Altered gut microbiota in Taiwanese A97S predominant transthyretin amyloidosis with polyneuropathy.

Increasing evidence suggests that gut microbiota alterations are related to development and phenotypes of many neuropsychiatric diseases. Here, we evaluated the fecal microbiota and its clinical correlates in patients with hereditary transthyretin amyloidosis (ATTRv) and polyneuropathy. Fecal microbiota from 38 ATTRv patients and 39 age-matched controls was analyzed by sequencing 16S V3-V4 ribosomal RNA, and its relationships with clinical characteristics of polyneuropathy and cardiomyopathy were explored. The familial amyloidotic polyneuropathy stage was stage I, II, and III in 13, 18, and 7 patients. 99mTc-PYP SPECT showed a visual score of 2 in 15 and 3 in 21 patients. The gut microbiota of ATTRv patients showed higher alpha diversity (ASV richness and Shannon effective numbers) and dissimilar beta diversity compared to controls. Relative abundance of microbiota was dominated by Firmicutes and decreased in Bacteroidetes in ATTRv patients than in controls. Patients with more myocardial amyloid deposition were associated with increased alpha diversity, and the abundance of Clostridia was significantly correlated with pathophysiology of polyneuropathy in ATTRv patients. These findings demonstrated alterations in the gut microbiota, especially Firmicutes, in ATTRv. The association between altered microbiota and phenotypes of cardiomyopathy and polyneuropathy might suggest potential contributions of gut microbiota to ATTRv pathogenesis.

The Potential Benefits of Quercetin for Brain Health: A Review of Anti-Inflammatory and Neuroprotective Mechanisms.

Neuroinflammation is a critical factor in developing and progressing numerous brain diseases, including neurodegenerative diseases. Chronic or excessive neuroinflammation can lead to neurotoxicity, causing brain damage and contributing to the onset and progression of various brain diseases. Therefore, understanding neuroinflammation mechanisms and developing strategies to control them is crucial for treating brain diseases. Studies have shown that neuroinflammation plays a vital role in the progression of neurodegenerative diseases, such as Alzheimer's (AD) and Parkinson's (PD), and stroke. Additionally, the effects of PM2.5 pollution on the brain, including neuroinflammation and neurotoxicity, are well-documented. Quercetin is a flavonoid, a plant pigment in many fruits, vegetables, and grains. Quercetin has been studied for its potential health benefits, including its anti-inflammatory, antioxidant, and anti-cancer properties. Quercetin may also have a positive impact on immune function and allergy symptoms. In addition, quercetin has been shown to have anti-inflammatory and neuroprotective properties and can activate AMP-activated protein kinase (AMPK), a cellular energy sensor that modulates inflammation and oxidative stress. By reducing inflammation and protecting against neuroinflammatory toxicity, quercetin holds promise as a safe and effective adjunctive therapy for treating neurodegenerative diseases and other brain disorders. Understanding and controlling the mechanisms of NF-κB and NLRP3 inflammasome pathways are crucial for preventing and treating conditions, and quercetin may be a promising tool in this effort. This review article aims to discuss the role of neuroinflammation in the development and progression of various brain disorders, including neurodegenerative diseases and stroke, and the impact of PM2.5 pollution on the brain. The paper also highlights quercetin's potential health benefits and anti-inflammatory and neuroprotective properties.

Objective Signal Analysis for Investigating Feasibility of Active Noise Cancellation in Hearing Screening.

With the development of active noise cancellation (ANC) technology, ANC has been used to mitigate the effects of environmental noise on audiometric results. However, objective evaluation methods supporting the accuracy of audiometry for ANC exposure to different levels of noise have not been reported. Accordingly, the audio characteristics of three different ANC headphone models were quantified under different noise conditions and the feasibility of ANC in noisy environments was investigated. Steady (pink noise) and non-steady noise (cafeteria babble noise) were used to simulate noisy environments. We compared the integrity of pure-tone signals obtained from three different ANC headphone models after processing under different noise scenarios and analyzed the degree of ANC signal correlation based on the Pearson correlation coefficient compared to pure-tone signals in quiet. The objective signal correlation results were compared with audiometric screening results to confirm the correspondence. Results revealed that ANC helped mitigate the effects of environmental noise on the measured signal and the combined ANC headset model retained the highest signal integrity. The degree of signal correlation was used as a confidence indicator for the accuracy of hearing screening in noise results. It was found that the ANC technique can be further improved for more complex noisy environments.

