Inhibition Of Glycogen Synthase Kinase-3β Research Articles

CHIR99021 causes inactivation of Tyrosine Hydroxylase and depletion of dopamine in rat brain striatum

CHIR99021, also known as laduviglusib or CT99021, is a Glycogen-synthase kinase 3β (GSK3β) inhibitor, which has been reported as a promising drug for cardiomyocyte regeneration or treatment of sensorial hearing loss. Since the activation of dopamine (DA) receptors regulates dopamine synthesis and they can signal through the β-arrestin pathway and GSK3β, we decided to check the effect of GSK3β inhibitors (CHIR99021, SB216763 and lithium ion) on the control of DA synthesis. Using ex vivo experiments with minces from rat brain striatum, we observed that CHIR99021, but not SB216763 or lithium, causes complete abrogation of both DA synthesis and accumulation, pointing to off-target effects of CHIR99021. This decrease can be attributed to tyrosine hydroxylase (TH) inhibition since the accumulation of l-DOPA in the presence of a DOPA decarboxylase inhibitor was similarly decreased. On the other hand, CHIR99021 caused a dramatic increase in the DOPAC/DA ratio, an indicator of DA metabolization, and hindered DA incorporation into striatum tissue. Tetrabenazine, an inhibitor of DA vesicular transport, also caused DA depletion and DOPAC/DA ratio increase to the same extent as CHIR99021. In addition, both CHIR99021 or SB216763, but not lithium, decreased TH phosphorylation in Ser19, but not in Ser31 or Ser40. These results demonstrate that CHIR99021 can lead to TH inactivation and DA depletion in brain striatum, opening the possibility of its use in DA-related disorders, and shows effects to be considered in future clinical trials. More work is needed to find the mechanism exerted by CHIR99021 on DA accumulation.

E-cadherin/β-catenin expression is conserved in human and rat erythropoiesis and marks stress erythropoiesis

AbstractE-cadherin is a crucial regulator of epithelial cell-to-cell adhesion and an established tumor suppressor. Aside epithelia, E-cadherin expression marks the erythroid cell lineage during human but not mouse hematopoiesis. However, the role of E-cadherin in human erythropoiesis remains unknown. Because rat erythropoiesis was postulated to reflect human erythropoiesis more closely than mouse erythropoiesis, we investigated E-cadherin expression in rat erythroid progenitors. E-cadherin expression is conserved within the erythroid lineage between rat and human. In response to anemia, erythroblasts in rat bone marrow (BM) upregulate E-cadherin as well as its binding partner β-catenin. CRISPR/Cas9–mediated knock out of E-cadherin revealed that E-cadherin expression is required to stabilize β-catenin in human and rat erythroblasts. Suppression of β-catenin degradation by glycogen synthase kinase 3β (GSK3β) inhibitor CHIR99021 also enhances β-catenin stability in human erythroblasts but hampers erythroblast differentiation and survival. In contrast, direct activation of β-catenin signaling, using an inducible, stable β-catenin variant, does not perturb maturation or survival of human erythroblasts but rather enhances their differentiation. Although human erythroblasts do not respond to Wnt ligands and direct GSK3β inhibition even reduces their survival, we postulate that β-catenin stability and signaling is mostly controlled by E-cadherin in human and rat erythroblasts. In response to anemia, E-cadherin–driven upregulation and subsequent activation of β-catenin signaling may stimulate erythroblast differentiation to support stress erythropoiesis in the BM. Overall, we uncover E-cadherin/β-catenin expression to mark stress erythropoiesis in rat BM. This may provide further understanding of the underlying molecular regulation of stress erythropoiesis in the BM, which is currently poorly understood.

MicroRNAs as Potential Regulators of GSK-3β in Renal Cell Carcinoma.

