Photobiomodulation Therapy: A Novel Therapeutic Approach to Alzheimer's Disease Made Possible by the Evidence of a Brain-Gut Interconnection.

The evidence of brain-gut interconnections in Alzheimer's disease (AD) opens novel avenues for the treatment of a pathology for which no definitive treatment exists. Gut microbiota and bacterial translocation may produce peripheral inflammation and immune modulation, contributing to brain amyloidosis, neurodegeneration, and cognitive deficits in AD. The gut microbiota can be used as a potential therapeutic target in AD. In particular, photobiomodulation (PBM) can affect the interaction between the microbiota and the immune system, providing a potential explanation for its restorative properties in AD-associated dysbiosis. PBM is a safe, non-invasive, non-ionizing, and non-thermal therapy that uses red or near-infrared light to stimulate the cytochrome c oxidase (CCO, complex IV), the terminal enzyme of the mitochondrial electron transport chain, resulting in adenosine triphosphate synthesis. The association of the direct application of PBM to the head with an abscopal and a systemic treatment through simultaneous application to the abdomen provides an innovative therapeutic approach to AD by targeting various components of this highly complex pathology. As a hypothesis, PBM might have a significant role in the therapeutic options available for the treatment of AD.

The visceral adipose tissue bacterial microbiota provides a signature of obesity based on inferred metagenomic functions.

BACKGROUND: Metabolic inflammation mediated obesity requires bacterial molecules to trigger immune and adipose cells leading to inflammation and adipose depot development. In addition to the well-established gut microbiota dysbiosis, a leaky gut has been identified in patients with obesity and animal models, characterized by the presence of a tissue microbiota in the adipose fat pads. METHODS: To determine its potential role, we sequenced the bacterial 16 S rRNA genes in the visceral adipose depot of patients with obesity. Taking great care (surgical, biochemical, and bioinformatic) to avoid environmental contaminants. We performed statistical discriminant analyses to identify specific signatures and constructed network of interactions between variables. RESULTS: The data showed that a specific 16SrRNA gene signature was composed of numerous bacterial families discriminating between lean versus patients with obesity and people with severe obesity. The main discriminant families were Burkholderiaceae, Yearsiniaceae, and Xanthomonadaceae, all of which were gram-negative. Interestingly, the Morganellaceae were totally absent from people without obesity while preponderant in all in patients with obesity. To generate hypotheses regarding their potential role, we inferred metabolic pathways from the 16SrRNA gene signatures. We identified several pathways associated with adenosyl-cobalamine previously described to be linked with adipose tissue development. We further identified chorismate biosynthesis, which is involved in aromatic amino-acid metabolism and could play a role in fat pad development. This innovative approach generates novel hypotheses regarding the gut to adipose tissue axis. CONCLUSIONS: This innovative approach generates novel hypotheses regarding the gut to adipose tissue axis in obesity and notably the potential role of tissue microbiota.

Human liver microbiota modeling strategy at the early onset of fibrosis.

BACKGROUND: Gut microbiota is involved in the development of liver diseases such as fibrosis. We and others identified that selected sets of gut bacterial DNA and bacteria translocate to tissues, notably the liver, to establish a non-infectious tissue microbiota composed of microbial DNA and a low frequency live bacteria. However, the precise set of bacterial DNA, and thereby the corresponding taxa associated with the early stages of fibrosis need to be identified. Furthermore, to overcome the impact of different group size and patient origins we adapted innovative statistical approaches. Liver samples with low liver fibrosis scores (F0, F1, F2), to study the early stages of the disease, were collected from Romania(n = 36), Austria(n = 10), Italy(n = 19), and Spain(n = 17). The 16S rRNA gene was sequenced. We considered the frequency, sparsity, unbalanced sample size between cohorts to identify taxonomic profiles and statistical differences. RESULTS: Multivariate analyses, including adapted spectral clustering with L1-penalty fair-discriminant strategies, and predicted metagenomics were used to identify that 50% of liver taxa associated with the early stage fibrosis were Enterobacteriaceae, Pseudomonadaceae, Xanthobacteriaceae and Burkholderiaceae. The Flavobacteriaceae and Xanthobacteriaceae discriminated between F0 and F1. Predicted metagenomics analysis identified that the preQ0 biosynthesis and the potential pathways involving glucoryranose and glycogen degradation were negatively associated with liver fibrosis F1-F2 vs F0. CONCLUSIONS: Without demonstrating causality, our results suggest first a role of bacterial translocation to the liver in the progression of fibrosis, notably at the earliest stages. Second, our statistical approach can identify microbial signatures and overcome issues regarding sample size differences, the impact of environment, and sets of analyses. TRIAL REGISTRATION: TirguMECCH ROLIVER Prospective Cohort for the Identification of Liver Microbiota, registration 4065/2014. Registered 01 01 2014.

