Aging-associated changes in microRNA expression profile of internal anal sphincter smooth muscle: Role of microRNA-133a

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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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 phospholipase A2 (group I secreted) in the genesis of basal tone in the internal anal sphincter smooth muscle

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00310.2007 ◽

2007 ◽

Vol 293 (5) ◽

pp. G979-G986 ◽

Cited By ~ 10

Author(s):

Márcio A. F. de Godoy ◽

Satish Rattan

Keyword(s):

Smooth Muscle ◽

Anal Sphincter ◽

Internal Anal Sphincter ◽

Critical Role ◽

Smooth Muscles ◽

Electrical Field Stimulation ◽

Group I ◽

Basal Tone ◽

Bromoenol Lactone

The role of phospholipase A2 (PLA2) in the genesis of basal tone in the internal anal sphincter (IAS) is not known. We determined the effects of PLA2 and inhibitors on the basal tone and intraluminal pressures (IASP) in the rat IAS vs. rectal smooth muscles (RSM). In addition, we determined the correlations between the IAS tone, PLA2 levels, and the actual enzymatic activity. Inhibition of PLA2 by 4-bromophenacyl bromide (universal inhibitor of PLA2) and MJ33 [selective inhibitor of secreted isoform of PLA2 (sPLA2)] caused concentration-dependent decrease in the IAS tone and in the IASP. Maximal decreases in the IAS tone and IASP by 4-bromophenacyl bromide and MJ33 were 58.8 ± 6.9 and 51.5 ± 6.3%, and 66.7 ± 5.1 and 79.8 ± 8.2%, respectively. The sPLA2 inhibitors were ∼100 times more potent in decreasing the IASP than the mean blood pressure. Conversely, the selective inhibitors of the cytosolic and calcium-independent PLA2 arachidonyl trifluoromethyl ketone and bromoenol lactone, respectively, produced no significant effect. The IAS had characteristically higher levels of sPLA2 activity (26.5 ± 4.9 μmol·min−1·ml−1) vs. the RSM (3.2 ± 0.4 μmol·min−1·ml−1), and higher levels of sPLA2 as shown by Western blot and RT-PCR. Interestingly, administration of sPLA2 transformed RSM into the tonic smooth muscle like that of the IAS: it developed basal tone and relaxed in response to the electrical field stimulation. From the present data, we conclude that sPLA2 plays a critical role in the genesis of tone in the IAS. PLA2 inhibitors may provide potential therapeutic target for treating anorectal motility disorders.

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Role of nitric oxide as a mediator of internal anal sphincter relaxation

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.1992.262.1.g107 ◽

1992 ◽

Vol 262 (1) ◽

pp. G107-G112 ◽

Cited By ~ 24

Author(s):

S. Rattan ◽

S. Chakder

Keyword(s):

Nitric Oxide ◽

Smooth Muscle ◽

Anal Sphincter ◽

Internal Anal Sphincter ◽

Inhibitory Action ◽

Dependent Manner ◽

Concentration Dependent Manner ◽

Inhibitory Neurotransmitter

The studies were performed in in vitro to examine the role of nitric oxide (NO) in nonadrenergic noncholinergic (NANC) nerve-mediated relaxation of the internal anal sphincter (IAS) smooth muscle strips of opossums. NO caused a concentration-dependent fall in the resting tension of the IAS. The inhibitory action of NO may be exerted directly on the IAS smooth muscle since it was not modified by the neurotoxin tetrodotoxin (1 x 10(-6) M), which abolished the neurally mediated fall in the IAS tension. The inhibitor of NO synthesis NG-nitro-L-arginine (L-NNA) produced concentration-dependent suppression of the neurally mediated fall in the IAS tension. The suppression of the neurally mediated IAS relaxation was stereoselective because D-NNA had no effect on the control responses. The suppressant action of L-NNA was selectively reversed by L-arginine in a concentration-dependent manner. The reversal was complete with 3 x 10(-4) M L-arginine. D-Arginine on the other hand, at the same concentration had no effect on L-NNA-suppressed IAS relaxation. Interestingly, the fall in the IAS tension caused by vasoactive intestinal polypeptide (VIP) (an inhibitory neurotransmitter in the IAS) was also inhibited by L-NNA (3 x 10(-5) M). From these data we conclude that NO or NO-like substances serve as important inhibitory mediators for the NANC nerve-mediated IAS relaxation. A part of the inhibitory action of VIP on the IAS involves NO-synthase pathway. The exact site of formation and release of NO or NO-like substances in response to NANC nerve stimulation remain to be investigated.

