Year: 2024
Type: article
Abstract: This research investigated the cytotoxicity of chitosan nanoparticles loaded with lysozyme, exploring their viability for biomedical applications. It focused on retinal pigment epithelial cells (ARPE-19), relevant to ocular diseases. DAPI and phalloidin staining techniques along with fluorescence microscopy were utilized to evaluate cellular morphological changes indicative of cytotoxicity. In the ongoing quest for effective and safe therapeutic alternatives, biomedical research has turned its attention to developing innovative drug delivery systems. In this context, chitosan nanoparticles loaded with lysozyme have emerged as a promising option due to their unique properties. This study aims to determine the toxicity of chitosan-based nanoparticles loaded with lysozyme, a naturally antimicrobial enzyme. Chitosan, a polymer derived from chitin, which has demonstrated to be a promising option for drug delivery due to its biocompatibility and drug-loading capacity, was used. Additionally, it was loaded with lysozyme, an enzyme with antimicrobial and anti-inflammatory properties, which could further enhance the therapeutic properties of the nanoparticles [1, 2]. Chitosan-carrageenan nanoparticles (Cs-Crg) and chitosan-carrageenan-lysozyme particles (CS-Crg-LZ) were synthesized using the ionic complexation method, with carrageenan as the cross-linking agent. This was based on a previous study by Hernandez et al. (2018), where CS-LZ and CS-LZ-Crg nanoparticles were obtained with sizes of 725±5.5 nm and 250±8.1 nm, respectively, analyzed by DLS. Additionally, in the same study, samples were analyzed via FT-IR to determine interactions between components, showing characteristic bands of chitosan, lysozyme, and carrageenan [3]. In this research, the biological activity of the synthesized particles was evaluated for their effect on subcellular structures of ARPE-19 cells, analyzed by fluorescence microscopy. They were incubated in 25 cm^3 culture flasks in DMEM (Dulbecco’s Modified Eagle’s Medium) supplemented with 5% fetal bovine serum, 100 U/mL penicillin, at 37°C in a 5% CO2 atmosphere and 38% humidity [4]. Subsequently, they were exposed to the synthesized particles to analyze their behavior and morphological changes at 0, 12, and 24 hours, with water as a control. DAPI staining enabled observation of cellular nuclei, while phalloidin staining facilitated visualization of the actin cytoskeleton (fig. 1). The absence of cellular morphological changes among the three treatments at different incubation times with the particles suggests that ARPE-19 cells maintained their structural and functional integrity after exposure to chitosan nanoparticles with lysozyme. The integrity of these cellular structures is crucial for normal cellular function, and the lack of alterations in their morphology indicates the absence of cytotoxicity. This is crucial for biomedical applications, as it implies that these nanoparticles can interact with target cells without causing significant harm [5]. Effect of Cs-Crg and CS-LZ particles and morphological changes of ARPE-19 cells in DMEM medium at 12 h. Visualization of actin cytoskeleton stained with phalloidin (red) and DNA stained with DAPI (blue). Water control.
Source: Microscopy and Microanalysis
Language: English
Subfield: Pharmaceutical Science
Domain: Life Sciences
Open Access status: bronze
OpenAlex ID: W4401005896