Eran Elinav

Intestinal Lamina Propria Dendritic Cell Subsets Have Different Origin and Functions

The intestinal immune system discriminates between tolerance toward the commensal microflora and robust responses to pathogens. Maintenance of this critical balance is attributed to mucosal dendritic cells (DCs) residing in organized lymphoid tissue and dispersed in the subepithelial lamina propria. In situ parameters of lamina propria DCs (lpDCs) remain poorly understood. Here, we combined conditional cell ablation and precursor-mediated in vivo reconstitution to establish that lpDC subsets have distinct origins and functions. CD103(+) CX(3)CR1(-) lpDCs arose from macrophage-DC precursors (MDPs) via DC-committed intermediates (pre-cDCs) through a Flt3L growth-factor-mediated pathway. CD11b(+) CD14(+) CX(3)CR1(+) lpDCs were derived from grafted Ly6C(hi) but not Ly6C(lo) monocytes under the control of GM-CSF. Mice reconstituted exclusively with CX(3)CR1(+) lpDCs when challenged in an innate colitis model developed severe intestinal inflammation that was driven by graft-derived TNF-alpha-secreting CX(3)CR1(+) lpDCs. Our results highlight the critical importance of the lpDC subset balance for robust gut homeostasis.

Pegylated leptin antagonist is a potent orexigenic agent: preparation and mechanism of activity.

Leptin, a pleiotropic adipokine, is a central regulator of appetite and weight and a key immunomodulatory protein. Although inborn leptin deficiency causes weight gain, it is unclear whether induced leptin deficiency in adult wild-type animals would be orexigenic. Previous work with a potent competitive leptin antagonist did not induce a true metabolic state of leptin deficiency in mice because of a short circulating half-life. In this study, we increased the half-life of the leptin antagonist by pegylation, which resulted in significantly increased bioavailability and retaining of antagonistic activity. Mice administered the pegylated antagonist showed a rapid and dramatic increase in food intake with weight gain. Resulting fat was confined to the mesenteric region with no accumulation in the liver. Serum cholesterol, triglyceride, and hepatic aminotransferases remained unaffected. Weight changes were reversible on cessation of leptin antagonist treatment. The mechanism of severe central leptin deficiency was found to be primarily caused by blockade of transport of circulating leptin across the blood-brain barrier with antagonisms at the arcuate nucleus playing a more minor role. Altogether we introduce a novel compound that induces central and peripheral leptin deficiency. This compound should be useful in exploring the involvement of leptin in metabolic and immune processes and could serve as a therapeutic for the treatment of cachexia.

Role of the intestinal microbiome in liver disease.

The liver integrates metabolic outcomes with nutrient intake while preventing harmful signals derived from the gut to spread throughout the body. Direct blood influx from the gastrointestinal tract through the portal vein makes the liver a critical firewall equipped with a broad array of immune cells and innate immune receptors that recognize microbial-derived products, microorganisms, toxins and food antigens that have breached the intestinal barrier. An overwhelming amount of evidence obtained in the last decade indicates that the intestinal microbiota is a key component of a wide variety of physiological processes, and alterations in the delicate balance that represents the intestinal bacterial communities are now considered important determinants of metabolic syndrome and immunopathologies. Moreover, it is now evident that the interaction between the innate immune system and the intestinal microbiota during obesity or autoimmunity promotes chronic liver disease progression and therefore it might lead to novel and individualized therapeutic approaches. In this review, we discuss a growing body of evidence that highlights the central relationship between the immune system, the microbiome, and chronic liver disease initiation and progression.

