Université Laval
Canada Excellence Research Chair on the
Microbiome-Endocannabinoidome Axis
in Metabolic Health

Focus articles

Several studies have been performed around the world on the gut microbiome-endocannabinoidome axis. The members of CERC-MEND have been busy finding relevant articles and reviewing them for your reading.

Mai 16, 2023

-Review by Hayatte-Dounia Mir, postdoctoral fellow and Prof. Vincenzo Di Marzo.

The human gut harbors a complex and dynamic microbial ecosystem including bacteria, archaea, fungi, and viruses, known as the gut microbiota. It plays a role in the host metabolism and its deregulation is potentially implicated in metabolic disorders such as obesity, diabetes and the metabolic syndrome. The latter disorder is characterized by a set of metabolic dysregulations such as insulin resistance, obesity, hypertension and dyslipidemia. The action pathways by which the gut microbiota influences the metabolic syndrome are not fully elucidated but some studies highlighted the potential implication of bacterial metabolites such as butyrate, which is known to improve metabolic processes of the host, hence potentially prevent the development of this syndrome.

The endocannabinoid system (ES) is a ubiquitous system with several physiological functions, making it a key component in the investigation of the metabolic syndrome. Stricto sensus, it is defined as the two receptors CB1 and CB2, their ligands anandamide and 2-AG, as well as four regulatory enzymes, DAGL and NAPE-PLD (anabolic) and MAGL and FAAH (catabolic). It has been demonstrated that ES dysregulation in adipose tissue induces adipogenesis as well as metabolic inflammation. Based on those observations, studies are ongoing to investigate the nature of the link between the gut microbiota and the ES in the context of the metabolic syndrome. Is the ES modulated by butyrate produced by the gut microbiota? Through what mechanisms? These are some of the questions that the study by Hwang et al. started to decipher.

The authors first exposed human intestinal cells to butyrate alone at several concentrations. Secondly, they used bacteria genetically modified in order to overproduce butyrate. The value of this second part of the study was to investigate whether microbially produced butyrate can regulate the endocannabinoid system, in order to potentially use in the future the butyrate-producing bacteria as probiotics destined to target populations with metabolic disorders. They used the supernatant of the genetically modified bacteria to treat the cells.

The treatment at different concentrations (from 1 to 10 mM) induced a reduction in the gene expression and protein levels of DAGL and NAPE-PLD (from 2 to 10 mM) and an increase in the gene expression and protein levels of MAGL (from 1 to 2 mM), but no change was observed for FAAH. Those observations showed that butyrate has the potential to reduce, in a dose-dependent manner, the intestinal levels of anandamide and 2-AG via the regulation of the enzymes implicated in their biosynthesis and degradation. Butyrate from the genetically modified bacteria showed the same trends for the four regulatory enzymes at the gene expression levels, but not always at the protein level (as was the case for DAGL). This might be explained by the implication of other bacterial metabolites present in the supernatant, that were not present in the first treatment with butyrate alone. Another explanation might be the presence of LPS in the supernatant, which is known to affect endocannabinoids biosynthesis or degradation.

To conclude, this work demonstrated that butyrate produced by the gut microbiota has the capacity to regulate the gene and protein expression of key enzymes of the ES, potentially regulating intestinal levels of endocannabinoids implicated in the metabolic syndrome. Additional studies would be necessary to verify whether this modulatory mechanism is also maintained in vivo, and whether the gene expression modulations of the enzymes results in changes in actual endocannabinoid levels. If so, this would allow very interesting possibilities for clinical research related to the metabolic syndrome, in particular the use of butyrate-overproducing bacteria as probiotics.

August 3rd, 2022

-Review by Gabrielle St-Arnaud, MSc student and Prof. Alain Veilleux.

Nowadays, depression affects more than 300 million people worldwide1. A better understanding of the pathophysiology of this disease is essential to develop therapeutic tools, as the underlying mechanisms of depression remain poorly defined. The heterogeneity of symptoms also represents a major challenge to the understanding of the disease. Several studies have highlighted certain processes that promote the development of depression, including an abnormal response to stress, as well as disturbances in neuromodulatory systems such as the endocannabinoid system (ECS). In the brain, the hippocampus appears to be a cerebral region particularly involved in the pathophysiology of depression. As the disease develops, the hippocampus gradually loses neurons and consequently decreases in volume, while chronic stress limits neurogenesis in this brain region.

Additionally, the gut microbiota is involved in a myriad of biological functions and plays a role in depression. It is now recognized as an integral part of brain physiology through the gut-brain axis4-7. In patients with depression, alterations in the gut microbiota have been characterized7,8. Recent studies have shown that, in the animal model of depression, supplementation with probiotics influenced positively the behavior of the animals9. In humans, interventions with probiotics appear to improve the patients’ mood10.

Thus, Chevalier et al. explored the role of the gut microbiota in an animal model of stress-induced depression, and its interactions with brain homeostasis, lipid metabolism, and endocannabinoids11. To conduct this study, the authors used the 8-week unpredictable chronic mild stress protocol (UCMS), a well-documented method to induce a depressive-like state in mice12.

In depressed mice, the hippocampus showed a decrease in the formation of new neurons, and a decrease in the survival of immature neurons. The depressed state of the mice alters the gut microbiota: a preponderance of the bacterial families Ruminococcaceae and Porphyromonodaceae develops, at the expense of the Lactobacillaceae family. The transfer of the intestinal microbiota from a depressed mouse to a recipient mouse was sufficient to induce the development of a depressive state. Metabolomics analyses indicated that microbiota transfer alters lipid metabolism in the hippocampus. Notably, microbiota transfer decreases levels of polyunsaturated fatty acids and arachidonic acid (AA) in the hippocampus. The decrease in the major precursors of endocannabinoid production was associated with a reduction in 2-arachidonylglycerol (2-AG) levels in the hippocampus, suggesting a decrease in cellular signaling of the endocannabinoid system, particularly via the CB1 receptor. Hence, the authors tested several solutions to reverse the depressive state of the mice. First, inhibition of the 2-AG-degrading enzyme, the monoacylglycerol lipase (MAGL), increases the levels of 2-AG in the hippocampus, which promotes the proliferation and differentiation of new neural stem cells. A diet supplementation in AA that also restores hippocampal 2-AG levels and helps to resolve the depression phenotype. Finally, probiotic supplementation with Lactobacillus plantarum reconstitutes the healthy gut microbiota of mice with a depression phenotype restores endocannabinoid and polyunsaturated fatty acid levels in the hippocampus. Hippocampal neurogenesis is also stimulated following the supplementation.

An important limitation of the study is that the authors only considered the hippocampus, whereas several other brain regions (i.e., cerebral cortex, striatum) are involved in the pathophysiology of depression13. It would be essential to continue the analyses considering these regions to better understand the mechanisms of depression.

Overall, this study reports a demonstration of cause and effect between the gut microbiota and depression and suggests mechanisms involving the gut-brain axis via lipid metabolism, the endocannabinoid system, and neurogenesis of hippocampal neurons. The development of alternative therapies involving diet and probiotics is of interest to improve the quality of life of patients living with depression.

Read the full article.

References

  1. Kupfer, D. J., Frank, E., & Phillips, M. L. (2012). Major depressive disorder : New clinical, neurobiological, and treatment perspectives. Lancet, 379(9820), 1045‑1055. https://doi.org/10.1016/S0140-6736(11)60602-8
  2. Sheline, Y. I., Wang, P. W., Gado, M. H., Csernansky, J. G., & Vannier, M. W. (1996). Hippocampal atrophy in recurrent major depression. Proceedings of the National Academy of Sciences of the United States of America, 93(9), 3908‑3913.
  3. Sahay, A., & Hen, R. (2007). Adult hippocampal neurogenesis in depression. Nature Neuroscience, 10(9), 1110‑1115. https://doi.org/10.1038/nn1969
  4. Belkaid, Y., & Hand, T. (2014). Role of the Microbiota in Immunity and inflammation. Cell, 157(1), 121‑141. https://doi.org/10.1016/j.cell.2014.03.011
  5. Cani, P. D. (2014). Metabolism in 2013 : The gut microbiota manages host metabolism. Nature Reviews. Endocrinology, 10(2), 74‑76. https://doi.org/10.1038/nrendo.2013.240
  6. Sharon, G., Sampson, T. R., Geschwind, D. H., & Mazmanian, S. K. (2016). The Central Nervous System and the Gut Microbiome. Cell, 167(4), 915‑932. https://doi.org/10.1016/j.cell.2016.10.027
  7. Jiang, H., Ling, Z., Zhang, Y., Mao, H., Ma, Z., Yin, Y., Wang, W., Tang, W., Tan, Z., Shi, J., Li, L., & Ruan, B. (2015). Altered fecal microbiota composition in patients with major depressive disorder. Brain, Behavior, and Immunity, 48, 186‑194. https://doi.org/10.1016/j.bbi.2015.03.016
  8. Naseribafrouei, A., Hestad, K., Avershina, E., Sekelja, M., Linløkken, A., Wilson, R., & Rudi, K. (2014). Correlation between the human fecal microbiota and depression. Neurogastroenterology and Motility: The Official Journal of the European Gastrointestinal Motility Society, 26(8), 1155‑1162. https://doi.org/10.1111/nmo.12378
  9. Yang, C., Fujita, Y., Ren, Q., Ma, M., Dong, C., & Hashimoto, K. (2017). Bifidobacterium in the gut microbiota confer resilience to chronic social defeat stress in mice. Scientific Reports, 7(1), 45942. https://doi.org/10.1038/srep45942
  10. Ng, Q. X., Peters, C., Ho, C. Y. X., Lim, D. Y., & Yeo, W.-S. (2018). A meta-analysis of the use of probiotics to alleviate depressive symptoms. Journal of Affective Disorders, 228, 13‑19. https://doi.org/10.1016/j.jad.2017.11.063
  11. Chevalier, G., Siopi, E., Guenin-Macé, L., Pascal, M., Laval, T., Rifflet, A., Boneca, I. G., Demangel, C., Colsch, B., Pruvost, A., Chu-Van, E., Messager, A., Leulier, F., Lepousez, G., Eberl, G., & Lledo, P.-M. (2020). Effect of gut microbiota on depressive-like behaviors in mice is mediated by the endocannabinoid system. Nature Communications, 11(1), 6363. https://doi.org/10.1038/s41467-020-19931-2
  12. Frisbee, J. C., Brooks, S. D., Stanley, S. C., & d’Audiffret, A. C. (2015). An Unpredictable Chronic Mild Stress Protocol for Instigating Depressive Symptoms, Behavioral Changes and Negative Health Outcomes in Rodents. Journal of Visualized Experiments : JoVE, 106, 53109. https://doi.org/10.3791/53109
  13. Pandya, M., Altinay, M., Malone, D. A., & Anand, A. (2012). Where in the Brain Is Depression? Current psychiatry reports, 14(6), 634‑642. https://doi.org/10.1007/s11920-012-0322-7

August 3rd, 2022

- Review by Tina Khalilzadehsabet, MSc. student and Prof. Cristoforo Silvestri

Within the general population, 15 to 20% experience anhedonia and amotivation1,2. Onset of a broad spectrum of clinical conditions such as psychosis, depression, and dementia might be associated with anhedonia and amotivation in healthy individuals as they precede the aforementioned clinical conditions3,4. Diminished gut-microbiome diversity has been shown to be linked with a range of mental disorders including depression, schizophrenia, and chronic fatigue which all express anhedonia and amotivation5,6,7,8. Based on an animal study of a depression model, the endocannabinoid system mediates the link between the gut microbiome and anhedonia9. These results are correlated with another study which demonstrated that anhedonic/amotivational syndrome can be induced in healthy subjects by acute and chronic administration of delta-9-tetrahydrocannabinol; while cannabidiol (CBD) prevents this, potentially by either counteracting THC’s activity at the Cannabinoid receptor 1 (CNR1/CB1) and/or as an inhibitor of anandamide (AEA) and 2-acylglicerol degradation10. Oral administration of palmitoylethanolamide (PEA), an endogenous equivalent of CBD, has shown anti-depressive effects according to a randomized trial’s results11. Given the fact that gut-microbiome composition affects PEA levels12, Minichino et al. (REF) hypothesized that a reduced gut-microbiome diversity can result in more severe anhedonia/amotivation through the endocannabinoid system intervention. Thus, the correlation between microbiota diversity and anhedonia/amotivation was expected to be mediated via reduced PEA in serum or increased PEA levels in stool.

