Deciphering the impact of the mitochondrial negative regulator MCJ on host-microbiota interactions in experimental ulcerative colitis

270 p.La colitis ulcerosa (CU) es un trastorno inflamatorio crónico que afecta al intestino. Aunque la causa es desconocida, es una enfermedad multifactorial que está desencadenada por el sistema inmune, factores medioambientales, variaciones genéticas y comunidad microbiana. En trabajos recientes se ha relacionado la disfunción mitocondrial con la EII. Además, se ha descrito que existe una interacción bidireccional entre las mitocondrias y la microbiota, que parece jugar un papel clave en la enfermedad. Por todo ello, con el fin de reducir la heterogeneidad clínica de los pacientes en este trabajo, hemos utilizado un modelo de ratón deficiente en MCJ (Methylation-controlled J protein), una proteína localizada en la membrana interna de la mitocondria que actúa como un regulador negativo del complejo I de la cadena respiratoria, para estudiar el papel de la disfunción mitocondrial dentro de la CU. La inducción de una colitis aguda en ratones deficientes en MCJ resultó en un fenotipo de enfermedad más grave, como consecuencia de la disfunción mitocondrial y una modificación de la composición microbiana. De hecho, se observó un aumento de la presencia de Ruminococcus gnavus, una especie comúnmente incrementada en la microbiota intestinal de pacientes con EII. Los ratones deficientes en MCJ mostraron en el colon una sobre expresión de Timp3, lo cual condujo a la inhibición de la actividad de TACE que es la enzima encargada de la liberación del Tnf unido a la membrana celular, confirmando los hallazgos descritos previamente en macrófagos derivados de médula ósea. Además, los análisis de expresión génica en pacientes que sufren CU también mostraron niveles reducidos de MCJ y una mayor expresión de TIMP3 en el colon. Nuestros resultados también implican los receptores Fc¿R de los macrófagos intestinales en la patogénesis de la CU y en el fallo a la respuesta a terapias biológicas a causa de la disfunción mitocondrial. Finalmente, la inducción de la colitis crónica demostró que el tejido del colon es capaz de adaptarse de una manera dinámica a los cambios del entorno adaptándose metabólicamente a las nuevas condiciones, desapareciendo el aumento de severidad por la falta de MCJ. En resumen, los datos obtenidos posibilitan el establecimiento de nuevas líneas de investigación para predecir en estadios tempranos la evolución de la enfermedad y la respuesta a la terapia con agentes anti-TNF en pacientes con CU. Por otra parte, este estudio nos ha permitido identificar posibles firmas microbianas en las heces asociadas con la progresión de la enfermedad y la respuesta a terapias, que podrían servir como biomarcadores predictivos, permitiendo la estratificación de los pacientes.CICbioGUN

Metalotioneinen mailak eta metalen banaketa Ag eta Cu pean mantendutako ostretan

Laburpena: Ikerlan hau itsasoaren kutsaduran zentratzen da, itsasoak arriskutsuak eta kaltegarriak diren substantziak jasotzen dituelako, bai ibaietako ekarpenarengatik, baita zuzeneko karguetatik. Organismo behale gisa ostrak erabiliko dira, izaki iragazleak izanik elikagaien irenstearen ondorioz medio urtsutik kutsatzaileak metatzen dituztelako eta gainera, ingurugiroan dauden mailen gainetik kontzentratzen dituztelako. Ostretan zilarra (Ag) eta kobrea (Cu) bezalako metalen eragina aztertuko da, horretarako, metalen aurreko esposizio biomarkatzaileak erabiliko dira. Besteak beste, autometalografia erabiliko da metalen mailak eta banaketa ostren ehunetan neurtzeko eta metalotioneinen mailak ere neurtuko dira. Ostrek metalekiko izan duten esposizio denborak 0, 6, 7, 13 eta 14 egunetakoak izan dira, bai itsasoko eta bai estuarioko uretan mantenduz. Emaitzak ikusita behatu da estuarioko laginek, normalean, metal kontzentrazio baxuagoak aurkezten dituztela, eta zenbait kasutan kontrolek metalen pean jarritako laginak baino metal kontzentrazio handiagoak erakutsi dituzte aste bat igaro eta gero. Bigarren astean, metal mailen jaitsiera bat behatu da. Beste alde batetik, metalotioneinetan neurtutako indukzio mailak oso baxuak izan dira. Metalen distribuzioa ehun desberdinetan aldez aurretik beste molusku batzuetan deskribatutakoen oso antzekoa izan da ostretan

