Product Citations: 261

Fig.5.A showing Western Blotting from the publication: STING inhibition enables efficient plasmid-based gene expression in primary vascular cells: A simple and cost-effective transfection protocol.
Fig.5.B showing Western Blotting from the publication: Indole-3-Propionic Acid, a Gut Microbiota-Derived Tryptophan Metabolite, Promotes Endothelial Dysfunction Impairing Purinergic-Induced Nitric Oxide Release in Endothelial Cells.
Fig.2.C showing Western Blotting from the publication: Polydatin improves vascular endothelial function by maintaining mitochondrial homeostasis under high glucose conditions.
Fig.6.C showing Western Blotting from the publication: Decreased expression of mitochondrial aminoacyl-tRNA synthetases causes downregulation of OXPHOS subunits in type 2 diabetic muscle.
Fig.6.D showing Western Blotting from the publication: Decreased expression of mitochondrial aminoacyl-tRNA synthetases causes downregulation of OXPHOS subunits in type 2 diabetic muscle.
Fig.1.C showing Western Blotting from the publication: Murine endothelial serine palmitoyltransferase 1 (SPTLC1) is required for vascular development and systemic sphingolipid homeostasis.
Fig.1.A showing Western Blotting from the publication: Murine endothelial serine palmitoyltransferase 1 (SPTLC1) is required for vascular development and systemic sphingolipid homeostasis.
Fig.2.K showing Immunohistochemistry-immunofluorescence in a Homo sapiens (Human) sample from the publication: Nrg1β Released in Remote Ischemic Preconditioning Improves Myocardial Perfusion and Decreases Ischemia/Reperfusion Injury via ErbB2-Mediated Rescue of Endothelial Nitric Oxide Synthase and Abrogation of Trx2 Autophagy.
Fig.6.B showing Western Blotting from the publication: Latent HIV-Exosomes Induce Mitochondrial Hyperfusion Due to Loss of Phosphorylated Dynamin-Related Protein 1 in Brain Endothelium.
Fig.6.T showing Western Blotting from the publication: Latent HIV-Exosomes Induce Mitochondrial Hyperfusion Due to Loss of Phosphorylated Dynamin-Related Protein 1 in Brain Endothelium.
Fig.5.D showing Western Blotting from the publication: BReast CAncer susceptibility gene 2 deficiency exacerbates oxidized LDL-induced DNA damage and endothelial apoptosis.
Fig.3.C showing Western Blotting from the publication: Nerve damage induced skeletal muscle atrophy is associated with increased accumulation of intramuscular glucose and polyol pathway intermediates.
Fig.4.E showing Western Blotting from the publication: Myofibrillar function differs markedly between denervated and dexamethasone-treated rat skeletal muscles: Role of mechanical load.
Fig.1.E showing Western Blotting from the publication: Peroxynitrite supports a metabolic reprogramming in merlin-deficient Schwann cells and promotes cell survival.
Fig.1.C showing Western Blotting in a Homo sapiens (Human) sample from the publication: DDAH1 regulates apoptosis and angiogenesis in human fetal pulmonary microvascular endothelial cells.
Fig.4.C showing Western Blotting in a Mus musculus (House mouse) sample from the publication: RELi protocol: Optimization for protein extraction from white, brown and beige adipose tissues.
Fig.4.B showing Western Blotting in a Mus musculus (House mouse) sample from the publication: RELi protocol: Optimization for protein extraction from white, brown and beige adipose tissues.
Fig.4.A showing Western Blotting in a Mus musculus (House mouse) sample from the publication: RELi protocol: Optimization for protein extraction from white, brown and beige adipose tissues.
Fig.8.A showing Western Blotting from the publication: Reduced caveolae density in arteries of SHR contributes to endothelial dysfunction and ROS production.
Fig.8.C showing Western Blotting from the publication: Reduced caveolae density in arteries of SHR contributes to endothelial dysfunction and ROS production.
Fig.7.B showing Western Blotting in a Rattus norvegicus (Rat) sample from the publication: Long-Term Regulation of Excitation-Contraction Coupling and Oxidative Stress in Cardiac Myocytes by Pirfenidone.
Fig.5.B showing Western Blotting from the publication: Endothelial AMP-Activated Kinase α1 Phosphorylates eNOS on Thr495 and Decreases Endothelial NO Formation.
Fig.6.A showing Western Blotting in a Rattus norvegicus (Rat) sample from the publication: Electrical Stimulation Prevents Preferential Skeletal Muscle Myosin Loss in Steroid-Denervation Rats.
Fig.6.D showing Western Blotting in a Rattus norvegicus (Rat) sample from the publication: Electrical Stimulation Prevents Preferential Skeletal Muscle Myosin Loss in Steroid-Denervation Rats.
Fig.2.D showing Western Blotting from the publication: Boysenberry polyphenol inhibits endothelial dysfunction and improves vascular health.
35 images found

Immune-mediated regeneration of cell-free vascular grafts in an ovine model.

In Npj Regenerative Medicine on 19 March 2025 by Nasiri, B., Das, A., et al.

We developed acellular tissue engineered vessels (ATEV) using small intestine submucosa (SIS) incorporating heparin and a novel protein named H2R5. ATEVs were implanted into the arterial circulation of an ovine animal model, demonstrating high primary patency rates over a period of three months. Implanted grafts were infiltrated by host cells, the majority of which were monocytes/macrophages (MC/MΦ), as demonstrated by scRNA sequencing and immunostaining. They also developed functional endothelial and medial layers that deposited new extracellular matrix leading to matrix remodeling and acquisition of mechanical properties that were similar to those of native arteries. Notably, during this short implantation time, ATEVs turned into functional neo-arteries, as evidenced by the development of the vascular contractile function. Our findings underscore the potential of H2R5-functionalized ATEVs as promising candidates for tissue replacement grafts in a large pre-clinical animal model and highlight the contribution of macrophages in vascular regeneration.
© 2025. The Author(s).

  • IHC
  • Immunology and Microbiology
  • Veterinary Research
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Simultaneous isolation and culture of endothelial colony-forming cells, endothelial cells and vascular smooth muscle cells from human umbilical cords.

In Molecular Biology Reports on 13 March 2025 by Burger, M. L., Menétrey, S., et al.

Regulation of the human umbilical circulation under physiological and pathological conditions remains poorly understood. We previously demonstrated that intrauterine growth restriction (IUGR) is associated with sex-specific alterations in the human umbilical circulation. Our data strongly suggest a differential contribution of subcellular compartmentation depending on fetal sex, vessel type and the presence of IUGR. We therefore developed a protocol to isolate and culture umbilical vascular cells to further investigate the relative contribution of each cell type and subcellular compartmentation to the human umbilical circulation regulation.
Human umbilical cords and cord blood were collected just after delivery. Mononuclear cells were recovered from cord blood using a Ficoll gradient and cultured to obtain endothelial colony-forming cells (ECFCs). Endothelial cells (ECs) were isolated from human umbilical vein (HUV) and arteries (HUAs) by collagenase/dispase digestion, and vascular smooth muscle cells (SMCs) by migration from vascular explants. All cell types were characterized by visualization, and by analysis of biomarkers using immunocytofluorescence and Western blot. ECFCs were also submitted to polychromatic flow cytometry analysis.
This protocol enables simultaneous isolation and culture of ECFCs, HUVECs, HUAECs, HUVSMCs and HUASMCs from the same umbilical cord. It is simpler, faster and more cost-effective than other previously published methods, with good success rates. This will be helpful to further investigate the regulatory mechanisms implicated in the human umbilical circulation under physiological and pathological conditions and to study the influence of fetal sex.
© 2025. The Author(s).

  • Biochemistry and Molecular biology
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Mural cell dysfunction contributes to diastolic heart failure by promoting endothelial dysfunction and vessel remodelling.

