CHD4 is essential for transcriptional repression and lineage progression in B lymphopoiesis.

Cell lineage specification is a tightly regulated process that is dependent on appropriate expression of lineage and developmental stage-specific transcriptional programs. Here, we show that Chromodomain Helicase DNA-binding protein 4 (CHD4), a major ATPase/helicase subunit of Nucleosome Remodeling and Deacetylase Complexes (NuRD) in lymphocytes, is essential for specification of the early B cell lineage transcriptional program. In the absence of CHD4 in B cell progenitors in vivo, development of these cells is arrested at an early pro-B-like stage that is unresponsive to IL-7 receptor signaling and unable to efficiently complete V(D)J rearrangements at Igh loci. Our studies confirm that chromatin accessibility and transcription of thousands of gene loci are controlled dynamically by CHD4 during early B cell development. Strikingly, CHD4-deficient pro-B cells express transcripts of many non-B cell lineage genes, including genes that are characteristic of other hematopoietic lineages, neuronal cells, and the CNS, lung, pancreas, and other cell types. We conclude that CHD4 inhibits inappropriate transcription in pro-B cells. Together, our data demonstrate the importance of CHD4 in establishing and maintaining an appropriate transcriptome in early B lymphopoiesis via chromatin accessibility.

YY1 plays an essential role at all stages of B-cell differentiation.

Ying Yang 1 (YY1) is a ubiquitously expressed transcription factor shown to be essential for pro-B-cell development. However, the role of YY1 in other B-cell populations has never been investigated. Recent bioinformatics analysis data have implicated YY1 in the germinal center (GC) B-cell transcriptional program. In accord with this prediction, we demonstrated that deletion of YY1 by Cγ1-Cre completely prevented differentiation of GC B cells and plasma cells. To determine if YY1 was also required for the differentiation of other B-cell populations, we deleted YY1 with CD19-Cre and found that all peripheral B-cell subsets, including B1 B cells, require YY1 for their differentiation. Transitional 1 (T1) B cells were the most dependent upon YY1, being sensitive to even a half-dosage of YY1 and also to short-term YY1 deletion by tamoxifen-induced Cre. We show that YY1 exerts its effects, in part, by promoting B-cell survival and proliferation. ChIP-sequencing shows that YY1 predominantly binds to promoters, and pathway analysis of the genes that bind YY1 show enrichment in ribosomal functions, mitochondrial functions such as bioenergetics, and functions related to transcription such as mRNA splicing. By RNA-sequencing analysis of differentially expressed genes, we demonstrated that YY1 normally activates genes involved in mitochondrial bioenergetics, whereas it normally down-regulates genes involved in transcription, mRNA splicing, NF-κB signaling pathways, the AP-1 transcription factor network, chromatin remodeling, cytokine signaling pathways, cell adhesion, and cell proliferation. Our results show the crucial role that YY1 plays in regulating broad general processes throughout all stages of B-cell differentiation.

HDAC inhibition imparts beneficial transgenerational effects in Huntington's disease mice via altered DNA and histone methylation.

Increasing evidence has demonstrated that epigenetic factors can profoundly influence gene expression and, in turn, influence resistance or susceptibility to disease. Epigenetic drugs, such as histone deacetylase (HDAC) inhibitors, are finding their way into clinical practice, although their exact mechanisms of action are unclear. To identify mechanisms associated with HDAC inhibition, we performed microarray analysis on brain and muscle samples treated with the HDAC1/3-targeting inhibitor, HDACi 4b. Pathways analyses of microarray datasets implicate DNA methylation as significantly associated with HDAC inhibition. Further assessment of DNA methylation changes elicited by HDACi 4b in human fibroblasts from normal controls and patients with Huntington's disease (HD) using the Infinium HumanMethylation450 BeadChip revealed a limited, but overlapping, subset of methylated CpG sites that were altered by HDAC inhibition in both normal and HD cells. Among the altered loci of Y chromosome-linked genes, KDM5D, which encodes Lys (K)-specific demethylase 5D, showed increased methylation at several CpG sites in both normal and HD cells, as well as in DNA isolated from sperm from drug-treated male mice. Further, we demonstrate that first filial generation (F1) offspring from drug-treated male HD transgenic mice show significantly improved HD disease phenotypes compared with F1 offspring from vehicle-treated male HD transgenic mice, in association with increased Kdm5d expression, and decreased histone H3 Lys4 (K4) (H3K4) methylation in the CNS of male offspring. Additionally, we show that overexpression of Kdm5d in mutant HD striatal cells significantly improves metabolic deficits. These findings indicate that HDAC inhibitors can elicit transgenerational effects, via cross-talk between different epigenetic mechanisms, to have an impact on disease phenotypes in a beneficial manner.

