William Yen

Nuclear Factor Kappa B Signaling Initiates Early Differentiation of Neural Stem Cells

Inflammatory mediators, many of which activate the signaling of nuclear factor kappa B (NFκB), have received increasing attention in the field of neurogenesis. NFκB signaling regulates neurite outgrowth and neural plasticity as well as the proliferation/apoptosis and terminal differentiation of neural stem cells (NSCs). Early neurogenesis from NSCs produces identical progeny through symmetric division and committed daughter cells through asymmetric division. Here, we show that NFκB signaling is required for NSC initial differentiation. The canonical IKKβ/IκBα/p65 pathway is activated during the initial stages of neural differentiation induced by treatment with TNFα or withdrawal of epidermal growth factor/basic fibroblast growth factor. NSC‐specific inhibition of NFκB in transgenic mice causes an accumulation of Nestin+/Sox2+/glial fibrillary acidic protein+ NSCs. Inhibition of NFκB signaling in vitro blocks differentiation and asymmetric division and maintains NSCs in an undifferentiated state. The induction of initial differentiation and asymmetry by NFκB signaling occurs through the inhibition of C/EBPβ expression. Our data reveal a novel function of NFκB signaling in early neurogenesis and provide insight into the molecular mechanisms underlying neurodevelopmental disorders and neurodegenerative diseases. STEM CELLS 2012;30:510–524

Cocaine-Induced Loss of White Matter Proteins in the Adult Mouse Nucleus Accumbens Is Attenuated by Administration of a β-Lactam Antibiotic during Cocaine Withdrawal

We report significantly decreased white matter protein levels in the nucleus accumbens in an adult mouse model of chronic cocaine abuse. Previous studies from human cocaine abuse patients show disruption of white matter and myelin loss, thus supporting our observations. Understanding the neuropathological mechanisms for white matter disruption in cocaine abuse patients is complicated by polydrug use and other comorbid factors, hindering the development of effective therapeutic strategies to ameliorate damage or compliment rehabilitation programs. In this context, our data further demonstrate that cocaine-induced loss of white matter proteins is absent in mice treated with the β-lactam antibiotic, ceftriaxone, during cocaine withdrawal. Other studies report that ceftriaxone, a glutamate transporter subtype-1 activator, is neuroprotective in murine models of multiple sclerosis, thereby demonstrating potential therapeutic properties for diseases with white matter loss. Cocaine-induced white matter abnormalities likely contribute to the cognitive, motor, and psychological deficits commonly afflicting cocaine abusers, yet the underlying mechanisms responsible for these changes remain unknown. Our observations describe an adult animal model for the study of cocaine-induced myelin loss for the first time, and highlight a potential pharmacological intervention to ameliorate cocaine-induced white matter loss.

Ceftriaxone attenuates locomotor activity induced by acute and repeated cocaine exposure in mice

Ceftriaxone (CTX) decreases locomotor activation produced by initial cocaine exposure and attenuates development of behavioral sensitization produced by repeated cocaine exposure. An important question that has not yet been answered is whether or not CTX reduces behavioral sensitization to cocaine in cases in which the antibiotic is administered only during the period of cocaine absence that follows repeated cocaine exposure and precedes reintroduction to cocaine. We investigated this question using C57BL/6 mice. Mice pretreated with cocaine (15mg/kg×14 days) and then challenged with cocaine (15mg/kg) after 30 days of cocaine absence displayed sensitization of locomotor activity. For combination experiments, CTX injected during the 30 days of cocaine absence attenuated behavioral sensitization produced by cocaine challenge. In the case in which CTX was injected together with cocaine for 14 days, development of behavioral sensitization to cocaine challenge was also reduced. CTX attenuated the increase in locomotor activity produced by acute cocaine exposure; however, its efficacy was dependent on the dose of cocaine as inhibition was detected against 30mg/kg, but not 15mg/kg, of cocaine. These results from mice indicate that CTX attenuates locomotor activity produced by acute and repeated cocaine exposure and counters cocaines locomotor activating properties in a paradigm in which the antibiotic is injected during the period of forced cocaine absence that follows repeated cocaine exposure.