Longitudinal data was collected from the TwinsUK cohort, including 786 twin pairs. Microbiota profiles were generated by sequencing stool samples with the Illumina MiSeq platform. Untargeted LC-MS was used to measure the PEA concentration in faeces and serum and anhedonia/amotivation measurement was performed using Hospital Anxiety and Depression Scale. Age, gender, obesity, unhealthy diet, use of antidepressants and technical confounders were considered as covariates. Initial correlation analysis found that alpha diversity negatively correlated with both fecal PEA levels and anhedonia/amotivation severity. The association between the three variables was determined using a multilevel mediation model with family structure set as the random intercept. Alpha diversity, PEA levels, and anhedonia/amotivation were set as predictor, mediator, and outcome respectively. The results of the mediation model indicated a statistically significant direct relationship between alpha diversity and anhedonia/amotivation (β = −0.37; 95%CI: −0.71 to −0.03; P = 0.03). PEA fecal, but not serum, levels were found to mediate this association, having significant indirect and total effects, whilst the predictor direct effect on anhedonia/amotivation was diminished (β = −0.25; 95%CI: −0.60 to 0.09; P = 0.16). The results of unadjusted and adjusted models for technical confounders, in addition to the covariates, were similar. Furthermore, the association between any specific microbial genera, faecal PEA and anhedonia/amotivation was investigated, identifying Blautia and Dorea as being associated with both. While two mediation models did not find that the PEA mediated the association between Blautia and Dorea abundance with anhedonia/amotivation, using Blautia as the predictor, suggested a partial mediation of PEA on the association between the relative abundance of Blautia and anhedonia/amotivation severity.

High alpha diversity represents a healthy gut microbiome13. However, the relationship between diminished gut microbiome diversity and clinical conditions expressing anhedonia/amotivation is not consistent between studies, perhaps due to the variation of examined clinical conditions being heterogeneous5. The results of a recent study showed that anhedonia/amotivation severity in psychosis, and not the psychotic illness per se, is associated with reduced gut microbiota diversity14.

Increased excretion of PEA may have caused higher faecal levels of this metabolite, which may contribute to the severity of anhedonia/amotivation. Indeed, PEA supplementation therapy showed positive effects on patients with anxiety11.

Moreover, PEA shows anti-inflammatory effects via peroxisome proliferator-activated receptors γ blockage15. In addition to the endocannabinoidome, the PEA effects on anhedonia/amotivation might be a result of an alternative biological pathway considering the association between mental health and inflammation. For example, PEA might improve gut-barrier function, preventing metabolic endotoxemia, while gut permeability increases as a result of reduced gut microbiota diversity16.

The current study suggests the gut-microbiome-endocannabinoidome interaction as a predictive factor for anhedonia/amotivation. This data supports the need to take into account the endocannabinoidome’s role as a mediator in gut-microbiome targeted therapeutics aimed at mental health issues including anhedonia/amotivation.

Read the full article.

References

  1. Werbeloff N, Dohrenwend BP, Yoffe R, van Os J, Davidson M, Weiser M. The association between negative symptoms, psychotic experiences and later schizophrenia: a population-based longitudinal study. PLoS ONE. 2015;10: e0119852.
  2. Dominguez MDG, Saka MC, Lieb R, Wittchen HU, van Os J. Early expression of negative/disorganized symptoms predicting psychotic experiences and subsequent clinical psychosis: a 10-year study. Am J Psychiatry 2010; 167:1075–1082.
  3. Mallet J, Guessoum SB, Tebeka S, le Strat Y, Dubertret C. Self-evaluation of negative symptoms in adolescent and young adult first psychiatric episodes. Prog Neuropsychopharmacol Biol Psychiatry 2020; 103:109988.
  4. Winograd-Gurvich C, Fitzgerald PB, Georgiou-Karistianis N, Bradshaw JL, White OB. Negative symptoms: a review of schizophrenia, melancholic depression and Parkinson’s disease. Brain Res Bull 2006; 70:312–321.
  5. Safadi JM, Quinton AMG, Lennox BR, Burnet PWJ, Minichino A. Gut dysbiosis in severe mental illness and chronic fatigue: a novel trans-diagnostic construct? A systematic review and meta-analysis. Mol Psychiatry 2021. https://doi.org/10.1038/s41380- 021-01032-1.
  6. Valles-Colomer M, Falony G, Darzi Y, Tigchelaar EF, Wang J, Tito RY, et al. The neuroactive potential of the human gut microbiota in quality of life and depression. Nat Microbiol 2019; 4:623–632.
  7. Kelly JR, Minuto C, Cryan JF, Clarke G, Dinan TG. The role of the gut microbiome in the development of schizophrenia. Schizophrenia Research. 2020. https://doi.org/10.1016/j.schres. 2020.02.010.
  8. Giloteaux L, Goodrich JK, Walters WA, Levine SM, Ley RE, Hanson MR. Reduced diversity and altered composition of the gut microbiome in individuals with myalgic encephalomyelitis/ chronic fatigue syndrome. Microbiome. 2016; 4:30.
  9. Chevalier G, Siopi E, Guenin-Macé L, Pascal M, Laval T, Rifflet A, et al. Effect of gut microbiota on depressive-like behaviors in mice is mediated by the endocannabinoid system. Nat Commun 2020; 11:1–15.
  10. Lawn W, Freeman TP, Pope RA, Joye A, Harvey L, Hindocha C, et al. Acute and chronic effects of cannabinoids on effort-related decision-making and reward learning: an evaluation of the cannabis ‘amotivational’ hypotheses. Psychopharmacology 2016; 233:3537–3552.
  11. Ghazizadeh-Hashemi M, Ghajar A, Shalbafan MR, GhazizadehHashemi F, Afarideh M, Malekpour F, et al. Palmitoylethanolamide as adjunctive therapy in major depressive disorder: a doubleblind, randomized and placebo-controlled trial. J Affect Disord 2018; 232:127–133.
  12. Couch DG, Cook H, Ortori C, Barrett D, Lund JN, O’Sullivan SE. Palmitoylethanolamide and cannabidiol prevent inflammation-induced hyperpermeability of the human gut in vitro and in vivo-a randomized, placebo-controlled, double-blind controlled trial. Inflamm Bowel Dis 2019; 25:1006–1018.
  13. Mosca A, Leclerc M, Hugot JP. Gut microbiota diversity and human diseases: should we reintroduce key predators in our ecosystem? Front Microbiol 2016; 7:455.
  14. Schwarz E, Maukonen J, Hyytiäinen T, Kieseppä T, Orešič M, Sabunciyan S, et al. Analysis of microbiota in first episode psychosis identifies preliminary associations with symptom severity and treatment response. Schizophrenia Res 2018; 192:398–403.
  15. Alhouayek M, Muccioli GG. Harnessing the anti-inflammatory potential of palmitoylethanolamide. Drug Discov Today 2014; 19:1632–1639.
  16. Sarkar A, Harty S, Johnson KVA, Moeller AH, Carmody RN, Lehto SM, et al. The role of the microbiome in the neurobiology of social behaviour. Biol Rev 2020; 95:1131–1166.

June 21, 2022

- Review by Nadine Leblanc, research professional, and Prof. Frédéric Raymond.

It is now recognized that bacteria play a role in the health of their host, and advances in the field are taking great strides. Bacteria have the ability to produce molecules such as metabolic regulators, hormones, neurotransmitters, and ligand-mimicking molecules. These molecules bind to cellular receptors to elicit a response from them. Considering that more than 2,400 bacterial genomes are associated with humans, and that more than 14,000 gene families are likely to contribute to the production of small metabolites, the possibilities of obtaining molecules that can affect human health are extremely numerous.

Fatty acid amides (FAAs), molecules combining a fatty acid with an amine, are involved in cell signaling. Anandamide, the first AAG to have been thoroughly studied, can, among other things, activate the G protein-coupled receptors (GPCRs) CB1 and CB2, the first involved, among others, in the feeling of pleasure and motivation as well as obesity, the second in immune function and obesity. Human cells produce AAGs via the hydrolysis of phospholipids or via N-acyltransferases, as do some gram-negative bacteria in the intestine. The manufacture of lipoamino acids (LAAs), a subfamily of FAAs is a process similar to non-ribosomal peptide synthesis (NRPS), a process consisting of modules (A, T and C, respectively adenylation, transport/thiolation and condensation module) integrating subunits of amino acids, and which bacteria use to generate, for example, regulators of inflammation or antimicrobial peptides. It is with this process in mind that Changs and collaborators set up a protocol combining bioinformatics and microbiology in order to detect bacterial molecules such as new LAAs or other FAAs likely to have an impact on the health of the host.

The first steps were to browse the databases of the human intestinal microbiome in order to detect the presence of metabolic pathways of interest. Eight were kept because they are ubiquitous and highly transcribed in the gut, and the compounds they produce have not been characterized. Three genes are consistently found in these pathways, genes that have sequences similar to the A T C modules found in NRPS. Then, different synthetic biology approaches made it possible to reconstitute the pathways, to understand the biosynthesis of LAAs and other FAAs, to determine the specificity of substrates of the enzymes involved in the pathways and to identify the predominance of production of such metabolites. These approaches consisted, among other things, in redrawing the metabolic pathways, introducing them into a plasmid in order to clone them in E coli, validating in culture then by LC-MS the production of the metabolites, and bringing together a panoply of substrates (fatty acids and amines) representative of possible sources present in the human gut. Due to their greater production, four FAAs were selected and placed in the presence of G protein receptors. For the four, several GPCRs were activated or inactivated. FAAs with an oelic acid portion show greater GPR119 and GPR132 activity. Other studies have reported that both receptors are activated by known FAAs, including two in humans, indicating that these receptors are promiscuous for various human and bacterial FAAs. Lauroyl tryptamine, one of the four FAAs tested, has the particularity of inhibiting the EBI2 receptor, which is associated with inflammatory bowel disease. Oleoyl dopamine appears to interact with the majority of GPCR receptors, including those involved in inflammation and inflammatory bowel disease, as does oleoyl tyramine, which activates receptors associated with inflammatory bowel disease but also colorectal cancer. The final stages of the study made it possible to validate that the bacterium was indeed able to produce FAAs, both when it is isolated and when it is in the intestinal matrix.

