Biology subjects

Lentini, G.

Publications and source records attributed to Lentini, G..

4 recordsLinked to original sources

bioRxiv2025

Revisiting Cysteine Protease Function in Trypanosoma cruzi: Implications for Parasite Egress and Differentiation

Chagas disease is a major global health concern affecting millions of people worldwide, with limited therapeutic options and no vaccine. The causative agent, Trypanosoma cruzi, is a unicellular eukaryotic parasite whose life cycle alternates between insect vectors and a wide range of mammalian hosts. In mammalian cells, parasite proliferation depends on iterative cycles of host cell invasion, intracellular multiplication, differentiation, and host cell rupture, releasing hundreds of infective parasites. The mechanisms governing the critical transition from replicative amastigotes to infective trypomastigotes (trypomastigogenesis) and subsequent egress remain poorly understood, largely due to the lack of robust analytical tools. Here, we combined real-time cellular impedance monitoring, stage-specific fluorescent parasites, ultrastructural expansion microscopy, and automated high-content imaging to dissect the late steps of the lytic cycle. We demonstrate that trypomastigogenesis is temporally coordinated with egress, ensuring the release of fully mature, infective trypomastigotes. Futhermore, we re-evaluate the effect of the cysteine protease inhibitor Z-Phe-Ala fluoromethyl ketone (Z-FA-FMK), previously reported as an egress inhibitor. Our results reveal that Z-FA-FMK primarily impairs trypomastigogenesis, causing an accumulation of amastigotes and blocking progression to mature trypomastigotes. This arrest delays egress and leads to the release of immature forms, highlighting the essential role of cysteine proteases in parasite differentiation. By decoupling trypomastigogenesis from egress, we provide experimental evidence that the late lytic cycle is a multi-step, tightly regulated process linking parasite differentiation with egress. Together, our work establishes a quantitative framework for dissecting lytic cycle termination and offers a versatile platform for phenotypic screening and drug discovery. ImportanceChagas disease, caused by Trypanosoma cruzi, affects millions worldwide and remains a major global health burden, causing chronic cardiac, digestive, and neurological complications. The disease, disproportionately impacting vulnerable populations, lacks effective treatments or a vaccine. Parasite replication and host cell exit are tightly linked, but the mechanisms driving the transition from intracellular replicative parasites to infective forms and their subsequent release upon host cell lysis are poorly understood. Using real-time monitoring, fluorescent parasites, and high-resolution imaging, we show that cysteine proteases are critical for parasite maturation and that their inhibition uncouples differentiation from egress, leading to the release of non-infective parasites. These findings reveal fundamental principles of parasite biology, provide a platform for drug discovery, and highlight new avenues to target Chagas disease at a critical stage of infection.

microbiology↗

bioRxiv2024

Molecular Insights into the Rescue Mechanism of an hERG Activator Against Severe LQT2 Mutations.

BackgroundMutations in the HERG potassium channel are a major cause of long QT syndrome type 2 (LQT2), which can lead to sudden cardiac death. The HERG channel plays a critical role in the repolarization of the myocardial action potential, and loss-of-function mutations prolong cardiac repolarization. MethodsIn this study, we investigated the efficacy and underlying molecular mechanism of ICA-105574, an HERG activator, in shortening the duration of cardiac repolarization in severe LQT2 variants. We characterized the efficacy of ICA-105574 in vivo, using an animal model to assess its ability to shorten the QT interval and in vitro, in cellular models mimicking severe HERG channel mutations (A561V, G628S, and L779P) to evaluate its impact in enhancing IKr current. Additionally, molecular dynamics simulations were used to investigate the molecular mechanism of ICA-105574 action. ResultsIn vivo, ICA-105574 significantly shortened the QT interval. LQT2 mutations drastically reduced IKr amplitude and suppressed tail currents in cellular models. ICA-105574 restored IKr in A561V and G628S. Finally, in silico data showed that ICA-105574 stabilizes a pattern of interactions similar to gain-of-function SQT1 mutations and can reverse the G628S modifications, through an allosteric network linking the binding site to the selectivity filter and the S5P turret helix, thereby restoring its K+ ion permeability. ConclusionsOur results support the development of HERG activators like ICA-105574 as promising pharmacological molecules against some severe LQT2 mutations and suggest that molecular dynamics simulations can be used to test the ability of molecules to modulate HERG function in silico, paving the way for the rational design of new HERG activators.

biophysics↗

bioRxiv2022

Camel urine limits proliferation and modifies cell morphology in human renal tumoral and non-tumoral cells

The widespread ethnomedical practice of dromedary urinotherapy as a remedy against various illnesses is well recognized in traditional dromedary countries, and multiple researchers tried to unravel its bioactive potential and provide scientific evidence through in vivo and in vitro experiments. None of these studies (i) measured urine osmolarity prior to bioactivity testing, which could deeply influence the results of in vitro tests, nor (ii) addressed issues related to cells morphological changes after exposure to camel urines. Taken together, the above aspects point to the need for a "good practice" to be shared by researchers in this field, in order to reduce the variability of in vitro testing of camel urine bioactivity. In this work, using a set of biological samples from animals differing in sex, age, and physiological status, we investigated, the antiproliferative activity of camel urine towards human non-tumoral (HK2) and tumoral renal cells (Caki-1), through cell viability and microscopy analysis, and taking the possible influence of osmolarity into account. We employed cell lines commonly used in toxicological research which, to the best of our knowledge, have not been previously exposed to camel urine. HK2 and Caki-1 cells tolerated well mannitol-induced hyperosmolarity up to 500 mOsm/L. Significant antiproliferative effects were observed only in Caki-1 cells, when exposed to urine solutions (diluted to <500 mOsm/L) from two males out of the ten tested samples, while effects on cell morphology (elongation) were observed only in HK2 cells, when exposed to urine solutions from six samples. The significant antiproliferative effect observed only in tumoral cells looks promising for forthcoming developments in the cancer treatment field. Finally, the presented approach may serve as a guide for future research in this specific, multidisciplinary field.

cell biology↗

bioRxiv2019

Targeting the Pregnane X Receptor Using Microbial Metabolite Mimicry

The human pregnane X receptor (PXR), a master regulator of drug metabolism, has important roles in intestinal homeostasis and abrogating inflammation. Existing PXR ligands have substantial off-target toxicity. Based on prior work that established microbial (indole) metabolites as PXR ligands, we proposed microbial metabolite mimicry as a novel strategy for drug discovery that allows to exploit previously unexplored parts of chemical space. Here we report functionalized indole-derivatives as first-in-class non-cytotoxic PXR agonists, as a proof-of-concept for microbial metabolite mimicry. The lead compound, FKK6, binds directly to PXR protein in solution, induces PXR specific target gene expression in, cells, human organoids, and mice. FKK6 significantly represses pro-inflammatory cytokine production cells and abrogates inflammation in mice expressing the human PXR gene. The development of FKK6 demonstrates for the first time that microbial metabolite mimicry is a viable strategy for drug discovery and opens the door to mine underexploited regions of chemical space.

biochemistry↗