Direct measurement of TRPV4 and PIEZO1 activity reveals multiple mechanotransduction pathways in chondrocytes.
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ABSTRACT:
SUBMITTER: Servin-Vences MR
PROVIDER: S-EPMC5279942 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Mechanical loading of joints plays a critical role in maintaining the health and function of articular cartilage. The mechanism(s) of chondrocyte mechanotransduction are not fully understood, but could provide important insights into new physical or pharmacologic therapies for joint diseases. Transient receptor potential vanilloid 4 (TRPV4), a Ca(2+)-permeable osmomechano-TRP channel, is highly expressed in articular chondrocytes, and loss of TRPV4 function is associated with joint arthropathy and osteoarthritis. The goal of this study was to examine the hypothesis that TRPV4 transduces dynamic compressive loading in articular chondrocytes. We first confirmed the presence of physically induced, TRPV4-dependent intracellular Ca(2+) signaling in agarose-embedded chondrocytes, and then used this model system to study the role of TRPV4 in regulating the response of chondrocytes to dynamic compression. Inhibition of TRPV4 during dynamic loading prevented acute, mechanically mediated regulation of proanabolic and anticatabolic genes, and furthermore, blocked the loading-induced enhancement of matrix accumulation and mechanical properties. Furthermore, chemical activation of TRPV4 by the agonist GSK1016790A in the absence of mechanical loading similarly enhanced anabolic and suppressed catabolic gene expression, and potently increased matrix biosynthesis and construct mechanical properties. These findings support the hypothesis that TRPV4-mediated Ca(2+) signaling plays a central role in the transduction of mechanical signals to support cartilage extracellular matrix maintenance and joint health. Moreover, these insights raise the possibility of therapeutically targeting TRPV4-mediated mechanotransduction for the treatment of diseases such as osteoarthritis, as well as to enhance matrix formation and functional properties of tissue-engineered cartilage as an alternative to bioreactor-based mechanical stimulation.
| S-EPMC3910592 | biostudies-literature
Project description:Osteoarthritis (OA) is a painful and debilitating condition of synovial joints without any disease-modifying therapies [A. M. Valdes, T. D. Spector, Nat. Rev. Rheumatol. 7, 23-32 (2011)]. We previously identified mechanosensitive PIEZO channels, PIEZO1 and PIEZO2, both expressed in articular cartilage, to function in chondrocyte mechanotransduction in response to injury [W. Lee et al., Proc. Natl. Acad. Sci. U.S.A. 111, E5114-E5122 (2014); W. Lee, F. Guilak, W. Liedtke, Curr. Top. Membr. 79, 263-273 (2017)]. We therefore asked whether interleukin-1-mediated inflammatory signaling, as occurs in OA, influences Piezo gene expression and channel function, thus indicative of maladaptive reprogramming that can be rationally targeted. Primary porcine chondrocyte culture and human osteoarthritic cartilage tissue were studied. We found that interleukin-1α (IL-1α) up-regulated Piezo1 in porcine chondrocytes. Piezo1 expression was significantly increased in human osteoarthritic cartilage. Increased Piezo1 expression in chondrocytes resulted in a feed-forward pathomechanism whereby increased function of Piezo1 induced excess intracellular Ca2+ at baseline and in response to mechanical deformation. Elevated resting state Ca2+ in turn rarefied the F-actin cytoskeleton and amplified mechanically induced deformation microtrauma. As intracellular substrates of this OA-related inflammatory pathomechanism, in porcine articular chondrocytes exposed to IL-1α, we discovered that enhanced Piezo1 expression depended on p38 MAP-kinase and transcription factors HNF4 and ATF2/CREBP1. CREBP1 directly bound to the proximal PIEZO1 gene promoter. Taken together, these signaling and genetic reprogramming events represent a detrimental Ca2+-driven feed-forward mechanism that can be rationally targeted to stem the progression of OA.
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Project description:Fibrosis is an irreversible, debilitating condition marked by the excessive production of extracellular matrix and tissue scarring that eventually results in organ failure and disease. Differentiation of fibroblasts to hypersecretory myofibroblasts is the key event in fibrosis. Although both soluble and mechanical factors are implicated in fibroblast differentiation, much of the focus is on TGF-β signaling, but to date, there are no specific drugs available for the treatment of fibrosis. In this review, we describe the role for TRPV4 mechanotransduction in cardiac and lung fibrosis, and we propose TRPV4 as an alternative therapeutic target for fibrosis.
