Metformin-induced oxidative stress inhibits LNCaP prostate cancer cell survival.

BACKGROUND: Preclinical and clinical studies over the past several decades have indicated the potential value of metformin, a widely utilized treatment for Type 2 diabetes, in prostate cancer therapy. Notably, these studies demonstrated metformin's pleiotropic effects on several molecular and metabolic pathways, such as androgen signaling, cell cycle, and cellular bioenergetics. In this study we investigated the role of metformin in regulating intracellular redox status and cell survival in LNCaP prostate cancer cells. METHODS AND RESULTS: The cytotoxic effects of metformin with or without the presence of SBI0206965 (AMPK inhibitor) on LNCaP cells were determined using MTT and trypan blue exclusion assays. Seahorse XP extracellular analysis, Liquid Chromatography/ Mass Spectrophotometry (LC/MS), and 2,7- and Dichlorofluoresin diacetate (DCFDA) assay were used to assess the effects of metformin on cellular bioenergetics, redox status, and redox-related metabolites. mRNA expression and protein concentration of redox-related enzymes were measured using Real Time-qPCR and ELISA assay, respectively. Independently of AMP-activated protein kinase, metformin exhibited a dose- and time-dependent inhibition of LNCaP cell survival, a response mitigated by glutathione or N-acetylcysteine (ROS scavengers) treatment. Notably, these findings were concomitant with a decline in ATP levels and the inhibition of oxidative phosphorylation. The results further indicated metformin's induction of reactive oxygen species, which significantly decreased glutathione levels and the ratio of reduced to oxidized glutathione, as well as the transsulfuration metabolite, cystathionine. Consistent with an induction of oxidative stress condition, metformin increased mRNA levels of the master redox transcription factor Nrf-2 (nuclear factor erythroid-derived 2-like), as well as transsulfuration enzymes cystathionine beta-synthase and cystathionase and GSH synthesis enzymes γ-glutamylcysteine synthetase and glutathione synthetase. CONCLUSION: Our findings highlight multiple mechanisms by which metformin-induced formation of reactive oxygen species may contribute to its efficacy in prostate cancer treatment, including promotion of oxidative stress, Nrf2 activation, and modulation of redox-related pathways, leading to its anti-survival action.

Combination microRNA-based cellular reprogramming with paclitaxel enhances therapeutic efficacy in a relapsed and multidrug-resistant model of epithelial ovarian cancer.

Most advanced-stage ovarian cancer patients, including those with epithelial ovarian cancer (EOC), develop recurrent disease and acquisition of resistance to chemotherapy, leading to limited treatment options. Decrease in Let7b miRNA levels in clinical ovarian cancer has been associated with chemoresistance, increased proliferation, invasion, and relapse in EOC. We have established a murine EOC relapsed model by administering paclitaxel (PTX) and stopping therapy to allow for tumor regrowth. Global microRNA profiling in the relapsed tumor showed significant downregulation of Let7b relative to untreated tumors. Here, we report the use of hyaluronic acid (HA)-based nanoparticle formulation that can deliver Let7b miRNA mimic to tumor cells and achieve cellular programming both in vitro and in vivo. We demonstrate that a therapeutic combination of Let7b miRNA and PTX leads to significant improvement in anti-tumor efficacy in the relapsed model of EOC. We further demonstrate that the combination therapy is safe for repeated administration. This novel approach of cellular reprogramming of tumor cells using clinically relevant miRNA mimic in combination with chemotherapy could enable more effective therapeutic outcomes for patients with advanced-stage relapsed EOC.

Decreased cortical Nrf2 gene expression in autism and its relationship to thiol and cobalamin status.

