Dataset Information

Curcumin effect on Acanthamoeba triangularis encystation under nutrient starvation.

ABSTRACT:

Background

Curcumin is an active compound derived from turmeric, Curcuma longa, and is known for its benefits to human health. The amoebicidal activity of curcumin against Acanthamoeba triangularis was recently discovered. However, a physiological change of intracellular pathways related to A. triangularis encystation mechanism, including autophagy in the surviving amoeba after curcumin treatment, has never been reported. This study aims to investigate the effect of curcumin on the survival of A. triangularis under nutrient starvation and nutrient-rich condition, as well as to evaluate the A. triangularis encystation and a physiological change of Acanthamoeba autophagy at the mRNA level.

Methods

In this study, A. triangularis amoebas were treated with a sublethal dose of curcumin under nutrient starvation and nutrient-rich condition and the surviving amoebas was investigated. Cysts formation and vacuolization were examined by microscopy and transcriptional expression of autophagy-related genes and other encystation-related genes were evaluated by real-time PCR.

Results

A. triangularis cysts were formed under nutrient starvation. However, in the presence of the autophagy inhibitor, 3-methyladenine (3-MA), the percentage of cysts was significantly reduced. Interestingly, in the presence of curcumin, most of the parasites remained in the trophozoite stage in both the starvation and nutrient-rich condition. In vacuolization analysis, the percentage of amoebas with enlarged vacuole was increased upon starvation. However, the percentage was significantly declined in the presence of curcumin and 3-MA. Molecular analysis of A. triangularis autophagy-related (ATG) genes showed that the mRNA expression of the ATG genes, ATG3, ATG8b, ATG12, ATG16, under the starvation with curcumin was at a basal level along the treatment. The results were similar to those of the curcumin-treated amoebas under a nutrient-rich condition, except AcATG16 which increased later. On the other hand, mRNA expression of encystation-related genes, cellulose synthase and serine proteinase, remained unchanged during the first 18 h, but significantly increased at 24 h post treatment.

Conclusion

Curcumin inhibits cyst formation in surviving trophozoites, which may result from its effect on mRNA expression of key Acanthamoeba ATG-related genes. However, further investigation into the mechanism of curcumin in A. triangularis trophozoites arrest and its association with autophagy or other encystation-related pathways is needed to support the future use of curcumin.

SUBMITTER: Boonhok R

PROVIDER: S-EPMC9261923 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundThe encystation of Acanthamoeba leads to the development of resilient cysts from vegetative trophozoites. This process is essential for the survival of parasites under unfavorable conditions. Previous studies have reported that, during the encystation of A. castellanii, the expression levels of encystation-related factors are upregulated. However, the regulatory mechanisms for their expression during the encystation process remains unknown. Proteins in the sirtuin family, which consists of nicotinamide adenine dinucleotide-dependent deacetylases, are known to play an important role in various cellular functions. In the present study, we identified the Acanthamoeba silent-information regulator 2-like protein (AcSir2) and examined its role in the growth and encystation of Acanthamoeba.MethodsWe obtained the full-length sequence for AcSir2 using reverse-transcription polymerase chain reaction. In Acanthamoeba transfectants that constitutively overexpress AcSir2 protein, SIRT deacetylase activity was measured, and the intracellular localization of AcSir2 and the effects on the growth and encystation of trophozoites were examined. In addition, the sirtuin inhibitor salermide was used to determine whether these effects were caused by AcSir2 overexpression RESULTS: AcSir2 was classified as a class-IV sirtuin. AcSir2 exhibited functional SIRT deacetylase activity, localized mainly in the nucleus, and its transcription was upregulated during encystation. In trophozoites, AcSir2 overexpression led to greater cell growth, and this growth was inhibited by treatment with salermide, a sirtuin inhibitor. When AcSir2 was overexpressed in the cysts, the encystation rate was significantly higher; this was also reversed with salermide treatment. In AcSir2-overexpressing encysting cells, the transcription of cellulose synthase was highly upregulated compared with that of control cells, and this upregulation was abolished with salermide treatment. Transmission electron microscope-based ultrastructural analysis of salermide-treated encysting cells showed that the structure of the exocyst wall and intercyst space was impaired and that the endocyst wall had not formed.ConclusionsThese results indicate that AcSir2 is a SIRT deacetylase that plays an essential role as a regulator of a variety of cellular processes and that the regulation of AcSir2 expression is important for the growth and encystation of A. castellanii.