Dysregulation of the Long Noncoding RNA X-Inactive-Specific Transcript Expression in Male Patients with Pulmonary Arterial Hypertension.

Pulmonary arterial hypertension (PAH) is a sex-biased disease with female gender as a significant risk factor. Recently, we reported that increased expression of the long noncoding RNA X-inactive-specific transcript (Xist), as induced by an intersectin-1s protein fragment with proliferative potential (EHITSN), may explain the sexual dimorphism of female pulmonary artery endothelial cells (ECs) and at least in part, the imbalance sex/ratio of PAH. Xist is essential for X-chromosome inactivation and dosage compensation of X-linked genes. Increased Xist expression was also detected in a subset of ECs and lung tissue samples of male patients with PAH. The role of different Xist expression levels in ECs of male patients with PAH (ECPAH) was studied in several lines of male ECPAH in conjunction with molecular, biochemical, morphologic, and functional approaches. Male ECPAH showed on average 10.3-fold increase in high Xist versus low Xist, a significant association between Xist levels and their proliferative potential, and a heterogeneous methylation of the Xist/Tsix locus. Interestingly, Xist up-regulation in male ECPAH decreases the expression of Klf2, via EHITSN interaction with EZH2, the catalytic subunit of the polycomb repressive complex 2. Moreover, the studies demonstrate that EHITSN-triggered p38/Elk1/c-Fos signaling is a pathologic mechanism central to ECPAH proliferation and the dynamic crosstalk with cell cycle regulatory proteins cyclin A1/cyclin D2 and Xist-EZH2-Klf2 interaction participate directly and differentially in establishing the proliferative profile of male ECPAH.

Prenol, but Not Vitamin C, of Fruit Binds to SARS-CoV-2 Spike S1 to Inhibit Viral Entry: Implications for COVID-19.

Fruit consumption may be beneficial for fighting infection. Although vitamin C is the celebrity component of fruit, its role in COVID-19 is unclear. Because spike S1 of SARS-CoV-2 binds to angiotensin-converting enzyme 2 (ACE2) on host cells to enter the cell and initiate COVID-19, using an α-screen-based assay, we screened vitamin C and other components of fruit for inhibiting the interaction between spike S1 and ACE2. We found that prenol, but neither vitamin C nor other major components of fruit (e.g., cyanidin and rutin), reduced the interaction between spike S1 and ACE2. Thermal shift assays indicated that prenol associated with spike S1, but not ACE2, and that vitamin C remained unable to do so. Although prenol inhibited the entry of pseudotyped SARS-CoV-2, but not vesicular stomatitis virus, into human ACE2-expressing HEK293 cells, vitamin C blocked the entry of pseudotyped vesicular stomatitis virus, not SARS-CoV-2, indicating the specificity of the effect. Prenol, but not vitamin C, decreased SARS-CoV-2 spike S1-induced activation of NF-κB and the expression of proinflammatory cytokines in human A549 lung cells. Moreover, prenol also decreased the expression of proinflammatory cytokines induced by spike S1 of N501Y, E484K, Omicron, and Delta variants of SARS-CoV-2. Finally, oral treatment with prenol reduced fever, decreased lung inflammation, enhanced heart function, and improved locomotor activities in SARS-CoV-2 spike S1-intoxicated mice. These results suggest that prenol and prenol-containing fruits, but not vitamin C, may be more beneficial for fighting against COVID-19.

Activation of PPARα Exhibits Therapeutic Efficacy in a Mouse Model of Juvenile Neuronal Ceroid Lipofuscinosis.

