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| Status |
Public on Jun 05, 2024 |
| Title |
The chondrocyte “mechanome”: Activation of the mechanosensitive ion channels TRPV4 and PIEZO1 drives unique transcriptional signatures |
| Organism |
Sus scrofa |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
The mechanosensitive ion channels Transient Receptor Potential Vanilloid 4 (TRPV4) and PIEZO1 transduce physiologic and supraphysiologic magnitudes of mechanical signals in the chondrocyte, respectively. TRPV4 activation promotes chondrogenesis, while PIEZO1 activation by supraphysiologic deformations drives cell death. The mechanisms by which activation of these channels discretely drives changes in gene expression to alter cell behavior remains to be determined. To date, no studies have contrasted the transcriptomic response to activation of these channels, nor has any published data attempted to correlate these transcriptomes to alterations in cellular function. This study used RNA sequencing to comprehensively investigate the transcriptomes associated with activation of TRPV4 or PIEZO1, revealing that TRPV4 and PIEZO drive distinct transcriptomes, but also exhibit unique co-regulated clusters of genes. Notably, activation of PIEZO1 through supraphysiologic deformation induced a transient inflammatory profile that overlapped with the interleukin (IL)-1-responsive transcriptome and contained genes associated with cartilage degradation and osteoarthritis progression. However, both TRPV4 and PIEZO1 were also shown to elicit anabolic effects. PIEZO1 expression promoted a pro-chondrogenic transcriptome under unloaded conditions, and daily treatment with PIEZO1 agonist Yoda1 significantly increased sulfated glycosaminoglycan deposition in vitro. These findings emphasize the presence of a broad “mechanome” with distinct effects of TRPV4 and PIEZO1 activation in chondrocytes, suggesting complex roles for PIEZO1 in both the physiologic and pathologic responses of chondrocytes. The identification of transcriptomic profiles unique to or shared by PIEZO1 and TRPV4 (distinct from IL-1-induced inflammation) could inform future therapeutic designs targeting these channels for the management and treatment of osteoarthritis.
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| Overall design |
To study the TRPV4-responsive transcriptome, primary chondrocytes were cast in 2% agarose to form tissue engineered cartilage constructs. Constructs were stimulated for 3 hours with cyclic, unconfined compressive loading (10% peak-to-peak deformation, 1 Hz) with or without TRPV4 antagonist GSK205 (10 uM) or treated with TRPV4 agonist GSK101 (1 nM) or 1 ng/mL of IL-1. Vehicle controls were exposed to DMSO (1:1000 in media) but left free swelling. For both TRPV4 and IL-1 stimulation groups, tissue constructs were taken down at 3 hours and 9 hours post stimulation onset, flash frozen, and stored at -80C (n=4 per group). To study the PIEZO1-responsive transcriptome, chondrocytes were plated at 500k cells per well in a 6 well plate and transfected (DharmaFECT Transfection Reagent, Dharmacon) with siRNA targeting PIEZO1 (siPIEZO1), a non-targeting control siRNA (siNTC), or left non-transfected (NT). All transfection groups were cast in 2% agarose to form tissue engineered cartilage constructs. Constructs were subjected to a single round of quasi-static unconfined compression to 80% deformation (supraphysiologic range) at 1.9% deformation/min (~0.73 um/sec), treated with PIEZO1 agonist Yoda1 (5 uM), or standard media with a DMSO vehicle control (1:1000) but left free swelling. In response to stimulation, tissue constructs were taken down at 3 hours and 9 hours post stimulation onset, flash frozen, and stored at -80C (n=4 per group).