Hesteetako hanturazko gaixotasuna: patogenia, tratamenduak eta mikrobiotan oinarritutako biomarkatzaileak

Inflammatory bowel disease (IBD) encompasses two types of idiopathic intestinal diseases, ulcerative colitis (UC) and Crohn’s disease (CD). Both are chronic, heterogeneous, and severe inflammatory disorders that primarily affect the intestine. IBD has emerged as a global disease with a sharp increase in worldwide incidence and prevalence. Although the specific underlying cause of UC is unknown, it is considered the result of a complex interaction between the microbiota, immune system, host genetics and environmental factors. The advent of new technologies has enabled the identification of disturbed microbiota composition and function and reduced microbial diversity in IBD pa-tients, termed dysbiosis. Obtained clinical and experimental data points dysbiosis as a key player in IBD pathogenesis, but it is still unclear whether it is the cause of consequence. Although a wide range of therapies are approved for use as treatment for IBD, there is no cure. TNF inhibitors are frequently used to induce clinical remission in severe patients, however a roughly one third of the patients may not respond, and another third may lose response over time. In addition, the cost associated with IBD treatments is increasing over time, mainly due the costs associated with the biologic treatment. Emerging evidence has pointed towards gut microbiota to find a set of biomarkers for diagnosis, and for prediction of disease severity and infliximab treatment response in IBD patients. The human fecal microbiota harbors promising and non-invasive biomarkers, which emphasizes its potential ability to stratify IBD patients and apply personalized therapy for optimal outcomes.; Hesteetako hanturazko gaixotasunak (HHG, ingelesez IBD) bi gaixotasun idiopatiko ezberdin biltzen ditu, ultzeradun kolitisa (UK) eta Crohn-en gaixotasuna (CG). Biak batez ere hesteari eragiten dioten gaixotasun kronikoak, heterogeneoak eta larriak dira. HHGa handitzen ari den gaixotasun globala da, eta haren intzidentzia eta prebalentzia areagotuz doa mundu osoan. UKaren kausa zehatza ezezaguna bada ere, uste da mikrobiotaren, sistema immunearen, ostalariaren genetikaren eta ingurumen-faktoreen arteko elkarrekintza konplexuaren ondorioz sortzen dela. Genomikako teknologia berriek HHG-pazienteetan mikrobiotaren osaeran eta funtzioan aldaketak identifikatzea ahalbidetu dute. Aldaketa horiek disbiosia bezala ezagutzen dira. Lortutako datu kliniko eta esperimentalak, disbiosia, HHGen funtsezko eragile gisa finkatzen dute, nahiz eta oraindik argi egon ez kausa edo ondorioa den. Gaur egun H HGaren aurkako tratamendu ugari dauden arren, ez dago sendabide-H HGaren aurkako tratamendu ugari dauden arren, ez dago sendabide- aurkako tratamendu ugari dauden arren, ez dago sendabide-rik. Paziente larrietan sintomatologia arintzeko, TNF inhibitzaileak dira gehien erabiltzen den tratamendua. Hala ere, pazienteen heren batek ez dio tratamenduari erantzuten eta beste heren batek denborarekin erantzuna galtzen du. Gainera, tratamenduei lotutako kostu ekonomikoa eten-gabe handitzen ari da, batez ere tratamendu biologikoen kostuengatik. HHGa duten gaixoen hesteetako mikrobiota baliagarri bilakatzen ari da gaixotasuna diagnostikatzeko, gaixotasunaren larritasuna zehazteko eta tratamenduarekiko erantzuna iragartzeko, hau da, mikrobiotaren osaera biomarkatzaile gisa erabil daiteke. Izan ere, giza gorotzetako mikrobiota biomarkatzaile esperantzagarri eta ez-inbaditzaile bihurtu da. Mikro-biotaren azterketak HHGa duten gaixoak sailkatzeko aukerak gehitzen ditu, eta tratamendu pertsonalizatuen aukera zabaldu