The authors conclude that metabolic pathways of Clostridia are indeed able to produce fatty acid amides that can interact with the health of the host.

Read the full article.

June 10, 2022

- Review by Besma Boubertakh, PhD student and Prof. Cristoforo Silvestri

April is the Irritable Bowel Syndrome (IBS) Awareness Month, thus we present you this paper at the farewell of April 2022.

In contrast to organic disorders, which can be concretely (structurally or biochemically) identified and diagnosed, functional (non-organic) disorders present multiple clinical manifestations, yet exhibit no substantial histological irregularities or characterized monitoring markers [1]. Alarmingly, functional gastrointestinal (GI) disorders (FGIDs) affect more than 40% of the world population [2], and irritable bowel syndrome (IBS) is the most common among them, with a global prevalence of around 12% [3]. The Rome IV criteria validate IBS diagnosis when its symptoms occur at least six months ahead of the diagnosis, and have existed during the last three months, and includes recurrent abdominalgia that is associated with defecation and bowel movement changes, such as diarrhea, constipation, or a mix of both, and possible abdominal bloating or distension [3, 4]. Thus, IBS diagnosis cannot be based on a typical structural or biochemical anomaly, nor characterized with specific markers, but is rather based on the patient’s diet, medical, surgical, and psychological history [3, 5], hence the complexity of this chronic disease.

Colonic biopsies from affected humans and dogs commonly divulge the absence or the mild presence of low-grade mucosal inflammation and other morphological injuries, and the origin can be acute infectious enteritis or more traditionally mental/physical anxiety/stress [6]. Dogs are the oldest domesticated animals, through their coexistence with humans for more than 30,000 years, which have allowed them to develop humanlike sophisticated emotions [7]. This could contribute to dogs GI dysmotility with certain similar clinical signs similar to those of IBS in humans, such as in chronic idiopathic large-bowel diarrhea (CILBD) [7]. Despite the high prevalence of these illnesses, as for other multi-factorial diseases, their mechanisms still require deeper investigation.

The endocannabinoid (eCB) system has been proven to hold numerous answers to numerous challenging medical research questions, as it is implicated in both the nervous and peripheral systems, such as the immune and digestive systems [8]. Indeed, it is tightly implicated in many GI physiological and pathological mechanisms, for example, by limiting the enteric hypermotility and hypersensitivity [9]. Mast cells also play a critical role at the GI level, such as through increasing intestinal peristalsis and epithelial permeability [10].

Based on the aforementioned considerations, Giacomo Rossi and colleagues in the study discussed herein [6] recruited two groups, namely a study group (SG) of 20 adult dogs diagnosed with CILBD/IBS, and a control group (CG) of five healthy dogs of similar age. They conducted mechanistic investigative comparisons between these two groups, and further evaluated the hypothesis of potential health improvement effects of a probiotic mixture (Slab51 bacterial blend; Sivoy®) that introduced 112 to 225 × 109 lyophilized bacteria/10 kg body weight/day over three months (T0 to T1, compared) in the drinking water of the SG. Whilst no significant differences in histology scores that evaluate the severity of colonic biopsy architectural distortion and epithelial alteration were identified, the canine chronic enteropathy clinical activity index (CCECAI) score  showed significantly decreased disease activity at T1 compared to T0. Likewise, T1 colonic mucosal mastocytes were significantly less numerous than at T0 (and were considerably lower in CG than SG), as shown by immunohistochemical analysis. On the other hand, cellular counts of eCB receptor (CB)+ cells and qPCR gene expression assessments showed that colonic CB1 and CB2 expression was significantly lower in the SG compared to CG and was considerably augmented by probiotic ingestion at T1 [6].

The authors suggested that new treatments of CILBD/IBS may be based on long-term probiotic therapy, which might significantly activate the eCB system in the colon. We support the authors' view that the eCB system might fight this pathology at different levels, such as through mediating the gut microflora communication with the host, or through regulating gastrointestinal motility, mucosal secretion, immunological responses, and the body reaction to physical and emotional stress. Future studies could add to the assessment of mastocyte numbers performed here, though, since these cells exhibit eCB receptor expression and activity, especially CB2, through which the eCB system has been reported to paracrinally and/or autocrinally control their degranulation [11].

It is important to remember some of the limitations in this study. For example, IBS has been reported to be more frequent in females [12] who may also respond differently to therapy [13], and similarly, the eCB system presents sex differences [14, 15]. However, such discrepancies were not explored in this study. Furthermore, the study groups did not present a statistically balanced design, as the SG was fourfold bigger than the CG. In addition, the dog breeds assessed, the number of females/males and those that were spayed/neutered, respectively, were different within and between the groups. This may have had a significant impact on the findings in this study; however, the statistically significant changes observed suggest that they occured independently of genetic variability between the breeds. In agreement with this, the utilized probiotic blend was not only efficient in this study, but also resulted in significant effects and similar outcomes on eCB receptor expression in a zebrafish model [16]. However, the specific and potential role of its component species has not been discussed in this paper.

This study presents very appealing results that suggest both a promising therapy and interesting mechanistic insights of CILBD in dogs, a relatively rarely investigated animal in laboratory research, which could also suggest projections to IBS treatment in humans. Finally, this paper mentioned, more than once, the similarity between the endocannabinoid and opioid receptor expression patterns in response to probiotics [6]. It is also known that they might present several similar strong effects on the gastrointestinal tract, such as reducing bowel motility, and this ought to productively guide researchers studying one of the two systems to learn from the other, as the endocannabinoid and opioid systems have many expression patterns and functional roles in common, and they interact in health and disease, such as to induce analgesic effects [17-20].

Read the full article.

References

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  2. Sperber AD, Bangdiwala SI, Drossman DA, Ghoshal UC, Simren M, Tack J, Whitehead WE, Dumitrascu DL, Fang X, Fukudo S, et al: Worldwide Prevalence and Burden of Functional Gastrointestinal Disorders, Results of Rome Foundation Global Study. Gastroenterology 2021, 160:99-114.e113.
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  6. Rossi G, Gioacchini G, Pengo G, Suchodolski JS, Jergens AE, Allenspach K, Gavazza A, Scarpona S, Berardi S, Galosi L, et al: Enterocolic increase of cannabinoid receptor type 1 and type 2 and clinical improvement after probiotic administration in dogs with chronic signs of colonic dysmotility without mucosal inflammatory changes. Neurogastroenterol Motil 2020, 32:e13717.
  7. Katayama M, Kubo T, Yamakawa T, Fujiwara K, Nomoto K, Ikeda K, Mogi K, Nagasawa M, Kikusui T: Emotional Contagion From Humans to Dogs Is Facilitated by Duration of Ownership. Frontiers in Psychology 2019, 10.
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  9. Izzo AA, Muccioli GG, Ruggieri MR, Schicho R: Endocannabinoids and the Digestive Tract and Bladder in Health and Disease. In Endocannabinoids. Edited by Pertwee RG. Cham: Springer International Publishing; 2015: 423-447
  10. O'Sullivan M, Clayton N, Breslin NP, Harman I, Bountra C, McLaren A, O'Morain CA: Increased mast cells in the irritable bowel syndrome. Neurogastroenterol Motil 2000, 12:449-457.
  11. Wang Z, Lu M, Ren J, Wu X, Long M, Chen L, Chen Z: Electroacupuncture inhibits mast cell degranulation via cannabinoid CB2 receptors in a rat model of allergic contact dermatitis. Acupunct Med 2019, 37:348-355.
  12. Kim YS, Kim N: Sex-Gender Differences in Irritable Bowel Syndrome. Journal of Neurogastroenterology and Motility 2018, 24:544-558.
  13. van Kessel L, Teunissen D, Lagro-Janssen T: Sex-Gender Differences in the Effectiveness of Treatment of Irritable Bowel Syndrome: A Systematic Review. Int J Gen Med 2021, 14:867-884.
  14. Blanton HL, Barnes RC, McHann MC, Bilbrey JA, Wilkerson JL, Guindon J: Sex differences and the endocannabinoid system in pain. Pharmacology Biochemistry and Behavior 2021, 202:173107.
  15. Levine A, Liktor-Busa E, Lipinski AA, Couture S, Balasubramanian S, Aicher SA, Langlais PR, Vanderah TW, Largent-Milnes TM: Sex differences in the expression of the endocannabinoid system within V1M cortex and PAG of Sprague Dawley rats. Biology of Sex Differences 2021, 12:60.
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April 19, 2022

- Review by Pejman Abbasi Pashaki, PhD student and Prof. Cristoforo Silvestri

Gut microbial (GM) communities are crucial for maintaining the stability of the body, including neuronal and immune systems. They mediate their influences through microbial metabolites derived from carbohydrates, proteins, lipids, and bile acids, as well as microbial-associated molecular patterns (MAMPs), molecules that are conserved in all microbial classes but are absent in the host, e.g., flagellin from bacteria (1-3). GM also affects the host by producing a wide range of neurotransmitters such as norepinephrine, serotonin, dopamine, and gamma-aminobutyric acid (GABA). On the contrary, they are also able to consume neurotransmitters for their own purposes, thus, microbiota are capable of influencing neurotransmitter levels through various mechanisms. Specialized cells, called enteroendocrine cells (EECs) in the gut sense GM and their behavior, responding to stimulation (by microbial associated factors) in various ways, including secretion of hormones or neuronal transmitters (1). The connection between EECs and neurons is exclusively synaptic(2, 4); they are directly connected to the central nervous system and convey nutritional information. EECs have been shown to be activated by short-chain fatty acids (SCFAs) and branched-chain fatty acids (BCFAs) via G-protein coupled receptors, however, EECs are activated by various stimuli in the gut to secrete hormones or neurotransmitters in a calcium-dependent manner(5). Microbial products such as indole, a degradation product of tryptophan, and isovalerate are able to activate EECs (6) that extend to spinal sensory nerves, resulting in the release of 5-hydroxytryptamine (5-HT; serotonin) (3). Extrinsic neurons in the gut include sensory nerve fibers from the nodal vagal ganglia and dorsal root ganglia, which terminate in the spinal cord and may affect the brain (7).