The prognosis of patients with advanced renal cell carcinoma (RCC) has improved with newer therapies, including molecular-targeted therapies and immuno-oncology agents. Despite these therapeutic advances, many patients with metastatic disease remain uncured. Inhibition of glycogen synthase kinase-3β (GSK-3β) is a promising new therapeutic strategy for RCC; however, the precise regulatory mechanism has not yet been fully elucidated. MicroRNAs (miRNAs) act as post-translational regulators of target genes, and we investigated the potential regulation of miRNAs on GSK-3β in RCC. We selected nine candidate miRNAs from three databases that could potentially regulate GSK-3β. Among these, hsa-miR-4465 (miR-4465) was downregulated in RCC cell lines and renal cancer tissues. Furthermore, luciferase assays revealed that miR-4465 directly interacted with the 3' untranslated region of GSK-3β, and Western blot analysis showed that overexpression of miR-4465 significantly decreased GSK-3β protein expression. Functional assays showed that miR-4465 overexpression significantly suppressed cell invasion of A498 and Caki-1 cells; however, cell proliferation and migration were suppressed only in Caki-1 and A498 cells, respectively, with no effect on cell cycle and apoptosis. In conclusion, miR-4465 regulates GSK-3β expression but does not consistently affect RCC cell function as a single molecule. Further comprehensive investigation of regulatory networks is required in this field.

Novel synergistic treatment for depression: involvement of GSK3β-regulated AMPA receptors in the prefrontal cortex of mice.

Previous evidence has suggested a vital role of glycogen synthase kinase 3β-mediated α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors trafficking in depression. Considering the antidepressant effect of α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors activation in the prefrontal cortex, we hypothesized that glycogen synthase kinase 3β-induced alterations in α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors function in the prefrontal cortex participate in depression. Herein, we confirmed that the levels of phosphorylated glycogen synthase kinase 3β and GluA1, the latter being a subunit of α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors, were decreased in the prefrontal cortex of the chronic social defeat stress model mice presenting with depressive-like behaviors. We then found that a glycogen synthase kinase 3β (p.S9A) point mutation downregulated GluA1 and induced depressive-like behaviors in mice, whereas an agonist of α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors, PF-4778574 (2mg/kg) did not reversed the molecular changes. On the other hand, the antidepressant effect of PF-4778574 was dose dependent, and the single administration of PF-4778574 at a lower dose (0.5mg/kg) or of the glycogen synthase kinase 3β inhibitor SB216763 (5 and 10mg/kg) did not evoke an antidepressant effect. In contrast, co-treatment with PF-4778574 (0.5mg/kg) and SB216763 (10mg/kg) led to antidepressant effects similar to those of PF-4778574 (2mg/kg). Our results suggest that glycogen synthase kinase 3β-induced α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors dysfunction in the prefrontal cortex is one of the key mechanisms of depression, and the combination of a lower dose of PF-4778574 with SB216763 shows potential as a novel synergistic treatment for depression.

Computational Simulations Reveal the Synergistic Action of Phytochemicals of Morus alba to Exert Anti-Alzheimer Activity via Inhibition of Acetylcholinesterase and Glycogen Synthase Kinase-3β

Morus alba Linn. is a popular medium-sized tree with a potent neuroprotective activity such as anti-Alzheimer. To date, it is not known which phytocompound of M. alba exerts anti-Alzheimer activity. Therefore, the present study was aimed to identify the phytochemicals of M. alba responsible for the anti-Alzheimer activity by targeting proteins such as acetylcholinesterase (AChE) and glycogen synthase kinase-3 beta (GSK-3β) that are associated in the pathogenesis of Alzheimer disease (AD). Molecular docking, molecular dynamics (MD) simulations and MM-GBSA calculations were performed for the study. Phytocompounds with code 32, 41, 67, and 70 showed the best glide gscore against AchE and GSK-3β. All these phytocompounds interacted with multiple crucial amino acids present at the active binding site of the proteins through conventional hydrogen bonds and hydrophobic interactions. MD simulations of 100 ns revealed stable conformation for all protein-ligand complexes. Based on the computational findings, we conclude that M. alba’s phytoconstituents (32, 41 67, and 70) exerts anti-Alzheimer potential by inhibiting AChE and GSK-3β.

