Photoexcited Toluidine Blue Inhibits Tau Aggregation in Alzheimer’s Disease

Abstract

The aggregates of microtubule-associated protein Tau are considered as a major hallmark of Alzheimer’s disease. Tau aggregates accumulate intracellularly leading to neuronal toxicity. Numerous approaches have been targeted against Tau protein aggregation, which include application of synthetic and natural compounds. Toluidine blue is a basic dye of phenothiazine family, which on irradiation with a 630 nm light gets converted into a photoexcited form, leading to generation of singlet oxygen species. Methylene blue is the parent compound of toluidine blue, which has been reported to be potent against tauopathy. In the present work, we studied the potency of toluidine blue and photoexcited toluidine blue against Tau aggregation. Biochemical and biophysical analyses using sodium dodecyl sulfate-polyacrylamide gel electrophoresis, ThS fluorescence, circular dichroism spectroscopy, and electron microscopy suggested that toluidine blue inhibited the aggregation of Tau in vitro. The photoexcited toluidine blue potentially dissolved the matured Tau fibrils, which indicated the disaggregation property of toluidine blue. The cell biology studies including the cytotoxicity assay and reactive oxygen species (ROS) production assay suggested toluidine blue to be a biocompatible dye as it reduced ROS levels and cell death. The photoexcited toluidine blue modulates the cytoskeleton network in cells, which was supported by immunofluorescence studies of neuronal cells. The studies in a UAS Tau E14 transgenic Drosophila model suggested that photoexcited toluidine blue was potent to restore the survival and memory deficits of Drosophila. The overall finding of our studies suggested toluidine blue to be a potent molecule in rescuing the Tau-mediated pathology by inhibiting its aggregation, reducing the cell death, and modulating the tubulin levels and behavioral characteristics of Drosophila. Thus, toluidine blue can be addressed as a potent molecule against Alzheimer’s disease.

Photodynamic exposure of Rose-Bengal inhibits Tau aggregation and modulates cytoskeletal network in neuronal cells

“…The cell viability was analyzed by methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay. The method was followed according to published protocol 20 . Neuro2a cells (ATCC CCL-131) were cultured in advanced DMEM F-12 media supplemented with 10% FBS and glutamine.…”

“…Fly cultures and crosses were carried out at 25 °C. The stocks has been maintained as per published protocol 20 , 64 .…”

“…The obtained eggs were transferred at a density of 50-eggs/6 mL of SM and allowed to develop till early third instar(as referred in 20 ). The early third instar larvae were removed from the SM vials and used in the feeding behaviour assay.…”

“…The ability to move against gravity and climb indicates the level of physical fitness of test animals(as referred in 20 ). The vertical climbing ability of male flies that emerged from different treatment bottles were assessed.…”

“…Moreover, metal complexes and metal nanoparticles are also reported to have potency against Tau aggregation 16 , 17 . Several model systems have been studied for screening therapeutic molecules, the in-vitro heparin-induced Tau aggregation and transgenic E14 Drosophila is widely examined as a model system for Tauopathy 18 – 20 . Rose Bengal (RB) is a xanthene dye, which is known to possess the property of photo-excitation and hence it has been widely used as a photo-sensitizer 21 .…”