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| Contributor(s) |
Nims R, Palmer D, Kassab J, Zhang B, Guilak F |
| Citation(s) |
38959010 |
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| Submission date |
Jun 05, 2024 |
| Last update date |
Sep 04, 2024 |
| Contact name |
Farshid Guilak |
| E-mail(s) |
guilak@wustl.edu
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| Organization name |
Washington University in St. Louis
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| Department |
Dept of Orthopedics
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| Lab |
Guilak Lab
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| Street address |
660 S Euclid Ave, Campus Box 8233
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| City |
Saint Louis |
| State/province |
MO |
| ZIP/Postal code |
63110 |
| Country |
USA |
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| Platforms (1) |
| GPL26351 |
Illumina NovaSeq 6000 (Sus scrofa) |
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| Samples (120)
|
| GSM8307210 |
GSK101 treatment, t=3 hours post-onset of stimulation, sample 1 |
| GSM8307211 |
GSK101 treatment, t=3 hours post-onset of stimulation, sample 2 |
| GSM8307212 |
GSK101 treatment, t=3 hours post-onset of stimulation, sample 3 |
| GSM8307213 |
GSK101 treatment, t=3 hours post-onset of stimulation, sample 4 |
| GSM8307214 |
GSK205 treatment, t=3 hours post-onset of stimulation, sample 1 |
| GSM8307215 |
GSK205 treatment, t=3 hours post-onset of stimulation, sample 2 |
| GSM8307216 |
GSK205 treatment, t=3 hours post-onset of stimulation, sample 3 |
| GSM8307217 |
GSK205 treatment, t=3 hours post-onset of stimulation, sample 4 |
| GSM8307218 |
DMSO vehicle control, t=3 hours post-onset of stimulation, sample 1 |
| GSM8307219 |
DMSO vehicle control, t=3 hours post-onset of stimulation, sample 2 |
| GSM8307220 |
DMSO vehicle control, t=3 hours post-onset of stimulation, sample 3 |
| GSM8307221 |
DMSO vehicle control, t=3 hours post-onset of stimulation, sample 4 |
| GSM8307222 |
IL-1 treatment, t=3 hours post-onset of stimulation, sample 1 |
| GSM8307223 |
IL-1 treatment, t=3 hours post-onset of stimulation, sample 2 |
| GSM8307224 |
IL-1 treatment, t=3 hours post-onset of stimulation, sample 3 |
| GSM8307225 |
IL-1 treatment, t=3 hours post-onset of stimulation, sample 4 |
| GSM8307226 |
Physiologic loading plus GSK205 treatment, t=3 hours post-onset of stimulation, sample 1 |
| GSM8307227 |
Physiologic loading plus GSK205 treatment, t=3 hours post-onset of stimulation, sample 2 |
| GSM8307228 |
Physiologic loading plus GSK205 treatment, t=3 hours post-onset of stimulation, sample 3 |
| GSM8307229 |
Physiologic loading plus GSK205 treatment, t=3 hours post-onset of stimulation, sample 4 |
| GSM8307230 |
Physiologic loading, t=3 hours post-onset of stimulation, sample 1 |
| GSM8307231 |
Physiologic loading, t=3 hours post-onset of stimulation, sample 2 |
| GSM8307232 |
Physiologic loading, t=3 hours post-onset of stimulation, sample 3 |
| GSM8307233 |
Physiologic loading, t=3 hours post-onset of stimulation, sample 4 |
| GSM8307234 |
non-transduced, DMSO vehicle control, t=3 hours post-onset of stimulation, sample 1 |
| GSM8307235 |
non-transduced, DMSO vehicle control, t=3 hours post-onset of stimulation, sample 2 |
| GSM8307236 |
non-transduced, DMSO vehicle control, t=3 hours post-onset of stimulation, sample 3 |
| GSM8307237 |
