Estudo da ação imunomoduladora de complexos de ciclodextrinas em modelo experimental de esclerose múltipla

Abstract

Multiple sclerosis (MS) is a chronic autoimmune disease that causes neurodegeneration in the central nervous system (CNS), for which conventional treatments are limited by high cost, adverse effects, and partial efficacy. In this context, the search for more effective, safe, and accessible alternatives has led to the investigation of Ursolic Acid (UA) and Rhodium (Rh) compounds. UA possesses well-established anti-inflammatory and immunomodulatory actions; however, its unfavorable physicochemical characteristics present a pharmacotechnical challenge. On the other hand, while Rh complexes with cyclodextrins have previously shown immunomodulatory effects in vitro and in vivo, their efficacy in MS models remained unexplored. Therefore, this study aimed to synthesize, characterize, and evaluate the immunomodulatory effect of a UA complex with poly-beta-cyclodextrin (UA/pβCD) and an Rh complex with hydroxypropyl-beta-cyclodextrin (Rh/HPβCD) in the Experimental Autoimmune Encephalomyelitis (EAE) model. The physicochemical characterization followed a strategic approach: the formation of Rh/HPβCD, building upon previous work, was confirmed by Fourier-transform infrared spectroscopy (FTIR) and thermal analysis (TGA/DTA). The novel UA/pβCD complex was subjected to an in-depth investigation to elucidate its supramolecular behavior, which included, in addition to FTIR and TGA/DTA, phase solubility studies, dynamic light scattering (DLS), and zeta potential analysis. The biological evaluation began in vitro with cytotoxicity (MTT) and nitric oxide (NO) production assays for UA/pβCD, and subsequently proceeded to the in vivo evaluation of both complexes in the EAE model using female C57BL/6 mice. The in-depth characterization of UA/pβCD revealed the formation of an inclusion complex with a 1:1 stoichiometry, resulting in increased UA solubility and the formation of stable colloidal nanoparticles in an aqueous medium. In vitro, the UA/pβCD complex reduced NO production in macrophages, proving more potent than uncomplexed UA and exhibiting no cytotoxicity at therapeutic concentrations. In vivo, both complexes, as well as their parent compounds, significantly attenuated the neurological disability score of EAE. Notably, uncomplexed Rh demonstrated the most rapid and potent action during the inflammatory peak of the disease. Ex vivo analyses of the CNS revealed a dual, compartmentalized action: in the brain, the treatments promoted clear immunomodulation, with a reduction in the pro-inflammatory cytokine IL-12p70 and inflammatory infiltrate. In contrast, in the spinal cord, they exerted a potent neuroprotective effect, decreasing demyelination even without reducing the local inflammatory infiltrate. In conclusion, the UA and Rh complexes attenuate the severity of EAE through a combined mechanism of central immunomodulation and direct neuroprotection. Rh emerged as a highly potent therapeutic agent, while the complexation of UA proved to be a successful pharmacotechnical strategy for both improving physicochemical properties and maintaining efficacy, thus warranting continued investigation into the development of new therapies for MS.