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The human PKD2 locus encodes Polycystin-2 (PC2), a TRPP channel that localises to several distinct cellular compartments, including the cilium. PKD2 mutations cause Autosomal Dominant Polycystic Kidney Disease (ADPKD) and affect many cellular pathways. Data underlining the importance of ciliary PC2 localisation in preventing PKD are limited because PC2 function is ablated throughout the cell in existing model systems. Here, we dissect the ciliary role of PC2 by analysing mice carrying a non-ciliary localising, yet channel-functional, PC2 mutation. Mutants develop embryonic renal cysts that appear indistinguishable from mice completely lacking PC2. Despite not entering the cilium in mutant cells, mutant PC2 accumulates at the ciliary base, forming a ring pattern consistent with distal appendage localisation. This suggests a two-step model of ciliary entry; PC2 first traffics to the cilium base before TOP domain dependent entry. Our results suggest that PC2 localisation to the cilium is necessary to prevent PKD., The molecular role of ciliary Polycystin-2 (PC2) in cyst formation and polycystic kidney disease (ADKPD) is unclear. Here, the authors identify a PC2 mutant lacking ciliary localisation but with active Ca2+ channel function in mice, that is sufficient to generate an ADPKD phenotype.

The role of mammalian cilia in cell signalling was first identified in embryonic development and subsequent analysis has revealed roles in multiple signalling pathways. We now understand that these developmental roles impact human health and this is evident in the class of ciliary diseases which we call the ciliopathies. By their nature cilia defects are usually pleiotropic, affecting more than one system. This often leads to early lethality, meaning that subsequent functions are harder to examine. Current studies are revealing previously unrealised cilia-related phenotypes later in embryonic development. Furthermore, they are exposing the importance of cell biology in understanding the mechanisms of cilia function. In this review, we discuss advances in the field.

Cigarette smoking has been identified as a major risk factor for osteoporosis decades ago. Several studies have shown a direct relationship between cigarette smoking, decreased bone mineral density, and impaired fracture healing. However, the mechanisms behind impaired fracture healing and cigarette smoking are yet to be elucidated. Migration and osteogenesis of mesenchymal stem/stromal cells (MSCs) into the fracture site play a vital role in the process of fracture healing. In human nicotine, the most pharmacologically active and major addictive component present in tobacco gets rapidly metabolized to the more stable cotinine. This study demonstrates that physiological concentrations of both nicotine and cotinine do not affect the osteogenic differentiation of MSCs. However, cigarette smoke exposure induces oxidative stress by increasing superoxide radicals and reducing intracellular glutathione in MSCs, negatively affecting osteogenic differentiation. Although, not actively producing reactive oxygen species (ROS) nicotine and cotinine inhibit catalase and glutathione reductase activity, contributing to an accumulation of ROS by cigarette smoke exposure. Coincubation with N-acetylcysteine or L-ascorbate improves impaired osteogenesis caused by cigarette smoke exposure by both activation of nuclear factor erythroid 2-related factor 2 (Nrf2) signaling and scavenging of ROS, which thus might represent therapeutic targets to support fracture healing in smokers.

Transforming growth factor β (TGF-β) is a critical regulator of bone density owing to its multiple effects on cell growth and differentiation. Recently, we have shown that TGF-β1 effectively blocks bone morphogenetic protein (BMP) induced maturation of osteoblasts by upregulating histone deacetylase (HDAC) activity. The current study aimed at investigating the effect of rhTGF-β1 treatment on the expression of specific HDACs and their cellular effects, e.g., microtubule structures (primary cilia) and mechanosensation. Exposure to TGF-β1 most significantly induced expression of HDAC6 both on gene and protein level. Being most abundant in the cytoplasm HDAC6 effectively deacetylates microtubule structures. Thus, TGF-β1-induced expression of HDAC6 led to deformation and shortening of primary cilia as well as to reduced numbers of ciliated cells. Primary cilia are described to sense mechanical stimuli. Thus, fluid flow was applied to the cells, which stimulated osteoblast function (AP activity and matrix mineralization). Compromised primary cilia in TGF-β1-treated cells were associated with reduced osteogenic function, despite exposure to fluid flow conditions. Chemical inhibition of HDAC6 with Tubacin restored primary cilium structure and length. These cells showed improved osteogenic function especially under fluid flow conditions. Summarizing our results, TGF-β1 impairs human osteoblast maturation partially via HDAC6-mediated distortion and/or shortening of primary cilia. This knowledge opens up new treatment options for trauma patients with chronically elevated TGF-β1-levels (e.g., diabetics), which frequently suffer from delayed fracture healing despite adequate mechanical stimulation.