GSK3β Inhibition Ameliorates Atherosclerotic Calcification.

Endothelial-mesenchymal transition (EndMT) drives endothelium to contribute to atherosclerotic calcification. In a previous study, we showed that glycogen synthase kinase-3β (GSK3β) inhibition induced β-catenin and reduced mothers against DPP homolog 1 (SMAD1) in order to redirect osteoblast-like cells towards endothelial lineage, thereby reducing vascular calcification in Matrix Gla Protein (Mgp) deficiency and diabetic Ins2Akita/wt mice. Here, we report that GSK3β inhibition or endothelial-specific deletion of GSK3β reduces atherosclerotic calcification. We also find that alterations in β-catenin and SMAD1 induced by GSK3β inhibition in the aortas of Apoe-/- mice are similar to Mgp-/- mice. Together, our results suggest that GSK3β inhibition reduces vascular calcification in atherosclerotic lesions through a similar mechanism to that in Mgp-/- mice.

Mango ginger (Curcuma amada Roxb.) may alleviate the effect of high-fat diet/streptozotocin-induced diabetes by activation of the GSK-3β/Fyn/Nrf2 pathway.

Mango ginger (MG) exhibits antioxidant, anti-inflammatory, and antihyperglycemic effects; however, the exact mechanism of action of MG extract in relation to its antidiabetic properties remains unclear. To investigate the potential antidiabetic effect of MG extract, we used a high-fat diet (HFD)/low-dose streptozotocin (STZ)-induced type 2 diabetic rat model. A total of 28 male Wistar rats were randomly divided into four groups: (i) Control, (ii) MG (50 mg/kg/day of MG extract), (iii) HFD + STZ (40 mg/kg i.p.), and (iv) HFD + STZ + MG. Following a 12-week administration of MG extract, significant reductions were observed in serum glucose, insulin, free fatty acid, cholesterol, and triglyceride levels in diabetic rats (p < .0001 for all). MG extract supplementation led to an increase in the total antioxidant capacity of the serum and a decrease in malondialdehyde (MDA) levels in both the serum and liver (p < .0001). Furthermore, hepatocellular fat accumulation was partially attenuated in the HFD + STZ + MG group. Notably, MG extract inhibited glycogen synthase kinase-3β (GSK-3β) in the liver (p < .01) and downregulated Fyn expression, resulting in elevated nuclear factor erythroid 2-related factor 2 (Nrf2) activity in the HFD + STZ + MG group compared to the HFD + STZ group (p < .05). The increased activity of Nrf2 in the HFD + STZ + MG group likely promoted the upregulation of heme oxygenase 1 (HO-1) in the liver (p < .0001). In conclusion, MG extract may exert antidiabetic effects by augmenting the antioxidant defense system through the regulation of GSK-3β/Fyn/Nrf2 in a rat model of type 2 diabetes.

Synthesis, Characterization and Biological Evaluation of Glycogen Synthase Kinase-3β Inhibitors as Antidiabetic Agents

Diabetes is a substantially growing unhealthiest in the globe that causes severe morbidity and mortality. Currently a day’s there’s an imperative for the invention, development and improvement of novel antidiabetic drug molecules. A chain of compound (5-imidazol-2-yl-4-phenylpyrimidin-2-yl) [2-(2- pyridyl amino) ethyl] amine were developed and synthesized by using suitable chemical synthetic techniques. These recently synthesized derivatives were characterized with the help of modern analytical techniques. This compound was evaluated for glycogen synthase kinase-3 enzyme inhibitor activities for managing of polygenic disease. GSK-3 inhibitors may advance and emerge as a novel strategy for the management of diabetes.