Functional human antibody CDR fusions as long-acting therapeutic endocrine agonists.

On the basis of the 3D structure of a bovine antibody with a well-folded, ultralong complementarity-determining region (CDR), we have developed a versatile approach for generating human or humanized antibody agonists with excellent pharmacological properties. Using human growth hormone (hGH) and human leptin (hLeptin) as model proteins, we have demonstrated that functional human antibody CDR fusions can be efficiently engineered by grafting the native hormones into different CDRs of the humanized antibody Herceptin. The resulting Herceptin CDR fusion proteins were expressed in good yields in mammalian cells and retain comparable in vitro biological activity to the native hormones. Pharmacological studies in rodents indicated a 20- to 100-fold increase in plasma circulating half-life for these antibody agonists and significantly extended in vivo activities in the GH-deficient rat model and leptin-deficient obese mouse model for the hGH and hLeptin antibody fusions, respectively. These results illustrate the utility of antibody CDR fusions as a general and versatile strategy for generating long-acting protein therapeutics.

An Epitope-Specific Respiratory Syncytial Virus Vaccine Based on an Antibody Scaffold.

Respiratory syncytial virus (RSV) is a leading cause of lower respiratory tract infections in children. We have generated an epitope-specific RSV vaccine by grafting a neutralizing epitope (F-epitope) in its native conformation into an immunoglobulin scaffold. The resulting antibody fusion exhibited strong binding affinity to Motavizumab, an RSV neutralizing antibody, and effectively induced potent neutralizing antibodies in mice. This work illustrates the potential of the immunoglobulin molecule as a scaffold to present conformationally constrained B-cell epitopes.

Selective histone deacetylase (HDAC) inhibition imparts beneficial effects in Huntington's disease mice: implications for the ubiquitin-proteasomal and autophagy systems.

We previously demonstrated that the histone deacetylase (HDAC) inhibitor, 4b, which preferentially targets HDAC1 and HDAC3, ameliorates Huntington's disease (HD)-related phenotypes in different HD model systems. In the current study, we investigated extensive behavioral and biological effects of 4b in N171-82Q transgenic mice and further explored potential molecular mechanisms of 4b action. We found that 4b significantly prevented body weight loss, improved several parameters of motor function and ameliorated Huntingtin (Htt)-elicited cognitive decline in N171-82Q transgenic mice. Pathways analysis of microarray data from the mouse brain revealed gene networks involving post-translational modification, including protein phosphorylation and ubiquitination pathways, associated with 4b drug treatment. Using real-time qPCR analysis, we validated differential regulation of several genes in these pathways by 4b, including Ube2K, Ubqln, Ube2e3, Usp28 and Sumo2, as well as several other related genes. Additionally, 4b elicited increases in the expression of genes encoding components of the inhibitor of kappaB kinase (IKK) complex. IKK activation has been linked to phosphorylation, acetylation and clearance of the Htt protein by the proteasome and the lysosome, and accordingly, we found elevated levels of phosphorylated endogenous wild-type (wt) Htt protein at serine 16 and threonine 3, and increased AcK9/pS13/pS16 immunoreactivity in cortical samples from 4b-treated mice. We further show that HDAC inhibitors prevent the formation of nuclear Htt aggregates in the brains of N171-82Q mice. Our findings suggest that one mechanism of 4b action is associated with the modulation of the ubiquitin-proteasomal and autophagy pathways, which could affect accumulation, stability and/or clearance of important disease-related proteins, such as Htt.

Epigenetic changes at gene promoters in response to immune activation in utero.