Juvenile neuronal ceroid lipofuscinosis (JNCL) is a fatal inherited neurodegenerative disease of children that occurs because of defective function of the lysosomal membrane glycoprotein CLN3. JNCL features glial activation and accumulation of autofluorescent storage material containing subunit c of mitochondrial ATP synthase (SCMAS), ultimately resulting into neuronal loss. Until now, no effective therapy is available for JNCL. This study underlines the possible therapeutic importance of gemfibrozil, an activator of peroxisome proliferator-activated receptor α (PPARα) and a lipid-lowering drug approved by the Food and Drug Administration in an animal model of JNCL. Oral gemfibrozil treatment reduced microglial and astroglial activation, attenuated neuroinflammation, restored the level of transcription factor EB (TFEB; the master regulator of lysosomal biogenesis), and decreased the accumulation of storage material SCMAS in somatosensory barrel field (SBF) cortex of Cln3Δex7/8 (Cln3ΔJNCL) mice of both sexes. Accordingly, gemfibrozil treatment also improved locomotor activities of Cln3ΔJNCL mice. While investigating the mechanism, we found marked loss of PPARα in the SBF cortex of Cln3ΔJNCL mice, which increased after gemfibrozil treatment. Oral gemfibrozil also stimulated the recruitment of PPARα to the Tfeb gene promoter in vivo in the SBF cortex of Cln3ΔJNCL mice, indicating increased transcription of Tfeb in the CNS by gemfibrozil treatment via PPARα. Moreover, disease pathologies aggravated in Cln3ΔJNCL mice lacking PPARα (Cln3ΔJNCLΔPPARα) and gemfibrozil remained unable to decrease SCMAS accumulation, reduce glial activation, and improve locomotor performance of Cln3ΔJNCLΔPPARα mice. These results suggest that activation of PPARα may be beneficial for JNCL and that gemfibrozil may be repurposed for the treatment of this incurable disease.SIGNIFICANCE STATEMENT Despite intense investigations, no effective therapy is available for JNCL, an incurable inherited lysosomal storage disorder. Here, we delineate that oral administration of gemfibrozil, a lipid-lowering drug, decreases glial inflammation, normalizes and/or upregulates TFEB, and reduces accumulation of autofluorescent storage material in SBF cortex to improve locomotor activities in Cln3Δex7/8 (Cln3ΔJNCL) mice. Aggravation of disease pathology in Cln3ΔJNCL mice lacking PPARα (Cln3ΔJNCLΔPPARα) and inability of gemfibrozil to decrease SCMAS accumulation, reduce glial activation, and improve locomotor performance of Cln3ΔJNCLΔPPARα mice delineates an important role of PPARα in this process. These studies highlight a new property of gemfibrozil and indicate its possible therapeutic use in JNCL patients.

Muscle-building supplement ß-hydroxy ß-methylbutyrate stimulates the maturation of oligodendroglial progenitor cells to oligodendrocytes.

Oligodendrocytes are the myelinating cells in the CNS and multiple sclerosis (MS) is a demyelinating disorder that is characterized by progressive loss of myelin. Although oligodendroglial progenitor cells (OPCs) should be differentiated into oligodendrocytes, for multiple reasons, OPCs fail to differentiate into oligodendrocytes in MS. Therefore, increasing the maturation of OPCs to oligodendrocytes may be of therapeutic benefit for MS. The ß-hydroxy ß-methylbutyrate (HMB) is a muscle-building supplement in humans and this study underlines the importance of HMB in stimulating the maturation of OPCs to oligodendrocytes. HMB treatment upregulated the expression of different maturation markers including PLP, MBP, and MOG in cultured OPCs. Double-label immunofluorescence followed by immunoblot analyses confirmed the upregulation of OPC maturation by HMB. While investigating mechanisms, we found that HMB increased the maturation of OPCs isolated from peroxisome proliferator-activated receptor ß-/- (PPARß-/- ) mice, but not PPARα-/- mice. Similarly, GW6471 (an antagonist of PPARα), but not GSK0660 (an antagonist of PPARß), inhibited HMB-induced maturation of OPCs. GW9662, a specific inhibitor of PPARγ, also could not inhibit HMB-mediated stimulation of OPC maturation. Furthermore, PPARα agonist GW7647, but neither PPARß agonist GW0742 nor PPARγ agonist GW1929, alone increased the maturation of OPCs. Finally, HMB treatment of OPCs led to the recruitment of PPARα, but neither PPARß nor PPARγ, to the PLP gene promoter. These results suggest that HMB stimulates the maturation of OPCs via PPARα and that HMB may have therapeutic prospects in remyelination.

