Project Summary
Our multidisciplinary team assembles basic and translational researchers with expertise in joint biology and
neuroscience, proposing a holistic approach to mapping the sensory innervation of murine and human knee
joints. We will use state-of-the-art imaging techniques, combined with transcriptomics to construct 3D models of
the sensory innervation of the knee, compose a cell atlas in which knee afferents are transcriptionally profiled at
a single cell resolution, and document the nerve-joint cell interactome at the transcriptional level. Our overarching
objective is to precisely describe the sensory innervation of the knee, and the dynamic changes occurring with
aging, joint injury, and osteoarthritis (OA). This will provide the Consortium with a rich anatomical and molecular
resource to study mechanisms underlying joint pain and guide the development of novel analgesic strategies.
Aim 1. Documenting the sensory innervation of the healthy and diseased mouse knee: Anatomical and molecular
perspectives. Using fluorescent reporter mice to label nociceptors, C-fiber subsets, and proprioceptors, we will
map the anatomical innervation of the mouse knee in (a) naïve mice of different ages; (b) after joint injury; (c) in
surgically induced OA. We will use ribbon scanning confocal and clearing-enabled lightsheet microscopy to
construct high-resolution 3-D anatomical models of joint innervation. We will backlabel knee-innervating afferents
and use spatial transcriptomics to describe their molecular phenotypes compared to other non-knee innervating
DRG neurons. Aim 2. Documenting the sensory innervation of the healthy and diseased human knee: Anatomical
and molecular perspectives. We will use a unique set of post mortem knee/DRG samples from (1) healthy knees,
age 20-40 (n=15/sex); (2) knees from donors over 70 (n=15/sex), in which we anticipate 80-90% to exhibit OA
pathology. Knee tissues will be collected in a standardized fashion, including synovium, osteochondral plugs
(medial tibial plateau), meniscus, ACL, fat pad, and quadriceps muscle. In each tissue, we will perform (1)
histopathology; (2) IHC for sensory innervation; (3) bulk and scRNAseq; (4) spatial transcriptomics. Matched
DRGs will be used for bulk RNAseq to identify differentially expressed genes (DEG) between the groups provide
information for ligand-receptor analysis. Aim 3. Identifying mediators in the knee synovium that drive disease-
associated neuroplasticity. (1) We will reconstruct the cellular interactome between synovial cells and DRG
neurons in mouse models of aging, joint injury, and OA using scRNAseq of matched synovium and DRG
samples. (2) We will compare patient reports of OA knee pain at the time of TKR to matched synovial histology,
including extent of lining hyperplasia, single-cell transcriptional changes, and innervation. Overall, this project
will provide the community with comprehensive databases of the neuro-articular environment, which can be
mined to (1) undertake mechanistic studies to inhibit pathological neuroplasticity and (2) identify and test new
druggable targets. This strategy will pave the way for the development of novel, targeted, non-addictive, and
safe analgesic therapeutics for the treatment of joint pain.