Blood Bacterial Profiles Associated With Human Immunodeficiency Virus Infection and Immune Recovery.

Human immunodeficiency virus (HIV) infection impairs mucosal immunity and leads to bacterial translocation, fueling chronic inflammation and disease progression. While this is well established, questions remain about the compositional profile of the translocated bacteria, and to what extent it is influenced by antiretroviral therapy (ART). Using 16S ribosomal DNA targeted sequencing and shotgun proteomics, we showed that HIV increases bacterial translocation from the gut to the blood. HIV increased alpha diversity in the blood, which was dominated by aerobic bacteria belonging to Micrococcaceae (Actinobacteria) and Pseudomonadaceae (Proteobacteria) families, and the number of circulating bacterial proteins was also increased. Forty-eight weeks of ART attenuated this phenomenon. We found that enrichment with Lactobacillales order, and depletion of Actinobacteria class and Moraxellaceae and Corynebacteriacae families, were significantly associated with greater immune recovery and correlated with several inflammatory markers. Our findings suggest that the molecular cross talk between the host and the translocated bacterial products could influence ART-mediated immune recovery.

Changes in blood microbiota profiles associated with liver fibrosis in obese patients: A pilot analysis.

The early detection of liver fibrosis among patients with nonalcoholic fatty liver disease (NAFLD) is an important clinical need. In view of the suggested role played by bacterial translocation in liver disease and obesity, we sought to investigate the relationship between blood microbiota and liver fibrosis (LF) in European cohorts of patients with severe obesity. We carried out a cross-sectional study of obese patients, well characterized with respect to the severity of the NAFLD, in the cohort FLORINASH. This cohort has been divided into a discovery cohort comprising 50 Spanish patients and then in a validation cohort of 71 Italian patients. Blood bacterial DNA was analyzed both quantitatively by 16S ribosomal DNA (rDNA) quantitative polymerase chain reaction and qualitatively by 16S rDNA targeted metagenomic sequencing and functional metagenome prediction. Spanish plasma bile acid contents were analyzed by liquid chromatography/mass spectrometry. The 16S rDNA concentration was significantly higher in patients of the discovery cohort with LF. By 16S sequencing, we found specific differences in the proportion of several bacterial taxa in both blood and feces that correlate with the presence of LF, thus defining a specific signature of the liver disease. Several secondary/primary bile acid ratios were also decreased with LF in the discovery cohort. We confirmed, in the validation cohort, the correlation between blood 16S rDNA concentration and LF, whereas we did not confirm the specific bacterial taxa signature, despite a similar trend in patients with more-severe fibrosis. CONCLUSION: Changes in blood microbiota are associated with LF in obese patients. Blood microbiota analysis provides potential biomarkers for the detection of LF in this population. (Hepatology 2016;64:2015-2027).

Comprehensive description of blood microbiome from healthy donors assessed by 16S targeted metagenomic sequencing.

