition from chondrogenesis to osteogenesis occurs, guaranteeing the robust mineral structure of the subchondral bone of the mandibular condyle7. Cellular changes in the unmineralized and mineralized regions ultimately lead to morphological and structural changes in the mandibular condyle and mandible. Maintenance of the homeostasis of all cellular regions of the MCC and the mineralization of the subchondral portion are essential to the health, load-bearing capacity, and integrity of the TMJ.
The multiple collagen transgenic mouse model (as described by Utreja et al.)8 is a great tool to use to understand changes in collagen expression because all transgenes are expressed in the MCC. For an in-depth histological evaluation, histological stains are used to study matrix deposition, mineralization, cell proliferation, and apoptosis, as well as protein expression at the different cell layers of the MCC.
In this manuscript, histological and morphometric analyses are used to evaluate cellular and structural changes in the MCC and subchondral bone of the mandibular condyle of mice. In addition, a cell quantification method, for analyzing fluorescent histological images and for mapping light microscope slides, is described. The compressive static TMJ loading method, which causes cellular and morphological changes at the MCC and subchondral bone9, is also illustrated to validate our methods.
The methods described here can be used to determine morphometric and histological changes in the mandibular condyle and mandible of rodents or to analyze other regions of endochondral ossification and the morphology of additional mineralized tissues.

<table><tbody><tr><td>MX20 Radiography System</td><td>Faxitron X-Ray LLC&amp;nbsp;</td><td></td><td></td></tr><tr><td>Digimizer Image software&amp;nbsp;</td><td>MedCalc Software</td><td></td><td></td></tr><tr><td>Shandon Cryomatrix embedding resin</td><td>Thermo Scientific</td><td>6769006</td><td></td></tr><tr><td>Manual microscope Axio Imager Z1</td><td>Carl Zeiss</td><td>208562</td><td></td></tr><tr><td>yellow fluorescent protein filter&amp;nbsp; - EYFP</td><td>Chroma Technology Corp</td><td>49003</td><td></td></tr><tr><td>cyan fluorescent protein filter - ECFP</td><td>Chroma Technology Corp</td><td>49001</td><td></td></tr><tr><td>red fluoresecent protein filter - Cy5</td><td>Chroma Technology Corp</td><td>49009</td><td></td></tr><tr><td>sodium acetate anhydrous</td><td>Sigma-Aldrich</td><td>S2889</td><td></td></tr><tr><td>sodium L-tartrate dibasic dihydrate&amp;nbsp;</td><td>Sigma-Aldrich</td><td>228729</td><td></td></tr><tr><td>sodium nitrite&amp;nbsp;</td><td>Sigma-Aldrich</td><td>237213</td><td></td></tr><tr><td>ELF97 substrate</td><td>Thermo Fisher Scientific</td><td>E6600</td><td></td></tr><tr><td>ClickiT EdU Alexa Fluor 594 HCS kit</td><td>Life Technologies</td><td>C10339&amp;nbsp;</td><td>includes EdU (5-ethynyl-2&#039;-deoxyuridine)&amp;nbsp;</td></tr><tr><td>DAPI (4&#039;,6-Diamidino-2-Phenylindole, Dihydrochloride)</td><td>Thermo Scientific</td><td>D1306</td><td></td></tr><tr><td>Sodium phosphate dibasic&amp;nbsp;</td><td>Sigma-Aldrich</td><td>S3264</td><td></td></tr><tr><td>Sodium phosphate monobasic&amp;nbsp;</td><td>Sigma-Aldrich</td><td>71505</td><td></td></tr><tr><td>Toluidine Blue O&amp;nbsp;</td><td>Sigma-Aldrich</td><td>T3260</td><td></td></tr><tr><td>Adobe Photoshop&amp;nbsp;</td><td>Adobe Systems Incorporated</td><td></td><td></td></tr><tr><td>Phosphate buffered saline tablets (PBS)</td><td>Research Products International</td><td>P32080-100T</td><td></td></tr><tr><td>CNA Beta III Nickel-Free Archwire</td><td>Ortho Organizers, Inc.</td><td></td><td></td></tr><tr><td>GraphPad Prism&amp;nbsp;</td><td>GraphPad Software, Inc.</td><td></td><td></td></tr></tbody></table>