In Cardiovascular Diabetology on 7 February 2025 by Grootaert, M. O. J., Pasut, A., et al.

Heart failure with preserved ejection fraction (HFpEF) is a complex cardiovascular disease associated with metabolic comorbidities. Microvascular dysfunction has been proposed to drive HFpEF, likely via endothelial cell (EC) dysfunction, yet the role of the mural cells herein has never been explored.
We used the diabetic db/db mouse given 1% salt as a new model of HFpEF and crossed then with PDGFRβtg/tg-CreERT2-EYFPtg/tg mice to label the mural cells. We combined single-cell RNA sequencing, NichetNet analysis and histology to determine the role of mural cell dysfunction in HFpEF.
Db/db mice given 1% salt for 8 weeks developed diastolic dysfunction preceded by capillary density loss, pericyte loss and vessel regression. At 4 weeks of salt, hearts of db/db mice already showed EC dysfunction associated with an anti-angiogenic signature, and an increase in pericyte-EC intracellular space. Db/db + salt hearts were further characterised by increased ACTA2 expression, arteriole wall thickening and vessel enlargement. NicheNet analysis on the single cell transcriptomic data revealed little signalling from the ECs to the mural cells; instead, mural cells signalled strongly to ECs. Mechanistically, pericyte dysfunction induces an EC growth arrest via TNFα-dependent paracrine signalling and downstream signalling through STAT1.
Mural cell dysfunction contributes to HFpEF by inducing coronary vessel remodelling, at least in part by reducing EC proliferation and inducing EC inflammation through TNFα-dependent paracrine signalling.
© 2025. The Author(s).

  • Cardiovascular biology
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Endothelial-Protective Actions of Diethylether Extract from Gentiana kochiana and Xanthone Gentiacaulein Against Oxidized LDL-Induced Injury-In Vitro Evaluation.

In International Journal of Molecular Sciences on 5 February 2025 by Tovilović-Kovačević, G., Zogović, N., et al.

Endothelial dysfunction is an early pathophysiological event in atherosclerosis. The endothelial-protective abilities of diethylether extract (EE) from the Gentiana kochiana (Gentianaceae) herb and its main component, xanthone aglycone gentiacaulein (GC), were evaluated in an oxidized low-density lipoprotein (oxLDL)-treated EA.hy926 endothelial cell line. The EE and GC actions were assessed using cell viability assays, flow cytometry, immunoblot, DPPH, NBT, TBARS, conjugated diene formation, and Griess tests. Both EE and GC prevented oxLDL-induced apoptosis by reducing intracellular reactive oxygen species levels, mitochondrial depolarization, and caspase activation in EA.hy926 cells. EE and GC dose-dependently diminished oxLDL-induced cellular lipid peroxidation. In cell-free conditions, EE moderately scavenged superoxide anions and had no affinity toward DPPH radicals, GC did not interact with either of investigated free radicals, while both EE and GC effectively delayed Cu²⁺-induced LDL oxidation. EE and GC upregulated oxLDL-suppressed protective Akt/CREB/eNOS and ERK signals and restored oxLDL-reduced nitric oxide levels. Therefore, EE and GC effectively counteract oxLDL-induced endothelial apoptosis by reducing oxidative stress, promoting mitochondrial recovery, and enhancing the prosurvival Akt/CREB/eNOS axis and ERK activity. Our study is the first to demonstrate that xanthone-rich EE from aerial parts of G. kochiana and xanthone GC alleviate oxLDL-induced endothelial cell injury, underscoring their potential for cardiovascular protection.

  • WB
  • Homo sapiens (Human)
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Fibrillin-1 Deficiency Perturbs Aortic Cholinergic Relaxation and Adrenergic Contraction in a Mouse Model of Early Onset Progressively Severe Marfan Syndrome.

In Journal of Vascular Research on 20 January 2025 by Cantalupo, A., Asano, K., et al.

The pathogenic role of nitric oxide (NO) signaling during development of thoracic aortic aneurysm (TAA) in Marfan syndrome (MFS) is currently unclear. We characterized vasomotor function and its relationship to the activity of the NO-generating enzymes in mice with early onset progressively severe MFS.
Wire myography, immunoblotting, measurements of aortic NO, and superoxide levels were used to compare vasomotor function, contractile protein levels, and the activity of endothelial and inducible NO synthase (eNOS and iNOS, respectively) in ascending thoracic aortas of Fbn1mgR/mgR mice relative to wild-type littermates.
Isometric force measurements of aortic rings from 16-day-old male Fbn1mgR/mgR mice revealed a significant reduction in acetylcholine-induced relaxation and increased phenylephrine (PE)-promoted contractility, associated with abnormally low eNOSSer1177 phosphorylation, decreased NO production, and augmented superoxide levels. Greater aortic contractility was associated with α1-adrenoceptor upregulation and normal levels of contractile proteins. While iNOS inhibition had no effect on vasomotor functions, mutant aortic rings preincubated with a nonspecific NOS inhibitor yielded a greater PE response, implying a significant contribution of endothelial dysfunction to aortic hypercontractility.
Impaired eNOS signaling disrupts aortic cholinergic relaxation and adrenergic contraction in MFS mice with dissecting TAA.
The pathogenic role of nitric oxide (NO) signaling during development of thoracic aortic aneurysm (TAA) in Marfan syndrome (MFS) is currently unclear. We characterized vasomotor function and its relationship to the activity of the NO-generating enzymes in mice with early onset progressively severe MFS.
Wire myography, immunoblotting, measurements of aortic NO, and superoxide levels were used to compare vasomotor function, contractile protein levels, and the activity of endothelial and inducible NO synthase (eNOS and iNOS, respectively) in ascending thoracic aortas of Fbn1mgR/mgR mice relative to wild-type littermates.
Isometric force measurements of aortic rings from 16-day-old male Fbn1mgR/mgR mice revealed a significant reduction in acetylcholine-induced relaxation and increased phenylephrine (PE)-promoted contractility, associated with abnormally low eNOSSer1177 phosphorylation, decreased NO production, and augmented superoxide levels. Greater aortic contractility was associated with α1-adrenoceptor upregulation and normal levels of contractile proteins. While iNOS inhibition had no effect on vasomotor functions, mutant aortic rings preincubated with a nonspecific NOS inhibitor yielded a greater PE response, implying a significant contribution of endothelial dysfunction to aortic hypercontractility.
Impaired eNOS signaling disrupts aortic cholinergic relaxation and adrenergic contraction in MFS mice with dissecting TAA.
© 2025 The Author(s). Published by S. Karger AG, Basel.

  • Cardiovascular biology
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STING inhibition enables efficient plasmid-based gene expression in primary vascular cells: A simple and cost-effective transfection protocol.

In PLoS One on 11 July 2024 by Yuan, S. & Straub, A. C.

Fig.5.A
Fig.5.A showing Western Blotting from the publication: STING inhibition enables efficient plasmid-based gene expression in primary vascular cells: A simple and cost-effective transfection protocol.
  • WB
Over-expressed eNOS residues in the physiological compartment and responses to VEGF signaling.HAECs were transfected with 0.2 μg pCIGX plasmids carrying no transgene (EV), wild-type eNOS (WT), or S1177A mutant eNOS (S1177A) with optimized MRT67307 treatment. 48 hours after transfection, cells wer... more
Over-expressed eNOS residues in the physiological compartment and responses to VEGF signaling.HAECs were transfected with 0.2 μg pCIGX plasmids carrying no transgene (EV), wild-type eNOS (WT), or S1177A mutant eNOS (S1177A) with optimized MRT67307 treatment. 48 hours after transfection, cells were deprived of serum and growth factors for 2 hours and treated with 20 ng/ml VEGF for 5 minutes. Western blot showed WT but not S1177A eNOS is phosphorylated by VEGF treatment (A) (* annotates p < 0.05, n = 3). Immunostaining shows over-expressed eNOS in GFP-positive cells co-localized with the plasma membrane and ER (B). WT but not S1177A eNOS over-expressing cells show increased phosphorylation of eNOS S1177. less

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Indole-3-Propionic Acid, a Gut Microbiota-Derived Tryptophan Metabolite, Promotes Endothelial Dysfunction Impairing Purinergic-Induced Nitric Oxide Release in Endothelial Cells.

