Cultured Nestin-Positive Cells from Postnatal Mouse Small Bowel Differentiate Ex Vivo into Neurons, Glia, and Smooth Muscle

AbstractThe cervix is responsible for maintaining pregnancy, and its timely remodeling is essential for the proper delivery of a baby. Cervical insufficiency, or “weakness”, may lead to preterm birth, which causes infant morbidities and mortalities worldwide. We used a mouse model of pregnancy and term labor, to examine the cervical structure by histology (Masson Trichome and Picrosirius Red staining), immunohistochemistry (Hyaluronic Acid Binding Protein/HABP), and ex-vivo MRI (T2-weighted and diffusion tensor imaging), focusing on two regions of the cervix (i.e., endocervix and ectocervix). Our results show that mouse endocervix has a higher proportion of smooth muscle cells and collagen fibers per area, with more compact tissue structure, than the ectocervix. With advanced gestation, endocervical changes, indicative of impending delivery, are manifested in fewer smooth muscle cells, expansion of the extracellular space, and lower presence of collagen fibers. MRI detected three distinctive zones in pregnant mouse endocervix: (1) inner collagenous layer, (2) middle circular muscular layer, and (3) outer longitudinal muscular layer. Diffusion MRI images detected changes in tissue organization as gestation progressed suggesting the potential application of this technique to non-invasively monitor cervical changes that precede the onset of labor in women at risk for preterm delivery.

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NTPDase1 Modulates Smooth Muscle Contraction in Mice Bladder by Regulating Nucleotide Receptor Activation Distinctly in Male and Female

Biomolecules ◽

10.3390/biom11020147 ◽

2021 ◽

Vol 11 (2) ◽

pp. 147

Author(s):

Romuald Brice Babou Kammoe ◽

Gilles Kauffenstein ◽

Julie Pelletier ◽

Bernard Robaye ◽

Jean Sévigny

Keyword(s):

Smooth Muscle ◽

Ex Vivo ◽

Smooth Muscle Contraction ◽

Receptor Activation ◽

Nucleoside Triphosphate ◽

Nucleotide Receptors ◽

Nerve Terminals ◽

Wild Type ◽

Nucleoside Triphosphate Diphosphohydrolase ◽

Bladder Contraction

Nucleotides released by smooth muscle cells (SMCs) and by innervating nerve terminals activate specific P2 receptors and modulate bladder contraction. We hypothesized that cell surface enzymes regulate SMC contraction in mice bladder by controlling the concentration of nucleotides. We showed by immunohistochemistry, enzymatic histochemistry, and biochemical activities that nucleoside triphosphate diphosphohydrolase-1 (NTPDase1) and ecto-5′-nucleotidase were the major ectonucleotidases expressed by SMCs in the bladder. RT-qPCR revealed that, among the nucleotide receptors, there was higher expression of P2X1, P2Y1, and P2Y6 receptors. Ex vivo, nucleotides induced a more potent contraction of bladder strips isolated from NTPDase1 deficient (Entpd1−/−) mice compared to wild type controls. The strongest responses were obtained with uridine 5′-triphosphate (UTP) and uridine 5′-diphosphate (UDP), suggesting the involvement of P2Y6 receptors, which was confirmed with P2ry6−/− bladder strips. Interestingly, this response was reduced in female bladders. Our results also suggest the participation of P2X1, P2Y2 and/or P2Y4, and P2Y12 in these contractions. A reduced response to the thromboxane analogue U46619 was also observed in wild type, Entpd1−/−, and P2ry6−/− female bladders showing another difference due to sex. In summary, NTPDase1 modulates the activation of nucleotide receptors in mouse bladder SMCs, and contractions induced by P2Y6 receptor activation were weaker in female bladders.

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Reduced biomechanical models for precision-cut lung-slice stretching experiments

Journal of Mathematical Biology ◽

10.1007/s00285-021-01578-2 ◽

2021 ◽

Vol 82 (5) ◽

Author(s):

Hannah J. Pybus ◽

Amanda L. Tatler ◽

Lowell T. Edgar ◽

Reuben D. O’Dea ◽

Bindi S. Brook

Keyword(s):

Smooth Muscle ◽

Muscle Contraction ◽

Ex Vivo ◽

Finite Thickness ◽

Biomechanical Model ◽

Local Stress ◽

Smooth Muscle Contraction ◽

Biomechanical Models ◽

Cytokine Activation ◽

Asymptotic Reduction

AbstractPrecision-cut lung-slices (PCLS), in which viable airways embedded within lung parenchyma are stretched or induced to contract, are a widely used ex vivo assay to investigate bronchoconstriction and, more recently, mechanical activation of pro-remodelling cytokines in asthmatic airways. We develop a nonlinear fibre-reinforced biomechanical model accounting for smooth muscle contraction and extracellular matrix strain-stiffening. Through numerical simulation, we describe the stresses and contractile responses of an airway within a PCLS of finite thickness, exposing the importance of smooth muscle contraction on the local stress state within the airway. We then consider two simplifying limits of the model (a membrane representation and an asymptotic reduction in the thin-PCLS-limit), that permit analytical progress. Comparison against numerical solution of the full problem shows that the asymptotic reduction successfully captures the key elements of the full model behaviour. The more tractable reduced model that we develop is suitable to be employed in investigations to elucidate the time-dependent feedback mechanisms linking airway mechanics and cytokine activation in asthma.

