Oral bacteria relative abundance in faeces increases due to gut microbiota depletion and is linked with patient outcomes.

The detection of oral bacteria in faecal samples has been associated with inflammation and intestinal diseases. The increased relative abundance of oral bacteria in faeces has two competing explanations: either oral bacteria invade the gut ecosystem and expand (the 'expansion' hypothesis), or oral bacteria transit through the gut and their relative increase marks the depletion of other gut bacteria (the 'marker' hypothesis). Here we collected oral and faecal samples from mouse models of gut dysbiosis (antibiotic treatment and DSS-induced colitis) and used 16S ribosomal RNA sequencing to determine the abundance dynamics of oral bacteria. We found that the relative, but not absolute, abundance of oral bacteria increases, reflecting the 'marker' hypothesis. Faecal microbiome datasets from diverse patient cohorts, including healthy individuals and patients with allogeneic haematopoietic cell transplantation or inflammatory bowel disease, consistently support the 'marker' hypothesis and explain associations between oral bacterial abundance and patient outcomes consistent with depleted gut microbiota. By distinguishing between the two hypotheses, our study guides the interpretation of microbiome compositional data and could potentially identify cases where therapies are needed to rebuild the resident microbiome rather than protect against invading oral bacteria.

IL-18-secreting multi-antigen targeting CAR T-cells eliminate antigen-low myeloma in an immunocompetent mouse model.

Multiple myeloma is a plasma cell malignancy that is currently incurable with conventional therapies. Following the success of CD19-targeted chimeric antigen receptor (CAR) T-cells in leukemia and lymphoma, CAR T-cells targeting B-cell maturation antigen (BCMA) more recently demonstrated impressive activity in relapsed and refractory myeloma patients. However, BCMA-directed therapy can fail due to low expression of BCMA on myeloma cells, suggesting that novel approaches to better address antigen-low disease may improve patient outcomes. We hypothesized that engineered secretion of the pro-inflammatory cytokine interleukin-18 (IL-18) and multi-antigen targeting could improve CAR T-cell activity against BCMA-low myeloma. In a syngeneic murine model of myeloma, CAR T-cells targeting the myeloma-associated antigens BCMA and B-cell activating factor (BAFF-R) failed to eliminate myeloma when these antigens were weakly expressed, whereas IL-18-secreting CAR T-cells targeting these antigens promoted myeloma clearance. IL-18-secreting CAR T-cells developed an effector-like T-cell phenotype, promoted interferon-gamma production, reprogrammed the myeloma bone marrow microenvironment through type I/II interferon signaling, and activated macrophages to mediate anti-myeloma activity. Simultaneous targeting of weakly expressed BCMA and BAFF-R with dual-CAR T-cells enhanced T-cell:target cell avidity, increased overall CAR signal strength, and stimulated anti-myeloma activity. Dual-antigen targeting augmented CAR T-cell secretion of engineered IL-18 and facilitated elimination of larger myeloma burdens in vivo. Our results demonstrate that combination of engineered IL-18 secretion and multi-antigen targeting can eliminate myeloma with weak antigen expression through distinct mechanisms.

Altered microbial bile acid metabolism exacerbates T cell-driven inflammation during graft-versus-host disease.

Microbial transformation of bile acids affects intestinal immune homoeostasis but its impact on inflammatory pathologies remains largely unknown. Using a mouse model of graft-versus-host disease (GVHD), we found that T cell-driven inflammation decreased the abundance of microbiome-encoded bile salt hydrolase (BSH) genes and reduced the levels of unconjugated and microbe-derived bile acids. Several microbe-derived bile acids attenuated farnesoid X receptor (FXR) activation, suggesting that loss of these metabolites during inflammation may increase FXR activity and exacerbate the course of disease. Indeed, mortality increased with pharmacological activation of FXR and decreased with its genetic ablation in donor T cells during mouse GVHD. Furthermore, patients with GVHD after allogeneic hematopoietic cell transplantation showed similar loss of BSH and the associated reduction in unconjugated and microbe-derived bile acids. In addition, the FXR antagonist ursodeoxycholic acid reduced the proliferation of human T cells and was associated with a lower risk of GVHD-related mortality in patients. We propose that dysbiosis and loss of microbe-derived bile acids during inflammation may be an important mechanism to amplify T cell-mediated diseases.

