Activity and Cell-Death Pathway in Leishmania infantum Induced by Sugiol: Vectorization Using Yeast Cell Wall Particles Obtained From Saccharomyces cerevisiae

Abstract

Visceral leishmaniasis, caused by Leishmania infantum , is a neglected tropical disease, to which efforts in the innovation of effective and affordable treatments remain limited, despite the rising incidence in several regions of the world. In this work, the antileishmanial effects of sugiol were investigated in vitro . This compound was isolated from the bark of Cupressus lusitanica and showed promising activity against L. infantum . In spite of the positive results, it is known that the compound is a poorly water-soluble diterpene molecule, which hinders further investigation, especially in preclinical animal studies. Thus, in an alternative delivery method, sugiol was entrapped in glucan-rich particles obtained from Saccharomyces cerevisiae yeast cell walls (YCWPs). To evaluate the activity of sugiol, the experiments were divided into two parts: (i) the in vitro investigation of antileishmanial activity of free sugiol against L. infantum promastigotes after 24, 48, and 72 h of treatment and (ii) the evaluation of antileishmanial activity of sugiol entrapped in glucan-rich particles against intracellular L. infantum amastigotes. Free sugiol induced the cell-death process in promastigotes, which was triggered by enhancing cytosolic calcium level and promoting the autophagy up to the first 24 h. Over time, the presence of autophagic vacuoles became rarer, especially after treatment with lower concentrations of sugiol, but other cellular events intensified, like ROS production, cell shrinkage, and phosphatidylserine exposure. Hyperpolarization of mitochondrial membrane potential was found at 72 h, induced by the mitochondria calcium uptake, causing an increase in ROS production and lipid peroxidation as a consequence. These events resulted in the cell death of promastigotes by secondary necrosis. Sugiol entrapped in glucan-rich particles was specifically recognized by dectin-1 receptor on the plasma membrane of macrophages, the main host cell of Leishmania spp. Electron micrographs revealed particles containing sugiol within the infected macrophages and these particles were active against the intracellular L. infantum amastigotes without affecting the host cell. Therefore, the YCWPs act like a Trojan horse to successfully deliver sugiol into the macrophage, presenting an interesting strategy to deliver water-insoluble drugs to parasitized cells.

PF-429242, a Subtilisin Inhibitor, Is Effective in vitro Against Leishmania infantum

“…Furthermore, PF-429242 treatment induced oxidative stress in L. infantum promastigotes. Hyperpolarization of ΔΨm is able to cause an increase of ROS production as was already observed during hyperglycemic conditions ( Yu et al, 2006 ) and in L. infantum promastigotes treated with sugiol, a compound extracted from the bark of Cupressus lusitanica ( Scariot et al, 2019 ). Thus, the hyperpolarization of ΔΨm could cause the increase in ROS production after treatment with PF-429242.…”

Total Phenolic Fraction (TPF) from Extra Virgin Olive Oil: Induction of apoptotic-like cell death in Leishmania spp. promastigotes and in vivo potential of therapeutic immunomodulation

“…At the early event of apoptosis, the plasma membrane still excludes viability assays such as PI, so single staining with annexin V serves as an early marker of apoptosis, while in the late stage of apoptosis, the membrane loses its integrity and the late apoptotic/secondary apoptotic cells, become positive for both annexin V and PI [ 55 , 62 ]. Consequently, the absence of PI staining signals membrane integrity while necrotic cells become PI-permeable even though they remain negative to annexin V. However, it should be acknowledged that the exposure of PS in Leishmania promastigotes as a marker of apoptosis-like cell death is still a controversial subject [ 63 ]. Research models based on the detection of PS exposure suggest that Leishmania parasites mimic mammalian apoptotic cells by exposing PS at their cell surface to trigger phagocytic uptake into host macrophages [ 64 – 67 ].…”

Yeast Cells in Microencapsulation. General Features and Controlling Factors of the Encapsulation Process

“…Another solution is to induce the precipitation of the active compound. For example, albeit with low efficiency, a scarcely water-soluble drug such as sugiol can be accumulated in YCWs by repeated cycles of exposure of YCWs to an organic solution (e.g., acetone) and subsequent drying [ 116 ]. The release of the active would then be controlled by its own solubility; for example, a moderately soluble molecule such as ibuprofen is rapidly released, reaching saturation within minutes [ 93 ].…”