Increasing evidence suggests that maternal infection increases the risk of psychiatric disorders, such as schizophrenia and autism in offspring. However, the molecular mechanisms associated with these effects are unclear. Here, we have studied epigenetic gene regulation in mice exposed to non-specific immune activation elicited by polyI:C injection to pregnant dams. Using Western blot analysis, we detected global hypoacetylation of histone H3, at lysine residues 9 and 14, and histone H4, at lysine residue 8, in the cortex from juvenile (∼24days of age) offspring exposed to polyI:C in utero, but not from adult (3months of age) offspring, which exhibit significant behavioral abnormalities. Accordingly, we detected robust deficits in the expression of genes associated with neuronal development, synaptic transmission and immune signaling in the cortex of polyI:C-exposed juvenile mice. In particular, we found that several genes in the glutamate receptor signaling pathway, including Gria1 and Slc17a7, showed decreases in promoter-specific histone acetylation, and corresponding gene expression deficits, in polyI:C-exposed offspring at both juvenile and adult ages. In contrast, the expression of these same genes, in addition to Disc1 and Ntrk3, was elevated in the hippocampus of juvenile mice, in concordance with elevated levels of promoter-specific histone acetylation. We suggest that these early epigenetic changes contribute to the delayed behavioral abnormalities that are observed in adult animals after exposure to polyI:C, and which resemble symptoms seen in schizophrenia and related disorders.

Histone deacetylase (HDAC) inhibitors targeting HDAC3 and HDAC1 ameliorate polyglutamine-elicited phenotypes in model systems of Huntington's disease.

We have previously demonstrated amelioration of Huntington's disease (HD)-related phenotypes in R6/2 transgenic mice in response to treatment with the novel histone deacetylase (HDAC) inhibitor 4b. Here we have measured the selectivity profiles of 4b and related compounds against class I and class II HDACs and have tested their ability to restore altered expression of genes related to HD pathology in mice and to rescue disease effects in cell culture and Drosophila models of HD. R6/2 transgenic and wild-type (wt) mice received daily injections of HDAC inhibitors for 3 days followed by real-time PCR analysis to detect expression differences for 13 HD-related genes. We find that HDACi 4b and 136, two compounds showing high potency for inhibiting HDAC3 were most effective in reversing the expression of genes relevant to HD, including Ppp1r1b, which encodes DARPP-32, a marker for medium spiny striatal neurons. In contrast, compounds targeting HDAC1 were less effective at correcting gene expression abnormalities in R6/2 transgenic mice, but did cause significant increases in the expression of selected genes. An additional panel of 4b-related compounds was tested in a Drosophila model of HD and in STHdhQ111 striatal cells to further distinguish HDAC selectivity. Significant improvement in huntingtin-elicited Drosophila eye neurodegeneration in the fly was observed in response to treatment with compounds targeting human HDAC1 and/or HDAC3. In STHdhQ111 striatal cells, the ability of HDAC inhibitors to improve huntingtin-elicited metabolic deficits correlated with the potency at inhibiting HDAC1 and HDAC3, although the IC50 values for HDAC1 inhibition were typically 10-fold higher than for inhibition of HDAC3. Assessment of HDAC protein localization in brain tissue by Western blot analysis revealed accumulation of HDAC1 and HDAC3 in the nucleus of HD transgenic mice compared to wt mice, with a concurrent decrease in cytoplasmic localization, suggesting that these HDACs contribute to a repressive chromatin environment in HD. No differences were detected in the localization of HDAC2, HDAC4 or HDAC7. These results suggest that inhibition of HDACs 1 and 3 can relieve HD-like phenotypes in model systems and that HDAC inhibitors targeting these isotypes might show therapeutic benefit in human HD.

Combined R-alpha-lipoic acid and acetyl-L-carnitine exerts efficient preventative effects in a cellular model of Parkinson's disease.