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Tissue stress from laparoscopic grasper use and bowel injury in humans: establishing intraoperative force boundaries

BMJ Surgery, Interventions, & Health Technologies ◽

10.1136/bmjsit-2021-000084 ◽

2021 ◽

Vol 3 (1) ◽

pp. e000084

Author(s):

Amanda Farah Khan ◽

Matthew Kenneth MacDonald ◽

Catherine Streutker ◽

Corwyn Rowsell ◽

James Drake ◽

...

Keyword(s):

Small Bowel ◽

Tissue Injury ◽

Ex Vivo ◽

Gastrointestinal Surgery ◽

Convenience Sample ◽

Thickness Change ◽

Novel Device ◽

Significant Difference ◽

Laparoscopic Grasper

ObjectivesWe aim to determine what threshold of compressive stress small bowel and colon tissues display evidence of significant tissue trauma during laparoscopic surgery.DesignThis study included 10 small bowel and 10 colon samples from patients undergoing routine gastrointestinal surgery. Each sample was compressed with pressures ranging from 100 kPa to 600 kPa. Two pathologists who were blinded to all study conditions, performed a histological analysis of the tissues. Experimentation: November 2018–February 2019. Analysis: March 2019–May 2020.SettingAn inner-city trauma and ambulatory hospital with a 40-bed inpatient general surgery unit with a diverse patient population.ParticipantsPatients were eligible if their surgery procured healthy tissue margins for experimentation (a convenience sample). 26 patient samples were procured; 6 samples were unusable. 10 colon and 10 small bowel samples were tested for a total of 120 experimental cases. No patients withdrew their consent.InterventionsA novel device was created to induce compressive “grasps” to simulate those of a laparoscopic grasper. Experimentation was performed ex-vivo, in-vitro. Grasp conditions of 0–600 kPa for a duration of 10 s were used.ResultsSmall bowel (10), M:F was 7:3, average age was 54.3 years. Colon (10), M:F was 1:1, average age was 65.2 years. All 20 patients experienced a significant difference (p<0.05) in serosal thickness post-compression at both 500 and 600 kPa for both tissue types. A logistic regression analysis with a sensitivity of 100% and a specificity of 84.6% on a test set of data predicts a safety threshold of 329–330 kPa.ConclusionsA threshold was discovered that corresponded to both significant serosal thickness change and a positive histological trauma score rating. This “force limit” could be used in novel sensorized laparoscopic tools to avoid intraoperative tissue injury.

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Myorelaxant Effect of the Dysphania ambrosioides Essential Oil on Sus scrofa domesticus Coronary Artery and Its Toxicity in the Drosophila melanogaster Model

Molecules ◽

10.3390/molecules26072041 ◽

2021 ◽

Vol 26 (7) ◽

pp. 2041

Author(s):

Luiz Jardelino de Lacerda Neto ◽

Andreza Guedes Barbosa Ramos ◽

Renata Evaristo Rodrigues da Silva ◽

Luís Pereira-de-Morais ◽

Fernanda Maria Silva ◽

...

Keyword(s):

Drosophila Melanogaster ◽

Smooth Muscle ◽

Coronary Artery ◽

Essential Oil ◽

Coronary Arteries ◽

Sus Scrofa ◽

Ex Vivo ◽

Alternative Methods ◽

Major Constituent ◽

Sus Scrofa Domesticus

Purpose: Alternative methods for the use of animals in research have gained increasing importance, due to assessments evaluating the real need for their use and the development of legislation that regulates the subject. The principle of the 3R’s (replacement, reduction and refinement) has been an important reference, such that in vitro, ex vivo and cord replacement methods have achieved a prominent place in research. Methods: Therefore, due to successful results from studies developed with these methods, the present study aimed to evaluate the myorelaxant effect of the Dysphania ambrosioides essential oil (EODa) using a Sus scrofa domesticus coronary artery model, and the toxicity of both the Dysphania ambrosioides essential oil and its major constituent, α-terpinene, against Drosophila melanogaster in toxicity and negative geotaxis assays. Results: The EODa relaxed the smooth muscle of swine coronary arteries precontracted with K+ and 5-HT in assays using Sus scrofa domesticus coronary arteries. The toxicity results presented LC50 values of 1.546 mg/mL and 2.282 mg/mL for the EODa and α-terpinene, respectively, thus showing the EODa and α-terpinene presented toxicity to these dipterans, with the EODa being more toxic. Conclusions: Moreover, the results reveal the possibility of using the EODa in vascular disease studies since it promoted the relaxation of the Sus scrofa domesticus coronary smooth muscle.

