The purpose of this study is to investigate the spacial expression pattern and functional significance of three key transcription factors related to bone and cartilage formation, namely, Sox9, Runx2, and Osterix in cartilages during the late development of mouse mandible. Immunohistochemical examinations of Sox9, Runx2, and Osterix were conducted in the mandibular cartilages of the 15 neonatal C57BL/6N mice. In secondary cartilages, both Sox9 and Runx2 were weakly expressed in the polymorphic cell zone, strongly expressed in the flattened cell zone and throughout the entire hypertrophic cell zone. Similarly, both transcriptional factors were weakly expressed in the uncalcified Meckel's cartilage while strongly expressed in the rostral cartilage. Meanwhile, Osterix was at an extremely low level in cells of the flattened cell zone and the upper hypertrophic cell zone in secondary cartilages. Surprisingly, Osterix was intensely expressed in hypertrophic chondrocytes in the center of the uncalcified Meckel's cartilage while moderately expressed in part of hypertrophic chondrocytes in the rostral process. Consequently, it is suggested that Sox9 is a main and unique positive regulator in the hypertrophic differentiation process of mandibular secondary cartilages, in addition to Runx2. Furthermore, Osterix is likely responsible for phenotypic conversion of Meckel's chondrocytes during its degeneration.

BackgroundRetinal ganglion cells are often classified into different subtypes according to their morphology or physiological functions. The axons of RGCs contain three major cytoskeletal components: actin filaments (F-actin); microtubules; and neurofilaments (NFs). The contents of these components vary among axons. Our objective was to classify axons into subtypes based on the contents of cytoskeletal components and study their distributions across the retina in normal rodent retinas.MethodsWhole-mounted retinas of female Wistar rats were stained with phalloidin to label F-actin, anti-β-tubulin monoclonal antibody to mark microtubules, and antineurofilament antibody to label NFs. A confocal laser scanning microscope was used to provide en face images of retinal nerve fiber bundles with a resolution of 0.24 μm/pixel. Staining intensity profiles across axons were obtained for each cytoskeletal component. Axonal subtypes were then determined from the relative contents, indicated by the staining intensity, of these components. Linear density was used to investigate topographical distribution of each subtype across the retina.ResultsNormal axons could be classified into seven subtypes – FMN, FM, FN, and MN subtypes, (in which at least two or three cytoskeletal components were intensely stained), and F, M, and N subtypes, (in which only one cytoskeletal component was intensely stained within an axon). The FMN subtype was the most abundant subtype. There was no preferential distribution of subtypes around the optic nerve head. However, the densities of the axonal subtypes that contained NFs were found significantly different in the central and peripheral retinal regions. Axonal sizes were subtype-dependent.ConclusionAxons of retinal ganglion cells can be classified into different subtypes, based on the contents of axonal cytoskeletal components. The classified subtypes will provide a new means to study selective damage of axonal ultrastructures in ocular neuropathic diseases.