Amelioration of Colitis by Genetically Engineered Murine Regulatory T Cells Redirected by Antigen-Specific Chimeric Receptor

BACKGROUND & AIMSnThe therapeutic application of regulatory T cells (Tregs) for the treatment of inflammatory diseases is limited by the scarcity of antigen-specific Tregs. A preferred approach to endow effector T cells (Teff) with a desired specificity uses chimeric immune receptors with antibody-type specificity. Accordingly, employing such chimeric immune receptors to redirect Tregs to sites of inflammation may be a useful therapeutic approach to alleviate a broad scope of diseases in which an uncontrolled inflammatory response plays a major role.nnnMETHODSnTo enable application of the approach in clinical setting, which requires the genetic modification of the patients own Tregs, we describe here a novel protocol that allows the efficient retroviral transduction and 2,4,6-trinitrophenol-specific expansion of murine naturally occurring regulatory T cells (nTregs), with a 2,4,6-trinitrophenol-specific tripartite chimeric receptor.nnnRESULTSnTransduced Tregs maintained their Foxp3 level, could undergo repeated expansion upon ex vivo encounter with their cognate antigen in a major histocompatibility complex-independent, costimulation-independent, and contact-dependent manner and specifically suppressed Teff cells. Adoptive transfer of small numbers of the transduced nTregs was associated with antigen-specific, dose-dependent amelioration of trinitrobenzenesulphonic acid colitis.nnnCONCLUSIONSnThis study demonstrates that nTregs can be efficiently transduced to express functional, antigen-specific chimeric receptors that enable the specific suppression of effector T cells both in vitro and in vivo. This approach may enable future cell-based therapeutic application in inflammatory bowel disease, as well as other inflammatory disorders.

Competitive inhibition of leptin signaling results in amelioration of liver fibrosis through modulation of stellate cell function

Leptin signaling is involved in T‐cell polarization and is required for profibrotic function of hepatic stellate cells (HSCs). Leptin‐deficient ob/ob mice do not develop liver fibrosis despite the presence of severe long‐standing steatohepatitis. Here, we blocked leptin signaling with our recently generated mouse leptin antagonist (MLA), and examined the effects on chronic liver fibrosis in vivo using the chronic thioacetamide (TAA) fibrosis model, and in vitro using freshly‐isolated primary HSCs. In the chronic TAA fibrosis model, leptin administration was associated with significantly enhanced liver disease and a 100% 5‐week to 8‐week mortality rate, while administration or coadministration of MLA markedly improved survival, attenuated liver fibrosis, and reduced interferon γ (IFN‐γ) levels. No significant changes in weight, serum cholesterol, or triglycerides were noted. In vitro administration of rat leptin antagonist (RLA), either alone or with leptin, to rat primary HSCs reduced leptin‐stimulated effects such as increased expression of α‐smooth muscle actin (α‐SMA), and activation of α1 procollagen promoter. Conclusion: Inhibition of leptin‐enhanced hepatic fibrosis may hold promise as a future antifibrotic therapeutic modality. (HEPATOLOGY 2009;49:278‐286.)

Redirection of Regulatory T Cells With Predetermined Specificity for the Treatment of Experimental Colitis in Mice

BACKGROUND & AIMSnTreatment with ex vivo expanded regulatory T cells (Tregs) is regarded as a promising therapeutic approach in inflammatory bowel disease but is hampered by impaired Treg accumulation and function at inflammatory sites. We aim to study whether antigen-specific redirected Tregs can overcome these limitations.nnnMETHODSnWe developed transgenic mice whose T cells, including Tregs, express chimeric receptor (CR) made of antibody variable region as recognition unit and T-cell stimulatory and costimulatory domains to activate specifically in response to the predetermined model antigen 2,4,6-trinitrophenol (TNP).nnnRESULTSnTNP-specific CR-bearing Tregs were potently and specifically activated by exogenous TNP and suppressed effector T cells in the absence of costimulatory B7-CD28 interaction. TNP-specific transgenic (Tg) mice were resistant to 2,4,6-trinitrobenzene sulphonic acid (TNBS) colitis but not to other hapten-mediated colitis. Adoptive transfer of CR-bearing Tregs to wild-type mice with TNBS colitis was associated with significant amelioration of colitis and improved survival. Although TNP-specific CR-bearing Tregs did not suppress oxazolone colitis, they cured it after addition of traces of TNBS to oxazolone-inflamed colons, demonstrating a bystander effect. In vivo imaging of adoptively transferred CR-bearing Tregs demonstrated that they preferentially migrate to TNBS-induced colonic mucosal lesions within hours of induction of colitis.nnnCONCLUSIONSnTregs can be redirected with specificity distinct from that of pathogenic lymphocytes, accumulate at colonic inflammatory lesions, and suppress effector T cells in a specific, nonmajor histocompatibility complex-restricted, and noncostimulatory-dependent manner, resulting in significant amelioration of colitis. Hopefully, this approach will lead to a novel therapy for inflammatory bowel disease, as well as other inflammatory diseases.

