The Modulation of Nrf-2/HO-1 Signaling Axis by Carthamus tinctorius L. Alleviates Vascular Inflammation in Human Umbilical Vein Endothelial Cells
Abstract
:1. Introduction
2. Materials and Methods
2.1. Preparation of ECT
2.2. Chemicals and Reagents for HPLC Analysis
2.3. HPLC Analysis of Six Marker Analytes in Flowers of C. tinctorius
2.4. Cell Cultures
2.5. Enzyme-Linked Immunsorbent Assay
2.6. Western Blot Analysis
2.7. Total mRNA Preparation and Reverse Transcription-Polymerase Chain Reaction (RT-PCR)
2.8. HL-60 Monocyte and HUVEC Adhesion Analysis
2.9. Intracellular ROS Production Assay
2.10. NF-κB Activation Assay
2.11. Immunofluorescence Microscopy
2.12. Statistical Analysis
3. Results
3.1. Quantification of Six Marker Compounds in ECT
3.2. Effect of ECT on TNF-α-Induced CAMs Expression in HUVEC
3.3. Effect of ECT on Leukocyte Adhesion to Vascular Endothelial Cells
3.4. Effect of ECT on TNF-α-Induced ROS/NF-κB Pathway
3.5. Involvement of HO-1/Nrf2 Pathway in ECT Protective Effect against Vascular Inflammation
4. Discussion
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Gene | Primer Nucleotide Sequence |
---|---|
ICAM-1 | Forward: 5′-CTCACCCGTGTACTGGACTC-3′ Reverse: 5′-CGCCGG AAAGCTGTAGATGG-3′ |
VCAM-1 | Forward: 5′-ATGCCTGGG AAGATGGTCGTGA-3′ Reverse: 5′-TGGAGCTGGTAGACCCTCGCTG-3′ |
E-selectin | Forward: 5′-ATCATCCTGCAACTTCACC-3′ Reverse: 5′-ACACCTCACCAAACCCTTC-3′ |
MCP-1 | Forward: 5’-CAGCCAGATGCAATCAATGC-3’ Reverse: 5’-GTGGTCCATGGAATCCTGAA-3’ |
β-actin | Forward: 5′-AGGGAGGCGT TCACCTCAGG-3′ Reverse 5′-AACTCCATCACCAGGCG TGGG-3′ |
Compound | Linear Range (μg/mL) | Regression Equation (y = ax + b) a | r2 | LOD (μg/mL) b | LOQ (μg/mL) c |
---|---|---|---|---|---|
Quercetin 3-rutinoside | 0.31–20.00 | y = 28,539.15x + 2768.62 | 1.0000 | 0.06 | 0.18 |
Quercetin 3-glycoside | 0.31–20.00 | y = 28,930.08x + 1776.94 | 1.0000 | 0.04 | 0.12 |
Kaempferol 3-rutinoside | 0.78–50.00 | y = 21,078.39x + 3828.68 | 1.0000 | 0.14 | 0.43 |
Hydroxysafflor yellow A | 1.56–100.00 | y = 33,781.29x + 3563.25 | 0.9999 | 0.19 | 0.57 |
Kaempferol 3-glycoside | 0.31–20.00 | y = 38,389.02x + 1978.30 | 1.0000 | 0.01 | 0.02 |
Bidenoside C | 0.31–20.00 | y = 18,983.02x + 1748.57 | 1.0000 | 0.01 | 0.04 |
Compound | Leaves | ||
---|---|---|---|
Mean (mg/g) | SD (×10−2) | RSD (%) | |
Quercetin 3-rutinoside | 0.02 | 0.04 | 2.60 |
Quercetin 3-glycoside | 0.05 | 0.06 | 1.25 |
Kaempferol 3-rutinoside | 0.10 | 0.19 | 1.81 |
Hydroxysafflor yellow A | 1.59 | 0.12 | 0.07 |
Kaempferol 3-glycoside | 0.02 | 0.06 | 2.44 |
Bidenoside C | 0.03 | 0.06 | 2.33 |
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Lee, Y.J.; Lee, Y.P.; Seo, C.S.; Choi, E.S.; Han, B.H.; Yoon, J.J.; Jang, S.H.; Jeong, C.G.; Mun, Y.J.; Kang, D.G.; et al. The Modulation of Nrf-2/HO-1 Signaling Axis by Carthamus tinctorius L. Alleviates Vascular Inflammation in Human Umbilical Vein Endothelial Cells. Plants 2021, 10, 2795. https://doi.org/10.3390/plants10122795
Lee YJ, Lee YP, Seo CS, Choi ES, Han BH, Yoon JJ, Jang SH, Jeong CG, Mun YJ, Kang DG, et al. The Modulation of Nrf-2/HO-1 Signaling Axis by Carthamus tinctorius L. Alleviates Vascular Inflammation in Human Umbilical Vein Endothelial Cells. Plants. 2021; 10(12):2795. https://doi.org/10.3390/plants10122795
Chicago/Turabian StyleLee, Yun Jung, Yong Pyo Lee, Chang Seob Seo, Eun Sik Choi, Byung Hyuk Han, Jung Joo Yoon, Se Hoon Jang, Chae Ghang Jeong, Yeun Ja Mun, Dae Gill Kang, and et al. 2021. "The Modulation of Nrf-2/HO-1 Signaling Axis by Carthamus tinctorius L. Alleviates Vascular Inflammation in Human Umbilical Vein Endothelial Cells" Plants 10, no. 12: 2795. https://doi.org/10.3390/plants10122795