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Supplementary Figure 2 from Negative Regulation of p53 by the Long Isoform of ErbB3 Binding Protein Ebp1 in Brain Tumors

Supplementary Figure 3 from Negative Regulation of p53 by the Long Isoform of ErbB3 Binding Protein Ebp1 in Brain Tumors

Supplementary Figure 6 from Negative Regulation of p53 by the Long Isoform of ErbB3 Binding Protein Ebp1 in Brain Tumors

Supplementary Figure 4 from Negative Regulation of p53 by the Long Isoform of ErbB3 Binding Protein Ebp1 in Brain Tumors

Supplementary Methods, Figure Legends 1-6 from Negative Regulation of p53 by the Long Isoform of ErbB3 Binding Protein Ebp1 in Brain Tumors

Supplementary Figure 5 from Negative Regulation of p53 by the Long Isoform of ErbB3 Binding Protein Ebp1 in Brain Tumors

The ErbB3 binding protein Ebp1 has been implicated in a number of human cancers. Ebp1 includes 2 isoforms, p48 and p42, that exhibit different cellular activities. Here we show that the larger p48 isoform is transforming and that it promotes cell growth, clonogenicity, and invasion in human glioblastoma (GBM). P48 overexpression in GBM cells facilitated tumorigenesis and enhanced tumor growth in mouse xenograft models. Human GBM tissues displayed elevated levels of p48 compared with surrounding normal tissues or low-grade tumors. Notably, p48 levels were inversely correlated with poor prognosis in GBM patients. We determined that p48 binds to the p53 E3 ligase HDM2, enhancing HDM2-p53 association and thereby promoting p53 polyubiquitination and degradation to reduce steady-state p53 levels and activity. Together, our findings suggest that p48 functions as an oncogene by promoting glioma tumorigenicity via interactions with HDM2 that contribute to p53 downregulation. Cancer Res; 70(23); 9730–41. ©2010 AACR.

Supplementary Figure 1 from Negative Regulation of p53 by the Long Isoform of ErbB3 Binding Protein Ebp1 in Brain Tumors

Cerebellar deficits with Purkinje cell (PCs) loss are observed in several neurologic disorders. However, the underlying mechanisms as to how the cerebellum is affected during development remain unclear. Here we demonstrated that specific inactivation of murine Ebp1 in the central nervous system causes a profound neuropathology characterized by reduced cerebellar volume and PCs loss with abnormal dendritic development, leading to phenotypes including motor defects and schizophrenia (SZ)-like behaviors. Loss of Ebp1 leads to untimely gene expression of Fbxw7, an E3 ubiquitin ligase, resulting in aberrant protein degradation of PTF1A, thereby eliciting cerebellar defects. Reinstatement of Ebp1, but not the Ebp1-E183Ter mutant found in SZ patients, reconstituted cerebellar architecture with increased PCs numbers and improved behavioral phenotypes. Thus, our findings indicate a crucial role for EBP1 in cerebellar development, and define a molecular basis for the cerebellar contribution to neurologic disorders such as SZ.

Butea monosperma (Lam.) Taub. has been applied to treat inflammatory, metabolic, and infectious diseases. However, the antiobesity effects of B. monosperma (Lam.) Taub. flower (BMF) and the underlying mechanisms have not been determined. In this study, we analyzed the various extraction procedures, investigated the antiobesity effects, and identified the main chemical constituents of BMF. The BMF was subjected to acid hydrolysis in 5% H2SO4 in methanol at 50°C for 48 h and partitioned with ethyl acetate. The acid-hydrolyzed BMF ethyl acetate extracts (BMFE) strongly induced the expression of uncoupling protein 1 (Ucp1) and other thermogenic genes in C3H10T1/2 adipocytes. Daily oral administration of 70 mg/kg BMFE (BMFE70) to mice with diet-induced obesity resulted in less body weight gain, increased glucose tolerance, higher rectal temperature, and increased oxygen consumption. Qualitative and quantitative analyses along with treatments in Akt1 knockout mouse embryonic fibroblasts indicate that butein is a major active ingredient of BMFE, which stimulates Ucp1 gene expression. These data show the effects of butein-containing B. monosperma flower extract on thermogenesis and energy expenditure, further suggesting the potential role of BMFE as a functional ingredient in obesity and related metabolic diseases.