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S29-7 THE ROLE OF S-PHASE KINASE-ASSOCIATED PROTEIN-2 (SKP2) IN THE REGULATION OF VASCULAR SMOOTH MUSCLE CELL MIGRATION AND APOPTOSIS IN VITRO AND NEOINTIMAL THICKENING IN VIVO

International Journal of Cardiology ◽

10.1016/s0167-5273(08)70479-0 ◽

2007 ◽

Vol 122 ◽

pp. S48 ◽

Cited By ~ 1

Author(s):

Yih-Jer Wu ◽

Andrew C. Newby ◽

Graciela B. Sala-Newby ◽

Kuo-Tung Hsu ◽

Sywoei Tseng ◽

...

Keyword(s):

Smooth Muscle ◽

Cell Migration ◽

Vascular Smooth Muscle ◽

Smooth Muscle Cell ◽

S Phase ◽

Neointimal Thickening ◽

Smooth Muscle Cell Migration

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Heparan sulfate side chains have a critical role in the inhibitory effects of perlecan on vascular smooth muscle cell response to arterial injury

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00654.2013 ◽

2014 ◽

Vol 307 (3) ◽

pp. H337-H345 ◽

Cited By ~ 6

Author(s):

Lara Gotha ◽

Sang Yup Lim ◽

Azriel B. Osherov ◽

Rafael Wolff ◽

Beiping Qiang ◽

...

Keyword(s):

Smooth Muscle ◽

Critical Role ◽

Arterial Injury ◽

Side Chains ◽

Wild Type ◽

Injury Response ◽

Migratory Response

Perlecan is a proteoglycan composed of a 470-kDa core protein linked to three heparan sulfate (HS) glycosaminoglycan chains. The intact proteoglycan inhibits the smooth muscle cell (SMC) response to vascular injury. Hspg2Δ3/Δ3 (MΔ3/Δ3) mice produce a mutant perlecan lacking the HS side chains. The objective of this study was to determine differences between these two types of perlecan in modifying SMC activities to the arterial injury response, in order to define the specific role of the HS side chains. In vitro proliferative and migratory activities were compared in SMC isolated from MΔ3/Δ3 and wild-type mice. Proliferation of MΔ3/Δ3 SMC was 1.5× greater than in wild type ( P < 0.001), increased by addition of growth factors, and showed a 42% greater migratory response than wild-type cells to PDGF-BB ( P < 0.001). In MΔ3/Δ3 SMC adhesion to fibronectin, and collagen types I and IV was significantly greater than wild type. Addition of DRL-12582, an inducer of perlecan expression, decreased proliferation and migratory response to PDGF-BB stimulation in wild-type SMC compared with MΔ3/Δ3. In an in vivo carotid artery wire injury model, the medial thickness, medial area/lumen ratio, and macrophage infiltration were significantly increased in the MΔ3/Δ3 mice, indicating a prominent role of the HS side chain in limiting vascular injury response. Mutant perlecan that lacks HS side chains had a marked reduction in the inhibition of in vitro SMC function and the in vivo arterial response to injury, indicating the critical role of HS side chains in perlecan function in the vessel wall.