In Int J Mol Sci on 16 March 2024 by Geddo, F., Antoniotti, S., et al.

Fig.5.B
Fig.5.B showing Western Blotting from the publication: Indole-3-Propionic Acid, a Gut Microbiota-Derived Tryptophan Metabolite, Promotes Endothelial Dysfunction Impairing Purinergic-Induced Nitric Oxide Release in Endothelial Cells.
  • WB
IPA regulates eNOS phosphorylation in BAE-1 cells. (a) Bar graph summarizing the p-eNOS/eNOS ratio normalized toward positive control (ATP) after treatment with 100 µM of ATP (5 min) and 1 µM of IPA (30 min and 24 h). Data in percentages are represented as mean ± SEM, * p < 0.05, ** p < 0.01, ***... more
IPA regulates eNOS phosphorylation in BAE-1 cells. (a) Bar graph summarizing the p-eNOS/eNOS ratio normalized toward positive control (ATP) after treatment with 100 µM of ATP (5 min) and 1 µM of IPA (30 min and 24 h). Data in percentages are represented as mean ± SEM, * p < 0.05, ** p < 0.01, *** p < 0.001. Values of the n = 4 independent experiments were as follows: CTRL: 29.36 ± 5.15; ATP: 98.85 ± 0.86; IPA 30 min: 39.06 ± 6.64; IPA 30 min + ATP: 67.45 ± 5.88; IPA 24 h: 28.57 ± 4.55; IPA 24 h + ATP: 73.41 ± 4.55. (b) Representative Western blot experiment showing the effect of ATP and IPA on eNOS phosphorylation. Comparison of β-actin intensity ensured equal protein loading. IPA = indole-3-propionic acid, ATP = adenosine triphosphate. less

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Polydatin improves vascular endothelial function by maintaining mitochondrial homeostasis under high glucose conditions.

In Sci Rep on 2 October 2023 by Shah, W., Zhao, Q., et al.

Fig.2.C
Fig.2.C showing Western Blotting from the publication: Polydatin improves vascular endothelial function by maintaining mitochondrial homeostasis under high glucose conditions.
  • WB
Effect of polydatin on morphological changes of aortic rings and viability of HUVECs under high-glucose condition. (A) Representative HE–stained thoracic aortas of normal glucose (NG) incubation (a), high glucose (HG) incubation (b), and polydatin (Poly, 10 µM) co-incubation with HG (Poly + HG) (... more
Effect of polydatin on morphological changes of aortic rings and viability of HUVECs under high-glucose condition. (A) Representative HE–stained thoracic aortas of normal glucose (NG) incubation (a), high glucose (HG) incubation (b), and polydatin (Poly, 10 µM) co-incubation with HG (Poly + HG) (c). Scale bar: 50 µm. (B) CCK-8 assay showed that the viability ratio was increased in HUVECs after Poly treatment under HG conditions (n = 5 per group). (C–E) Representative bands (C) and summarized data (D, E) of eNOS and iNOS protein expression in NG, HG, and Poly + HG groups (n = 6 per group). Data are expressed as mean ± SEM. *P < 0.05 versus NG, △P < 0.05 versus HG. less

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Decreased expression of mitochondrial aminoacyl-tRNA synthetases causes downregulation of OXPHOS subunits in type 2 diabetic muscle.

In Redox Biol on 1 May 2023 by López-Soldado, I., Torres, A. G., et al.

Fig.6.C
Fig.6.C showing Western Blotting from the publication: Decreased expression of mitochondrial aminoacyl-tRNA synthetases causes downregulation of OXPHOS subunits in type 2 diabetic muscle.
  • WB
Skeletal muscle gene expression of iNOS and eNOS in obese and diabetic mice and their respective matched control group. iNOS and eNOS protein expression in mitochondria isolated from skeletal muscle of obese and diabetic mice and their respective matched control group.Real-time PCR or Western blo... more
Skeletal muscle gene expression of iNOS and eNOS in obese and diabetic mice and their respective matched control group. iNOS and eNOS protein expression in mitochondria isolated from skeletal muscle of obese and diabetic mice and their respective matched control group.Real-time PCR or Western blot was performed in skeletal muscle biopsies. Ob/ob are obese mice, db/db are diabetic mice, and their respective controls are C57BL/6 and db/+. (A) Skeletal muscle gene expression of iNOS and eNOS in obese mice. (B) Skeletal muscle gene expression of iNOS and eNOS in diabetic mice. (C) iNOS and eNOS protein expression in enriched mitochondrial fractions from skeletal muscle of obese mice. (D) iNOS and eNOS protein expression in enriched mitochondrial fractions from skeletal muscle of diabetic mice. Data are mean ± SEM. Statistical analyses comparing C57BL6 vs ob/ob or db/+ vs db/db mice were performed by unpaired t-test *p < 0.05. n = 6–8/group. less

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Decreased expression of mitochondrial aminoacyl-tRNA synthetases causes downregulation of OXPHOS subunits in type 2 diabetic muscle.

In Redox Biol on 1 May 2023 by López-Soldado, I., Torres, A. G., et al.

Fig.6.D
Fig.6.D showing Western Blotting from the publication: Decreased expression of mitochondrial aminoacyl-tRNA synthetases causes downregulation of OXPHOS subunits in type 2 diabetic muscle.
  • WB
Skeletal muscle gene expression of iNOS and eNOS in obese and diabetic mice and their respective matched control group. iNOS and eNOS protein expression in mitochondria isolated from skeletal muscle of obese and diabetic mice and their respective matched control group.Real-time PCR or Western blo... more
Skeletal muscle gene expression of iNOS and eNOS in obese and diabetic mice and their respective matched control group. iNOS and eNOS protein expression in mitochondria isolated from skeletal muscle of obese and diabetic mice and their respective matched control group.Real-time PCR or Western blot was performed in skeletal muscle biopsies. Ob/ob are obese mice, db/db are diabetic mice, and their respective controls are C57BL/6 and db/+. (A) Skeletal muscle gene expression of iNOS and eNOS in obese mice. (B) Skeletal muscle gene expression of iNOS and eNOS in diabetic mice. (C) iNOS and eNOS protein expression in enriched mitochondrial fractions from skeletal muscle of obese mice. (D) iNOS and eNOS protein expression in enriched mitochondrial fractions from skeletal muscle of diabetic mice. Data are mean ± SEM. Statistical analyses comparing C57BL6 vs ob/ob or db/+ vs db/db mice were performed by unpaired t-test *p < 0.05. n = 6–8/group. less

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Murine endothelial serine palmitoyltransferase 1 (SPTLC1) is required for vascular development and systemic sphingolipid homeostasis.

In Elife on 5 October 2022 by Kuo, A., Checa, A., et al.

