Journal: Experimental and Therapeutic Medicine

Article Title: Sinomenine activation of Nrf2 signaling prevents inflammation and cerebral injury in a mouse model of ischemic stroke

doi: 10.3892/etm.2021.10079

Figure Lengend Snippet: SINO treatment relieves MCAO-induced cerebral injuries. (A) Representative micrographs of H&E staining of brain sections (x20 magnification; scale bar, 50 µm). (B) Brain water content analysis in brain tissues from Sham (sham-operated, n=6), SINO (treatment only, n=6), MCAO (MCAO-operated, n=6) and SINO/MCAO (MCAO operated plus SINO treatment, n=6). (C) Western blot analysis of n-Nrf2, HO-1 and NQO1 protein expression levels from brain tissue. The samples from two randomly selected brains in each group were presented. (D) Quantification of brain protein expression levels. All the experiments were repeated at least three times. The data are presented as the mean ± SEM. * P<0.05 and ** P<0.01 vs. Sham; # P<0.05 and ## P<0.01 vs. MCAO. HO-1, heme oxygenase-1; MCAO, middle cerebral artery occlusion; n-Nrf2, nuclear-nuclear factor-erythroid 2-related factor; NQO1, NAD(P)H: Quinoneoxidoreductase 1; SINO, sinomenine.

Article Snippet: The following antibodies were used in the present study: SINO (MedChemExpress); ML385 (MedChemExpress); Nrf2, Keap1, NAD(P)H: Quinoneoxidoreductase (NQO1), Lamin B and HO-1 antibodies (Abcam); β-actin antibodies (Bioworld Technology, Inc.); NF-κB p65, Phosphorylated (p-)IκBα and IκBα antibodies (Cell Signaling Technology, Inc.); HRP-conjugated secondary antibodies (Bioworld Technology, Inc.); and Dylight488 conjugated goat anti-rabbit IgG secondary antibodies (Bioworld Technology, Inc.).

Techniques: Staining, Western Blot, Expressing

Journal: Experimental and Therapeutic Medicine

Article Title: Sinomenine activation of Nrf2 signaling prevents inflammation and cerebral injury in a mouse model of ischemic stroke

doi: 10.3892/etm.2021.10079

Figure Lengend Snippet: SINO treatment activates the Nrf2 signaling pathway. (A) BV2 cells were treated with a serial of doses of SINO for 24 h. HO-1 and NQO1 protein expression levels were assayed using western blotting. (B) BV2 cells were treated with SINO (200 µM) for various durations. HO-1 and NQO1 protein expression levels were assayed using western blotting. (C) BV2 cells were treated with SINO (200 µM) for 4 h. n-Nrf2 and t-Nrf2 protein expression levels were assayed using western blotting. Nuclear protein Lamin B and β-actin were used as controls. (D) BV2 cells were treated with serial doses of SINO for 24 h. The mRNA expression levels of HO-1 and NQO1 were measured using reverse transcription-semi-quantitative PCR. (E) Cell viability assay. BV2 cells were treated with various doses of SINO for 24 h. The cell viability was assayed by a CCK-8 kit. All the experiments were repeated at least three times. * P<0.05 and ** P<0.01 vs. Control HO-1, heme oxygenase-1; NQO1, NAD(P)H: Quinoneoxidoreductase 1; n-, nuclear; Nrf2; nuclear factor-erythroid 2-related factor; SINO, sinomenine; t-, total.

Article Snippet: The following antibodies were used in the present study: SINO (MedChemExpress); ML385 (MedChemExpress); Nrf2, Keap1, NAD(P)H: Quinoneoxidoreductase (NQO1), Lamin B and HO-1 antibodies (Abcam); β-actin antibodies (Bioworld Technology, Inc.); NF-κB p65, Phosphorylated (p-)IκBα and IκBα antibodies (Cell Signaling Technology, Inc.); HRP-conjugated secondary antibodies (Bioworld Technology, Inc.); and Dylight488 conjugated goat anti-rabbit IgG secondary antibodies (Bioworld Technology, Inc.).