Resveratrol Mitigates Oxygen and Glucose Deprivation-Induced Inflammation, NLRP3 Inflammasome, and Oxidative Stress in 3D Neuronal Culture.

Oxygen glucose deprivation (OGD) can produce hypoxia-induced neurotoxicity and is a mature in vitro model of hypoxic cell damage. Activated AMP-activated protein kinase (AMPK) regulates a downstream pathway that substantially increases bioenergy production, which may be a key player in physiological energy and has also been shown to play a role in regulating neuroprotective processes. Resveratrol is an effective activator of AMPK, indicating that it may have therapeutic potential as a neuroprotective agent. However, the mechanism by which resveratrol achieves these beneficial effects in SH-SY5Y cells exposed to OGD-induced inflammation and oxidative stress in a 3D gelatin scaffold remains unclear. Therefore, in the present study, we investigated the effect of resveratrol in 3D gelatin scaffold cells to understand its neuroprotective effects on NF-κB signaling, NLRP3 inflammasome, and oxidative stress under OGD conditions. Here, we show that resveratrol improves the expression levels of cell viability, inflammatory cytokines (TNF-α, IL-1ß, and IL-18), NF-κB signaling, and NLRP3 inflammasome, that OGD increases. In addition, resveratrol rescued oxidative stress, nuclear factor-erythroid 2 related factor 2 (Nrf2), and Nrf2 downstream antioxidant target genes (e.g., SOD, Gpx GSH, catalase, and HO-1). Treatment with resveratrol can significantly normalize OGD-induced changes in SH-SY5Y cell inflammation, oxidative stress, and oxidative defense gene expression; however, these resveratrol protective effects are affected by AMPK antagonists (Compounds C) blocking. These findings improve our understanding of the mechanism of the AMPK-dependent protective effect of resveratrol under 3D OGD-induced inflammation and oxidative stress-mediated cerebral ischemic stroke conditions.

Cardiomyopathy correlates to nerve damage in p.A117S late-onset transthyretin amyloid polyneuropathy.

OBJECTIVE: Late-onset hereditary transthyretin amyloidosis with polyneuropathy (ATTRv-PN) is often associated with heart involvement. Recent advances in cardiac imaging allow the detection of cardiac amyloidosis. This study aimed to explore cardiomyopathy by cardiac imaging and its clinical correlates with polyneuropathy in late-onset ATTRv-PN. METHODS: Polyneuropathy was assessed by intraepidermal nerve fiber (IENF) density, nerve conduction study (NCS), autonomic function tests, quantitative sensory testing, and clinical questionnaires. Cardiomyopathy was evaluated by echocardiography, 99m Tc-pyrophosphate (PYP) single-photon emission computed tomography (SPECT) imaging, cardiac magnetic resonance imaging (CMR), and serum Pro-B-type natriuretic peptide. Healthy controls and patients with Brugada syndrome were enrolled for comparison of CMR. RESULTS: Fifty late-onset ATTRv-PN patients (38 men, 46 with p. A117S mutation), aged 63.7 ± 5.5 years, of polyneuropathy disability stage 1-4 were enrolled. All patients presented polyneuropathy in NCS, and 74.5% of patients had reduced IENF density in distal legs. All patients showed significant radiotracer uptake in the heart on 99m Tc-PYP SPECT imaging, and 87.8% of patients had abnormally increased left ventricular (LV) septum thickness on echocardiography. CMR showed longer myocardial native T1, larger extracellular volume, greater LV mass index, and higher LV mass to end-diastolic volume ratio in ATTRv-PN patients than healthy controls and patients with Brugada syndrome. These CMR parameters were associated with skin denervation, absent sympathetic skin responses, elevated thermal thresholds, worsened NCS profiles, and functional deficits of polyneuropathy. INTERPRETATION: Late-onset ATTRv-PN coexisted with cardiomyopathy regardless of the clinical severity of polyneuropathy. The cardiac amyloid burden revealed by CMR was correlated with pathophysiology and clinical disability of nerve degeneration.

GSH-AuNP anti-oxidative stress, ER stress and mitochondrial dysfunction in amyloid-beta peptide-treated human neural stem cells.