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Project description:Group-2 innate-lymphoid cells (ILC2s) are critical mediators of the type-2 immune responses in multiple lung pathologies. We show that Piezo1, a mechanosensitive ion channel, plays a key role in regulating ILC2 functions by linking mechanical cues to biochemical signaling pathways. Both murine and human ILC2s strongly express Piezo1, and its activation by Yoda1 selectively enhances IL-13 production through calcium influx, which activates the mTOR-S6K pathway. This pathway leads to translational reprogramming, favoring IL-13 translation. Piezo1-deficient in ILC2s impairs this process, reducing IL-13 levels and resulting in attenuated lung inflammation and fibrosis in mouse models of IL-33- or Alternaria alternata-induced airway inflammation and bleomycin-induced fibrosis. These findings position Piezo1 as a critical mediator of ILC2-driven type-2 immune responses and highlight its potential as a therapeutic target for lung diseases characterized by excessive inflammation. This streamlined understanding of Piezo1 function improves focus on its mechanistic role in lung pathology.
2024-09-30 | GSE278085 | GEO
Project description:BackgroundMutations in PIEZO1 (Piezo type mechanosensitive ion channel component 1) cause human lymphatic malformations. We have previously uncovered an ORAI1 (ORAI calcium release-activated calcium modulator 1)-mediated mechanotransduction pathway that triggers lymphatic sprouting through Notch downregulation in response to fluid flow. However, the identity of its upstream mechanosensor remains unknown. This study aimed to identify and characterize the molecular sensor that translates the flow-mediated external signal to the Orai1-regulated lymphatic expansion.MethodsVarious mutant mouse models, cellular, biochemical, and molecular biology tools, and a mouse tail lymphedema model were employed to elucidate the role of Piezo1 in flow-induced lymphatic growth and regeneration.ResultsPiezo1 was found to be abundantly expressed in lymphatic endothelial cells. Piezo1 knockdown in cultured lymphatic endothelial cells inhibited the laminar flow-induced calcium influx and abrogated the flow-mediated regulation of the Orai1 downstream genes, such as KLF2 (Krüppel-like factor 2), DTX1 (Deltex E3 ubiquitin ligase 1), DTX3L (Deltex E3 ubiquitin ligase 3L,) and NOTCH1 (Notch receptor 1), which are involved in lymphatic sprouting. Conversely, stimulation of Piezo1 activated the Orai1-regulated mechanotransduction in the absence of fluid flow. Piezo1-mediated mechanotransduction was significantly blocked by Orai1 inhibition, establishing the epistatic relationship between Piezo1 and Orai1. Lymphatic-specific conditional Piezo1 knockout largely phenocopied sprouting defects shown in Orai1- or Klf2- knockout lymphatics during embryo development. Postnatal deletion of Piezo1 induced lymphatic regression in adults. Ectopic Dtx3L expression rescued the lymphatic defects caused by Piezo1 knockout, affirming that the Piezo1 promotes lymphatic sprouting through Notch downregulation. Consistently, transgenic Piezo1 expression or pharmacological Piezo1 activation enhanced lymphatic sprouting. Finally, we assessed a potential therapeutic value of Piezo1 activation in lymphatic regeneration and found that a Piezo1 agonist, Yoda1, effectively suppressed postsurgical lymphedema development.ConclusionsPiezo1 is an upstream mechanosensor for the lymphatic mechanotransduction pathway and regulates lymphatic growth in response to external physical stimuli. Piezo1 activation presents a novel therapeutic opportunity for preventing postsurgical lymphedema. The Piezo1-regulated lymphangiogenesis mechanism offers a molecular basis for Piezo1-associated lymphatic malformation in humans.
| S-EPMC9308715 | biostudies-literature
Project description:The growth factor TGFβ and the mechanosensitive calcium-permeable cation channel TRPV4 are both important for the development and maintenance of many tissues. Although TRPV4 and TGFβ both affect core cellular functions, how their signals are integrated is unknown. Here we show that pharmacological activation of TRPV4 significantly increased the canonical response to TGFβ stimulation in chondrocytes. Critically, this increase was only observed when TRPV4 was activated after, but not before TGFβ stimulation. The increase was prevented by pharmacological TRPV4 inhibition or knockdown and is calcium/CamKII dependent. RNA-seq analysis after TRPV4 activation showed enrichment for the TGFβ signalling pathway and identified JUN and SP1 as key transcription factors involved in this response. TRPV4 modulation of TGFβ signalling represents an important pathway linking mechanical signalling to tissue development and homeostasis.
| S-EPMC8064313 | biostudies-literature
Project description:Mechanical force is a fundamental regulator of bone development and homeostasis. Mechanosensitive osteocytes are the most abundant bone cells that form interconnected dendrites to respond to mechanical stimuli and interact with the bone-forming osteoblasts and the bone-remodeling osteoclasts. However, the molecular mechanisms underlying osteocyte maturation and dendrite formation remain unclear. By generating a Piezo1 "knock-out" osteocyte cell line, we identified a key role of Piezo1-mediated mechanotransduction in osteocyte differentiation. QRT-PCR analysis revealed delayed osteocyte differentiation in the Piezo1 KO cells relative to WT cells. By performing bulk RNA sequencing of WT and Piezo1 KO OCY454 cells at early (D1), intermediate (D14), and late (D28) stages of differentiation allowed for the identification of key signaling pathways in driving normal osteocyte dfiferentiation, as well as those regulated by Piezo1 mechanotransduction.