TRPA1, an ionotropic receptor and the only member of the ankyrin family, is a nociceptor for pain and neuroinflammation that can be activated by environmental factors such as temperatures below 17°C, chemical and inflammatory factors. There are few endocannabinoids that appear to modulate TRPA1, but phyto- and synthetic cannabinoids have been reported to activate this channel (8). Investigating the effects of tryptophan catabolites produced by the gut microbiota on zebrafish Trpa1was the basis of the study by Yan et al(9). They constructed transgenic zebrafish to monitor the calcium activity of intestinal EECs in real time. In this line, they exposed the transgenic zebrafish to 11 live bacterial strains, and Edwardsiella tarda (E. tarda) was the only strain that stimulated EEC cells. An analysis of the transcriptome of zebrafish EECs cells revealed that they are significantly enriched in 192 genes mainly responsible for hormone secretion, chemical synaptic transmission, and neuropeptide signaling. Comparative profiling analysis showed that 24% of these genes are conserved in humans and 40% in mice. The conserved EEC genes are responsible for coding hormones, transcription factors, G protein-coupled receptors, and ion channels (10). Trpa1 is expressed in EECs, intestinal mesenchymal cells, and sensory neurons, the data showed that the zebrafish genome encodes two trpa1 paralogs, trpa1a and trpa1b, however, only trpa1b is expressed by a subset of EECs. To confirm that Trpa1 is indispensable for EEC activation, the study used a Trpa1 antagonist that remarkably impaired E. tarda's ability to stimulate EECs. This finding suggests that Trpa1 is required for the interaction between the microorganism and the host via EECs. Furthermore, they hypothesized that Trpa1 signalling may act as a warning signal for the exposure to pathogens and trigger the gut to expel the abnormality. The use of a Cre-loxP construct to eliminate trpa1 expression from EECs confirmed that EEC Trpa1 alone is responsible for the interaction between E. tarda and the host. To investigate the mechanism of the interaction between EEC Trpa1 and the gut microbiome, they applied optopharmacological methods to control EEC Trpa1 activation by pre-treating zebrafish with Optovin, a chemical that specifically activates Trpa1 only in the presence of UV light. UV light activation increased EEC Ca2+ levels in the wild-type (WT) but no trpa1b-/- larva and resulted in increased velocity of intestinal motility Similar results were obtained by microgavage of live E. tarda. Although formalin-killed E. tarda failed to activate downstream receptors, a cell-free supernatant has the potential to activate Trpa1b receptors through various indole ring-containing tryptophan derivatives namely indole, tryptophol (IEt), and indole-3-carboxaldehyde (IAld). In general, indole and IAld, but not other types of tryptophan derivatives, are sufficient to activate EECs in a Trpa1b-dependent manner. The group also documented that in zebrafish, as in mammals, vagal sensory ganglion cells transmit information directly to the vagal sensory region in the hindbrain. The information is sensed in the gut, in a region between the pancreas and the liver.

In summary, tryptophan, an essential amino taken from diet or synthesized by gut microbes, can be microbially degraded and produces a wide range of metabolites such as indole.  Yan et al. show that E. tarda has this potential and is able to alter gut behavior, such as motility, through Trpa1-expressing EECs. In humans, as in mice, TRPA1-expressing EECs are abundant in the small intestine but not in the colon, while on the contrary, microbially-derived tryptophan metabolites are restricted to the colon and are largely absent in the small intestine under normal physiological conditions.  Therefore, TRPA1-mediated regulation on gut motility, may not be significant under normal physiological conditions, but rather represents a protective host detection mechanism for tryptophan metabolite accumulation, which presents during exotic microbial overgrowth in the small intestine, or invasion by specific microbes. Nevertheless, Yan et al. raise the possibility that activation of the vagal sensory-ganglia-hindbrain pathway by gut microbes promotes neurological features associated with the gut microbiota, namely gut motility, and potentially, appetite and metabolism as well as influencing emotional behavior and cognitive function.

 

Read the full article.

References

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    10. Roberts GP, Larraufie P, Richards P, Kay RG, Galvin SG, Miedzybrodzka EL, et al. Comparison of Human and Murine Enteroendocrine Cells by Transcriptomic and Peptidomic Profiling. Diabetes. 2019;68(5):1062-72.

 

February 23, 2022

- Review by Mehdi Zineddine Messaoudene, MSc student and Prof. Cristoforo Silvestri

The gut microbiota plays an essential role in maintaining host health. Dysfunction of this symbiotic microbial ecosystem can lead to the development of various human diseases such as inflammatory bowl disease (IBD) [1]. Indeed, the interaction between the host and gut microbiota controls gut homeostasis and inflammatory responses through mechanisms involving the modulation of gut-resident regulatory T cells (Tregs), which express the transcription factor FOXP3 [2].

Prostaglandins (PGs) are bioactive lipid mediators generated from arachidonic acid by cyclooxygenase (COXs) and specific PG synthases that signal in an autocrine and/or paracrine manner. The inflammation mediator PGE2 is present in most tissues at biologically functional nanomolar levels at steady state, and its levels are increased at sites of inflammation [3]. Polymorphisms in the PTGER4 gene that encodes the PGE2 receptor EP4 are associated with its overexpression and a more severe disease phenotype in IBD patients [5]. Tang et al, 2012 revealed that mouse and human intestines express EP4 genes (Ptger4 and PTGER4, respectively) at remarkably higher levels than other PG receptors, indicating that the PGE2 -EP4 pathway may play a more important role than other PG signals in the gut.  Furthermore, hyperactivation of the PGE2 pathway is involved in several inflammatory pathologies by promoting the production of interferon-γ (IFN-γ), T helper 1 (TH1) cells, and interleukin-17 (IL-17)-producing TH17 cells [6].  Crittenden and colleagues set out to determine the role of PGE2 in the control of the accumulation of Tregs in the gut; they report that endogenous PGE2 negatively regulates intestinal Treg responses and intestinal inflammation by affecting the composition of the gut microbiota and modulating mononuclear phagocyte (MNP) function and regulating type I interferon signaling.

Upon performing a detailed analysis of the production of PGs and the expression of their receptors, the authors surmised that the PGE2 -EP4 pathway may play a more important role than other PG signals in the gut. Administration of a non-selective COX inhibitor to mice specifically increased the number RORγt+Foxp3+Tregs, but not RORγt-Foxp3+Tregs, in the colon, and that co-administration of an EP4 agonist counteracted this increase, despite the agonist not being able to decrease RORγt+Foxp3+Tregs in naïve tissues. This indicated that PGE2-EP4 signaling is able to decrease the accumulation of RORγt+Foxp3+Tregs in the colon, which are able to inhibit intestinal inflammation with greater suppressive potential than RORγt-Foxp3+Tregs [7].

Given that it has been reported that gut microbiota regulates the development of intestinal Tregs, particularly the RORγt+Foxp3+Tregs subtype [8], and that use of NSAIDs has been reported to induce changes in the composition of the gut microflora in humans and rodents [9], the above results suggested to the authors that commensal microbiota may be involved in the PGE 2-dependent control of intestinal Tregs. Indeed, they found that in both antibiotic-treated mice and genetically modified mice that have muted responses to microbiota Treg level changes in response to COX inhibition and EP4 agonism were muted. Furthermore, the authors were able to identify changes in the gut microbiota of mice in response to COX inhibition, including an increase in short-chain fatty acid (SCFA)-producing bacteria, which were partially reverted by EP4 agonism.  In conjunction with these changes, the authors also identified alterations in SCFA levels in the cecum of treated mice.  Furthermore, faecal transplantation from treated mice induced similar changes in Treg levels in recipient mice, as had been observed in the donor mice.  These data together support the notion that PGE 2-EP4 signaling reduces intestinal Treg accumulation, at least partially, via the modulation of SCFA-producing microbiota.

Monocyte-derived MNPs mediate the generation of intestinal Foxp3+Tregs by producing soluble mediators such as type I IFNs [10][11]. In fact, upon further investigation, the authors found that PGE2-EP4 modulation of gut microbiota suppresses gut resident MNPs, inhibiting their type I IFN production and signaling and that EP4 signaling in MNPs are responsible for PGE 2 inhibition of colonic RORγt+Tregs.

In conclusion, Crittenden and colleagues have identified a critical role for gut microbiota and MNPs PGE2-EP4-mediated regulation of Treg accumulation within the colon. Their results provide an explanation for the association between PTGER4 (EP4) gene polymorphisms and IBD susceptibility and suggest a potential therapeutic strategy to treat intestinal inflammation by targeting the PGE2-EP4-microbiota-MNP-Treg cascade.

Read the full article.

References

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  7. B.-H. Yang, S. Hagemann, P. Mamareli, U. Lauer, U. Hoffmann, M. Beckstette, L. Föhse, I. Prinz, J. Pezoldt, S. Suerbaum, T. Sparwasser, A. Hamann, S. Floess, J. Huehn, M. Lochner, Foxp3(+) T cells expressing RORt represent a stable regulatory T-cell effector lineage with enhanced suppressive capacity during intestinal inflammation. Mucosal Immunol. 9, 444–457 (2016).
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  1. Y. Yun, H.-N. Kim, S. E. Kim, Y. Chang, S. Ryu, H. Shin, S.-Y. Woo, H.-L. Kim, The effect of probiotics, antibiotics, and antipyretic analgesics on gut microbiota modification. J. Bacteriol. Virol. 47, 64 (2017).
  2. J. L. Coombes, K. R. R. Siddiqui, C. V. Arancibia-Cárcamo, J. Hall, C.-M. Sun, Y. Belkaid, F. Powrie, A functionally specialized population of mucosal CD103+ DCs induces Foxp3+ regulatory T cells via a TGF- and retinoic acid-dependent mechanism. J. Exp. Med. 204, 1757–1764 (2007).
  3. Y. Tanaka, H. Nagashima, K. Bando, L. Lu, A. Ozaki, Y. Morita, S. Fukumoto, N. Ishii, S. Sugawara, Oral CD103−CD11b+ classical dendritic cells present sublingual antigen and induce Foxp3+ regulatory T cells in draining lymph nodes. Mucosal Immunol. 10, 79–90 (2017).

February 15, 2022

-Review by Nayudu Nallabelli, PhD candidate and Prof. Vincenzo Di Marzo.

Atherosclerosis is a major health threat, characterized by the formation of a fibrous plaque or atheroma in the arterial intima, and with a close association with aging. Aging is an individual risk for the development of atherosclerosis1, since it increases the expression of cell adhesion molecules and proinflammatory cytokines, which in turn contribute to atherogenic inflammation. Aging-induced chronic systemic inflammatory responses play an important role in the development of atherosclerosis (AS) but their precise mechanisms are not known. Aged individuals are prone to develop inflammation, which increases levels of proinflammatory markers and the condition known as inflammaging2.

Several mechanisms such as cellular senescence, genetic predisposition to diseases, NLRP3 inflammasome activation, oxidative stress, central obesity, increased gut permeability and changes of microbiota composition contribute to inflammaging3. The imbalance of homeostasis between bacteria and the host can lead to a disease state, resulting in the accumulation of toxic substances and reduction in short-chain fatty acids (SCFAs). Aging is a risk factor for many diseases such as cancer, cardiovascular disease, diabetes, and neurodegenerative disorders4. Therefore, studying the role of aging in AS is key to understanding the underlying metabolic changes during atherosclerotic progression. The development and progression of AS is a complex process regulated by many factors, including arachidonic acid-derived lipid mediators5.