Nuclear factor erythroid 2-related factor 2 (Nrf2) promotes expression of a large number of antioxidant genes and multiple studies have described oxidative stress and impaired methylation in autism spectrum disorder (ASD), including decreased brain levels of methylcobalamin(III) (MeCbl). Here we report decreased expression of the Nrf2 gene (NFE2L2) in frontal cortex of ASD subjects, as well as differences in other genes involved in redox homeostasis. In pooled control and ASD correlation analyses, hydroxocobalamin(III) (OHCbl) was inversely correlated with NFE2L2 expression, while MeCbl and total cobalamin abundance were positively correlated with NFE2L2 expression. Levels of methionine, S-adenosylmethionine (SAM), S-adenosylhomocysteine (SAH) and cystathionine were positively correlated with NFE2L2 expression, while homocysteine (HCY) was negatively correlated. The relationship between Nrf2 activity and cobalamin was further supported by a bioinformatics-based comparison of cobalamin levels in different tissues with expression of a panel of 40 Nrf2-regulated genes, which yielded a strong correlation. Lastly, Nrf2-regulated gene expression was also correlated with expression of intracellular cobalamin trafficking and processing genes, such as MMADHC and MTRR. These findings highlight a previously unrecognized relationship between the antioxidant-promoting role of Nrf2 and cobalamin status, which is dysfunctional in ASD.

Alterations in Gene Expression of Renin-Angiotensin System Components and Related Proteins in Colorectal Cancer.

MATERIALS AND METHODS: Quantitative expression of the RNA of these 17 genes in normal and cancerous tissues obtained using chip arrays from the public functional genomics data repository, Gene Expression Omnibus (GEO) application, was compared statistically. RESULTS: Expression of four genes, AGT (angiotensinogen), ENPEP (aminopeptidase A) MME (neprilysin), and PREP (prolyl endopeptidase), was significantly upregulated in CRC specimens. Expression of REN (renin), THOP (thimet oligopeptidase), NLN (neurolysin), PRCP (prolyl carboxypeptidase), ANPEP (aminopeptidase N), and MAS1 (Mas receptor) was downregulated in CRC specimens. CONCLUSIONS: Presuming gene expression parallel protein expression, these results suggest that increased production of the angiotensinogen precursor of angiotensin (ANG) peptides, with the reduction of the enzymes that metabolize it to ANG II, can lead to accumulation of angiotensinogen in CRC tissues. Downregulation of THOP, NLN, PRCP, and MAS1 gene expression, whose proteins contribute to the ACE2/ANG 1-7/Mas axis, suggests that reduced activity of this RAS branch could be permissive for oncogenicity. Components of the RAS may be potential therapeutic targets for treatment of CRC.

Nutritional interventions for autism spectrum disorder.

Autism spectrum disorder (ASD) is an increasingly prevalent neurodevelopmental disorder with considerable clinical heterogeneity. With no cure for the disorder, treatments commonly center around speech and behavioral therapies to improve the characteristic social, behavioral, and communicative symptoms of ASD. Gastrointestinal disturbances are commonly encountered comorbidities that are thought to be not only another symptom of ASD but to also play an active role in modulating the expression of social and behavioral symptoms. Therefore, nutritional interventions are used by a majority of those with ASD both with and without clinical supervision to alleviate gastrointestinal and behavioral symptoms. Despite a considerable interest in dietary interventions, no consensus exists regarding optimal nutritional therapy. Thus, patients and physicians are left to choose from a myriad of dietary protocols. This review, summarizes the state of the current clinical and experimental literature on nutritional interventions for ASD, including gluten-free and casein-free, ketogenic, and specific carbohydrate diets, as well as probiotics, polyunsaturated fatty acids, and dietary supplements (vitamins A, C, B6, and B12; magnesium and folate).

Methylation-related metabolic effects of D4 dopamine receptor expression and activation.

D4 dopamine receptor (D4R) activation uniquely promotes methylation of plasma membrane phospholipids, utilizing folate-derived methyl groups provided by methionine synthase (MS). We evaluated the impact of D4R expression on folate-dependent phospholipid methylation (PLM) and MS activity, as well as cellular redox and methylation status, in transfected CHO cells expressing human D4R variants containing 2, 4, or 7 exon III repeats (D4.2R, D4.4R, D4.7R). Dopamine had no effect in non-transfected CHO cells, but increased PLM to a similar extent for both D4.2R- and D4.4R-expressing cells, while the maximal increase was for D4.7R was significantly lower. D4R expression in CHO cells decreased basal MS activity for all receptor subtypes and conferred dopamine-sensitive MS activity, which was greater with a higher number of repeats. Consistent with decreased MS activity, D4R expression decreased basal levels of methylation cycle intermediates methionine, S-adenosylmethionine (SAM), and S-adenosylhomocysteine (SAH), as well as cysteine and glutathione (GSH). Conversely, dopamine stimulation increased GSH, SAM, and the SAM/SAH ratio, which was associated with a more than 2-fold increase in global DNA methylation. Our findings illustrate a profound influence of D4R expression and activation on MS activity, coupled with the ability of dopamine to modulate cellular redox and methylation status. These previously unrecognized signaling activities of the D4R provide a unique link between neurotransmission and metabolism.