Amyloid-beta (Aß) peptides have a role in the pathogenesis of Alzheimer's disease (AD) and are thought to promote oxidative stress, endoplasmic reticulum (ER) stress and mitochondrial deficiency, causing neuronal loss in the AD brain. The potential applications of glutathione conjugated gold nanoparticles (GSH-AuNPs) suggest they might have therapeutic value. Several studies have demonstrated that the effects of nanoparticles could provide protective roles in AD. Here, we showed that GSH-AuNPs mediate the viability of human neural stem cells (hNSCs) with Aß, which was correlated with decreased caspase 3 and caspase 9. Importantly, hNSCs co-treated with GSH-AuNPs were significantly protected from Aß-induced oxidative stress, as detected using the DCFH-DA, DHE, and MitoSOX staining assays. Furthermore, hNSCs co-treated with GSH-AuNPs were significantly protected from the Aß-induced reduction in the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and Nrf2 downstream antioxidant target genes (SOD-1, SOD-2, Gpx, Catalase, and HO-1). In addition, GSH-AuNPs rescued the expression levels of ER stress-associated genes (Bip, CHOP, and ASK1) in Aß-treated hNSCs. GSH-AuNPs normalized ER calcium and mitochondrial cytochrome c homeostasis in Aß-treated hNSCs. Furthermore, treatment with GSH-AuNPs restored the levels of ATP, D-loop, mitochondrial mass, basal respiration, ATP-linked reparation, maximal respiration capacity, COX activity, mitochondrial membrane potential, and mitochondrial genes (PGC1α, NRF-1 and Tfam) in Aß-treated hNSCs. Taken together, these findings extend our understanding of the protective effects of GSH-AuNPs against oxidative stress, ER stress and mitochondrial dysfunction in hNSCs with Aß.

Exposure to PM2.5 induces neurotoxicity, mitochondrial dysfunction, oxidative stress and inflammation in human SH-SY5Y neuronal cells.

Ambient air pollution is a global public health issue. Recent evidence suggests that exposure to fine aerosolized particulate matter (PM) as small as ≤2.5 microns (PM2.5) is neurotoxic to brain structures. Many studies also suggest exposure to PM2.5 may cause neurotoxicity and affect brain function. However, the molecular mechanisms by which PM2.5 exerts these effects are not fully understood. Thus, we evaluated the hypothesis that PM2.5 exposure exerts its neurotoxic effects via increased oxidative and inflammatory cellular damage and mitochondrial dysfunction using human SH-SY5Y neuronal cells. Here, we show PM2.5 exposure significantly decreases viability, and increases caspase 3 and 9 protein expression and activity in SH-SY5Y cells. In addition, PM2.5 exposure decreases SH-SY5Y survival, disrupts cell and mitochondrial morphology, and significantly decreases ATP levels, D-loop levels, and mitochondrial mass and function (maximal respiratory function, COX activity, and mitochondrial membrane potential) in SH-SY5Y cells. Moreover, SH-SY5Y cells exposed to PM2.5 have significantly decreased mRNA and protein expression levels of survival genes (CREB and Bcl-2) and neuroprotective genes (PPARγ and AMPK). We further show SH-SY5Y cells exposure to PM2.5 induces significant increases in the levels of oxidative stress, and expression levels of the inflammatory mediator's TNF-α, IL-1ß, and NF-κB. Taken together, these results provide the first evidence of the biochemical, molecular and morphological effects of PM2.5 on human neuronal SH-SY5Y cells, and support our hypothesis that increased mitochondrial disruption, oxidative stress and inflammation are critical mediators of its neurotoxic effects. These findings further improve our understanding of the neuronal cell impact of PM2.5 exposure, and may be useful in the design of strategies for the treatment and prevention of human neurodegenerative disorders.

Early changes of nerve integrity in preclinical carriers of hereditary transthyretin Ala117Ser amyloidosis with polyneuropathy.

