Potential of dietary polyphenols for protection from age-related decline and neurodegeneration: a role for gut microbiota?

Many epidemiological studies have shown the beneficial effects of a largely plant-based diet, and the strong association between the consumption of a Mediterranean-type diet with healthy aging including a lower risk of cognitive decline. The Mediterranean diet is characterized by a high intake of olive oil, fruits and vegetables and is rich in dietary fiber and polyphenols - both of which have been postulated to act as important mediators of these benefits. Polyphenols are large molecules produced by plants to protect them from environmental threats and injury. When ingested by humans, as little as 5% of these molecules are absorbed in the small intestine with the majority metabolized by the gut microbiota into absorbable simple phenolic compounds. Flavan-3-ols, a type of flavonoid, contained in grapes, berries, pome fruits, tea, and cocoa have been associated with many beneficial effects on several risk factors for cardiovascular disease, cognitive function and brain regions involved in memory formation. Both preclinical and clinical studies suggest that these brain and heart benefits can be attributed to endothelial vascular effects and anti-inflammatory properties among others. More recently the gut microbiota has emerged as a potential modulator of the aging brain and intriguingly polyphenols have been shown to alter microbiota composition and be metabolized by different microbial species. However, there is a need for well controlled studies in large populations to identify predictors of response, particularly given the vast inter-individual variation of human gut microbiota.

The microbiome regulates amygdala-dependent fear recall.

The amygdala is a key brain region that is critically involved in the processing and expression of anxiety and fear-related signals. In parallel, a growing number of preclinical and human studies have implicated the microbiome-gut-brain in regulating anxiety and stress-related responses. However, the role of the microbiome in fear-related behaviours is unclear. To this end we investigated the importance of the host microbiome on amygdala-dependent behavioural readouts using the cued fear conditioning paradigm. We also assessed changes in neuronal transcription and post-transcriptional regulation in the amygdala of naive and stimulated germ-free (GF) mice, using a genome-wide transcriptome profiling approach. Our results reveal that GF mice display reduced freezing during the cued memory retention test. Moreover, we demonstrate that under baseline conditions, GF mice display altered transcriptional profile with a marked increase in immediate-early genes (for example, Fos, Egr2, Fosb, Arc) as well as genes implicated in neural activity, synaptic transmission and nervous system development. We also found a predicted interaction between mRNA and specific microRNAs that are differentially regulated in GF mice. Interestingly, colonized GF mice (ex-GF) were behaviourally comparable to conventionally raised (CON) mice. Together, our data demonstrates a unique transcriptional response in GF animals, likely because of already elevated levels of immediate-early gene expression and the potentially underlying neuronal hyperactivity that in turn primes the amygdala for a different transcriptional response. Thus, we demonstrate for what is to our knowledge the first time that the presence of the host microbiome is crucial for the appropriate behavioural response during amygdala-dependent memory retention.

The antimicrobial capacity of embalming solutions: a comparative study.

AIMS: Infectious health risks are associated with handling human cadavers and to decrease such risks, cadavers are embalmed using different chemicals. The aim of this study is to quantify the amount of micro-organisms present in different regions of human cadavers before embalming, after embalming and over a period of 8 months. METHODS AND RESULTS: Human cadavers were embalmed using Thiel, formalin, Genelyn and the Imperial College London soft-preservation (ICL-SP) solution with two cadavers per technique. Sterile swabs were used to collect samples from different regions. Samples were collected every 2 months. All cadavers had a high number of microbial colonies before embalming. While no colonies were detected on formalin and Genelyn embalmed cadavers post-embalming, the number of colonies decreased significantly in Thiel-embalmed cadavers and stayed relatively the same in ICL-SP-embalmed cadavers. CONCLUSIONS: Formalin-embalmed cadavers showed the strongest disinfecting abilities followed by Thiel-embalmed cadavers, then Genelyn-embalmed cadavers and finally by ICL-SP cadavers. SIGNIFICANCE AND IMPACT OF THE STUDY: This study highlights how under researched this area is and the evident variation in the antimicrobial abilities of different embalming solutions on the cadaver as a whole and within different regions of the same cadaver.

A Survey of tooth morphology teaching methods employed in the United Kingdom and Ireland.