Microglial activation is closely associated with neuroinflammatory pathologies. The nucleotide-binding and oligomerization domain-like receptor containing a pyrin domain 3 (NLRP3) inflammasomes are highly organized intracellular sensors of neuronal alarm signaling. NLRP3 inflammasomes activate nuclear factor kappa-B (NF-κB) and reactive oxygen species (ROS), which induce inflammatory responses. Moreover, NLRP3 dysfunction is a common feature of chronic inflammatory diseases. The present study investigated the effect of a novel thiazol derivative, N-cyclooctyl-5-methylthiazol-2-amine hydrobromide (KHG26700), on inflammatory responses in lipopolysaccharide (LPS)-treated BV-2 microglial cells. KHG26700 significantly attenuated the expression of several pro-inflammatory cytokines, including tumor necrosis factor-α, interleukin-1β, and interleukin-6, in these cells, as well as the LPS-induced increases in NLRP3, NF-κB, and phospho-IkBα levels. KHG26700 also suppressed the LPS-induced increases in protein levels of autophagy protein 5 (ATG5), microtubule- associated protein 1 light chain 3 (LC3), and beclin-1, as well as downregulating the LPS-enhanced levels of ROS, lipid peroxidation, and nitric oxide. These results suggest that the anti-inflammatory effects of KHG26700 may be due, at least in part, to the regulation of the NLRP3-mediated signaling pathway during microglial activation. [BMB Reports 2021; 54(11): 557-562].

Acetylation of α-tubulin lysine 40 (αK40) contributes to microtubule (MT) stability and is essential for neuronal development and function, whereas excessive αK40 deacetylation is observed in neurodegenerative disorders including Alzheimer’s disease (AD). Here we identified inhibitor of DNA binding 2 (Id2) as a novel MT-binding partner that interacts with α-tubulin and enhances αK40 acetylation, leading to MT polymerization in the neurons. Commensurate with our finding that the low levels of Id2 expression along with a reduced αK40 acetylation in the postmortem human AD patient and 5X-FAD, AD model mice brain, Id2 upregulation in the hippocampus of 5X-FAD, which exhibit high levels of Sirt2 expression, increased αK40 acetylation and reconstitutes axon growth. Hence our study suggests that Id2 is critical for maintaining MT stability during neural development and the potential of Id2 to counteract pathogenic Sirt2 activity in AD.

Chemoresistance is a major cause of treatment failure in many cancers. However, the life cycle of cancer cells as they respond to and survive environmental and therapeutic stress is understudied. In this study, we utilized a microfluidic device to induce the development of doxorubicin-resistant (DOXR) cells from triple negative breast cancer (TNBC) cells within 11 days by generating gradients of DOX and medium.In vivochemoresistant xenograft models, an unbiased genome-wide transcriptome analysis, and a patient data/tissue analysis all showed that chemoresistance arose from failed epigenetic control of the nuclear protein-1 (NUPR1)/histone deacetylase 11 (HDAC11) axis, and high Nupr1 expression correlated with poor clinical outcomes. These results suggest that the chip can rapidly induce resistant cells that increase tumor heterogeneity and chemoresistance, highlighting the need for further studies on the epigenetic control of the NUPR1/HDAC11 axis in TNBC.

ErbB3-binding protein 1 (EBP1) is a multifunctional protein associated with neural development. Loss of Ebp1 leads to upregulation of the gene silencing unit suppressor of variegation 3-9 homolog 1 (Suv39H1)/DNA (cytosine 5)-methyltransferase (DNMT1). EBP1 directly binds to the promoter region of DNMT1, repressing DNA methylation, and hence, promoting neural development. In the current study, we showed that EBP1 suppresses histone methyltransferase activity of Suv39H1 by promoting ubiquitin-proteasome system (UPS)-dependent degradation of Suv39H1. In addition, we showed that EBP1 directly interacts with Suv39H1, and this interaction is required for recruiting the E3 ligase MDM2 for Suv39H1 degradation. Thus, our findings suggest that EBP1 regulates UPS-dependent degradation of Suv39H1 to govern proper heterochromatin assembly during neural development. [BMB Reports 2021; 54(8): 413-418].