Apoptosis of Kinetin Riboside in Colorectal Cancer Cells Occurs by Promoting β-Catenin Degradation.

Kinetin riboside is a naturally produced cytokinin that displays strong antiproliferative activity in various human cancer cells. However, the mechanism of chemoprevention in colorectal cancer cells has not been elucidated. We used a cell-based reporter system to identify kinetin riboside as an antagonist of the Wnt/β-catenin pathway, which is aberrantly upregulated in colorectal cancer. Kinetin riboside suppressed β-catenin response transcription (CRT) by accelerating the degradation of intracellular β-catenin via a proteasomal degradation pathway. Pharmacological inhibition of glycogen synthase kinase-3β did not affect CRT downregulation. Kinetin riboside decreased the intracellular β-catenin levels in colorectal cancer cells with mutations in adenomatous polyposis coli (APC) and β-catenin. Consistently, kinetin riboside repressed expression of c-Myc and cyclin D1, β-catenin/T-cell factor (TCF)-dependent genes, and inhibited the proliferation of colorectal cancer cells. In addition, kinetin riboside stimulated apoptosis, as measured by an increase in annexin V-FITC-stained cells. These findings suggest that kinetin riboside exerts its anti-cancer activity by promoting β-catenin degradation and has significant potential as a chemopreventive agent for colorectal cancer cells.

Elraglusib (formerly 9-ING-41) possesses potent anti-lymphoma properties which cannot be attributed to GSK3 inhibition

Elraglusib (formerly 9-ING-41) is an ATP-competitive inhibitor of glycogen synthase kinase-3β (GSK3β) undergoing clinical trials for the treatment of various cancers including non-Hodgkin lymphoma (NHL). The drug reduces proliferation of several NHL cell lines and has efficacy in xenograft models of the disease. To confirm the importance of its action on GSK3β, we treated 3 lymphoma cell lines with selective, structurally distinct GSK3 inhibitors: CT99021, SB216763, LY2090314, tideglusib, and elraglusib. Stabilization of β-catenin and reduced phosphorylation of CRMP2, two validated targets of GSK3, were used as functional read-outs for GSK3 inhibition. CT99021, SB216763, and LY2090314 failed to reduce proliferation or viability in any cell line at concentrations that stabilized β-catenin and reduced CRMP2 phosphorylation. There was partial reduction of CRMP2 phosphorylation but no significant effect on β-catenin at cytotoxic doses of elraglusib. There was no indication of GSK3 inhibition at doses of tideglusib that affected cell viability and apoptosis. Cell-free kinase screening confirmed several other targets of elraglusib, distinct from the GSK3 inhibitors with no anti-lymphoma actions, including PIM kinases and MST2. These data question GSK3 as the target of elraglusib in lymphoma, and hence the utility of GSK3 expression as a ‘stand-alone’, therapeutic biomarker in NHL.7CsbmemxsJeEQuU9mL3kzMVideo

Synthesis and biological evaluation of thieno[3,2-c]pyrazol-3-amine derivatives as potent glycogen synthase kinase 3β inhibitors for Alzheimer’s disease

Glycogen synthase kinase 3β (GSK-3β) is a potential target for anti-Alzheimer’s disease (AD) drug development. In this study, a series of novel thieno[3,2-c]pyrazol-3-amine derivatives was synthesized and evaluated as potential GSK-3β inhibitors by structure-based drug design. The thieno[3,2-c]pyrazol-3-amine derivative 54 with a 4-methylpyrazole moiety which interacted with Arg141 by π-cation interaction was identified as a potent GSK-3β inhibitor with an IC50 of 3.4 nM and an acceptable kinase selectivity profile. In the rat primary cortical neurons, compound 54 showed neuroprotective effects on Aβ-induced neurotoxicity. Western blot analysis indicated that 54 inhibited GSK-3β by up-regulating the expression of phosphorylated GSK-3β at Ser9 and down-regulating the expression of phosphorylated GSK-3β at Tyr216. Meanwhile, 54 decreased tau phosphorylation at Ser396 in a dose-dependent way. In astrocytes and microglia cells, 54 inhibited the expression of inducible nitric oxide synthase (iNOS), indicating that 54 showed an anti-neuroinflammatory effect. In the AlCl3-induced zebrafish AD model, 54 significantly ameliorated the AlCl3-induced dyskinesia, demonstrating its anti-AD activity in vivo.