PINCH in the cellular stress response to tau-hyperphosphorylation.

Particularly interesting new cysteine- histidine- rich protein (PINCH) is an adaptor protein that our data have shown is required for neurite extension under stressful conditions. Our previous studies also report that PINCH is recalled by neurons showing decreased levels of synaptodendritic signaling proteins such as MAP2 or synaptophysin in the brains of human immunodeficiency virus (HIV) patients. The current study addressed potential role(s) for PINCH in neurodegenerative diseases. Mass spectrometry predicted the interaction of PINCH with Tau and with members of the heat shock response. Our in vitro data confirmed that PINCH binds to hyperphosphorylated (hp) Tau and to E3 ubiquitin ligase, carboxy-terminus of heat shock-70 interacting protein. Silencing PINCH prior to induction of hp-Tau resulted in more efficient clearance of accumulating hp-Tau, suggesting that PINCH may play a role in stabilizing hp-Tau. Accumulation of hp-Tau is implicated in more than 20 neuropathological diseases including Alzheimers disease (AD), frontotemporal dementia (FTD), and human immunodeficiency virus encephalitis (HIVE). Analyses of brain tissues from HIVE, AD and FTD patients showed that PINCH is increased and binds to hp-Tau. These studies address a new mechanism by which AD and HIV may intersect and identify PINCH as a contributing factor to the accumulation of hyperphosphorylated Tau.

Evidence for involvement of NFBP in processing of ribosomal RNA.

Ribosomal RNA (rRNA) in vertebrates is initially transcribed as a single 47S precursor which is modified by the addition of 2′‐O‐methyl ribose moieties, pseudouridines, and methyl groups, followed by cleavage at several sites to produce the mature 28S, 18S, and 5.8S rRNAs. Cleavage of the rRNA precursor to generate the 18S rRNA is mediated by a ribonucleoprotein (RNP) complex termed the processome containing U3, a box C/D small nucleolar RNA (snoRNA), and at least 28 cellular proteins. We previously identified a novel human RNA binding protein, NF‐κB binding protein (NFBP), which is the human homolog of Rrp5p, a protein component of the yeast U3 processome. Here, we show that NFBP colocalizes with and coprecipitates U3 in the nucleolus. We also demonstrate that NFBP is essential for the generation of 18S rRNA as maturation of the 18S rRNA is repressed in the absence of NFBP. Using Northern blot analyses, we further show that NFBP is specifically necessary for cleavages at sites A0, 1, and 2, as unprocessed intermediate forms of rRNA accumulated in the absence of NFBP. J. Cell. Physiol. 214: 381–388, 2008.

Cocaine decreases expression of neurogranin via alterations in thyroid receptor/retinoid X receptor signaling

J. Neurochem. (2012) 121, 302–313.

Cocaine Induces Nuclear Export and Degradation of Neuronal Retinoid X Receptor-γ via a TNF-α/JNK- Mediated Mechanism

Cocaine abuse represents an immense societal health and economic burden for which no effective treatment currently exists. Among the numerous intracellular signaling cascades impacted by exposure to cocaine, increased and aberrant production of pro-inflammatory cytokines in the CNS has been observed. Additionally, we have previously reported a decrease in retinoid-X-receptor-gamma (RXR-γ) in brains of mice chronically exposed to cocaine. Through obligate heterodimerization with a number of nuclear receptors, RXRs serve as master regulatory transcription factors, which can potentiate or suppress expression of a wide spectrum of genes. Little is known about the regulation of RXR levels, but previous studies indicate cellular stressors such as cytokines negatively regulate levels of RXRs in vitro. To evaluate the mechanism underlying the cocaine-induced decreases in RXR-γ levels observed in vivo, we exposed neurons to cocaine in vitro and examined pathways which may contribute to disruption in RXR signaling, including activation of stress pathways by cytokine induction. In these studies, we provide the first evidence that cocaine exposure disrupts neuronal RXR-γ signaling in vitro by promoting its nuclear export and degradation. Furthermore, we demonstrate this effect may be mediated, at least in part, by cocaine-induced production of TNF-α and its downstream effector c-Jun-NH-terminal kinase (JNK). Findings from this study are therefore applicable to both cocaine abuse and to pathological conditions characterized by neuroinflammatory factors, such as neurodegenerative disease.