The roles of the two isoforms of ErbB3-binding protein 1 (Ebp1) in cellular function and its regulation in disease and development is a stimulating area in current fields of biology, such as neuroscience, cancer biology, and structural biology. Over the last two decades, a growing body of studies suggests have suggested different functions for the EBP1 isoforms in various cancers, along with their specific binding partners in the ubiquitin-proteasome system. Owing to the specific cellular context or spatial/temporal expression of the EBP1 isoforms, either transcriptional repression or the activation function of EBP1 has been proposed, and epigenetic regulation by p48 EBP1 has also been observed during in the embryo development, including in brain development and neurologic disorders, such as schizophrenia, in using an Ebp1 knockout mouse model. Here, we review recent findings that have shaped our current understanding of the emerging function of EBP1 isoforms in cellular events and gene expression, from development to disease., Cell biology: deciphering differences between “sibling” proteins A pair of proteins that originate from a common gene exert strikingly different effects on embryonic development as well as tumor growth and progression. RNA transcripts generated from the PA2G4 gene can undergo enzymatic processing to yield two different protein products, p42 EB1 and p48 EB1. These proteins differ by the presence or absence of 54 amino acids at one end, and Jee-Yin Ahn at the Sungkyunkwan University School of Medicine, Suwon, South Korea, and colleagues have reviewed current insights into the functional consequences of this difference. The two proteins bind to distinct sets of molecular partners. The p48 form appears to regulate a host of genes involved in brain development, but also appears to drive cancerous growth in various tumors. In contrast, p42 is scarcer during development, and appears to inhibit tumor formation.

Although Krüppel-associated box domain-containing zinc-finger proteins (K-ZNFs) may be associated with sophisticated gene regulation in higher organisms, the physiological functions of most K-ZNFs remain unknown. The Zfp212 protein was highly conserved in mammals and abundant in the brain; it was mainly expressed in the cerebellum (Cb). Zfp212 (mouse homolog of human ZNF212) knockout (Zfp212-KO) mice showed a reduction in survival rate compared to wild-type mice after 20 months of age. GABAergic Purkinje cell degeneration in the Cb and aberrant locomotion were observed in adult Zfp212-KO mice. To identify genes related to the ataxia-like phenotype of Zfp212-KO mice, 39 ataxia-associated genes in the Cb were monitored. Substantial alterations in the expression of ataxin 10, protein phosphatase 2 regulatory subunit beta, protein kinase C gamma, and phospholipase D3 (Pld3) were observed. Among them, Pld3 alone was tightly regulated by Flag-tagged ZNF212 overexpression or Zfp212 knockdown in the HT22 cell line. The Cyclic Amplification and Selection of Targets assay identified the TATTTC sequence as a recognition motif of ZNF212, and these motifs occurred in both human and mouse PLD3 gene promoters. Adeno-associated virus-mediated introduction of human ZNF212 into the Cb of 3-week-old Zfp212-KO mice prevented Purkinje cell death and motor behavioral deficits. We confirmed the reduction of Zfp212 and Pld3 in the Cb of an alcohol-induced cerebellar degeneration mouse model, suggesting that the ZNF212–PLD3 relationship is important for Purkinje cell survival.

Akt signaling is an important regulator of neural development, but the distinctive function of Akt isoforms in brain development presents a challenge. Here we show Siah1 as an ubiquitin ligase that preferentially interacts with Akt3 and facilitates ubiquitination and degradation of Akt3. Akt3 is enriched in the axonal shaft and branches but not growth cone tips, where Siah1 is prominently present. Depletion of Siah1 enhanced Akt3 levels in the soma and axonal tips, eliciting multiple branching. Brain-specific somatic mutation in Akt3-E17K escapes from Siah1-mediated degradation and causes improper neural development with dysmorphic neurons. Remarkably, coexpression of Siah1 with Akt3-WT restricted disorganization of neural development is caused by Akt3 overexpression, whereas forced expression of Siah1 with the Akt3-E17K mutant fails to cope with malformation of neural development. These findings demonstrate that Siah1 limits Akt3 turnover during brain development and that this event is essential for normal organization of the neural network.