BACKGROUND: Tooth morphology is a central component of the dental curriculum and is applicable to all dental specialities. Traditional teaching methods are being supplemented with innovative strategies to tailor teaching and accommodate the learning styles of the recent generation of students. METHODS: An online survey was compiled and distributed to the staff involved in teaching tooth morphology in the United Kingdom and Ireland to assess the importance of tooth morphology in the dentistry curriculum and the methodologies employed in teaching. RESULTS: The results of the survey show that tooth morphology constitutes a small module in the dental curriculum. It is taught in the first 2 years of the dental curriculum but is applicable in the clinical years and throughout the dental career. Traditional teaching methods, lecture and practical, are being augmented with innovative teaching including e-learning via virtual learning environment, tooth atlas and e-books leading to blended learning. The majority of the schools teach both normal dental anatomy and morphologic variations of dental anatomy and utilise plastic teeth for practical and examination purposes. Learning the 3D aspects of tooth morphology was deemed important by most of the respondents who also agreed that tooth morphology is a difficult topic for the students. CONCLUSION: Despite being core to the dental curriculum, overall minimal time is dedicated to the delivery of tooth morphology, creating a reliance on the student to learn the material. New forms of delivery including computer-assisted learning tools should help sustain learning and previously acquired knowledge.

Evaluation of an animation tool developed to supplement dental student study of the cranial nerves.

INTRODUCTION: The structure/function of the cranial nerves is a core topic for dental students. However, due to the perceived complexity of the subject, it is often difficult for students to develop a comprehensive understanding of key concepts using textbooks and models. It is accepted that the acquisition of anatomical knowledge can be facilitated by visualisation of structures. This study aimed to develop and assess a novel cranial nerve animation as a supplemental learning aid for dental students. MATERIALS AND METHODS: A multidisciplinary team of anatomists, neuroscientists and a computer scientist developed a novel animation depicting the cranial nerves. The animation was viewed by newly enrolled first-year dental students, graduate entry dental students (year 1) and dental hygiene students (year 1). A simple life scenario employing the use of the cranial nerves was developed using a cartoon-type animation with a viewing time of 3.58 minutes. The animation was developed with emphasis on a life scenario. The animation was placed online for 2 weeks with open access or viewed once in a controlled laboratory setting. Questionnaires were designed to assess the participants' attitude towards the animation and their knowledge of the cranial nerves before and after visualisation. This study was performed before the delivery of core lectures on the cranial nerves. RESULTS: Our findings indicate that the use of the animation can act as a supplemental tool to improve student knowledge of the cranial nerves. Indeed, data indicate that a single viewing of the animation, in addition to 2-week access to the animation, can act as a supplemental learning tool to assist student understanding of the structure and function of cranial nerves. The animation significantly enhanced the student's opinion that their cranial nerve knowledge had improved. From a qualitative point of view, the students described the animation as an enjoyable and useful supplement to reading material/lectures and indicated that the animation was a useful tool in understanding the cranial nerves. CONCLUSION: Overall, these findings indicate that an animation demonstrating the cranial nerves in a simple, everyday functional scenario may act as a learning aid in the study of cranial nerves.

Imaging of oxygen gradients in giant umbrella cells: an ex vivo PLIM study.

O2 plays a pivotal role in aerobic metabolism and regulation of cell and tissue function. Local differences and fluctuations in tissue O2 levels are well documented; however, the physiological significance of O2 microgradients, particularly at the subcellular level, remains poorly understood. Using the cell-penetrating phosphorescent O2 probe Pt-Glc and confocal fluorescence microscopy, we visualized O2 distribution in individual giant (>100-µm) umbrella cells located superficially in the urinary bladder epithelium. We optimized conditions for in vivo phosphorescent staining of the inner surface of the mouse bladder and subsequent ex vivo analysis of excised live tissue. Imaging experiments revealed significant (≤85 µM) and heterogeneous deoxygenation within respiring umbrella cells, with radial O2 gradients of up to 40 µM across the cell, or ∼0.6 µM/µm. Deeply deoxygenated (5-15 µM O2) regions were seen to correspond to the areas enriched with polarized mitochondria. Pharmacological activation of mitochondrial respiration decreased oxygenation and O2 gradients in umbrella cells, while inhibition with antimycin A dissipated the gradients and caused gradual reoxygenation of the tissue to ambient levels. Detailed three-dimensional maps of O2 distribution potentially can be used for the modeling of intracellular O2-dependent enzymatic reactions and downstream processes, such as hypoxia-inducible factor signaling. Further ex vivo and in vivo studies on intracellular and tissue O2 gradients using confocal imaging can shed light on the molecular mechanisms regulating O2-dependent (patho)physiological processes in the bladder and other tissues.