Mitochondrial dysfunction and oxidative damage are highly involved in the pathogenesis of Parkinson's disease (PD). Some mitochondrial antioxidants/nutrients that can improve mitochondrial function and/or attenuate oxidative damage have been implicated in PD therapy. However, few studies have evaluated the preventative effects of a combination of mitochondrial antioxidants/nutrients against PD, and even fewer have sought to optimize the doses of the combined agents. The present study examined the preventative effects of two mitochondrial antioxidant/nutrients, R-alpha-lipoic acid (LA) and acetyl-L-carnitine (ALC), in a chronic rotenone-induced cellular model of PD. We demonstrated that 4-week pretreatment with LA and/or ALC effectively protected SK-N-MC human neuroblastoma cells against rotenone-induced mitochondrial dysfunction, oxidative damage and accumulation of alpha-synuclein and ubiquitin. Most notably, we found that when combined, LA and ALC worked at 100-1000-fold lower concentrations than they did individually. We also found that pretreatment with combined LA and ALC increased mitochondrial biogenesis and decreased production of reactive oxygen species through the up-regulation of the peroxisome proliferator-activated receptor-gamma coactivator 1alpha as a possible underlying mechanism. This study provides important evidence that combining mitochondrial antioxidant/nutrients at optimal doses might be an effective and safe prevention strategy for PD.

A milk-based wolfberry preparation prevents prenatal stress-induced cognitive impairment of offspring rats, and inhibits oxidative damage and mitochondrial dysfunction in vitro.

Lycium barbarum (Fructus Lycii, Wolfberry, or Gouqi) belongs to the Solanaceae. The red-colored fruits of L. barbarum have been used for a long time as an ingredient in Chinese cuisine and brewing, and also in traditional Chinese herbal medicine for improving health. However, its effects on cognitive function have not been well studied. In the present study, prevention of a milk-based wolfberry preparation (WP) on cognitive dysfunction was tested in a prenatal stress model with rats and the antioxidant mechanism was tested by in vitro experiments. We found that prenatal stress caused a significant decrease in cognitive function (Morris water maze test) in female offspring. Pretreatment of the mother rats with WP significantly prevented the prenatal stress-induced cognitive dysfunction. In vitro studies showed that WP dose-dependently scavenged hydroxyl and superoxide radicals (determined by an electron spin resonance spectrometric assay), and inhibited FeCl(2)/ascorbic acid-induced dysfunction in brain tissue and tissue mitochondria, including increases in reactive oxygen species and lipid peroxidation and decreases in the activities of complex I, complex II, and glutamate cysteine ligase. These results suggest that dietary supplementation with WP may be an effective strategy for preventing the brain oxidative mitochondrial damage and cognitive dysfunction associated with prenatal stress.

Lipoamide protects retinal pigment epithelial cells from oxidative stress and mitochondrial dysfunction.

alpha-Lipoic acid (LA) has been widely studied as an agent for preventing and treating various diseases associated with oxidative disruption of mitochondrial functions. To investigate a related mitochondrial antioxidant, we compared the effects of lipoamide (LM), the neutral amide of LA, with LA for measures of oxidative damage and mitochondrial dysfunction in a human retinal pigment epithelial (RPE) cell line. Acrolein, a major component of cigarette smoke and a product of lipid peroxidation, was used to induce oxidative mitochondrial damage in RPE cells. Overall, using comparable concentrations, LM was more effective than LA at preventing acrolein-induced mitochondrial dysfunction and oxidative stress. Relative to LA, LM improved ATP levels, membrane potentials, and activities of mitochondrial complexes I, II, and V and dehydrogenases that had been decreased by acrolein exposure. LM reduced acrolein-induced oxidant generation, calcium levels, protein oxidation, and DNA damage to a greater degree than LA. And, total antioxidant capacity, glutathione content, glutathione S-transferase, and superoxide dismutase activities and expression of nuclear factor-E2-related factor 2 were increased by LM relative to LA. These results suggest that LM is a more potent mitochondrial-protective agent and antioxidant than LA in protecting RPE from oxidative damage.

High doses of nicotinamide prevent oxidative mitochondrial dysfunction in a cellular model and improve motor deficit in a Drosophila model of Parkinson's disease.

