Twenty minutes after the stress procedure, male rats were lethally anaesthetized (pentobarbital sodium 100 mg/kg ip (Ceva)). Immediately after sacrifice, sections of the colon, cut along the mesenteric border, were mounted in Ussing chambers (Easymount). All segments were properly adjusted to fit the entire exposed tissue surface area corresponding to 0.5 cm2. Chambers were filled with 5 mL of Krebs‐Henseleit solution (Sigma) maintained at 37°C and continuously gassed with 95% O2/5% CO2. Transepithelial resistance (TER) was monitored throughout the experiment to assess the viability of the tissue.26 Paracellular permeability was assessed by measuring mucosal‐to‐serosal flux of fluorescein isothiocyanate (FITC)‐labeled 4 kDa dextran (Sigma). After 10‐minute equilibrium period, 500 µL of the Krebs‐Henseleit solution was replaced by 500 µL of the FITC‐labeled 4 kDa dextran (2.2 mg/mL as the final concentration) in the mucosal side. After 1 hour, fluorescence was measured in the serosal buffer side by a fluorimeter (wavelength of 540 nm, Tecan). Results were expressed as the flux of dextran crossing the epithelial barrier (nmol/cm2/h).
Krebs henseleit solution
Manufactured by Merck Group
Sourced in United States
Krebs-Henseleit solution is a buffered salt solution used in various laboratory and experimental applications. It is designed to simulate the extracellular fluid composition in the body, making it a widely used medium for maintaining and studying isolated tissues or organs ex vivo.
Automatically generated - may contain errors
Lab products found in correlation
Fitc labeled dextran 4 kda, by Merck Group (2 mentions)
Fluorimeter, by Tecan (1 mentions)
Sodium pentobarbital, by Ceva (1 mentions)
U46619, by Merck Group (1 mentions)
Grass s48 stimulator, by Natus (1 mentions)
Model 403a, by Aurora Scientific (1 mentions)
Powerlab, by ADInstruments (1 mentions)
Serotonin, by Merck Group (1 mentions)
Apocynin, by Merck Group (1 mentions)
Anti mnsod, by Enzo Life Sciences (1 mentions)
Anti phospho enos ser1177, by Cell Signaling Technology (1 mentions)
Urethane, by Merck Group (1 mentions)
Anti catalase, by Merck Group (1 mentions)
Anti enos, by BD (1 mentions)
Anti α actin, by Abcam (1 mentions)
Anti p47phox, by Merck Group (1 mentions)
Anti gpx, by Abcam (1 mentions)
Acetylcholine, by Merck Group (1 mentions)
Sodium nitroprusside, by Merck Group (1 mentions)
Fitc dextran, by Tecan (1 mentions)
7 protocols using krebs henseleit solution
1
Colon Epithelial Barrier Permeability Assessment
2
Euthanasia and Tissue Harvesting in Rat Reproduction Study
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The study protocol obtained approval from the Regional Directorate for Health, Animal and Environment Protection, an accredited organization in France. Additionally, it was also approved by the French Ministry of Research under the reference number APAFIS#8207-2016121322563594 v2. These regulatory approvals ensured that the study adhered to the necessary ethical and scientific standards regarding animal welfare and research conduct.
At PND60, the offspring rats were euthanized using an injection of (1 mL·kg−1; 200 mg·mL−1 solution) sodium pentobarbital from EXAGON (Axience, Pantin, France). After euthanasia, the reproductive organs were dissected and placed directly into Krebs Henseleit solution from Sigma Aldrich (Saint Quentin Fallavier, France). This solution is commonly used to maintain the viability and functionality of isolated tissues during experimental procedures (Figure 2 ).
At PND60, the offspring rats were euthanized using an injection of (1 mL·kg−1; 200 mg·mL−1 solution) sodium pentobarbital from EXAGON (Axience, Pantin, France). After euthanasia, the reproductive organs were dissected and placed directly into Krebs Henseleit solution from Sigma Aldrich (Saint Quentin Fallavier, France). This solution is commonly used to maintain the viability and functionality of isolated tissues during experimental procedures (
3
Aortic Endothelial Relaxation Measurement
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The measurement of endothelial relaxation functioning was conducted as previously described (3 (link)). In brief, the thoracic aorta was immediately dissected and immersed in chilled Krebs-Henseleit solution (Sigma-Aldrich; Merck KGaA) at 37°C and aerated with 95% O2 and 5% CO2 (pH 7.4). The aortic rings were then pre-contracted with U46619 (Sigma-Aldrich; Merck KGaA; 30 nM). Endothelium-dependent and -independent vasodilation were determined using ACh (Selleck Chemicals; 10−9-10−5 M) and SNP (Selleck Chemicals; 10−10-10−6 M), respectively.
4
Isometric Force Measurement of Engineered Muscle Fibers
5
Muscle Contractile Properties Measured Ex Vivo
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As previously described, EDL muscle contractile properties were measured ex vivo [22 (link)]. Each muscle strip (approximately 1 cm in length) was prepared on ice and rapidly mounted in a tissue chamber containing Krebs-Henseleit solution (Sigma, Saint Louis, MO, USA), which was bubbled with a gas mixture of 95% O2–5% CO2. Muscle extremities were held in spring clips and attached to an electromagnetic force transducer for the measurements. The maximum tetanic forces and the force-frequency curves were used to evaluate muscle force-generating capacity.
6
Evaluation of Antioxidant Enzyme Expression
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NP and LP diets were purchased from Pragsoluções Biociências (Jaú, SP, Brazil) and taurine (2-aminoethanesulfonic acid) was from Botica Ouro da Mata Compounding Pharmacy (Campinas, SP, Brazil). MnTMPyP [Mn(III) tetrakis(1-methyl-4-pyridyl)porphyrin] pentachloride was from Calbiochem (San Diego, CA, USA). Components of Krebs-Henseleit solution, urethane, serotonin, acetylcholine, sodium nitroprusside, SOD, and apocynin were purchased from Sigma-Aldrich (Saint Louis, MO, USA). Primary antibodies: anti-eNOS from BD Transduction Laboratories (Franklin Lakes, NJ, USA); anti-phospho-eNOS (Ser 1177) and anti-CuZn-SOD from Cell Signaling Technology (Beverly, MA, USA); anti-EC-SOD and anti-Mn-SOD from Enzo Life Sciences (Farmingdale, NY, EUA); anti-p47phox and anti-catalase from Sigma-Aldrich (St Louis, MO, USA); anti-gp91phox from Millipore (Millipore Corporation, Bedford, MA, USA); anti-GPX and anti-α-actin from Abcam (Cambridge, MA, USA).
7
Evaluating Intestinal Permeability via Ussing Chambers
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Sections of ileum and colon were mounted in Ussing chambers (Easymount; Hamden) having a flux area of in order to evaluate the intestinal permeability as previously described.25 (link) Briefly, intestinal tissues were maintained at 37°C in oxygenated Krebs-Henseleit solution (Sigma-Aldrich). Mucosal-to-serosal transport of macromolecules was determined by adding fluorescein isothiocyanate (FITC)-labeled 4 kDa dextran (Sigma-Aldrich) in the luminal side of the chamber. After 1 h, fluorescence was measured in the serosal buffer with a fluorimeter (Tecan Spark), and the results were expressed as the flux of FITC dextran crossing of epithelium per hour ( ). Data are reported as the means of triplicate measurements.
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