Functionalized Exosomes Promote Neural Differentiation of P19 Cells by Activating the Wnt Signaling Pathway.

Global population growth and population aging continue to accelerate, and lead to a further increase in the risk of neurological diseases worldwide. Extracellular vesicles secreted by mesenchymal stem cells carry many proteins, lipids, and genetic material that mediate cell-to-cell communication and improve therapeutic outcomes for neurological disorders. Stem cells from human exfoliated deciduous teeth are considered a suitable cell source for tissue regeneration, which exerts therapeutic effects via the secretion of exosomes. This study was performed to assess the effect of functionalized exosomes on neural differentiation of embryonic carcinoma cell line P19. We stimulated stem cells from human exfoliated deciduous teeth with the glycogen synthase kinase-3β inhibitor TWS119 and then extracted its exosomes. P19 cells were induced to differentiate using functionalized exosomes, and the biological functions and involved signaling pathways of differentially expressed genes were analyzed by RNA-sequencing. Immunofluorescence techniques detected neuronal specific markers. It was found that TWS119 activated the Wnt signaling pathway in stem cells from human exfoliated deciduous teeth. RNA-sequencing showed that upregulated differentially expressed genes in the functionalized exosome-treated group were responsible for cell differentiation, neurofilament, and structural constituent of the synapse. Kyoto Encyclopedia of Genes and Genomes enrichment analysis revealed that the functionalized exosome-treated group activated the Wnt signaling pathway. Immunofluorescence showed that functionalized exosomes induced neurite outgrowth in P19 cells. Our results demonstrated that functionalized exosomes promoted neural differentiation of P19 cells by activating the Wnt signaling pathway.

Lithium chloride attenuates sepsis-induced acute respiratory distress syndrome via decreasing the expressions of TNF-α, IL-6, CXCL1, and pulmonary edema

Abstract Sepsis is a life-threatening illness caused by an excessive host response to infection. Acute respiratory distress syndrome (ARDS), a serious complication of sepsis, is characterized by pulmonary edema and alveolar barrier injury. However, there is no effective drug proven to reduce the mortality of ARDS yet. Lithium chloride (LiCl) is an inhibitor of glycogen synthase kinase-3β (GSK-3β). Recent studies reveal that GSK-3β inhibition exhibits anti-inflammatory effects in sepsis thereby ameliorating lipopolysaccharide (LPS)-induced acute lung injury. Given that GSK-3β inhibition has been investigated for various diseases, whether LiCl exerting a novel therapeutic effect for treating endotoxemia-induced ARDS remains unclear. Here, we investigated the effects of LiCl on endotoxemia-induced lung inflammation, respiratory failure, and the pulmonary barrier integrity of respiratory membrane. Mice intraperitoneally injected with LPS could induce a significant lymphopenia, lung inflammation with amount immune cells infiltration, and high mobility group box 1 (HMGB1) expression in lung supernatants. LiCl treatment ameliorated lung inflammation and lung function decline in LPS mice. In addition, LPS-induced TNF-α, IL-6, as well as CXCL1 were significantly lessened after LiCl treatment. The measurements of E-cadherin/VE-cadherin expression in lung tissues and IgM in bronchoalveolar lavage fluids indicated that LiCl could partly ameliorate LPS-mediated loss of pulmonary barrier integrity. Accordingly, LiCl effectively attenuated sepsis-caused lung inflammation, lung function decline, and alveolar barrier injury, which suggested that LiCl might act as a potential repurposing drug for treating ARDS in future.

