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| Status |
Public on Oct 28, 2016 |
| Title |
Title: Aging-associated changes in microRNA expression profile of internal anal sphincter smooth muscle: Role of microRNA-133a |
| Platform organisms |
Rattus norvegicus; synthetic construct |
| Sample organism |
Rattus norvegicus |
| Experiment type |
Expression profiling by array
Non-coding RNA profiling by array
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| Summary |
A comprehensive –omic, computational, and physiological approach was employed to examine the (previously unexplored) role of microRNAs (miRNAs) as regulators of IAS smooth muscle contractile phenotype and basal tone. MicroRNA profiling, genome wide expression, validation and network analyses were employed to assess changes in mRNA and miRNA expression in IAS smooth muscles from young vs. aging rats. Multiple miRNAs, including rno-miR-1, rno-miR-340-5p, rno-miR-185, rno-miR-199a-3p, rno-miR-200c, rno-miR-200b, rno-miR-31, rno-miR-133a and rno-miR-206 were found to be up-regulated in aging IAS. qRT-PCR confirmed the up-regulated expression of these miRNAs and down regulation of multiple, predicted targets (Eln, Col3a1, Col1a1, Zeb2, Myocd, SRF, Smad1, Smad2, RhoA/ROCK2, Fn1, Sm22-v2, Klf4, and Acta2) involved in regulation of SM contractility. Subsequent studies demonstrated an aging-associated increase in the expression of miR-133a, corresponding decreases in RhoA, ROCK2, MYOCD, SRF and SM22α protein expression, RhoA-signaling, and a decrease in basal and agonist (U-46619 (thromboxane A2 analog))-induced increase in the IAS tone. Moreover, in vitro transfection of miR-133a caused a dose-dependent increase of IAS tone in strips, which was reversed by anti-miR-133a. Lastly, in vivo perianal injection of anti-miR-133a reversed the loss of IAS tone associated with age. This work establishes the important regulatory effect of miRNA-133a on basal and agonist-stimulated IAS tone. Moreover, reversal of age-associated loss of tone via anti-miR delivery strongly implicates miR dysregulation as a causal factor in the aging-associated decrease in IAS tone, and suggests miR-133a is feasible therapeutic target in aging-associated rectoanal incontinence.
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| Overall design |
Three rats (Fischer 344) provided by the National Institutes of Aging of three different age groups (6 months, 18 months, and 26 months of age) were used for these studies. The smooth muscle cells (SMCs) from the internal anal sphincter (IAS) from above different age groups were isolated using collagenase dispersion method. The cells were subjected to RNA collection; and RNA from these samples were used for mRNA microarray analysis. For another sets of studies, from another batch of Fischer 344 rats (of 6 months, 18 months, and 26 months of age), the IAS SMCs were isolated and their miRNA fractions using miRVana miRNA Isolation kit following the manufacturer’s protocol were collected for miRNA microarray analysis.
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| Contributor(s) |
Singh J, Boopathi E, Addya S, Rigoutsos I, B.Penn R, Rattan S |
| Citation(s) |
27634012 |
| Submission date |
Mar 17, 2016 |
| Last update date |
May 02, 2017 |
| Contact name |
Satish Rattan |
| E-mail(s) |
satish.rattan@jefferson.edu
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| Phone |
215-955-5614
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| Organization name |
Thomas Jefferson University
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| Department |
Medicine
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| Lab |
GI
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| Street address |
1025 Walnut St., Room 320 College
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| City |
Philadelphia |
| State/province |
PA |
| ZIP/Postal code |
19107 |
| Country |
USA |
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| Platforms (2) |
| GPL6247 |
[RaGene-1_0-st] Affymetrix Rat Gene 1.0 ST Array [transcript (gene) version] |
| GPL14613 |
[miRNA-2] Affymetrix Multispecies miRNA-2 Array |
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| Samples (6)
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| Relations |
| BioProject |
PRJNA315564 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE79350_RAW.tar |
11.2 Mb |
(http)(custom) |
TAR (of CEL) |
| Processed data included within Sample table |
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