Increasing evidence point to the relevance of intestinal disfunction and changes in the microbiome composition during chronic liver disease. More specifically, recent studies have highlighted that ...cholestatic diseases associate with a reduction in the microbiome diversity in patients. Still, the dynamics of the changes in the microbiome composition observed, as well as their implication in contributing to the pathogenesis of this disease remain largely undefined. Hence, experimental mouse models resembling the human pathogenesis are crucial to move forward our understanding on the mechanisms underpinning cholestatic disease and to enable the development of effective therapeutics. Our results show that the bile duct ligation (BDL) experimental model of cholestasis leads to rapid and significant changes in the microbiome diversity, with more than 100 OTUs being significantly different in faecal samples obtained from WT mice at 3 days and 7 days after BDL when compared to control animals. Changes in the microbial composition in mice after BDL included the enrichment of Akkermansia, Prevotella, Bacteroides and unclassified Ruminococcaceae in parallel with a drastic reduction of the presence of Faecalibacterium prausnitzii. In conclusion, our results support that bile duct ligation induces changes in the microbiome that partly resemble the gut microbial changes observed during human cholestatic disease.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Background and Aims
Mounting evidence supports an association between cholestatic liver disease and changes in the composition of the microbiome. Still, the role of the microbiome in the pathogenesis ...of this condition remains largely undefined.
Approach and Results
To address this, we have used two experimental models, administering alpha‐naphtylisocyanate or feeding a 0.1% 3,5‐diethoxycarbonyl‐1,4‐dihydrocollidine diet, to induce cholestatic liver disease in germ‐free mice and germ‐free mice conventionalized with the microbiome from wild‐type, specific pathogen‐free animals. Next, we have inhibited macrophage activation by depleting these cells using clodronate liposomes and inhibiting the inflammasome with a specific inhibitor of NOD‐, LRR‐, and pyrin domain‐containing protein 3. Our results demonstrate that cholestasis, the accumulation of bile acids in the liver, fails to promote liver injury in the absence of the microbiome in vivo. Additional in vitro studies supported that endotoxin sensitizes hepatocytes to bile‐acid–induced cell death. We also demonstrate that during cholestasis, macrophages contribute to promoting intestinal permeability and to altered microbiome composition through activation of the inflammasome, overall leading to increased endotoxin flux into the cholestatic liver.
Conclusions
We demonstrate that the intestinal microbiome contributes to cholestasis‐mediated cell death and inflammation through mechanisms involving activation of the inflammasome in macrophages.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Aerobic bacterial methane synthesis Wang, Qian; Alowaifeer, Abdullah; Kerner, Patricia ...
Proceedings of the National Academy of Sciences,
07/2021, Volume:
118, Issue:
27
Journal Article
Peer reviewed
Open access
Reports of biogenic methane (CH
) synthesis associated with a range of organisms have steadily accumulated in the literature. This has not happened without controversy and in most cases the process ...is poorly understood at the gene and enzyme levels. In marine and freshwater environments, CH
supersaturation of oxic surface waters has been termed the "methane paradox" because biological CH
synthesis is viewed to be a strictly anaerobic process carried out by O
-sensitive methanogens. Interest in this phenomenon has surged within the past decade because of the importance of understanding sources and sinks of this potent greenhouse gas. In our work on Yellowstone Lake in Yellowstone National Park, we demonstrate microbiological conversion of methylamine to CH
and isolate and characterize an
sp. capable of this activity. Furthermore, we identify and clone a gene critical to this process (encodes pyridoxylamine phosphate-dependent aspartate aminotransferase) and demonstrate that this property can be transferred to
with this gene and will occur as a purified enzyme. This previously unrecognized process sheds light on environmental cycling of CH
, suggesting that O
-insensitive, ecologically relevant aerobic CH
synthesis is likely of widespread distribution in the environment and should be considered in CH
modeling efforts.
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BFBNIB, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
is a flagellated gut anaerobic bacterium belonging to the
family within the Firmicutes phylum. A significant decrease of
colonization in the gut of ulcerative colitis patients has recently been ...demonstrated. In this work, we have investigated the mechanisms of
-host cross talk using both murine and
models.
The complete genome sequence of
A2-183 was determined. C3H/HeN germ-free mice were mono-colonized with
, and the host-microbe interaction was studied using histology, transcriptome analyses and FACS. Further investigations were performed
and using the TLR5KO and DSS-colitis murine models.
In the bacterium,
, host gut colonization upregulated genes involved in conjugation/mobilization, metabolism, motility, and chemotaxis. In the host cells, bacterial colonization upregulated genes related to antimicrobial peptides, gut barrier function, toll-like receptors (TLR) signaling, and T cell biology. CD4
CD25
FoxP3
T cell numbers increased in the
of both mono-associated and conventional mice treated with
. Treatment with the
bacterium provided protection against DSS-induced colitis. The role of flagellin in host-bacterium interaction was also investigated.
Mono-association of mice with
bacteria results in specific bidirectional gene expression patterns. A set of genes thought to be important for host colonization are induced in
, while the host cells respond by strengthening gut barrier function and enhancing Treg population expansion, possibly
TLR5-flagellin signaling. Our data reveal the immunomodulatory properties of
that could be useful for the control and treatment of gut inflammation.