Fig.1.C
Fig.1.C showing Western Blotting from the publication: Murine endothelial serine palmitoyltransferase 1 (SPTLC1) is required for vascular development and systemic sphingolipid homeostasis.
  • WB
Ablation of Sptlc1 in EC reduces SL content in both EC and non-EC cell population of the lung.(A) Scheme of lung EC and non-EC isolation procedure used in this figure. (B) Representative flow cytometry analysis for determination of lung non-EC and EC population purities. Non-EC is identified as C... more
Ablation of Sptlc1 in EC reduces SL content in both EC and non-EC cell population of the lung.(A) Scheme of lung EC and non-EC isolation procedure used in this figure. (B) Representative flow cytometry analysis for determination of lung non-EC and EC population purities. Non-EC is identified as CD45+/-; CD31- cells (Q1 + Q4). EC is identified as CD45-; CD31 + cells (Q3) (n=4 for both EC and non-EC). (C) EC and non-EC from Ctrl and Sptlc1 ECKO lungs were analyzed by Western Blotting for SPTLC1 and eNOS expressions. eNOS was used as an EC marker to demonstrate purity of each population. β-actin was used as an internal control. Relative fold change of SPTLC1 to Ctrl in EC and non-EC population were normalized to β-actin and quantified in (D) an (E) (n=3 for both genotypes). SL content in Ctrl and Sptlc1 KO EC was determined by LC-MS/MS, including dhC16-Cer, dhSph, dhS1P, Sph and S1P in (F), total Cer content in (G), and SM in (H) (n=5 for Ctrl EC, n=4 for Sptlc1 KO EC). SL content in Ctrl and Sptlc1 KO non-EC was determined by LC-MS/MS, including dhC16-Cer, dhSph, dhS1P, Sph, and S1P in (I), total Cer content in (J), and total SM content in (K) (n=5 for Ctrl non-EC, n=4 for Sptlc1 KO non-EC). Values of lipidomic results can be accessed in Figure 1—source data 1. Data are expressed as mean ± SD. Statistical significance was determined by unpaired t test. *p<0.05; **p<0.01. ***p<0.001. ns, non-significant.Figure 1—source data 1.Levels of dhC16-Cer, dhSph, dhS1P, Sph, S1P, Cer, and SM in lung EC and non-EC populations of Ctrl and Sptlc1 ECKO mice.Figure 1—source data 2.Original blots of EC and non-EC from Ctrl and Sptlc1 ECKO lungs.Levels of dhC16-Cer, dhSph, dhS1P, Sph, S1P, Cer, and SM in lung EC and non-EC populations of Ctrl and Sptlc1 ECKO mice.Original blots of EC and non-EC from Ctrl and Sptlc1 ECKO lungs.Sptlc1 ECKO mice exhibit similar body weight and blood chemistry as control mice.(A) Schematic diagram illustrates generation of Sptlc1 ECKO mice. (B) Body weight was measured at indicated time after tamoxifen injection in Ctrl and Sptlc1 ECKO mice (n=3 for Ctrl, n=3–4 for Sptlc1 ECKO). (C) Blood chemistry analysis was performed using Ctrl and Sptlc1 ECKO serum via VETSCAN VS2 Chemistry Analyzer (n=4 for Ctrl, n=3 for Sptlc1 ECKO). Statistical significance in (B) was determined by One way-ANOVA test. Statistical significance in (C) was determined by unpaired t test. Data are expressed as mean ± SD. ns, non-significant.SPTLC1 expressions are not affected in bone marrow of Sptlc1 ECKO mice.(A) Bone marrow cells isolated from Ctrl and Sptlc1 ECKO mice were analyzed by western blotting for SPTLC1 expressions. β-actin was used as an internal control (n=3 for Ctrl, n=3 for Sptlc1 ECKO). (B) Relative fold change to Ctrl samples was quantified. Statistical significance was determined by unpaired t test. Data are expressed as mean ± SD. ns, non-significant.Figure 1—figure supplement 2—source data 1.Original blots of lysates from Ctrl and Sptlc1 ECKO bone marrows.Original blots of lysates from Ctrl and Sptlc1 ECKO bone marrows.Cer and SM are reduced in Sptlc1 KO EC while they are mildly affected in non-EC.(A) Lung EC Cer with different chain lengths in Ctrl and Sptlc1 ECKO mice were measured by LC-MS/MS (n=5 for Ctrl EC, n=4 for Sptlc1 KO EC). (B) Lung EC SMs with different chain lengths in Ctrl and Sptlc1 ECKO mice were measured by LC-MS/MS (n=5 for Ctrl EC, n=4 for Sptlc1 KO EC). (C) Lung non-EC Cer with different chain lengths in Ctrl and Sptlc1 ECKO mice were measured by LC-MS/MS (n=5 for Ctrl non-EC, n=4 for Sptlc1 KO non-EC). (D) Lung non-EC SMs with different chain lengths in Ctrl and Sptlc1 ECKO mice were measured by LC-MS/MS (n=5 for Ctrl non-EC, n=4 for Sptlc1 KO non-EC). Statistical significance was determined by unpaired t test. Data are expressed as mean ± SD. *p<0.05. **p<0.01. ***p<0.001. ns, nonsignificant.eNOS expressions and phosphorylation of eNOS are not affected in Sptlc1 KO EC.(A) Lung EC isolated from Ctrl and Sptlc1 ECKO mice were analyzed by Western Blotting for eNOS, phospho-eNOS and SPTLC1 expressions. β-actin was used as an internal control (n=4 for Ctrl, n=4 for Sptlc1 ECKO). (B) Relative fold change to β-actin signal was quantified. Statistical significance was determined by unpaired t test. Data are expressed as mean ± SD. ns, non-significant.Figure 1—figure supplement 4—source data 1.Original blots of EC and non-EC from Ctrl and Sptlc1 ECKO lungs.Original blots of EC and non-EC from Ctrl and Sptlc1 ECKO lungs. less

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Murine endothelial serine palmitoyltransferase 1 (SPTLC1) is required for vascular development and systemic sphingolipid homeostasis.

In Elife on 5 October 2022 by Kuo, A., Checa, A., et al.

Fig.1.A
Fig.1.A showing Western Blotting from the publication: Murine endothelial serine palmitoyltransferase 1 (SPTLC1) is required for vascular development and systemic sphingolipid homeostasis.
  • WB
eNOS expressions and phosphorylation of eNOS are not affected in Sptlc1 KO EC.(A) Lung EC isolated from Ctrl and Sptlc1 ECKO mice were analyzed by Western Blotting for eNOS, phospho-eNOS and SPTLC1 expressions. β-actin was used as an internal control (n=4 for Ctrl, n=4 for Sptlc1 ECKO). (B) Relat... more
eNOS expressions and phosphorylation of eNOS are not affected in Sptlc1 KO EC.(A) Lung EC isolated from Ctrl and Sptlc1 ECKO mice were analyzed by Western Blotting for eNOS, phospho-eNOS and SPTLC1 expressions. β-actin was used as an internal control (n=4 for Ctrl, n=4 for Sptlc1 ECKO). (B) Relative fold change to β-actin signal was quantified. Statistical significance was determined by unpaired t test. Data are expressed as mean ± SD. ns, non-significant.Figure 1—figure supplement 4—source data 1.Original blots of EC and non-EC from Ctrl and Sptlc1 ECKO lungs.Original blots of EC and non-EC from Ctrl and Sptlc1 ECKO lungs. less

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Nrg1β Released in Remote Ischemic Preconditioning Improves Myocardial Perfusion and Decreases Ischemia/Reperfusion Injury via ErbB2-Mediated Rescue of Endothelial Nitric Oxide Synthase and Abrogation of Trx2 Autophagy.

In Arterioscler Thromb Vasc Biol on 1 August 2021 by Kundumani-Sridharan, V., Subramani, J., et al.