non-transduced, DMSO vehicle control, t=3 hours post-onset of stimulation, sample 4 |
| GSM8307238 |
non-transduced, supraphysiologic loading, t=3 hours post-onset of stimulation, sample 1 |
| GSM8307239 |
non-transduced, supraphysiologic loading, t=3 hours post-onset of stimulation, sample 2 |
| GSM8307240 |
non-transduced, supraphysiologic loading, t=3 hours post-onset of stimulation, sample 3 |
| GSM8307241 |
non-transduced, supraphysiologic loading, t=3 hours post-onset of stimulation, sample 4 |
| GSM8307242 |
non-transduced, Yoda1 treatment, t=3 hours post-onset of stimulation, sample 1 |
| GSM8307243 |
non-transduced, Yoda1 treatment, t=3 hours post-onset of stimulation, sample 2 |
| GSM8307244 |
non-transduced, Yoda1 treatment, t=3 hours post-onset of stimulation, sample 3 |
| GSM8307245 |
non-transduced, Yoda1 treatment, t=3 hours post-onset of stimulation, sample 4 |
| GSM8307246 |
transduced with non-targeting siRNA, DMSO vehicle control, t=3 hours post-onset of stimulation, sample 1 |
| GSM8307247 |
transduced with non-targeting siRNA, DMSO vehicle control, t=3 hours post-onset of stimulation, sample 2 |
| GSM8307248 |
transduced with non-targeting siRNA, DMSO vehicle control, t=3 hours post-onset of stimulation, sample 3 |
| GSM8307249 |
transduced with non-targeting siRNA, DMSO vehicle control, t=3 hours post-onset of stimulation, sample 4 |
| GSM8307250 |
transduced with non-targeting siRNA, supraphysiologic loading, t=3 hours post-onset of stimulation, sample 1 |
| GSM8307251 |
transduced with non-targeting siRNA, supraphysiologic loading, t=3 hours post-onset of stimulation, sample 2 |
| GSM8307252 |
transduced with non-targeting siRNA, supraphysiologic loading, t=3 hours post-onset of stimulation, sample 3 |
| GSM8307253 |
transduced with non-targeting siRNA, supraphysiologic loading, t=3 hours post-onset of stimulation, sample 4 |
| GSM8307254 |
transduced with non-targeting siRNA, Yoda1 treatment, t=3 hours post-onset of stimulation, sample 1 |
| GSM8307255 |
transduced with non-targeting siRNA, Yoda1 treatment, t=3 hours post-onset of stimulation, sample 2 |
| GSM8307256 |
transduced with non-targeting siRNA, Yoda1 treatment, t=3 hours post-onset of stimulation, sample 3 |
| GSM8307257 |
transduced with non-targeting siRNA, Yoda1 treatment, t=3 hours post-onset of stimulation, sample 4 |
| GSM8307258 |
transduced with PIEZO1-targeting siRNA, DMSO vehicle control, t=3 hours post-onset of stimulation, sample 1 |
| GSM8307259 |
transduced with PIEZO1-targeting siRNA, DMSO vehicle control, t=3 hours post-onset of stimulation, sample 2 |
| GSM8307260 |
transduced with PIEZO1-targeting siRNA, DMSO vehicle control, t=3 hours post-onset of stimulation, sample 3 |
| GSM8307261 |
transduced with PIEZO1-targeting siRNA, DMSO vehicle control, t=3 hours post-onset of stimulation, sample 4 |
| GSM8307262 |
transduced with PIEZO1-targeting siRNA, supraphysiologic loading, t=3 hours post-onset of stimulation, sample 1 |
| GSM8307263 |
transduced with PIEZO1-targeting siRNA, supraphysiologic loading, t=3 hours post-onset of stimulation, sample 2 |
| GSM8307264 |
transduced with PIEZO1-targeting siRNA, supraphysiologic loading, t=3 hours post-onset of stimulation, sample 3 |
| GSM8307265 |
transduced with PIEZO1-targeting siRNA, supraphysiologic loading, t=3 hours post-onset of stimulation, sample 4 |
| GSM8307266 |
transduced with PIEZO1-targeting siRNA, Yoda1 treatment, t=3 hours post-onset of stimulation, sample 1 |