BACKGROUND: Recent studies have revealed that the blood of healthy humans is not as sterile as previously supposed. The objective of this study was to provide a comprehensive description of the microbiome present in different fractions of the blood of healthy individuals. STUDY DESIGN AND METHODS: The study was conducted in 30 healthy blood donors to the French national blood collection center (Établissement Français du Sang). We have set up a 16S rDNA quantitative polymerase chain reaction assay as well as a 16S targeted metagenomics sequencing pipeline specifically designed to analyze the blood microbiome, which we have used on whole blood as well as on different blood fractions (buffy coat [BC], red blood cells [RBCs], and plasma). RESULTS: Most of the blood bacterial DNA is located in the BC (93.74%), and RBCs contain more bacterial DNA (6.23%) than the plasma (0.03%). The distribution of 16S DNA is different for each fraction and spreads over a relatively broad range among donors. At the phylum level, blood fractions contain bacterial DNA mostly from the Proteobacteria phylum (more than 80%) but also from Actinobacteria, Firmicutes, and Bacteroidetes. At deeper taxonomic levels, there are striking differences between the bacterial profiles of the different blood fractions. CONCLUSION: We demonstrate that a diversified microbiome exists in healthy blood. This microbiome has most likely an important physiologic role and could be implicated in certain transfusion-transmitted bacterial infections. In this regard, the amount of 16S bacterial DNA or the microbiome profile could be monitored to improve the safety of the blood supply.

Cross-alteration of murine skin and tick microbiome concomitant with pathogen transmission after Ixodes ricinus bite.

BACKGROUND: Ticks are major vectors of diseases affecting humans such as Lyme disease or domestic animals such as anaplasmosis. Cross-alteration of the vertebrate host skin microbiome and the tick microbiome may be essential during the process of tick feeding and for the mechanism of pathogen transmission. However, it has been poorly investigated. METHODS: We used mice bitten by field-collected ticks (nymphs and adult ticks) in different experimental conditions to investigate, by 16S rRNA gene metabarcoding, the impact of blood feeding on both the mouse skin microbiome and the tick microbiome. We also investigated by PCR and 16S rRNA gene metabarcoding, the diversity of microorganisms transmitted to the host during the process of tick bite at the skin interface and the dissemination of the pathogen in host tissues (blood, heart, and spleen). RESULTS: Most of the commensal bacteria present in the skin of control mice were replaced during the blood-feeding process by bacteria originating from the ticks. The microbiome of the ticks was also impacted by the blood feeding. Several pathogens including tick-borne pathogens (Borrelia/Borreliella, Anaplasma, Neoehrlichia, Rickettsia) and opportunistic bacteria (Williamsia) were transmitted to the skin microbiome and some of them disseminated to the blood or spleen of the mice. In the different experiments of this study, skin microbiome alteration and Borrelia/Borreliella transmission were different depending on the tick stages (nymphs or adult female ticks). CONCLUSIONS: Host skin microbiome at the bite site was deeply impacted by the tick bite, to an extent which suggests a role in the tick feeding, in the pathogen transmission, and a potentially important impact on the skin physiopathology. The diversified taxonomic profiles of the tick microbiome were also modified by the blood feeding. Video Abstract.

Blood, Gut, and Oral Microbiome in Kidney Transplant Recipients.

Background and Objective: Recent reports describe the existence of a blood microbiome profile not associated with an infection state. Given the high impact that the dysbiotic human microbiome appears to have in chronic kidney disease and, in particular, in the outcome of kidney transplant recipients (KTRs), we aimed to explore the variations and correlations of the gut, oral, and blood microbiome of recipients, 3 months after kidney transplantation. Materials and Methods: We conducted a cross-sectional study where the microbiome of stool, saliva, and blood collected from recipients 3 months after kidney transplantation (N = 6) was analyzed by polymerase chain reaction (PCR) amplification and sequencing of the V3-V4 hypervariable regions of the 16S rRNA gene using MiSeq Illumina® technology. Results: Blood of KTRs harbors a distinct low-abundance microbiome dominated by Proteobacteria and Firmicutes. Gut and oral microbiome of KTRs also present distinct profiles. The existence of a proportion of shared operational taxonomic units among the different body sites is reported, mainly classified as Proteobacteria and Firmicutes. Conclusions: This study provides evidence of existence a blood microbiome in KTRs, different from the gut and the oral microbiome profiles, with a small number of operational taxonomic units representing a shared microbiome. The clinical relevance of this observation should be further explored in these patients.

