Spectroscopic Analysis of the Extracellular Matrix in Naked Mole-Rat Temporomandibular Joints

Abstract

Naked mole-rats are extremely long-living rodents with a maximum lifespan of 37 years, and their cellular aging and tissue aging are almost nonexistent. Therefore, in this study, we aim to analyze the extracellular matrix of the temporomandibular joint (TMJ) of naked mole-rats at the molecular level and explore the molecules involved in anti-aging and their localization. Micro-computed tomography (CT) scans revealed increased mineral density and wear of the mandibular condyle in aged mice. Conversely, CT scans did not reveal wear of the mandibular condyle in naked mole-rats, and histological analysis did not reveal wear of the articular disk. Using various spectroscopies and artificial intelligence (AI), we found that the articular disk of naked mole-rats is composed of a cartilage-like layer with hyaluronic acid and collagen fibers with varying orientations, which is thought to have relieved mechanical stress and have protected the mandibular condyle. These results suggest that not only the amount, but also the spatial distribution of the extracellular matrix is important for the anti-aging properties of the TMJ, and may contribute to elucidating the pathology of TMJ disorders and other degenerative conditions and developing therapeutic drugs.

Keywords: anti-aging; extracellular matrix; glycosaminoglycan; hyaluronic acid; naked mole-rat.

Figure 1

(a,c) Representative three-dimensional reconstructions and (b,d) plain radiographs of the skulls of a naked mole-rat and aged mouse.

Figure 2

Representative three-dimensional reconstructions of (a,b) the sagittal sections of the mandibular condyle, (c,d) BMD, and (e,f) trabecular or cortical bone. Yellow: spongy bone, Green: cortical bone.

Figure 3

Representative micrographs of (a,b) HE, (c,d) Masson trichrome, and (e,f) safranin O-stained sagittal sections of the TMJ. Arrow heads indicate cartilaginous tissues.

Figure 4

Representative microscopic images of HE-stained cross-sections of naked mole-rat articular disk (high magnification). Solid arrows and dotted arrows indicate cartilaginous tissues and collagen fiber, respectively. The black box indicates the fibrous layer of the mandibular condylar cartilage.

Figure 5

Representative FTIR spectra in the zones of the (a) bone and (b) articular disk for the naked mole-rat and aged mouse. The 1386 and 1740 cm⁻¹ peaks (arrows) indicate the symmetric deformation of CH₃ in glycosaminoglycans (GAGs) and the O-C=O stretching in phospholipids and GAGs, respectively.

Figure 6

(a,b) Bright-field optical micrograph and related synchrotron radiation-based Fourier transform infrared (SR-FTIR) image of (c,d) collagen type I (Col I), (e,f) apatite (HAp), (g,h) glycosaminoglycans (GAGs), and (i,j) collagen type II (Col II) structures in a microscopic portion within the TMJ.

Figure 7

Averaged Raman spectra in the zones of the (a) bone and (b) articular disk for the naked mole-rat and aged mouse. The arrow and dotted lines indicate 960, 1389, 1670, and 1740 cm⁻¹.

Figure 8

(a) Averaged Raman spectra in the finger-print region of the articular disk for the naked mole-rat and aged mouse. (b) Raman spectra in the high-frequency wavenumber interval 2700~3200 cm⁻¹ after normalization of the naked mole-rat and aged mouse. The red dashed line and the blue dashed line indicate at 1660 and 1740 cm⁻¹.

Figure 9

Bright-field optical micrograph and related Raman images of HAp, GAGs, Col I, and phospholipid structures in a microscopic portion within the TMJ for the (a) naked mole-rat and (b) aged mouse. The arrows and the dashed box indicate bone and cartilage, respectively.

Figure 10

(a,b) Representative micrographs of Masson trichrome (MT) staining in sagittal sections of the TMJ. (c,d) Spatial images obtained using cluster analysis using AI, consistent with the tissue images. (e,f) Averaged spectra of the clusters obtained in (c,d) for the (a,c,e) naked mole-rat and (b,d,f) aged mouse. AI emphasizes the differences in these overall spectra, even though the differences seem small in a qualitative manner. Detailed differences are attributed to signals at ~1000 cm⁻¹ and the slope from 2000 cm⁻¹ to 4000 cm⁻¹.