Fig.2.K
Fig.2.K showing Immunohistochemistry-immunofluorescence in a Homo sapiens (Human) sample from the publication: Nrg1β Released in Remote Ischemic Preconditioning Improves Myocardial Perfusion and Decreases Ischemia/Reperfusion Injury via ErbB2-Mediated Rescue of Endothelial Nitric Oxide Synthase and Abrogation of Trx2 Autophagy.
  • IHC-IF
  • Homo sapiens (Human)
Remote ischemic preconditioning (RIPC) improved cardiac perfusion via Nrg1 (neuregulin 1)-dependent eNOS (endothelial nitric oxide synthase) activation. A, Mice were subjected to sham or myocardial ischemia/reperfusion (I/R) or RIPC+I/R surgery, and at the end of the reperfusion period, the nonta... more
Remote ischemic preconditioning (RIPC) improved cardiac perfusion via Nrg1 (neuregulin 1)-dependent eNOS (endothelial nitric oxide synthase) activation. A, Mice were subjected to sham or myocardial ischemia/reperfusion (I/R) or RIPC+I/R surgery, and at the end of the reperfusion period, the nontargeted contrast agent was injected to mice via femoral vein while collecting B-mode/contrast mode images of the heart from long-axis view using Vevo 3100. To block the function of circulating Nrg1 in mice, neutralizing anti-Nrg1 antibodies (150 µg/kg) was administered to mice before RIPC and then subjected to RIPC+I/R injury. B, Contrast intensities in the left ventricle (LV) anterior wall downstream to the ligation site were quantitated and plotted over time. Loss of peak height and the rightward shift of peak indicates hypoperfusion. C, From the rate of contrast agent intensity change, the perfusion index was calculated. Percentage of perfusion was calculated based on the perfusion index assuming its level in the sham group is 100% and blotted as a bar graph (n=5) and shown as a heat map over the anterior LV in A. Values are means±SEM (n=5 mice). *P<0.01 vs sham, **P<0.01 vs I/R, §P<0.01 vs RIPC+I/R (ANOVA). D, Left coronary artery from mice were isolated, and after indicated treatments, NO formation was detected by electron spin resonance (EPR) spectrometry using NO spin trap Fe2+-(N-methyl-D-glucamine dithiocarbamate) 2 (Fe-[MGD]2) as described in methods. E, NO-Fe(MGD) 2 spin count was quantified and plotted as a bar graph (n=5). *P<0.01 vs sham; **P<0.01 vs I/R (ANOVA). F–H, HMVECs were exposed to 2 h hypoxia followed by 1-hour reoxygenation, and the medium was collected (preconditioned medium [PM]). HCAECs were treated with PM for the indicated period, lysed, and cell lysates were collected. An equal amount of cell lysates were analyzed for eNOS activity (F) and plotted as a bar graph (n=3). *P<0.01 vs control (ANOVA). An equal amount of cell lysates were analyzed for eNOS phosphorylation at Ser 1177 by Western blotting using its phospho-specific antibodies and normalized with total eNOS levels (G), or the cell lysates were immunoprecipitated with anti-eNOS antibodies, and the immunoprecipitates were analyzed by Western blotting using anti-phospho-tyrosine specific antibodies (H). I–J, To determine the role of HMVEC-released Nrg1 in PM in the activation of eNOS, the collected PM was pretreated with neutralizing anti-Nrg1 antibodies and incubated with human coronary artery endothelial cell (HCAEC) for 1 h, and cell lysates were collected and analyzed for eNOS tyrosine phosphorylation by Western blotting (I) or eNOS activity (J). K, Mice were injected control IgG or neutralizing anti-Nrg1 antibodies via the tail vein. After 24 h, they were subjected to hindlimb RIPC followed by myocardial I/R, and heart from these mice was sectioned below the LAD ligation point and analyzed for eNOS phosphorylation at Tyr81 by immunofluorescence staining. Scale bar=50 mm. L, Green fluorescence localized to the coronary artery endothelial cells were quantitated (n=5), and the results were plotted as a bar graph. *P<0.05 vs I/R, **P<0.05 vs control IgG+I/R (ANOVA). less

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Latent HIV-Exosomes Induce Mitochondrial Hyperfusion Due to Loss of Phosphorylated Dynamin-Related Protein 1 in Brain Endothelium.

In Mol Neurobiol on 1 June 2021 by Chandra, P. K., Rutkai, I., et al.

Fig.6.B
Fig.6.B showing Western Blotting from the publication: Latent HIV-Exosomes Induce Mitochondrial Hyperfusion Due to Loss of Phosphorylated Dynamin-Related Protein 1 in Brain Endothelium.
  • WB
HIV-exosomes contain HIV-1 Tat protein and viral Tat protein reduce p-DRP1, and p-eNOS expression in primary HBMVECs. a HIV-1 Tat protein in U1 Exo, U937 Exo, and U1 cell lysate was detected (20 μg protein) by western blotting. Recombinant HIV-Tat protein (50 ng) was used as positive control. b R... more
HIV-exosomes contain HIV-1 Tat protein and viral Tat protein reduce p-DRP1, and p-eNOS expression in primary HBMVECs. a HIV-1 Tat protein in U1 Exo, U937 Exo, and U1 cell lysate was detected (20 μg protein) by western blotting. Recombinant HIV-Tat protein (50 ng) was used as positive control. b Representative western blot of p-eNOS and total eNOS in cells exposed to U1 Exo. c Quantitative analysis of p-eNOS and total eNOS. d Representative western blot, and e quantitative analysis of p-DRP1 and total DRP1 in cells exposed to recombinant HIV-Tat protein. f Representative western blot, and g quantitative analysis of p-eNOS and total eNOS in cells exposed to recombinant HIV-Tat protein. Error bars show mean ± SD, n = 3 independent experiments, and significant changes are presented as p values (*p < 0.05) less

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Latent HIV-Exosomes Induce Mitochondrial Hyperfusion Due to Loss of Phosphorylated Dynamin-Related Protein 1 in Brain Endothelium.

In Mol Neurobiol on 1 June 2021 by Chandra, P. K., Rutkai, I., et al.

Fig.6.T
Fig.6.T showing Western Blotting from the publication: Latent HIV-Exosomes Induce Mitochondrial Hyperfusion Due to Loss of Phosphorylated Dynamin-Related Protein 1 in Brain Endothelium.
  • WB
HIV-exosomes contain HIV-1 Tat protein and viral Tat protein reduce p-DRP1, and p-eNOS expression in primary HBMVECs. a HIV-1 Tat protein in U1 Exo, U937 Exo, and U1 cell lysate was detected (20 μg protein) by western blotting. Recombinant HIV-Tat protein (50 ng) was used as positive control. b R... more
HIV-exosomes contain HIV-1 Tat protein and viral Tat protein reduce p-DRP1, and p-eNOS expression in primary HBMVECs. a HIV-1 Tat protein in U1 Exo, U937 Exo, and U1 cell lysate was detected (20 μg protein) by western blotting. Recombinant HIV-Tat protein (50 ng) was used as positive control. b Representative western blot of p-eNOS and total eNOS in cells exposed to U1 Exo. c Quantitative analysis of p-eNOS and total eNOS. d Representative western blot, and e quantitative analysis of p-DRP1 and total DRP1 in cells exposed to recombinant HIV-Tat protein. f Representative western blot, and g quantitative analysis of p-eNOS and total eNOS in cells exposed to recombinant HIV-Tat protein. Error bars show mean ± SD, n = 3 independent experiments, and significant changes are presented as p values (*p < 0.05) less

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BReast CAncer susceptibility gene 2 deficiency exacerbates oxidized LDL-induced DNA damage and endothelial apoptosis.

In Physiol Rep on 1 July 2020 by Singh, S., Nguyen, H., et al.

Fig.5.D
Fig.5.D showing Western Blotting from the publication: BReast CAncer susceptibility gene 2 deficiency exacerbates oxidized LDL-induced DNA damage and endothelial apoptosis.
  • WB
Endothelial cell‐specific loss of BRCA2 exacerbates oxLDL‐induced endothelial dysfunction. (a) Representative micrographs and (b) quantification showing capillary‐like tube formation 5 hr post‐oxLDL (100 μg/ml) treatment to scrambled control‐ or siBRCA2‐ transfected HUVECs for 48 hr. Six‐wells/gr... more
Endothelial cell‐specific loss of BRCA2 exacerbates oxLDL‐induced endothelial dysfunction. (a) Representative micrographs and (b) quantification showing capillary‐like tube formation 5 hr post‐oxLDL (100 μg/ml) treatment to scrambled control‐ or siBRCA2‐ transfected HUVECs for 48 hr. Six‐wells/group. ***p < .001 versus scrambled control, $p < .05 versus scrambled control + oxLDL. (c) qPCR for eNOS and (d) immunoblotting for (p)eNOS, eNOS, and GAPDH in HUVECs transfected with either scrambled control or siBRCA2 for 48 hr and then treated with oxLDL (100 μg/ml) for 24 hr. *, ***p < .05, 0.001 versus scrambled control, $$p < .01 versus scrambled control + oxLDL, p = .078 versus scrambled control less

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Nerve damage induced skeletal muscle atrophy is associated with increased accumulation of intramuscular glucose and polyol pathway intermediates.

