Role of Ciliary Protein Intraflagellar Transport Protein 88 in the Regulation of Cartilage Thickness and Osteoarthritis Development in Mice

Abstract

Objective. Mechanical and biologic cues drive cellular signaling in cartilage development, health, and disease. Primary cilia proteins, which are implicated in the transduction of biologic and physiochemical signals, control cartilage formation during skeletal development. This study was undertaken to assess the influence of the ciliary protein intraflagellar transport protein 88 (IFT88) on postnatal cartilage from mice with conditional knockout of the Ift88 gene (Ift88-KO).Methods. Ift88 fl/fl and aggrecanCre ERT2 mice were crossed to create a strain of cartilage-specific Ift88-KO mice (aggrecanCre ERT2 ;Ift88 fl/fl ). In these Ift88-KO mice and Ift88 fl/fl control mice, tibial articular cartilage thickness was assessed by histomorphometry, and the integrity of the cartilage was assessed using Osteoarthritis Research Society International (OARSI) damage scores, from adolescence through adulthood. In situ mechanisms of cartilage damage were investigated in the microdissected cartilage sections using immunohistochemistry, RNAScope analysis, and quantitative polymerase chain reaction. Osteoarthritis (OA) was induced in aggrecanCre ERT2 ;Ift88 fl/fl mice and Ift88 fl/fl control mice using surgical destabilization of the medial meniscus (DMM). Following tamoxifen injection and DMM surgery, the mice were given free access to exercise on a wheel.Results. Deletion of Ift88 resulted in progressive reduction in the thickness of the medial tibial cartilage in adolescent mice, as well as marked atrophy of the cartilage in mice during adulthood. In aggrecanCre ERT2 ;Ift88 fl/fl mice at age 34 weeks, the median thickness of the medial tibial cartilage was 89.42 μm (95% confidence interval [95% CI] 84.00-93.49), whereas in Ift88 fl/fl controls at the same age, the median cartilage thickness was 104.00 μm (95% CI 100.30-110.50; P < 0.0001). At all time points, the median thickness of the calcified cartilage was reduced. In some mice, atrophy of the medial tibial cartilage was associated with complete, spontaneous degradation of the cartilage. Following DMM, aggrecanCre ERT2 ;Ift88 fl/fl mice were found to have increased OARSI scores of cartilage damage. In articular cartilage from maturing mice, atrophy was not associated with obvious increases in aggrecanase-mediated destruction or chondrocyte hypertrophy. Of the 44 candidate genes analyzed, only Tcf7l2 expression levels correlated with Ift88 expression levels in the microdissected cartilage. However, RNAScope analysis revealed that increased hedgehog (Hh) signaling (as indicated by increased expression of Gli1) was associated with the reductions in Ift88 expression in the tibial cartilage from Ift88-deficient mice. Wheel exercise restored both the articular cartilage thickness and levels of Hh signaling in these mice.Conclusion. Our results in a mouse model of OA demonstrate that IFT88 performs a chondroprotective role in articular cartilage by controlling the calcification of cartilage via maintenance of a threshold of Hh signaling during physiologic loading.

Isolation, profiling, and tracking of extracellular vesicle cargo in Caenorhabditis elegans

“…Taken together, our data establish the cilium as a cellular location for ENPP- Article in cilia, mammalian ENPP1 loss of function manifests signs of ciliopathies, such as accelerated ossification, hearing impairment, and altered insulin metabolism and immune response. 39,40 To validate the IL2-enriched category of EV cargo candidates (Category I; Figure 2F), we focused on another nucleic acid binding protein, minichromosome maintenance complex component 3 (MCM-3) licensing factor, and generated transgenic animals expressing MCM-3::mScarlet, driven by its native promoter. MCM-3::mScarlet showed nuclear localization in the developing embryo, consistent with the known function of the MCM proteins in DNA replication licensing and DNA helicase activity of the MCM-2-7 complex 41 (Figure S5A).…”

Role of Primary Cilia in Skeletal Disorders

“…Deletion of IFT88 in cartilage causes several OA phenotypes with increased expression levels of degeneration markers, including MMP13, collagen type X, Adamts5, and Runx2 [ 8 ]. Similarly, the Bardet-Biedl syndrome 1 (Bbs1), Bbs2, or Bbs6 gene mutation mouse model developed OA-like cartilage abnormalities including proteoglycan loss, small surface fibrillation, marked atrophy of the cartilage, and increased MMP13 expression [ 42 , 43 ]. Moreover, IFT88 deletion following surgical destabilization of the medial meniscus was found to have increased OARSI scores of cartilage damage mouse [ 43 ].…”

“…Similarly, the Bardet-Biedl syndrome 1 (Bbs1), Bbs2, or Bbs6 gene mutation mouse model developed OA-like cartilage abnormalities including proteoglycan loss, small surface fibrillation, marked atrophy of the cartilage, and increased MMP13 expression [ 42 , 43 ]. Moreover, IFT88 deletion following surgical destabilization of the medial meniscus was found to have increased OARSI scores of cartilage damage mouse [ 43 ]. All these studies suggested that primary cilia are essential for cartilage development and prevent its degeneration.…”

Roles of the calcified cartilage layer and its tissue engineering reconstruction in osteoarthritis treatment

“…Pharmaceutically, the long-term application of dexamethasone increases the calcium content of CCL in both murine model and OA patients, which can also predispose OA and accelerate OA progression ( Chen et al, 2021 ). In the contrast, a list of molecules that perform chondroprotective roles through inhibition of cartilage calcification is also found, including ciliary protein intraflagellar transport protein 88 (IFT88) ( Coveney et al, 2022 ), betaine ( Yajun et al, 2021 ), gasotransmitter hydrogen sulfide (H 2 S) ( Nasi et al, 2021 ), and Tougu Xiaotong capsule ( Li et al, 2013 ). Phosphocitrate (PC) and its analogs can restrain endochondral ossification by suppressing the hypertrophic phenotype ( Hartlev et al, 2018 ).…”