BACKGROUND AND PURPOSE: Disease-modifying therapies provide new horizons for hereditary transthyretin amyloidosis with polyneuropathy (ATTRv-PN) to slow neuropathic progression. Initiating treatment at the earliest time requires biomarkers reflecting both small- and large-fiber degeneration in carriers. METHODS: This study included examinations of pathology (intraepidermal nerve fiber [IENF] density), physiology (nerve conduction studies, autonomic function test, and nerve excitability), and psychophysics (thermal thresholds) in carriers to compare to healthy controls and asymptomatic diabetic patients. RESULTS: There were 43 carriers (44.2 ± 11.4 years, p.Ala117Ser in 42 carriers), 43 controls (43.4 ± 12.7 years) including 26 noncarrier families, and 50 asymptomatic diabetic patients (58.1 ± 9.5 years). Carriers had lower IENF densities than controls and similar densities as diabetic patients. Median nerve conduction parameters, especially distal motor latency, were the most frequent neurophysiological abnormality in carriers, could differentiate carriers from controls and diabetic patients, were correlated with IENF densities in carriers but not in controls and diabetic patients, and were correlated with nerve excitability parameters in carriers but not in controls. Fifteen carriers (34.9%) with electrophysiological evidence of median nerve entrapment at the wrist had lower IENF densities and more abnormal conduction parameters than carriers without. We defined nerve dysfunction index-the ratio of median distal motor latency to IENF density-which differentiated carriers from controls. CONCLUSIONS: In late-onset ATTRv-PN carriers with predominant p.Ala117Ser, median conduction parameters were the most common neurophysiological abnormalities and served as surrogate signatures of small- and large-fiber impairment. Combination of median distal motor latency and IENF density can reflect early neuropathy in carriers.

Involvement of MicroRNA-296 in the Inhibitory Effect of Epigallocatechin Gallate against the Migratory Properties of Anoikis-Resistant Nasopharyngeal Carcinoma Cells.

Short noncoding endogenous RNAs, including microRNAs (miRNAs), are associated with the development and metastasis of multiple cancers. Epigallocatechin gallate (EGCG), the most active and abundant polyphenol in green tea, plays a crucial role in the modulation of miRNA expression, which is related to changes in cancer progression. In the present study, we explore whether EGCG exerts its suppressive effects on nasopharyngeal carcinoma (NPC) cells through miRNA regulation. The anoikis-resistant sphere-forming NPC cells grown under anchorage-independent conditions exhibit enhanced migratory properties, which were inhibited by EGCG treatment. The miR-296 level was lower in the anoikis-resistant cells than in the monolayer parental cells; however, miR-296 was significantly upregulated after EGCG treatment. We demonstrate that miR-296 is involved in the inhibitory effects of EGCG on the anoikis-resistant NPC cells through the downregulation of signal transducer and activator of transcription 3 (STAT3) activation. Our study is the first to demonstrate that EGCG inhibited the migratory properties of anoikis-resistant cells by modulating the expression of miRNA in NPC cells. Our results indicate the novel effects of EGCG on miRNA regulation to inhibit an invasive phenotype of NPC as well as the regulatory role of miR-296.

Neuroprotective effects of resveratrol against oxygen glucose deprivation induced mitochondrial dysfunction by activation of AMPK in SH-SY5Y cells with 3D gelatin scaffold.

Ischemic stroke arising from the sudden blockage of arteries in the brain, is a common and serious brain damaging problem worldwide, often leading to disability or death. The oxygen glucose deprivation (OGD) model was created to improve understanding of hypoxia- and hypoglycemia-induced neuronal cell injury, and provide an in vitro surrogate to assess novel treatments for cerebral hypoxia-ischemia. AMP-activated protein kinase (AMPK) is a critical neuroprotective regulator of energy homeostasis, metabolism and cell survival. However, the neuroprotective mechanisms by which AMPK achieves these beneficial effects in human SH-SY5Y neural cells exposed to OGD are still not well understood. Resveratrol is a potent activator of AMPK suggesting it may have therapeutic potential as a neuroprotective agent. Therefore, we hypothesized the AMPK activator resveratrol protects against OGD-mediated impairment of human SH-SY5Y neuronal cells. The novelty of the experiment using a 3D gelatin scaffold cell culture assay, we have tested the potential of 3D systems to mimic the endogenous neuronal environment and have applied these systems to study the effect of OGD on neuronal cells with/without resveratrol. Here we show resveratrol reverses, via AMPK-dependent downregulation of caspase 3 and 9 activity, the OGD-mediated decreases in SH-SY5Y cell viability on a 3D gelatin scaffold. In addition, treatment with OGD decreases mRNA levels of AMPK and the neuroprotective genes (Bcl-2 and CREB); however, co-treatment with resveratrol significantly normalizes these effects. Importantly, resveratrol improves the expression of AMPK and p-AMPK in OGD-exposed SH-SY5Y cells. Resveratrol also significantly rescues SH-SY5Y cells from OGD-mediated mitochondrial deficiency (lower D-loop level, mitochondrial mass, maximal respiratory function, COX activity, and mitochondrial membrane potential). Resveratrol also rescues the transcript expression levels of PGC1α and mitochondrial genes (NRF-1 and Tfam) in OGD-treated SH-SY5Y cells. These findings extend our mechanistic understanding of the central role of AMPK in OGD-related neuronal impairment, and may serve as basis for implementing new therapeutic strategies in the treatment of ischemic stroke.