Diversity and phylogeny of the tick-borne bacterial genus Candidatus Allocryptoplasma (Anaplasmataceae).

The family Anaplasmataceae includes tick-borne bacteria of major public and veterinary health interest, as best illustrated by members of the genera Anaplasma and Ehrlichia. Recent epidemiological surveys have also reported on the presence of a novel putative genus in the Anaplasmataceae, Candidatus Allocryptoplasma, previously described as Candidatus Cryptoplasma in the western black-legged tick, Ixodes pacificus. However, the genetic diversity of Ca. Allocryptoplasma and its phylogenetic relationship with other Anaplasmataceae remain unclear. In this study, we developed a multi-locus sequence typing approach, examining the DNA sequence variation at five genes of Ca. Allocryptoplasma found in ticks. Combining this multi-locus sequence typing and genetic data available on public databases, we found that substantial genetic diversity of Ca. Allocryptoplasma is present in Ixodes, Amblyomma and Haemaphysalis spp. ticks on most continents. Further analyses confirmed that the Ca. Allocryptoplasma of ticks, the Ca. Allocryptoplasma of lizards and some Anaplasma-like bacteria of wild mice cluster into a monophyletic genus, divergent from all other genera of the family Anaplasmataceae. Candidatus Allocryptoplasma appears as a sister genus of Anaplasma and, with the genera Ehrlichia and Neoehrlichia, they form a monophyletic subgroup of Anaplasmataceae associated with tick-borne diseases. The detection of genetically distinct Ca. Allocryptoplasma in ticks of significant medical or veterinary interest supports the hypothesis that it is an emergent genus of tick-borne pathogens of general concern.

Title: Diversité et phylogénie du genre bactérien transmis par les tiques Candidatus Allocryptoplasma (Anaplasmataceae). Abstract: La famille des Anaplasmataceae comprend des bactéries transmises par les tiques qui présentent un intérêt majeur pour la santé publique et vétérinaire, comme les membres des genres Anaplasma et Ehrlichia. Des surveillances épidémiologiques récentes ont également signalé la présence d'un nouveau genre putatif dans les Anaplasmataceae, Candidatus Allocryptoplasma, initialement décrit comme Ca. Cryptoplasma chez une tique nord-américaine, Ixodes pacificus. Cependant, la diversité génétique des bactéries Ca. Allocryptoplasma et leurs relations phylogénétiques avec d'autres Anaplasmataceae restent méconnues. Dans cette étude, nous avons développé une approche de typage génétique multi-locus, en examinant la variation nucléotidique pour cinq gènes de bactéries Ca. Allocryptoplasma détectées chez les tiques. En combinant ce typage génétique multi-locus et les données génétiques disponibles dans les bases de données publiques, nous avons mis en évidence qu'une diversité génétique substantielle des bactéries Ca. Allocryptoplasma est présente chez les tiques des genres Ixodes, Amblyomma et Haemaphysalis sur la plupart des continents. Des analyses complémentaires confirment que les bactéries Ca. Allocryptoplasma des tiques, les bactéries Ca. Allocryptoplasma de lézards et des bactéries Anaplasma-like de souris sauvages se regroupent dans un genre monophylétique, divergent de tous les autres genres de la famille Anaplasmataceae. Candidatus Allocryptoplasma apparaît comme un genre frère d'Anaplasma et, avec les genres Ehrlichia et Neoehrlichia, ces trois genres forment un sous-groupe monophylétique d'Anaplasmataceae associé aux maladies transmises par les tiques. La détection de Ca. Allocryptoplasma dans des tiques d'intérêt médical et vétérinaire soutient l'hypothèse qu'il s'agit d'un genre émergent d'agents pathogènes majeurs.

Deficiency of histone variant macroH2A1.1 is associated with sexually dimorphic obesity in mice.

