4 hydroxynonenal 4 hne

Manufactured by Merck Group

Sourced in United States

4-hydroxynonenal (4-HNE) is a naturally occurring lipid peroxidation product. It is an organic compound that serves as a marker of oxidative stress in biological systems.

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Lab products found in correlation

8 protocols using 4 hydroxynonenal 4 hne

Modulating Pluripotent Stem Cell Response to Oxidative Stress

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Cells from the WT ESC line, CCE, and mutant cells derived from this cell line, were cultured as previously described (Kashyap et al., 2011 (link)). Cells were treated with 40 mM EtOH, 1 mM acetaldehyde (AcH) (Calbiochem, San Diego, CA), 2 mM N-acetylcysteine (NAc) (Sigma, St. Louis, MO) or 1 μM 4-hydroxynonenal (4-HNE) (Sigma). AcH was aliquoted from a freshly opened bottle and tubes were stored at −20°C for no more than 2 months. Each aliquot was prepared for a single use. NAc was made fresh in water and pH-adjusted to pH=7.4 for each dosage administered and replaced in the medium every 12 hours. For experiments in which only EtOH was used as a reagent, we administered EtOH to fresh medium twice daily, every 12 hours, for 48 hours, with the final reagent change completed 8 hours prior to harvest. For experiments in which AcH or 4-HNE were used, reagents were changed three times daily, 6–8 hours apart. 1 × 10³ units/ml of LIF were added to the medium for all experiments. ESCs were seeded in 6-well plates to collect mRNA or protein, and 24 well plates for proliferation assays. Cell numbers were counted using an electron particle counter (Coulter Z1, Beckman Coulter, Brea, CA).

Serio R.N., Lu C., Gross S.S, & Gudas L.J. (2019). Different effects of knockouts in ALDH2 and ACSS2 on embryonic stem cell differentiation. Alcoholism, clinical and experimental research, 43(9), 1859-1871.

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Reagents for Biochemical Assays

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Streptozotocin (STZ), 4-nitrophenyl-N-acetyl-β-D-glucosaminide, 4-hydroxynonenal (4-HNE), and 2,4-dinitrophenylhydrazine (DNPH) were purchased from Sigma (St. Louis, MO, USA). All other chemicals used were of the highest analytical grade available.

Arellano-Buendía A.S., Tostado-González M., García-Arroyo F.E., Cristóbal-García M., Loredo-Mendoza M.L., Tapia E., Sánchez-Lozada L.G, & Osorio-Alonso H. (2016). Anti-Inflammatory Therapy Modulates Nrf2-Keap1 in Kidney from Rats with Diabetes. Oxidative Medicine and Cellular Longevity, 2016, 4693801.

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Evaluation of Kidney Injury Markers

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Sodium cholate hydrate, cholesterol, D-(+)-glucose, D-(−)-fructose, 2,4-dinitrophenylhydrazine (DNPH), and 4-hydroxynonenal (4HNE) were purchased from Sigma (St. Louis, MO, USA). Antibodies to nephrin (sc-376522), NGAL (sc-515876), IL-1β (sc-52012), IL-6 (sc-57315), TNF-α (sc-52746), and actin (sc-8432) were acquired from Santa Cruz Biotechnology, Inc., (Dallas, TX, USA). KIM-1 (GTX85067), Nrf2 (GTX103322), Keap1 (GTX60660), NFκB p65 (GTX102090), IκB (GTX82797), and PCNA (GTX100539) were from GeneTex (Irvine, CA, USA). Goat anti-rabbit and goat anti-mouse IgG-HRP were purchased from Cell Signaling Technology (Danvers, MA, USA). All other chemicals used were of the highest purity available.

Arellano Buendia A.S., Juárez Rojas J.G., García-Arroyo F., Aparicio Trejo O.E., Sánchez-Muñoz F., Argüello-García R., Sánchez-Lozada L.G., Bojalil R, & Osorio-Alonso H. (2023). Antioxidant and anti-inflammatory effects of allicin in the kidney of an experimental model of metabolic syndrome. PeerJ, 11, e16132.

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Oxidative Stress in Human Lens Cells

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A human lens epithelial cell line was provided by Yang Chunbo (Tianjin Medical University, Eye Institute). The cells were grown in Eagle’s minimum essential medium containing 10% fetal bovine serum. For transient transfection, plasmids were transfected into cells with Lipofectamine 2000 (Invitrogen) according to the manufacturer’s protocol [16 (link)]. We used 4-hydroxynonenal (4-HNE, Sigma) at a working concentration of 10 µM as a stressor to induce oxidative stress, incubated the cells for 18 h, and collected them for further use [17 (link)].

Teng H., Hong Y., Cao J., Li H., Tian F., Sun J., Wen K., Han G., Whelchel A., Zhang X., Li X, & Dong L. (2022). Senescence marker protein30 protects lens epithelial cells against oxidative damage by restoring mitochondrial function. Bioengineered, 13(5), 12955-12971.