TLR2 and TLR4 interact with sulfide system in the modulation of mouse colonic motility

Background H2S is a neuromodulator that may inhibit intestinal motility. H2S production in colon is yielded by cystathionine beta-synthase (CBS) and cystathionine gamma-lyase (CSE) enzymes and sulfate-reducing bacteria (SRB). Toll-like receptors (TLRs) recognize intestinal microbiota. The aim of this work was to evaluate the influence of TLR2 and TLR4 on the endogenous and SRB-mediated synthesis of H2S and its consequences on the colonic motility of mouse. Methods Muscle contractility studies were performed in colon from WT, Tlr2(-/-), and Tlr4(-/-) mice. The mRNA levels of TLR2, TLR4, CBS, CSE, and SRB were measured by real-time PCR. Free sulfide levels in colon and feces were determined by colorimetric assays. Results NaHS and GYY4137, donors of H2S, reduced the contractility of colon. Aminooxyacetic acid (AOAA), inhibitor of CBS, and D-L propargylglycine (PAG), inhibitor of CSE, increased the contractility of colon. In vivo treatment with NaHS or GYY4137 inhibited the spontaneous contractions and upregulated TLR2 expression. The in vivo activation of TLR4 with lipopolysaccharide increased the contractile response to PAG, mRNA levels of CSE, and the free sulfide levels of H2S in colon. In Tlr2(-/-) and Tlr4(-/-) mice, the contractions induced by AOAA and PAG and mRNA levels of CBS and CSE were lower with respect to WT mice. Deficiency of TLR2 or TLR4 provokes alterations in free sulfide levels and SRB of colon. Conclusions and Inferences Our study demonstrates interaction between TLR2 and TLR4 and the sulfide system in the regulation of colonic motility and contributes to the pathophysiology knowledge of intestinal motility disorders

Peripheral Blood Mononuclear Cells (PBMC) Microbiome is Not Affected by Colon Microbiota in Healthy Goats

BACKGROUND: The knowledge about blood circulating microbiome and its functional relevance in healthy individuals remains limited. An assessment of changes in the circulating microbiome was performed by sequencing peripheral blood mononuclear cells (PBMC) bacterial DNA from goats supplemented or not in early life with rumen liquid transplantation. RESULTS: Most of the bacterial DNA associated to PBMC was identified predominantly as Proteobacteria (55%) followed by Firmicutes (24%), Bacteroidetes (11%) and Actinobacteria (8%). The predominant genera found in PBMC samples were Pseudomonas, Prevotella, Sphingomonas, Acinetobacter, Corynebacterium and Ruminococcus. Other genera such as Butyrivibrivio, Bifidobacterium, Dorea and Coprococcus were also present in lower proportions. Several species known as blood pathogens or others involved in gut homeostasis such as Faecalibacterium prausnitzii were also identified. However, the PBMC microbiome phylum composition differed from that in the colon of goats (P≤0.001), where Firmicutes was the predominant phylum (83%). Although, rumen liquid administration in early-life altered bacterial community structure and increased Tlr5 expression (P=0.020) in colon pointing to higher bacterial translocation, less than 8% of OTUs in colon were also observed in PBMCs. CONCLUSIONS: Data suggest that in physiological conditions, PBMC microbiome differs from and is not affected by colon gut microbiota in small ruminants. Although, further studies with larger number of animals and covering other animal tissues are required, results point to a common circulating bacterial profile on mammals being phylum Proteobacteria, and genera Pseudomonas and Prevotella the most abundants. All suggest that PBMC microbiome in healthy ruminants could be implicated in homeostatic condition. This study expands our knowledge about PBMC microbiome contribution to health in farm animals.This work was supported by grants from the Spanish Ministry of Science and Innovation (MCI) co-financed with FEDER funds [AGL2017-86757- to LA, AGL2017-86938-R to DRY]. Other contributions were SAF2015-65327-R to JA and SAF2015-73549-JIN to HR. LA is a Ramón y Cajal fellow [RYC-2013-13666] from the Spanish Ministry of Science and Innovation. APC is a recipient of a fellowship from the University of the Basque Country. We thank the MCI for the Severo Ochoa Excellence accreditation (SEV-2016-0644) and the Basque Department of Industry, Tourism and Trade (Etortek and Elkartek programs