Intestinal epithelial cells play a fundamental role in maintaining homeostasis. Shedding of intestinal cells in a controlled manner is critical to maintenance of barrier function. Barrier function is ...maintained during this shedding process by a redistribution of tight junctional proteins to facilitate closure of the gap left by the shedding cell. However, despite the obvious importance of epithelial cell shedding to gut health, a central question is how the extrusion of epithelial cells is achieved, enabling barrier integrity to be maintained in the healthy gut and restored during inflammation remains largely unanswered. Recent studies have provided evidence that excessive epithelial cell shedding and loss of epithelial barrier integrity is triggered by exposure to lipopolysaccharide or tumor necrosis factor alpha. Subsequent studies have provided evidence of the involvement of specific cellular components and signaling mechanisms as well as the functionality of microbiota that can be either detrimental or beneficial for intestinal barrier integrity. This review will focus on the evidence and decipher how the signaling systems through which the mucosal immune system and microbiota can regulate epithelial cell shedding and how these mechanisms interact to preserve the viability of the epithelium.
Hematopoietic stem cells (HSCs) undergo rapid expansion in response to stress stimuli. Here we investigate the bioenergetic processes which facilitate the HSC expansion in response to infection. We ...find that infection by Gram-negative bacteria drives an increase in mitochondrial mass in mammalian HSCs, which results in a metabolic transition from glycolysis toward oxidative phosphorylation. The initial increase in mitochondrial mass occurs as a result of mitochondrial transfer from the bone marrow stromal cells (BMSCs) to HSCs through a reactive oxygen species (ROS)-dependent mechanism. Mechanistically, ROS-induced oxidative stress regulates the opening of connexin channels in a system mediated by phosphoinositide 3-kinase (PI3K) activation, which allows the mitochondria to transfer from BMSCs into HSCs. Moreover, mitochondria transfer from BMSCs into HSCs, in the response to bacterial infection, occurs before the HSCs activate their own transcriptional program for mitochondrial biogenesis. Our discovery demonstrates that mitochondrial transfer from the bone marrow microenvironment to HSCs is an early physiologic event in the mammalian response to acute bacterial infection and results in bioenergetic changes which underpin emergency granulopoiesis.
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BFBNIB, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
Viral structural proteins can have multiple activities. Antivirals that target structural proteins have potential to exhibit multiple antiviral mechanisms. Hepatitis B virus (HBV) core protein (Cp) ...is involved in most stages of the viral life cycle; it assembles into capsids, packages viral RNA, is a metabolic compartment for reverse transcription, interacts with nuclear trafficking machinery, and disassembles to release the viral genome into the nucleus. During nuclear localization, HBV capsids bind to host importins (e.g., Impβ) via Cp's C-terminal domain (CTD); the CTD is localized to the interior of the capsid and is transiently exposed on the exterior. We used HAP12 as a representative Cp allosteric modulator (CpAM), a class of antivirals that inappropriately stimulates and misdirects HBV assembly and deforms capsids. CpAM impact on other aspects of the HBV life cycle is poorly understood. We investigate how HAP12 influences the interactions between empty or RNA-filled capsids with Impβ and trypsin
. We show that HAP12 can modulate CTD accessibility and capsid stability, depending on the saturation of HAP12-binding sites. We demonstrate that Impβ synergistically contributes to capsid disruption at high levels of HAP12 saturation, using electron microscopy to visualize the disruption and rearrangement of Cp dimers into aberrant complexes. However, RNA-filled capsids resist the destabilizing effects of HAP12 and Impβ. In summary, we show host protein-induced catalysis of capsid disruption, an unexpected additional mechanism of action for CpAMs. Potentially, untimely capsid disassembly can hamper the HBV life cycle and also cause the virus to become vulnerable to host innate immune responses.
The HBV core, an icosahedral complex of 120 copies of the homodimeric core (capsid) protein with or without packaged nucleic acid, is transported to the host nucleus by its interaction with host importin proteins. Importin-core interaction requires the core protein C-terminal domain, which is inside the capsid, to "flip" to the capsid exterior. Core protein-directed drugs that affect capsid assembly and stability have been developed recently. We show that these molecules can, synergistically with importins, disrupt capsids. This mechanism of action, synergism with host protein, has the potential to disrupt the virus life cycle and activate the innate immune system.
The intestinal epithelial monolayer, at the boundary between microbes and the host immune system, plays an important role in the development of inflammatory bowel disease (IBD), particularly as a ...target and producer of pro-inflammatory TNF. Chronic overexpression of TNF leads to IBD-like pathology over time, but the mechanisms driving early pathogenesis events are not clear. We studied the epithelial response to inflammation by combining mathematical models with in vivo experimental models resembling acute and chronic TNF-mediated injury. We found significant villus atrophy with increased epithelial cell death along the crypt-villus axis, most dramatically at the villus tips, in both acute and chronic inflammation. In the acute model, we observed overexpression of TNF receptor I in the villus tip rapidly after TNF injection and concurrent with elevated levels of intracellular TNF and rapid shedding at the tip. In the chronic model, sustained villus atrophy was accompanied by a reduction in absolute epithelial cell turnover. Mathematical modelling demonstrated that increased cell apoptosis on the villus body explains the reduction in epithelial cell turnover along the crypt-villus axis observed in chronic inflammation. Cell destruction in the villus was not accompanied by changes in proliferative cell number or division rate within the crypt. Epithelial morphology and immunological changes in the chronic setting suggest a repair response to cell damage although the villus length is not recovered. A better understanding of how this state is further destabilised and results in clinical pathology resembling IBD will help identify suitable pathways for therapeutic intervention.
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