Techniques: Expressing, Western Blot, Reverse Transcription, Real-time Polymerase Chain Reaction, Viability Assay, CCK-8 Assay, Control

Journal: Experimental and Therapeutic Medicine

Article Title: Sinomenine activation of Nrf2 signaling prevents inflammation and cerebral injury in a mouse model of ischemic stroke

doi: 10.3892/etm.2021.10079

Figure Lengend Snippet: SINO treatment regulates microglia polarization and inflammation in an Nrf2-dependent manner. BV2 cells were stimulated with OGD for 4 h followed by treatment with SINO (200 µM) for 12 h. The mRNA expression levels of (A) M1 markers (IL-6/NOS2) and (B) M2 markers (IL-10/Arg-1) were determined using RT-sqPCR. BV2 cells were pretreated with ML385 (5 µM) for 48 h to inhibit Nrf2 expression. Cells were then stimulated with OGD for 4 h followed by treatment with SINO (200 µM) for 12 h. The mRNA expression levels of (C) M1 and (D) M2 markers were measured using RT-sqPCR. The data are presented as the mean ± SEM of three independent experiments. * P<0.05 and ** P<0.01 vs. Control; ## P<0.01 vs. OGD. Arg-1, arginase-1; NOS2, nitric oxide synthase 2; OGD, oxygen and glucose deprivation; SINO, sinomenine; RT-sqPCR, reverse transcription-semi-quantitative PCR.

Article Snippet: The following antibodies were used in the present study: SINO (MedChemExpress); ML385 (MedChemExpress); Nrf2, Keap1, NAD(P)H: Quinoneoxidoreductase (NQO1), Lamin B and HO-1 antibodies (Abcam); β-actin antibodies (Bioworld Technology, Inc.); NF-κB p65, Phosphorylated (p-)IκBα and IκBα antibodies (Cell Signaling Technology, Inc.); HRP-conjugated secondary antibodies (Bioworld Technology, Inc.); and Dylight488 conjugated goat anti-rabbit IgG secondary antibodies (Bioworld Technology, Inc.).

Techniques: Expressing, Control, Reverse Transcription, Real-time Polymerase Chain Reaction

Journal: Experimental and Therapeutic Medicine

Article Title: Sinomenine activation of Nrf2 signaling prevents inflammation and cerebral injury in a mouse model of ischemic stroke

doi: 10.3892/etm.2021.10079

Figure Lengend Snippet: SINO treatment regulates microglia inflammation in an Nrf2-dependent manner. (A) BV2 cells were pretreated with ML385 (5 µM) for 48 h to inhibit Nrf2 expression. Cells were then stimulated with OGD for 4 h followed by treatment with SINO (200 µM) for 12 h. The protein expression levels of p-IκBα and total IκBα were analyzed using western blotting. (B) Immuno-fluorescent staining of cells was performed using an NF-κB p65 primary antibody and Dylight488 conjugated secondary antibody (middle row of panels). The cells were also stained with DAPI (top row of panels) and merged with NF-κB images (lower row of panels). The data are presented as the mean ± SEM of three independent experiments. ** P<0.01 vs. Control; ## P<0.01 vs. OGD. OGD, oxygen and glucose deprivation; p-, phosphorylated; SINO, sinomenine.

Article Snippet: The following antibodies were used in the present study: SINO (MedChemExpress); ML385 (MedChemExpress); Nrf2, Keap1, NAD(P)H: Quinoneoxidoreductase (NQO1), Lamin B and HO-1 antibodies (Abcam); β-actin antibodies (Bioworld Technology, Inc.); NF-κB p65, Phosphorylated (p-)IκBα and IκBα antibodies (Cell Signaling Technology, Inc.); HRP-conjugated secondary antibodies (Bioworld Technology, Inc.); and Dylight488 conjugated goat anti-rabbit IgG secondary antibodies (Bioworld Technology, Inc.).

Techniques: Expressing, Western Blot, Staining, Control

Journal: Journal of Veterinary Science

Article Title: Epigallocatechin-3-gallate suppresses hemin-aggravated colon carcinogenesis through Nrf2-inhibited mitochondrial reactive oxygen species accumulation

doi: 10.4142/jvs.22097

Figure Lengend Snippet: WST, water-soluble tetrazolium salt; EGCG, epigallocatechin-3-gallate; Nrf2, nuclear factor erythroid-2-related factor 2. * p < 0.05.

