Name: a morphometric and cellular analysis method for the murine mandibular condyle
Uploaded: 2018-01-11T12:30:00
Description: Watch this Scientific Journal Video about A Morphometric and Cellular Analysis Method for the Murine Mandibular Condyle at JoVE.com

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.

The institutional animal care committee of the University of Connecticut Health Center approved all animal procedures.
1. Compressive Static TMJ Loading: Mouth Forced Open
Note: Four-week-old transgenic mice harboring fluorescent reporters for collagen (Col2a1XCol10a1), kindly provided by Dr. David Rowe (University of Connecticut), were used for the experiments described in this manuscript (n = 8; 4 males and 4 females). The Col2a1 cyan (blue) transgene is expressed i...

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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.

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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, ...

Watch this Scientific Journal Video about A Morphometric and Cellular Analysis Method for the Murine Mandibular Condyle at JoVE.com

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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