Induction of IL-2 by interleukin-12 p40 homodimer and IL-12, but not IL-23, in microglia and macrophages: Implications for multiple sclerosis.

The level of IL-2 increases markedly in serum and central nervous system (CNS) of patients with multiple sclerosis (MS) and animals with experimental allergic encephalomyelitis (EAE). However, mechanisms by which IL-2 is induced under autoimmune demyelinating conditions are poorly understood. The present study underlines the importance of IL-12p40 homodimer (p402), the so-called biologically inactive molecule, in inducing the expression of IL-2 in mouse BV-2 microglial cells, primary mouse and human microglia, mouse peritoneal macrophages, RAW264.7 macrophages, and T cells. Interestingly, we found that p402 and IL-12p70 (IL-12), but not IL-23, dose-dependently induced the production of IL-2 and the expression of IL-2 mRNA in microglial cells. Similarly, p402 also induced the activation of IL-2 promoter in microglial cells and RAW264.7 cells. Among various stimuli tested, p402 was the most potent stimulus followed by IFN-γ, bacterial lipopolysaccharide, HIV-1 gp120, and IL-12 in inducing the activation of IL-2 promoter in microglial cells. Moreover, p402, but not IL-23, increased NFATc2 mRNA expression and the transcriptional activity of NFAT. Furthermore, induction of IL-2 mRNA expression by over-expression of p40, but not by p19, cDNA indicated that p40, but not p19, is responsible for the induction of IL-2 mRNA in microglia. Finally, by using primary microglia from IL to 12 receptor ß1 deficient (IL-12Rß1-/-) and IL-12 receptor ß2 deficient (IL-12Rß2-/-) mice, we demonstrate that p402 induces the expression of IL-2 via IL-12Rß1, but not IL-12Rß2. In experimental autoimmune encephalomyelitis, an animal model of MS, neutralization of p402 by mAb a3-1d led to decrease in clinical symptoms and reduction in IL-2 in T cells and microglia. These results delineate a new biological function of p402, which is missing in the so-called autoimmune cytokine IL-23, and raise the possibility of controlling increased IL-2 and the disease process of MS via neutralization of p402.

Treadmill workout activates PPARα in the hippocampus to upregulate ADAM10, decrease plaques and improve cognitive functions in 5XFAD mouse model of Alzheimer's disease.

Although liver is rich in peroxisome proliferator-activated receptor α (PPARα), recently we have described the presence of PPARα in hippocampus where it is involved in non-amyloidogenic metabolism of amyloid precursor protein (APP) via ADAM10, decreasing amyloid plaques and improving memory and learning. However, mechanisms to upregulate PPARα in vivo in the hippocampus are poorly understood. Regular exercise has multiple beneficial effects on human health and here, we describe the importance of regular mild treadmill exercise in upregulating PPARα in vivo in the hippocampus of 5XFAD mouse model of Alzheimer's disease. We also demonstrate that treadmill exercise remained unable to stimulate ADAM10, reduce plaque pathology and improve cognitive functions in 5XFADΔPPARα mice (5XFAD mice lacking PPARα). On the other hand, treadmill workout increased ADAM10, decreased plaque pathology and protected memory and learning in 5XFADΔPPARß mice (5XFAD mice lacking PPARß). Moreover, the other PPAR (PPARγ) also did not play any role in the transcription of ADAM10 in vivo in the hippocampus of treadmill exercised 5XFAD mice. These results underline an important role of PPARα in which treadmill exercise remains unable to exhibit neuroprotection in the hippocampus in the absence of PPARα.