In the study by Sun et al., a group of young (five-week-old, YM) and aged (32-week-old, OM) male apoE-/- mice with a high-fat diet (HFD) were used as models, and age-matched male wild-type C57BL/6J (WT) mice used as controls (YC and OC). ApoE knockout mice have been created by homologous recombination in embryonic stem (ES) cells6. Aged apoE-/- mice suffered from severe AS lesions compared to their younger counterparts and exhibited increased and decreased levels of lipopolysaccharide (LPS) and SCFA production, respectively.

The authors analysed SCFA production as well as the gut microbiota profile from fecal samples, and serum was used to study the lipid profile and the levels of inflammatory cytokines. Cytokines were measured using multiplex kits and AS lesions and atherosclerotic plaque areas were analysed by high frequency ultrasound technology and Oil Red O and HE Staining. The results demonstrated that the OM and YM groups showed elevated levels of TG and LDL cholesterol compared to aged matched controls, while HDL cholesterol levels were significantly lower. Serum levels of pro-inflammatory cytokines, including GM-CSF, IFN-g, IL-7, and TNF-a, were significantly higher in the OM and YM groups than those in the YC and OC groups, while anti-inflammatory cytokine IL-10 levels were significantly decreased in aged mice.

Disturbed gut microflora and decreased SCFA levels could induce a systemic inflammatory response through the activation of the intestinal immune system7. Short-chain fatty acid (SCFA)-producing bacteria such as Bacteriodes decreased in apoE-/- mice. SCFAs play an important role in maintaining intestinal mucosal barrier integrity. Gut flora alterations and metabolite changes might be associated with inflammaging during the advancement of AS. Interestingly, there was a significant decrease in the quantities of beneficial microbes belonging to the families Ruminococcaceae-UCG-014, and an increase in Lachnospiraceae-FCS020, Ruminococcaceae UCG-009, Acetatifactor, Lachnoclostridium, and Lactobacillus gasseri in the feces of aged mice with respect to their younger counterparts.

Decreased intestinal bacteria such as Ruminococcaceae-UCG-014 were negatively correlated with the production of inflammatory factors and LPS in serum, whereas increased intestinal bacteria Lachnospiraceae-FCS020, Ruminococcaceae-UCG-009, Acetatifactor, Lachnoclostridium, and Lactobacillus gasseri were positively correlated with these factors8. A HFD can induce an imbalance of the intestinal flora, especially an increase in the abundance of endotoxin-producing Desulfovibrio, which damages gut barrier function and results in high levels of circulating LPS. An aging-induced imbalance of intestinal flora seems thus closely related to the development of AS.

The serum metabolome results showed that arachidonic acid (AA) metabolism was markedly elevated in aged AS mice compared with their younger counterparts. AA metabolites through different pathways, such as LTB4, PGF2a, and 20-HETE, were significantly increased in the serum of aged AS mice compared their younger counterparts. Thus, aging causes arachidonic acid metabolic dysfunction that potentially exacerbates the progression of AS, and further alterations in gut microbes and metabolites may contribute to inflammatory responses and dysregulated immune function in aged AS mice.

This study reveals the impact of aging on changes in the gut microflora and metabolic profiles, and its association in the progression of atherosclerosis. Aged AS mice showed more severe AS lesions and elevated inflammatory state than their younger AS mice. These findings may provide new insights for how to delay AS development in aged patients. However, additional studies would be needed to prove the link between gut microflora and atherosclerosis mechanistically.

Read the full article.

References

  1. Wang, J. C. & Bennett, M. Aging and atherosclerosis: mechanisms, functional consequences, and potential therapeutics for cellular senescence. Circ. Res. 111, 245–259 (2012).
  2. Rea IM, Gibson DS, McGilligan V, McNerlan SE, Alexander HD, Ross OA. Age and age-related diseases: role of inflammation triggers and cytokines. Front Immunol. 2018; 9:586.
  3. Grebe, A., Hoss, F., and Latz, E. (2018). NLRP3 Inflammasome and the IL-1 Pathway in Atherosclerosis. Circ. Res. 122 (12), 1722–1740. doi: 10.1161/ CIRCRESAHA.118.311362
  4. Niccoli, T., and L. Partridge. 2012. Ageing as a risk factor for disease. Curr. Biol. 22:R741–R752. https://doi.org/10.1016/j.cub.2012.07.024
  5. Huang CC, Chang MT, Leu HB, Yin WH, Tseng WK, Wu YW, et al. Association of arachidonic acid-derived lipid mediators with subsequent onset of acute myocardial infarction in patients with coronary artery disease. Sci Rep. 2020;10.
  6. Plump, A. S. et al. Severe hypercholesterolemia and atherosclerosis in apolipoprotein E-deficient mice created by homologous recombination in ES cells. Cell 71, 343–353 (1992).
  7. Bartolomaeus H, Balogh A, Yakoub M, et al.. The short-chain fatty acid propionate protects from hypertensive cardiovascular damage [published online December 4, 2018].Circulation. doi: 10.1161/CIRCULATIONAHA.118.036652
  8. Kazemian N, Mahmoudi M, Halperin F, Wu JC, Pakpour S. Gut microbiota and cardiovascular disease: opportunities and challenges. Microbiome. 2020;8(1):1–17.

February 10, 2022.

-Review by Fredy Alexander Guevara Agudelo, PhD candidate and Prof. Frédéric Raymond.

The endocannabinoid system (ECS) is composed of signaling lipids known as endocannabinoids, which contribute to the regulation of various processes in the body, such as immune response, communication between cells, appetite, and metabolism, among others 1. Endocannabinoid production occurs through the lipolysis of host cell membrane phospholipids. Endocannabinoids activate G-protein-coupled receptors CB1 and CB2 to stimulate a cellular response. A set of host membrane associated enzymes degrade endocannabinoids to terminate the signaling 2.

In addition to the multiple functions in which the ECS is involved, studies have demonstrated interactions with the gut microbiome 3. Recent studies have shown that enzymes used for the degradation of endocannabinoids also existed in bacterial genomes and could hydrolyze 2-arachidonoyl glycerol (2-AG) in vitro 4. Thus, bacteria could have the potential to respond to endocannabinoids in the intestinal environment, which could influence the severity of the infection. Through a descriptive and mechanistic approach, Melissa Ellermann and collaborators reported that elevated levels of the endocannabinoid 2-AG were associated to attenuated disease infection of enterohemorrhagic Citrobacter rodentium (CR). Their work also investigated how 2-AG directly modulated the virulence of pathogenic Enterobacteriaceae.

For this purpose, the authors tested the susceptibility to CR infection of mice with elevated 2-AG. The enzyme monoacylglycerol lipase (Mgll) degrades 2-AG into arachidonic acid (AA) and glycerol. Knockout mice for the enzyme Mgll (Mgll KO) exhibit increased 2-AG levels in various organs 5. These Mgll KO mice developed significantly attenuated intestinal disease in response to CR. The attenuated pathology observed in Mgll KO mice corresponded with decreased colonic expression of some pro-inflammatory cytokines, such as Nos2, Lcn2, and Mip2a, which positively correlated with CR disease progression and severity. These results showed that Mgll KO mice developed less severe intestinal disease in response to CR infection.

With these results, the authors established the hypothesis that 2-AG mediated protective effects against CR infection in Mgll KO mice. However, since other monoacylglycerols (MAG) can serve as substrates for Mgll, genetic disruption of Mgll could potentially alter the profile of other biologically active lipids in addition to 2-AG 6. Reduced hydrolysis of 2-AG by Mgll may decrease AA generation in certain tissues, which in turn modulates inflammation. Therefore, they also evaluated whether genetic elimination of Mgll altered AA concentration in the colon. The authors found that AA was not increased in the colon of Mgll KO mice, even in mice infected with CR. Colonic levels of other detectable MAGs also remained unchanged in PBS-treated or infected Mgll wild-type (WT) and KO mice. These suggest that Mgll deficiency primarily alters colonic 2-AG levels and does not impact the levels of AA or other MAGs in the colon. Thus, elevated 2-AG likely mediated the protective effects against enteric infection observed in Mgll KO mice.

The authors verify if this process could affect pathogen virulence, which could potentially aggravate inflammation and disease. The authors studied whether the reduced pathology observed in infected Mgll KO mice corresponded with reduced intestinal CR loads. CR fecal loads were significantly reduced in Mgll KO mice relative to WT controls. Sex seemed to contribute to the variability in fecal elimination of CR and tissue colonization in Mgll KO mice. A higher proportion of female Mgll KO mice were culture-negative for CR compared to males, which corresponded with a tendency for increased 2-AG levels in the colon of females. This suggested that Mgll KO mice with elevated 2-AG levels eliminated CR infection earlier than Mgll WT mice, which would coincide with the mitigation of inflammation and pathology observed in Mgll KO mice.

Melissa Ellermann and collaborators then investigated whether Mgll KO mice showed changes in immune function. Deletion of Mgll did alter the expression of some immune markers in the colon of untreated adult mice, in particular Lcn2, Il10 and Il12b, which play a role in modulating the host response to enteric infection. Therefore, they employed a pharmacological inhibitor JZL184 to increase 2-AG levels in the colon of WT mice, as previously reported by Taschier et.al 7. Similar to Mgll KO mice, cecal CR loads decreased in JZL184-treated WT mice compared to vehicle controls, which corresponded to attenuated cecal pathology. This suggested that, as a result of Mgll dysfunction, increased 2-AG attenuates CR infection, corresponding with faster pathogen clearance and attenuated disease.

The 2-AG endocannabinoid signaling is manifested through activation of host cannabinoid CB1 or CB2 receptors, which stimulates intracellular signaling that modulates gastrointestinal physiology 7. However, chronic Mgll blockade results in CB1 receptor inactivation. To this aim, the authors studied whether CB2 blockade with the inhibitor AM630 could reverse the protective effects of JZL184 observed with CR infection. Fecal loads of CR, colonization of cecal tissues, and corresponding cecal pathology were comparable between WT mice with JZL184 or JZL184 and AM630 together. These results suggest that the protective effects of 2-AG on CR infection are not mediated by host CB2 receptor participation.

They then assessed whether compositional or functional changes in the microbiota could explain the reduced CR infection observed in Mgll KO mice. Community profiling of the microbiota revealed that before CR infection, the microbiota between WT and Mgll KO mice did not differ significantly in composition. To determine whether functional differences in the microbiota could occur as a result of the high levels of 2-AG, the authors performed fecal microbiota transplantation (FMT) experiments. Germ-free WT recipient mice were transfected with Mgll WT or KO donor microbiota and then subjected to CR infection. Pathogen loads in feces and colon tissues did not differ between Mgll WT or KO MFT recipients. Taken together, these results suggest that the microbiota is unlikely to be involved in the more rapid CR elimination and disease attenuation observed in Mgll KO mice.