Cross-Generational Transmission of Early Life Stress Effects on HPA Regulators and Bdnf Are Mediated by Sex, Lineage, and Upbringing.

Early life stress (ELS) is a potent developmental disruptor and increases the risk for psychopathology. Various forms of ELS have been studied in both humans and rodents, and have been implicated in altered DNA methylation, gene transcription, stress hormone levels, and behavior. Although recent studies have focused on stress-induced epigenetic changes, the extent to which ELS alters HPA axis function and stress responsivity across generations, whether these effects are sex-specific, and how lineage interacts with upbringing to impact these effects, remain unclear. To address these points, two generations of rodents were utilized, with the first generation subjected to ELS via maternal separation, and the second to a balanced cross-fostering paradigm. We hypothesized that ELS would disrupt normative development in both generations, manifesting as altered methylation and expression of genes associated with stress signaling pathways (Nr3c1, Nr3c2, and Bdnf), blunted corticosterone (CORT), and anxiety-like behaviors. Additionally, we expected deficits in the second generation to be modulated by caretaking environment and for the pattern of results to differ between the sexes. Results suggest that direct exposure to ELS leads to sex-specific effects on gene regulation and HPA functioning in adulthood, with maternal separation leading to increases in Bdnf methylation in both sexes, decreases in Bdnf expression in females, and decreases in Nr3c1 methylation in males, as well as blunted CORT and less anxiety-like behavior in females. These alterations converged with caretaking to impart perturbations upon the subsequent generation. Across sex, ELS lineage led to decreased methylation of Nr3c1, and increased methylation of Bdnf. In fostered animals, upbringing by a previously stressed mother interacted with offspring lineage to impact methylation of Nr3c1 and Bdnf. Upbringing was also implicated in altered anxiety-like behavior in males, and baseline CORT levels in females. Such effects may correspond with observed alterations in maternal behavior across groups. In conclusion, ELS conferred enduring sex-specific alterations, both first-hand and trans-generationally via lineage and upbringing. Importantly, lineage of cross-fostered pups was sufficient to normalize or disturb maternal behavior of foster-dams, an observation requiring further elucidation. These results have implications for multi-generational effects of ELS in humans and may motivate early interventions.

Alterations in DNA Methylation Status Associated with Gulf War Illness.

Gulf War Illness (GWI) affects about 25% of Persian Gulf veterans with a cluster of chronic symptoms, including immune dysfunction and neurological issues. Recent studies implicate gene expression changes in immune function to be associated with GWI. Since DNA methylation can regulate such changes in gene expression, and disruption of DNA methylation pattern is implicated in various immune and neurological diseases, we aimed to study the DNA methylation patterns in peripheral blood mononuclear cells from GWI patients. Global DNA methylation levels were similar in GWI patients and controls. However, the genome-wide microarray technology detected 10,767 differentially methylated CpG sites across gene regulatory elements and within coding regions. Approximately 88% of them were hypermethylated in GWI patients. The separate analysis found 776 differentially methylated gene promoters (DMP), which were predominantly hypermethylated. Pyrosequencing validation confirmed microarray results. Functional analysis revealed that majority of the DMPs belonged to genes responsible for metabolism and immune system. This is the first pilot human study characterizing genome-wide epigenetic changes associated with GWI. It suggests a significant contribution of epigenetic dysfunction in GWI. Moreover, it supports the dysregulation of immune function in GWI. Lastly, it suggests studies with the larger cohort to validate our findings.

Brain iron loading impairs DNA methylation and alters GABAergic function in mice.

