Anti-Acanthamoeba activity of a semi-synthetic mangostin derivative and its ability in removal of Acanthamoeba triangularis WU19001 on contact lens

Garcinia mangostana L., also known as the mangosteen tree, is a native medicinal plant in Southeast Asia having a wide variety of pharmacologically active compounds, including xanthonoid mangostin. In this study, we examined the pharmacological activities of the selected semi-synthetic mangostin derivative, namely, amoebicidal activity, encystation inhibition, excystation activity, and removal capacity of adhesive Acanthamoeba from the surface of contact lens (CL). Among the three derivatives, C1 exhibited promising anti-Acanthamoeba activity against Acanthamoeba triangularis WU19001 trophozoites and cysts. SEM images displayed morphological changes in Acanthamoeba trophozoites, including the loss of acanthopodia, pore formation in the cell membrane, and membrane damage. In addition, the treated cyst was shrunken and adopted an irregular flat cyst shape. Under a fluorescence microscope, acridine orange and propidium iodide (AO/PI) staining revealed C1 induced condensation of cytoplasm and chromatin with the loss of cell volume in the treated trophozoites, while calcofluor white staining demonstrated the leakage of cell wall in treated cysts, leading to cell death. Interestingly, at the concentration ranges in which C1 showed the anti-Acanthamoeba effects (IC50 values ranging from 0.035–0.056 mg/mL), they were not toxic to Vero cells. C1 displayed the highest inhibitory effect on A. triangularis encystation at 1/16×MIC value (0.004 mg/mL). While C1 demonstrated the excystation activity at 1/128×MIC value with a high rate of 89.47%. Furthermore, C1 exhibited the removal capacity of adhesive Acanthamoeba from the surface of CL comparable with commercial multipurpose solutions (MPSs). Based on the results obtained, C1 may be a promising lead agent to develop a therapeutic for the treatment of Acanthamoeba infections and disinfectant solutions for CL

Targeting Acanthamoeba proteins interaction with flavonoids of Propolis extract by in vitro and in silico studies for promising therapeutic effects

Background: Propolis is a natural resinous mixture produced by bees. It provides beneficial effects on human health in the treatment/management of many diseases. The present study was performed to demonstrate the anti-Acanthamoeba activity of ethanolic extracts of Propolis samples from Iran. The interactions of the compounds and essential proteins of Acanthamoeba were also visualized through docking simulation. Methods: The minimal inhibitory concentrations (MICs) of Propolis extract against Acanthamoeba trophozoites and cysts was determined in vitro. In addition, two-fold dilutions of each of agents were tested for encystment, excystment and adhesion inhibitions. Three major compounds of Propolis extract such as chrysin, tectochrysin and pinocembrin have been selected in molecular docking approach to predict the compounds that might be responsible for encystment, excystment and adhesion inhibitions of A. castellanii. Furthermore, to confirm the docking results, molecular dynamics (MD) simulations were also carried out for the most promising two ligand-pocket complexes from docking studies. Results: The minimal inhibitory concentrations (MICs) 62.5 and 125 µg/mL of the most active Propolis extract were assessed in trophozoites stage of Acanthamoeba castellanii ATCC30010 and ATCC50739, respectively. At concentrations lower than their MICs values (1/16 MIC), Propolis extract revealed inhibition of encystation. However, at 1/2 MIC, it showed a potential inhibition of excystation and anti-adhesion. The molecular docking and dynamic simulation revealed the potential capability of Pinocembrin to form hydrogen bonds with A. castellanii Sir2 family protein (AcSir2), an encystation protein of high relevance for this process in Acanthamoeba. Conclusions: The results provided a candidate for the development of therapeutic drugs against Acanthamoeba infection. In vivo experiments and clinical trials are necessary to support this claim

Effects of Kaempferia parviflora Wall. Ex. Baker on electrophysiology of the swine hearts

Background & objectives: Pre-clinical studies in swine have demonstrated that a supratherapeutic concentration of sildenafil citrate decreased defibrillation efficacy and facilitated cardiac arrhythmia. We therefore, decided to investigate the effects of Kaempferia parviflora (KP) extract on these parameters in the swine heart. The underlying assumption was that in the heart, KP might be producing effects similar to sildanafil citrate as KP has long been used in southeast Asian traditional medicine to correct erectile dysfunction. Methods: The study was conducted as the defibrillation study, and ventricular fibrillation (VF) induction study. In both studies, the defibrillation threshold (DFT), the upper limit of vulnerability (ULV) and VF threshold were determined before and after KP extract administration. Results: In both studies KP extract at high concentrations (100 and 50 mg/kg) significantly increased the DFT and ULV, without altering the VF threshold. At these concentrations, systolic and diastolic blood pressures were also attenuated. Interpretation & conclusions: High concentrations of KP extract attenuated defibrillation efficacy and increased cardiac vulnerability to arrhythmia in a normal swine heart. When used in appropriate concentrations, its blood pressure lowering effect may be useful in hypertensive states. Further studies need to be done to elucidate its mechanism of action

Ligation of Na, K ATPase β3 subunit on monocytes by a specific monoclonal antibody mediates T cell hypofunction