Cyclodextrin mediated delivery of NF-κB and SRF siRNA reduces the invasion potential of prostate cancer cells in vitro.

Prostate cancer is the most common cancer in men of the western world. To date, no effective treatment exists for metastatic prostate cancer and consequently, there is an urgent need to develop new and improved therapeutics. In recent years, the therapeutic potential of RNA interference (RNAi) has been extensively explored in a wide range of diseases including prostate cancer using numerous gene delivery vectors. The aims of this study were to investigate the ability of a non-viral modified cyclodextrin (CD) vector to deliver siRNA to the highly metastatic PC-3 prostate cancer cell line, to quantify the resulting knockdown of the two target genes (RelA and SRF) and to study the effects of the silencing on metastasis. Data from a Matrigel in vitro invasion assay indicated that the silencing of the target genes achieved by the CD vector resulted in significant reductions (P=0.0001) in the metastatic potential of these cells. As the silencing of these target genes was shown not to have a negative impact on cell viability, we hypothesise that the mechanism of invasion inhibition is due, in part, to the significant reduction observed (P⩽0.0001) in the level of pro-inflammatory cytokine, MMP9, which is known to be implicated in the metastasis of prostate cancer.

Genomics of schizophrenia: time to consider the gut microbiome?

Research into the genomics of schizophrenia promises much, but so far is resplendent with failures to replicate, and has yielded little of therapeutic potential. Within our bodies resides a dynamic population of gut microbes forming a symbiotic superorganism comprising a myriad of bacteria of approximately 10(14) cells, containing 100 times the number of genes of the human genome and weighing approximately the same as the human brain. Recent preclinical investigations indicate that these microbes majorly impact on cognitive function and fundamental behavior patterns, such as social interaction and stress management. We are pivotally dependent on the neuroactive substances produced by such bacteria. The biological diversity of this ecosystem is established in the initial months of life and is highly impacted upon by environmental factors. To date, this vast quantity of DNA has been largely ignored in schizophrenia research. Perhaps it is time to reconsider this omission.

A sustained hypothalamic-pituitary-adrenal axis response to acute psychosocial stress in irritable bowel syndrome.

BACKGROUND: Despite stress being considered a key factor in the pathophysiology of the functional gastrointestinal (GI) disorder irritable bowel syndrome (IBS), there is a paucity of information regarding the ability of IBS patients to respond to acute experimental stress. Insights into the stress response in IBS could open the way to novel therapeutic interventions. To this end, we assessed the response of a range of physiological and psychological parameters to the Trier Social Stress Test (TSST) in IBS. METHOD: Thirteen female patients with IBS and 15 healthy female age-matched control participants underwent a single exposure to the TSST. Salivary cortisol, salivary C-reactive protein (CRP), skin conductance level (SCL), GI symptoms, mood and self-reported stress were measured pre- and post-exposure to the TSST. RESULTS: The hypothalamic-pituitary-adrenal (HPA) axis response to the TSST was sustained in IBS, as shown by a greater total cortisol output throughout (p = 0.035) and higher cortisol levels measured by an area under the curve with respect to ground (AUCG) analysis (p = 0.044). In IBS patients, GI symptoms increased significantly during the recovery period following exposure to the TSST (p = 0.045). Salivary CRP and SCL activity showed significant changes in relation to stress but with no differential effect between experimental groups. CONCLUSIONS: Patients with IBS exhibit sustained HPA axis activity, and an increase in problematic GI symptoms in response to acute experimental psychosocial stress. These data pave the way for future interventional studies aimed at identifying novel therapeutic approaches to modulate the HPA axis and GI symptom response to acute psychosocial stress in IBS.