Promotion of rhBMP2 and rhBMP7 for the routine use to support fracture healing has been hampered by high costs, safety concerns and reasonable failure rates, imposing restrictions in its clinical use. Since there is little debate regarding its treatment potential, there is rising need for a better understanding of the mode of action of these BMPs to overcome its drawbacks and promote more efficacious treatment strategies for bone regeneration. Recently, BMP9, owing to its improved osteogenic potential, is gaining attention as a promising therapeutic alternative. Our study aimed at identifying specific gene expression patterns which may predict and explain individual responses to rhBMP7 and rhBMP9 treatments. Therefore, we investigated the effect of rhBMP7 and rhBMP9 on primary human osteoblasts from 110 donors and corresponding THP-1-derived osteoclasts. This was further compared with each other and our reported data on rhBMP2 response. Based on the individual donor response, we found three donor groups profiting from rhBMP treatment either directly via stimulation of osteoblast function or viability and/or indirectly via inhibition of osteoclasts. The response on rhBMP7 treatment correlated with expression levels of the genes BAMBI, SOST, Noggin, Smad4 and RANKL, while the response of rhBMP9 correlated to the expression levels of Alk6, Endoglin, Smurf1, Smurf2, SOST and RANKL in these donors. Noteworthy, rhBMP9 treatment showed significantly increased osteogenic activity (AP activity and Smad nuclear translocation) when compared to the two clinically used rhBMPs. Based on patient's respective expression profiles, clinical application of rhBMP9 either solely or in combination with rhBMP2 and/or rhBMP7 can become a promising new approach to fit the patient's needs to promote fracture healing.

Several studies have explored the negative effects of cigarette smoke on bone healing; however, the complex pathogenesis still remains unclear. One crucial and primary factor determining effective fracture repair is the recruitment and differentiation of mesenchymal stem cells (MSCs) into bone-forming cells. Recently, primary cilia, microtubule-based sensory organelles, have been shown to be critical in lineage commitment and differentiation of MSCs. Our present study indicates that exposure to cigarette smoke extract (CSE 0.1–10%) impaired osteogenic differentiation of human mesenchymal stem cell line (SCP-1) and interestingly, also affected primary cilia distribution and integrity in these cells during the differentiation. Furthermore, significant amounts of free radicals generated by CSE could be causative of primary cilia loss since treatment with 0.01% of hydrogen peroxide, a prime free radical in CSE, destroyed primary cilia in these cells. The debilitated differentiation of CSE-exposed SCP-1 cells also correlated with the significantly reduced expression of transcription factor and target genes of primary cilia-specific hedgehog signalling, a key player in osteogenic differentiation. As a treatment strategy, co-incubation of the CSE-exposed SCP-1 cells with the antioxidant resveratrol (1 µM) had a protective effect as it significantly reduced free radical production, protected the primary cilia and enhanced osteogenic differentiation. The current study shows for the first time that cigarette smoke affects primary cilia in human MSCs during osteogenic differentiation and treatment with resveratrol could reverse the effects and enhance differentiation, thus opening up potential therapeutic alternatives to treat fracture healing in smokers, in particularly, when delayed fracture healing is assumed.

Transforming growth factor β (TGF-β) is a critical regulator of bone density owing to its multiple effects on cell growth and differentiation. Recently, we have shown that TGF-βExposure to TGF-β