a morphometric and cellular analysis method for the murine mandibular condyle

The temporomandibular joint (TMJ) has the capacity to adapt to external stimuli, and loading changes can affect the position of condyles, as well as the structural and cellular components of the mandibular condylar cartilage (MCC). This manuscript describes methods for analyzing these changes and a method for altering the loading of the TMJ in mice (i.e., compressive static TMJ loading). The structural evaluation illustrated here is a simple morphometric approach that uses the Digimizer software and is performed in radiographs of small bones. In addition, the analysis of cellular changes leading to alterations in collagen expression, bone remodeling, cell division, and proteoglycan distribution in the MCC is described. The quantification of these changes in histological sections - by counting the positive fluorescent pixels using image software and measuring the distance mapping and stained area with Digimizer - is also demonstrated. The methods shown here are not limited to the murine TMJ, but could be used on additional bones of small experimental animals and in other regions of endochondral ossification.

Descriptive statistics were performed to examine the distribution of morphometric measurements (mandibular length, condylar length, condylar width) and histological analyses. Outcomes were compared between the loaded group (i.e., mice subjected to compressive loading with the beta titanium spring) and the control group (i.e., matching control mice that did not receive any procedure). Statistically significant differences between means were determined by unpaired t-test, ...

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A Morphometric and Cellular Analysis Method for the Murine Mandibular Condyle

This manuscript presents methods for analyzing morphometric and cellular changes within the mandibular condyle of rodents.

The temporomandibular joint (TMJ) has the capacity to adapt to external stimuli, and loading changes can affect the position of condyles, as well as the ...

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8.2K Views.  University of Connecticut Health Center. This manuscript presents methods for analyzing morphometric and cellular changes within the mandibular condyle of rodents.

Video: A Morphometric and Cellular Analysis Method for the Murine Mandibular Condyle

Visualization of Chondrocyte Intercalation and Directional Proliferation via Zebrabow Clonal Cell Analysis in the Embryonic Meckel&#8217;s Cartilage

Development of the vertebrate craniofacial structures requires precise coordination of cell migration, proliferation, adhesion and differentiation. Patterning of the Meckel&#39;s cartilage, a first pharyngeal arch derivative, involves the migration of cranial neural crest (CNC) cells and the progressive partitioning, proliferation and organization of differentiated chondrocytes. Several studies have described CNC migration during lower jaw&#160;morphogenesis, but the details of how the chondrocytes achieve organization in the growth and extension of Meckel&#8217;s cartilage remains unclear. The sox10 restricted and chemically induced Cre recombinase-mediated recombination generates permutations of distinct fluorescent proteins (RFP, YFP and CFP), thereby creating a multi-spectral labeling of progenitor cells and their progeny, reflecting distinct clonal populations. Using confocal time-lapse photography, it is possible to observe the chondrocytes behavior during the development of the zebrafish Meckel&#8217;s cartilage.
Multispectral cell labeling enables scientists to demonstrate extension of the Meckel&#8217;s chondrocytes.&#160;During extension phase of the Meckel&#8217;s cartilage, which prefigures the mandible, chondrocytes intercalate to effect extension as they stack in an organized single-cell layered row. Failure of this organized intercalating process to mediate cell extension provides the cellular mechanistic explanation for hypoplastic mandible that we observe in mandibular malformations.

Cell organization of craniofacial bones has long been hypothesized but never directly visualized. Multi-spectral cell labeling and in vivo live imaging allows visualization of dynamic cell behavior in zebrafish lower jaw. Here, we detail the protocol to manipulate Zebrabow transgenic fish and directly observe cell intercalation and morphological changes of chondrocytes in the Meckel&#8217;s cartilage.

Development of the vertebrate craniofacial structures requires precise coordination of cell migration, proliferation, adhesion and differentiation. ...