Iron deficiency is closely associated with altered GABA metabolism and affective behavior. While mutation in the hemochromatosis ( HFE) gene disrupts iron homeostasis and promotes oxidative stress that increases the risk of neurodegeneration, it is largely unknown whether HFE mutation modifies GABAergic homeostasis and emotional behavior. The goal of our study was to investigate the impact of HFE on GABAergic neurochemistry and redox-epigenetic regulation in the brain using H67D HFE-mutant mice that recapitulates the H63D-HFE mutation in humans. H67D mice displayed elevated redox-active iron levels in the brain by 32% compared to age-matched wild-type mice. Moreover, the H67D brain had increased isoprostane and decreased glutathione, indicating elevated oxidative stress. Additionally, the H67D brain had decreased global methylation and attenuated DNA methyltransferase (DNMT) activity. Direct addition of iron to purified DNMT in vitro decreased enzyme activity in a concentration-dependent manner. Last, H67D mice exhibited decreased anxiety-like behavior, which was associated with increased expression of the GABAA receptor α2 subunits by 93%, and these changes were also observed in H67D mice fed a low-iron diet. Taken together, our results suggest a putative role of HFE in regulating labile iron status in the brain, and mutation in H67D perturbs redox-methylation status, contributing to GABAergic dysfunction.-Ye, Q., Trivedi, M., Zhang, Y., Böhlke, M., Alsulimani, H., Chang, J., Maher, T., Deth, R., Kim, J. Brain iron loading impairs DNA methylation and alters GABAergic function in mice.

Oxycodone Exposure: A Magnetic Resonance Imaging Study in Response to Acute and Chronic Oxycodone Treatment in Rats.

The present study was designed to use blood-oxygen-level dependent (BOLD) imaging to "fingerprint" the change in activity in response to oxycodone (OXY) in drug naïve rats before and after repeated exposure to OXY. It was hypothesized that repeated exposure to OXY would initiate adaptive changes in brain organization that would be reflected in an altered response to opioid exposure. Male rats exposed to OXY repeatedly showed conditioned place preference, evidence of drug-seeking behavior and putative neuroadaptation. As these studies were done on awake rats we discovered it was not possible to image rats continuously exposed to OXY due to motion artifact judged to be withdrawal while in the scanner. To circumvent this problem manganese-enhanced MRI (MEMRI) was used to map the distributed integrated activity pattern resulting from continuous OXY exposure. Rats were administered OXY (2.5 mg/kg, i.p.) during image acquisition and changes in BOLD signal intensity were recorded and the activation and deactivation of integrated neural circuits involved in olfaction and motivation were identified. Interestingly, the circuitry of the mesencephalic dopaminergic system showed little activity to the first exposure of OXY. In the MEMRI study, rats received OXY treatments (2.5 mg/kg, twice daily) for four consecutive days following intraventricular MnCl2. Under isoflurane anesthesia, T1-weighted images were acquired and subsequently analyzed showing activity in the forebrain limbic system, ventral striatum, accumbens, amygdala and hippocampus. These results show brain activity is markedly different when OXY is presented to drug naïve rats versus rats with prior, repeated exposure to drug.

Identification of Myalgic Encephalomyelitis/Chronic Fatigue Syndrome-associated DNA methylation patterns.

BACKGROUND: Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is a complex condition involving multiple organ systems and characterized by persistent/relapsing debilitating fatigue, immune dysfunction, neurological problems, and other symptoms not curable for at least 6 months. Disruption of DNA methylation patterns has been tied to various immune and neurological diseases; however, its status in ME/CFS remains uncertain. Our study aimed at identifying changes in the DNA methylation patterns that associate with ME/CFS. METHODS: We extracted genomic DNA from peripheral blood mononuclear cells from 13 ME/CFS study subjects and 12 healthy controls and measured global DNA methylation by ELISA-like method and site-specific methylation status using Illumina MethylationEPIC microarrays. Pyrosequencing validation included 33 ME/CFS cases and 31 controls from two geographically distant cohorts. RESULTS: Global DNA methylation levels of ME/CFS cases were similar to those of controls. However, microarray-based approach allowed detection of 17,296 differentially methylated CpG sites in 6,368 genes across regulatory elements and within coding regions of genes. Analysis of DNA methylation in promoter regions revealed 307 differentially methylated promoters. Ingenuity pathway analysis indicated that genes associated with differentially methylated promoters participated in at least 15 different pathways mostly related to cell signaling with a strong immune component. CONCLUSIONS: This is the first study that has explored genome-wide epigenetic changes associated with ME/CFS using the advanced Illumina MethylationEPIC microarrays covering about 850,000 CpG sites in two geographically distant cohorts of ME/CFS cases and matched controls. Our results are aligned with previous studies that indicate a dysregulation of the immune system in ME/CFS. They also suggest a potential role of epigenetic de-regulation in the pathobiology of ME/CFS. We propose screening of larger cohorts of ME/CFS cases to determine the external validity of these epigenetic changes in order to implement them as possible diagnostic markers in clinical setting.

