Gene therapy rescues cilia defects and restores olfactory function in a mammalian ciliopathy model

Department of Pharmacology, University of Michigan, Ann Arbor, Michigan, USA.
Nature medicine (Impact Factor: 27.36). 09/2012; 18(9). DOI: 10.1038/nm.2860
Source: PubMed


Cilia are evolutionarily conserved microtubule-based organelles that are crucial for diverse biological functions, including motility, cell signaling and sensory perception. In humans, alterations in the formation and function of cilia manifest clinically as ciliopathies, a growing class of pleiotropic genetic disorders. Despite the substantial progress that has been made in identifying genes that cause ciliopathies, therapies for these disorders are not yet available to patients. Although mice with a hypomorphic mutation in the intraflagellar transport protein IFT88 (Ift88(Tg737Rpw) mice, also known as ORPK mice) have been well studied, the relevance of IFT88 mutations to human pathology is unknown. We show that a mutation in IFT88 causes a hitherto unknown human ciliopathy. In vivo complementation assays in zebrafish and mIMCD3 cells show the pathogenicity of this newly discovered allele. We further show that ORPK mice are functionally anosmic as a result of the loss of cilia on their olfactory sensory neurons (OSNs). Notably, adenoviral-mediated expression of IFT88 in mature, fully differentiated OSNs of ORPK mice is sufficient to restore ciliary structures and rescue olfactory function. These studies are the first to use in vivo therapeutic treatment to reestablish cilia in a mammalian ciliopathy. More broadly, our studies indicate that gene therapy is a viable option for cellular and functional rescue of the complex ciliary organelle in established differentiated cells.

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Available from: Dyke P Mcewen
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    • "Treatment of anosmia due to ciliary defects in olfactory sensory neurons is a potential target for gene delivery, via intranasal injection. AV-mediated IFT88 transduction in fully differentiated olfactory sensory neurons in Ift88 mutant mice saw restoration of ciliary structures and rescue of olfactory function (McIntyre et al., 2012). Lentivirus has recently been proven beneficial for gene therapy against childhood lysosomal disorders (Aiuti et al., 2013; Biffi et al., 2013); however, lentivirus has not yet been widely utilized for gene delivery in ciliopathies except for in vitro studies. "
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    ABSTRACT: Primary cilia were the largely neglected nonmotile counterparts of their better-known cousin, the motile cilia. For years these nonmotile cilia were considered evolutionary remnants of little consequence to cellular function. Fast forward 10 years and we now recognize primary cilia as key integrators of extracellular ligand-based signaling and cellular polarity, which regulate neuronal cell fate, migration, differentiation, as well as a host of adult behaviors. Important future questions will focus on structure-function relationships, their roles in signaling and disease and as areas of target for treatments.
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    • "On neurons, disruption of the primary cilium or ciliary proteins revealed that they have an important role in regulating satiation and object recognition through unknown mechanisms [16-18]. In addition, cilia are known to be important for sight and smell [16,19-21] and recently it was demonstrated that they regulate pathways involved in adult neurogenesis and migration of newborn neurons [22]. However, the ubiquitous nature of primary cilia on most neurons in the central nervous system (CNS) was unexpected [23]. "
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    ABSTRACT: Cilia are found on nearly every cell type in the mammalian body, and have been historically classified as either motile or immotile. Motile cilia are important for fluid and cellular movement; however, the roles of non-motile or primary cilia in most tissues remain unknown. Several genetic syndromes, called the ciliopathies, are associated with defects in cilia structure or function and have a wide range of clinical presentations. Much of what we know about the formation and maintenance of cilia comes from model systems like C. elegans and Chalmydomonas. Studies of mammalian cilia in live tissues have been hampered by difficulty visualizing them. To facilitate analyses of mammalian cilia function we generated an inducible CiliaGFP mouse by targeting mouse cDNA encoding a cilia-localized protein somatostatin receptor 3 fused to GFP (Sstr3::GFP) into the ROSA26 locus. In this system, Sstr3::GFP is expressed from the ubiquitous ROSA26 promoter after Cre mediated deletion of an upstream Neo cassette flanked by lox P sites. Fluorescent cilia labeling was observed in a variety of live tissues and after fixation. Both cell-type specific and temporally regulated cilia labeling were obtained using multiple Cre lines. The analysis of renal cilia in anesthetized live mice demonstrates that cilia commonly lay nearly parallel to the apical surface of the tubule. In contrast, in more deeply anesthetized mice the cilia display a synchronized, repetitive oscillation that ceases upon death, suggesting a relationship to heart beat, blood pressure or glomerular filtration. The ability to visualize cilia in live samples within the CiliaGFP mouse will greatly aid studies of ciliary function. This mouse will be useful for in vivo genetic and pharmacological screens to assess pathways regulating cilia motility, signaling, assembly, trafficking, resorption and length control and to study cilia regulated physiology in relation to ciliopathy phenotypes.
    Full-text · Article · Jul 2013 · Cilia
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    • "Overall, our findings implicate the role of IFT80 in chondrocyte differentiation and provide direct evidence for IFT80 regulation in the Hh and Wnt/b-catenin signaling pathways. Most recently, McIntyre [69] et al. found that adenoviral-mediated expression of IFT88 can restore ciliary structures and rescue olfactory function in IFT88 mutant mice. This study provides a proof and an effective therapeutic option by using IFT proteins to rescue of the complex ciliary organelle in IFT/Cilia and Hh signaling related human diseases. "
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    ABSTRACT: Partial mutation of intraflagellar transport 80 (IFT80) in humans causes Jeune asphyxiating thoracic dystrophy (JATD) and short-rib polydactyly (SRP) syndrome type III. These diseases are autosomal recessive chondrodysplasias that share clinical similarities, including shortened long bones and constricted thoracic cage. However, the role and mechanism of IFT80 in the regulation of chondrocyte differentiation and function remain largely unknown. We hypothesize that IFT80 is required for the formation and function of cilia and plays a critical role in chondrogenic differentiation by regulating Hedgehog (Hh) and Wingless (Wnt) signaling pathways. To test this hypothesis, we first analyzed the IFT80 expression pattern and found that IFT80 was predominantly expressed in growth plate chondrocytes and during chondrogenic differentiation. Silencing IFT80 impaired cilia formation and chondrogenic differentiation in mouse bone marrow derived stromal cells (BMSCs), and decreased the expression of chondrocyte marker genes - collagen II and aggrecan. Additionally, silencing IFT80 down-regulated Hh signaling activity whereas up-regulated Wnt signaling activity. The overexpression of Gli2 in IFT80-silenced cells promoted chondrogenesis and recovered the chondrogenic deficiency from IFT80 silencing. Overall, our results demonstrate that IFT80 is essential for chondrocyte differentiation by regulating the Hh and Wnt signaling pathways.
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