Dysregulation of epigenetic machinery can cause a variety of neurological disorders associated with cognitive abnormalities. In the hippocampus of postmortem Schizophrenia (SZ) patients, the most notable finding is the deregulation of GAD67 along with differential regulation of epigenetic factors associated with glutamate decarboxylase 67 (GAD67) expression. As we previously reported, ErbB3-binding protein 1 (EBP1) is a potent epigenetic regulator. EBP1 can induce repression of Dnmt1, a well-studied transcriptional repressor of GAD67. In this study, we investigated whether EBP1 contributes to the regulation of GAD67 expression in the hippocampus, controlling epigenetic machinery. In accordance with SZ-like behaviors in Ebp1(+/&minus, ) mice, heterozygous deletion of EBP1 led to a dramatic reduction of GAD67 expression, reflecting an abnormally high level of Dnmt1. Moreover, we found that EBP1 binds to the promoter region of HDAC1, which leads to histone deacetylation of GAD67, and suppresses histone deacetylase 1 (HDAC1) expression, inversely mirroring an unusually high level of HDAC1 in Ebp1(+/&minus, ) mice. However, EBP1 mutant (p.Glu 183 Ter) found in SZ patients did not elevate the expression of GAD67, failing to suppress Dnmt1 and/or HDAC1 expression. Therefore, this data supports the hypothesis that a reduced amount of EBP1 may contribute to an etiology of SZ due to a loss of transcriptional inhibition of epigenetic repressors, leading to a decreased expression of GAD67.

Stimulation of white adipose tissue (WAT) browning is considered as a potential approach to treat obesity and metabolic diseases. Our previous studies have shown that phytochemical butein can stimulate WAT browning through induction of Prdm4 in adipocytes. Here, we investigated the effects of butein on diet-induced obesity and its underlying molecular mechanism. Treatment with butein prevented weight gains and improved metabolic profiles in diet-induced obese mice. Butein treatment groups also displayed higher body temperature, increased energy expenditure, and enhanced expression of thermogenic genes in adipose tissue. Butein also suppressed body weight gains and improved glucose and insulin tolerance in mice housed at thermoneutrality (30 °C). These effects were associated with adipose-selective induction of Prdm4, suggesting the role of Prdm4 in butein-mediated anti-obese effects. To directly assess the in vivo role of Prdm4, we generated aP2-Prdm4 transgenic mouse lines overexpressing Prdm4 in adipose tissues. Adipose-specific transgenic expression of Prdm4 recapitulated the butein’s actions in stimulating energy expenditure, cold tolerance, and thermogenic gene expression, resulting in prevention of obesity and improvement of metabolism. Mechanistically, direct inhibition of PI3Kα activity followed by selective suppression of its downstream Akt1 mirrored butein’s effect on Ucp1 expression and oxygen consumption. In addition, effects of butein were completely abolished in Akt1 KO mouse embryonic fibroblasts. Together, these studies demonstrate the role of butein in obesity and metabolic diseases, further highlighting that adipose PI3Kα–Akt1–Prdm4 axis is a regulator of energy expenditure.

We aimed to assess the anti-inflammatory and antioxidative properties of KHG26792, a novel azetidine derivative, in amyloid β (Aβ)-treated primary microglial cells. KHG26792 attenuated the Aβ-induced production of inflammatory mediators such as IL-6, IL-1β, TNF-α, and nitric oxide. The levels of protein oxidation, lipid peroxidation, ROS, and NADHP oxidase enhanced by Aβ were also downregulated by KHG26792 treatment. The effects of KHG26792 against the Aβ-induced increases in inflammatory cytokine levels and oxidative stress were achieved by increasing the phosphorylation of Akt/ GSK-3β signaling and by decreasing the Aβ-induced translocation of NF-κB. Our results provide novel insights into the use of KHG26792 as a potential agent against Aβ toxicity, including its role in the reduction of inflammation and oxidative stress. Nevertheless, further investigations of cellular signaling are required to clarify the in vivo effects of KHG26792 against Aβ-induced toxicity. [BMB Reports 2017; 50(12): 634-639].

Mesenchymal stem cell (MSC) transplantation represents the next breakthrough in the treatment of currently intractable and devastating neonatal disorders with complex multifactorial etiologies, including bronchopulmonary dysplasia, hypoxic ischemic encephalopathy, and intraventricular hemorrhage. Absent engraftment and direct differentiation of transplanted MSCs, and the "hit-and-run" therapeutic effects of these MSCs suggest that their pleiotropic protection might be attributable to paracrine activity via the secretion of various biologic factors rather than to regenerative activity. The transplanted MSCs, therefore, exert their therapeutic effects not by acting as "stem cells," but rather by acting as "paracrine factors factory." The MSCs sense the microenvironment of the injury site and secrete various paracrine factors that serve several reparative functions, including antiapoptotic, anti-inflammatory, antioxidative, antifibrotic, and/or antibacterial effects in response to environmental cues to enhance regeneration of the damaged tissue. Therefore, the therapeutic efficacy of MSCs might be dependent on their paracrine potency. In this review, we focus on recent investigations that elucidate the specifically regulated paracrine mechanisms of MSCs by injury type and discuss potential strategies to enhance paracrine potency, and thus therapeutic efficacy, of transplanted MSCs, including determining the appropriate source and preconditioning strategy for MSCs and the route and timing of their administration.