RNaseT2 knockout rats exhibit hippocampal neuropathology and deficits in memory.

RNASET2 deficiency in humans is associated with infant cystic leukoencephalopathy, which causes psychomotor impairment, spasticity and epilepsy. A zebrafish mutant model suggests that loss of RNASET2 function leads to neurodegeneration due to the accumulation of non-degraded RNA in the lysosomes. The goal of this study was to characterize the first rodent model of RNASET2 deficiency. The brains of 3- and 12-month-old RNaseT2 knockout rats were studied using multiple magnetic resonance imaging modalities and behavioral tests. While T1- and T2-weighted images of RNaseT2 knockout rats exhibited no evidence of cystic lesions, the prefrontal cortex and hippocampal complex were enlarged in knockout animals. Diffusion-weighted imaging showed altered anisotropy and putative gray matter changes in the hippocampal complex of the RNaseT2 knockout rats. Immunohistochemistry for glial fibrillary acidic protein (GFAP) showed the presence of hippocampal neuroinflammation. Decreased levels of lysosome-associated membrane protein 2 (LAMP2) and elevated acid phosphatase and ß-N-acetylglucosaminidase (NAG) activities indicated that the RNASET2 knockout rats likely had altered lysosomal function and potential defects in autophagy. Object recognition tests confirmed that RNaseT2 knockout rats exhibited memory deficits. However, the Barnes maze, and balance beam and rotarod tests indicated there were no differences in spatial memory or motor impairments, respectively. Overall, patients with RNASET2 deficiency exhibited a more severe neurodegeneration phenotype than was observed in the RNaseT2 knockout rats. However, the vulnerability of the knockout rat hippocampus as evidenced by neuroinflammation, altered lysosomal function and cognitive defects indicates that this is still a useful in vivo model to study RNASET2 function.

Evidence of Neurobiological Changes in the Presymptomatic PINK1 Knockout Rat.

BACKGROUND: Genetic models of Parkinson's disease (PD) coupled with advanced imaging techniques can elucidate neurobiological disease progression, and can help identify early biomarkers before clinical signs emerge. PTEN-induced putative kinase 1 (PINK1) helps protect neurons from mitochondrial dysfunction, and a mutation in the associated gene is a risk factor for recessive familial PD. The PINK1 knockout (KO) rat is a novel model for familial PD that has not been neuroradiologically characterized for alterations in brain structure/function, alongside behavior, prior to 4 months of age. OBJECTIVE: To identify biomarkers of presymptomatic PD in the PINK1 -/- rat at 3 months using magnetic resonance imaging techniques. METHODS: At postnatal weeks 12-13; one month earlier than previously reported signs of motor and cognitive dysfunction, this study combined imaging modalities, including assessment of quantitative anisotropy across 171 individual brain areas using an annotated MRI rat brain atlas to identify sites of gray matter alteration between wild-type and PINK1 -/- rats. RESULTS: The olfactory system, hypothalamus, thalamus, nucleus accumbens, and cerebellum showed differences in anisotropy between experimental groups. Molecular analyses revealed reduced levels of glutathione, ATP, and elevated oxidative stress in the substantia nigra, striatum and deep cerebellar nuclei. Mitochondrial genes encoding proteins in Complex IV, along with mRNA levels associated with mitochondrial function and genes involved in glutathione synthesis were reduced. Differences in brain structure did not align with any cognitive or motor impairment. CONCLUSIONS: These data reveal early markers, and highlight novel brain regions involved in the pathology of PD in the PINK1 -/- rat before behavioral dysfunction occurs.

Neuro-Immune Influences on Health Outcomes: A Nutritional Perspective.

No Abstract Available.

Alterations in Gene Expression of Components of the Renin-Angiotensin System and Its Related Enzymes in Lung Cancer.