Mitochondrial complex I dysfunction alters the balance of soluble and membrane-bound TNF during chronic experimental colitis.

Inflammatory bowel disease (IBD) is a complex, chronic, relapsing and heterogeneous disease induced by environmental, genomic, microbial and immunological factors. MCJ is a mitochondrial protein that regulates the metabolic status of macrophages and their response to translocated bacteria. Previously, an acute murine model of DSS-induced colitis showed increased disease severity due to MCJ deficiency. Unexpectedly, we now show that MCJ-deficient mice have augmented tumor necrosis factor alpha converting enzyme (TACE) activity in the context of chronic inflammation. This adaptative change likely affects the balance between soluble and transmembrane TNF and supports the association of the soluble form and a milder phenotype. Interestingly, the general shifts in microbial composition previously observed during acute inflammation were absent in the chronic model of inflammation in MCJ-deficient mice. However, the lack of the mitochondrial protein resulted in increased alpha diversity and the reduction in critical microbial members associated with inflammation, such as Ruminococcus gnavus, which could be associated with TACE activity. These results provide evidence of the dynamic metabolic adaptation of the colon tissue to chronic inflammatory changes mediated by the control of mitochondrial function

Microbioma de las células mononucleares de sangre periférica en cabras

1 página.- Trabajo presentado a las: XIX Jornadas sobre Producción Animal AIDA. On line. Zaragoza, España, 1-2 junio 2021.Financiado por los proyectos AGL2017-86757 y AGL2017-86938-R

Borrelia burgdorferi infection induces long-term memory-like responses in macrophages with tissue-wide consequences in the heart

Lyme carditis is an extracutaneous manifestation of Lyme disease characterized by episodes of atrioventricular block of varying degrees and additional, less reported cardiomyopathies. The molecular changes associated with the response to Borrelia burgdorferi over the course of infection are poorly understood. Here, we identify broad transcriptomic and proteomic changes in the heart during infection that reveal a profound down-regulation of mitochondrial components. We also describe the long-term functional modulation of macrophages exposed to live bacteria, characterized by an augmented glycolytic output, increased spirochetal binding and internalization, and reduced inflammatory responses. In vitro, glycolysis inhibition reduces the production of tumor necrosis factor (TNF) by memory macrophages, whereas in vivo, it produces the reversion of the memory phenotype, the recovery of tissue mitochondrial components, and decreased inflammation and spirochetal burdens. These results show that B. burgdorferi induces long-term, memory-like responses in macrophages with tissue-wide consequences that are amenable to be manipulated in vivo.Supported by grants from the Spanish Ministry of Science, Innovation and Universities (MCIU) co-financed with FEDER funds (SAF2015-65327-R and RTI2018-096494-B-100 to JA; BFU2016-76872-R to EB, AGL2017-86757-R to LA, SAF2017-87301-R to MLMC, SAF2015-64111-R to AP, SAF2015-73549-JIN to HR), Instituto de Salud Carlos III (PIE13/0004 to AP), the Basque Government Department of Health (2015111117 to LA), the Basque Foundation for Innovation and Health Research (BIOEF), through the EiTB Maratoia grant BIO15/CA/016/BS to MLMC, the regional Government of Andalusia co-funded by CEC and FEDER funds (Proyectos de Excelencia P12-CTS-2232) and Fundación Domingo Martínez (to AP). LA is supported by the Ramon y Cajal program (RYC-2013-13666). DB, MMR and TMM are recipients of MCIU FPI fellowships. ACG and AP are recipients of fellowships form the Basque Government. APC is a recipient of a fellowship from the University of the Basque Country. We thank the MCIU for the Severo Ochoa Excellence accreditation (SEV-2016-0644), the Basque Department of Industry, Tourism and Trade (Etortek and Elkartek programs), the Innovation Technology Department of the Bizkaia Province and the CIBERehd network. DB and JA are supported by a grant from the Jesús de Gangoiti Barrera Foundation