Living Donor Whole and Partial Liver Grafts, Deceased Donor Whole Liver and SPLIT: Outcome Comparison.

BACKGROUND: Currently, graft options for pediatric liver transplantation (PLT) include whole (WL) and partial (P) grafts, in the form of either deceased donor transplantation (DD) or living donor liver transplantation (LD). WL transplants from LD are commonly referred to as domino LT. The objective of this manuscript is to compare the outcomes of PLT performed with each of the available graft options. METHODS: Retrospective cohort study from Jan. 2010 to Dec. 2022. The variables included data on the recipients' preoperative clinical status, intraoperative technical aspects, post-operative complications, and survival studies. There were 4 groups: SPLIT (17), DD-WL (55), LD-WL (824), and LD-P (22). RESULTS: The median age and BW of the recipients was smaller in SPLIT, LD-P, and LD-WL compared to DDT-WL groups. HVOO (HR 15.87, 95% CI 1.89-133.06, P = 0.01), retransplantation (HR 7.94, 95% CI 2.63-24.02, P < 0.01), and malignancies (HR 3.08, 95% CI 1.29-7.37, P = 0.01) were independently associated with decreased patient survival. HAT (HR 27.54, 95% CI 10.44-72.68, P < 0.01) and malignancies (HR 2.42, 95% CI 1.10-5.34, P = 0.03) increased the risk of graft loss. The overall survival in this series was 91.4% (mean follow-up of 74.3 months). Patient and graft survival were not different among groups. CONCLUSION: HAT and malignancies were associated with reduced graft survival. Whole liver from living donors with MSUD presented 100% patient survival at 120 months. Even without statistical differences in survival among the studied groups, LD-P and LD-WL recipients presented a trend towards better outcomes. LEVEL OF EVIDENCE: LEVEL III.

Combination of in silico and cell culture strategies to predict biomaterial performance: Effects of sintering temperature on the biological properties of hydroxyapatite.

Advances in methodologies to evaluate biomaterials brought an explosive growth of data, ensuing computational challenges to better analyzing them and allowing for high-throughput profiling of biological systems cost-efficiently. In this sense, we have applied bioinformatics tools to better understand the biological effect of different sintering temperatures of hydroxyapatite (abbreviated HA; at 1100, 1150, and 1250°C) on osteoblast performance. To do, we have better analyzed an earlier deposited study, in which the access code is E-MTAB-7219, which the authors have explored different in silico tools on this purpose. In this study, differential gene expression analyses were performed using the gene set variation analysis (GSVA) algorithm from the transcriptomes respecting the thermal changes of HA, which were validated using exclusively in vitro strategies. Furthermore, in silico approaches elected biomarkers during cell behavior in response to different sintering temperatures of HA, and it was further validated using cell culture and qPCR technologies. Altogether, the combination of those strategies shows the capacity of sintered HA at 1250°C to present a better performance in organizing an adequate microenvironment favoring bone regeneration, angiogenesis and material resorption stimulus once it has promoted higher involvement of genes such as CDK2, CDK4 (biomarkers of cell proliferation), p15, Osterix gene (related with osteogenic differentiation), RANKL (related with osteoclastogenesis), VEGF gene (related with angiogenesis), and HIF1α (related with hypoxia microenvironment). Altogether, the combination of in silico and cell culture strategies shows the capacity of sintered HA at 1250°C in guaranteeing osteoblast differentiation and it can be related in organizing an adequate microenvironment favoring bone regeneration, angiogenesis, and material resorption stimulus.