Nicotinamide, the principal form of niacin (vitamin B3), has been proposed to be neuroprotective in Parkinson's disease. However, the effects and mechanisms of nicotinamide on motor function in animals and on mitochondrial function in cellular systems have not been well studied. We hypothesized that niacin-derived NAD(P)H as antioxidants and enzyme cofactors could inhibit oxidative damage and improve mitochondrial function and thus protect neurodegeneration and improve motor function. In the present study, the effects of nicotinamide on mitochondrial function and oxidative stress were studied in a 1-methyl-4-phenylpyridinium (MPP(+))-induced cellular model of Parkinson's disease, and the effects of improving motor dysfunction were studied in an alpha-synuclein transgenic Drosophila Parkinson's model. Mitochondrial function was tested by measuring the activity of mitochondrial complex I and alpha-ketoglutarate dehydrogenase, and oxidative damage was tested by measuring reactive oxygen species, DNA damage (8-oxo-7,8-dihydro-2'-deoxyguanosine and Comet assay), and protein oxidation (protein carbonyls) levels. Nicotinamide at a relatively higher concentration, that is, 100-fold of the level in the cell culture medium (101 mg/L), significantly protected SK-N-MC human neuroblastoma cells from an MPP(+)-induced decrease in cell viability, complex I and alpha-ketoglutarate dehydrogenase activity, and an increase in oxidant generation, DNA damage, and protein oxidation. In the Drosophila model, nicotinamide at 15 and 30 mg/100 g diet significantly improved climbing ability. These results suggest that nutritional supplementation of nicotinamide at high doses decreases oxidative stress and improves mitochondrial and motor function in cellular and/or Drosophila models and may be an effective strategy for preventing and ameliorating Parkinson's disease.

Polyhydroxylated fullerene derivative C(60)(OH)(24) prevents mitochondrial dysfunction and oxidative damage in an MPP(+) -induced cellular model of Parkinson's disease.

To find effective agents for Parkinson's disease (PD) prevention and therapy, we examined the protective effects of the polyhydroxylated fullerene derivative C(60)(OH)(24) in a 1-methyl-4-phenylpyridinium (MPP(+)) -induced acute cellular PD model in human neuroblastoma cells and the free radical scavenging effects in this model with an electron spin resonance (ESR) spectrometer. Pretreatment with C(60)(OH)(24) at concentrations greater than 20 microM showed significant protective effects on MPP(+) -induced loss in cell viability, decreases in mitochondrial function (including mitochondrial membrane potential and activities of complex I and II), and increases in the levels of reactive oxygen species and oxidative damage to DNA and proteins. In addition, C(60)(OH)(24) acts as a phase 2 enzyme inducer to protect cells from MPP(+) -induced decreases in expression of nuclear factor-E2-related factor 2, expression and activity of gamma-glutamyl cysteine ligase and level of glutathione. The ESR study showed that C(60)(OH)(24) is a powerful radical scavenger for superoxide, hydroxyl, and lipid radicals. These data suggest that C(60)(OH)(24) is a mitochondrial protective antioxidant with direct radical scavenging activity and indirect antioxidant inducing activity.

Hydroxytyrosol protects retinal pigment epithelial cells from acrolein-induced oxidative stress and mitochondrial dysfunction.

Hydroxytyrosol (HTS) is a natural polyphenol abundant in olive oil. Increasing evidence indicates HTS has beneficial effect on human health for preventing various diseases. In the present study, we investigated the protective effects of HTS on acrolein-induced toxicity in human retinal pigment epithelial cell line, ARPE-19, a cellular model of smoking- and age-related macular degeneration. Acrolein, a major component of the gas phase cigarette smoke and also a product of lipid peroxidation in vivo, at 75 µmol/L for 24 h caused significant loss of cell viability, oxidative damage (increase in oxidant generation and oxidative damage to proteins and DNA, decrease in antioxidants and antioxidant enzymes, and also inactivation of the Keap1/Nrf2 pathway), and mitochondrial dysfunction (decrease in membrane potential, activities of mitochondrial complexes, viable mitochondria, oxygen consumption, and factors for mitochondrial biogenesis, and increase in calcium). Pre-treatment with HTS dose dependently and also time dependently protected the ARPE-19 cells from acrolein-induced oxidative damage and mitochondrial dysfunction. A short-term pre-treatment with HTS (48 h) required > 75 µmol/L for showing protection while a long-term pre-treatment (7 days) showed protective effect from 5 µmol/L on. The protective effect of HTS in this model was as potent as that of established mitochondria-targeting antioxidant nutrients. These results suggest that HTS is also a mitochondrial-targeting antioxidant nutrient and that dietary administration of HTS may be an effective measure in reducing and or preventing cigarette smoke-induced or age-related retinal pigment epithelial degeneration, such as age-associated macular degeneration.

Identification of a critical site in Wld(s): essential for Nmnat enzyme activity and axon-protective function.

