Functional evidence for a role of GABA receptors in modulating nerve activities of circular smooth muscle from rat colon in vitro

Hyperplasia of smooth muscle contributes to the thickening of the intestinal wall that is characteristic of inflammation, but the mechanisms of growth control are unknown. Nitric oxide (NO) from enteric neurons expressing neuronal NO synthase (nNOS) might normally inhibit intestinal smooth muscle cell (ISMC) growth, and this was tested in vitro. In ISMC from the circular smooth muscle of the adult rat colon, chemical NO donors inhibited [3H]thymidine uptake in response to FCS, reducing this to baseline without toxicity. This effect was inhibited by the guanylyl cyclase inhibitor ODQ and potentiated by the phosphodiesterase-5 inhibitor zaprinast. Inhibition was mimicked by 8-bromo (8-Br)-cGMP, and ELISA measurements showed increased levels of cGMP but not cAMP in response to sodium nitroprusside. However, 8-Br-cAMP and cilostamide also showed inhibitory actions, suggesting an additional role for cAMP. Via a coculture model of ISMC and myenteric neurons, immunocytochemistry and image analysis showed that innervation reduced bromodeoxyuridine uptake by ISMC. Specific blockers of nNOS (7-NI, NAAN) significantly increased [3H]thymidine uptake in response to a standard stimulus, showing that nNOS activity normally inhibits ISMC growth. In vivo, nNOS axon number was reduced threefold by day 1 of trinitrobenzene sulfonic acid-induced rat colitis, preceding the hyperplasia of ISMC described earlier in this model. We conclude that NO can inhibit ISMC growth primarily via a cGMP-dependent mechanism. Functional evidence that NO derived from nNOS causes inhibition of ISMC growth in vitro predicts that the loss of nNOS expression in colitis contributes to ISMC hyperplasia in vivo.

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

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00290.2016 ◽

2016 ◽

Vol 311 (5) ◽

pp. G964-G973 ◽

Cited By ~ 12

Author(s):

Jagmohan Singh ◽

Ettickan Boopathi ◽

Sankar Addya ◽

Benjamin Phillips ◽

Isidore Rigoutsos ◽

...

Keyword(s):

Smooth Muscle ◽

Anal Sphincter ◽

Internal Anal Sphincter ◽

Smooth Muscles ◽

Smooth Muscle Contractility ◽

Network Analyses ◽

Rhoa Signaling

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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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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Abstract WMP31: Dedifferentiation of Smooth Muscle Cells and its Potential Contribution to Pathogenesis of Intracranial Aneurysms

Stroke ◽

10.1161/str.51.suppl_1.wmp31 ◽

2020 ◽

Vol 51 (Suppl_1) ◽

Author(s):

Mieko Oka ◽

Nobuhiko Ohno ◽

Takakazu Kawamata ◽

Tomohiro Aoki

Keyword(s):

Endothelial Cells ◽

Smooth Muscle ◽

Smooth Muscle Cells ◽

Inflammatory Responses ◽

Muscle Cells ◽

Inflammatory Factors ◽

Potential Contribution ◽

Electron Microscopic

Introduction: Intracranial aneurysm (IA) affects 1 to 5 % in general public and becomes the primary cause of subarachnoid hemorrhage, the most severe form of stroke. However, currently, no drug therapy is available for IAs to prevent progression and rupture of lesions. Elucidation of mechanisms underlying the disease is thus mandatory. Considering the important role of vascular smooth muscle cells (SMCs) in the maintenance of stiffness of arterial walls and also in the pathogenesis of atherosclerosis via mediating inflammatory responses, we in the present study analyzed morphological or phenotypical changes of SMCs during the disease development in the lesions. Methods: We subjected rats to an IA model in which lesions are induced by increase of hemodynamic force loading on intracranial arterial bifurcations and performed histopathological analyses of induced lesions including the electron microscopic examination. We then immunostained specimens from induced lesions to explore factors responsible for dedifferentiation or migration of SMCs. In vitro study was also done to examine effect of some candidate factors on dedifferentiation or migration of cultured SMCs. Results: We first found the accumulation of SMCs underneath the endothelial cell layer mainly at the neck portion of the lesion. These cells was positive for the embryonic form of myosin heavy chain, a marker for the dedifferentiated SMCs, and the expression of pro-inflammatory factors like TNF-α. In immunostaining to explore the potential factor regulating the dedifferentiation of SMCs, we found that Platelet-derived growth factor-BB (PDGF-BB) was expressed in endothelial cells at the neck portion of IA walls. Consistently, recombinant PDGF-BB could promote the dedifferentiate of SMCs and chemo-attracted them in in vitro. Finally, in the stenosis model of the carotid artery, PDGF-BB expression was induced in endothelial cells in which high wall shear stress was loaded and the dedifferentiation of SMCs occurred there. Conclusions: The findings from the present study imply the role of dedifferentiated SMCs partially recruited by PDGF-BB from endothelial cells in the formation of inflammatory microenvironment at the neck portion of IA walls, leading to the progression of the disease.

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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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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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Neural regulation of in vitro giant contractions in the rat colon

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.2001.281.1.g275 ◽

2001 ◽

Vol 281 (1) ◽

pp. G275-G282 ◽

Cited By ~ 27

Author(s):

Asensio Gonzalez ◽

Sushil K. Sarna

Keyword(s):

Smooth Muscle ◽

Smooth Muscle Cells ◽

Sch 23390 ◽

Circular Muscle ◽

Electrical Field Stimulation ◽

Muscle Cells ◽

Enteric Neurons ◽

Rat Colon ◽

Spontaneous Contractions

The rat middle colon spontaneously generates regularly occurring giant contractions (GCs) in vitro. We investigated the neurohumoral and intracellular regulation of these contractions in a standard muscle bath. cGMP content was measured in strips and single smooth muscle cells. The circular muscle strips generated spontaneous GCs. Their amplitude and frequency were significantly increased by tetrodotoxin (TTX), ω-conotoxin, N ω-nitro-l-arginine (l-NNA), and the dopamine D1 receptor antagonist Sch-23390. The GCs were unaffected by hexamethonium, atropine, and antagonists of serotonergic (5-HT1–4), histaminergic (H1–2), and tachykininergic (NK1–2) receptors but enhanced by NK3receptor antagonism. The guanylate cyclase inhibitor 1H-[1,2,4]oxidiazolo[4,3-a]quinoxalin-1-one (ODQ) also enhanced GCs to the same extent as TTX and l-NNA, and each of the three agents prevented the effects of the others. GCs were abolished by electrical field stimulation, S-nitroso- N-acetyl-penicillamine, and 8-bromo-cGMP. BAY-K-8644 and apamin enhanced the GCs, but they were abolished by D-600. Basal cGMP content in strips was decreased by TTX,l-NNA, or ODQ, but these treatments had no effect on cGMP content of enzymatically dissociated single smooth muscle cells. We conclude that spontaneous contractions in the rat colonic muscle strips are not generated by cholinergic, serotonergic, or histaminergic input. Constitutive release of nitric oxide from enteric neurons sustains cGMP synthesis in the colonic smooth muscle to suppress spontaneous in vitro GCs.

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