Developmental Biology

Laser Capture Microdissection of Mouse Embryonic Cartilage and Bone for Gene Expression Analysis

Laser capture microdissection (LCM) is a powerful tool to isolate specific cell types or regions of interest from heterogeneous tissues. The cellular and molecular complexity of skeletal elements increases with development. Tissue heterogeneity, such as at the interface of cartilaginous and osseous elements with each other or with surrounding tissues, is one obstacle to the study of developing cartilage and bone. Our protocol provides a rapid method of tissue processing and isolation of cartilage and bone that yields high quality RNA for gene expression analysis. Fresh frozen tissues of mouse embryos are sectioned and brief cresyl violet staining is used to visualize cartilage and bone with colors distinct from surrounding tissues. Slides are then rapidly dehydrated, and cartilage and bone are isolated subsequently by LCM. The minimization of exposure to aqueous solutions during this process maintains RNA integrity. Mouse Meckel&rsquo;s cartilage and mandibular bone at E16.5 were successfully collected and gene expression analysis showed differential expression of marker genes for osteoblasts, osteocytes, osteoclasts, and chondrocytes. High quality RNA was also isolated from a range of tissues and embryonic ages. This protocol details sample preparation for LCM including cryoembedding, sectioning, staining and dehydrating fresh frozen tissues, and precise isolation of cartilage and bone by LCM resulting in high quality RNA for transcriptomic analysis.

This protocol describes laser capture microdissection for the isolation of cartilage and bone from fresh frozen sections of the mouse embryo. Cartilage and bone can be rapidly visualized by cresyl violet staining and collected precisely to yield high quality RNA for transcriptomic analysis.

Laser capture microdissection (LCM) is a powerful tool to isolate specific cell types or regions of interest from heterogeneous tissues. The cellular and ...

Dissection, MicroCT Scanning and Morphometric Analyses of the Baculum

Modern morphometrics provides powerful methods to quantify size and shape variation. A basic requirement is a list of coordinates that define landmarks; however such coordinates must represent homologous structures across specimens. While many biological objects consist of easily identified landmarks to satisfy the assumption of homology, many lack such structures. One potential solution is to mathematically place semi-landmarks on an object that represent the same morphological region across specimens. Here, we illustrate a recently developed pipeline to mathematically define semi-landmarks from the mouse baculum (penis bone). Our methods should be applicable to a wide range of objects.

Many biological structures lack easily definable landmarks, making it difficult to apply modern morphometric methods. Here we illustrate methods to study the mouse baculum (a bone in the penis), including dissection and microCT scanning, followed by computational methods to define semi-landmarks that are used to quantify size and shape variation.

Modern morphometrics provides powerful methods to quantify size and shape variation. A basic requirement is a list of coordinates that define landmarks; ...

Bioengineering

Analysis of Craniomaxillofacial Malformations in Mice Using Three-dimensional Microcomputed Tomography

To model craniofacial malformations caused by vitamin A deficiency (VAD), we expressed a dominant-negative retinoid receptor mutation in osteoblasts to specifically inhibit RAR transcriptional activity in mice. This approach allowed us to investigate the effects of VAD on cranial hypomineralization, mandibular deformity, and clavicular hypoplasia in clinical cases. In this study, microcomputed tomography (microCT) scanning of the craniomaxillofacial region of mice represented a valuable tool for studying the growth and development of this animal model. The manual estimation of images is both time-consuming and inaccurate. Hence, here, we present a straightforward, efficient, and accurate approach for segmenting and quantifying the microCT images of each craniomaxillofacial bone. MicroCT software was used to slice the mandible, frontal bone, parietal bone, nasal bone, premaxilla, maxilla, interparietal bone, and occipital bone of mice and measure their corresponding lengths and widths. This segmentation method can be applied to study growth and development in developmental biology, biomedicine, and other related sciences and allows researchers to analyze the effects of genetic mutations on individual craniofacial bones.

Here, we describe in depth a method, which is based on microcomputed tomography, to segment and measure 3D models of craniomaxillofacial bones in mice, for better assessment of craniomaxillofacial bone development in mice than what is possible with current methods.

To model craniofacial malformations caused by vitamin A deficiency (VAD), we expressed a dominant-negative retinoid receptor mutation in osteoblasts to ...