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Role of endothelial cells in regulating hemoglobin-induced changes in lymphatic pumping

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.1992.263.6.h1880 ◽

1992 ◽

Vol 263 (6) ◽

pp. H1880-H1887 ◽

Cited By ~ 1

Author(s):

R. M. Elias ◽

J. Eisenhoffer ◽

M. G. Johnston

Keyword(s):

Endothelial Cells ◽

Smooth Muscle ◽

Inhibitory Effect ◽

Direct Inhibitory Effect ◽

Induced Changes ◽

Lymphatic Pumping ◽

Pumping Activity

Studies with a sheep isolated duct preparation in vivo demonstrated that the route of administration of hemoglobin was important in demonstrating its inhibitory effect on lymphatic pumping. With autologous oxyhemoglobin administered intravenously (final plasma concentration 5 x 10(-5) M), pumping was not inhibited. However, the addition of oxyhemoglobin (5 x 10(-5) M) into the reservoir (lumen of the duct) resulted in > 95% inhibition of pumping. The extraluminal administration of oxyhemoglobin (10(-5) M) to bovine mesenteric lymphatics in vitro resulted in a 40% inhibition of pumping, whereas the introduction of oxyhemoglobin (10(-5) M) into the lumen of the vessels suppressed pumping 95%. In vessels mechanically denuded of endothelium, intraluminal oxyhemoglobin inhibited pumping 50%. These results suggested that oxyhemoglobin depressed pumping through an effect on both smooth muscle and endothelium. Once pumping was inhibited with oxyhemoglobin administration, stimulation of the duct with elevations in transmural pressure restored pumping activity when endothelial cells were present. However, in the absence of endothelium, pumping decreased with increases in distending pressures. We conclude that oxyhemoglobin has a direct inhibitory effect on lymphatic smooth muscle. The ability of oxyhemoglobin to alter the pressure range over which the lymph pump operates appears to be dependent on an intact endothelium.

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Decreased airway narrowing and smooth muscle contraction in hyperresponsive pigs

Journal of Applied Physiology ◽

10.1152/japplphysiol.00150.2002 ◽

2002 ◽

Vol 93 (4) ◽

pp. 1296-1300 ◽

Cited By ~ 12

Author(s):

Debra J. Turner ◽

Peter B. Noble ◽

Matthew P. Lucas ◽

Howard W. Mitchell

Keyword(s):

Smooth Muscle ◽

Airway Hyperresponsiveness ◽

Porcine Model ◽

Smooth Muscle Contraction ◽

Airway Wall ◽

Muscle Contractility ◽

Smooth Muscle Contractility ◽

Airway Narrowing

Increased smooth muscle contractility or reduced smooth muscle mechanical loads could account for the excessive airway narrowing and hyperresponsiveness seen in asthma. These mechanisms were investigated by using an allergen-induced porcine model of airway hyperresponsiveness. Airway narrowing to electric field stimulation was measured in isolated bronchial segments, over a range of transmural pressures (0–20 cmH2O). Contractile responses to ACh were measured in bronchial segments and in isolated tracheal smooth muscle strips isolated from control and test (ovalbumin sensitized and challenged) pigs. Test airways narrowed less than controls ( P < 0.0001). Test pigs showed reduced contractility to ACh, both in isolated bronchi ( P < 0.01) and smooth muscle strips ( P < 0.01). Thus isolated airways from pigs exhibiting airway hyperresponsiveness in vivo are hyporesponsive in vitro. The decreased narrowing in bronchi from hyperresponsive pigs may be related to decreased smooth muscle contractility. These data suggest that mechanisms external to the airway wall may be important to the hyperresponsive nature of sensitized lungs.