Selective Inhibition of the Interaction between SARS-CoV-2 Spike S1 and ACE2 by SPIDAR Peptide Induces Anti-Inflammatory Therapeutic Responses.

Many patients with coronavirus disease 2019 in intensive care units suffer from cytokine storm. Although anti-inflammatory therapies are available to treat the problem, very often, these treatments cause immunosuppression. Because angiotensin-converting enzyme 2 (ACE2) on host cells serves as the receptor for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), to delineate a SARS-CoV-2-specific anti-inflammatory molecule, we designed a hexapeptide corresponding to the spike S1-interacting domain of ACE2 receptor (SPIDAR) that inhibited the expression of proinflammatory molecules in human A549 lung cells induced by pseudotyped SARS-CoV-2, but not vesicular stomatitis virus. Accordingly, wild-type (wt), but not mutated (m), SPIDAR inhibited SARS-CoV-2 spike S1-induced activation of NF-κB and expression of IL-6 and IL-1ß in human lung cells. However, wtSPIDAR remained unable to reduce activation of NF-κB and expression of proinflammatory molecules in lungs cells induced by TNF-α, HIV-1 Tat, and viral dsRNA mimic polyinosinic-polycytidylic acid, indicating the specificity of the effect. The wtSPIDAR, but not mutated SPIDAR, also hindered the association between ACE2 and spike S1 of SARS-CoV-2 and inhibited the entry of pseudotyped SARS-CoV-2, but not vesicular stomatitis virus, into human ACE2-expressing human embryonic kidney 293 cells. Moreover, intranasal treatment with wtSPIDAR, but not mutated SPIDAR, inhibited lung activation of NF-κB, protected lungs, reduced fever, improved heart function, and enhanced locomotor activities in SARS-CoV-2 spike S1-intoxicated mice. Therefore, selective targeting of SARS-CoV-2 spike S1-to-ACE2 interaction by wtSPIDAR may be beneficial for coronavirus disease 2019.

IL-12 p40 monomer is different from other IL-12 family members to selectively inhibit IL-12Rß1 internalization and suppress EAE.

Multiple sclerosis (MS) is the most common human demyelinating disease of the central nervous system. The IL-12 family of cytokines has four members, which are IL-12 (p40:p35), IL-23 (p40:p19), the p40 monomer (p40), and the p40 homodimer (p402). Since all four members contain p40 in different forms, it is important to use a specific monoclonal antibody (mAb) to characterize these molecules. Here, by using such mAbs, we describe selective loss of p40 in serum of MS patients as compared to healthy controls. Similarly, we also observed decrease in p40 and increase in IL-12, IL-23, and p402 in serum of mice with experimental autoimmune encephalomyelitis (EAE), an animal model of MS, as compared to control mice. Interestingly, weekly supplementation of mouse and human recombinant p40 ameliorated clinical symptoms and disease progression of EAE. On the other hand, IL-12, IL-23, and p402 did not exhibit such inhibitory effect. In addition to EAE, p40 also suppressed collagen-induced arthritis in mice. Using IL-12Rß1-/-, IL-12Rß2-/-, and IL-12Rß1+/-/IL-12Rß2-/- mice, we observed that p40 required IL-12Rß1, but not IL-12Rß2, to suppress EAE. Interestingly, p40 arrested IL-12-, IL-23-, or p402-mediated internalization of IL-12Rß1, but neither IL-12Rß2 nor IL-23R, protected regulatory T cells, and suppressed Th1 and Th17 biasness. These studies identify p40 as an anti-autoimmune cytokine with a biological role different from IL-12, IL-23, and p402 in which it attenuates autoimmune signaling via suppression of IL-12Rß1 internalization, which may be beneficial in patients with MS and other autoimmune disorders.

Upregulation of DJ-1 in Dopaminergic Neurons by a Physically-Modified Saline: Implications for Parkinson's Disease.