Cognitive performance in irritable bowel syndrome: evidence of a stress-related impairment in visuospatial memory.

BACKGROUND: Central nervous system (CNS) dysfunction is a prominent feature of the functional gastrointestinal (GI) disorder, irritable bowel syndrome (IBS). However, the neurobiological and cognitive consequences of key pathophysiological features of IBS, such as stress-induced changes in hypothalamic-pituitary-adrenal (HPA)-axis functioning, is unknown. Our aim was to determine whether IBS is associated with cognitive impairment, independently of psychiatric co-morbidity, and whether cognitive performance is related to HPA-axis function. METHOD: A cross-sectional sample of 39 patients with IBS, a disease control group of 18 patients with Crohn's disease (CD) in clinical remission and 40 healthy age- and IQ-matched control participants were assessed using the Paired Associates Learning (PAL), Intra-Extra Dimensional Set Shift (IED) and Spatial Working Memory (SWM) tests from the Cambridge Neuropsychological Test Automated Battery (CANTAB) and a computerized Stroop test. HPA-axis function was determined by measuring the cortisol awakening response (CAR). RESULTS: IBS patients exhibited a subtle visuospatial memory deficit at the PAL six- pattern stage (p = 0.03), which remained after psychiatric co-morbidity was controlled for (p = 0.04). Morning cortisol levels were lower in IBS (p = 0.04) and significantly associated with visuospatial memory performance within IBS only (p = 0.02). CONCLUSIONS: For the first time, altered cognitive function on a hippocampal-mediated test of visuospatial memory, which was related to cortisol levels and independent of psychiatric co-morbidity, has been identified in IBS. Visuospatial memory impairment may be a common, but currently neglected, component of IBS. Further elucidation of the nature of this impairment may lead to a greater understanding of the underlying pathophysiology of IBS, and may provide novel therapeutic approaches.

The microbiome-gut-brain axis during early life regulates the hippocampal serotonergic system in a sex-dependent manner.

Bacterial colonisation of the intestine has a major role in the post-natal development and maturation of the immune and endocrine systems. These processes are key factors underpinning central nervous system (CNS) signalling. Regulation of the microbiome-gut-brain axis is essential for maintaining homeostasis, including that of the CNS. However, there is a paucity of data pertaining to the influence of microbiome on the serotonergic system. Germ-free (GF) animals represent an effective preclinical tool to investigate such phenomena. Here we show that male GF animals have a significant elevation in the hippocampal concentration of 5-hydroxytryptamine and 5-hydroxyindoleacetic acid, its main metabolite, compared with conventionally colonised control animals. Moreover, this alteration is sex specific in contrast with the immunological and neuroendocrine effects which are evident in both sexes. Concentrations of tryptophan, the precursor of serotonin, are increased in the plasma of male GF animals, suggesting a humoral route through which the microbiota can influence CNS serotonergic neurotransmission. Interestingly, colonisation of the GF animals post weaning is insufficient to reverse the CNS neurochemical consequences in adulthood of an absent microbiota in early life despite the peripheral availability of tryptophan being restored to baseline values. In addition, reduced anxiety in GF animals is also normalised following restoration of the intestinal microbiota. These results demonstrate that CNS neurotransmission can be profoundly disturbed by the absence of a normal gut microbiota and that this aberrant neurochemical, but not behavioural, profile is resistant to restoration of a normal gut flora in later life.

Priming for health: gut microbiota acquired in early life regulates physiology, brain and behaviour.

UNLABELLED: The infant gut microbiome is dynamic, and radical shifts in composition occur during the first 3 years of life. Disruption of these developmental patterns, and the impact of the microbial composition of our gut on brain and behaviour, has attracted much recent attention. Integrating these observations is an important new research frontier. CONCLUSION: Early-life perturbations of the developing gut microbiota can impact on the central nervous system and potentially lead to adverse mental health outcomes.

Little things on which happiness depends: microRNAs as novel therapeutic targets for the treatment of anxiety and depression.