Article Snippet: Nrf2 (NBP1-32822) and Keap1 (NBP2-03319) antibodies were purchased from Novus Biologicals (Centennial, CO, USA).

Techniques:

Journal: Food Science & Nutrition

Article Title: Trans‐Fatty Acids ( TFA ) Induced Vascular Injury Through the Regulation of the Sirt1‐Ppargc1a‐Nfe2l2 Signaling Pathway in Male Rats

doi: 10.1002/fsn3.70975

Figure Lengend Snippet: Effect of trans‐fatty acids on Sirt1/Ppargc1a/Nfe2l2 pathway of rat blood vessels. (A) Western blotting results of Silent information regulator 1 (Sirt1), Peroxisome proliferator‐activated receptor gamma coactivator 1‐alpha (Ppargc1a), and nuclear factor erythroid 2‐related factor 2 (Nfe2l2). (B−D) Semi‐quantitative analyses of protein expression. Results are shown as mean ± SD ( n = 6). * p < 0.05 by contrast with ND group; # p < 0.05 by contrast with LTD group. ND, normal diet; LTD, 1% trans‐fatty acids diet; HTD, 4% trans‐fatty acids diet.

Article Snippet: The following primary antibodies were employed in the study: anti‐rabbit Bcl2 (Signalway Antibody, China), anti‐rabbit Bax (Cell Signaling Technology, USA), anti‐rabbit Ppargc1a (Affinity, China), anti‐rabbit Casp3 (GeneTex, USA), anti‐rabbit Cleaved Casp3 (Arigo Biolaboratories, China), anti‐rabbit Nfe2l2 (Medical & Biological Laboratories, China), anti‐rabbit Sirt1 (OriGene, USA), and anti‐rabbit β‐actin (Abcam, China).

Techniques: Western Blot, Expressing

Journal: The Journal of biological chemistry

Article Title: Nuclear factor Nrf2 promotes glycosidase OGG1 expression by activating the AKT pathway to enhance leukemia cell resistance to cytarabine.

doi: 10.1016/j.jbc.2022.102798

Figure Lengend Snippet: Figure 1. High expression of Nrf2 associates with AML relapse. A, analysis of gene expression profile of Nrf2 in AML samples (n = 173) and matched normal samples (n = 70) according to GEPIA website. B, the expression level of Nrf2 mRNA in normal (n = 17), complete remission (n = 33), and relapsed AML patients (n = 46) was identified by RT–PCR. C, the expression of Nrf2 was detected by RT–PCR in the same AML patients before and after relapse (n = 8). D, Western blot was adopted for detecting the expression level of Nrf2 in normal (n = 9), complete remission (n = 9), and relapsed (n = 9) AML samples.

Article Snippet: The Nrf2 antibody was acquired from the abcam, and the OGG1 antibody was purchased from the Novus.

Techniques: Expressing, Gene Expression, Reverse Transcription Polymerase Chain Reaction, Western Blot

Journal: The Journal of biological chemistry

Article Title: Nuclear factor Nrf2 promotes glycosidase OGG1 expression by activating the AKT pathway to enhance leukemia cell resistance to cytarabine.

doi: 10.1016/j.jbc.2022.102798

Figure Lengend Snippet: Figure 2. High expression of OGG1 associates with high expression of Nrf2 in AML. A–D, RT–PCR analysis of the expressions of OGG1, XRCC1, AP site endonuclease 1 (APE1), and DNA polymerase β (POL-β) in Nrf2-low (n = 29) and Nrf2-high (n = 26) groups. E, the gene expression profiles of OGG1 in AML samples (n = 173) and matched normal samples (n = 70) were analyzed according to GEPIA website. F, RT–PCR analysis of OGG1 expression in normal (n = 28), complete remission (n = 29), and relapsed (n = 26) AML patients. G, the expression of OGG1 in the same AML patients before and after relapse was

Article Snippet: The Nrf2 antibody was acquired from the abcam, and the OGG1 antibody was purchased from the Novus.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Gene Expression

Journal: The Journal of biological chemistry

Article Title: Nuclear factor Nrf2 promotes glycosidase OGG1 expression by activating the AKT pathway to enhance leukemia cell resistance to cytarabine.