P48 Ebp1 is expressed in rapidly proliferating cells such as cancer cells and accelerates cell growth and survival. However, its expression pattern and role in central nervous system development have not been studied. Here, we demonstrated the spatiotemporal expression pattern of p48 Ebp1 during embryonic development and the postnatal period. During embryonic development, p48 Ebp1 was highly expressed in the brain. Expression gradually decreased after birth but was still more abundant than p42 expression after birth. Strikingly, we found that p48 Ebp1 was expressed in a cell type specific manner in neurons but not astrocytes. Moreover, p48 Ebp1 physically interacted with beta tubulin but not alpha tubulin. This fits with its accumulation in distal microtubule growth cone regions. Furthermore, in injured hippocampal slices, p48 Ebp1 introduction promoted axon regeneration. Thus, we speculate that p48 Ebp1 might contribute to microtubule dynamics acting as an MAP and promotes CNS axon regeneration. [BMB Reports 2017; 50(3): 126-131].

Because ischaemic stroke is one of the most common brain disorders, diverse effective therapies are urgently required. Recent studies reported a variety of azetidine-based scaffolds for the development of central nervous system-focused lead-like libraries. However, their mechanisms of action and in vivo functions remain unclear. Here, we investigated the potential mechanism and beneficial effects of 3-(naphthalen-2-yl(propoxy)methyl)azetidine hydrochloride (KHG26792), a novel azetidine derivative, on ischaemia/reperfusion (I/R) brain injury. We adapted a mouse brain ischaemia model induced by 2 h of middle cerebral artery occlusion followed by 24 h of reperfusion. We measured apoptotic cell death, inflammatory mediators, free radical generation, and anti-oxidative enzymes activities. We also measured the mitochondrial ATP level and Na+, K+-ATPase and cytochrome c oxidase activities. Using western blotting, we analysed the protein levels of inducible NOS, hypoxia-upregulated protein 1, PTEN-induced putative kinase, uncoupling protein 2, p-Akt, MMP-3, and full-length receptor for advanced glycation end-products (RAGE). KHG26792 significantly improved neurological deficits and brain oedema and suppressed I/R-induced apoptosis. KHG26792 significantly attenuated I/R-induced inflammation and oxidative stress by upregulating SOD and catalase activity, GSH, p-Akt, mitochondrial ATP, Na+, K+-ATPase, cytochrome c oxidase, and soluble RAGE and downregulating iNOS, HYOUP1, and MMP-3, suggesting a potential anti-inflammatory and antioxidant role of KHG26792. This is the first study to show that KHG26792 can protect mouse brains against I/R injury by inhibiting apoptotic damage, modulating inflammation, scavenging free radicals, ameliorating oxidative stress, and improving the energy metabolism of the brain, although the clinical relevance of our findings remains unknown.

ErbB3-binding protein 1 (EBP1) is implicated in diverse cellular functions, including apoptosis, cell proliferation, and differentiation. Here, by generating genetic inactivation of Ebp1 mice, we identified the physiological roles of EBP1 in vivo. Loss of Ebp1 in mice caused aberrant organogenesis, including brain malformation, and death between E13.5 and 15.5 owing to severe hemorrhages, with massive apoptosis and cessation of cell proliferation. Specific ablation of Ebp1 in neurons caused structural abnormalities of brain with neuron loss in [Nestin-Cre; Ebp1(flox/flox)] mice. Notably, global methylation increased with high levels of the gene-silencing unit Suv39H1/DNMT1 in Ebp1-deficient mice. EBP1 repressed the transcription of Dnmt1 by binding to its promoter region and interrupted DNMT1-mediated methylation at its target gene, Survivin promoter region. Reinstatement of EBP1 into embryo brain relived gene repression and rescued neuron death. Our findings uncover an essential role for EBP1 in embryonic development and implicate its function in transcriptional regulation.