Bacteria and bacteriophage consortia are associated with protective intestinal metabolites in patients receiving stem cell transplantation.

The microbiome is a predictor of clinical outcome in patients receiving allogeneic hematopoietic stem cell transplantation (allo-SCT). Microbiota-derived metabolites can modulate these outcomes. How bacteria, fungi and viruses contribute to the production of intestinal metabolites is still unclear. We combined amplicon sequencing, viral metagenomics and targeted metabolomics from stool samples of patients receiving allo-SCT (n = 78) and uncovered a microbiome signature of Lachnospiraceae and Oscillospiraceae and their associated bacteriophages, correlating with the production of immunomodulatory metabolites (IMMs). Moreover, we established the IMM risk index (IMM-RI), which was associated with improved survival and reduced relapse. A high abundance of short-chain fatty acid-biosynthesis pathways, specifically butyric acid via butyryl-coenzyme A (CoA):acetate CoA-transferase (BCoAT, which catalyzes EC 2.8.3.8) was detected in IMM-RI low-risk patients, and virome genome assembly identified two bacteriophages encoding BCoAT as an auxiliary metabolic gene. In conclusion, our study identifies a microbiome signature associated with protective IMMs and provides a rationale for considering metabolite-producing consortia and metabolite formulations as microbiome-based therapies.

Dietary cystine restriction increases the proliferative capacity of the small intestine of mice.

Currently, over 88 million people are estimated to have adopted a vegan or vegetarian diet. Cysteine is a semi-essential amino acid, which availability is largely dependent on dietary intake of meat, eggs and whole grains. Vegan/vegetarian diets are therefore inherently low in cysteine. Sufficient uptake of cysteine is crucial, as it serves as substrate for protein synthesis and can be converted to taurine and glutathione. We found earlier that intermolecular cystine bridges are essential for the barrier function of the intestinal mucus layer. Therefore, we now investigate the effect of low dietary cystine on the intestine. Mice (8/group) received a high fat diet with a normal or low cystine concentration for 2 weeks. We observed no changes in plasma methionine, cysteine, taurine or glutathione levels or bile acid conjugation after 2 weeks of low cystine feeding. In the colon, dietary cystine restriction results in an increase in goblet cell numbers, and a borderline significant increase mucus layer thickness. Gut microbiome composition and expression of stem cell markers did not change on the low cystine diet. Remarkably, stem cell markers, as well as the proliferation marker Ki67, were increased upon cystine restriction in the small intestine. In line with this, gene set enrichment analysis indicated enrichment of Wnt signaling in the small intestine of mice on the low cystine diet, indicative of increased epithelial proliferation. In conclusion, 2 weeks of cystine restriction did not result in apparent systemic effects, but the low cystine diet increased the proliferative capacity specifically of the small intestine and induced the number of goblet cells in the colon.

Immune-Related Colitis Is Associated with Fecal Microbial Dysbiosis and Can Be Mitigated by Fecal Microbiota Transplantation.

Colitis induced by treatment with immune-checkpoint inhibitors (ICI), termed irColitis, is a substantial cause of morbidity complicating cancer treatment. We hypothesized that abnormal fecal microbiome features would be present at the time of irColitis onset and that restoring the microbiome with fecal transplant from a healthy donor would mitigate disease severity. Herein, we present fecal microbiota profiles from 18 patients with irColitis from a single center, 5 of whom were treated with healthy-donor fecal microbial transplantation (FMT). Although fecal samples collected at onset of irColitis had comparable α-diversity to that of comparator groups with gastrointestinal symptoms, irColitis was characterized by fecal microbial dysbiosis. Abundances of Proteobacteria were associated with irColitis in multivariable analyses. Five patients with irColitis refractory to steroids and biologic anti-inflammatory agents received healthy-donor FMT, with initial clinical improvement in irColitis symptoms observed in four of five patients. Two subsequently exhibited recurrence of irColitis symptoms following courses of antibiotics. Both received a second "salvage" FMT that was, again, followed by clinical improvement of irColitis. In summary, we observed distinct microbial community changes that were present at the time of irColitis onset. FMT was followed by clinical improvements in several cases of steroid- and biologic-agent-refractory irColitis. Strategies to restore or prevent microbiome dysbiosis in the context of immunotherapy toxicities should be further explored in prospective clinical trials.