Parkinson's disease (PD) is the second most common neurodegenerative disorder in human and loss-of-functions DJ-1 mutations are associated with a familial form of early onset PD. Functionally, DJ-1 (PARK7), a neuroprotective protein, is known to support mitochondria and protect cells from oxidative stress. Mechanisms and agents by which the level of DJ-1 could be increased in the CNS are poorly described. RNS60 is a bioactive aqueous solution created by exposing normal saline to Taylor-Couette-Poiseuille flow under high oxygen pressure. Recently we have described neuroprotective, immunomodulatory and promyelinogenic properties of RNS60. Here we delineate that RNS60 is also capable of increasing the level of DJ-1 in mouse MN9D neuronal cells and primary dopaminergic neurons, highlighting another new neuroprotective effect of RNS60. While investigating the mechanism we found the presence of cAMP response element (CRE) in DJ-1 gene promoter and stimulation of CREB activation in neuronal cells by RNS60. Accordingly, RNS60 treatment increased the recruitment of CREB to the DJ-1 gene promoter in neuronal cells. Interestingly, RNS60 treatment also induced the enrollment of CREB-binding protein (CBP), but not the other histone acetyl transferase p300, to the promoter of DJ-1 gene. Moreover, knockdown of CREB by siRNA led to the inhibition of RNS60-mediated DJ-1 upregulation, indicating an important role of CREB in DJ-1 upregulation by RNS60. Together, these results indicate that RNS60 upregulates DJ-1 in neuronal cells via CREB-CBP pathway. It may be of benefit for PD and other neurodegenerative disorders.

Gemfibrozil Protects Dopaminergic Neurons in a Mouse Model of Parkinson's Disease via PPARα-Dependent Astrocytic GDNF Pathway.

Parkinson's disease (PD) is the most common neurodegenerative movement disorder in humans. Despite intense investigations, effective therapies are not yet available to halt the progression of PD. Gemfibrozil, a Food and Drug Administration-approved lipid-lowering drug, is known to decrease the risk of coronary heart disease by increasing the level of high-density lipoprotein cholesterol and decreasing the level of low-density lipoprotein cholesterol. This study underlines the importance of gemfibrozil in protecting dopaminergic neurons in an animal model of PD. Oral administration of the human equivalent dose of gemfibrozil protected tyrosine hydroxylase (TH)-positive dopaminergic neurons in the substantia nigra pars compacta and TH fibers in the striatum of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-insulted mice of both sexes. Accordingly, gemfibrozil also normalized striatal neurotransmitters and improved locomotor activities in MPTP-intoxicated mice. Gemfibrozil-mediated protection of the nigrostriatal and locomotor activities in WT but not PPARα-/- mice from MPTP intoxication suggests that gemfibrozil needs the involvement of peroxisome proliferator-activated receptor α (PPARα) in protecting dopaminergic neurons. While investigating further mechanisms, we found that gemfibrozil stimulated the transcription of glial-derived neurotrophic factor (GDNF) gene in astrocytes via PPARα and that gemfibrozil protected nigral neurons, normalized striatal fibers and neurotransmitters, and improved locomotor activities in MPTP-intoxicated Gfafcre mice, but not GdnfΔastro mice lacking GDNF in astrocytes. These findings highlight the importance of the PPARα-dependent astroglial GDNF pathway in gemfibrozil-mediated protection of dopaminergic neurons in an animal model of PD and suggest the possible therapeutic use of gemfibrozil in PD patients.SIGNIFICANCE STATEMENT Increasing the level of glial cell-derived neurotrophic factor (GDNF) in the brain is important for the protection of dopamine neurons in Parkinson's disease (PD). Although gene manipulation and GDNF protein infusion into the brain are available options, it seems from the therapeutic angle that the best option would be to stimulate/induce the production of GDNF in vivo in the brain of PD patients. Here, we delineate that gemfibrozil, a lipid-lowering drug, stimulates GDNF in astrocytes via peroxisome proliferator-activated receptor α (PPARα). Moreover, gemfibrozil protected nigral neurons, normalized striatal fibers and neurotransmitters, and improved locomotor activities from MPTP toxicity via the PPARα-dependent astroglial GDNF pathway. These studies highlight a new property of gemfibrozil and suggest its possible therapeutic use in PD patients.

