IIluminando novas fronteiras: explorando o potencial fotossensibilizador de espécies de Passiflora no controle de infecções por Staphylococcus aureus resistente à meticilina em camundongos senescentes

Abstract

INTRODUCTION: Antimicrobial Photodynamic Therapy (aPDT) has emerged as a potential alternative for the treatment of multidrug-resistant bacterial skin infections, such as those caused by methicillin-resistant Staphylococcus aureus (MRSA), which pose a high risk in elderly individuals. This approach is based on the use of a chemical agent known as a photosensitizer (PS), considered one of the main components of aPDT. OBJECTIVE: This study aimed to evaluate the photosensitizing activity of Passiflora genus extracts, both incorporated and not incorporated into an emulsion, for controlling MRSA infection in vitro and in vivo using a senescence model. METHODOLOGY: The research methodology comprised in vitro and in vivo stages, in addition to the formulation and characterization of a topical emulsion. Initially, in the in vitro assays, the light absorption spectrum of Passiflora extracts and the effective dose for their photoactivation were determined. Additionally, the cytotoxicity of these extracts on eukaryotic cells was evaluated, and the intracellular localization of the butanolic fraction (BF) of P. cincinnata in MRSA was investigated. Subsequently, in the in vivo study, senescent animals intradermically infected in the ear with MRSA were randomly distributed into three experimental groups: Control (receiving only the vehicle); Treated with nonphotoactivated P. cincinnata butanolic fraction (BF); and Treated with photoactivated P. cincinnata BF (APDT). Throughout the in vivo study, cytokine levels in retromaxillary lymph nodes, animal body weight, bacterial load at the infection site, and the degree of ear tissue inflammation were measured. Finally, for the developed topical formulation, a preliminary stability test was conducted, followed by its physicochemical characterization. RESULTS: Passiflora extracts were considered non-cytotoxic to HUVEC (Human Umbilical Vein Endothelial Cells) cell cultures. P. edulis and P. alata extracts showed photosensitizing capacity at a concentration of 1000 µg/mL. The butanolic fraction of P. cincinnata exhibited significant photosensitizing activity at a concentration of 100 µg/mL. Meanwhile, the crude extract (CE) demonstrated this activity at concentrations lower than BF, at 5-50 µg/mL. Isoorientin, the major compound present in P. cincinnata BF, did not show as significant a reduction as P. cincinnata CE and BF. Animals treated with P. cincinnata BF showed better control of bacterial load, less leukocyte infiltration, and less weight loss throughout the MRSA infection process. Furthermore, the APDT group showed a more distinct pattern of cytokine correlations and influence, with a greater number of cytokines exhibiting negative correlations, for example, both among pro-inflammatory cytokines and with the anti-inflammatory cytokine IL-10. Dendrogram analysis and Principal Component Analysis showed that leukocyte levels in the ear are important for the course of infection among the groups, with differences regarding treated or untreated animals. Zeta potential analysis of MRSA revealed that the butanolic fraction of P. cincinnata does not significantly alter the bacterial membrane potential. In the developed formulation, an oil-in-water emulsion compatible system, statistically significant variations did not show changes that would compromise the formulation. The fluidity index values indicated pseudoplastic fluid behavior. Additionally, the emulsion showed thixotropic behavior, a typical characteristic of non-Newtonian fluids. CONCLUSION: The results are promising, given the limited understanding of APDT in senescent animals, and highlight P. cincinnata as a potential photosensitizer against MRSA. This research lays the foundation for developing targeted APDT strategies using Passiflora species and emphasizes the need to explore new photosensitizers derived from underexplored plants. Furthermore, the standardization of a topical formulation, demonstrating stability at room temperature and suitable rheological properties, paves the way for the development of a new therapeutic approach for MRSA skin infections, especially in elderly individuals.