The chimeric Wld(s) protein consisting of the N-terminal 70 amino acids of Ufd2 and the complete sequence of nicotinamide mononucleotide adenylyltransferase1 (Nmnat1), delays Wallerian degeneration in Wld(s) mice. Although Nmnat1 enzyme activity was showed to be critical for the function of Wld(s) protein, the expected phenotype was not observed in Nmnat1 transgenic mice. To further check whether Nmnat1 enzyme activity is involved, we aligned sequences of eukaryotic Nmnats, and found that Phe in helix A is highly conserved not only in various species, but also in different homologues. The Phe is a residue located near to the highly conserved GXFXPX(T/H)XXH motif and resides in the same helix as the last His of this conserved motif. To investigate the role of the conserved Phe in Nmnat activity, we made the point mutation of Phe. The Phe28 mutation of mouse Nmnat1 in Wld(s) completely abolished its Nmnat enzyme activity. To study the role of mutant Wld(s) in axon degeneration, herpes viruses were packaged to infect cultured SCGs. We found that the mutant Wld(s) failed to protect axon degeneration from morphological changes, microtubule integration and neurofilament degradation. Therefore, we have identified a Phe residue that critical for both enzyme activity of Nmnat and the axon-protective function of Wld(s), and further confirmed that Nmnat1 enzyme activity is required in Wld(s) function.

The Effects of Pharmacological Inhibition of Histone Deacetylase 3 (HDAC3) in Huntington's Disease Mice.

An important epigenetic modification in Huntington's disease (HD) research is histone acetylation, which is regulated by histone acetyltransferase and histone deacetylase (HDAC) enzymes. HDAC inhibitors have proven effective in HD model systems, and recent work is now focused on functional dissection of the individual HDAC enzymes in these effects. Histone deacetylase 3 (HDAC3), a member of the class I subfamily of HDACs, has previously been implicated in neuronal toxicity and huntingtin-induced cell death. Hence, we tested the effects of RGFP966 ((E)-N-(2-amino-4-fluorophenyl)-3-(1-cinnamyl-1H-pyrazol-4-yl)acrylamide), a benzamide-type HDAC inhibitor that selectively targets HDAC3, in the N171-82Q transgenic mouse model of HD. We found that RGFP966 at doses of 10 and 25 mg/kg improves motor deficits on rotarod and in open field exploration, accompanied by neuroprotective effects on striatal volume. In light of previous studies implicating HDAC3 in immune function, we measured gene expression changes for 84 immune-related genes elicited by RGFP966 using quantitative PCR arrays. RGFP966 treatment did not cause widespread changes in cytokine/chemokine gene expression patterns, but did significantly alter the striatal expression of macrophage migration inhibitory factor (Mif), a hormone immune modulator associated with glial cell activation, in N171-82Q transgenic mice, but not WT mice. Accordingly, RGFP966-treated mice showed decreased glial fibrillary acidic protein (GFAP) immunoreactivity, a marker of astrocyte activation, in the striatum of N171-82Q transgenic mice compared to vehicle-treated mice. These findings suggest that the beneficial actions of HDAC3 inhibition could be related, in part, with lowered Mif levels and its associated downstream effects.

Rational Design of Dual Agonist-Antibody Fusions as Long-acting Therapeutic Hormones.

Recent studies have suggested that modulation of two or more signaling pathways can achieve substantial weight loss and glycemic stability. We have developed an approach to the generation of bifunctional antibody agonists that activate leptin receptor and GLP-1 receptor. Leptin was fused into the complementarity determining region 3 loop of the light chain alone, or in combination with exendin-4 (EX4) fused at the N-terminus of the heavy chain of Herceptin. The antibody fusions exhibit similar or increased in vitro activities on their cognate receptors, but 50-100-fold longer circulating half-lives in rodents compared to the corresponding native peptides/proteins. The efficacy of the leptin/EX4 dual antibody fusion on weight loss, especially fat mass loss, was enhanced in ob/ob mice and DIO mice compared to the antibody fusion of either EX4 or leptin alone. This work demonstrates the versatility of this combinatorial fusion strategy for generating dual antibody agonists with long half-lives.

