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Ethyl acetate fraction from Nymphaea hybrida Peck modulates inflammatory responses in LPS-stimulated RAW 264.7 cells and acute inflammation murine models.
J Ethnopharmacol. 2021 Apr 06; 269:113698.JE

Abstract

ETHNOPHARMACOLOGICAL RELEVANCE

Nymphaea hybrida Peck is used as a traditional medicinal herb for treating pain and inflammatory diseases, and known for its ornamental value and as a hot drink. However, the effects of N. hybrida polar fractions on lipopolysaccharide (LPS)-induced in vitro inflammation model and acute inflammation murine models have yet to be evaluated.

AIM OF THE STUDY

The aim of this study was to elucidate the anti-inflammatory effects of N. hybrida ethanol extract (NHE) and its polar fractions: petroleum ether (PE), methylene chloride (MC), ethyl acetate (EA), methanol (ME), and water (WA). The underlying molecular mechanisms of active fraction in LPS-stimulated RAW 264.7 murine macrophages were further investigated.

MATERIAL AND METHODS

Fractions with potential anti-inflammatory effects were screened using direct nitric oxide (NO) radical scavenging and cyclooxygenase (COX)-2 inhibition assays in vitro. The anti-inflammatory properties of potential fraction were evaluated in LPS-stimulated RAW264.7 cells, xylene-induced ear edema, carrageenan-induced paw edema and xylene-induced Evans blue exudation of acute inflammation murine models. The regulation of nuclear factor-κB (NF-κB) and mitogen-activated protein kinase (MAPK) signaling pathways were investigated using western blotting and immunofluorescence.

RESULTS

Compared to other polar fractions, NHE-EA displayed higher phenol and flavonoid content, and exerted greater activity in direct NO radical scavenging and COX-2 inhibition assay in vitro. NHE-EA markedly decreased the levels of inflammatory mediators, NO and prostaglandin E2 (PGE2), by suppressing the over-expression of inducible nitric oxide synthase (iNOS) and COX-2 in LPS-stimulated RAW264.7 cells. The NHE-EA fraction dose-dependently alleviated over-elevation of LPS-associated intracellular calcium and decreased the abnormal secretion of pro-inflammatory cytokines, tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), IL-6, and interferon-γ (IFN-γ). The combination with NHE-EA effectively attenuated the activation and nuclear translocation of NF-κB p65, and the phosphorylation of extracellular signal-regulated kinases (ERK), c-Jun N-terminal kinases (JNK), and p38 kinases of MAPK pathways. NHE-EA could significantly ameliorate the degree of swelling of the mice ear and paw, the skin exudation of Evans blue and the excessive secretion of inflammatory cytokines.

CONCLUSION

Our results demonstrated that NHE-EA was the most active polar fraction of N. hybrida extracts. It inhibited the LPS-associated inflammatory response by blocking the activation of NF-κB and MAPKs pathways in RAW264.7 cells. It also effectively alleviated the inflammatory response of acute inflammation. These results indicated the role of NHE-EA as adjuvants and their potential role in alternative strategy for the treatment of inflammatory diseases.

Authors+Show Affiliations

Zhejiang Provincial Engineering Technology Research Center of Marine Biomedical Products, School of Food and Pharmacy, Zhejiang Ocean University, 1 South Haida Road, Zhoushan, 316000, People's Republic of China. Electronic address: zw.zhang1997@zjou.edu.cn.Zhejiang Provincial Engineering Technology Research Center of Marine Biomedical Products, School of Food and Pharmacy, Zhejiang Ocean University, 1 South Haida Road, Zhoushan, 316000, People's Republic of China. Electronic address: S18070700050@zjou.edu.cn.Zhejiang Provincial Engineering Technology Research Center of Marine Biomedical Products, School of Food and Pharmacy, Zhejiang Ocean University, 1 South Haida Road, Zhoushan, 316000, People's Republic of China. Electronic address: thq_22326@163.com.Zhejiang Provincial Engineering Technology Research Center of Marine Biomedical Products, School of Food and Pharmacy, Zhejiang Ocean University, 1 South Haida Road, Zhoushan, 316000, People's Republic of China. Electronic address: zyddy1028@163.com.Zhejiang Provincial Engineering Technology Research Center of Marine Biomedical Products, School of Food and Pharmacy, Zhejiang Ocean University, 1 South Haida Road, Zhoushan, 316000, People's Republic of China. Electronic address: hekang7072@163.com.ZhouShan Academy of Agriculture Sciences, Zhoushan, 316022, China.Zhejiang Provincial Engineering Technology Research Center of Marine Biomedical Products, School of Food and Pharmacy, Zhejiang Ocean University, 1 South Haida Road, Zhoushan, 316000, People's Republic of China. Electronic address: fmyu@zjou.edu.cn.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