The different functions of the two isoforms of ErbB3 binding protein 1 (Ebp1), p48 and p42, have recently become the focus of interest as they reveal contradictory roles in cell growth promoting ability. The conformational change that crystal structure of p42 was shown to lack α helices at the amino-terminus present in p48 represents the differential binding partners and protein modifications of two Ebp1 isoforms. N-terminal specific phosphorylation by CDK2 and deregulation of the p53 tumor suppressor through specific interaction with HDM2 and Akt activation is postulated to contribute to p48-mediated tumorigenesis. The short isoform p42 Ebp1, which is actual binding partner of ErbB3 has been implicated as a tumor suppressor with many binding partners such as Rb, HDAC2, Sin3A and the p85 subunit of PI3K with HSP70/CHIP, inhibiting its own antiproliferative activity or inhibiting PI3K activity. The aim of the current review is to provide a summary on distinctive cellular functions of two Ebp1 proteins and their molecular partners that might be responsible for the unique functions of each isoform of Ebp1.

B23, also known as nucleophosmin (NPM), is multifunctional protein directly implicated in cell proliferation, cell cycle progression, and cell survival. In the current study, in addition to confirming its anti-apoptotic function in neuronal survival, we demonstrated that the spatial-temporal expression profile of B23 during development of hippocampal neurons is high in the embryonic stage, down-regulated after birth, and preferentially localized at the tips of growing neuritis and branching points. Overexpression of B23 promotes axon growth with abundant branching points in growing hippocampal neurons, but depletion of B23 impairs axon growth, leading to neuronal death. Following injury to the trisynaptic path in hippocampal slice, overexpression of B23 remarkably increased the number and length of regenerative fibers in the mossy fiber path. Our study suggests that B23 expression in developing neurons is essential for neuritogenesis and axon growth and that up-regulation of B23 may be a strategy for enhancing the reconstitution of synaptic paths after injury to hippocampal synapses.

Background Human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) have been shown to prevent brain damage and improve neurocognition following intraventricular hemorrhage (IVH). However, the molecular mechanisms underlying the effects of hUCB-MSCs are still elusive. Thus, as the hippocampus is essential for learning, memory, and cognitive functions and is intimately involved in the ventricular system, making it a potential site of IVH-induced injury, we determined the molecular basis of the effects of hUCB-derived MSCs on hippocampal neurogenesis and the recovery of hippocampal neural circuits after IVH in a rodent model. Methods We inflicted severe IVH injury on postnatal day 4 (P4) in rats. After confirmation of successful induction of IVH using MRI (P5), intracerebroventricular administration of MSCs (ICV-MSC) was performed at 2 days post-injury (P6). For hippocampal synaptic determination, a rat entorhinal-hippocampus (EH) organotypic slice co-culture (OSC) was performed using day 3 post-IVH brains (P7) with or without ICV-MSCs. A similar strategy of experiments was applied to those rats receiving hUCB-MSC transfected with BDNF-Si-RNA for knockdown of BDNF or scrambled siRNA controls after IVH. The molecular mechanism of the MSCs effects on neurogenesis and the attenuation of neuron death was determined by evaluation of BDNF-TrkB-Akt-CREB signaling axis. Results We showed that treatment with hUCB-MSCs attenuated neuronal loss and promoted neurogenesis in the hippocampus, an area highly vulnerable to IVH-induced brain injury. hUCB-MSCs activate BDNF-TrkB receptor signaling, eliciting intracellular activation of Akt and/or Erk and subsequent phosphorylation of CREB, which is responsible for promoting rat BDNF transcription. In addition to the beneficial effects of neuroprotection and neurogenesis, hUCB-MSCs also contribute to the restoration of impaired synaptic circuits in the hippocampus and improve neurocognitive functions in IVH-injured neonatal rat through BDNF-TrkB-CREB signaling axis activation. Conclusions Our data suggest that hUCB-MSCs possess therapeutic potential for treating neuronal loss and neurocognitive dysfunction in IVH through the activation of intracellular TrkB-CREB signaling that is invoked by hUCB-MSC-secreted BDNF. Electronic supplementary material The online version of this article (10.1186/s13287-018-1052-5) contains supplementary material, which is available to authorized users.