Enabling the Methanol Economy: Opportunities and Challenges for Heterogeneous Catalysis in the Production of Liquid Fuels via Methanol.

ConspectusThirty years ago, George A. Olah proposed the concept of the methanol economy, where methanol replaces fossil fuels as a means of energy storage, ground transportation fuel, and raw material for the manufacture of other carbon-based products. Over the years, with rising global warming concerns, the concept has evolved. A special interest is devoted to the development of catalytic processes that allow the transformation of carbon dioxide, via methanol, into CO2 neutral liquid hydrocarbons. These products could replace the oil-based fuels currently used by combustion engines. The rapid depletion of such fuels would avoid a considerable amount of CO2 emissions during the current energy transition.Over the past decade, we have focused on different key processes that should allow for maximal atom efficiency and, therefore, minimal energy consumption in a field, CO2 valorization, that can easily become a zero-sum game. In this Account, we highlight the importance of catalyst design to overcome the process challenges in the production of liquid fuels from methanol. Additionally, progress in multifunctional catalysts able to directly convert, in one single reactor, CO2 to liquid fuels is also discussed in detail. This integrated option is of particular interest since it allows an important decrease in operational units while increasing throughput by converting, in situ, a thermodynamically limited intermediate.

Enhanced clinical assessment of hematologic malignancies through routine paired tumor and normal sequencing.

Genomic profiling of hematologic malignancies has augmented our understanding of variants that contribute to disease pathogenesis and supported development of prognostic models that inform disease management in the clinic. Tumor only sequencing assays are limited in their ability to identify definitive somatic variants, which can lead to ambiguity in clinical reporting and patient management. Here, we describe the MSK-IMPACT Heme cohort, a comprehensive data set of somatic alterations from paired tumor and normal DNA using a hybridization capture-based next generation sequencing platform. We highlight patterns of mutations, copy number alterations, and mutation signatures in a broad set of myeloid and lymphoid neoplasms. We also demonstrate the power of appropriate matching to make definitive somatic calls, including in patients who have undergone allogeneic stem cell transplant. We expect that this resource will further spur research into the pathobiology and clinical utility of clinical sequencing for patients with hematologic neoplasms.

Agency effects on the binding of event elements in episodic memory.

Representing events in episodic memory in a coherent manner requires that their constituent elements are bound together. So far, only few moderators of these binding processes have been identified. Here we investigate whether the presence of an agentic element in an event facilitates binding. The results from six experiments provided no evidence for a facilitating effect of agency on the binding of event elements. In addition, binding effects were only found when event elements were presented simultaneously, but not when they were presented sequentially pairwise, contrary to previous findings. The results suggest that the presence of an agentic element in an event does not, or only to a very limited extent, contribute to the formation of coherent memory representations and that additional processes may be required when binding event elements across temporarily divided encoding episodes. These findings add to a growing body of research regarding moderators and processes relevant for the binding of event elements in episodic memory. Explanations of these findings and directions for future research are discussed.

Circulating microbial cell-free DNA is increased during neutropenia after hematopoietic stem cell transplantation.

