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Hypericum erectum alcoholic extract inhibits Toxoplasma growth and Entamoeba encystation: an exploratory study on the anti-protozoan potential

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Abstract

Hypericum erectum is an important ethnobotanical medicine in East Asian tradition. To explore the anti-parasitic potential of H. erectum, inhibitory effects on the growth of intracellular parasite Toxoplasma and on the encystation of intestinal parasite Entamoeba were examined. The constituents in H. erectum alcoholic extracts and fractions separated by solvent-partitioning were analysed by high resolution LC–MS. Toxoplasma gondii growth inhibition assay was performed using GFP-labelled T. gondii strain PTG-GFP by measuring the fluorescence intensity. Anti-Toxoplasma drug pyrimethamine was used as a positive control. T. gondii-induced immune reaction was assessed by quantitative PCR and fluorescence microscopy, using co-culture of PTG-GFP and monocyte-macrophage cell line Raw264. The inhibitory effect on the encystation of Entamoeba invadens was measured by flow-cytometry, where paromomycin was used as a positive control. H. erectum methanol (MeOH) extract (50 µg/mL) and ethyl acetate (EtOAc) fraction (50 µg/mL) inhibited the growth of T. gondii, while 50%MeOH extract and hydrophilic fractions were ineffective. Co-culture with T. gondii reduced the viability of macrophages, however macrophages were protected in the presence of H. erectum MeOH extract or EtOAc fraction (above 10 µg/mL). The MeOH extract and EtOAc fraction also effectively suppressed the encystation of E. invadens at 1 mg/mL. Hypericine, a major constituent in MeOH extract and EtOAc fraction, inhibited T. gondii growth and E. invadens encystation. Our results demonstrated that H. erectum effectively inhibited Toxoplasma growth and Entamoeba encystation. These activities are partly mediated by hypericin. In addition, it was suggested the extract and fraction may protect innate immune cells from Toxoplasma-induced damages, thereby enhancing parasite clearance. Further investigation is warranted to address the in vivo effectiveness of H. erectum as an anti-protozoal medicine.

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Acknowledgements

We thank Dr. Yasunobu Miyake (Saga University) and Dr. Akihiro Sekine (Chiba University) who provided their expertise that assisted this study, and all our colleagues who created the supportive work environment. This work was funded by Grants-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology of Japan 19K07520 and 17KT0124 to K.N., 17K15676 to K.H., and 19K07839 to N.S., AMED-J-PRIDE (JP18fm0208025) to F.M. and H.Y. This work was also supported by the Naito Foundation to F.M. The flow cytometric analysis was performed using a MACSQuant Analyzer at the Analytical Research Center for Experimental Sciences, Saga University.

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Author notes

  1. Noriko Shinjyo

    Present address: Department of Infection and Host Defense, Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba, 260-8670, Japan

Authors and Affiliations

  1. Division of Molecular and Cellular Immunoscience, Department of Biomolecular Sciences, Faculty of Medicine, Saga University, 5-1-1 Nabeshima, Saga, 849-8501, Japan

    Noriko Shinjyo, Fumika Mi-ichi & Hiroki Yoshida

  2. Saga Prefectural Institute of Public Health and Pharmaceutical Research, 1-20 Hacchounawate, Saga, 849-0925, Japan

    Hideyuki Nakayama

  3. Department of Applied Biological Sciences, Faculty of Agriculture, Saga University, 1 Honjo, Saga, 840-8502, Japan

    Kanji Ishimaru

  4. Department of Infection and Host Defense, Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba, 260-8670, Japan

    Kenji Hikosaka & Kazumi Norose

  5. School of Tropical Medicine and Global Health, Nagasaki University, 1-12-4 Sakamoto, Nagasaki, 852-8523, Japan

    Noriko Shinjyo

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Shinjyo, N., Nakayama, H., Ishimaru, K. et al. Hypericum erectum alcoholic extract inhibits Toxoplasma growth and Entamoeba encystation: an exploratory study on the anti-protozoan potential. J Nat Med 74, 294–305 (2020). https://doi.org/10.1007/s11418-019-01369-6

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