By contrast to the finding of the non-difference of the composition of microbiota between WT and Mgll KO in an infection model, Dione and collaborators observed that Mgll KO mice exhibited a different gut microbiome profile compared to WT mice, both under a normal chow diet and, more significantly, following an obesogenic high fat diet (HFD). According to their results, mice lacking the Mgll enzyme showed an altered gut microbiome that could be linked with their obesity-resistant phenotype. Roseburia increased significantly in abundance only in the WT mice after 8 weeks on the HFD and then returned to the baseline after 22 weeks. Lactobacillaceae increased in WT after 22 weeks on the HFD. Ruminococcaceae and Lachnospiraceae increased in WT mice, but only at 8 weeks, while levels in Mgll KO mice remained constant. In contrast, Prevotellaceae remained unchanged in WT mice but were significantly decreased by HFD in Mgll KO mice. Microbial families differentially abundant in Mgll KO mice were similarly affected in cultures supplemented with 2-AG, providing further evidence for the potential impact of 2-AG on the microbiome 5.

Pharmacological blockade of Mgll resulted in the accumulation of 2-AG in colon tissue 8. Mice with elevated 2-AG levels were less susceptible to CR infection. Specifically, increased host 2-AG levels promoted earlier elimination of the pathogen while minimally impacting its initial establishment. This effect was most pronounced in female Mgll KO mice, which interestingly showed a trend toward increased 2-AG in the colon compared to males. Sex-dependent differences in ECS have been described previously 9. The authors reported observations of sexual differences with ECS manipulations in the context of colitis as a novel finding 8.

Taken together, the findings presented in this study introduce the potential for larger effects of 2-AG in modulating bacterial function. Besides, 2-AG may also modulate other aspects of bacterial function that affect host-commensal interactions and host susceptibility to disease. Modulation of host immune function by 2-AG may be especially important during opportunistic infections. Future studies are needed to unravel the complexity of these interactions.

Read the full article.

References

  1. Silvestri, C. & Di Marzo, V. The endocannabinoid system in energy homeostasis and the etiopathology of metabolic disorders. Cell Metab. 17, 475–490 (2013).
  2. Blankman, J. L., Simon, G. M. & Cravatt, B. F. A Comprehensive Profile of Brain Enzymes that Hydrolyze the Endocannabinoid 2-Arachidonoylglycerol. Chem. Biol. 14, 1347–1356 (2007).
  3. Cani, P. D. et al. Endocannabinoids — at the crossroads between the gut microbiota and host metabolism. Nat. Rev. Endocrinol. 12, 133–143 (2016).
  4. Aman, K., Vanessa, S. & Arturo, C. Indole Signaling at the Host-Microbiota-Pathogen Interface. MBio 10, e01031-19 (2021).
  5. Dione, N. et al. Mgll Knockout Mouse Resistance to Diet-Induced Dysmetabolism Is Associated with Altered Gut Microbiota. Cells 9, (2020).
  6. Nomura, D. K. et al. Monoacylglycerol Lipase Regulates a Fatty Acid Network that Promotes Cancer Pathogenesis. Cell 140, 49–61 (2010).
  7. Taschler, U., Hasenoehrl, C., Storr, M. & Schicho, R. Cannabinoid Receptors in Regulating the GI Tract: Experimental Evidence and Therapeutic Relevance BT - Gastrointestinal Pharmacology. in (ed. Greenwood-Van Meerveld, B.) 343–362 (Springer International Publishing, 2017). doi:10.1007/164_2016_105.
  8. Ellermann, M. et al. Endocannabinoids Inhibit the Induction of Virulence in Enteric Pathogens. Cell 183, 650-665.e15 (2020).
  9. Wagner, E. J. Sex differences in cannabinoid-regulated biology: A focus on energy homeostasis. Front. Neuroendocrinol. 40, 101–109 (2016).

 

 

 

 

 

February 9, 2022

-Review by Sophie Castonguay-Paradis, PhD candidate and Prof. Alain Veilleux.

The metabolic health benefits of the Mediterranean diet (MED) are well documented. Many studies agree on the many health benefits provided by consuming this type of diet (1). Med diet is mainly composed of fruits, vegetables, cereal grains, nuts, legumes, olive oil and includes a moderate consumption of fish and other meats, dairy products and red wine (2). The macronutrients thus consumed are complex carbohydrates, unsaturated fats and vegetable proteins. On the other hand, the "Western" diet, which favours refined carbohydrates, saturated fats and animal proteins, has been shown to be detrimental to health (3). Although there is no doubt about the benefits of the MED diet, the precise mechanisms involved in its positive effects on metabolic health remain to be explored. One hypothesis is that the gut microbiota and the endocannabinoidome, known as the gut microbiome-endocannabinoidome axis, may play a role in this relationship. The endocannabinoidome include a large family of lipid molecules called endocannabinoidome mediators that collectively play an important role in homeostasis. The intake of certain dietary components, including fatty acids, is associated with an increase of the circulating profile of these mediators (4-6). The composition of the gut microbiota is also strongly influenced by the host's diet. It is therefore crucial to better understand the link between the metabolic benefits of the MED diet, the gut microbiota and the endocannabinoidome mediators.

Tagliamonte et al. examined changes in gut microbiota composition and circulating endocannabinoidome mediator levels in response to MED diet consumption (7).

The randomized clinical trial was conducted over 8 weeks in which 82 overweight or obese participants were asked to consume either their usual diet (control group) or an isocaloric MED diet. Adherence to the diet was measured every 2 weeks using food diaries while plasma and stool samples were collected at weeks 0, 4 and 8.

Results showed changes in the microbiome-endocannabinoidome axis following MED diet consumption. Indeed, circulating levels of anandamide (AEA) were decreased, whereas the ratios of N‑oleoyl‑ethanolamine (OEA)/N‑palmitoyl‑ethanolamine (PEA) and OEA/AEA were increased in the MED diet group. The composition of the gut microbiota was altered in the MED diet group showing an increase in fiber metabolizing bacteria such as Faecalibacterium prausnitzii and several members of the Lachnospiraceae family. The relative abundance of Akkermansia muciniphila, known to be inversely related to obesity and its complications (8) was also increased following consumption of the MED diet. Interestingly, changes in gut microbiota composition were proportional to MED diet adherence. AEA/PEA ratio was also associated with beneficial changes in plasma cholesterol levels and insulin resistance. Furthermore, depending on the initial composition of the gut microbiota, some individuals also positive effects on insulin resistance and inflammatory status, suggesting that some individuals would be more likely to respond favourably to nutritional interventions.

Several food groups were strongly associated with NAEs (AEA, OEA, PEA, N‑linoleoyl‑ethanolamine (LEA)) as well as 2-arachidonoyl-glycerol (2-AG). However, food groups that define the MED diet (e.g., legumes and fish) and those that define the "Western" diet (e.g., meats and sugar-sweetened beverages) were positively associated with NAEs, which seems quite contradictory but was not addressed by the authors.

Considering that the study was aimed at self-modification of the diet, an important limitation is that not all participants achieved the same degree of adherence to the MED diet. Although this limitation is inherent to nutritional studies, it would have been appropriate to better coach participants or to provide them more MED diet food components, especially fresh foods, to decrease interindividual variability.

This study demonstrates the key role of the endocannabinoidome and the gut microbiota in the relationship between diet and gut health. This relationship is very important considering that it could be a step towards personalize nutrition.

Read the full article.

References

  1. Estruch R, Ros E, Salas-Salvadó J, Covas M-I, Corella D, Arós F, Gómez-Gracia E, Ruiz-Gutiérrez V, Fiol M, Lapetra J, et al. Primary Prevention of Cardiovascular Disease with a Mediterranean Diet Supplemented with Extra-Virgin Olive Oil or Nuts. N Engl J Med 2018;378:e34.
  2. Davis C, Bryan J, Hodgson J, Murphy K. Definition of the Mediterranean Diet; A Literature Review. Nutrients Multidisciplinary Digital Publishing Institute; 2015;7:9139–53.
  3. Zinöcker MK, Lindseth IA. The Western Diet–Microbiome-Host Interaction and Its Role in Metabolic Disease. Nutrients Multidisciplinary Digital Publishing Institute; 2018;10:365.
  4. Castonguay-Paradis S, Lacroix S, Rochefort G, Parent L, Perron J, Martin C, Lamarche B, Raymond F, Flamand N, Di Marzo V, et al. Dietary fatty acid intake and gut microbiota determine circulating endocannabinoidome signaling beyond the effect of body fat. Sci Rep 2020;10:15975.
  5. Banni S, Carta G, Murru E, Cordeddu L, Giordano E, Sirigu AR, Berge K, Vik H, Maki KC, Di Marzo V, et coll. Krill oil significantly decreases 2-arachidonoylglycerol plasma levels in obese subjects. Nutr Metab (Lond) 2011;8:7.
  6. Pu S, Eck P, Jenkins DJA, Connelly PW, Lamarche B, Kris-Etherton PM, West SG, Liu X, Jones PJH. Interactions between dietary oil treatments and genetic variants modulate fatty acid ethanolamides in plasma and body weight composition. British Journal of Nutrition 2016;115:1012–23.
  7. Tagliamonte S, Laiola M, Ferracane R, Vitale M, Gallo MA, Meslier V, Pons N, Ercolini D, Vitaglione P. Mediterranean diet consumption affects the endocannabinoid system in overweight and obese subjects: possible links with gut microbiome, insulin resistance and inflammation. Eur J Nutr 2021
  8. Cani PD, de Vos WM. Next-Generation Beneficial Microbes: The Case of Akkermansia muciniphila. Front Microbiol 2017;8.

 

February 7, 2022

-Review by Briscia Anaid Tinoco Mar, PhD candidate and Prof. Vincenzo Di Marzo.

The intestinal microbiota represents a central regulator of host metabolic processes. Among its functions, the microbiota transforms dietary components not digestible by host enzymes into bioavailable compounds and enables their metabolism1. This process includes, among others, depolymerization or fermentation of complex carbohydrates on the intestinal lumen, thereby producing short-chain fatty acids (SCFA) such as acetate (FA 2:0), propionate (FA 3:0), and butyrate (FA 4:0)2. Once produced, SCFA pass through the intestinal epithelium, reach the liver via portal, are transformed, and continue their function. However, SCFA per se also play signaling functions in the host3.

SCFA production relies on the gut microbiota composition, which largely depends on the host's diet and habits. Therefore, dietary changes, antibiotic use, stress, and specific pathologies result in a dysbiosis, leading to SCFA levels variation4,5.

Previously, it was established that gut microbiota and SCFA production are involved in both hepatic lipid metabolism and blood lipid levels6.

However, to maintain its integrity, metabolism and function, the intestinal epithelium employs SCFA as an energetic substrate7. Liebisch et al. (2021) set out to determine the microbiota's effects on the intestinal lipidome. Their main objective was to provide quantitative lipidomic data from the small and large intestine tissue in germ-free mice (GF) and specific pathogen-free (SPF) mice. For this purpose, the use of GF provides a scenario that mimics the absence of any microorganism in the body, whereas SPF are free of a specific list of disease-causing pathogens and other opportunistic microorganisms8–11. With their work, Liebisch et al. revealed significant differences in free cholesterol content in the colon and ileum, and differences in glycerophospholipid species proportions, between GF and SPF mice. Thus, the presence of microbiota might increase colon membrane fluidity by modifying these components11.