Inhibition of GSK-3β Enhances Osteoblast Differentiation of Human Mesenchymal Stem Cells through Wnt Signalling Overexpressing Runx2.

Small-molecule-inhibitor-based bone differentiation has been recently exploited as a novel approach to regulating osteogenesis-related signaling pathways. In this study, we identified 1-Azakenpaullone, a highly selective inhibitor of glycogen synthase kinase-3β (GSK-3β), as a powerful inducer of osteoblastic differentiation and mineralization of human mesenchymal stem cells (MSCs). GSK-3β is a serine-threonine protein kinase that plays a major role in different disease development. GSK-3β is a key regulator of Runx2 activity in osteoblastic formation. We evaluated alkaline phosphatase activity and staining assays to assess osteoblast differentiation and Alizarin Red staining to assess the mineralization of cultured human MSCs. Gene expression profiling was assessed using an Agilent microarray platform, and bioinformatics were performed using Ingenuity Pathway Analysis software. Human MSCs treated with 1-Azakenpaullone showed higher ALP activity, increased in vitro mineralized matrix formation, and the upregulation of osteoblast-specific marker gene expression. Global gene expression profiling of 1-Azakenpaullone-treated human MSCs identified 1750 upregulated and 2171 downregulated mRNA transcripts compared to control cells. It also suggested possible changes in various signaling pathways, including Wnt, TGFβ, and Hedgehog. Further bioinformatics analysis employing Ingenuity Pathway Analysis recognized significant enrichment in the 1-Azakenpaullone-treated cells of genetic networks involved in CAMP, PI3K (Complex), P38 MAPK, and HIF1A signaling and functional categories associated with connective tissue development. Our results suggest that 1-Azakenpaullone significantly induced the osteoblastic differentiation and mineralization of human MSCs mediated by the activation of Wnt signaling and the nuclear accumulation of β-catenin, leading to the upregulation of Runx2, a key transcription factor that ultimately promotes the expression of osteoblast-specific genes. Thus, 1-Azakenpaullone could be used as an osteo-promotor factor in bone tissue engineering.

AR-A014418, a glycogen synthase kinase-3β inhibitor, mitigates lipopolysaccharide-induced inflammation in rat dental pulp stem cells via NLR family pyrin domain containing 3 inflammasome impairment

Background/purposeCell pyroptosis and gingival inflammation have been implicated in periodontitis progression. Our previous study revealed that AR-A014418, a pharmacological inhibitor of glycogen synthase kinase-3β (GSK-3β), can enhance the migratory and osteogenic differentiation abilities of rat dental pulp stem cells (rDPSCs). The present study aimed to explore the effect of AR on the inflammation of rDPSCs. Materials and methodsThe primary rDPSCs were isolated and identified by flow cytometry, as well as Oil red O and Alizarin Red S staining. The rDPSCs were cultured and exposed to lipopolysaccharide (LPS) before treating them with different concentrations of AR-A014418. The cell viability was detected using the CCK-8 assay. The generation and secretion of pro-inflammatory cytokines (IL-18, TNF-α, L-1β, and IL-6) were examined by qPCR and ELISA, respectively. To investigate the activation of the NLRP3 inflammasome, the expression levels of pro-caspase 1, cleaved caspase 1, as well as NLRP3 were analyzed by western blotting and immunofluorescence, respectively. ResultsIn the rDPSCs, LPS prohibited cell viability and enhanced the generation and secretion of pro-inflammatory cytokines. LPS upregulated NLRP3 and cleaved caspase-1 protein levels and promoted ASC speck formation in the rDPSCs. AR-A014418 administration effectively blocked the LPS-induced inflammation of the rDPSCs in a dose-dependent way. Mechanistically, AR-A014418 significantly restrained the up-regulation of NLRP3 and cleaved caspase-1 in LPS-treated rDPSCs. ConclusionCollectively, our findings suggest that AR-A014418 significantly mitigates LPS-induced inflammation of rDPSCs by blocking the activation of the NLRP3 inflammasome.