33338590

Citation

Zhang, Zhuangwei, et al. "Ethyl Acetate Fraction From Nymphaea Hybrida Peck Modulates Inflammatory Responses in LPS-stimulated RAW 264.7 Cells and Acute Inflammation Murine Models." Journal of Ethnopharmacology, vol. 269, 2021, p. 113698.
Zhang Z, Jiang S, Tian H, et al. Ethyl acetate fraction from Nymphaea hybrida Peck modulates inflammatory responses in LPS-stimulated RAW 264.7 cells and acute inflammation murine models. J Ethnopharmacol. 2021;269:113698.
Zhang, Z., Jiang, S., Tian, H., Zeng, Y., He, K., Lin, L., & Yu, F. (2021). Ethyl acetate fraction from Nymphaea hybrida Peck modulates inflammatory responses in LPS-stimulated RAW 264.7 cells and acute inflammation murine models. Journal of Ethnopharmacology, 269, 113698. https://doi.org/10.1016/j.jep.2020.113698
Zhang Z, et al. Ethyl Acetate Fraction From Nymphaea Hybrida Peck Modulates Inflammatory Responses in LPS-stimulated RAW 264.7 Cells and Acute Inflammation Murine Models. J Ethnopharmacol. 2021 Apr 6;269:113698. PubMed PMID: 33338590.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Ethyl acetate fraction from Nymphaea hybrida Peck modulates inflammatory responses in LPS-stimulated RAW 264.7 cells and acute inflammation murine models. AU - Zhang,Zhuangwei, AU - Jiang,Shuoqi, AU - Tian,Hengqun, AU - Zeng,Yu, AU - He,Kang, AU - Lin,Lin, AU - Yu,Fangmiao, Y1 - 2020/12/15/ PY - 2020/09/07/received PY - 2020/12/11/revised PY - 2020/12/11/accepted PY - 2020/12/19/pubmed PY - 2021/7/21/medline PY - 2020/12/18/entrez KW - Anti-inflammation KW - Extracts KW - MAPKs KW - NF-κB KW - RAW264.7 cells SP - 113698 EP - 113698 JF - Journal of ethnopharmacology JO - J Ethnopharmacol VL - 269 N2 - ETHNOPHARMACOLOGICAL RELEVANCE: Nymphaea hybrida Peck is used as a traditional medicinal herb for treating pain and inflammatory diseases, and known for its ornamental value and as a hot drink. However, the effects of N. hybrida polar fractions on lipopolysaccharide (LPS)-induced in vitro inflammation model and acute inflammation murine models have yet to be evaluated. AIM OF THE STUDY: The aim of this study was to elucidate the anti-inflammatory effects of N. hybrida ethanol extract (NHE) and its polar fractions: petroleum ether (PE), methylene chloride (MC), ethyl acetate (EA), methanol (ME), and water (WA). The underlying molecular mechanisms of active fraction in LPS-stimulated RAW 264.7 murine macrophages were further investigated. MATERIAL AND METHODS: Fractions with potential anti-inflammatory effects were screened using direct nitric oxide (NO) radical scavenging and cyclooxygenase (COX)-2 inhibition assays in vitro. The anti-inflammatory properties of potential fraction were evaluated in LPS-stimulated RAW264.7 cells, xylene-induced ear edema, carrageenan-induced paw edema and xylene-induced Evans blue exudation of acute inflammation murine models. The regulation of nuclear factor-κB (NF-κB) and mitogen-activated protein kinase (MAPK) signaling pathways were investigated using western blotting and immunofluorescence. RESULTS: Compared to other polar fractions, NHE-EA displayed higher phenol and flavonoid content, and exerted greater activity in direct NO radical scavenging and COX-2 inhibition assay in vitro. NHE-EA markedly decreased the levels of inflammatory mediators, NO and prostaglandin E2 (PGE2), by suppressing the over-expression of inducible nitric oxide synthase (iNOS) and COX-2 in LPS-stimulated RAW264.7 cells. The NHE-EA fraction dose-dependently alleviated over-elevation of LPS-associated intracellular calcium and decreased the abnormal secretion of pro-inflammatory cytokines, tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), IL-6, and interferon-γ (IFN-γ). The combination with NHE-EA effectively attenuated the activation and nuclear translocation of NF-κB p65, and the phosphorylation of extracellular signal-regulated kinases (ERK), c-Jun N-terminal kinases (JNK), and p38 kinases of MAPK pathways. NHE-EA could significantly ameliorate the degree of swelling of the mice ear and paw, the skin exudation of Evans blue and the excessive secretion of inflammatory cytokines. CONCLUSION: Our results demonstrated that NHE-EA was the most active polar fraction of N. hybrida extracts. It inhibited the LPS-associated inflammatory response by blocking the activation of NF-κB and MAPKs pathways in RAW264.7 cells. It also effectively alleviated the inflammatory response of acute inflammation. These results indicated the role of NHE-EA as adjuvants and their potential role in alternative strategy for the treatment of inflammatory diseases. SN - 1872-7573 UR - https://www.unboundmedicine.com/medline/citation/33338590/Ethyl_acetate_fraction_from_Nymphaea_hybrida_Peck_modulates_inflammatory_responses_in_LPS_stimulated_RAW_264_7_cells_and_acute_inflammation_murine_models_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0378-8741(20)33586-8 DB - PRIME DP - Unbound Medicine ER -