Aspirin binds to PPARα to stimulate hippocampal plasticity and protect memory.

Despite its long history, until now, no receptor has been identified for aspirin, one of the most widely used medicines worldwide. Here we report that peroxisome proliferator-activated receptor alpha (PPARα), a nuclear hormone receptor involved in fatty acid metabolism, serves as a receptor of aspirin. Detailed proteomic analyses including cheminformatics, thermal shift assays, and TR-FRET revealed that aspirin, but not other structural homologs, acts as a PPARα ligand through direct binding at the Tyr314 residue of the PPARα ligand-binding domain. On binding to PPARα, aspirin stimulated hippocampal plasticity via transcriptional activation of cAMP response element-binding protein (CREB). Finally, hippocampus-dependent behavioral analyses, calcium influx assays in hippocampal slices and quantification of dendritic spines demonstrated that low-dose aspirin treatment improved hippocampal plasticity and memory in FAD5X mice, but not in FAD5X/Ppara-null mice. These findings highlight a property of aspirin: stimulating hippocampal plasticity via direct interaction with PPARα.

Aspirin Induces Lysosomal Biogenesis and Attenuates Amyloid Plaque Pathology in a Mouse Model of Alzheimer's Disease via PPARα.

Lysosomes play a central role in cellular homeostasis by regulating the cellular degradative machinery. Because aberrant lysosomal function has been associated with multiple lysosomal storage and neurodegenerative disorders, enhancement of lysosomal clearance has emerged as an attractive therapeutic strategy. Transcription factor EB (TFEB) is known as a master regulator of lysosomal biogenesis and, here, we reveal that aspirin, one of the most widely used medications in the world, upregulates TFEB and increases lysosomal biogenesis in brain cells. Interestingly, aspirin induced the activation of peroxisome proliferator-activated receptor alpha (PPARα) and stimulated the transcription of Tfeb via PPARα. Finally, oral administration of low-dose aspirin decreased amyloid plaque pathology in both male and female 5X familial Alzheimer's disease (5XFAD) mice in a PPARα-dependent fashion. This study reveals a new function of aspirin in stimulating lysosomal biogenesis via PPARα and suggests that low-dose aspirin may be used in lowering storage materials in Alzheimer's disease and lysosomal storage disorders.SIGNIFICANCE STATEMENT Developing drugs for the reduction of amyloid ß containing senile plaques, one of the pathological hallmarks of Alzheimer's disease (AD), is an important area of research. Aspirin, one of the most widely used medications in the world, activates peroxisome proliferator-activated receptor alpha (PPARα) to upregulate transcription factor EB and increase lysosomal biogenesis in brain cells. Accordingly, low-dose aspirin decreases cerebral plaque load in a mouse model of Alzheimer's disease via PPARα. These results reveal a new mode of action of aspirin that may be beneficial for AD and lysosomal storage disorders.

Cinnamic acid activates PPARα to stimulate Lysosomal biogenesis and lower Amyloid plaque pathology in an Alzheimer's disease mouse model.

The response of the lysosomes, the waste clearance machinery of the cell, to different environmental stimuli is coordinated by a gene network with a master regulator Transcription factor EB (TFEB) at the core. Disruption of multiple facets of the lysosomal and autophagic network has been linked to various neurodegenerative and lysosomal storage disorders, making TFEB an attractive therapeutic target to rescue or augment lysosomal function under pathological scenario. In this study, we demonstrate that cinnamic acid, a naturally occurring plant-based product, induces lysosomal biogenesis in mouse primary brain cells via upregulation of TFEB. We delineate that cinnamic acid activates the nuclear hormone receptor PPARα to transcriptionally upregulate TFEB and stimulate lysosomal biogenesis. Moreover, using in-silico and biochemical approaches we established that cinnamic acid serves as a potent ligand for peroxisome proliferator-activated receptor α (PPARα). Finally, cinnamic acid treatment in male and female 5× Familial Alzheimer's disease (5XFAD) mice remarkably reduced cerebral amyloid-beta plaque burden and improved memory via PPARα. Therefore, stimulation of lysosomal biogenesis by cinnamic acid may have therapeutic implications for treatment of Alzheimer's disease and other lysosomal disorders originating from accumulation of toxic protein aggregates.