Cantilever Bending of Murine Femoral Necks

Fractures in the femoral neck are a common occurrence in individuals with osteoporosis. Many mouse models have been developed to assess disease states and therapies, with biomechanical testing as a primary outcome measure. However, traditional biomechanical testing focuses on torsion or bending tests applied to the midshaft of the long bones. This is not typically the site of high-risk fractures in osteoporotic individuals. Therefore, a biomechanical testing protocol was developed that tests the femoral necks of murine femurs in cantilever bending loading to replicate better the types of fractures experienced by osteoporosis patients. Since the biomechanical outcomes are highly dependent on the flexural loading direction relative to the femoral neck, 3D printed guides were created to maintain a femoral shaft at an angle of 20&#176; relative to the loading direction. The new protocol streamlined the testing by reducing variability in alignment (21.6&#176; &plusmn; 1.5&#176;, COV = 7.1%, n = 20) and improved reproducibility in the measured biomechanical outcomes (average COV = 26.7%). The new approach using the 3D printed guides for reliable specimen alignment improves rigor and reproducibility by reducing the measurement errors due to specimen misalignment, which should minimize sample sizes in mouse studies of osteoporosis.

The present protocol describes the development of a reproducible testing platform for murine femoral necks in a cantilever bending set-up. Custom 3D printed guides were used to consistently and rigidly fix the femurs in optimal alignment.

Fractures in the femoral neck are a common occurrence in individuals with osteoporosis. Many mouse models have been developed to assess disease states and ...

Standardized Histomorphometric Evaluation of Osteoarthritis in a Surgical Mouse Model

One of the most prevalent joint disorders in the United States, osteoarthritis (OA) is characterized by progressive degeneration of articular cartilage, primarily in the hip and knee joints, which results in significant impacts on patient mobility and quality of life. To date, there are no existing curative therapies for OA able to slow down or inhibit cartilage degeneration. Presently, there is an extensive body of ongoing research to understand OA pathology and discover novel therapeutic approaches or agents that can efficiently slow down, stop, or even reverse OA. Thus, it is crucial to have a quantitative and reproducible approach to accurately evaluate OA-associated pathological changes in the joint cartilage, synovium, and subchondral bone. Currently, OA severity and progression are primarily assessed using the Osteoarthritis Research Society International (OARSI) or Mankin scoring systems. In spite of the importance of these scoring systems, they are semiquantitative and can be influenced by user subjectivity. More importantly, they fail to accurately evaluate subtle, yet important, changes in the cartilage during the early disease states or early treatment phases. The protocol we describe here uses a computerized and semiautomated histomorphometric software system to establish a standardized, rigorous, and reproducible quantitative methodology for the evaluation of joint changes in OA. This protocol presents a powerful addition to the existing systems and allows for more efficient detection of pathological changes in the joint.

The current protocol establishes a rigorous and reproducible method for quantification of morphological joint changes that accompany osteoarthritis. Application of this protocol can be valuable in monitoring disease progression and evaluating therapeutic interventions in osteoarthritis.

One of the most prevalent joint disorders in the United States, osteoarthritis (OA) is characterized by progressive degeneration of articular cartilage, ...

Medicine

3D Imaging of PDL Collagen Fibers during Orthodontic Tooth Movement in Mandibular Murine Model

Orthodontic tooth movement is a complex biological process of altered soft and hard tissue remodeling as a result of external forces. In order to understand these complex remodeling processes, it is critical to study the tooth and periodontal tissues within their 3D context and therefore minimize any sectioning and tissue artefacts. Mouse models are often utilized in developmental and structural biology, as well as in biomechanics due to their small size, high metabolic rate, genetics and ease of handling. In principle this also makes them excellent models for dental related studies. However, a major impediment is their small tooth size, the molars in particular. This paper is aimed at providing a step by step protocol for generating orthodontic tooth movement and two methods for 3D imaging of the periodontal ligament fibrous component of a mouse mandibular molar. The first method presented is based on a micro-CT setup enabling phase enhancement&#160;imaging of fresh collagen tissues. The second method is a bone clearing method using ethyl cinnamate that enables imaging through the bone without sectioning and preserves endogenous fluorescence. Combining this clearing method with reporter mice like Flk1-Cre;TdTomato provided a first of its kind opportunity to image the 3D vasculature in the PDL and alveolar bone.

We present a protocol for generating orthodontic tooth movement in mice and methods for 3D visualization of the collagen fibers and blood vessels of periodontal ligament without sectioning.

Orthodontic tooth movement is a complex biological process of altered soft and hard tissue remodeling as a result of external forces. In order to ...