Obesity has a major socio-economic health impact. There are profound sex differences in adipose tissue deposition and obesity-related conditions. The underlying mechanisms driving sexual dimorphism in obesity and its associated metabolic disorders remain unclear. Histone variant macroH2A1.1 is a candidate epigenetic mechanism linking environmental and dietary factors to obesity. Here, we used a mouse model genetically depleted of macroH2A1.1 to investigate its potential epigenetic role in sex dimorphic obesity, metabolic disturbances and gut dysbiosis. Whole body macroH2A1 knockout (KO) mice, generated with the Cre/loxP technology, and their control littermates were fed a high fat diet containing 60% of energy derived from fat. The diet was administered for three months starting from 10 to 12 weeks of age. We evaluated the progression in body weight, the food intake, and the tolerance to glucose by means of a glucose tolerance test. Gut microbiota composition, visceral adipose and liver tissue morphology were assessed. In addition, adipogenic gene expression patterns were evaluated in the visceral adipose tissue. Female KO mice for macroH2A1.1 had a more pronounced weight gain induced by high fat diet compared to their littermates, while the increase in body weight in male mice was similar in the two genotypes. Food intake was generally increased upon KO and decreased by high fat diet in both sexes, with the exception of KO females fed a high fat diet that displayed the same food intake of their littermates. In glucose tolerance tests, glucose levels were significantly elevated upon high fat diet in female KO compared to a standard diet, while this effect was absent in male KO. There were no differences in hepatic histology. Upon a high fat diet, in female adipocyte cross-sectional area was larger in KO compared to littermates: activation of proadipogenic genes (ACACB, AGT, ANGPT2, FASN, RETN, SLC2A4) and downregulation of antiadipogenic genes (AXIN1, E2F1, EGR2, JUN, SIRT1, SIRT2, UCP1, CCND1, CDKN1A, CDKN1B, EGR2) was detected. Gut microbiota profiling showed increase in Firmicutes and a decrease in Bacteroidetes in females, but not males, macroH2A1.1 KO mice. MacroH2A1.1 KO mice display sexual dimorphism in high fat diet-induced obesity and in gut dysbiosis, and may represent a useful model to investigate epigenetic and metabolic differences associated to the development of obesity-associated pathological conditions in males and females.

Mucolipin-3 regulates luminal calcium, acidification, and membrane fusion in the endosomal pathway.

Mucolipin-3 (MCOLN3) is a pH-regulated Ca(2+) channel that localizes to the endosomal pathway. Gain-of-function mutation in MCOLN3 causes the varitint-waddler (Va) phenotype in mice, which is characterized by hearing loss, vestibular dysfunction, and coat color dilution. The Va phenotype results from a punctual mutation (A419P) in the pore region of MCOLN3 that locks the channel in an open conformation causing massive entry of Ca(2+) inside cells and inducing cell death by apoptosis. Overexpression of wild-type MCOLN3 produces severe alterations of the endosomal pathway, including enlargement and clustering of endosomes, delayed EGF receptor degradation, and impaired autophagosome maturation, thus suggesting that MCOLN3 plays an important role in the regulation of endosomal function. To understand better the physiological role of MCOLN3, we inhibited MCOLN3 function by expression of a channel-dead dominant negative mutant (458DD/KK) or by knockdown of endogenous MCOLN3. Remarkably, we found that impairment of MCOLN3 activity caused a significant accumulation of luminal Ca(2+) in endosomes. This accumulation led to severe defects in endosomal acidification as well as to increased endosomal fusion. Our findings reveal a prominent role for MCOLN3 in regulating Ca(2+) homeostasis at the endosomal pathway and confirm the importance of luminal Ca(2+) for proper acidification and membrane fusion.

Gut microbiota dysbiosis of type 2 diabetic mice impairs the intestinal daily rhythms of GLP-1 sensitivity.