GSK3β Inhibition Reduced Vascular Calcification in Ins2Akita/+ Mice.

Endothelial-mesenchymal transition (EndMT) drives the endothelium to contribute to vascular calcification in diabetes mellitus. In our previous study, we showed that glycogen synthase kinase-3β (GSK3β) inhibition induces β-catenin and reduces mothers against DPP homolog 1 (SMAD1) to direct osteoblast-like cells toward endothelial lineage, thereby reducing vascular calcification in Matrix Gla Protein (Mgp) deficiency. Here, we report that GSK3β inhibition reduces vascular calcification in diabetic Ins2Akita/wt mice. Cell lineage tracing reveals that GSK3β inhibition redirects endothelial cell (EC)-derived osteoblast-like cells back to endothelial lineage in the diabetic endothelium of Ins2Akita/wt mice. We also find that the alterations in β-catenin and SMAD1 by GSK3β inhibition in the aortic endothelium of diabetic Ins2Akita/wt mice are similar to Mgp-/- mice. Together, our results suggest that GSK3β inhibition reduces vascular calcification in diabetic arteries through a similar mechanism to that in Mgp-/- mice.

Detection of natural compounds by virtual screening, molecular docking and dynamics studies and evaluation of their effects on tau level in vitro Alzheimer’s model

In Alzheimer’s disease (AD), neurofibrillary tangles are composed of hyperphosphorylated tau protein, and tau hyperphosphorylation reduces microtubule binding. Many protein kinases are thought to be involved in tau hyperphosphorylation. Based on the fact that tau hyperphosphorylation can be prevented by inhibition of glycogen synthase kinase-3β (GSK-3β), which is one of the tau kinases, the effectiveness of potential GSK-3β inhibitors determined by virtual screening, molecular docking, and dynamics simulations studies on Alzheimer’s pathology has been examined and its role in neurodegeneration has been investigated by studies. Neomangiferin was determined as the most effective molecule according to the results of studies with potential compounds determined by virtual screening and molecular docking to be GSK-3β inhibitors in the in vitro Alzheimer’s model created by neuronal differentiation studies. Neomangiferin has been shown to have a protective role in induced neurodegeneration by the MTT method and Real Time Cell Analysis. It has been determined that Neomangiferin inhibits GSK-3β and reduces the level of phosphorylated tau. In summary, our findings suggested Neomangiferin can be a therapeutic candidate for AD treatment. Communicated by Ramaswamy H. Sarma

Adhesive Hydrogel Patch-Mediated Combination Drug Therapy Induces Regenerative Wound Healing through Reconstruction of Regenerative Microenvironment.

Regenerative wound healing involves the scarless wound healing as observed in fetal skin. Multiple features of regenerative wound healing have been well studied; however, the practical application of pro-regenerative materials to recapitulate the regenerative wound healing in adult skins has not yet been achieved. In this study, the authors identified that their novel pro-regenerative material, pyrogallol-functionalized hyaluronic acid (HA-PG) patches in combination with protein transduction domain-fused Dishevelled (Dvl)-binding motif (PTD-DBM), a peptide inhibiting the CXXC-type zinc finger protein 5 (CXXC5)-Dvl interaction, promoted regenerative wound healing in mice. The HA-PG patches loaded with this competitor peptide and valproic acid (VPA), a glycogen synthase kinase 3β (GSK3β) inhibitor, significantly inhibited scar formation during wound healing. The HA-PG patches with PTD-DBM and/or VPA inhibit the expression of differentiated cell markers such as α-smooth muscle actin (α-SMA) while inducing the expression of stem cell markers such as CD105 and Nestin. Moreover, Collagen III, an important factor for regenerative healing, is critically induced by the HA-PG patches with PTD-DBM and/or VPA, as also seen in VPA-treated Cxxc5-/- mouse fibroblasts. Overall, these findings suggest that the novel regeneration-promoting material can be utilized as a potential therapeutic agent to promote both wound healing and scar attenuation.

