A comparison of the contractile properties of smooth muscle from pig urethra and internal anal sphincter

A comprehensive genomic and proteomic, computational, and physiological approach was employed to examine the (previously unexplored) role of microRNAs (miRNAs) as regulators of internal anal sphincter (IAS) smooth muscle contractile phenotype and basal tone. miRNA 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 upregulated in aging IAS. qPCR confirmed the upregulated expression of these miRNAs and downregulation of multiple, predicted targets ( Eln, Col3a1, Col1a1, Zeb2, Myocd, Srf, Smad1, Smad2, Rhoa/Rock2, Fn1, Tagln v2, Klf4, and Acta2) involved in regulation of smooth muscle 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 A2analog)]-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. Last, 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 that miR-133a is a feasible therapeutic target in aging-associated rectoanal incontinence.

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Use of Smooth Muscle Plasty After Intersphincteric Rectal Resection to Replace a Partially Resected Internal Anal Sphincter: Long-Term Follow-Up

Diseases of the Colon & Rectum ◽

10.1007/dcr.0b013e3181b55507 ◽

2009 ◽

Vol 52 (11) ◽

pp. 1895-1901 ◽

Cited By ~ 19

Author(s):

Osman Krand ◽

Tunç Yalti ◽

Gurkan Tellioglu ◽

Melih Kara ◽

Ibrahim Berber ◽

...

Keyword(s):

Smooth Muscle ◽

Anal Sphincter ◽

Internal Anal Sphincter ◽

Rectal Resection ◽

Long Term Follow Up

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CGRP and ANF cause relaxation of opossum internal anal sphincter via different mechanisms

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.1991.260.5.g764 ◽

1991 ◽

Vol 260 (5) ◽

pp. G764-G769 ◽

Cited By ~ 4

Author(s):

S. Rattan ◽

C. Moummi ◽

S. Chakder

Keyword(s):

Smooth Muscle ◽

Anal Sphincter ◽

Cyclic Nucleotides ◽

Internal Anal Sphincter ◽

Camp Content ◽

Calcitonin Gene ◽

Before And After ◽

Camp And Cgmp

This investigation examined and compared the role of cyclic nucleotides in the mediation of internal anal sphincter (IAS) relaxation caused by the addition of neuropeptide calcitonin gene-related peptide (CGRP) and atrial natriuretic factor (ANF). The studies were performed in vitro on smooth muscle strips of opossum IAS. The relaxation produced by CGRP and ANF was examined before and after the addition of tetrodotoxin (TTX) (1 x 10(-6)M). At this concentration, TTX did not have any significant effect on the relaxation produced by either CGRP or ANF, suggesting that these peptides act directly on the smooth muscle. Addition of CGRP (3 x 10(-6) M) produced the maximal relaxation and significantly increased cAMP content without changing cGMP. On the other hand, addition of ANF (3 x 10(-6) M) caused a similar fall in IAS tension that was accompanied by a significant elevation in cGMP without any change in cAMP content. The rises in the levels of cyclic nucleotides preceded the onset of fall in the resting tension of IAS. Our results demonstrate that CGRP and ANF relax isolated strips of opossum IAS by their action directly at the smooth muscle and that this relaxation is associated with an increase in cAMP and cGMP, respectively. The studies suggest the presence of both cAMP and cGMP pathways in the IAS and that the relaxation of IAS smooth muscle in response to different peptides may occur via a specific intracellular biochemical pathway.

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Role of differentially expressed microRNA-139-5p in the regulation of phenotypic internal anal sphincter smooth muscle tone

Scientific Reports ◽

10.1038/s41598-017-01550-5 ◽

2017 ◽

Vol 7 (1) ◽

Cited By ~ 3

Author(s):

Jagmohan Singh ◽

Ipsita Mohanty ◽

Sankar Addya ◽

Benjamin Phillips ◽

Hwan Mee Yong ◽

...

Keyword(s):

Smooth Muscle ◽

Anal Sphincter ◽

Internal Anal Sphincter ◽

Muscle Tone ◽

Differentially Expressed ◽

Smooth Muscle Tone

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Mechanism of internal anal sphincter smooth muscle relaxation by phorbol 12,13-dibutyrate

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.2001.280.6.g1341 ◽

2001 ◽

Vol 280 (6) ◽

pp. G1341-G1350 ◽

Cited By ~ 10

Author(s):

Sushanta Chakder ◽

D. N. K. Sarma ◽

Satish Rattan

Keyword(s):

Smooth Muscle ◽

Smooth Muscle Cells ◽

Anal Sphincter ◽

Internal Anal Sphincter ◽

Inhibitory Action ◽

Muscle Tone ◽

Muscle Relaxation ◽

Muscle Cells ◽

Smooth Muscle Relaxation ◽

Smooth Muscle Tone

We investigated the mechanism of the inhibitory action of phorbol 12,13-dibutyrate (PDBu), one of the typical protein kinase C (PKC) activators, in in vitro smooth muscle strips and in isolated smooth muscle cells of the opossum internal anal sphincter (IAS). The inhibitory action of PDBu on IAS smooth muscle (observed in the presence of guanethidine + atropine) was partly attenuated by tetrodotoxin, suggesting that a part of the inhibitory action of PDBu is via the nonadrenergic, noncholinergic neurons. A major part of the action of PDBu in IAS smooth muscle was, however, via its direct action at the smooth muscle cells, accompanied by a decrease in free intracellular Ca2+ concentration ([Ca2+]i) and inhibition of PKC translocation. PDBu-induced IAS smooth muscle relaxation was unaffected by agents that block Ca2+ mobilization and Na+-K+-ATPase. The PDBu-induced fall in basal IAS smooth muscle tone and [Ca2+]i resembled that induced by the Ca2+ channel blocker nifedipine and were reversed specifically by the Ca2+ channel activator BAY K 8644. We speculate that a major component of the relaxant action of PDBu in IAS smooth muscle is caused by the inhibition of Ca2+ influx and of PKC translocation to the membrane. The specific role of PKC downregulation and other factors in the phorbol ester-mediated fall in basal IAS smooth muscle tone remain to be determined.