Neurite growth is controlled by a complex molecular signaling network that regulates filamentous actin (F-actin) dynamics at the growth cone. The evolutionarily conserved ezrin, radixin, and moesin family of proteins tether F-actin to the cell membrane when phosphorylated at a conserved threonine residue and modulate neurite outgrowth. Here we show that Akt binds to and phosphorylates a threonine 573 residue on radixin. Akt-mediated phosphorylation protects radixin from ubiquitin-dependent proteasomal degradation, thereby enhancing radixin protein stability, which permits proper neurite outgrowth and growth cone formation. Conversely, the inhibition of Akt kinase or disruption of Akt-dependent phosphorylation reduces the binding affinity of radixin to F-actin as well as lowers radixin protein levels, resulting in decreased neurite outgrowth and growth cone formation. Our findings suggest that Akt signaling regulates neurite outgrowth by stabilizing radixin interactions with F-actin, thus facilitating local F-actin dynamics.

Although the short isoform of ErbB3-binding protein 1 (Ebp1), p42 has been considered to be a potent tumor suppressor in a number of human cancers, whether p42 suppresses tumorigenesis of lung cancer cells has never been clarified. In the current study we investigated the tumor suppressor role of p42 in non-small cell lung cancer cells. Our data suggest that the expression level of p42 is inversely correlated with the cancerous properties of NSCLC cells and that ectopic expression of p42 is sufficient to inhibit cell proliferation, anchorage-independent growth, and invasion as well as tumor growth in vivo. Interestingly, p42 suppresses Akt activation and overexpression of a constitutively active form of Akt restores the tumorigenic activity of A549 cells that is ablated by exogenous p42 expression. Thus, we propose that p42 Ebp1 functions as a potent tumor suppressor of NSCLC through interruption of Akt signaling. [BMB Reports 2015; 48(3): 159-165]

We previously reported the role of vascular endothelial growth factor (VEGF) secreted by mesenchymal stem cells (MSCs) in protecting against neonatal hyperoxic lung injuries. Recently, the paracrine protective effect of MSCs was reported to be primarily mediated by extracellular vesicle (EV) secretion. However, the therapeutic efficacy of MSC-derived EVs and the role of the VEGF contained within EVs in neonatal hyperoxic lung injury have not been elucidated. The aim of the study was to determine whether MSC-derived EVs attenuate neonatal hyperoxic lung injury and, if so, whether this protection is mediated via the transfer of VEGF. We compared the therapeutic efficacy of MSCs, MSC-derived EVs with or without VEGF knockdown, and fibroblast-derived EVs in vitro with a rat lung epithelial cell line challenged with H2O2 and in vivo with newborn Sprague-Dawley rats exposed to hyperoxia (90%) for 14 days. MSCs (1 × 105 cells) or EVs (20 µg) were administered intratracheally on postnatal day 5. The MSCs and MSC-derived EVs, but not the EVs derived from VEGF-knockdown MSCs or fibroblasts, attenuated the in vitro H2O2-induced L2 cell death and the in vivo hyperoxic lung injuries, such as impaired alveolarization and angiogenesis, increased cell death, and activated macrophages and proinflammatory cytokines. PKH67-stained EVs were internalized into vascular pericytes (22.7%), macrophages (21.3%), type 2 epithelial cells (19.5%), and fibroblasts (4.4%) but not into vascular endothelial cells. MSC-derived EVs are as effective as parental MSCs for attenuating neonatal hyperoxic lung injuries, and this protection was mediated primarily by the transfer of VEGF., Neonatal lung damage: Vesicles as vehicles for treatment Membrane-bound sacs called vesicles, secreted by stem cells, carry a protein that might prevent oxygen damaging the lungs of newborn infants. This damage is particularly prevalent in babies born prematurely. The ‘mesenchymal’ stem cells generate cells of connective tissue, blood and blood vessels. They produce vascular endothelial growth factor (VEGF), a protein known to allow the stem cells to protect against neonatal lung injuries caused by oxygen. In studies with rats, Jee-Yin Ahn, Yun Sil Chang and colleagues at Sungkyunkwan University in South Korea, found secreted vesicles carrying VEGF are as effective as the whole stem cells. They also confirmed that the effect was specifically due to the VEGF. The research extends understanding of the protective action of VEGF and suggests the vesicles should be explored for their therapeutic potential in newborn infants.