Cinnamic Acid Protects the Nigrostriatum in a Mouse Model of Parkinson's Disease via Peroxisome Proliferator-Activated Receptorα.

Parkinson's disease (PD) is the second most common devastating human neurodegenerative disorder and despite intense investigation, no effective therapy is available for PD. Cinnamic acid, a naturally occurring aromatic fatty acid of low toxicity, is a precursor for the synthesis of a huge number of plant substances. This study highlights the neuroprotective effect of cinnamic acid in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD. Oral administration of cinnamic acid protected tyrosine hydroxylase (TH)-positive dopaminergic neurons in the substantia nigra pars compacta (SNpc) and TH fibers in the striatum of MPTP-insulted mice. Accordingly, oral cinnamic acid also normalized striatal neurotransmitters and improved locomotor activities in MPTP-intoxicated mice. While investigating mechanisms, we found that cinnamic acid induced the activation of peroxisome proliferator-activated receptor α (PPARα), but not PPARß, in primary mouse astrocytes. Cinnamic acid mediated protection of the nigrostriatal system and locomotor activities in WT and PPARß (-/-), but not PPARα (-/-) mice from MPTP intoxication suggests that cinnamic acid requires the involvement of PPARα in protecting dopaminergic neurons in this model of PD. This study delineates a new function of cinnamic acid in protecting dopaminergic neurons via PPARα that could be beneficial for PD.

Identification and characterization of PPARα ligands in the hippocampus.

Peroxisome proliferator-activated receptor-α (PPARα) regulates hepatic fatty acid catabolism and mediates the metabolic response to starvation. Recently we found that PPARα is constitutively activated in nuclei of hippocampal neurons and controls plasticity via direct transcriptional activation of CREB. Here we report the discovery of three endogenous PPARα ligands-3-hydroxy-(2,2)-dimethyl butyrate, hexadecanamide, and 9-octadecenamide-in mouse brain hippocampus. Mass spectrometric detection of these compounds in mouse hippocampal nuclear extracts, in silico interaction studies, time-resolved FRET analyses, and thermal shift assay results clearly indicated that these three compounds served as ligands of PPARα. Site-directed mutagenesis studies further revealed that PPARα Y464 and Y314 are involved in binding these hippocampal ligands. Moreover, these ligands activated PPARα and upregulated the synaptic function of hippocampal neurons. These results highlight the discovery of hippocampal ligands of PPARα capable of modulating synaptic functions.

Upregulation of Myelin Gene Expression by a Physically-Modified Saline via Phosphatidylinositol 3-Kinase-Mediated Activation of CREB: Implications for Multiple Sclerosis.