2024-09-20 | GSE277264 | GEO
Project description:Cystic fibrosis (CF) is a life-shortening genetic disease caused by a mutation in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Despite reports of CFTR expression on endothelial cells, pulmonary vascular perturbations, and perfusion deficit in CF patients, the mechanism of pulmonary vascular disease in CF remains unclear. Here, we describe loss of small pulmonary blood vessels in CF patients with severe lung disease. Using a vessel-on-a-chip model, we establish a shear stress-dependent mechanism of endothelial barrier failure in CF involving calcium-permeable mechanosensitive channel TRPV4. Furthermore, we demonstrate that CFTR deficiency downregulates the function of PIEZO1, another calcium-permeable mechanosensitive channel involved in angiogenesis and wound repair, and further exacerbates loss of small pulmonary blood vessels. We show that CFTR directly interacts with PIEZO1 and enhances its function, and that CFTR deficiency reduces PIEZO1 activity. Our study identifies key cellular targets to mitigate loss of small pulmonary blood vessels in CF.
2024-09-18 | GSE244214 | GEO
Project description:The mechanoelectrical transducer (MET) channels located at the stereocilia tip of cochlear hair cells are crucial to convert the mechanical energy of sound into receptor potentials, but the identity of its pore-forming subunits remains uncertain. Piezo1, which has been identified in the transcriptome of mammalian cochlear hair cells, encodes a transmembrane protein that forms mechanosensitive channels in other tissues. We investigated the properties of the MET channel in outer hair cells (OHCs) of Piezo1 mice (postnatal day 6-9). The MET current was elicited by deflecting the hair bundle of OHCs using sinewave and step stimuli from a piezo-driven fluid jet. Apical and basal OHCs were investigated because the properties of the MET channel vary along the cochlea. We found that the maximal MET current amplitude and the resting open probability of the MET channel in OHCs were similar between Piezo1(+/-) haploinsufficient mice and wild-type littermates. The sensitivity to block by the permeant MET channel blocker dihydrostreptomycin was also similar between the two genotypes. Finally, the anomalous mechano-gated current, which is activated by sheer force and which is tip-link independent, was unaffected in OHCs from Piezo1(+/-) haploinsufficient mice. Our results suggest that Piezo1 is unlikely to be a component of the MET channel complex in mammalian cochlear OHCs.
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Project description:Elevated pressure in the pancreatic gland is the central cause of pancreatitis following abdominal trauma, surgery, endoscopic retrograde cholangiopancreatography, and gallstones. In the pancreas, excessive intracellular calcium causes mitochondrial dysfunction, premature zymogen activation, and necrosis, ultimately leading to pancreatitis. Although stimulation of the mechanically activated, calcium-permeable ion channel Piezo1 in the pancreatic acinar cell is the initial step in pressure-induced pancreatitis, activation of Piezo1 produces only transient elevation in intracellular calcium that is insufficient to cause pancreatitis. Therefore, how pressure produces a prolonged calcium elevation necessary to induce pancreatitis is unknown. We demonstrate that Piezo1 activation in pancreatic acinar cells caused a prolonged elevation in intracellular calcium levels, mitochondrial depolarization, intracellular trypsin activation, and cell death. Notably, these effects were dependent on the degree and duration of force applied to the cell. Low or transient force was insufficient to activate these pathological changes, whereas higher and prolonged application of force triggered sustained elevation in intracellular calcium, leading to enzyme activation and cell death. All of these pathological events were rescued in acinar cells treated with a Piezo1 antagonist and in acinar cells from mice with genetic deletion of Piezo1. We discovered that Piezo1 stimulation triggered transient receptor potential vanilloid subfamily 4 (TRPV4) channel opening, which was responsible for the sustained elevation in intracellular calcium that caused intracellular organelle dysfunction. Moreover, TRPV4 gene-KO mice were protected from Piezo1 agonist- and pressure-induced pancreatitis. These studies unveil a calcium signaling pathway in which a Piezo1-induced TRPV4 channel opening causes pancreatitis.
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