doi: 10.1016/j.jbc.2022.102798

Figure Lengend Snippet: Figure 3. High expression of Nrf2 in Ara-C-resistant AML cell lines. A, U937 and U937R cells were treated with different concentrations of Ara-C (4, 16, 64, 192, 386, 578, 768, and 1536 μM) for 24 h, and the cell viability was detected by CCK-8 colorimetry. B, THP-1 and THP-1R cells were treated with different concentrations of Ara-C (0.5, 4, 64, 192, 386, 578, 768, and 1536 μM) for 24 h, and the cell viability was detected by CCK-8 colorimetry. C, after U937, U937R, THP-1, and THP-1R were cocultured with Ara-C (2 μM) for 24 h, the apoptosis rate of AML cell lines was detected by FCM. D, analysis of apoptosis rate.

Article Snippet: The Nrf2 antibody was acquired from the abcam, and the OGG1 antibody was purchased from the Novus.

Techniques: Expressing, CCK-8 Assay, Colorimetric Assay

Journal: The Journal of biological chemistry

Article Title: Nuclear factor Nrf2 promotes glycosidase OGG1 expression by activating the AKT pathway to enhance leukemia cell resistance to cytarabine.

doi: 10.1016/j.jbc.2022.102798

Figure Lengend Snippet: Figure 4. High expression of Nrf2 decreases the sensitivity of leukemic cells to Ara-C. A, Western blot was used to verify the upregulation of Nrf2 in AML- sensitive celllines (U937andTHP-1). B, the relative grayvalues of Nrf2expression. C, Western blotwas appliedtodetect thedownregulation ofNrf2in AML-resistant cell lines (U937R andTHP-1R). D, the relativegray values of Nrf2 expression. E, RT–PCR verifiedthe expression ofmRNA after Nrf2 regulation.F, FCM was employed todetect the apoptosis rate of AML-sensitive cell lines cocultured with Ara-C (2 μM) for 24 h after Nrf2 overexpression. G, after downregulation of Nrf2, the apoptosis rate of AML drug–resistant cell lines cocultured with Ara-C (100 μM) for 24 h was detected by FCM. Each experiment was repeated three times independently and expressed as mean ± SD. *p < 0.05, **p < 0.01, and ***p < 0.001. AML, acute myeloid leukemia; Ara-C, cytarabine; FCM, flow cytometry; Nrf2, nuclear factor E2–related factor 2.

Article Snippet: The Nrf2 antibody was acquired from the abcam, and the OGG1 antibody was purchased from the Novus.

Techniques: Expressing, Western Blot, Reverse Transcription Polymerase Chain Reaction, Over Expression, Cytometry

Journal: The Journal of biological chemistry

Article Title: Nuclear factor Nrf2 promotes glycosidase OGG1 expression by activating the AKT pathway to enhance leukemia cell resistance to cytarabine.

doi: 10.1016/j.jbc.2022.102798

Figure Lengend Snippet: Figure 5. Overexpression of Nrf2 promotes the expression of OGG1 to mediate drug resistance in AML cells. A, detection of OGG1 expression in normal (bone marrow mononuclear cells from normal healthy donors) and AML-sensitive cell lines (U937 and THP-1) and drug-resistant cell lines (U937R and THP-1R) by IF staining. Scale bars represent 20 μm. B, quantification of OGG1 relative fluorescence intensity in normal and leukemia cells by ImageJ. C, Western blot was employed to detect OGG1 protein expression after upregulation of Nrf2. D, the relative gray values of Nrf2 and OGG1 expressions.

Article Snippet: The Nrf2 antibody was acquired from the abcam, and the OGG1 antibody was purchased from the Novus.