The commensal bacterium Lactiplantibacillus plantarum imprints innate memory-like responses in mononuclear phagocytes

Gut microbiota is a constant source of antigens and stimuli to which the resident immune system has developed tolerance. However, the mechanisms by which mononuclear phagocytes, specifically monocytes/macrophages, cope with these usually pro-inflammatory signals are poorly understood. Here, we show that innate immune memory promotes anti-inflammatory homeostasis, using as model strains of the commensal bacterium Lactiplantibacillus plantarum. Priming of monocytes/macrophages with bacteria, especially in its live form, enhances bacterial intracellular survival and decreases the release of pro-inflammatory signals to the environment, with lower production of TNF and higher levels of IL-10. Analysis of the transcriptomic landscape of these cells shows downregulation of pathways associated with the production of reactive oxygen species (ROS) and the release of cytokines, chemokines and antimicrobial peptides. Indeed, the induction of ROS prevents memory-induced bacterial survival. In addition, there is a dysregulation in gene expression of several metabolic pathways leading to decreased glycolytic and respiratory rates in memory cells. These data support commensal microbe-specific metabolic changes in innate immune memory cells that might contribute to homeostasis in the gut.Supported by grants from the Spanish Ministry of Science, Innovation and Universities (MCIU) co-financed with FEDER funds (RTI2018-096494-B-100 to JA; BFU2016-76872-R to EB; AGL2017-86757-R to LA; SAF2015-73549-JIN to HR; SAF2016–77433-R and PID2019-110240RB-I00 to RPR). AP is supported by a Postdoctoral Fellowship from the Basque Government. DB and TMM are recipients of MCIU FPI fellowships. APC is a recipient of a fellowship from the University of the Basque Country. LA and RPR are supported by the Ramon y Cajal program from the Spanish Ministry of Economy and Competitiveness. We thank the MCIU for the Severo Ochoa Excellence accreditation (SEV-2016-0644), the Basque Department of Industry, Tourism and Trade (Etortek and Elkartek programs) and the Innovation Technology Department of the Bizkaia Province. This work was further supported by grants from the Jesús de Gangoiti Barrera Foundation.Peer reviewe

The outcome of boosting mitochondrial activity in alcohol-associated liver disease is organ-dependent.