Destabilization of the Medial Meniscus and Cartilage Scratch Murine Model of Accelerated Osteoarthritis

Osteoarthritis is the most prevalent musculoskeletal disease in people over 45, leading to an increasing economic and societal cost. Animal models are used to mimic many aspects of the disease. The present protocol describes the destabilization and cartilage scratch model (DCS) of post-traumatic osteoarthritis. Based on the widely used destabilization of the medial meniscus (DMM) model, DCS introduces three scratches on the cartilage surface. The current article highlights the steps to destabilize the knee by transecting the medial meniscotibial ligament followed by three intentional superficial scratches on the articular cartilage. The possible analysis methods by dynamic weight-bearing, microcomputed tomography, and histology are also demonstrated. While the DCS model is not recommended for studies that focus on the effect of osteoarthritis on the cartilage, it enables the study of osteoarthritis development in a shorter time window, with special focus on (1) osteophyte formation, (2) osteoarthritic and injury pain, and (3) the effect of cartilage damage in the whole joint.

The present protocol describes the controlled microblade scratches on the surface of the articular cartilage after destabilizing the mouse knee by cutting the medial miniscotibial ligament. This animal model presents an accelerated form of osteoarthritis (OA) suitable for studying osteophyte formation, osteosclerosis, and early-stage pain.

Osteoarthritis is the most prevalent musculoskeletal disease in people over 45, leading to an increasing economic and societal cost. Animal models are ...

A Murine Maxillary Orthodontic Model Guide to Bone Remodeling Analysis

The Establishment of a Murine Maxillary Orthodontic Model

Due to the lack of reproducible protocols for establishing a murine maxillary orthodontic model, we present a reliable and reproducible protocol to provide researchers with a feasible tool to analyze mechanical loading-associated bone remodeling. This study presents a detailed flowchart in addition to different types of schematic diagrams, operation photos, and videos. We performed this protocol on 11 adult wide-type C57/B6J mice and harvested samples on postoperative days 3, 8, and 14. The micro-CT and histopathological data have proven the success of tooth movements coupled with bone remodeling using this protocol. Furthermore, according to the micro-CT results on days 3, 8, and 14, we have divided bone modeling into three stages: preparation stage, bone resorption stage, and bone formation stage. These stages are expected to help researchers concerned with different stages to set sample collection time reasonably. This protocol can equip researchers with a tool to carry out regenerative analysis of bone remodeling.

Author Spotlight: Insights into an Efficient Murine Maxillary Orthodontic Model Protocol

Here we demonstrate step-by-step a manageable, orthodontic tooth movement protocol operated on a murine maxillary model. With the explicit explanation of each step and visual demonstration, researchers can master this model and apply it to their experimental needs with a few modifications.

Due to the lack of reproducible protocols for establishing a murine maxillary orthodontic model, we present a reliable and reproducible protocol to ...

Investigating Orthodontic Tooth Movement and Bone Adaptation Mechanisms in Mice

Studying Orthodontic Tooth Movement in Mice

Orthodontic tooth movement (OTM) represents a dynamic process in which the alveolar bone undergoes resorption at compression sites and deposition at tension sites, orchestrated by osteoclasts and osteoblasts, respectively. This mechanism serves as a valuable model for studying various aspects of bone adaptation, including root resorption and the cellular response to mechanical force stimuli. The protocol outlined here offers a straightforward approach to investigate OTM, establishing 0.35 N as the optimal force in a mouse model employing a nickel-titanium (NiTi) coil spring. Utilizing micro-computed tomography analysis, we quantified OTM by assessing the discrepancy in the linear distance at the cement-enamel junction. The evaluation also included an analysis of orthodontic-induced inflammatory root resorption, assessing parameters such as root mineral density and the percentage of root volume per total volume. This comprehensive protocol contributes to advancing our understanding of bone remodeling processes and enhancing the ability to develop effective orthodontic treatment strategies.

Here, we present a protocol for studying orthodontic tooth movement (OTM), serving as a suitable model for investigating the mechanisms of bone adaptation, root resorption, and the response of bone cells to mechanical stimuli. This comprehensive guide provides detailed information on the OTM model, micro-computed tomography acquisition, and subsequent analysis.

Orthodontic tooth movement (OTM) represents a dynamic process in which the alveolar bone undergoes resorption at compression sites and deposition at ...