Techniques: Over Expression, Expressing, Staining, Western Blot

Journal: The Journal of biological chemistry

Article Title: Nuclear factor Nrf2 promotes glycosidase OGG1 expression by activating the AKT pathway to enhance leukemia cell resistance to cytarabine.

doi: 10.1016/j.jbc.2022.102798

Figure Lengend Snippet: Figure 7. Nrf2 overexpression promotes OGG1 expression by activating AKT signaling pathway. A, GeneMania protein–protein interaction network of OGG1, Nrf2, and AKT. B and C, Western blot was applied to identify the expressions of Nrf2, OGG1, AKT, p-AKT, Cleaved-caspase 9, and Total-caspase 9 proteins in AML-sensitive cell lines after overexpression of Nrf2. D and E, Western blot was adopted for detecting the expression of Nrf2, OGG1, AKT, p-AKT, Cleaved-caspase 9, and Total-caspase 9 proteins in AML drug–resistant cell lines after downregulation of Nrf2. Each experiment was repeated three times independently and denoted as mean ± SD. *p < 0.05, **p < 0.01, and ***p < 0.001. Nrf2, nuclear factor E2–related factor 2; OGG1, 8-hydroxyguanine DNA glycosidase.

Article Snippet: The Nrf2 antibody was acquired from the abcam, and the OGG1 antibody was purchased from the Novus.

Techniques: Over Expression, Expressing, Western Blot

Journal: The Journal of biological chemistry

Article Title: Nuclear factor Nrf2 promotes glycosidase OGG1 expression by activating the AKT pathway to enhance leukemia cell resistance to cytarabine.

doi: 10.1016/j.jbc.2022.102798

Figure Lengend Snippet: Figure 8. Inhibition of AKT signaling pathway to reduce OGG1 expression. A, after U937 cells were treated with 2 μM MK-2206 for 24 h, the protein expression levels of Nrf2, OGG1, AKT, p-AKT, Cleaved-caspase 9, and Total-caspase 9 in Nrf2 overexpression and EV1 groups were detected by Western blot. B, the relative gray value of related proteins. C, after THP-1 cells were treated with 2 μM MK-2206 for 24 h, the protein expression levels of Nrf2, OGG1, AKT, p-AKT, Cleaved-caspase 9, and Total-caspase 9 in Nrf2 overexpression and EV1 groups were detected by Western blot. D, the relative gray values of related

Article Snippet: The Nrf2 antibody was acquired from the abcam, and the OGG1 antibody was purchased from the Novus.

Techniques: Inhibition, Expressing, Over Expression, Western Blot

Journal: The Journal of biological chemistry

Article Title: Nuclear factor Nrf2 promotes glycosidase OGG1 expression by activating the AKT pathway to enhance leukemia cell resistance to cytarabine.

doi: 10.1016/j.jbc.2022.102798

Figure Lengend Snippet: Figure 9. Downregulation of Nrf2 reduces the risk of drug resistance in vivo. A and B, tumor volume of mice after subcutaneous injection of U937R (n = 4), U937R-EV (n = 4), U937R-si-Nrf2 (n = 3), and U937R-EV + Ara-C (n = 4) and U937R-si-Nrf2 + Ara-C (n = 3) cells after 35 days growth. C, changes of tumor weight after subcutaneousinjectionofU937R(n=4),U937R-EV(n=4),U937R-si-Nrf2(n=3),andU937R-EV+Ara-C(n=4)andU937R-si-Nrf2+Ara-C(n=3)cells.DandE,volumegrowth curveofxenografttumor.F,analysisanddrawingofsurvivalcurveofxenograftedtumormicebyKaplan–Meiermethod.G–J,IHCwasappliedtoidentifytheexpressionsof Nrf2andOGG1insubcutaneoustransplantedtumortissuesections.Thescalebarsfromlefttorightrepresent100and50μm,respectively.*p< 0.05,**p< 0.01,and***p< 0.001. Ara-C, cytarabine; EV, empty vector; IHC, immunohistochemistry; Nrf2, nuclear factor E2–related factor 2; OGG1, 8-hydroxyguanine DNA glycosidase.

Article Snippet: The Nrf2 antibody was acquired from the abcam, and the OGG1 antibody was purchased from the Novus.