Additionally, the authors found that microbiota inhibit polyunsaturated acid (PUFA) metabolism since GF exhibited increased PUFAs species, such as arachidonic acid (FA 20: n-6) and docosahexaenoic acid (FA 22:6 n-3), and low levels of palmitic (FA 16:0), palmitoleic (FA 16:1) and oleic (FA 18:1 n-9) acids in the ileum. In addition, GF mice showed an increase in the expression of genes relevant for PUFA elongation and desaturation, a condition found in several pathologies12,13. These results are very relevant to the research activities of CERC-MEND, since fatty acids, and PUFA in particular, are a strong determinant of the endogenous levels of endocannabinoidome mediators, which play several roles in regulating host function14.

Together, these findings provide a big opportunity for further research on the intestinal lipidome. It is indispensable to determine the effects on intestinal lipid metabolism under those conditions already known to affect SCFA production (e.g., HFHS diets, high fiber, polyphenol supplementation1,2) and other pathologies (e.g. obesity, diabetes)15,16. The use of cell cultures and, particularly, intestinal organoid would be a formidable tool to determine mechanisms involved in the effect of SCFA on intestinal lipid metabolism17.

Read the full article.

References

  1. Kawabata, K., Yoshioka, Y. & Terao, J. Role of intestinal microbiota in the bioavailability and physiological functions of dietary polyphenols. Molecules 24, 370 (2019).
  2. Bishehsari, F. et al. Dietary fiber treatment corrects the composition of gut microbiota, promotes SCFA production, and suppresses colon carcinogenesis. Genes 9, (2018).
  3. Mirzaei, R. et al. Dual role of microbiota-derived short-chain fatty acids on host and pathogen. Biomed. Pharmacother. 145, 112352 (2022).
  4. Rau, M. et al. Fecal SCFAs and SCFA‐producing bacteria in gut microbiome of human NAFLD as a putative link to systemic T‐cell activation and advanced disease. United Eur. Gastroenterol. J. 6, 1496–1507 (2018).
  5. De la Cuesta-Zuluaga, J. et al. Higher fecal short-chain fatty acid levels are associated with gut microbiome dysbiosis, obesity, hypertension and cardiometabolic disease risk factors. Nutrients 11, (2019).
  6. LeBlanc, J. G. et al. Beneficial effects on host energy metabolism of short-chain fatty acids and vitamins produced by commensal and probiotic bacteria. Microb. Cell Factories 16, 79 (2017).
  7. Parada Venegas, D. et al. Short Chain Fatty Acids (SCFAs)-mediated gut epithelial and immune regulation and its relevance for inflammatory bowel diseases. Front. Immunol. 10, 277 (2019).
  8. Manca, C. et al. Germ-free mice exhibit profound gut microbiota-dependent alterations of intestinal endocannabinoidome signaling. J. Lipid Res. 61, 70–85 (2020).
  9. Luczynski, P. et al. Growing up in a bubble: using germ-free animals to assess the influence of the gut microbiota on brain and behavior. Int. J. Neuropsychopharmacol. 19, pyw020 (2016).
  10. Uzbay, T. Germ-free animal experiments in the gut microbiota studies. Curr. Opin. Pharmacol. 49, 6–10 (2019).
  11. Liebisch, G., Plagge, J., Höring, M., Seeliger, C. & Ecker, J. The effect of gut microbiota on the intestinal lipidome of mice. Int. J. Med. Microbiol. 311, 151488 (2021).
  12. Resch, C. et al. The Influence of diet and sex on the gut microbiota of lean and obese JCR:LA-cp rats. Microorganisms 9, (2021).
  13. Wenzel, T. J., Gates, E. J., Ranger, A. L. & Klegeris, A. Short-chain fatty acids (SCFAs) alone or in combination regulate select immune functions of microglia-like cells. Mol. Cell. Neurosci. 105, 103493 (2020).
  14. Castonguay-Paradis, S. et al. Dietary fatty acid intake and gut microbiota determine circulating endocannabinoidome signaling beyond the effect of body fat. Sci. Rep. 10, 15975 (2020).
  15. Veilleux, A. et al. Intestinal lipid handling: evidence and implication of insulin signaling abnormalities in human obese subjects. Arterioscler. Thromb. Vasc. Biol. 34, 644–653 (2014).
  16. Veilleux, A. et al. Altered intestinal functions and increased local inflammation in insulin-resistant obese subjects: a gene-expression profile analysis. BMC Gastroenterol. 15, 119 (2015).
  17. Pearce, S. C. et al. Intestinal enteroids recapitulate the effects of short-chain fatty acids on the intestinal epithelium. PLOS ONE 15, e0230231 (2020).

 

September 27, 2021

-Review by Isabelle Bourdeau-Julien, PhD candidate and Prof. Frédéric Raymond.

The gut shelters billions of microorganisms consuming and rejecting different molecules that are absorbed and directed to the bloodstream [1]. The importance of the contribution of the gut microbiota to host metabolism is well recognized, but the mechanisms regulating this interaction is still poorly understood [2]. Indeed, the large variety of bacterial metabolic functions and cross-feeding interactions between bacteria makes the gut microbiota hard to study [3]. Using a mechanistic approach, Cohen and colleagues report that the gut microbiota would affect host metabolism through GPCR activity [4].

G-protein coupled receptors (GPCRs) are the largest family membrane receptors in eukaryote and the most studied drug targets. GPCR ligands include N-acyl amides, a class of molecules with numerous possible combinations of amine head groups and acyl tails. They modulate a large variety of metabolic functions involved in glucose metabolism, inflammation, lipid metabolism, neuronal activity, satiety, appetite, gastrointestinal motility, etc. [5, 6]. Their importance for the metabolism is also demonstrated by their implication in many diseases such as diabetes, obesity, cancer, inflammatory bowel disease and others [7]. Moreover, it was reported that, in 2017, clinical trials for new drugs targeting GPCRs were mostly aimed at obesity and diabetes [8].

Recent evidence suggests that GPCR and bioactive lipid metabolism would be important for the interaction between the gut microbiota and host metabolism. Among GPCR ligands, endocannabinoidome mediators has been strongly associated to the gut microbiota and metabolic diseases [9, 10]. Also, commendamide is a long-chain N-acyl amide produced by gut bacteria that can interact with GPR132 (G2A) [11]. Cohen and colleagues provide further evidence by finding gut microbiota N-acyl amides that interact with eukaryotic G-protein coupled receptors.

First, they used bioinformatic analysis of the human microbiota sequencing data to identify potential GPCR-active N-acyl amides encoded by gut microorganisms. With a BLASTN search of N-acyl synthase (NAS) genes in the Human Microbiome Project, they identified 143 human microbial (hm) NAS genes. Out of the 44 hm-NAS tested in E. Coli cultures, 31 produced N-acyl amides that could be grouped in six families based on the amine head group and the fatty acid tail; (1) N-acyl glycine; (2) N-acyloxyacyl lysine; (3) N-acyloxyacyl glutamine; (4) N-acyl lysine/ornithine; (5) N-acyl alanine; (6) N-acyl serinol.

Looking at their distribution in different body sites, they observed an enrichment of hm-NAS genes in bacteria of the gastrointestinal tract. Even within the gastrointestinal tract, some region harbors different patterns of hm-NAS genes corresponding to specific N-acyl amide families. For example, in stool samples, genes encoding for N-acyl glycine (1) are highly enriched compared to the other N-acyl amide families. Although their expression level differs between individuals, most hm-NAS genes are found in 90% of individuals except for the genes responsible for the production of the N-acyl amides from two families (3 and 5) which are very little or not detected. Thus, NAS-genes are highly prevalent in human gut microbiome.

Then, to validate the potential of the bacterial N-acyl amide to interact with GPCRs, the major metabolites of each gene families were assayed for agonist and antagonist activity against 240 human GPCRs. Strong and specific agonist interactions were observed for N-palmitoyl serinol (6) with GPR119, N-3-hydroxypalmitoyl ornithine (4) with S1PR4 and N-myristoyl alanine (5) with GPR132 (G2A). Specific antagonist interaction was observed for N-acyloxyacyl glutamine (3) with prostaglandin receptor PTGIR and PTGER4 was antagonized by some N-acyl amides including N-acyloxyacyl glutamine (3). Thus, in addition to having hm-NAS genes, the bacterial N-acyl amide produced can interact with GPCRs.

Bacterial N-acyl amides have structural and functional similarities with endogenous human GPCR ligands. The clearest overlap is observed between OEA and 2-OG, ligands for the endocannabinoid receptor GPR119, and N-oleoyl serinol (6). Beyond structural similarities, Cohen and al. show the bacterial ligand N-oleoyl serinol (6) induce a higher activation of GPR119 and GLP-1 secretion by GLUTag cells than the human ligands. In mice, colonization with E.coli producing N-oleoyl serinols (6) decreased blood glucose and increased secretion of the hormones GLP-1 and insulin. Thus, GPR119 bacterial agonist can regulate metabolic hormones and glucose homeostasis as efficiently as human ligands.

Cohen and colleagues suggest that the relationship between the gut microbiota and host metabolism would be regulated through GPCR-associated signaling. Indeed, the authors provide strong evidence for the potential of the gut microbiota to modulate many physiological processes through the production of GPCR ligands. However, there are some limitations to the study. First, by looking at the gene abundance of hm-NAS genes from the 6 main families in different body sites, the N-acyl serinol (6) family was not detected in stool samples. Yet, the metabolite interacting with GPR119 and regulating metabolic hormones and glucose homeostasis as efficiently as human ligands is part of the N-acyl serinol (6) family. Thus, further studies will be needed to investigate which hm-NAS genes are present in human stool bacteria, their expression level, and the presence of N-acyl amides. Also, as the authors have pointed out, it would be interesting to look at co-localization of GPCR with hm-NAS gene expression in gastrointestinal niches to confirm the potential interactions.

The authors suggest a commensal relationship between gut microbiota and the host where the beneficial bacteria evolved to mimic our signaling molecules. The relationship between the gut microbiota and the host can be seen a mutualism interaction, being beneficial for both the bacteria and the host. Furthermore, organisms from mutualistic relationships tend to co-evolve [12]. In fact, several evidence show that the gut microbiota and the host co-evolved, among others with the immune system [13]. Therefore, we can hypothesize that humans have evolved to express GPCRs interacting with bacterial ligands in tissues where the microbiota is abundant. Also, bacteria use metabolites for communication between microorganisms. Among these, N-acyl-homoserine lactones (AHLs) are used for quorum-sensing and are detected by LuxR family proteins in prokaryote [14]. As bacterial N-acyl amides have structural similarities with AHLs, they could be used for bacterial communication. It would be useful to test the microorganism’s response to bacterial N-acyl amides. It would also be interesting to compare the presence and expression of genes responsible for N-acyl amide production in the bacteria of the microbiota to bacteria from other non-human associated ecosystems. In this regard, Cohen and colleagues point out that the bacterial metabolites N-acyl ornithine, lysine and glutamines are natural produced by soil bacteria [15, 16]. In plants, it has been shown that bacterial quorum-sensing AHLs regulate Arabidopsis root growth through two receptors, whose expression level is dependent on the bacterial metabolites [17]. This way, the interaction of the gut microbiota and human host through GPCR metabolism could be the result of co-evolution. Although, an important research effort would be necessary to test this hypothesis.