<div><p>T cells play a crucial role in orchestrating body immune responses. T cell hyperfunction, however, leads to inflammation and induction of autoimmune diseases. Understanding of T cell regulation mechanisms and successful modulation of T cell responses is beneficial in treatment of disease associated to T cell hyperresponsiveness. Our previous study indicated that monoclonal antibody (mAb) P-3E10, a mAb to Na, K ATPase β3 subunit, inhibited anti-CD3-induced PBMC proliferation. In the current study, we further investigated the mechanism of mAb P-3E10 in the induction of T cell hypofunction. We demonstrated that mAb P-3E10 decreased T cell proliferation and Th1, Th2 and Th17 cytokine production. Monocytes were the cells playing a key role in mediation of mAb P-3E10 induced T cell hypofunction. The inhibition of T cell activation by mAb P-3E10 required cell contact between monocytes and T cells. The mAb P-3E10 induced the down-expression level of MHC class II and CD86 and increased IL-6, IL-10 and TNF-α production of monocytes. We concluded that ligation of the Na, K ATPase β3 subunit on monocytes by mAb P-3E10 arbitrated T cell hypofunction. This mAb might be a promising novel immunotherapeutic antibody for the treatment of hyperresponsive T cell associated diseases.</p></div

Ligation of Na, K ATPase β3 subunit on monocytes downregulates MHC class II and CD86 expressions and upregulates IL-6, IL-10 and TNF-α production.

<p>(A) PBMCs were stimulated with anti-CD3 mAb in the absence (medium control) or presence of mAb P-3E10 or isotype-matched control mAb. The surface expression levels of MHC class I (HLA-ABC) (<i>n</i> = 4), MHC class II (HLA-DR) (<i>n</i> = 3) and CD86 (<i>n</i> = 4) on CD14⁺ monocytes were determined by flow cytometry. The relative geometric mean fluorescence intensity (GeoMFI of specific marker mAb staining/GeoMFI of isotype-matched control mAb staining) was normalized to medium control as 1. Unpaired t-test was used in the comparison. *P<0.05. **P<0.01. (B) PBMCs were stimulated with anti-CD3 mAb or kept unstimulated in the absence (medium control) or presence of mAb P-3E10 or isotype-matched control mAb. The % IL-6, % IL-10 and % TNF-α producing CD14⁺ monocytes in the indicated conditions are shown as the representative result from one of the four individuals. (C) The data were analyzed as the percentage of cytokine positive cells (% specific cytokine staining—% isotype-matched control staining). The percentage of cytokine positive cells were normalized to isotype-matched mAb treated control of each activation condition as 100%. The data are expressed as mean ± SE (<i>n</i> = 4). Unpaired t-test was performed. *P<0.05. ***P<0.001. The flow cytometric data were illustrated in the Supporting Information (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0199717#pone.0199717.s004" target="_blank">S4 Fig</a>).</p

The inhibition of T cell activation by mAb P-3E10.

<p>(A) PBMCs were activated with anti-CD3 mAb OKT3 or kept unstimulated in the presence of mAb P-3E10 or mAb 13M (isotype-matched control mAb) or medium alone. Flow cytometric data were expressed in histograms showing the percentage of divided cells in each condition using CFSE proliferation assay. (B) The percentage of divided cells upon OKT3-induced PBMC proliferation in the presence of indicated concentrations of mAb P-3E10 or isotype-matched control mAb is shown. Each individual data was normalized relative to its medium control as 100%. Unpaired <i>t</i>-test was used for comparison, *<i>P</i><0.05. ***<i>P</i><0.001. (C) After stimulation as was described in (A), CD3⁺ T cells were gated. The percentage of the indicated cytokine producing T cells was determined and is shown as mean ± SE from three individuals. The data were exhibited as the percentage of cytokine positive cells by normalization to medium control of OKT3-stimulated condition as 100%. Two-way ANOVA followed by Tukey’s test was used for comparison, *<i>P</i><0.05. **<i>P</i><0.01. ***<i>P</i><0.001. The flow cytometric data were illustrated in the Supporting Information (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0199717#pone.0199717.s002" target="_blank">S2 Fig</a>).</p

Targeting Acanthamoeba proteins interaction with flavonoids of Propolis extract by in vitro and in silico studies for promising therapeutic effects [version 2; peer review: 2 approved]

Background: Propolis is a natural resinous mixture produced by bees. It provides beneficial effects on human health in the treatment/management of many diseases. The present study was performed to demonstrate the anti-Acanthamoeba activity of ethanolic extracts of Propolis samples from Iran. The interactions of the compounds and essential proteins of Acanthamoeba were also visualized through docking simulation. Methods: The minimal inhibitory concentrations (MICs) of Propolis extract against Acanthamoeba trophozoites and cysts was determined in vitro. In addition, two-fold dilutions of each of agents were tested for encystment, excystment and adhesion inhibitions. Three major compounds of Propolis extract such as chrysin, tectochrysin and pinocembrin have been selected in molecular docking approach to predict the compounds that might be responsible for encystment, excystment and adhesion inhibitions of A. castellanii. Furthermore, to confirm the docking results, molecular dynamics (MD) simulations were also carried out for the most promising two ligand-pocket complexes from docking studies. Results: The minimal inhibitory concentrations (MICs) 62.5 and 125 µg/mL of the most active Propolis extract were assessed in trophozoites stage of Acanthamoeba castellanii ATCC30010 and ATCC50739, respectively. At concentrations lower than their MICs values (1/16 MIC), Propolis extract revealed inhibition of encystation. However, at 1/2 MIC, it showed a potential inhibition of excystation and anti-adhesion. The molecular docking and dynamic simulation revealed the potential capability of Pinocembrin to form hydrogen bonds with A. castellanii Sir2 family protein (AcSir2), an encystation protein of high relevance for this process in Acanthamoeba. Conclusions: The results provided a candidate for the development of therapeutic drugs against Acanthamoeba infection. In vivo experiments and clinical trials are necessary to support this claim