Techniques: In Vivo, Injection, Plasmid Preparation, Immunohistochemistry

Journal: Scientific Reports

Article Title: RNA sequencing of corneas from two keratoconus patient groups identifies potential biomarkers and decreased NRF2-antioxidant responses

doi: 10.1038/s41598-020-66735-x

Figure Lengend Snippet: Canonical pathways significantly altered in KC and KJ. Grey bars show total number of genes in the pathway (scale on top X axis), with blue and red bars showing the numbers of down-regulated and up-regulated genes (scale on bottom X axis). The p value next to each bar was calculated for that pathway by the IPA Core Analysis. “Acute Phase” is the Acute Phase Response Signaling pathway; “NRF2-mediated” is NRF2-mediated Oxidative Stress Response pathway; “ECM related” is the Hepatic Fibrosis / Hepatic Stellate Cell Activation pathway; “Rheumatoid Arthritis” is the Role of Macrophages, Fibroblasts and Endothelial Cells in Rheumatoid Arthritis pathway; and, “Osteoarthritis” includes genes associated with osteoarthritis.

Article Snippet: The following primary antibodies were used from Biorbyt Research Products: NRF2 (orb128433, 1:50), KEAP1 (orb48426, 1:100), Hsp40 (orb520080, 1:100), and GAS1 (orb414757, 1:100) and RXRA antibody were used from Novus Biologicals (NBP2–75653, 1:50).

Techniques: Activation Assay

Journal: Scientific Reports

Article Title: RNA sequencing of corneas from two keratoconus patient groups identifies potential biomarkers and decreased NRF2-antioxidant responses

doi: 10.1038/s41598-020-66735-x

Figure Lengend Snippet: NRF2 target genes significantly altered in KCN corneas. ( A ) Decreased NRF2 target gene expression in KC and KJ RNA seq. ( B ) Mechanism of NRF2 regulation by KEAP1; the latter binds to NRF2 and CUL3 for NRF2-ubiquitination and degradation to maintain low levels of NRF2 under homeostatic conditions. Under oxidative stress, KEAP1 dissociates from NRF2 allowing its increase and upregulation of target genes.

Article Snippet: The following primary antibodies were used from Biorbyt Research Products: NRF2 (orb128433, 1:50), KEAP1 (orb48426, 1:100), Hsp40 (orb520080, 1:100), and GAS1 (orb414757, 1:100) and RXRA antibody were used from Novus Biologicals (NBP2–75653, 1:50).

Techniques: Targeted Gene Expression, RNA Sequencing, Ubiquitin Proteomics

Journal: Scientific Reports

Article Title: RNA sequencing of corneas from two keratoconus patient groups identifies potential biomarkers and decreased NRF2-antioxidant responses

doi: 10.1038/s41598-020-66735-x

Figure Lengend Snippet: KEAP1 and NRF2 immunostaining in DN and KCN corneas. ( A ) KEAP1 staining is decreased in KCN corneas, with focally increased staining in some basal epithelial cells (inset), whereas in DN corneas KEAP1 shows staining of all epithelial layers (inset). ( B ) NRF2 shows very little to no staining of KCN corneas and these were all cytoplasmic (inset), while DN sections show stronger staining of epithelial cells and some nuclear staining (inset) DAPI nuclear staining shown in blue. IF staining of additional KCN and DN cornea sections are shown in Supplemental Fig. . Scale bar: 50 µm.

Article Snippet: The following primary antibodies were used from Biorbyt Research Products: NRF2 (orb128433, 1:50), KEAP1 (orb48426, 1:100), Hsp40 (orb520080, 1:100), and GAS1 (orb414757, 1:100) and RXRA antibody were used from Novus Biologicals (NBP2–75653, 1:50).

Techniques: Immunostaining, Staining

Journal: Antioxidants

Article Title: Ellagic Acid Alleviates Oxidative Stress by Mediating Nrf2 Signaling Pathways and Protects against Paraquat-Induced Intestinal Injury in Piglets

doi: 10.3390/antiox11020252

Figure Lengend Snippet: Primer sequences 1 .

Article Snippet: Membranes were sealed for 10 min with a rapid blocking solution (Beyotime Institute of Biotechnology, Shanghai, China) at room temperature and incubated for 2 h with the primary antibodies including NQO1 (1:1000; Proteintech; Chicago, IL, USA), Nrf2 (1:600; Abcam; Cambridge, UK), HO-1(1:800; Abcam; Cambridge, UK), Recombinant Kelch Like ECH Associated Protein 1 (Keap1) (1:800; Proteintech; Chicago, IL, USA), and β-actin (1:1000; Cell Signaling Technology; Danvers, MA, USA), along with the secondary antibody horseradish peroxidase-conjugated goat anti-rabbit immunoglobulin G (1:5000; ZSGB, Biological Technology, Beijing, China).