Anxiety and depression are devastating mental illnesses that are a significant public health concern. Selective serotonin-reuptake inhibitors are the first-line treatment strategy for these disorders, which despite being a significant advantage over older treatments, are hampered by a limited efficacy in a significant subset of patients, delayed onset of action and side effects that affect compliance. Thus, there is much impetus to develop novel therapeutic strategies. However, this goal can only be rationally realised with a better understanding of the molecular pathophysiology of these disorders. MicroRNAs (miRNAs) are a newly discovered class of gene-expression regulators that may represent a novel class of therapeutic targets to treat a variety of disorders including psychiatric diseases. miRNAs are heavily involved in regulating many physiological processes including those fundamental to the functioning of the central nervous system. Evidence collected to date has already demonstrated that miRNA-expression levels are altered in patients suffering from depression and anxiety and in pre-clinical models of psychological stress. Furthermore, increasing evidence suggests that psychoactive agents including antidepressants and mood stabilisers utilise miRNAs as downstream effectors. Altering miRNA levels has been shown to alter behaviour in a therapeutically desirable manner in pre-clinical models. This review aims to outline the evidence collected to date demonstrating miRNAs role in anxiety and depression, the potential advantages of targeting these small RNA molecules as well as some of the hurdles that will have to be overcome to fully exploit their therapeutic potential.

Disruption of the microbiota-gut-brain axis is a defining characteristic of the α-Gal A (-/0) mouse model of Fabry disease.

Fabry disease (FD) is an X-linked metabolic disease caused by a deficiency in α-galactosidase A (α-Gal A) activity. This causes accumulation of glycosphingolipids, especially globotriaosylceramide (Gb3), in different cells and organs. Neuropathic pain and gastrointestinal (GI) symptoms, such as abdominal pain, nausea, diarrhea, constipation, and early satiety, are the most frequent symptoms reported by FD patients and severely affect their quality of life. It is generally accepted that Gb3 and lyso-Gb3 are involved in the symptoms; nevertheless, the origin of these symptoms is complex and multifactorial, and the exact mechanisms of pathogenesis are still poorly understood. Here, we used a murine model of FD, the male α-Gal A (-/0) mouse, to characterize functionality, behavior, and microbiota in an attempt to elucidate the microbiota-gut-brain axis at three different ages. We provided evidence of a diarrhea-like phenotype and visceral hypersensitivity in our FD model together with reduced locomotor activity and anxiety-like behavior. We also showed for the first time that symptomology was associated with early compositional and functional dysbiosis of the gut microbiota, paralleled by alterations in fecal short-chain fatty acid levels, which partly persisted with advancing age. Interestingly, most of the dysbiotic features suggested a disruption of gut homeostasis, possibly contributing to accelerated intestinal transit, visceral hypersensitivity, and impaired communication along the gut-brain axis.

Region specific decrease in glial fibrillary acidic protein immunoreactivity in the brain of a rat model of depression.

A growing body of evidence from human postmortem and animal studies suggests that deficits in glial cell (particularly astrocytes) density and function, in limbic regions of the brain contribute to the etiology of depressive disorders. Despite the widespread use of Wistar-Kyoto (WKY) rat strain as a model of depression and stress susceptibility, there is a paucity of data examining whether alterations in brain astrocytic population are present in the model. In the present study, we investigated the expression of the astrocytic markers glial fibrillary acidic protein (GFAP) in various brain regions in WKY rats in comparison to Sprague-Dawley rats. A significant deficit in GFAP-immunoreactive cells was found in the prefrontal cortex region (infralimbic, prelimbic and anterior cingulate cortex), in the basolateral amygdala as well as in the hippocampus (CA3 and dentate gyrus) in WKY rat brain. No statistical difference was found in the other brain regions analyzed (insular cortex, somatosensory cortex, CA1 and callosal white matter). No difference was found in the total density of astrocytes (assessed by s-100beta immunoreactivity), neurons (determined by NeuN expression) or in the total number of cells in the regions of interest. A slight increase in the intensity of s-100beta immunoreactivity was observed. The lower expression of GFAP in WKY rats was further confirmed by Western-blot analysis. These results suggest that specific astrocytic deficits in GFAP expression in corticolimbic circuits may be a general correlate of depressive-like behavior in animal models in addition to human major depression. Moreover, they suggest that glial physiology may become a therapeutic target in depression and other stress-related conditions.

mGluR7 facilitates extinction of aversive memories and controls amygdala plasticity.