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Evaluating Cell Death Pathways

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All-trans-retinal (atRAL) (TRC, Canada), 4-Hydroxynonenal (4-HNE) (Sigma, Burlington, MA), 2-(4-Amino-3-methylphenyl)benzothiazole (DF 203) (eMolecules, San Diego, CA), Z-VAD (R&D, Minneapolis, MN), Necrostatin-1 (Biovision, Waltham, MA), 5,11-Dihydroindolo[3,2-b]carbazole-6-carboxaldehyde (FICZ), and kynurenic acid (Tocris, Minneapolis, MN) were used in this study. atRAL was stored and manipulated with limited light exposure.

Schustak J., Han H., Bond K., Huang Q., Saint-Geniez M, & Bao Y. (2024). Phenotypic high-throughput screening identifies aryl hydrocarbon receptor agonism as common inhibitor of toxin-induced retinal pigment epithelium cell death. PLOS ONE, 19(4), e0301239.

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Assessing Autophagy and Protein Regulation

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For assessing effects on autophagy, SF were silenced for CBP or p300 and treated with TNF as described above. Bafilomycin A1 (100 nM, Tocris Bioscience) was added for 4 h prior to harvesting the cells. For assessing the regulation of CBP and p300 protein expression, SF were treated with TNF (10 ng/ml), 4-hydroxynonenal (4-HNE, 5 µM, Sigma Aldrich), or a combination of both for 48 h, or with H₂O₂ (0,1 and 0,3 mM, Sigma Aldrich) for 24 h. Controls were treated with matched amounts of ethanol.

Krošel M., Gabathuler M., Moser L., Maciukiewicz M., Züllig T., Seifritz T., Tomšič M., Distler O., Ospelt C, & Klein K. (2023). The histone acetyl transferases CBP and p300 regulate stress response pathways in synovial fibroblasts at transcriptional and functional levels. Scientific Reports, 13, 17112.

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Investigating Oxidative Stress Markers

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CdCl₂ was purchased from Alfa Aesar (Tewksbury, MA, USA). The antibodies against atrial natriuretic peptide (ANP), collagen 1a1 (COL1A1), β-actin, catalase (CAT), superoxide dismutase 2 (SOD2), and NADPH dehydrogenase quinone 1 (NQO-1) were purchased from Santa Cruz Biotechnologies (Dallas, TX, USA). The antibodies for transforming growth factor β1 (TGF-β1) and fibronectin (FN) were from Abcam (Cambridge, MA, USA). Plasminogen activator inhibitor-1 (PAI-1) antibody was purchased from BD (Franklin Lakes, NJ, USA). The 3-nitrotyrosine (3-NT) and 4-hydroxynonenal (4-HNE) antibodies were from Millipore (Billerica, CA, USA) and Alpha Diagnostic International (San Antonio, TX, USA), respectively. Antibodies for metallothionein (MT) were from DakoCytomation (Carpinteria, CA, USA). All other chemicals were of the highest purity commercially available.

Liang Y., Young J.L., Kong M., Tong Y., Qian Y., Freedman J.H, & Cai L. (2019). Gender Differences in Cardiac Remodeling Induced by a High-Fat Diet and Lifelong, Low-Dose Cadmium Exposure. Chemical research in toxicology, 32(6), 1070-1081.

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Histological and Immunohistochemical Analysis of Embryonic Tissues

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The embryos were fixed in 4% paraformaldehyde (PFA) in phosphate buffered saline (PBS, pH 7.4) at 4°C for 16 hours, followed by dehydration through a series of concentrations of ethanol and embedding in paraffin wax through mediation steps of xylene. Tissue sections at 6 μm thickness were cut using a microtome.
Tissue sections were dewaxed in xylene and rehydrated through a reverse ethanol concentration series to water. For histological preparation, the sections were stained with hematoxylin and eosin (Junqueira and Carneiro, 2003 ).
For immunohistochemistry, the sections were boiled in a citrate buffer (10 mM Citrate Acid, pH 6.0) for 5 minutes to unmask antigens, equilibrated in PBS (pH 7.4), incubated with primary antibodies recognizing 4-hydroxynonenal (4-HNE; Millipore, Temecula, CA), 3-nitrotyrosine (3-NT, Cell Signaling Technology, Danvers, MA), protein disulfide isomerase (PDI; Cell Signaling Technology), phosphorylated (p-) calnexin (Abcam, Cambridge, MA), Inhibin βA (EMD Biosciences, Gibbstown, NJ), and p-Smad3 (Cell Signaling Technology) (Zhao, 2012 (link)).
Signals were detected using secondary antibodies conjugated with alkaline phosphatase (AP; Santa Cruz Biotechnology, Santa Cruz, CA) and its substrates (nitro-blue tetrazolium and 5-bromo-4-chloro-3’-indolyphosphate). Tissue sections were counterstained with fast red.

, & Zhao Z. (2014). TGFβ and Wnt in Cardiac Outflow Tract Defects in Offspring of Diabetic Pregnancies. Birth defects research. Part B, Developmental and reproductive toxicology, 101(5), 364-370.

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