In Sci Rep on 5 February 2020 by Langer, H. T., Afzal, S., et al.

Fig.3.C
Fig.3.C showing Western Blotting from the publication: Nerve damage induced skeletal muscle atrophy is associated with increased accumulation of intramuscular glucose and polyol pathway intermediates.
  • WB
Protein levels associated with membrane damage, cytoskeletal integrity, inflammation and autophagy are increased in nerve damaged TA, while oxidative activity appears to be decreased. Rat TA was subjected to western blot anaylsis of proteins associated with muscle damage. (A) Dysferlin, AnnexinA2... more
Protein levels associated with membrane damage, cytoskeletal integrity, inflammation and autophagy are increased in nerve damaged TA, while oxidative activity appears to be decreased. Rat TA was subjected to western blot anaylsis of proteins associated with muscle damage. (A) Dysferlin, AnnexinA2, Cyclophilin, MLIM and LC3B are significantly upregulated in nerve damaged TA, while NOS (pan) levels are substantially decreased with nerve damage. (B) Bands corresponding to the western blot quantification in (A) (n = 5). The cropped pictures displayed above show the entire width of the membrane and all samples included for several different proteins. In order to allow for probing of multiple proteins with distinct molecular weight simultaneously, the membranes where cut horizontally at the appropriate heights. All band intensities were normalized to total protein content of the gel as assessed via coomassie blue staining. (C) Bands corresponding to the western blot quantification in (A) (n = 6). These blots were added during a later stage of the manuscript. All band intensities were normalized to total protein content of the gel as assessed via stain-free technology. *p < 0.05, **p < 0.01, ***p < 0.001. less

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Myofibrillar function differs markedly between denervated and dexamethasone-treated rat skeletal muscles: Role of mechanical load.

In PLoS One on 10 October 2019 by Yamada, T., Ashida, Y., et al.

Fig.4.E
Fig.4.E showing Western Blotting from the publication: Myofibrillar function differs markedly between denervated and dexamethasone-treated rat skeletal muscles: Role of mechanical load.
  • WB
Neuromuscular electrical stimulation (ES) training prevents the upregulation of ROS/RNS producing proteins in DEN muscles.Representative immunoblots illustrating the levels of NADPH oxidase (NOX2/gp91phox), NOX4 (A, B), neuronal nitric oxide synthase (nNOS), and endotherial NOS (eNOS) (E, F) in w... more
Neuromuscular electrical stimulation (ES) training prevents the upregulation of ROS/RNS producing proteins in DEN muscles.Representative immunoblots illustrating the levels of NADPH oxidase (NOX2/gp91phox), NOX4 (A, B), neuronal nitric oxide synthase (nNOS), and endotherial NOS (eNOS) (E, F) in whole muscle protein of gastrocnemius muscles from control (CNT), denervation (DEN), and dexamethasone (DEX) rats with or without ES training. The levels of NOX2/gp91phox (C), NOX4 (D), nNOS (G), and eNOS (H) expression were normalized to the total protein content seen in the stain free images. Bars show the mean and SEM results from 5–9 muscles per group. Statistical significance was set at P<0.05: main effect of *treatment (i.e., DEN and DEX) and #ES; †interaction of treatment and ES; difference versus aCNT and bDEN. less

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Peroxynitrite supports a metabolic reprogramming in merlin-deficient Schwann cells and promotes cell survival.

In J Biol Chem on 26 July 2019 by Pestoni, J. C., Klingeman Plati, S., et al.

Fig.1.E
Fig.1.E showing Western Blotting from the publication: Peroxynitrite supports a metabolic reprogramming in merlin-deficient Schwann cells and promotes cell survival.
  • WB
Tyrosine nitration is increased in NF2 schwannoma cells.A, representative IR Western blots showing nitrotyrosine staining of vestibular schwannoma samples from three NF2 patients (VS 1 to 3), and human (I) and mouse (MSC) WT- and MD-Schwann cells. α-Tubulin and β-actin were used as loading contro... more
Tyrosine nitration is increased in NF2 schwannoma cells.A, representative IR Western blots showing nitrotyrosine staining of vestibular schwannoma samples from three NF2 patients (VS 1 to 3), and human (I) and mouse (MSC) WT- and MD-Schwann cells. α-Tubulin and β-actin were used as loading controls. B, quantitation of nitrotyrosine levels in HSC (n = 4). C, loss of merlin expression in both human and mouse MD-Schwann confirmed by IR Western blotting. D–F, representative Western blots showing the levels of: D, nNOS (n = 3–4); E, inducible and endothelial NOS (iNOS and eNOS, respectively, n = 4–5); and F, MnSOD (n = 7–8) in human and mouse WT- and MD-Schwann cells. Homogenate from human WT-Schwann cells treated with 1 μm lipopolysaccharide (LPS) for 24 h was used as positive control for iNOS expression, and brain homogenate as control for nNOS and eNOS expression. Columns represent the mean ± S.D. of the respective Western blotting band quantitation normalized against α-tubulin or β-actin and expressed relative to WT-Schwann cells. *, p < 0.05 versus WT by either Student's t test or Mann-Whitney test. less

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DDAH1 regulates apoptosis and angiogenesis in human fetal pulmonary microvascular endothelial cells.

In Physiol Rep on 1 July 2019 by Trittmann, J. K., Almazroue, H., et al.

Fig.1.C
Fig.1.C showing Western Blotting in a Homo sapiens (Human) sample from the publication: DDAH1 regulates apoptosis and angiogenesis in human fetal pulmonary microvascular endothelial cells.
  • WB
  • Homo sapiens (Human)
siRNA‐mediated knockdown of DDAH1 in hfPMVEC resulted in less NO production with no significant change in eNOS protein levels. (A) DDAH1 siRNA transfection of hfPMVECs resulted in effective knockdown of DDAH1 protein levels. hfPMVECs were transfected with either scramble siRNA or DDAH1 siRNA for ... more
siRNA‐mediated knockdown of DDAH1 in hfPMVEC resulted in less NO production with no significant change in eNOS protein levels. (A) DDAH1 siRNA transfection of hfPMVECs resulted in effective knockdown of DDAH1 protein levels. hfPMVECs were transfected with either scramble siRNA or DDAH1 siRNA for 24 h, recovered for 24 h, and protein isolated. Representative western blot and bar graph of densitometry data for DDAH1 protein levels normalized to β‐actin (n = 6 in each group). *P < 0.05, DDAH1 siRNA different from scramble. (B) DDAH1 knockdown with siRNA resulted in lower NO production than scramble transfected cells. hfPMVECs were transfected with either scramble siRNA or DDAH1 siRNA for 24 h, washed and allowed to recover for 24 h. Cell were incubated for an additional 24 h and cell media harvested for determination of nitrites using a chemiluminescence NO analyzer. Nitrite levels were normalized to protein concentration in each plate (n = 3 in each group). *P < 0.05, DDAH1 siRNA different from scramble. (C) DDAH1 siRNA transfection of hfPMVECs resulted in no significant change of eNOS protein levels. The protein isolated under B above was used in western blotting for eNOS protein levels. Representative western blot and bar graph of densitometry data for eNOS protein level normalized to β‐actin (n = 6 in each group). less

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RELi protocol: Optimization for protein extraction from white, brown and beige adipose tissues.