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Autocrine regulation of internal anal sphincter tone by renin-angiotensin system: comparison with phasic smooth muscle

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00115.2005 ◽

2005 ◽

Vol 289 (6) ◽

pp. G1164-G1175 ◽

Cited By ~ 22

Author(s):

Márcio A. F. De Godoy ◽

Satish Rattan

Keyword(s):

Smooth Muscle ◽

Anal Sphincter ◽

Internal Anal Sphincter ◽

Renin Angiotensin System ◽

Control Mechanisms ◽

Ang Ii ◽

Angiotensin System ◽

Renin Angiotensin ◽

Autocrine Regulation ◽

Basal Tone

The myogenic control mechanisms that govern the basal tone in the internal anal sphincter (IAS) are not known. The present studies determined the autocrine regulation of ANG II in the IAS. The studies were performed in the freshly isolated smooth muscle cells (SMC) of the IAS. We determined the presence of ANG II precursor angiotensinogen (Angen), and the enzymes that convert it into ANG II, using functional, molecular biology, and immunocytochemical studies in rats. ANG II levels in the SMC were determined using ELISA. The IAS SMC generate ANG II at a rate severalfold higher than those from the adjoining smooth muscle of rectum (RSM). RT-PCR data show that IAS exclusively expresses significant higher levels of renin, Angen, and angiotensin-converting enzyme (ACE). These data were confirmed using Western blot analyses and immunocytochemistry. In the IAS SMC, H-77 (10 μM; renin inhibitor) and captopril (1 μM; ACE inhibitor) decreased the basal as well as Angen-increased levels of ANG II. The following functional data corroborate the role of renin-angiotensin system (RAS) in the IAS tone. Angen produced concentration-dependent shortening of the IAS SMC that was inhibited by H-77 and captopril. In addition, H-77 or captopril caused a concentration-dependent fall in the IAS tone vs. nontonic tissues. Basal tone in IAS is partially under the autocrine control of cellular RAS evident by the expression of mRNA coding Angen, renin, and ACE and translation to the respective proteins in the SMC.

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Successful implantation of physiologically functional bioengineered mouse internal anal sphincter

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00269.2009 ◽

2010 ◽

Vol 299 (2) ◽

pp. G430-G439 ◽

Cited By ~ 20

Author(s):

Shreya Raghavan ◽

Eiichi A. Miyasaka ◽

Mohamed Hashish ◽

Sita Somara ◽

Robert R. Gilmont ◽

...

Keyword(s):

Smooth Muscle ◽

Smooth Muscle Cells ◽

Anal Sphincter ◽

Intracellular Signaling ◽

Internal Anal Sphincter ◽

Muscle Cells ◽

Nitric Oxide Donor ◽

Proof Of Concept ◽

Smooth Muscle Tissue ◽

Basal Tone

We have previously developed bioengineered three-dimensional internal anal sphincter (IAS) rings from circular smooth muscle cells isolated from rabbit and human IAS. We provide proof of concept that bioengineered mouse IAS rings are neovascularized upon implantation into mice of the same strain and maintain concentric smooth muscle alignment, phenotype, and IAS functionality. Rings were bioengineered by using smooth muscle cells from the IAS of C57BL/6J mice. Bioengineered mouse IAS rings were implanted subcutaneously on the dorsum of C57BL/6J mice along with a microosmotic pump delivering fibroblast growth factor-2. The mice remained healthy during the period of implantation, showing no external signs of rejection. Mice were killed 28 days postsurgery and implanted IAS rings were harvested. IAS rings showed muscle attachment, neovascularization, healthy color, and no external signs of infection or inflammation. Assessment of force generation on harvested IAS rings showed the following: 1) spontaneous basal tone was generated in the absence of external stimulation; 2) basal tone was relaxed by vasoactive intestinal peptide, nitric oxide donor, and nifedipine; 3) acetylcholine and phorbol dibutyrate elicited rapid-rising, dose-dependent, sustained contractions repeatedly over 30 min without signs of muscle fatigue; and 4) magnitudes of potassium chloride-induced contractions were 100% of peak maximal agonist-induced contractions. Our preliminary results confirm the proof of concept that bioengineered rings are neovascularized upon implantation. Harvested rings maintain smooth muscle alignment and phenotype. Our physiological studies confirm that implanted rings maintain 1) overall IAS physiology and develop basal tone, 2) integrity of membrane ionic characteristics, and 3) integrity of membrane associated intracellular signaling transduction pathways for contraction and relaxation by responding to cholinergic, nitrergic, and VIP-ergic stimulation. IAS smooth muscle tissue could thus be bioengineered for the purpose of implantation to serve as a potential graft therapy for dysfunctional internal anal sphincter in fecal incontinence.

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Sa1349 Bimodal Effect of Oxidative Stress in Internal Anal Sphincter (IAS) Smooth Muscle

Gastroenterology ◽

10.1016/s0016-5085(15)30989-6 ◽

2015 ◽

Vol 148 (4) ◽

pp. S-299

Author(s):

Sumit Kumar ◽

Jagmohan Singh ◽

Satish C. Rattan

Keyword(s):

Oxidative Stress ◽

Smooth Muscle ◽

Anal Sphincter ◽

Internal Anal Sphincter

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