Mapping cell diversity and dynamics in inflammatory temporomandibular joint osteoarthritis with pain at single-cell resolution
Supawadee Jariyasakulroj, Yang Shu, Ziying Lin, Jingyi Chen, Qing Chang, Pao-Fen Ko, Jian-Fu Chen
Supawadee Jariyasakulroj, Yang Shu, Ziying Lin, Jingyi Chen, Qing Chang, Pao-Fen Ko, Jian-Fu Chen
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Research Article Cell biology Inflammation

Mapping cell diversity and dynamics in inflammatory temporomandibular joint osteoarthritis with pain at single-cell resolution

Abstract

Temporomandibular joint (TMJ) osteoarthritis with pain is a highly prevalent disorder affecting patients’ quality of life. A comprehensive understanding of cell type diversity and its dynamics in painful TMJ osteoarthritis (TMJOA) is lacking. Here, we utilized an inflammatory TMJOA mouse model via intra-articular injection of CFA. TMJOA mice exhibited cartilage remodeling, bone loss, synovitis, increased osteoarthritis (OA) score, and orofacial pain, recapitulating hallmark symptoms in patients. Single-cell transcriptomic profiling of the TMJ was performed in conjunction with mouse genetic labeling, tissue clearing, light sheet and confocal 3D imaging, multiplex RNAscope, and immunodetection. We visualized, reconstructed, and analyzed the distribution and density of nociceptive innervation of TMJ at single-axon levels. We systematically mapped the heterogeneity and anatomical position of blood endothelial cells, synovial fibroblasts, and immune cells, including Cx3cr1-positive barrier macrophages. Importantly, TMJOA mice exhibited enhanced neurovascular coupling, sublining fibroblast hyperplasia, inflammatory immune cell expansion, disrupted signaling-dependent cell-cell interaction, and a breakdown of the sandwich-like organization consisting of synovial barrier macrophages and fibroblasts. By utilizing a mouse model with combined TMJ pain history and OA, we reveal the cellular diversity, anatomical structure, and cell dynamics of the TMJ at single-cell resolution, which facilitate our understanding and potential targeting of TMJOA.

Authors

Supawadee Jariyasakulroj, Yang Shu, Ziying Lin, Jingyi Chen, Qing Chang, Pao-Fen Ko, Jian-Fu Chen

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Figure 8

The functional importance of Igf1 signaling in barrier macrophages in TMJOA.

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The functional importance of Igf1 signaling in barrier macrophages in TM...
(A and B) The expression of Cx3cr1-YFP at the synovial membranes. Arrows indicate Cx3cr1+ macrophage lining cells (green). DAPI stains nuclei. Scale bar: 100 μm. (C) NetVisual_circle plot for the cell-cell interaction between major cell types in the TMJ. (D) Feature plots showing the expression of Cx3cr1 and Igf1 genes that are overlaid on the UMAP plot of CFA TMJ. (E and F) Confocal imaging of sagittal TMJ sections stained with Cx3cr1-YFP (green) in conjunction with RNAscope of Prg4 (red) and Thy1 (white). DAPI stains nuclei (blue). Images in F (63× objective) are enlargements of boxed regions in image E (4× objective). Scale bars: 100 μm. (G) Confocal imaging of sagittal TMJ sections stained with Cx3cr1-YFP in control and CFA mice. DAPI stains nuclei (blue). Scale bar: 100 μm. (H) Schematic of Cx3cr1 cells together with Iba1 macrophages and Ly6b neutrophils in control and CFA TMJ. (I and J) Confocal imaging of sagittal whole TMJ or anterior TMJ sections stained with antibodies against TUBB3 (red) and CGRP (green). DAPI stains nuclei (blue). Scale bars: 100 μm. (K) Quantification of the TUBB3+CGRP+ area fraction. (L) A schematic diagram of dynamic changes in cell type and anatomical position in the TMJ under healthy conditions and TMJOA with pain. All data are represented as mean ± SEM. Student’s t test. N = 3–5 mice, *P < 0.05.