In MethodsX on 14 May 2019 by Diaz Marin, R., Crespo-Garcia, S., et al.

Fig.4.C
Fig.4.C showing Western Blotting in a Mus musculus (House mouse) sample from the publication: RELi protocol: Optimization for protein extraction from white, brown and beige adipose tissues.
  • WB
  • Mus musculus (House mouse)
Assessment of quality and uniformity of adipose tissue proteins extracted with the CST and RELi methods. A–C) Western blot analysis of adipose tissue cellular compartments: Cytosol (β-actin ˜42 KDa), Stromal vascular fraction (eNOS ˜140 KDa), Mitochondria (BCL2 ˜26 KDa), Lipid associated proteins... more
Assessment of quality and uniformity of adipose tissue proteins extracted with the CST and RELi methods. A–C) Western blot analysis of adipose tissue cellular compartments: Cytosol (β-actin ˜42 KDa), Stromal vascular fraction (eNOS ˜140 KDa), Mitochondria (BCL2 ˜26 KDa), Lipid associated proteins (Perilipin A ˜68 KDa), Nuclear protein (Histone H3 ˜15 KDa) in WAT, BgAT and BAT extracts; C (Cytosol), SVF (Stromal vascular fraction), M (Mitochondria), LD (Lipid droplet associated), N (Nuclear). less

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RELi protocol: Optimization for protein extraction from white, brown and beige adipose tissues.

In MethodsX on 14 May 2019 by Diaz Marin, R., Crespo-Garcia, S., et al.

Fig.4.B
Fig.4.B showing Western Blotting in a Mus musculus (House mouse) sample from the publication: RELi protocol: Optimization for protein extraction from white, brown and beige adipose tissues.
  • WB
  • Mus musculus (House mouse)
Assessment of quality and uniformity of adipose tissue proteins extracted with the CST and RELi methods. A–C) Western blot analysis of adipose tissue cellular compartments: Cytosol (β-actin ˜42 KDa), Stromal vascular fraction (eNOS ˜140 KDa), Mitochondria (BCL2 ˜26 KDa), Lipid associated proteins... more
Assessment of quality and uniformity of adipose tissue proteins extracted with the CST and RELi methods. A–C) Western blot analysis of adipose tissue cellular compartments: Cytosol (β-actin ˜42 KDa), Stromal vascular fraction (eNOS ˜140 KDa), Mitochondria (BCL2 ˜26 KDa), Lipid associated proteins (Perilipin A ˜68 KDa), Nuclear protein (Histone H3 ˜15 KDa) in WAT, BgAT and BAT extracts; C (Cytosol), SVF (Stromal vascular fraction), M (Mitochondria), LD (Lipid droplet associated), N (Nuclear). less

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RELi protocol: Optimization for protein extraction from white, brown and beige adipose tissues.

In MethodsX on 14 May 2019 by Diaz Marin, R., Crespo-Garcia, S., et al.

Fig.4.A
Fig.4.A showing Western Blotting in a Mus musculus (House mouse) sample from the publication: RELi protocol: Optimization for protein extraction from white, brown and beige adipose tissues.
  • WB
  • Mus musculus (House mouse)
Assessment of quality and uniformity of adipose tissue proteins extracted with the CST and RELi methods. A–C) Western blot analysis of adipose tissue cellular compartments: Cytosol (β-actin ˜42 KDa), Stromal vascular fraction (eNOS ˜140 KDa), Mitochondria (BCL2 ˜26 KDa), Lipid associated proteins... more
Assessment of quality and uniformity of adipose tissue proteins extracted with the CST and RELi methods. A–C) Western blot analysis of adipose tissue cellular compartments: Cytosol (β-actin ˜42 KDa), Stromal vascular fraction (eNOS ˜140 KDa), Mitochondria (BCL2 ˜26 KDa), Lipid associated proteins (Perilipin A ˜68 KDa), Nuclear protein (Histone H3 ˜15 KDa) in WAT, BgAT and BAT extracts; C (Cytosol), SVF (Stromal vascular fraction), M (Mitochondria), LD (Lipid droplet associated), N (Nuclear). less

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Reduced caveolae density in arteries of SHR contributes to endothelial dysfunction and ROS production.

In Sci Rep on 30 April 2019 by Potje, S. R., Grando, M. D., et al.

Fig.8.A
Fig.8.A showing Western Blotting from the publication: Reduced caveolae density in arteries of SHR contributes to endothelial dysfunction and ROS production.
  • WB
(A,C) Representative western blot of eNOS dimer/monomer distribution in unboiled samples (top blot) and GAPDH or α-tubulin (bottom blot; loading control) in human umbilical vein endothelial cells (HUVECs) treated with methyl-β-cyclodextrin (10 mM dextrin for 60 min) or not (Control). (B,D) Basal ... more
(A,C) Representative western blot of eNOS dimer/monomer distribution in unboiled samples (top blot) and GAPDH or α-tubulin (bottom blot; loading control) in human umbilical vein endothelial cells (HUVECs) treated with methyl-β-cyclodextrin (10 mM dextrin for 60 min) or not (Control). (B,D) Basal changes in eNOS dimer and monomer induced by dextrin, in the presence of tetrahydrobiopterin (BH4, 100 μM), L-arginine (L-arg, 1 mM), or BH4 plus L-arg or not. We processed images of blots changing brightness and contrast and we applied equally over the entire image and controls. (E,F) NO determination with the DAF-2DA probe in HUVECs treated with dextrin or not (Control) in the presence of BH4, L-arg, L-NAME (1 mM), or BH4 plus L-arg. (G) Reactive oxygen species (ROS) detected with the DHE probe in the presence of BH4, L-arg, BH4 plus L-arg, L-NAME, or A23187 (5 μM) in HUVECs treated with dextrin or not. Bars are reported as the mean ± SEM of independent experiments (n = 4–5). Values of blank were not use to perform statistics. *p < 0.05 statistical differences between Dextrin versus control group. #p < 0.05 between dextrin (in the presence of BH4, or L-arg, or BH4 plus L-arg) and only BH4versus Dextrin group. &p < 0.05 between L-NAME, dextrin plus L-NAME or A23187versus Control group. less

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Reduced caveolae density in arteries of SHR contributes to endothelial dysfunction and ROS production.

In Sci Rep on 30 April 2019 by Potje, S. R., Grando, M. D., et al.

Fig.8.C
Fig.8.C showing Western Blotting from the publication: Reduced caveolae density in arteries of SHR contributes to endothelial dysfunction and ROS production.
  • WB
(A,C) Representative western blot of eNOS dimer/monomer distribution in unboiled samples (top blot) and GAPDH or α-tubulin (bottom blot; loading control) in human umbilical vein endothelial cells (HUVECs) treated with methyl-β-cyclodextrin (10 mM dextrin for 60 min) or not (Control). (B,D) Basal ... more
(A,C) Representative western blot of eNOS dimer/monomer distribution in unboiled samples (top blot) and GAPDH or α-tubulin (bottom blot; loading control) in human umbilical vein endothelial cells (HUVECs) treated with methyl-β-cyclodextrin (10 mM dextrin for 60 min) or not (Control). (B,D) Basal changes in eNOS dimer and monomer induced by dextrin, in the presence of tetrahydrobiopterin (BH4, 100 μM), L-arginine (L-arg, 1 mM), or BH4 plus L-arg or not. We processed images of blots changing brightness and contrast and we applied equally over the entire image and controls. (E,F) NO determination with the DAF-2DA probe in HUVECs treated with dextrin or not (Control) in the presence of BH4, L-arg, L-NAME (1 mM), or BH4 plus L-arg. (G) Reactive oxygen species (ROS) detected with the DHE probe in the presence of BH4, L-arg, BH4 plus L-arg, L-NAME, or A23187 (5 μM) in HUVECs treated with dextrin or not. Bars are reported as the mean ± SEM of independent experiments (n = 4–5). Values of blank were not use to perform statistics. *p < 0.05 statistical differences between Dextrin versus control group. #p < 0.05 between dextrin (in the presence of BH4, or L-arg, or BH4 plus L-arg) and only BH4versus Dextrin group. &p < 0.05 between L-NAME, dextrin plus L-NAME or A23187versus Control group. less

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Long-Term Regulation of Excitation-Contraction Coupling and Oxidative Stress in Cardiac Myocytes by Pirfenidone.

