Previous studies have demonstrated the presence of microbial DNA in the fetal environment. However, it remains unclear whether this DNA represents viable bacteria and how it relates to the maternal microbiota across different body sites. We studied the microbiota of human and mouse dyads to understand these relationships, localize bacteria in the fetus, and demonstrate bacterial viability. In human preterm and full-term mother-infant dyads at the time of Cesarean delivery, the oral cavity and meconium of newborn infants born as early as 24 weeks of gestation contained a microbiota that was predicted to originate from in utero sources including the placenta. Using operative deliveries of pregnant mice under highly controlled, sterile conditions in the laboratory, composition, visualization, and viability of bacteria in the in utero compartment and fetal intestine were demonstrated by 16S rRNA gene sequencing, fluorescence in situ hybridization, and bacterial culture. The composition and predicted source of the fetal gut microbiota shifted between mid- and late gestation. Cultivatable bacteria in the fetal intestine were found during mid-gestation but not late gestation. Our results demonstrate a dynamic, viable mammalian fetal microbiota during in utero development.
Noelle Younge, Jessica R. McCann, Julie Ballard, Catherine Plunkett, Suhail Akhtar, Félix Araújo-Pérez, Amy Murtha, Debra Brandon, Patrick C. Seed
Chagas disease is a life-long pathology resulting from Trypanosoma cruzi infection. It represents one of the most frequent causes of heart failure and sudden death in Latin America. Herein we provide evidence that aerobic glycolytic pathway activation in monocytes drives nitric oxide (NO) production, triggering tyrosine nitration (TN) on CD8 T cells and dysfunction in patients with chronic Chagas disease. Monocytes from patients exhibited higher frequency of hypoxia inducible factor (HIF)-1α and increased expression of its target genes/proteins. Non-classical monocytes are expanded in patients´ peripheral blood and represent an important source of NO. Monocytes entail CD8 T cell surface nitration since both the frequency of non-classical monocytes and that of NO-producing monocytes, positively correlated with the percentage of TN+ lymphocytes. Inhibition of glycolysis in (in vitro) infected peripheral blood mononuclear cells decreased the inflammatory properties of monocytes/macrophages diminishing the frequency of IL-1β- and NO-producing cells. In agreement, glycolysis inhibition reduced the percentage of TN+CD8 T cells improving their functionality. Altogether, these results clearly evidence that glycolysis governs oxidative stress on monocytes and modulates monocyte-T cell interplay in human chronic Chagas disease. Understanding the pathological immune mechanisms that sustains inflammatory environment in human pathology is key to design improved therapies.
Liliana María Sanmarco, Natalia Eberhardt, Gastón Bergero, Luz Piedad Quebrada Palacio, Pamela Martino Adami, Laura Marina Visconti, Ángel Ramón Minguez, Yolanda Hernández-Vasquez, Eugenio Antonio Carrera Silva, Laura Morelli, Miriam Postan, Maria Pilar Aoki
Long-term survivors post hematopoietic stem cell transplantation are at high risk of infection which accounts for one-third of all deaths. Little is known about the cause of inferior host defense after immune cell reconstitution. Here, we exploited a murine syngeneic bone marrow transplantation (BMT) model of late infection with gammaherpesvirus 68 (MHV-68) to determine the role of conventional dendritic cell (cDC) trafficking in adaptive immunity in BMT mice. Post infection, the expression of chemokine Ccl21 in the lung is reduced and the migration of cDCs into lung draining lymph nodes (dLNs) is impaired in BMT mice, limiting the opportunity for cDCs to prime Th cells in the dLNs. While cDC subsets are redundant in priming Th1 cells, Notch2 functions in cDC2s are required for priming increased Th17 responses in BMT mice and cDC1s can lessen this activity. Importantly, Th17 cells can be primed both in the lungs and dLNs, allowing for increased Th17 responses without optimum cDC trafficking in BMT mice. Taken together, impaired cDC trafficking in BMT mice reduces protective Th1 responses and allows increased pathogenic Th17 responses. Thus, we have revealed a previously unknown mechanism for BMT procedures to cause long-term inferior immune responses to herpes viral infection.
Carol A. Wilke, Matthew M. Chadwick, Paul R. Chan, Bethany B. Moore, Xiaofeng Zhou
Chromatin modifiers act to coordinate gene expression changes critical to neuronal differentiation from neural stem/progenitor cells (NSPCs). Lysine-specific methyltransferase 2D (KMT2D) encodes a histone methyltransferase that promotes transcriptional activation, and is frequently mutated in cancers and in the majority (>70%) of patients diagnosed with the congenital, multisystem intellectual disability (ID) disorder Kabuki syndrome 1 (KS1). Critical roles for KMT2D are established in various non-neural tissues, but the effects of KMT2D loss in brain cell development have not been described. We conducted parallel studies of proliferation, differentiation, transcription, and chromatin profiling in KMT2D-deficient human and mouse models to define KMT2D-regulated functions in neurodevelopmental contexts, including adult-born hippocampal NSPCs in vivo and in vitro. We report cell-autonomous defects in proliferation, cell cycle, and survival, accompanied by early NSPC maturation in several KMT2D-deficient model systems. Transcriptional suppression in KMT2D-deficient cells indicated strong perturbation of hypoxia-responsive metabolism pathways. Functional experiments confirmed abnormalities of cellular hypoxia responses in KMT2D-deficient neural cells, and accelerated NSPC maturation in vivo. Together, our findings support a model in which loss of KMT2D function suppresses expression of oxygen-responsive gene programs important to neural progenitor maintenance, resulting in precocious neuronal differentiation in a mouse model of KS1.