Lipoamide Acts as an Indirect Antioxidant by Simultaneously Stimulating Mitochondrial Biogenesis and Phase II Antioxidant Enzyme Systems in ARPE-19 Cells.

In our previous study, we found that pretreatment with lipoamide (LM) more effectively than alpha-lipoic acid (LA) protected retinal pigment epithelial (RPE) cells from the acrolein-induced damage. However, the reasons and mechanisms for the greater effect of LM than LA are unclear. We hypothesize that LM, rather than the more direct antioxidant LA, may act more as an indirect antioxidant. In the present study, we treated ARPE-19 cells with LA and LM and compared their effects on activation of mitochondrial biogenesis and induction of phase II enzyme systems. It is found that LM is more effective than LA on increasing mitochondrial biogenesis and inducing the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and its translocation to the nucleus, leading to an increase in expression or activity of phase II antioxidant enzymes (NQO-1, GST, GCL, catalase and Cu/Zn SOD). Further study demonstrated that mitochondrial biogenesis and phase II enzyme induction are closely coupled via energy requirements. These results suggest that LM, compared with the direct antioxidant LA, plays its protective effect on oxidative damage more as an indirect antioxidant to simultaneously stimulate mitochondrial biogenesis and induction of phase II antioxidant enzymes.

Disease Modifying Potential of Glatiramer Acetate in Huntington's Disease.

BACKGROUND: Deficiencies in brain-derived-neurotrophic-factor have been implicated in the pathogenesis of Huntington's disease (HD). OBJECTIVE: Glatiramer acetate, an FDA- approved drug used for the treatment of multiple sclerosis, has been shown to increase brain-derived-neurotrophic-factor levels in immune cells; hence, we investigated whether it could have similar effects in striatal cells. METHODS: Wild-type and HD striatal cells were treated with glatiramer acetate for 48 hrs. HD transgenic and wild-type mice were injected with glatiramer acetate (1.5 to 1.7 mg/mouse) for five days. These treatments were followed by protein measurements for brain-derived-neurotrophic-factor. RESULTS: Glatiramer acetate elicited concentration-dependent increases in brain-derived-neurotrophic-factor protein levels in wild-type and HD striatal cells and in striatal tissue from N171-82Q transgenic mice. Glatiramer acetate also improved metabolic activity of HD striatal cells, and significantly reduced the early hyperactivity phenotype exhibited by N171-82Q transgenic mice. CONCLUSIONS: These findings suggest that glatiramer acetate may represent a useful therapeutic approach for HD. The excellent safety and tolerability record of this compound makes it an ideal candidate for drug repurposing efforts.

Maternal docosahexaenoic acid feeding protects against impairment of learning and memory and oxidative stress in prenatally stressed rats: possible role of neuronal mitochondria metabolism.

AIMS: Docosahexaenoic acid (22:6n-3; DHA) is known to play a critical role in postnatal brain development. However, no study has been performed to investigate its preventive effect on prenatal stress-induced behavioral and molecular alterations in offspring. In the present study, rats were exposed to restraint stress on days 14-20 of pregnancy, three times a day, 2 hours each time; DHA was given at the doses of 100 and 300 mg/kg/day for two weeks. RESULTS: We showed that prenatal restraint stress caused (1) learning and memory impairment, (2) BDNF mRNA level decrease, (3) oxidative damage to proteins, (4) enhanced expression of nitric oxide synthase and apoptosis, and (5) abnormalities in mitochondrial metabolism that included changes in mitochondrial complexes I-V, and enhancement of expression of proteins involved in mitochondrial fusion/fission (Mfn-1, Mfn-2, Drp-1) and autophagy (Atg3, Atg7, Beclin-1, p-Akt, and p-mTOR) in the hippocampus of offspring. INNOVATION: Besides the well-known role in child brain development, we reported the novel finding of DHA in protecting prenatal stress-induced cognitive dysfunction involving the modulation of mitochondrial function and dynamics. CONCLUSION: Maternal feeding of DHA significantly prevented prenatal stress-induced impairment of learning and memory and normalized the biomarkers of oxidative damage, apoptosis, and mitochondrial metabolism in the hippocampus of both male and female offspring. These results suggest that maternal feeding of DHA exerts preventive effects on prenatal stress-induced brain dysfunction and that modulation of mitochondrial metabolism may play critical role in DHA protection.