An increase in central nervous system (CNS) remyelination and a decrease in CNS inflammation are important steps to halt the progression of multiple sclerosis (MS). RNS60 is a bioactive aqueous solution generated by subjecting normal saline to Taylor-Couette-Poiseuille flow under elevated oxygen pressure. Recently we have demonstrated that RNS60 exhibits anti-inflammatory properties. Here, we describe promyelinating property of RNS60. RNS60, but not normal saline (NS), RNS10.3 (TCP-modified saline without excess oxygen) or PNS60 (saline containing excess oxygen without TCP modification), stimulated the expression of myelin-specific genes and proteins (myelin basic protein, MBP; myelin oligodendrocyte glycoprotein, MOG and proteolipid protein, PLP) in primary mouse oligodendroglia and mixed glial cells. While investigating the mechanisms, we found that RNS60 treatment induced the activation of cAMP response element binding protein (CREB) in oligodendrocytes, ultimately leading to the recruitment of CREB to the promoters of myelin-specific genes. Furthermore, activation of type 1A p110ß/α, but not type 1B p110γ, phosphatidylinositol-3 (PI-3) kinase by RNS60 together with abrogation of RNS60-mediated activation of CREB and upregulation of myelin genes by LY294002 (a specific inhibitor of PI-3 kinase) suggest that RNS60 upregulates the activation of CREB and the expression of myelin-specific molecules in oligodendrocytes via activation of PI3 kinase. These results highlight a novel promyelinating property of RNS60, which may be of benefit for MS and other demyelinating disorders.

Activation of peroxisome proliferator-activated receptor α induces lysosomal biogenesis in brain cells: implications for lysosomal storage disorders.

Lysosomes are ubiquitous membrane-enclosed organelles filled with an acidic interior and are central to the autophagic, endocytic, or phagocytic pathway. In contrast to its classical function as the waste management machinery, lysosomes are now considered to be an integral part of various cellular signaling processes. The diverse functionality of this single organelle requires a very complex and coordinated regulation of its activity with transcription factor EB (TFEB), a master regulator of lysosomal biogenesis, at its core. However, mechanisms by which TFEB is regulated are poorly understood. This study demonstrates that gemfibrozil, an agonist of peroxisome proliferator-activated receptor (PPAR) α, alone and in conjunction with all-trans-retinoic acid is capable of enhancing TFEB in brain cells. We also observed that PPARα, but not PPARß and PPARγ, is involved in gemfibrozil-mediated up-regulation of TFEB. Reporter assay and chromatin immunoprecipitation studies confirmed the recruitment of retinoid X receptor α, PPARα, and PGC1α on the PPAR-binding site on the Tfeb promoter as well. Subsequently, the drug-mediated induction of TFEB caused an increase in lysosomal protein and the lysosomal abundance in cell. Collectively, this study reinforces the link between lysosomal biogenesis and lipid metabolism with TFEB at the crossroads. Furthermore, gemfibrozil may be of therapeutic value in the treatment of lysosomal storage disorders in which autophagy-lysosome pathway plays an important role.

Sodium Benzoate, a Metabolite of Cinnamon and a Food Additive, Upregulates Ciliary Neurotrophic Factor in Astrocytes and Oligodendrocytes.

Ciliary neurotrophic factor (CNTF) is a promyelinating trophic factor that plays an important role in multiple sclerosis (MS). However, mechanisms by which CNTF expression could be increased in the brain are poorly understood. Recently we have discovered anti-inflammatory and immunomodulatory activities of sodium benzoate (NaB), a metabolite of cinnamon and a widely-used food additive. Here, we delineate that NaB is also capable of increasing the mRNA and protein expression of CNTF in primary mouse astrocytes and oligodendrocytes and primary human astrocytes. Accordingly, oral administration of NaB and cinnamon led to the upregulation of astroglial and oligodendroglial CNTF in vivo in mouse brain. Induction of experimental allergic encephalomyelitis, an animal model of MS, reduced the level of CNTF in the brain, which was restored by oral administration of cinnamon. While investigating underlying mechanisms, we observed that NaB induced the activation of protein kinase A (PKA) and H-89, an inhibitor of PKA, abrogated NaB-induced expression of CNTF. The activation of cAMP response element binding (CREB) protein by NaB, the recruitment of CREB and CREB-binding protein to the CNTF promoter by NaB and the abrogation of NaB-induced expression of CNTF in astrocytes by siRNA knockdown of CREB suggest that NaB increases the expression of CNTF via the activation of CREB. These results highlight a novel myelinogenic property of NaB and cinnamon, which may be of benefit for MS and other demyelinating disorders.