Prophylactic effects of cucurbitacin B in the EAE Model of multiple sclerosis by adjustment of STAT3/IL-23/IL-17 axis and improvement of neuropsychological symptoms.

Multiple sclerosis (MS) is an autoimmune disease that affects the central nervous system. Although remarkable progress has been made in treating MS, current therapies are less effective in protecting against the progression of the disease. Since cucurbitacins have shown an extreme range of pharmacological properties, in this study, we aimed to investigate the prophylactic effect of cucurbitacin B (CuB) in the experimental MS model. Experimental autoimmune encephalomyelitis (EAE) induced by subcutaneous immunization of MOG35-55 in C57BL/6 mice. CuB interventions (0.5 and 1 mg/kg, i.p.) were performed every other day from the first day of EAE induction. Assessment of clinical scores and motor function, inflammatory responses, and microglial activation were assessed by qRT-PCR, western blotting, and immunohistochemical (IHC) analyses. CuB (1 mg/kg) significantly decreased the population of CD45+ (P < 0.01), CD11b+ (P < 0.01) and CD45+/CD11b+ (P < 0.05) cells in cortical lesions of EAE mice. In addition, activation of STAT3 (P < 0.001), expression of IL-17 A and IL-23 A (both mRNA and protein), and transcription of Iba-1 significantly decreased. On the contrary, CuB (1 mg/kg) significantly increased the transcription of MBP and Olig-2. Furthermore, a significant decrease in the severity of EAE (P < 0.05), and an improvement in motor function (P < 0.05) and coordination (P < 0.05) were observed after treatment with a high dose of CuB. Our results suggest that CuB may have a wide-ranging effect on autoimmune responses in MS via a reduction in STAT3 activation, microgliosis, and adaptation of the IL-23/IL-17 axis. Further studies are needed to investigate the exact effect of CuB in glial cells and its efficiency and bioavailability in other neuroinflammatory diseases.

α-Synuclein E46K Mutation and Involvement of Oxidative Stress in a Drosophila Model of Parkinson's Disease.

Parkinson's disease (PD) is an age-associated neurodegenerative condition in which some genetic variants are known to increase disease susceptibility on interaction with environmental factors inducing oxidative stress. Different mutations in the SNCA gene are reported as the major genetic contributors to PD. E46K mutation pathogenicity has not been investigated as intensive as other SNCA gene mutations including A30P and A53T. In this study, based on the GAL4-UAS binary genetic tool, transgenic Drosophila melanogaster flies expressing wild-type and E46K-mutated copies of the human SNCA gene were constructed. Western blotting, immunohistochemical analysis, and light and confocal microscopy of flies' brains were undertaken along with the survival rate measurement, locomotor function assay, and ethanol and paraquat (PQ) tolerance to study α-synuclein neurotoxicity. Biochemical bioassays were carried out to investigate the activity of antioxidant enzymes and alterations in levels of oxidative markers following damages induced by human α-synuclein to the neurons of the transgenic flies. Overexpression of human α-synuclein in the central nervous system of these transgenic flies led to disorganized ommatidia structures and loss of dopaminergic neurons. E46K α-synuclein caused remarkable climbing defects, reduced survivorship, higher ethanol sensitivity, and increased PQ-mediated mortality. A noticeable decline in activity of catalase and superoxide dismutase enzymes besides considerable increase in the levels of lipid peroxidation and reactive oxygen species was observed in head capsule homogenates of α-synuclein-expressing flies, which indicates obvious involvement of oxidative stress as a causal factor in SNCA E46K neurotoxicity. In all the investigations, E46K copy of the SNCA gene was found to impose more severe defects when compared to wild-type SNCA. It can be concluded that the constructed Drosophila models developed PD-like symptoms that facilitate comparative studies of molecular and cellular pathways implicated in the pathogenicity of different α-synuclein mutations.

Sepantronium Bromide (YM155), A Small Molecule Survivin Inhibitor, Promotes Apoptosis by Induction of Oxidative Stress, Worsens the Behavioral Deficits and Develops an Early Model of Toxic Demyelination: In Vivo and In-Silico Study.

Cuprizone (cup) model targets oligodendrocytes (OLGs) degeneration and is frequently used for the mechanistic understanding of de- and remyelination. Improperly, this classic model is time-consuming and the extent of brain lesions and behavioral deficits are changeable (both temporally and spatially) within a mouse strain. We aimed to offer an alternative, less time-consuming, and more reproducible cup model. Mice (C57BL/6) were treated with cup (400 mg kg-1 day-1/gavage) for three consecutive weeks to induce OLGs degeneration with or without YM155 (1 mg kg-1 day-1) to examine the effects of this molecule in cup neurotoxicity. Co-administration of cup and YM155 (cuYM) accelerated the intrinsic apoptosis of mature OLGs (MOG positive cells) through the upregulation of caspase-9 and caspase-3. In addition to the stimulation of oxidative stress via reduction of glutathione peroxidase and induction of malondialdehyde, behavioral deficits in both Open-field and Rota-rod tests were worsened by cuYM. In the cuYM group, the expression of BIRC5, BIRC4 and NAIP was reduced, but no significant changes were observed in the abundance of the other inhibitor of apoptosis proteins (cIAP1 and cIAP2) in comparison with the cup group. Moreover, in silico analysis validated that YM155 directly interrupts the binding sites of certain transcription factors, such as krüppel-like family (Klf), specificity proteins (SPs), myeloid zinc fingers (MZFs), zinc finger proteins (ZNFPs), and transcription factor activating enhancer-binding proteins (TFAPs), on the promoters of target genes. In conclusion, this modified model promotes cup-induced redox and apoptosis signaling, elevates behavioral deficits, saves time, minimizes variations, and can be employed for early evaluation of novel neuroprotective agents in oligodendropathies.