Techniques:

Journal: Antioxidants

Article Title: Ellagic Acid Alleviates Oxidative Stress by Mediating Nrf2 Signaling Pathways and Protects against Paraquat-Induced Intestinal Injury in Piglets

doi: 10.3390/antiox11020252

Figure Lengend Snippet: Protein abundances of Nrf2 signaling pathway of cytosol and nucleus of jejunal and ileal mucosa in piglets. ( A ) Nuclear factor erythroid 2-related factor 2 (Nrf2), Recombinant Kelch Like ECH Associated Protein 1 (Keap1), heme oxygenase-1 (HO-1), and quinone oxidoreductase 1 (NQO1) protein abundances in cytosol of jejunal and ileal mucosa. ( B ) Nrf2, HO-1, and NQO1 protein abundances in nucleus of jejunal and ileal mucosa. PQ = 4 mg/kg paraquet; EL = 0.005% ellagic acid + 4 mg/kg paraquet; EM = 0.01% ellagic acid + 4 mg/kg paraquet; EH = 0.02% ellagic acid + 4 mg/kg paraquet. n = 8. Data are shown as mean ± SEM. * means the difference was significant when compared to the control group. ns means the difference was not significant when compared to the control group. a–c Values with different lowercase letters are significantly different among PQ, EL, EM, and EH groups ( p < 0.05).

Article Snippet: Membranes were sealed for 10 min with a rapid blocking solution (Beyotime Institute of Biotechnology, Shanghai, China) at room temperature and incubated for 2 h with the primary antibodies including NQO1 (1:1000; Proteintech; Chicago, IL, USA), Nrf2 (1:600; Abcam; Cambridge, UK), HO-1(1:800; Abcam; Cambridge, UK), Recombinant Kelch Like ECH Associated Protein 1 (Keap1) (1:800; Proteintech; Chicago, IL, USA), and β-actin (1:1000; Cell Signaling Technology; Danvers, MA, USA), along with the secondary antibody horseradish peroxidase-conjugated goat anti-rabbit immunoglobulin G (1:5000; ZSGB, Biological Technology, Beijing, China).

Techniques: Recombinant, Control

Journal: Frontiers in Nutrition

Article Title: Cannabis oil modulates liver alterations and endocannabinoid system changes in a female rat model of diet-induced MASLD

doi: 10.3389/fnut.2026.1770150

Figure Lengend Snippet: Liver NrF2 and NFκB p65 expression in female rats fed a reference diet (RD), sucrose-rich diet (SRD) or SRD with cannabis oil (SRD + Ca). (A) Representative photomicrographs of immunocytochemical staining NrF2 in the liver sections of rats. Decreased levels of nuclear and cytoplasmic positive markers are observed in the SRD group. Scale bar 50 μm. (B) Quantitative immunohistochemical analysis of liver NrF2 expression expressed as integrated optical density (IOD). (C) Representative photomicrographs of immunocytochemical staining NFκB p65 in the liver sections of rats. Increased levels of nuclear and cytoplasmic positive markers are observed in the SRD group. Scale bar 50 μm. (D) Quantitative immunohistochemical analysis of NFκB p65 expression in the liver. Data are expressed as mean ± SEM ( n = 6). Statistical differences were evaluated by one-way ANOVA followed by the Newman–Keuls post hoc test (* P < 0.05).

Article Snippet: The samples were incubated in a humid chamber first with a specific primary antibody for 4-HNE (mouse monoclonal antibody; Catalog # MAB3249; R&D Systems), NrF2 (mouse monoclonal antibody; sc-365949; Santa Cruz Biotechnology), NF-κB p65 (mouse monoclonal antibody; sc-8008; Santa Cruz Biotechnology) and TGF-β1 (mouse monoclonal antibody; sc-52893; Santa Cruz Biotechnology) (for 14–16 h at 4°C) and then with biotin-conjugated secondary antibody (anti-mouse, 1:100 dilution, Sigma) for 30 min at room temperature.

Techniques: Expressing, Cannabis, Staining, Immunohistochemical staining