Design, Structure-Activity Relationships, and In Vivo Evaluation of Potent and Brain-Penetrant Imidazo[1,2-b]pyridazines as Glycogen Synthase Kinase-3β (GSK-3β) Inhibitors.

Glycogen synthase kinase-3 (GSK-3) is a serine/threonine kinase that regulates numerous cellular processes, including metabolism, proliferation, and cell survival. Due to its multifaceted role, GSK-3 has been implicated in a variety of diseases, including Alzheimer's disease, type 2 diabetes, cancer, and mood disorders. GSK-3β has been linked to the formation of the neurofibrillary tangles associated with Alzheimer's disease that arise from the hyperphosphorylation of tau protein. The design and synthesis of a series of imidazo[1,2-b]pyridazine derivatives that were evaluated as GSK-3β inhibitors are described herein. Structure-activity relationship studies led to the identification of potent GSK-3β inhibitors. In vivo studies with 47 in a triple-transgenic mouse Alzheimer's disease model showed that this compound is a brain-penetrant, orally bioavailable GSK-3β inhibitor that significantly lowered levels of phosphorylated tau.

The Role of Fucoxanthin as a Potent Nrf2 Activator via Akt/GSK-3β/Fyn Axis against Amyloid-β Peptide-Induced Oxidative Damage

Increasing evidence is suggesting that amyloid-β peptide (Aβ), a characteristic of Alzheimer's disease (AD), induces oxidative stress and mitochondrial dysfunction, leading to neuronal death. This study aimed to demonstrate the antioxidant and anti-apoptotic effects of fucoxanthin, a major marine carotenoid found in brown algae, against neuronal injury caused by Aβ. Non-toxic dose range of fucoxanthin (0.1-5 µM) were selected for the neuroprotective study against Aβ25-35. The PC12 cells were pretreated with different concentrations of fucoxanthin for 1 h before being exposed to 10 µM Aβ25-35 for another 24 h. The present results showed that fucoxanthin inhibited Aβ25-35-induced cell death by recovering cell cycle arrest and decreasing intracellular reactive oxygen species (ROS) level. The compound enhanced mitochondrial recovery and regulated apoptosis related proteins including B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (Bax) from Aβ25-35-induced oxidative stress. Concomitantly, fucoxanthin increased the expression of nuclear factor E2-related factor 2 (Nrf2) and its downstream phase II detoxifying enzymes including NADPH: quinone oxidoreductase-1 (NQO-1), glutamate cysteine ligase modifier subunit (GCLm), and thioredoxin reductase 1 (TrxR1), whereas it decreased the expression of cytoplasmic Kelch-like ECH-associated protein 1 (Keap1). Moreover, pretreatment of fucoxanthin reduced Fyn phosphorylation via protein kinase B (Akt)-mediated inhibition of glycogen synthase kinase-3β (GSK-3β), which increased the nuclear localization of Nrf2, suggesting that the compound enhanced Nrf2 expression by the activation of upstream kinase as well as the dissociation of the Nrf2-Keap1 complex. Further validation with a specific phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 demonstrated that the fucoxanthin-mediated Nrf2 antioxidant defense system was directly associated with the Akt/GSK-3β/Fyn signaling pathway. In silico simulation revealed that the oxygen groups of fucoxanthin participated in potent interactions with target markers in the Nrf2 signaling pathway, which may affect the biological activity of target markers. Taken together, the present results demonstrated that the preventive role of fucoxanthin on Aβ-stimulated oxidative injury and apoptosis via Akt/GSK-3β/Fyn signaling pathway. This study would provide a useful approach for potential intervention for AD prevention.