We used a next-generation sequencing platform to characterize microbial cell-free DNA (mcfDNA) in plasma samples from patients undergoing allogeneic hematopoietic stem cell transplantation (allo-HCT). In this observational study, we sought to characterize plasma mcfDNA in order to explore its potential association with the immunologic complications of transplantation. We compared serially collected patient samples with plasma collected from healthy control subjects. We observed changes in total mcfDNA burden in the plasma after transplantation, which was most striking during the early posttransplant neutropenic phase. This elevation could be attributed to a number of specific bacterial taxa, including Veillonella, Bacteroides, and Prevotella (genus level). For an additional cohort of patients, we compared the data of mcfDNA from plasma with 16s-ribosomal RNA sequencing data from stool samples collected at matched time points. In a number of patients, we confirmed that mcfDNA derived from specific microbial taxa (eg, Enterococcus) could also be observed in the matched stool sample. Quantification of mcfDNA may generate novel insights into mechanisms by which the intestinal microbiome influences systemic cell populations and, thus, has been associated with outcomes for patients with cancer.

Intestinal microbiota controls graft-versus-host disease independent of donor-host genetic disparity.

Acute graft-versus-host disease (aGVHD) remains a major limitation of allogeneic stem cell transplantation (SCT), and severe intestinal manifestation is the major cause of early mortality. Intestinal microbiota control MHC class II (MHC-II) expression by ileal intestinal epithelial cells (IECs) that promote GVHD. Here, we demonstrated that genetically identical mice of differing vendor origins had markedly different intestinal microbiota and ileal MHC-II expression, resulting in discordant GVHD severity. We utilized cohousing and antibiotic treatment to characterize the bacterial taxa positively and negatively associated with MHC-II expression. A large proportion of bacterial MHC-II inducers were vancomycin sensitive, and peri-transplant oral vancomycin administration attenuated CD4+ T cell-mediated GVHD. We identified a similar relationship between pre-transplant microbes, HLA class II expression, and both GVHD and mortality in a large clinical SCT cohort. These data highlight therapeutically tractable mechanisms by which pre-transplant microbial taxa contribute to GVHD independently of genetic disparity.

Tissue-specific features of the T cell repertoire after allogeneic hematopoietic cell transplantation in human and mouse.

T cells are the central drivers of many inflammatory diseases, but the repertoire of tissue-resident T cells at sites of pathology in human organs remains poorly understood. We examined the site-specificity of T cell receptor (TCR) repertoires across tissues (5 to 18 tissues per patient) in prospectively collected autopsies of patients with and without graft-versus-host disease (GVHD), a potentially lethal tissue-targeting complication of allogeneic hematopoietic cell transplantation, and in mouse models of GVHD. Anatomic similarity between tissues was a key determinant of TCR repertoire composition within patients, independent of disease or transplant status. The T cells recovered from peripheral blood and spleens in patients and mice captured a limited portion of the TCR repertoire detected in tissues. Whereas few T cell clones were shared across patients, motif-based clustering revealed shared repertoire signatures across patients in a tissue-specific fashion. T cells at disease sites had a tissue-resident phenotype and were of donor origin based on single-cell chimerism analysis. These data demonstrate the complex composition of T cell populations that persist in human tissues at the end stage of an inflammatory disorder after lymphocyte-directed therapy. These findings also underscore the importance of studying T cell in tissues rather than blood for tissue-based pathologies and suggest the tissue-specific nature of both the endogenous and posttransplant T cell landscape.

High-resolution analyses of associations between medications, microbiome, and mortality in cancer patients.

Discerning the effect of pharmacological exposures on intestinal bacterial communities in cancer patients is challenging. Here, we deconvoluted the relationship between drug exposures and changes in microbial composition by developing and applying a new computational method, PARADIGM (parameters associated with dynamics of gut microbiota), to a large set of longitudinal fecal microbiome profiles with detailed medication-administration records from patients undergoing allogeneic hematopoietic cell transplantation. We observed that several non-antibiotic drugs, including laxatives, antiemetics, and opioids, are associated with increased Enterococcus relative abundance and decreased alpha diversity. Shotgun metagenomic sequencing further demonstrated subspecies competition, leading to increased dominant-strain genetic convergence during allo-HCT that is significantly associated with antibiotic exposures. We integrated drug-microbiome associations to predict clinical outcomes in two validation cohorts on the basis of drug exposures alone, suggesting that this approach can generate biologically and clinically relevant insights into how pharmacological exposures can perturb or preserve microbiota composition. The application of a computational method called PARADIGM to a large dataset of cancer patients' longitudinal fecal specimens and detailed daily medication records reveals associations between drug exposures and the intestinal microbiota that recapitulate in vitro findings and are also predictive of clinical outcomes.

