Glycogen synthase kinase GSK is

Glycogen synthase kinase-3β (GSK-3β) is an evolutionarily conserved serine/threonine kinase that plays multifaceted role in diverse cellular and neurophysiological processes [8]. GSK-3β is regulated by inhibitory serine and stimulatory tyrosine phosphorylation on Ser9 and Tyr216 respectively [9]. Dysregulation of GSK-3β has been implicated in diabetes [10], mood disorders [11] and Alzheimer's disease (AD) [12]. Active GSK-3β in CGS 21680 stimulates the molecular machinery responsible for amyloid deposition and microglia-mediated neuroinflammation [13], and is pro-apoptotic [14]. Conversely, treatment with GSK 3β inhibitor has been reported to prevent Aβ accumulation in transgenic mice over-expressing GSK3β [12]. Inhibition of GSK-3β using short interference RNA (SiRNA) reduced NO production, but increased expression of IL-10 in both BV-2 cells and primary rat microglia cultures [15]. Since vast number of signaling pathways converge on GSK-3β, GSK-3β has turned out to be an important therapeutic target. Indirubin extracted from the plants such as Polygonum tinctorium and Isatis indigotica or the gastropod molluscs of the Muricidae family (Hexaplus trunculus and Morula granulata) has been used as an active ingredient in traditional Chinese medicine to prevent inflammatory diseases and leukemia [16,17]. Indirubin derivatives have been shown to inhibit cyclin dependent kinase-5 and GSK3β activities [18]. Studies reveal that indirubin-3′-monoxime (IMX) can pass blood brain barrier (BBB) [19], can attenuate amyloid toxicity and neuronal apoptosis in vitro [20,21] and prevent memory deficits in vivo [22]. The underlying molecular mechanisms behind the neuroprotective effects of IMX in rodent models still remain largely unknown. The present study aims to explore the extent of neuropathological changes induced by a combination of high fat-high fructose diet, and to resolve the effects of IMX intervention on amyloid deposition and glial cell-mediated neuroinflammation and apoptosis.
Materials and methods
Results
Discussion Previous studies from our laboratory have clearly documented that HFFD causes body weight gain and brings out metabolic changes (elevated levels of glucose, insulin, triglycerides, free fatty acids, total cholesterol, TNF-α and IL-6) in circulation and peripheral tissues [[24], [25], [26], [27], [28]]. Hematoxyin and eosin stained liver and kidney sections of HFFD mice displayed inflammatory infiltration of liver and kidney tubules and capsules (data not shown). Neuronal function is compromised during calorie excess and in the long-term may potentiate neurodegeneration. Of late, HFD has been employed to study the pathogenesis of neurodegenerative diseases in metabolically challenged rodents [3,29] and few studies have also demonstrated the detrimental effects of sucrose/fructose to brain [[30], [31], [32]]. Diet rich in fat or simple sugars stimulate amyloid deposition and glial cell activation in brain [1,[33], [34], [35]], which are the hallmarks of AD [36]. Gut dysbiosis is one mechanism through which HFFD may promote neuroinflammation. Perturbations in gut microbial diversity (changes in Firmicutes vs Baceteriodetes ratio) can alter gut epithelial integrity [37,38], that can release proinflammatory cytokines and microbial endotoxins into systemic circulation [39]. Endotoxaemia and inflammation cause leaky blood brain barrier, accentuating neuroinflammatory response via toll-like receptors that contributes to Aβ deposition and glial cell activation [40]. Aβ accumulation provokes glial cell activation in mice hippocampus [41], and exert proinflammatory functions [42,43]. Aβ triggers transformation of quiescent glial cells to a reactive phenotype which clear Aβ aggregates and dead or dying neurons, but sustained activation result in loss of its phagocytic function, elevated secretion of neurotoxic substances and stimulation of Aβ synthesis [44]. Thus, Aβ deposition rewound neuronal injury, which in turn generates more Aβ resulting in a vicious cycle. Concomitant to amyloid deposition, intense DAB staining for immunohistochemical localization of GFAP and CD-68 in brain of HFFD mice was observed flaunting glial cell activation and proliferation.