Pparα deficiency inhibits the proliferation of neuronal and glial precursors in the zebrafish central nervous system.

BACKGROUND: Many molecules and signaling pathways involved in neural development play a role in neurodegenerative diseases and brain tumor progression. Peroxisome proliferator-activated receptor (PPAR) proteins regulate the differentiation of tissues and the progression of many diseases. However, the role of these proteins in neural development is unclear. RESULTS: We examined the function of Pparα in the neural development of zebrafish. Two duplicate paralogs for mammalian PPARA/Ppara, namely pparaa and pparab, are present in the zebrafish genome. Both pparaa and pparab are expressed in the developing central nervous system in zebrafish embryos. Inhibiting the function of Pparα by using either the PPARα/Pparα antagonist GW6471 or pparaa or pparab truncated constructs produced identical phenotypes, which were sufficient to reduce the proliferation of neuronal and glial precursor cells without affecting the formation of neural progenitors. CONCLUSIONS: We demonstrated that both Pparαa and Pparαb proteins are essential regulators of the proliferation of neuronal and glial precursors. This study provides a better understanding of the functions of PPARα/Pparα in neural development and further expands our knowledge of the potential role of PPARα/Pparα in neurological disorders and brain tumors. Developmental Dynamics 247:1264-1275, 2018. © 2018 Wiley Periodicals, Inc.

MicroRNA-328 inhibits migration and epithelial-mesenchymal transition by targeting CD44 in nasopharyngeal carcinoma cells.

BACKGROUND: MicroRNAs (miRNAs) play crucial roles in various types of cancers, particularly in tumor development, migration, and progression. Dysregulation of miR-328 was reported to occur in some types of human malignancies, however, the role of miR-328 in nasopharyngeal carcinoma (NPC) and its potential involvement in metastasis remain undetermined. METHODS: The invasion capacity of NPC sphere-forming cells was evaluated by in vitro cell migration assays. Differential miRNAs expression was examined in NPC sphere-forming cells compared to parental monolayer cells using miRNA array analysis. The role of miR-328 in regulating NPC cells migratory properties was analyzed after miR-328 mimics transfection. The expression of E-cadherin and CD44 was analyzed by flow cytometry. CD44 was examined as a target of miR-328 through luciferase reporter assays and Western blotting. RESULTS: Here, we report that NPC TW01 and TW06 sphere-forming cells exhibited increased migratory ability in comparison with parental monolayer cells. Sphere-forming cells had significantly lower levels of miR-328, as observed using miRNA arrays and confirmed through real-time polymerase chain reaction. Overexpression of miR-328 induced by transfection with synthetic miR-328 mimics decreased the migration of NPC sphere-forming cells. The inhibitory effects were associated with increased expression of E-cadherin and the downregulated expression of mesenchymal markers such as N-cadherin, Snail, and vimentin. Moreover, our results demonstrated that miR-328 suppressed NPC cell migration and inhibited the epithelial-mesenchymal transition process directly through a binding site on the CD44 3' untranslated region. CONCLUSION: miR-328, a previously unrecognized miRNA, may serve as a potential prognostic marker and therapeutic target for NPC.

The neuroprotective role of rosiglitazone in advanced glycation end product treated human neural stem cells is PPARgamma-dependent.