Formation and extinction of aversive memories in the mammalian brain are insufficiently understood at the cellular and molecular levels. Using the novel metabotropic glutamate receptor 7 (mGluR7) agonist AMN082, we demonstrate that mGluR7 activation facilitates the extinction of aversive memories in two different amygdala-dependent tasks. Conversely, mGluR7 knockdown using short interfering RNA attenuated the extinction of learned aversion. mGluR7 activation also blocked the acquisition of Pavlovian fear learning and its electrophysiological correlate long-term potentiation in the amygdala. The finding that mGluR7 critically regulates extinction, in addition to acquisition of aversive memories, demonstrates that this receptor may be relevant for the manifestation and treatment of anxiety disorders.

Gamma-aminobutyric acid-producing lactobacilli positively affect metabolism and depressive-like behaviour in a mouse model of metabolic syndrome.

Metabolic and neuroactive metabolite production represents one of the mechanisms through which the gut microbiota can impact health. One such metabolite, gamma-aminobutyric acid (GABA), can modulate glucose homeostasis and alter behavioural patterns in the host. We previously demonstrated that oral administration of GABA-producing Lactobacillus brevis DPC6108 has the potential to increase levels of circulating insulin in healthy rats. Therefore, the objective of this study was to assess the efficacy of endogenous microbial GABA production in improving metabolic and behavioural outcomes in a mouse model of metabolic dysfunction. Diet-induced obese and metabolically dysfunctional mice received one of two GABA-producing strains, L. brevis DPC6108 or L. brevis DSM32386, daily for 12 weeks. After 8 and 10 weeks of intervention, the behavioural and metabolic profiles of the mice were respectively assessed. Intervention with both L. brevis strains attenuated several abnormalities associated with metabolic dysfunction, causing a reduction in the accumulation of mesenteric adipose tissue, increased insulin secretion following glucose challenge, improved plasma cholesterol clearance and reduced despair-like behaviour and basal corticosterone production during the forced swim test. Taken together, this exploratory dataset indicates that intervention with GABA-producing lactobacilli has the potential to improve metabolic and depressive- like behavioural abnormalities associated with metabolic syndrome in mice.

Dimensional thinking in psychiatry in the era of the Research Domain Criteria (RDoC).

The biological mechanisms underlying psychiatric diagnoses are not well defined. Clinical diagnosis based on categorical systems exhibit high levels of heterogeneity and co-morbidity. The Research Domain Criteria (RDoC) attempts to reconceptualize psychiatric disorders into transdiagnostic functional dimensional constructs based on neurobiological measures and observable behaviour. By understanding the underlying neurobiology and pathophysiology of the relevant processes, the RDoC aims to advance biomarker development for disease prediction and treatment response. This important evolving dimensional framework must also consider environmental factors. Emerging evidence suggests that gut microbes (microbiome) play a physiological role in brain diseases by modulating neuroimmune, neuroendocrine and neural signalling pathways between the gut and the brain. The integration of the gut microbiome signature as an additional dimensional component of the RDoC may enhance precision psychiatry.

Regulation of behaviour by the nuclear receptor TLX.

The orphan nuclear receptor Tlx (Nr2e1) is a key regulator of both embryonic and adult hippocampal neurogenesis. Several different mouse models have been developed which target Tlx in vivo including spontaneous deletion models (from birth) and targeted and conditional knockouts. Although some conflicting findings have been reported, for the most part studies have demonstrated that Tlx is important in regulating processes that underlie neurogenesis, spatial learning, anxiety-like behaviour and interestingly, aggression. More recent data have demonstrated that disrupting Tlx during early life induces hyperactivity and that Tlx plays a role in emotional regulation. Moreover, there are sex- and age-related differences in some behaviours in Tlx knockout mice during adolescence and adulthood. Here, we discuss the role of Tlx in motor-, cognitive-, aggressive- and anxiety-related behaviours during adolescence and adulthood. We examine current evidence which provides insight into Tlx during neurodevelopment, and offer our thoughts on the function of Tlx in brain and behaviour. We further hypothesize that Tlx is a key target in understanding the emergence of neurobiological disorders during adolescence and early adulthood.