The gut-brain-beta cell glucagon-like peptide-1 (GLP-1)-dependent axis and the clock genes both control insulin secretion. Evidence shows that a keystone of this molecular interaction could be the gut microbiota. We analyzed in mice the circadian profile of GLP-1 sensitivity on insulin secretion and the impact of the autonomic neuropathy, antibiotic treated in different diabetic mouse models and in germ-free colonized mice. We show that GLP-1sensitivity is maximal during the dark feeding period, i.e., the postprandial state. Coincidently, the ileum expression of GLP-1 receptor and peripherin is increased and tightly correlated with a subset of clock gene. Since both are markers of enteric neurons, it suggests a role in the gut-brain-beta cell GLP-1-dependent axis. We evaluated the importance of gut microbiota dysbiosis and found that the abundance of ileum bacteria, particularly Ruminococcaceae and Lachnospiraceae, oscillated diurnally, with a maximum during the dark period, along with expression patterns of a subset of clock genes. This diurnal pattern of circadian gene expression and Lachnospiraceae abundance was also observed in two separate mouse models of gut microbiota dysbiosis and of autonomic neuropathy with impaired GLP-1 sensitivity (1.high-fat diet-fed type 2 diabetic, 2.antibiotic-treated/germ-free mice). Our data show that GLP-1 sensitivity relies on specific pattern of intestinal clock gene expression and specific gut bacteria. This new statement opens opportunities to treat diabetic patient with GLP-1-based therapies by using on a possible pre/probiotic co-treatment to improve the time-dependent efficiency of these therapies.

Imbalanced gut microbiota fuels hepatocellular carcinoma development by shaping the hepatic inflammatory microenvironment.

Hepatocellular carcinoma (HCC) is a leading cause of cancer-related deaths worldwide, and therapeutic options for advanced HCC are limited. Here, we observe that intestinal dysbiosis affects antitumor immune surveillance and drives liver disease progression towards cancer. Dysbiotic microbiota, as seen in Nlrp6-/- mice, induces a Toll-like receptor 4 dependent expansion of hepatic monocytic myeloid-derived suppressor cells (mMDSC) and suppression of T-cell abundance. This phenotype is transmissible via fecal microbiota transfer and reversible upon antibiotic treatment, pointing to the high plasticity of the tumor microenvironment. While loss of Akkermansia muciniphila correlates with mMDSC abundance, its reintroduction restores intestinal barrier function and strongly reduces liver inflammation and fibrosis. Cirrhosis patients display increased bacterial abundance in hepatic tissue, which induces pronounced transcriptional changes, including activation of fibro-inflammatory pathways as well as circuits mediating cancer immunosuppression. This study demonstrates that gut microbiota closely shapes the hepatic inflammatory microenvironment opening approaches for cancer prevention and therapy.

The calcium channel mucolipin-3 is a novel regulator of trafficking along the endosomal pathway.

The varitint-waddler phenotype in mice is caused by gain-of-function mutations in mucolipin-3 (MCOLN3), a member of the mucolipin family of ion channels. These mice are characterized by defects in pigmentation, hearing loss and vestibular defects, suggesting that MCOLN3 might play a role in melanosome trafficking and hair cell maturation. Recent evidence has shown that MCOLN3 is a Ca(2+)-permeable channel and its activity is regulated by pH. Here we show that MCOLN3 primarily localizes to early and late endosomes in human epithelial cells. This distribution at the less acidic portions of the endocytic pathway is consistent with the reported inactivation of the channel by low pH. Furthermore, overexpression of MCOLN3 causes dramatic alterations in the endosomal pathway, including enlargement of Hrs-positive endosomes, delayed degradation of epidermal growth factor (EGF) and EGF receptor (EGFR) and defective autophagosome maturation, whereas depletion of endogenous MCOLN3 enhances EGFR degradation. Finally, we found that endosomal pH is higher in cells overexpressing MCOLN3 and propose a model in which Ca(2+) release from endosomes mediated by MCOLN3 might be important for efficient endosomal acidification. Therefore, MCOLN3 is a novel Ca(2+) channel that plays a crucial role in the regulation of cargo trafficking along the endosomal pathway.