Hyperglycemia is accompanied by an accelerated formation rate of advanced glycation end products (AGEs), which is associated with the pathogenesis of diabetic neuronal deficits. Peroxisome proliferator-activated receptor gamma (PPARγ) belongs to a family of ligand-activated nuclear receptors and its ligands are known to control many physiological, pathological and inflammatory pathways. Weinvestigated the hypothesis that the PPARγ agonist (rosiglitazone) would abrogate AGEs-mediated neurotoxic effects on human neural stem cells (hNSCs), by whichAGEs may play a role in diabetic-related neuronal impairment. Here, we show that rosiglitazone treatment increases cell viability of hNSCs via downregulation of caspase 3 activity. These rescue effects were extended in our studies showingrosiglitazone-mediated activation of PPARγ reversed the expression levels of two neuroprotective factors (Bcl-2 and PGC1α) that were downregulated in hNSCs exposed to AGEs alone. The stimulation of mitochondrial function and anti-oxidative stress by rosiglitazone was associated with activation of the PGC1α pathway by up-regulation of mitochondrial (NRF-1 and Tfam) and oxidative defense (SOD1, SOD2 and Gpx1) genes. Moreover, rosiglitazone significantly normalized the inflammatory responses (TNF-α and IL-1ß), NF-κB (p65), and inflammatory genes (iNOS and COX-2) in the hNSCs treated with AGEs. This neuroprotective effect of rosiglitazone was effectively blocked by PPARγ-specific antagonist (GW9662), demonstrating that the action of rosiglitazone was mediated by at PPARγ-dependent pathway. Collectively, these novel findings show AGEs induce neurotoxic effects in hNSCs, and provide important mechanistic insights that may explain the increased risk of neuronal impairment deficits in diabetic patients. More importantly, these data show rosiglitazone-mediated activation of PPARγ-dependent signaling is neuroprotective in AGE-treated hNSCs, and suggests PPARγ ligands may be useful in the therapeutic management of patients with neurodegenerative diseases.

STAT3 mediates resistance to anoikis and promotes invasiveness of nasopharyngeal cancer cells.

Nasopharyngeal carcinoma (NPC), a tumor arising from the epithelial cells of the nasopharynx, is endemic in Southeast Asia and Taiwan. The prognosis of NPC patients with local recurrence and metastasis is poor. Resistance to anoikis is a primary characteristic of tumor cells that metastasize. However, the mechanism through which NPC cells resist anoikis and are able to metastasize has not been fully elucidated. In the present study, the acquisition of anoikis resistance was analyzed in the TW01 and TW06 human NPC cell lines growing under anchorage-independent conditions. A considerable number of TW01 and TW06 cells was found to be resistant to anoikis and exhibited a higher capability of migration and invasion. These anoikis-resistant NPC cells exhibited significantly increased expression of signal transducer and activation of transcription 3 (Stat3) compared with adherent cells. Furthermore, blockade of STAT3 expression by STAT3 inhibitors or STAT3 silencing significant increased anoikis in anoikis-resistant NPC cells. Moreover, silencing STAT3 not only reduced the capacity of NPC cells to resist anoikis, but also reversed their invasive properties. The expression of epithelial­to-mesenchymal transition­related proteins and CD44 was also significantly decreased following STAT3 knockdown. The results of the present study established that STAT3 mediates anoikis resistance, with enhanced cell migration and invasion of NPC cells, and that activation of STAT3 may increase metastatic capacity, indicating the crucial role of STAT3 in conferring anoikis resistance and enhanced invasive properties to NPC cells.

Activation of AMPK is neuroprotective in the oxidative stress by advanced glycosylation end products in human neural stem cells.

Advanced glycosylation end products (AGEs) formation is correlated with the pathogenesis of diabetic neuronal damage, but its links with oxidative stress are still not well understood. Metformin, one of the most widely used anti-diabetic drugs, exerts its effects in part by activation of AMP-activated protein kinase (AMPK). Once activated, AMPK regulates many pathways central to metabolism and energy balance including, glucose uptake, glycolysis and fatty acid oxidation. AMPK is also present in neurons, but its role remains unclear. Here, we show that AGE exposure decreases cell viability of human neural stem cells (hNSCs), and that the AMPK agonist metformin reverses this effect, via AMPK-dependent downregulation of RAGE levels. Importantly, hNSCs co-treated with metformin were significantly rescued from AGE-induced oxidative stress, as reflected by the normalization in levels of reactive oxygen species. In addition, compared to AGE-treated hNSCs, metformin co-treatment significantly reversed the activity and mRNA transcript level changes of SOD1/2 and Gpx. Furthermore, hNSCs exposed to AGEs had significantly lower mRNA levels among other components of normal cellular oxidative defenses (GSH, Catalase and HO-1), which were all rescued by co-treatment with metformin. This metformin-mediated protective effect on hNSCs for of both oxidative stress and oxidative defense genes by co-treatment with metformin was blocked by the addition of an AMPK antagonist (Compound C). These findings unveil the protective role of AMPK-dependent metformin signaling during AGE mediated oxidative stress in hNSCs, and suggests patients undergoing AGE-mediated neurodegeneration may benefit from the novel therapeutic use of metformin.