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Release of epithelium-derived PGE2 from canine trachea after antigen inhalation

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.1998.274.2.l220 ◽

1998 ◽

Vol 274 (2) ◽

pp. L220-L225 ◽

Cited By ~ 4

Author(s):

I. McGrogan ◽

L. J. Janssen ◽

J. Wattie ◽

P. M. O’Byrne ◽

E. E. Daniel

Keyword(s):

Smooth Muscle ◽

Electrical Field Stimulation ◽

Tracheal Smooth Muscle ◽

Organ Bath ◽

Field Stimulation ◽

Electrical Field ◽

Concentration Response Curve

To investigate the role of prostaglandin (PG) E2 in allergen-induced hyperresponsiveness, dogs inhaled either the allergen Ascaris suum or vehicle (Sham). Twenty-four hours after inhalation, some animals exposed to allergen demonstrated an increased responsiveness to acetylcholine challenge in vivo (Hyp-Resp), whereas others did not (Non-Resp). Strips of tracheal smooth muscle, either epithelium intact or epithelium denuded, were suspended on stimulating electrodes, and a concentration-response curve to carbachol (10−9 to 10−5 M) was generated. Tissues received electrical field stimulation, and organ bath fluid was collected to determine PGE2content. With the epithelium present, all three groups contracted similarly to 10−5 M carbachol, whereas epithelium-denuded tissues from animals that inhaled allergen contracted more than tissues from Sham dogs. In response to electrical field stimulation, Hyp-Resp tissues contracted less than Sham tissues in the presence of epithelium and more than Sham tissues in the absence of epithelium. PGE2release in the muscle bath was greater in Non-Resp tissues than in Sham or Hyp-Resp tissues when the epithelium was present. Removal of the epithelium greatly inhibited PGE2release. We conclude that tracheal smooth muscle is hyperresponsive in vitro after in vivo allergen exposure only when the modulatory effect of the epithelium, largely through PGE2 release, is removed.

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Activation of heme oxygenase-1 by Ginkgo biloba extract differentially modulates endothelial and smooth muscle-like progenitor cells for vascular repair

Scientific Reports ◽

10.1038/s41598-019-53818-7 ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 1

Author(s):

Tao-Cheng Wu ◽

Jia-Shiong Chen ◽

Chao-Hung Wang ◽

Po-Hsun Huang ◽

Feng-Yen Lin ◽

...

Keyword(s):

Bone Marrow ◽

Smooth Muscle ◽

Progenitor Cells ◽

Heme Oxygenase ◽

Neointimal Hyperplasia ◽

Vascular Repair ◽

Vascular Progenitor Cells

AbstractVascular progenitors such as endothelial progenitor cells (EPCs) and smooth muscle-like progenitor cells (SMPCs) may play different roles in vascular repair. Ginkgo biloba extract (GBE) is an exogenous activator of heme oxygenase (HO)-1, which has been suggested to improve vascular repair; however, the detailed mechanisms have yet to be elucidated. This study aimed to investigate whether GBE can modulate different vascular progenitor cells by activating HO-1 for vascular repair. A bone marrow transplantation mouse model was used to evaluate the in vivo effects of GBE treatment on wire-injury induced neointimal hyperplasia, which is representative of impaired vascular repair. On day 14 of GBE treatment, the mice were subjected to wire injury of the femoral artery to identify vascular reendothelialization. Compared to the mice without treatment, neointimal hyperplasia was reduced in the mice that received GBE treatment for 28 days in a dose-dependent manner. Furthermore, GBE treatment increased bone marrow-derived EPCs, accelerated endothelial recovery, and reduced the number of SMPCs attached to vascular injury sites. The effects of GBE treatment on neointimal hyperplasia could be abolished by co-treatment with zinc protoporphyrin IX, an HO-1 inhibitor, suggesting the in vivo role of HO-1. In this in vitro study, treatment with GBE activated human early and late EPCs and suppressed SMPC migration. These effects were abolished by HO-1 siRNA and an HO-1 inhibitor. Furthermore, GBE induced the expression of HO-1 by activating PI3K/Akt/eNOS signaling in human late EPCs and via p38 pathways in SMPCs, suggesting that GBE can induce HO-1 in vitro through different molecular mechanisms in different vascular progenitor cells. Accordingly, GBE could activate early and late EPCs, suppress the migration of SMPCs, and improve in vivo vascular repair after mechanical injury by activating HO-1, suggesting the potential role of pharmacological HO-1 activators, such as GBE, for vascular protection in atherosclerotic diseases.

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