OBJECTIVES: The study assessed the existence and significance of associations between the expression of fifteen renin-angiotensin system component genes and lung adenocarcinoma. MATERIALS AND METHODS: NCBI's built-in statistical tool, GEO2R, was used to calculate Student's t-tests for the associations found in a DNA expression study of adenocarcinoma and matched healthy lung tissue samples. The raw data was processed with GeneSpring™ and then used to generate figures with and without Sidak's multiple comparison correction. RESULTS: Ten genes were found to be significantly associated with adenocarcinoma. Seven of these associations remained statistically significant after correction for multiple comparisons. Notably, AGTR2, which encodes the AT2 angiotensin II receptor subtype, was significantly underexpressed in adenocarcinoma tissue (p < 0.01). AGTR1, ACE, ENPEP, MME, and PRCP, which encode the AT1 angiotensin II receptor, angiotensin-converting enzyme, aminopeptidase N, neprilysin, and prolylcarboxypeptidase, respectively, were also underexpressed. AGT, which encodes angiotensinogen, the angiotensin peptide precursor, was overexpressed in adenocarcinoma tissue. CONCLUSION: The results suggest an association between the expression of the genes for renin-angiotensin system-related proteins and adenocarcinoma. While further research is necessary to conclusively demonstrate a link between the renin-angiotensin system and lung cancers, the results suggest that the renin-angiotensin system plays a role in the pathology of adenocarcinoma.

Short-term sleep deprivation leads to decreased systemic redox metabolites and altered epigenetic status.

Sleep is critical for repair as well as the rejuvenation processes in the body and many of these functions are regulated via underlying cellular metabolic homeostasis. Changes in sleep pattern are reported to alter such metabolic function resulting in altered disease susceptibility or behavior. Here, we measured the extent to which overnight total sleep deprivation (SD) in young adult humans can influence systemic (plasma-derived) redox-metabolism including the major antioxidant, glutathione as well as DNA methylation levels. Nineteen participants (n = 19, µ age = 21, SD = 3.09) underwent morning testing before and after overnight total SD. Biochemical measures before and after SD revealed that glutathione, ATP, cysteine, and homocysteine levels were significantly reduced following one night of sleep deprivation (all p's < 0.01). Parallel to the well-recognized fact that sleep deprivation (maintaining wakefulness) uses up metabolic reserves, we observed that morning cortisol levels were blunted after sleep deprivation. There were no significant correlations between self-reported or actigraphy-measured sleep and the biochemical measurements, strongly indicating that prior sleep behavior did not have any direct influence on the biochemical measures taken at baseline or after sleep deprivation. Results from the current investigation supports the previous literature implicating the induction of oxidative stress and ATP depletion with sleep deprivation. Furthermore, such altered antioxidant status can also induce downstream epigenetic changes. Although we did not measure the specific genes that were altered under the influence of such sleep deprivation, such epigenetic changes could potentially contribute towards disease predisposition.

MicroRNA-34a Encapsulated in Hyaluronic Acid Nanoparticles Induces Epigenetic Changes with Altered Mitochondrial Bioenergetics and Apoptosis in Non-Small-Cell Lung Cancer Cells.

Therapies targeting epigenetic changes for cancer treatment are in Phase I/II trials; however, all of these target only nuclear DNA. Emerging evidence suggests presence of methylation marks on mitochondrial DNA (mtDNA); but their contribution in cancer is unidentified. Expression of genes encoded on mtDNA are altered in cancer cells, along with increased glycolytic flux. Such glycolytic flux and elevated reactive oxygen species is supported by increased antioxidant; glutathione. MicroRNA-34a can translocate to mitochondria, mediate downstream apoptotic effects of tumor suppressor P53, and inhibit the antioxidant response element Nrf-2, resulting in depleted glutathione levels. Based on such strong rationale, we encapsulated microRNA-34a in our well-established Hyaluronic-Acid nanoparticles and delivered to cisplatin-sensitive and cisplatin-resistant A549-lung adenocarcinoma cells. Successful delivery and uptake in cells resulted in altered ATP levels, decreased glycolytic flux, Nrf-2 and glutathione levels, ultimately resulting in caspase-3 activation and apoptosis. Most important were the concurrent underlying molecular changes in epigenetic status of D-loop on the mtDNA and transcription of mtDNA-encoded genes. Although preliminary, we provide a novel therapeutic approach in form of altered mitochondrial bioenergetics and redox status of cancer cells with underlying changes in epigenetic status of mtDNA that can subsequently results in induction of cancer cell apoptosis.