Spatiotemporal expression of foxo4, foxo6a, and foxo6b in the developing brain and retina are transcriptionally regulated by PI3K signaling in zebrafish.

The forkhead box subclass O (FoxO) family of proteins is a group of highly evolutionary conserved transcription factors that regulate various cellular processes and embryonic development. Dysregulated expressions of FOXO genes have been identified in numerous tumors and genetic disorders. The expression of FOXO/Foxo, particularly FOXO4/Foxo4 and FOXO6/Foxo6, in the developing nervous system has not been fully characterized. Here, we identified zebrafish foxo4, foxo6a, and foxo6b homologs and demonstrated that all three genes were expressed in the developing nervous system. foxo4, foxo6a, and foxo6b displayed ubiquitous expression in the brain and later in distinct brain tissues. In addition, these three genes were expressed in different retinal layers in a time-dependent manner. Furthermore, the mRNA expression of all three genes was significantly downregulated after treatment with a selective PI3-kinase (PI3K) inhibitor, LY294002. Our results suggest that foxo4, foxo6a, and foxo6b play important roles in the developing brain and retina and that the transcriptional levels of these genes are regulated by PI3-kinase signaling.

Metformin activation of AMPK-dependent pathways is neuroprotective in human neural stem cells against Amyloid-beta-induced mitochondrial dysfunction.

Alzheimer's disease (AD) is the general consequence of dementia and is diagnostic neuropathology by the cumulation of amyloid-beta (Aß) protein aggregates, which are thought to promote mitochondrial dysfunction processes leading to neurodegeneration. AMP-activated protein kinase (AMPK), a critical regulator of energy homeostasis and a major player in lipid and glucose metabolism, is potentially implied in the mitochondrial deficiency of AD. Metformin, one of the widespread used anti- metabolic disease drugs, use its actions in part by stimulation of AMPK. While the mechanisms of AD are well established, the neuronal roles for AMPK in AD are still not well understood. In the present study, human neural stem cells (hNSCs) exposed to Aß had significantly reduced cell viability, which correlated with decreased AMPK, neuroprotective genes (Bcl-2 and CREB) and mitochondria associated genes (PGC1α, NRF-1 and Tfam) expressions, as well as increased activation of caspase 3/9 activity and cytosolic cytochrome c. Co-treatment with metformin distinct abolished the Aß-caused actions in hNSCs. Metformin also significantly rescued hNSCs from Aß-mediated mitochondrial deficiency (lower D-loop level, mitochondrial mass, maximal respiratory function, COX activity, and mitochondrial membrane potential). Importantly, co-treatment with metformin significantly restored fragmented mitochondria to almost normal morphology in the hNSCs with Aß. These findings extend our understanding of the central role of AMPK in Aß-related neuronal impairment. Thus, a better understanding of AMPK might assist in both the recognition of its critical effects and the implementation of new therapeutic strategies in the treatment of AD.

Rosiglitazone activation of PPARγ-dependent pathways is neuroprotective in human neural stem cells against amyloid-beta-induced mitochondrial dysfunction and oxidative stress.