Giovanni A. Carosso, Leandros Boukas, Jonathan J. Augustin, Ha Nam Nguyen, Briana L. Winer, Gabrielle H. Cannon, Johanna D. Robertson, Li Zhang, Kasper D. Hansen, Loyal A. Goff, Hans T. Bjornsson
Aberrant accumulation and activation of eosinophils and potentially mast cells (MCs) contribute to the pathogenesis of eosinophilic gastrointestinal diseases (EGIDs), including eosinophilic esophagitis (EoE), gastritis (EG), and gastroenteritis (EGE). Current treatment options such as diet restriction and corticosteroids have limited efficacy and are often inappropriate for chronic use. One promising new approach is to deplete eosinophils and inhibit MCs with a monoclonal antibody (mAb) against Siglec-8, an inhibitory receptor selectively expressed on MCs and eosinophils. Here, we characterize MCs and eosinophils from human EG and EoE biopsies using flow cytometry and evaluate the effects of an anti-Siglec-8 mAb using a novel Siglec-8 transgenic mouse model in which EG/EGE was induced by ovalbumin sensitization and intragastric challenge. Mast cells and eosinophils were significantly increased and activated in human EG and EoE biopsies compared to healthy controls. Similar observations were made in EG/EGE mice. In Siglec-8 transgenic mice, anti-Siglec-8 mAb administration significantly reduced eosinophils and MCs in the stomach, small intestine, and mesenteric lymph nodes, and decreased levels of inflammatory mediators. In summary, these findings suggest a role for both MCs and eosinophils in EGID pathogenesis and support the evaluation of anti-Siglec-8 as a therapeutic approach that targets both eosinophils and MCs.
Bradford A. Youngblood, Emily C. Brock, John Leung, Rustom Falahati, Bruce S. Bochner, Henrik S. Rasmussen, Kathryn Peterson, Christopher Bebbington, Nenad Tomasevic
Excessive vascular remodeling is characteristic of hemophilic arthropathy (HA) and may contribute to joint bleeding and the progression of HA. Mechanisms for pathological vascular remodeling after hemophilic joint bleeding are unknown. In hemophilia, activation of thrombin-activatable fibrinolysis inhibitor (TAFI) is impaired, which contributes to joint bleeding and may also underlie the aberrant vascular remodeling. Here, hemophilia A (FVIII-deficient) mice or TAFI-deficient mice with transient (antibody-induced) hemophilia A were used to determine the role of FVIII and TAFI in vascular remodeling after joint bleeding. Excessive vascular remodeling and vessel enlargement persisted in FVIII-deficient and TAFI-deficient but not in transient hemophilia WT mice after similar joint bleeding. TAFI-overexpression in FVIII-deficient mice prevented abnormal vessel enlargement and vascular leakage. Age-related vascular changes were observed with FVIII or TAFI deficiency, and correlated positively with bleeding severity after injury, supporting increased vascularity as a major contributor to joint bleeding. Antibody-mediated inhibition of uPA also prevented abnormal vascular remodeling, suggesting that TAFI’s protective effects include inhibition of uPA-mediated plasminogen activation. In conclusion, the functional TAFI deficiency in hemophilia drives maladaptive vascular remodeling in the joints after bleeding. These new mechanistic insights allow targeted development of new strategies to normalize vascularity and control re-bleeding in HA.
Tine Wyseure, Tingyi Yang, Jenny Y. Zhou, Esther J. Cooke, Bettina Wanko, Merissa Olmer, Ruchi Agashe, Yosuke Morodomi, Niels Behrendt, Martin Lotz, John Morser, Annette von Drygalski, Laurent O. Mosnier
CD8+ tumor-infiltrating lymphocytes (TILs) correlate with relapse-free survival (RFS) in most cancer types, including breast cancer. However, subset composition, functional status, and spatial location of CD8+ TILs in relation to RFS in human breast tumors remain unclear. Spatial tissue analysis via quantitative immunofluorescence showed that infiltration of CD8+ T cells into cancer islands is more significantly associated with RFS than CD8+ T cell infiltration into either tumor stroma or total tumor. Localization into cancer islands within tumors is mediated by expression of the integrin CD103, which is a marker for tissue resident memory T cells (TRMs). Analysis of fresh tumor samples revealed that CD8+ TRMs are functionally similar to other CD8+ TILs, suggesting that the basis of their protective effect is their spatial distribution rather than functional differences. Indeed, CD103+ TRMs, as compared to CD103- CD8+ TILs, are enriched within cancer islands and CD8+ TRM proximity to cancer cells drives the association of CD8+ TIL densities with RFS. Together, these findings reveal the importance of cancer island localized CD8+ TRMs in surveillance of the breast tumor microenvironment and as a critical determinant of RFS in breast cancer patients.