The Multifarious Functions of Leukotrienes in Bone Metabolism.

Leukotrienes (LTs) are derived from arachidonic acid metabolism by the 5-lipoxygenase (5-LO) enzyme. The production of LTs is stimulated in the pathogenesis of rheumatoid arthritis (RA), osteoarthritis, and periodontitis, with a relevant contribution to bone resorption. However, its role in bone turnover, particularly the suppression of bone formation by modulating the function of osteoclasts and osteoblasts, remains unclear. We investigated the effects of LTs on bone metabolism and their impact on osteogenic differentiation and osteoclastogenesis using a 5-LO knockout (KO) mouse model. Results from micro-computed tomography (µCT) analysis of femur from 8-week-old 5-LO-deficient mice showed increased cortical bone and medullary region in females and males and decreased trabecular bone in females. In the vertebra, we observed increased marrow area in both females and males 5-LO KO and decreased trabecular bone only in females 5-LO KO. Immunohistochemistry (IHC) analysis showed higher levels of osteogenic markers tissue-nonspecific alkaline phosphatase (TNAP) and osteopontin (OPN) and lower expression of osteoclastogenic marker tartrate-resistant acid phosphatase (TRAP) in the femurs of 5-LO KO mice versus wild-type (WT). Alkaline phosphatase activity and mineralization assay results showed that the 5-LO absence enhances osteoblasts differentiation and mineralization but decreases the proliferation. Alkaline phosphatase (ALP), Bglap, and Sp7 gene expression were higher in 5-LO KO osteoblasts compared to WT cells. Eicosanoids production was higher in 5-LO KO osteoblasts except for thromboxane 2, which was lower in 5-LO-deficient mice. Proteomic analysis identified the downregulation of proteins related to adenosine triphosphate (ATP) metabolism in 5-LO KO osteoblasts, and the upregulation of transcription factors such as the adaptor-related protein complex 1 (AP-1 complex) in long bones from 5-LO KO mice leading to an increased bone formation pattern in 5-LO-deficient mice. We observed enormous differences in the morphology and function of osteoclasts with reduced bone resorption markers and impaired osteoclasts in 5-LO KO compared to WT osteoclasts. Altogether, these results demonstrate that the absence of 5-LO is related to the greater osteogenic profile. © 2023 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

The TaxUMAP atlas: Efficient display of large clinical microbiome data reveals ecological competition in protection against bacteremia.

Longitudinal microbiome data provide valuable insight into disease states and clinical responses, but they are challenging to mine and view collectively. To address these limitations, we present TaxUMAP, a taxonomically informed visualization for displaying microbiome states in large clinical microbiome datasets. We used TaxUMAP to chart a microbiome atlas of 1,870 patients with cancer during therapy-induced perturbations. Bacterial density and diversity were positively associated, but the trend was reversed in liquid stool. Low-diversity states (dominations) remained stable after antibiotic treatment, and diverse communities had a broader range of antimicrobial resistance genes than dominations. When examining microbiome states associated with risk for bacteremia, TaxUMAP revealed that certain Klebsiella species were associated with lower risk for bacteremia localize in a region of the atlas that is depleted in high-risk enterobacteria. This indicated a competitive interaction that was validated experimentally. Thus, TaxUMAP can chart comprehensive longitudinal microbiome datasets, enabling insights into microbiome effects on human health.