In Front Physiol on 9 January 2019 by Monsalvo-Villegas, A., Osornio-Garduño, D. S., et al.

Fig.7.B
Fig.7.B showing Western Blotting in a Rattus norvegicus (Rat) sample from the publication: Long-Term Regulation of Excitation-Contraction Coupling and Oxidative Stress in Cardiac Myocytes by Pirfenidone.
  • WB
  • Rattus norvegicus (Rat)
Dual regulation of nNOS expression by PFD. (A) Normalized immunoreactivity that was detected in myocytes exposed to primary antibodies against NCX, eNOS, and nNOS (β-tubulin was the loading control). Bars represent the average values from a total of three to four independent experiments. ∗p < 0.0... more
Dual regulation of nNOS expression by PFD. (A) Normalized immunoreactivity that was detected in myocytes exposed to primary antibodies against NCX, eNOS, and nNOS (β-tubulin was the loading control). Bars represent the average values from a total of three to four independent experiments. ∗p < 0.05. (B) Examples of western blots that were analyzed to generate data shown in (A). less

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Endothelial AMP-Activated Kinase α1 Phosphorylates eNOS on Thr495 and Decreases Endothelial NO Formation.

In Int J Mol Sci on 13 September 2018 by Zippel, N., Loot, A. E., et al.

Fig.5.B
Fig.5.B showing Western Blotting from the publication: Endothelial AMP-Activated Kinase α1 Phosphorylates eNOS on Thr495 and Decreases Endothelial NO Formation.
  • WB
Effect of AMPK activation on eNOS activity and phosphorylation. Cultured human endothelial cells were incubated with solvent (Sol), thiopental (1 mmol/L, 5–60 min), or bradykinin (BK; 1 µmol/L, 2 min). Thereafter, the phosphorylation of (A) AMPK and (B) phosphorylation of eNOS at Ser1177 and Thr4... more
Effect of AMPK activation on eNOS activity and phosphorylation. Cultured human endothelial cells were incubated with solvent (Sol), thiopental (1 mmol/L, 5–60 min), or bradykinin (BK; 1 µmol/L, 2 min). Thereafter, the phosphorylation of (A) AMPK and (B) phosphorylation of eNOS at Ser1177 and Thr495 were assessed. Bar graphs summarize the data obtained in four to five different cell batches; * p < 0.05, ** p < 0.01 versus solvent treatment. less

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Electrical Stimulation Prevents Preferential Skeletal Muscle Myosin Loss in Steroid-Denervation Rats.

In Front Physiol on 28 August 2018 by Yamada, T., Himori, K., et al.

Fig.6.A
Fig.6.A showing Western Blotting in a Rattus norvegicus (Rat) sample from the publication: Electrical Stimulation Prevents Preferential Skeletal Muscle Myosin Loss in Steroid-Denervation Rats.
  • WB
  • Rattus norvegicus (Rat)
Expressions of redox-related proteins are upregulated in S-D muscles. Representative immunoblots illustrating the levels of NOX (NOX2/gp91phox), NOX4 (A), neuronal NOS (nNOS), and endotherial NOS (eNOS) (D) of gastrocnemius muscles in control (CNT) and steroid-denervation (S-D) rats with or witho... more
Expressions of redox-related proteins are upregulated in S-D muscles. Representative immunoblots illustrating the levels of NOX (NOX2/gp91phox), NOX4 (A), neuronal NOS (nNOS), and endotherial NOS (eNOS) (D) of gastrocnemius muscles in control (CNT) and steroid-denervation (S-D) rats with or without ES treatment. The levels of NOX2/gp91phox(B), NOX4 (C), nNOS (E), and eNOS (F) expressions were normalized to the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) content. Bars show the mean and SEM results from 5 to 6 muscles per group. ∗P < 0.05 vs. CNT, #P < 0.05 vs. S-D. less

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Electrical Stimulation Prevents Preferential Skeletal Muscle Myosin Loss in Steroid-Denervation Rats.

In Front Physiol on 28 August 2018 by Yamada, T., Himori, K., et al.

Fig.6.D
Fig.6.D showing Western Blotting in a Rattus norvegicus (Rat) sample from the publication: Electrical Stimulation Prevents Preferential Skeletal Muscle Myosin Loss in Steroid-Denervation Rats.
  • WB
  • Rattus norvegicus (Rat)
Expressions of redox-related proteins are upregulated in S-D muscles. Representative immunoblots illustrating the levels of NOX (NOX2/gp91phox), NOX4 (A), neuronal NOS (nNOS), and endotherial NOS (eNOS) (D) of gastrocnemius muscles in control (CNT) and steroid-denervation (S-D) rats with or witho... more
Expressions of redox-related proteins are upregulated in S-D muscles. Representative immunoblots illustrating the levels of NOX (NOX2/gp91phox), NOX4 (A), neuronal NOS (nNOS), and endotherial NOS (eNOS) (D) of gastrocnemius muscles in control (CNT) and steroid-denervation (S-D) rats with or without ES treatment. The levels of NOX2/gp91phox(B), NOX4 (C), nNOS (E), and eNOS (F) expressions were normalized to the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) content. Bars show the mean and SEM results from 5 to 6 muscles per group. ∗P < 0.05 vs. CNT, #P < 0.05 vs. S-D. less

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Boysenberry polyphenol inhibits endothelial dysfunction and improves vascular health.

In PLoS One on 15 August 2018 by Furuuchi, R., Shimizu, I., et al.

Fig.2.D
Fig.2.D showing Western Blotting from the publication: Boysenberry polyphenol inhibits endothelial dysfunction and improves vascular health.
  • WB
Boysenberry polyphenol inhibits metabolic stress-induced endothelial dysfunction.Human umbilical vein endothelial cells (HUVECs) were treated with BSA (Con group), palmitic acid (500 μM) (PA group(6hr)), or PA (500 μM) + BP (10 μg/ml) (PA+BP group). A. DHE staining of HUVECs in Con, PA, and PA+BP... more
Boysenberry polyphenol inhibits metabolic stress-induced endothelial dysfunction.Human umbilical vein endothelial cells (HUVECs) were treated with BSA (Con group), palmitic acid (500 μM) (PA group(6hr)), or PA (500 μM) + BP (10 μg/ml) (PA+BP group). A. DHE staining of HUVECs in Con, PA, and PA+BP groups (Scale bar = 100 μm). The right graph shows the relative fluorescence intensity of DHE (n = 4, 4, and 4). B. Western blot analysis of p53 expression in HUVECs. The right panel displays quantification of p53 relative to the β-actin loading control (n = 6, 6, and 6). C. DAR-4M staining of HUVECs to detect nitric oxide (Scale bar = 100 μm). The right graph shows the relative fluorescence intensity (n = 4, 4, and 4). D. Western blot analysis of eNOS dimer, eNOS monomer, and α-tubulin in HUVECs. The right panel shows quantification of the eNOS dimer/ monomer ratio adjusted for α-tubulin (n = 3, 3, and 3). Data were analyzed by 2-way ANOVA, followed by Tukey’s multiple comparison test. *P < 0.05; **P < 0.01. Values represent the mean ± SEM. less

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