Read the full article.

References

[1]       K. Oliphant and E. Allen-Vercoe, "Macronutrient metabolism by the human gut microbiome: major fermentation by-products and their impact on host health," (in eng), Microbiome, vol. 7, no. 1, p. 91, 06 2019, doi: 10.1186/s40168-019-0704-8.

[2]       Z. Y. Kho and S. K. Lal, "The Human Gut Microbiome - A Potential Controller of Wellness and Disease," (in eng), Front Microbiol, vol. 9, p. 1835, 2018, doi: 10.3389/fmicb.2018.01835.

[3]       D. Ríos-Covián, P. Ruas-Madiedo, A. Margolles, M. Gueimonde, C. G. de Los Reyes-Gavilán, and N. Salazar, "Intestinal Short Chain Fatty Acids and their Link with Diet and Human Health," (in eng), Front Microbiol, vol. 7, p. 185, 2016, doi: 10.3389/fmicb.2016.00185.

[4]       L. J. Cohen et al., "Commensal bacteria make GPCR ligands that mimic human signalling molecules," (in eng), Nature, vol. 549, no. 7670, pp. 48-53, 09 2017, doi: 10.1038/nature23874.

[5]       S. Lacroix et al., "Rapid and Concomitant Gut Microbiota and Endocannabinoidome Response to Diet-Induced Obesity in Mice," (in eng), mSystems, vol. 4, no. 6, Dec 2019, doi: 10.1128/mSystems.00407-19.

[6]       A. S. Husted, M. Trauelsen, O. Rudenko, S. A. Hjorth, and T. W. Schwartz, "GPCR-Mediated Signaling of Metabolites," (in eng), Cell Metab, vol. 25, no. 4, pp. 777-796, Apr 04 2017, doi: 10.1016/j.cmet.2017.03.008.

[7]       P. D. Cani et al., "Endocannabinoids--at the crossroads between the gut microbiota and host metabolism," (in eng), Nat Rev Endocrinol, vol. 12, no. 3, pp. 133-43, Mar 2016, doi: 10.1038/nrendo.2015.211.

[8]       A. S. Hauser, M. M. Attwood, M. Rask-Andersen, H. B. Schiöth, and D. E. Gloriam, "Trends in GPCR drug discovery: new agents, targets and indications," (in eng), Nat Rev Drug Discov, vol. 16, no. 12, pp. 829-842, Dec 2017, doi: 10.1038/nrd.2017.178.

[9]       C. Rousseaux et al., "Lactobacillus acidophilus modulates intestinal pain and induces opioid and cannabinoid receptors," (in eng), Nat Med, vol. 13, no. 1, pp. 35-7, Jan 2007, doi: 10.1038/nm1521.

[10]     V. Di Marzo and C. Silvestri, "Lifestyle and Metabolic Syndrome: Contribution of the Endocannabinoidome," (in eng), Nutrients, vol. 11, no. 8, Aug 20 2019, doi: 10.3390/nu11081956.

[11]     L. J. Cohen et al., "Functional metagenomic discovery of bacterial effectors in the human microbiome and isolation of commendamide, a GPCR G2A/132 agonist," (in eng), Proc Natl Acad Sci U S A, vol. 112, no. 35, pp. E4825-34, Sep 01 2015, doi: 10.1073/pnas.1508737112.

[12]     L. P. Medeiros, G. Garcia, J. N. Thompson, and P. R. Guimarães, "The geographic mosaic of coevolution in mutualistic networks," (in eng), Proc Natl Acad Sci U S A, vol. 115, no. 47, pp. 12017-12022, 11 20 2018, doi: 10.1073/pnas.1809088115.

[13]     H. Y. Cheng, M. X. Ning, D. K. Chen, and W. T. Ma, "Interactions Between the Gut Microbiota and the Host Innate Immune Response Against Pathogens," (in eng), Front Immunol, vol. 10, p. 607, 2019, doi: 10.3389/fimmu.2019.00607.

[14]     L. Keller and M. G. Surette, "Communication in bacteria: an ecological and evolutionary perspective," (in eng), Nat Rev Microbiol, vol. 4, no. 4, pp. 249-58, Apr 2006, doi: 10.1038/nrmicro1383.

[15]     X. Zhang, S. M. Ferguson-Miller, and G. E. Reid, "Characterization of ornithine and glutamine lipids extracted from cell membranes of Rhodobacter sphaeroides," (in eng), J Am Soc Mass Spectrom, vol. 20, no. 2, pp. 198-212, Feb 2009, doi: 10.1016/j.jasms.2008.08.017.

[16]     E. K. Moore et al., "Lysine and novel hydroxylysine lipids in soil bacteria: amino acid membrane lipid response to temperature and pH in Pseudopedobacter saltans," (in eng), Front Microbiol, vol. 6, p. 637, 2015, doi: 10.3389/fmicb.2015.00637.

[17]     G. Jin et al., "Two G-protein-coupled-receptor candidates, Cand2 and Cand7, are involved in Arabidopsis root growth mediated by the bacterial quorum-sensing signals N-acyl-homoserine lactones," (in eng), Biochem Biophys Res Commun, vol. 417, no. 3, pp. 991-5, Jan 20 2012, doi: 10.1016/j.bbrc.2011.12.066.

June 16, 2021

-Review by Volatiana Rakotoarivelo, postdoctoral fellow and Prof. Nicolas Flamand.

During obesity, the morphological changes induced by adipocyte hypertrophy leads to hypoxia and oxidative stress [1-3]. As a result, stressed adipocytes begin to secrete proinflammatory cytokines and chemokines resulting in chronic and low-grade inflammation [4]. This inflammation is induced by activation of the immune system in tissues such as adipose tissue [5], muscle [6], the liver and pancreas [7]. This chronic inflammation is believed to contribute to the development of obesity-related-diseases, such as type 2 diabetes, hypertension and cardiometabolic diseases[4].

Oxylipins (OXLs) are bioactive lipid metabolites derived from polyunsaturated fatty acids (PUFAs) that are implicated in the inflammatory response as well as in the resolution process and play an important role in the establishment of chronic inflammation. The OXLs derived from arachidonic acid (ARA) such as prostaglandins, leukotrienes and thromboxanes may act as proinflammatory mediators whereas lipoxins derived from ARA can be implicated in the resolution of inflammation [10]. OXLs may act as signaling molecules involved in inflammatory processes associated with obesity [8, 9].

At the same time, the gut microbiota plays an important role in obesity and associated diseases. Dysbiosis of the gut microbiota contributes to proinflammatory signalling through pattern recognition receptor (PRR) activation to induce an inflammatory response [11], and other processes.

Recently, Avila-Roman and colleagues described how the composition of the gut microbiota influences plasma OXLs in a manuscript in Clinical Nutrition [12]. In this study, Wistars rats were fed either a standard chow diet (STD) or a hypercaloric cafeteria diet (CAF), which results in the development of metabolic syndrome, for 5 weeks. Two additional groups of rats fed with the STD diet and the CAF diet were treated with a cocktail of antibiotics (ABX) administered in drinking water for the last two weeks.

First, the authors observed that the CAF diet induced obesity and glucose intolerance compared to STD diet. In addition, ABX administration reduced gut microbiota diversity but not CAF-induced obesity/weight gain. ABX administration to either STD or CAF-fed rats resulted in a significant increase of the relative abundance of Proteobacteria, which are associated with inflammation, and a significant decrease in that of Bacteroidetes. In contrast, ABX specifically decreased Actinobacteria, Deferribacteraceae and Verrucomicrobia, which are associated with inflammation in STD-fed rats, while it decreased Spirochaetes in CAF-fed rats.

In addition, the authors observed drastically different profiles of plasma OXLs between the CAF- and STD-fed rats, which could be explained by the nutritional composition of the CAF diet, which was enriched in fat with higher levels of PUFAs. The authors then based their analysis on the diet type; principal coordinate analysis revealed that ABX treatment did not affect the overall profile of OXLs in STD-fed rats but did in CAF-fed rats. However, ABX-dependent changes in specific OXLs in STD-fed rats: an increase in 4-HDHA and 8-HEPE levels and a decrease in 15(R)-Lipoxin A4/A5 levels. In CAF-fed rats, a significant increase in proinflammatory OXLs, (11(12)-DiHETE, 9-HETE, LTB4 and PG D2), as well as in anti-inflammatory OXLs (11-HEPE, 15(S)-HEPE, 10-HDHA and 13-HDHA) were observed in response to ABX.

Finally, the authors correlated the relative abundance of gut bacteria with OXLs levels. Proteobacteria and Bacteroidetes were the major phyla altered by the ABX treatment. Bacteroidetes, showed negative correlations with most of the plasma OXLs (16- HDHA, 8-HEPE, LTB4 and PGD2). It is noteworthy that 16-HDHA and 8-HEPE, which are derived from the w3-PUFAs DHA and EPA respectively, are associated with anti-inflammatory effects. LTB4 and PGD2 are derived from ARA, an omega 6-PUFA linked to proinflammatory effects. Both negative and positive correlations were observed with Proteobacteria and a significant positive correlation with LTB4 was particularly notable.

Based on these observations, Avila-Roman and colleagues propose that dysbiosis of the gut alters plasma OXLs levels in obesity as well as in healthy conditions. They suggested that the gut microbiota may regulate lipid metabolism and affect the inflammatory process mediated by OXLs.  This is the first demonstrated link between microbiota dysbiosis and plasma levels of OXLs. They also propose OXLs and gut microbiota as new biomarkers for chronic low-grade inflammation as well as metabolic profiling.

However, readers should also consider the limitations of the current study. First, it is important remember that the study only found associations, and does not provide evidence for direct causality.  Further studies will be required to elucidate the specific mechanisms involved in the reported apparent alteration in lipid metabolism. In addition, it is important to consider that dysbiosis of the gut microbiota [13], as well as intestinal bioactive lipids [14], regulate intestinal permeability, which is an important parameter in the establishment of inflammation associated with obesity.  Indeed the authors did not assess the inflammatory state of the animals within this study beyond assessing changes in OXL levels.

Finally, the literature remains unclear regarding the use of inflammatory mediators as biomarkers of metabolic diseases. Whereas Hotamisligil et al. proposed TNFa as well as other pro-inflammatory cytokines as being strongly linked to obesity and the development of diabetes [15-18], later this assertion has been repeatedly challenged [19]. Indeed, the comparison between tissue and circulating concentrations of inflammatory mediators remains debated since the chronic inflammation associated with obesity is initiated by the activation of innate immune cells present in tissues, such as resident macrophages in adipose tissue [20]. It is therefore important to assess the importance of bioactive lipids in the activity of metabolic tissues such as adipose tissue, liver or muscle.

Read the full article.

References

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