Antiarthritic Effects of a Root Extract from Harpagophytum procumbens DC: Novel Insights into the Molecular Mechanisms and Possible Bioactive Phytochemicals
Abstract
:1. Introduction
2. Materials and Methods
2.1. Harpagophytum Procumbens Extract
2.2. Phytochemical Analysis
2.2.1. Determination of Total Polyphenols, Tannins, and Flavonoids
2.2.2. Solid-Phase Microextraction (SPME)
2.2.3. Gas Chromatography–Mass Spectrometry (GC–MS)
2.3. Human Tissue
2.4. Immunohistochemistry
2.5. Human Primary Cell Isolation
2.6. Immunofluorescence
2.7. Densitometric Analysis
2.8. Cell Treatment
2.9. Cell Viability
2.10. RNA Extraction and Reverse Transcription
2.11. Quantitative Real-Time PCR
2.12. FAAH Inhibition
2.13. Statistical Analysis
3. Results
3.1. Phytochemical Characterization
3.2. Expression of Cannabinoid Receptors in OA Synovial Membrane
3.3. Effects of Harpagophytum Extracts on Synoviocyte Cell Viability
3.4. Mechanism of Action of Harpagophytum Extracts on FLSs
3.5. Inhibition of Fatty Acid Anandamide Hydrolase
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Gene | Primer Sequences |
---|---|
CB1 NM_016083 | Forward: 5′-TTCCTTCTTGTGAAGGCACTG-3′ Reverse: 5′-TCTTGACCGTGCTCTTGATGC-3′ |
CB2 NM_001841 | Forward: 5′-ATGCTGTGCCTCATCAACTC-3′ Reverse: 5′-CTCACACACTTCTTCCAGTG-3′ |
FAAH NM_001441 | Forward: 5′-CAGCTTTCCTCAGCAACATG-3′ Reverse: 5′-CAATCACGGTTTTGCGGTAC-3′ |
18S NM_003286 | Forward: 5′-CGCCGCTAGAGGTGAAATTC-3′ Reverse: 5′-CATTCTTGGCAAATGCTTTCG-3′ |
Compounds | HPEDMSO | HPEEtOH100 | HPEEtOH50 | HPEH2O |
---|---|---|---|---|
µg/mg of Dry Extract (Mean ± SE) | ||||
Polyphenols (TAE) | 99.6 ± 0.05 ***§ | 69.6 ± 0.05 | 97.9 ± 0.01 ***§ | 84.2 ± 0.04 ** |
Tannins (TAE) | 14.2 ± 0.02 ** | 9.2 ± 0.04 | 9.2 ± 0.02 | 17.9 ± 0.03 *** |
Flavonoids (QE) | 113.0± 5.1 ***§§ | 71.3 ± 6.6 §§ | 75.1 ± 6.1 §§ | 37.6 ± 4.3 |
Flavonols (QE) | 43.8 ± 2.4 ***§§ | 27.6 ± 3.2 | 30.1 ± 2.0 § | 25.1 ± 2.0 |
No. 1 | Compound 2 | LRI 3 | LRI 4 | MS 5 | HPEDMSO (%) | HPEEtOH100 (%) | HPEEtOH50 (%) | HPEH2O (%) |
---|---|---|---|---|---|---|---|---|
1 | β-pinene | 978 | 974 | + | - | - | 5.1 | - |
2 | thymol | 1308 | 1310 | + | 0.8 | - | - | - |
3 | eugenol | 1339 | 1344 | + | 6.0 | 14.6 | 51.6 | - |
4 | α-copaene | 1381 | 1387 | + | - | 6.3 | 3.7 | - |
5 | β-caryophyllene | 1412 | 1416 | + | 77.4 | 77.1 | 38.6 | - |
6 | isoeugenol | 1432 | 1439 | + | - | - | 1.0 | - |
7 | α-humulene | 1450 | 1454 | + | 10.0 | 0.8 | - | - |
8 | δ-cadinene | 1528 | 1530+ | + | 5.8 | 1.3 | - | - |
Total (%) | 100.0 | 100.0 | 100.0 |
Harpagophytum procumbens Root Extract | IC50 (CL) μg/mL |
---|---|
HPEDMSO | 94.7 (23.8–97.5) |
HPEEtOH100 | 65.5 (19.2–87.9) * |
HPEEtOH50 | 73.8 (24.5–94.3) |
HPEH2O | - |
JZL 195 | 0.03 (0.01–0.06) * |
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Mariano, A.; Di Sotto, A.; Leopizzi, M.; Garzoli, S.; Di Maio, V.; Gullì, M.; Dalla Vedova, P.; Ammendola, S.; Scotto d’Abusco, A. Antiarthritic Effects of a Root Extract from Harpagophytum procumbens DC: Novel Insights into the Molecular Mechanisms and Possible Bioactive Phytochemicals. Nutrients 2020, 12, 2545. https://doi.org/10.3390/nu12092545
Mariano A, Di Sotto A, Leopizzi M, Garzoli S, Di Maio V, Gullì M, Dalla Vedova P, Ammendola S, Scotto d’Abusco A. Antiarthritic Effects of a Root Extract from Harpagophytum procumbens DC: Novel Insights into the Molecular Mechanisms and Possible Bioactive Phytochemicals. Nutrients. 2020; 12(9):2545. https://doi.org/10.3390/nu12092545
Chicago/Turabian StyleMariano, Alessia, Antonella Di Sotto, Martina Leopizzi, Stefania Garzoli, Valeria Di Maio, Marco Gullì, Pietro Dalla Vedova, Sergio Ammendola, and Anna Scotto d’Abusco. 2020. "Antiarthritic Effects of a Root Extract from Harpagophytum procumbens DC: Novel Insights into the Molecular Mechanisms and Possible Bioactive Phytochemicals" Nutrients 12, no. 9: 2545. https://doi.org/10.3390/nu12092545
APA StyleMariano, A., Di Sotto, A., Leopizzi, M., Garzoli, S., Di Maio, V., Gullì, M., Dalla Vedova, P., Ammendola, S., & Scotto d’Abusco, A. (2020). Antiarthritic Effects of a Root Extract from Harpagophytum procumbens DC: Novel Insights into the Molecular Mechanisms and Possible Bioactive Phytochemicals. Nutrients, 12(9), 2545. https://doi.org/10.3390/nu12092545