BACKGROUND AND AIMS Alcohol-associated liver disease (ALD) accounts for 70% of liver-related deaths in Europe, with no effective approved therapies. Although mitochondrial dysfunction is one of the earliest manifestations of alcohol-induced injury, restoring mitochondrial activity remains a problematic strategy due to oxidative stress. Here, we identify methylation-controlled J protein (MCJ) as a mediator for ALD progression and hypothesize that targeting MCJ may help in recovering mitochondrial fitness without collateral oxidative damage. APPROACH AND RESULTS C57BL/6 mice [wild-type (Wt)] Mcj knockout and Mcj liver-specific silencing (MCJ-LSS) underwent the NIAAA dietary protocol (Lieber-DeCarli diet containing 5% (vol/vol) ethanol for 10 days, plus a single binge ethanol feeding at day 11). To evaluate the impact of a restored mitochondrial activity in ALD, the liver, gut, and pancreas were characterized, focusing on lipid metabolism, glucose homeostasis, intestinal permeability, and microbiota composition. MCJ, a protein acting as an endogenous negative regulator of mitochondrial respiration, is downregulated in the early stages of ALD and increases with the severity of the disease. Whole-body deficiency of MCJ is detrimental during ALD because it exacerbates the systemic effects of alcohol abuse through altered intestinal permeability, increased endotoxemia, and dysregulation of pancreatic function, which overall worsens liver injury. On the other hand, liver-specific Mcj silencing prevents main ALD hallmarks, that is, mitochondrial dysfunction, steatosis, inflammation, and oxidative stress, as it restores the NAD + /NADH ratio and SIRT1 function, hence preventing de novo lipogenesis and improving lipid oxidation. CONCLUSIONS Improving mitochondrial respiration by liver-specific Mcj silencing might become a novel therapeutic approach for treating ALD.This work was supported by grants from Ministerio de Ciencia e Innovación, Programa Retos-Colaboración RTC2019-007125-1 (for Jorge Simon and Maria Luz Martinez-Chantar); Ministerio de Economía, Industria y Competitividad, Retos a la Sociedad AGL2017- 86927R (for F.M.); Instituto de Salud Carlos III, Proyectos de Investigación en Salud DTS20/00138 and DTS21/00094 (for Jorge Simon and Maria Luz Martinez-Chantar, and Asis Palazon. respectively); Instituto de Salud Carlos III, Fondo de Investigaciones Sanitarias co-founded by European Regional Development Fund/European Social Fund, “Investing in your future” PI19/00819, “Una manera de hacer Europa” FIS PI20/00765, and PI21/01067 (for Jose J. G. Marin., Pau Sancho-Bru,. and Mario F. Fraga respectively); Departamento de Industria del Gobierno Vasco (for Maria Luz Martinez-Chantar); Asturias Government (PCTI) co-funding 2018-2023/ FEDER IDI/2021/000077 (for Mario F. Fraga.); Ministerio de Ciencia, Innovación y Universidades MICINN: PID2020-117116RB-I00, CEX2021-001136-S PID2020-117941RB-I00, PID2020-11827RB-I00 and PID2019-107956RA-100 integrado en el Plan Estatal de Investigación Científica y Técnica y Innovación, cofinanciado con Fondos FEDER (for Maria Luz Martinez-Chantar, Francisco J Cubero., Yulia A Nevzorova and Asis Palazon); Ayudas Ramón y Cajal de la Agencia Estatal de Investigación RY2013-13666 and RYC2018- 024183-I (for Leticia Abecia and Asis Palazon); European Research Council Starting Grant 804236 NEXTGEN-IO (for Asis Palazon); The German Research Foundation SFB/TRR57/P04, SFB1382-403224013/ A02 and DFG NE 2128/2-1 (for Francisco J Cubero and Yulia A Nevzorova); National Institute of Health (NIH)/National Institute of Alcohol Abuse and Alcoholism (NIAAA) 1U01AA026972-01 (For Pau Sancho-Bru); Junta de Castilla y León SA074P20 (for Jose J. G. Marin); Junta de Andalucía, Grupo PAIDI BIO311 (for Franz Martin); CIBERER Acciones Cooperativas y Complementarias Intramurales ACCI20-35 (for Mario F. Fraga); Ministerio de Educación, Cultura y Deporte FPU17/04992 (for Silvia Ariño); Fundació Marato TV3 201916-31 (for Jose J. G. Marin.); Ainize Pena-Cearra is a fellow of the University of the Basque Country (UPV/ EHU); BIOEF (Basque Foundation for Innovation and Health Research); Asociación Española contra el Cáncer (Maria Luz Martinez-Chantar and Teresa C. Delgado.); Fundación Científica de la Asociación Española Contra el Cáncer (AECC Scientific Foundation) Rare Tumor Calls 2017 (for Maria Luz Martinez-Chantar); La Caixa Foundation Program (for Maria Luz Martinez-Chantar); Proyecto Desarrollo Tecnologico CIBERehd (for Maria Luz Martinez-Chantar); Ciberehd_ISCIII_MINECO is funded by the Instituto de Salud Carlos III.S