| GSM8307267 |
transduced with PIEZO1-targeting siRNA, Yoda1 treatment, t=3 hours post-onset of stimulation, sample 2 |
| GSM8307268 |
transduced with PIEZO1-targeting siRNA, Yoda1 treatment, t=3 hours post-onset of stimulation, sample 3 |
| GSM8307269 |
transduced with PIEZO1-targeting siRNA, Yoda1 treatment, t=3 hours post-onset of stimulation, sample 4 |
| GSM8307270 |
GSK101 treatment, t=9 hours post-onset of stimulation, sample 1 |
| GSM8307271 |
GSK101 treatment, t=9 hours post-onset of stimulation, sample 2 |
| GSM8307272 |
GSK101 treatment, t=9 hours post-onset of stimulation, sample 3 |
| GSM8307273 |
GSK101 treatment, t=9 hours post-onset of stimulation, sample 4 |
| GSM8307274 |
GSK205 treatment, t=9 hours post-onset of stimulation, sample 1 |
| GSM8307275 |
GSK205 treatment, t=9 hours post-onset of stimulation, sample 2 |
| GSM8307276 |
GSK205 treatment, t=9 hours post-onset of stimulation, sample 3 |
| GSM8307277 |
GSK205 treatment, t=9 hours post-onset of stimulation, sample 4 |
| GSM8307278 |
DMSO vehicle control, t=9 hours post-onset of stimulation, sample 1 |
| GSM8307279 |
DMSO vehicle control, t=9 hours post-onset of stimulation, sample 2 |
| GSM8307280 |
DMSO vehicle control, t=9 hours post-onset of stimulation, sample 3 |
| GSM8307281 |
DMSO vehicle control, t=9 hours post-onset of stimulation, sample 4 |
| GSM8307282 |
IL-1 treatment, t=9hours post-onset of stimulation, sample 1 |
| GSM8307283 |
IL-1 treatment, t=9hours post-onset of stimulation, sample 2 |
| GSM8307284 |
IL-1 treatment, t=9hours post-onset of stimulation, sample 3 |
| GSM8307285 |
IL-1 treatment, t=9hours post-onset of stimulation, sample 4 |
| GSM8307286 |
Physiologic loading plus GSK205 treatment, t=9 hours post-onset of stimulation, sample 1 |
| GSM8307287 |
Physiologic loading plus GSK205 treatment, t=9 hours post-onset of stimulation, sample 2 |
| GSM8307288 |
Physiologic loading plus GSK205 treatment, t=9 hours post-onset of stimulation, sample 3 |
| GSM8307289 |
Physiologic loading plus GSK205 treatment, t=9 hours post-onset of stimulation, sample 4 |
| GSM8307290 |
Physiologic loading, t=9 hours post-onset of stimulation, sample 1 |
| GSM8307291 |
Physiologic loading, t=9 hours post-onset of stimulation, sample 2 |
| GSM8307292 |
Physiologic loading, t=9 hours post-onset of stimulation, sample 3 |
| GSM8307293 |
Physiologic loading, t=9 hours post-onset of stimulation, sample 4 |
| GSM8307294 |
non-transduced, DMSO vehicle control, t=9 hours post-onset of stimulation, sample 1 |
| GSM8307295 |
non-transduced, DMSO vehicle control, t=9 hours post-onset of stimulation, sample 2 |
| GSM8307296 |
non-transduced, DMSO vehicle control, t=9 hours post-onset of stimulation, sample 3 |
| GSM8307297 |
non-transduced, DMSO vehicle control, t=9 hours post-onset of stimulation, sample 4 |
| GSM8307298 |
non-transduced, supraphysiologic loading, t=9 hours post-onset of stimulation, sample 1 |
| GSM8307299 |
non-transduced, supraphysiologic loading, t=9 hours post-onset of stimulation, sample 2 |
| GSM8307300 |
non-transduced, supraphysiologic loading, t=9 hours post-onset of stimulation, sample 3 |
| GSM8307301 |
non-transduced, supraphysiologic loading, t=9 hours post-onset of stimulation, sample 4 |
| GSM8307302 |
non-transduced, Yoda1 treatment, t=9 hours post-onset of stimulation, sample 1 |
| GSM8307303 |
non-transduced, Yoda1 treatment, t=9 hours post-onset of stimulation, sample 2 |
| GSM8307304 |