Dysregulation of PINCH signaling in mesial temporal epilepsy

Mounting evidence suggests that inflammation is important in epileptogenesis. Particularly Interesting New Cysteine Histidine-rich (PINCH) protein is a highly conserved, LIM-domain protein known to interact with hyperphosphorylated Tau. We assessed PINCH expression in resected epileptogenic human hippocampi and further explored the relationships among PINCH, hpTau and associated kinases. Resected hippocampal tissue from 7 patients with mesial temporal lobe epilepsy (MTLE) was assessed by Western analyses to measure levels of PINCH and hyperphosphorylated Tau, as well as changes in phosphorylation levels of associated kinases AKT and GSK3β in comparison to normal control tissue. Immunolabeling was also conducted to evaluate PINCH and hpTau patterns of expression, co-localization and cell-type specific expression. Hippocampal PINCH was increased by 2.6 fold in the epilepsy cases over controls and hpTau was increased 10 fold over control. Decreased phospho-AKT and phospho-GSK3β in epilepsy tissue suggested involvement of this pathway in MTLE. PINCH and hpTau co-localized in some neurons in MTLE tissue. While PINCH was expressed by both neurons and astrocytes in MTLE tissue, hpTau was extracellular or associated with neurons. PINCH was absent from the serum of control subjects but readily detectable from the serum of patients with chronic epilepsy. Our study describes the expression of PINCH and points to AKT/GSK3β signaling dysregulation as a possible pathway in hpTau formation in MTLE. In view of the interactions between hpTau and PINCH, understanding the role of PINCH in MTLE may provide increased understanding of mechanisms leading to inflammation and MTLE epileptogenesis and a potential biomarker for drug-resistant epilepsy.

609 Tumor Necrosis Factor-α and Lymphotoxin-α1β2 Induce Enteric Neuronal Differentiation Through the NfκB Signaling Pathways

recorded from 12 rats 10 hrs after cisplatin (5 mg/kg). Basal firings increased to 9.5±3.5 impulses/20s. 12/45 neurons responded to SP. SP 5 and 10 μg/kg increased firings to 27 ± 4, and 64±7 impulses/20 s, respectively. 7/12 SP sensitive neurons also responded to 5HT. Luminal perfusion of 10-7M increased firings to 17±3 impulses/20 s. Combined 10-7M 5-HT and 5, 10 μg/kg of SP enhanced neuronal firings to 60 ± 5, and 133±11 impulses/ 20 s, respectively. Western blot studies demonstrated a 1.1 fold increase in NK1 receptor expression but no change in 5HT3 receptor in the NG 10 hrs following cisplatin. Similar increase in vagal NK1 receptor expression was observed with perfusion of 5-HT (10-5M) for 5 hrs. Administration PCPA, a 5-HT synthesis inhibitor suppressed cisplatin-induced increased in NK1 receptor and completely abolished the enhanced neuronal responses to SP. Conclusion: Anti-cancer therapy with cisplatin causes plastic changes in the NG characterized by upregulation of NK1 receptor in the vagal NG. This is mediated by the 5-HT pathways. Enhanced activation of NK1 receptor in the vagal afferent neurons and its interaction with 5-HT appear to be responsible for the delay emesis observed in anti-cancer chemotherapy.