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SMALL BOWEL ISCHEMIA/REPERFUSION (II/R) ALTERS COLONIC SMOOTH MUSCLE FUNCTION.

Shock ◽

10.1097/00024382-199806001-00042 ◽

1998 ◽

Vol 9 (Supplement) ◽

pp. 13

Author(s):

DT Dempsey ◽

BS Myers ◽

JP Ryan ◽

J Carroll ◽

SI Myers

Keyword(s):

Smooth Muscle ◽

Small Bowel ◽

Muscle Function ◽

Ischemia Reperfusion ◽

Bowel Ischemia ◽

Smooth Muscle Function

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MO583IN VITRO AND EX VIVO EXPERIMENTS OF VASCULAR CALCIFICATION

Nephrology Dialysis Transplantation ◽

10.1093/ndt/gfab086.0021 ◽

2021 ◽

Vol 36 (Supplement_1) ◽

Author(s):

Jana Holmar ◽

Heidi Noels ◽

Joachim Jankowski ◽

Setareh Orth-Alampour

Keyword(s):

Smooth Muscle ◽

Vascular Smooth Muscle ◽

Smooth Muscle Cells ◽

Vascular Calcification ◽

Vascular Smooth Muscle Cells ◽

Incubation Time ◽

Fetal Calf Serum ◽

Ex Vivo ◽

Calf Serum

Abstract Background and Aims Vascular calcification (VC) is one major complication in patients with chronic kidney disease whereas a misbalance in calcium and phosphate metabolism plays a crucial role. The mechanisms underlying VC have not been entirely revealed to date. Therefore are the studies aiming at the identification and characterization of the mediators/uremic toxins involved in VC ongoing and highly relevant. However, currently many different protocols being used in the studies of vascular calcification processes. This complicates the comparison of study outcomes, composing systematic reviews, and meta-analyses. Moreover, the reproducibility of data is hampered, and the efficiency in calcification research through the lack of a standardized protocol is reduced. In this study, we developed a standardized operating protocol for in vitro and ex vivo approaches to aiming at the comparability of these studies. Method We analysed in vitro and ex vivo experimental conditions to study VC. Vascular smooth muscle cells (HAoSMCs) were used for in vitro experiments and aortas from Wistar rats were used for ex vivo experiments. The influence of the following conditions was studied in detail: • Phosphate and calcium concentrations in calcifying media. • Incubation time. • Fetal calf serum (FCS) concentration. The degree of calcification was estimated by quantification of calcium concentrations that were normalized to protein content (in vitro) or to the dry weight of the aortic ring (ex vivo). Additionally, the aortic rings were stained using the von Kossa method. Optimal conditions for investigating medial vascular calcification were detected and summarized in the step-by-step protocol. Results We were able to demonstrate that the degree and the location of VC in vascular smooth muscle cells and aortic rings were highly dependent on the phosphate and CaCl2 concentration in the medium as well as the incubation time. Furthermore, the VC was reduced upon increasing fetal calf serum concentration in the medium. An optimized protocol for studying vascular calcification in vitro and ex vivo was developed and validated. The final protocol (Figure 1) presented will help to standardize in vitro and ex vivo approaches to investigate the processes of vascular calcification. Conclusion In the current study, we developed and validated a standardized operating protocol for systematic in vitro and ex vivo analyses of medial calcification, which is essential for the comparability of the results of future studies.

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Axial Stretch Increases Cell Proliferation in Arteries in Organ Culture

10.1115/imece2000-2516 ◽

2000 ◽

Author(s):

Hai-Chao Han ◽

Raymond P. Vito ◽

Kristin Michael ◽

David N. Ku

Keyword(s):

Cell Proliferation ◽

Smooth Muscle ◽

Organ Culture ◽

Vascular Function ◽

Culture System ◽

Carotid Arteries ◽

Ex Vivo ◽

Axial Stretch ◽

Organ Culture System ◽

Physiological Flow

Abstract To study the effect of axial stretch on vascular function and wall remodeling, porcine carotid arteries were cultured under conditions of physiological flow and elevated axial stretch in an ex vivo organ culture system. Smooth muscle cell proliferation was measured by bromodeoxyuridine index. Results showed that cell proliferation was significantly increased in the highly stretched arteries when compared to the normally stretched arteries. This may indicate the feasibility of stimulating new arterial growth by stretching natural arteries.

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