non-transduced, Yoda1 treatment, t=9 hours post-onset of stimulation, sample 3 |
| GSM8307305 |
non-transduced, Yoda1 treatment, t=9 hours post-onset of stimulation, sample 4 |
| GSM8307306 |
transduced with non-targeting siRNA, DMSO vehicle control, t=9 hours post-onset of stimulation, sample 1 |
| GSM8307307 |
transduced with non-targeting siRNA, DMSO vehicle control, t=9 hours post-onset of stimulation, sample 2 |
| GSM8307308 |
transduced with non-targeting siRNA, DMSO vehicle control, t=9 hours post-onset of stimulation, sample 3 |
| GSM8307309 |
transduced with non-targeting siRNA, DMSO vehicle control, t=9 hours post-onset of stimulation, sample 4 |
| GSM8307310 |
transduced with non-targeting siRNA, supraphysiologic loading, t=9 hours post-onset of stimulation, sample 1 |
| GSM8307311 |
transduced with non-targeting siRNA, supraphysiologic loading, t=9 hours post-onset of stimulation, sample 2 |
| GSM8307312 |
transduced with non-targeting siRNA, supraphysiologic loading, t=9 hours post-onset of stimulation, sample 3 |
| GSM8307313 |
transduced with non-targeting siRNA, supraphysiologic loading, t=9 hours post-onset of stimulation, sample 4 |
| GSM8307314 |
transduced with non-targeting siRNA, Yoda1 treatment, t=9 hours post-onset of stimulation, sample 1 |
| GSM8307315 |
transduced with non-targeting siRNA, Yoda1 treatment, t=9 hours post-onset of stimulation, sample 2 |
| GSM8307316 |
transduced with non-targeting siRNA, Yoda1 treatment, t=9 hours post-onset of stimulation, sample 3 |
| GSM8307317 |
transduced with non-targeting siRNA, Yoda1 treatment, t=9 hours post-onset of stimulation, sample 4 |
| GSM8307318 |
transduced with PIEZO1-targeting siRNA, DMSO vehicle control, t=9 hours post-onset of stimulation, sample 1 |
| GSM8307319 |
transduced with PIEZO1-targeting siRNA, DMSO vehicle control, t=9 hours post-onset of stimulation, sample 2 |
| GSM8307320 |
transduced with PIEZO1-targeting siRNA, DMSO vehicle control, t=9 hours post-onset of stimulation, sample 3 |
| GSM8307321 |
transduced with PIEZO1-targeting siRNA, DMSO vehicle control, t=9 hours post-onset of stimulation, sample 4 |
| GSM8307322 |
transduced with PIEZO1-targeting siRNA, supraphysiologic loading, t=9 hours post-onset of stimulation, sample 1 |
| GSM8307323 |
transduced with PIEZO1-targeting siRNA, supraphysiologic loading, t=9 hours post-onset of stimulation, sample 2 |
| GSM8307324 |
transduced with PIEZO1-targeting siRNA, supraphysiologic loading, t=9 hours post-onset of stimulation, sample 3 |
| GSM8307325 |
transduced with PIEZO1-targeting siRNA, supraphysiologic loading, t=9 hours post-onset of stimulation, sample 4 |
| GSM8307326 |
transduced with PIEZO1-targeting siRNA, Yoda1 treatment, t=9 hours post-onset of stimulation, sample 1 |
| GSM8307327 |
transduced with PIEZO1-targeting siRNA, Yoda1 treatment, t=9 hours post-onset of stimulation, sample 2 |
| GSM8307328 |
transduced with PIEZO1-targeting siRNA, Yoda1 treatment, t=9 hours post-onset of stimulation, sample 3 |
| GSM8307329 |
transduced with PIEZO1-targeting siRNA, Yoda1 treatment, t=9 hours post-onset of stimulation, sample 4 |
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| Relations |
| BioProject |
PRJNA1120429 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE269159_PIEZO1_TRPV4_Mechanome_RNA_Seq_Gene_Counts.csv.csv.gz |
4.8 Mb |
(ftp)(http) |
CSV |
SRA Run Selector |
| Raw data are available in SRA |
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