RESUMEN

The dysregulation of pH has been linked to the onset of chronic conditions, such as cancer and neurological diseases. Consequently, the development of a highly sensitive tool for intracellular pH sensing is imperative to investigate the interplay between pH and the biochemical changes accompanying disease pathogenesis. Here, we present the development of a ratiometric fluorescent nanoprobe, NpRhoDot, designed for precisely measuring pH levels. We demonstrate its efficacy in sensitively reporting intracellular pH in monolayer A549 lung cancer cells, primary fibroblast cells, and 3D tumor spheroids derived from the DLD-1 colorectal adenocarcinoma cell line. NpRhoDot leverages a novel design, where stable carbon dots are functionalized with a pH-responsive ratiometric fluorescent probe comprising a naphthalimide-rhodamine moiety, NpRho1. This design confers NpRhoDot with the high pH sensitivity characteristics of organic fluorescent probes, along with excellent photostability up to 1 h and biocompatibility of carbon dots. Through one-photon and two-photon fluorescence microscopy, we validate the reliability of NpRhoDot for biosensing intracellular pH in monolayer and three-dimensional tumor models from pH 4 to 7.

Asunto(s)

Carbono , Colorantes Fluorescentes , Puntos Cuánticos , Humanos , Concentración de Iones de Hidrógeno , Carbono/química , Colorantes Fluorescentes/química , Puntos Cuánticos/química , Línea Celular Tumoral , Esferoides Celulares/patología , Células A549 , Rodaminas/química

RESUMEN

The COVID-19 pandemic has had a devastating impact on global health, highlighting the need to understand how the SARS-CoV-2 virus damages the lungs in order to develop effective treatments. Recent research has shown that patients with COVID-19 experience severe oxidative damage to various biomolecules. We propose that the overproduction of reactive oxygen species (ROS) in SARS-CoV-2 infection involves an interaction between copper ions and the virus's spike protein. We tested two peptide fragments, Ac-ELDKYFKNH-NH2 (L1) and Ac-WSHPQFEK-NH2 (L2), derived from the spike protein of the Wuhan strain and the ß variant, respectively, and found that they bind Cu(II) ions and form a three-nitrogen complexes at lung pH. Our research demonstrates that these complexes trigger the overproduction of ROS, which can break both DNA strands and transform DNA into its linear form. Using A549 cells, we demonstrated that ROS overproduction occurs in the mitochondria, not in the cytoplasm. Our findings highlight the importance of the interaction between copper ions and the virus's spike protein in the development of lung damage and may aid in the development of therapeutic procedures.

Asunto(s)

COVID-19 , Humanos , SARS-CoV-2/metabolismo , Cobre/química , Especies Reactivas de Oxígeno , Glicoproteína de la Espiga del Coronavirus/química , Pandemias

RESUMEN

A fluorescent, naphthalimide-based, NADH mimic has been synthesised as a reversible, biocompatible, "on-off" probe for the detection of changes in intracellular redox environment (both oxidation and reduction). Interconversion was confirmed by means of electrochemistry and also 1H NMR, UV-vis and fluorescence spectroscopy. The reversibility was also successfully detected in A549 cells under simulated redox stress.