Neuronal cell impairment, such as that induced by amyloid-beta (Aß) protein, is a process with limited therapeutic interventions and often leads to long-term neurodegeneration common in disorders such as Alzheimer's disease. Interestingly, peroxisome proliferator-activated receptor gamma (PPARγ) is a ligand-activated nuclear receptor whose ligands control many physiological and pathologic processes, and may be neuroprotective. We hypothesized that rosiglitazone, a PPARγ agonist, would prevent Aß-mediated effects in human neural stem cells (hNSCs). Here, we show that rosiglitazone reverses, via PPARγ-dependent downregulation of caspase 3 and 9 activity, the Aß-mediated decreases in hNSC cell viability. In addition, Aß decreases hNSC messenger RNA (mRNA) levels of 2 neuroprotective factors (Bcl-2 and CREB), but co-treatment with rosiglitazone significantly rescues these effects. Rosiglitazone co-treated hNSCs also showed significantly increased mitochondrial function (reflected by levels of adenosine triphosphate and Mit mass), and PPARγ-dependent mRNA upregulation of PGC1α and mitochondrial genes (nuclear respiratory factor-1 and Tfam). Furthermore, hNSCs co-treated with rosiglitazone were significantly rescued from Aß-induced oxidative stress and correlates with reversal of the Aß-induced mRNA decrease in oxidative defense genes (superoxide dismutase 1, superoxide dismutase 2, and glutathione peroxidase 1). Taken together, these novel findings show that rosiglitazone-induced activation of PPARγ-dependent signaling rescues Aß-mediated toxicity in hNSCs and provide evidence supporting a neuroprotective role for PPARγ activating drugs in Aß-related diseases such as Alzheimer's disease.

Cardiac Fas-Dependent and Mitochondria-Dependent Apoptotic Pathways in a Transgenic Mouse Model of Huntington's Disease.

Huntington's disease is an autosomal dominant neurodegenerative disease caused by a CAG repeat expansion in the huntingtin gene. Heart disease is the second leading cause of death in patients with Huntington's disease. This study was to evaluate whether cardiac Fas-dependent and mitochondria-dependent apoptotic pathways are activated in transgenic mice with Huntington's disease. Sixteen Huntington's disease transgenic mice (HD) and sixteen wild-type (WT) littermates were studied at 10.5 weeks of age. The cardiac characteristics, myocardial architecture, and two major apoptotic pathways in the excised left ventricle from mice were measured by histopathological analysis, Western blotting, and TUNEL assays. The whole heart weight and the left ventricular weight decreased significantly in the HD group, as compared to the WT group. Abnormal myocardial architecture, enlarged interstitial spaces, and more cardiac TUNEL-positive cells were observed in the HD group. The key components of Fas-dependent apoptosis (TNF-alpha, TNFR1, Fas ligand, Fas death receptors, FADD, activated caspase-8, and activated caspase-3) and the key components of mitochondria-dependent apoptosis (Bax, Bax-to-Bcl-2 ratio, cytosolic cytochrome c, activated caspase-9, and activated caspase-3) increased significantly in the hearts of the HD group. Cardiac Fas-dependent and mitochondria-dependent apoptotic pathways were activated in transgenic mice with Huntington's disease, which might provide one of possible mechanisms to explain why patients with Huntington's disease will develop heart failure.

Rosiglitazone activation of PPARγ-dependent signaling is neuroprotective in mutant huntingtin expressing cells.

Peroxisome proliferator-activated receptor gamma (PPARγ) is a crucial transcription factor for neuroprotection in several brain diseases. Using a mouse model of Huntington's Disease (HD), we recently showed that PPARγ not only played a major function in preventing HD, but also oral intake of a PPARγ agonist (thiazolidinedione, TZD) significantly reduced the formation of mutant Huntingtin (mHtt) aggregates in the brain (e.g., cortex and striatum). The molecular mechanisms by which PPARγ exerts its HD neuroprotective effects remain unresolved. We investigated whether the PPARγ agonist (rosiglitazone) mediates neuroprotection in the mHtt expressing neuroblastoma cell line (N2A). Here we show that rosiglitazone upregulated the endogenous expression of PPARγ, its downstream target genes (including PGC1α, NRF-1 and Tfam) and mitochondrial function in mHtt expressing N2A cells. Rosiglitazone treatment also significantly reduced mHtt aggregates that included ubiquitin (Ub) and heat shock factor 1 (HSF1), as assessed by a filter-retardation assay, and increased the levels of the functional ubiquitin-proteasome system (UPS), HSF1 and heat shock protein 27/70 (HSP27/70) in N2A cells. Moreover, rosiglitazone treatment normalized endoplasmic reticulum (ER) stress sensors Bip, CHOP and ASK1, and significantly increased N2A cell survival. Taken together, these findings unveil new insights into the mechanisms by which activation of PPARγ signaling protects against the HD-mediated neuronal impairment. Further, our data also support the concept that PPARγ may be a novel therapeutic target for treating HD.