The activation of microglia is crucially associated with the neurodegeneration observed in many neuroinflammatory pathologies, such as multiple sclerosis, Parkinson's disease, and Alzheimer's disease. We have examined various thiazole derivatives with the goal of developing new anti-neuroinflammatory drugs. Thiazole derivatives are attractive candidates for drug development, because they are efficiently synthesized and active against a number of disease organisms and conditions, including neurodegenerative disorders. The present study investigated the effects of a new compound, N-adamantyl-4-methylthiazol-2-amine (KHG26693), against lipopolysaccharide (LPS)-induced inflammation in cultured BV-2 microglial cells. KHG26693 suppressed several inflammatory responses in LPS-activated cells, as evidenced by decreased levels of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), hydrogen peroxide (H(2)O(2)), reactive oxygen species (ROS), nitric oxide (NO), and lipid peroxidation. These anti-inflammatory/antioxidative actions occurred as a result of the downregulation of NADPH oxidase (NOX), inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2) content, but not as a result of the upregulation of superoxide dismutase (SOD) or catalase activity. The pharmacological properties of KHG26693 were also facilitated via inhibition of both the cluster of differentiation 14 (CD14)/toll-like receptor 4 (TLR4)-dependent nuclear factor kappa B (NF-κB) signaling pathway and extracellular signal-regulated kinase (ERK) phosphorylation. Furthermore, KHG26693 successfully blocked the migration of LPS-activated microglia, most likely by modulating the ERK pathway. Taken together, these results demonstrate that the anti-inflammatory and antioxidative actions of KHG26693 are mediated, at least in part, through the control of microglial activation.

The long isoform of ErbB3 binding protein 1 (Ebp1), p48, strongly promotes tumorigenesis of glioblastoma, accelerating cell proliferation and transformation, while the short isoform, p42, which lacks the N-terminal 54 amino acids, inhibits tumor growth. However, it is unclear if the N-terminal domain of p48 regulates the oncogenic function of p48. Here, we show that p48, but not p42, interacts with cyclin-dependent kinase 2 (CDK2) through its N-terminal domain, resulting in the specific phosphorylation of serine 34 of p48. Overexpression of wild-type p48 greatly enhanced tumor cell growth, whereas phospho-ablated mutant S34A of p48, which is mutated at the CDK2 phosphorylation site, antagonizes cell proliferation and transformation. Moreover, phospho-ablated mutant S34A abrogated the ability of p48 to accelerate tumor cell growth in a mouse engraft model. Thus, our findings indicate that p48Ebp1 acts as an oncoprotein through selective interaction and/or modification of the N-terminal domain that does not exist in its short isoform p42.

Cucurbitacin B, a member of the cucurbitaceae family, can act as a STAT3 signaling inhibitor to regulate the growth of hepatocellular carcinoma. STAT3 signaling has been shown to inhibit adipocyte differentiation through C/EBPα and PPARγ. Based on these studies, we hypothesized that cucurbitacin B would prevent PPARγ mediated adipocyte differentiation through STAT3 signaling. To test this hypothesis, mesenchymal C3H10T1/2 and 3T3-L1 preadipocyte cells were treated with a sub-cytotoxic concentration of cucurbitacin B. Cucurbitacin B treatment inhibits lipid accumulation and expression of adipocyte markers including PPARγ and its target genes in a dose-dependent manner. Cucurbitacin B treatment impairs STAT3 signaling as manifested by reduced phosphorylation of STAT3 and suppression of STAT3 target gene expression in preadipocytes. The anti-adipogenic effects of cucurbitacin B are significantly blunted in cells with STAT3 silenced by introducing small interfering RNA. Finally, our data show that cucurbitacin I, another cucurbitacin family member, also inhibits adipocyte differentiation by suppressing STAT3 signaling. Together, our data suggest the possibility of utilizing cucurbitacins as a new strategy to treat metabolic diseases and implicate STAT3 as a new target for the development of functional foods and drugs.

Severe intraventricular hemorrhage (IVH) remains a major cause of mortality and long-term neurologic morbidities in premature infants, despite recent advances in neonatal intensive care medicine. Several preclinical studies have demonstrated the beneficial effects of mesenchymal stem cell (MSC) transplantation in attenuating brain injuries resulting from severe IVH. Because there currently exists no effective intervention for severe IVH, the therapeutic potential of MSC transplantation in this intractable and devastating disease is creating excitement in this field. This review summarizes recent progress in stem cell research for treating neonatal brain injury due to severe IVH, with a particular focus on preclinical data concerning important issues, such as mechanism of protective action and determining optimal source, route, timing, and dose of MSC transplantation, and on the translation of these preclinical study results to a clinical trial.