Colt A. Egelston, Christian Avalos, Travis Y. Tu, Anthony Rosario, Roger Wang, Shawn Solomon, Gayathri Srinivasan, Michael S. Nelson, Yinghui Huang, Min Hui Lim, Diana L. Simons, Ting-Fang He, John H. Yim, Laura Kruper, Joanne Mortimer, Susan Yost, Weihua Guo, Christopher Ruel, Paul H. Frankel, Yuan Yuan, Peter P. Lee
There is increased interest in whether bariatric surgeries such as Roux-en-Y gastric bypass (RYGB) achieve their profound weight-lowering effects in morbidly obese individuals through the brain. Hypothalamic inflammation is a well-recognized etiologic factor in obesity pathogenesis and so represents a potential target of RYGB, but clinical evidence in support of this is limited. We therefore assessed hypothalamic T2-weighted signal intensities (T2W SI) and fractional anisotropy (FA) values, two validated radiologic measures of brain inflammation, in relation to BMI and fat mass as well as circulating inflammatory (C-reactive peptide - CrP) and metabolic markers in a cohort of 27 RYGB patients at baseline, 6 months and 12 months after surgery. We found that RYGB progressively increased hypothalamic T2W SI values while it progressively decreased hypothalamic FA values. Regression analyses further revealed that this could be most strongly linked to plasma CrP levels which independently predicted hypothalamic FA values when adjusting for age, sex, fat mass and diabetes diagnosis. These findings suggest that RYGB has a major time-dependent impact on hypothalamic inflammation status possibly by attenuating peripheral inflammation. They also suggest that hypothalamic FA values may provide a more specific radiologic measure of hypothalamic inflammation than more commonly used T2W SI values.
Mohammed K. Hankir, Michael Rullmann, Florian Seyfried, Sven Preusser, Sindy Poppitz, Stefanie Heba, Kostantinos Gousias, Jana Hoyer, Tatjana Schütz, Arne Dietrich, Karsten Müller, Burkhard Pleger
Calorie restriction (CR) improved healthspan in two longitudinal studies in nonhuman primates (NHPs), yet only the University of Wisconsin (UW) study demonstrated an increase in survival in CR monkeys relative to controls; the National Institute on Aging (NIA) study did not. Here, analysis of left ventricle samples showed that CR did not reduce cardiac fibrosis relative to controls. However, there was a 5.9-fold increase of total fibrosis in UW hearts, compared to NIA. Diet composition was a prominent difference between the studies; therefore, we used the NHP diets to characterize diet-associated molecular and functional changes in the hearts of mice. Consistent with the findings from the NHP samples, mice fed UW or a modified NIA diet with increased sucrose and fat developed greater cardiac fibrosis compared to the NIA diet, and transcriptomics analysis revealed diet-induced activation of myocardial oxidative phosphorylation and cardiac muscle contraction pathways.
Niranjana Natarajan, Ana Vujic, Jishnu Das, Annie C. Wang, Krystal K. Phu, Spencer H. Kiehm, Elisabeth M. Ricci-Blair, Anthony Y. Zhu, Kelli L. Vaughan, Ricki J. Colman, Julie A. Mattison, Richard T. Lee
Tumor-infiltrating B-cells (TIL-B) in breast cancer (BC) have previously been associated with improved clinical outcomes; however, their role(s) in tumor immunity is not currently well known. This study confirms and extends the correlation between higher TIL-B densities and positive outcomes through an analysis of HER2-positive and triple-negative BC patients from the BIG 02-98 clinical trial (10yr mean follow-up). Fresh tissue analyses identify an increase in TIL-B density in untreated primary BC compared to normal breast tissues, which is associated with global, CD4+ and CD8+ TIL, higher tumor grades, higher proliferation and hormone receptor negativity. All B-cell differentiation stages are detectable but significant increases in memory TIL-B are consistently present. BC with higher infiltrates are specifically characterized by germinal center TIL-B, which in turn are correlated with TFH TIL and antibody-secreting TIL-B principally located in tertiary lymphoid structures. Some TIL-B also interact directly with tumor cells. Functional analyses reveal TIL-B are responsive to BCR stimulation ex vivo, express activation markers and produce cytokines and immunoglobulins despite reduced expression of the antigen-presenting molecules HLA-DR and CD40. Overall, these data support the concept that ongoing humoral immune responses are generated by TIL-B and help to generate effective anti-tumor immunity at the tumor site.
Soizic Garaud, Laurence Buisseret, Cinzia Solinas, Chunyan Gu-Trantien, Alexandre de Wind, Gert Van den Eynden, Celine Naveaux, Jean-Nicolas Lodewyckx, Anaïs Boisson, Hugues Duvillier, Ligia Craciun, Lieveke Ameye, Isabelle Veys, Marianne Paesmans, Denis Larsimont, Martine Piccart-Gebhart, Karen Willard-Gallo
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