Redox-Related Epigenetic Mechanisms in Glioblastoma: Nuclear Factor (Erythroid-Derived 2)-Like 2, Cobalamin, and Dopamine Receptor Subtype 4.

Glioblastoma is an exceptionally difficult cancer to treat. Cancer is universally marked by epigenetic changes, which play key roles in sustaining a malignant phenotype, in addition to disease progression and patient survival. Studies have shown strong links between the cellular redox state and epigenetics. Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) is a redox-sensitive transcription factor that upregulates endogenous antioxidant production, and is aberrantly expressed in many cancers, including glioblastoma. Methylation of DNA and histones provides a mode of epigenetic regulation, and cobalamin-dependent reactions link the redox state to methylation. Antagonists of dopamine receptor subtype 4 (D4 receptor) were recently shown to restrict glioblastoma stem cell growth by downregulating trophic signaling, resulting in inhibition of functional autophagy. In addition to stimulating glioblastoma stem cell growth, D4 receptors have the unique ability to catalyze cobalamin-dependent phospholipid methylation. Therefore, D4 receptors represent an important node in a molecular reflex pathway involving Nrf2 and cobalamin, operating in conjunction with redox status and methyl group donor availability. In this article, we describe the redox-related effects of Nrf2, cobalamin metabolism, and the D4 receptor on the regulation of the epigenetic state in glioblastoma.

Clinical evaluation of glutathione concentrations after consumption of milk containing different subtypes of ß-casein: results from a randomized, cross-over clinical trial.

This study reports the plasma glutathione concentrations in a double-blind, randomized, controlled, 2 × 2 cross-over study in which healthy participants consumed conventional milk (2 × 250 mL per day) containing both A1 and A2 types of ß-casein, or milk containing only A2 type ß-casein. Beta-casomorphin-7 (BCM-7), a peptide uniquely derived from the A1 type of ß-casein, was previously reported to downregulate glutathione expression in human gut epithelial and neuronal cell lines by limiting cysteine uptake. The current human study demonstrates that consumption of milk containing only A2 ß-casein was associated with a greater increase in plasma glutathione concentrations compared with the consumption of milk containing both ß-casein types, and did not increase plasma BCM-7 concentrations compared with the washout diet in the study participants. Thus, milk containing only A2 ß-casein and not A1 ß-casein has the potential to promote the production of the antioxidant glutathione in humans. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov; identifier: NCT02406469.

Differential neurogenic effects of casein-derived opioid peptides on neuronal stem cells: implications for redox-based epigenetic changes.

Food-derived peptides, such as ß-casomorphin BCM7, have potential to cross the gastrointestinal tract and blood-brain barrier and are associated with neurological disorders and neurodevelopmental disorders. We previously established a novel mechanism through which BCM7 affects the antioxidant levels in neuronal cells leading to inflammatory consequences. In the current study, we elucidated the effects of casein-derived peptides on neuronal development by using the neurogenesis of neural stem cells (NSCs) as an experimental model. First, the transient changes in intracellular thiol metabolites during NSC differentiation (neurogenesis) were investigated. Next, the neurogenic effects of food-derived opioid peptides were measured, along with changes in intracellular thiol metabolites, redox status and global DNA methylation levels. We observed that the neurogenesis of NSCs was promoted by human BCM7 to a greater extent, followed by A2-derived BCM9 in contrast to bovine BCM7, which induced increased astrocyte formation. The effect was most apparent when human BCM7 was administered for 1day starting on 3days postplating, consistent with immunocytochemistry. Furthermore, neurogenic changes regulated by bovine BCM7 and morphine were associated with an increase in the glutathione/glutathione disulfide ratio and a decrease in the S-adenosylmethionine/S-adenosylhomocysteine ratio, indicative of changes in the redox and the methylation states. Finally, bovine BCM7 and morphine decreased DNA methylation in differentiating NSCs. In conclusion, these results suggest that food-derived opioid peptides and morphine regulated neurogenesis and differentiation of NSCs through changes in the redox state and epigenetic regulation.