Antibiotic prophylaxis for spontaneous bacterial peritonitis in cirrhotic patients with ascites, without gastro‐intestinal bleeding

BACKGROUNDnSpontaneous bacterial peritonitis is frequent among cirrhotic patients, associated with significant morbidity and mortality. Selective intestinal decontamination employing antibiotics is a proposed prophylactic measure. While data regarding this modality among cirrhotic patients with gastrointestinal bleeding exist, there is insufficient data synthesis regarding cirrhotic patients with ascites and no gastrointestinal bleeding.nnnOBJECTIVESnTo assess whether antibiotic prophylaxis decreases spontaneous bacterial peritonitis and mortality among cirrhotic patients with ascites and no gastrointestinal bleeding.nnnSEARCH STRATEGYnWe identified relevant randomised trials by searching trial registries of The Cochrane Hepato-Biliary Group and The Cochrane Collaboration, medical literature search engines, and reviewing all literature we found on the topic until February 2009.nnnSELECTION CRITERIAnWe searched for randomised clinical trials assessing prophylactic treatment among adult cirrhotic patients with ascites and no gastrointestinal bleeding, comparing antibiotic therapy with no intervention, placebo, or with another antibiotic regimen.nnnDATA COLLECTION AND ANALYSISnThree independent authors searched for and collected the trials and extracted relevant data. Four other independent authors validated the findings and assessed them. The studies were assessed for design, patient and intervention characteristics, and quality. A meta-analysis was performed to estimate measures of association between antibiotic prophylaxis and spontaneous bacterial peritonitis or mortality.nnnMAIN RESULTSnNine trials were included in the review. Seven trials, comparing antibiotics to placebo or no treatment, were meta-analysed. Systematic bias in design or publication is suggested by trial results. The randomisation results suggest that the probability that true randomisation took place in all trials is very small and the report of most trials regarding design was poor. The proportion of participants with spontaneous bacterial peritonitis varied between the trials from 15% to 50%. The calculated relative risks (95% confidence interval) of spontaneous bacterial peritonitis and mortality among patients treated with antibiotics compared with no treatment/placebo were 0.20 (0.11 to 0.37) and 0.61 (0.43 to 0.87). There were very few reports of adverse events.nnnAUTHORS CONCLUSIONSnThe pooled estimates suggest that antibiotic prophylaxis might be prudent among cirrhotic patients with ascites and no gastrointestinal bleeding. However, poor trial methodology and report coupled with findings suggesting systematic bias in publication and design reflect the fragility of these findings. Potential hazard to society and the patients themselves from resistant pathogens should be considered when promoting long-lasting antibiotic prophylaxis. It seems that recommending antibiotic prophylaxis is still far from being a substantiated prevention strategy. Trials of better design, well reported, and of longer follow-up are greatly needed.

Metagenomic cross-talk: the regulatory interplay between immunogenomics and the microbiome

The human microbiome, often referred to as the ‘second genome’, encompasses up to 100-fold more genes than the host genome. In contrast to the human genome, the microbial genome is flexible and amenable to change during the host’s lifetime. As the composition of the microbial metagenome has been associated with the development of human disease, the mechanisms controlling the composition and function of the metagenome are of considerable interest and therapeutic potential. In the past few years, studies have revealed how the host immune system is involved in determining the microbial metagenome, and, in turn, how the microbiota regulates gene expression in the immune system. This species-specific bidirectional interaction is required for homeostatic health, whereas aberrations in the tightly controlled regulatory circuits that link the host immunogenome and the microbial metagenome drive susceptibility to common human diseases. Here, we summarize some of the major principles orchestrating this cross-talk between microbial and host genomes, with a special focus on the interaction between the intestinal immune system and the gut microbiome. Understanding the reciprocal genetic and epigenetic control between host and microbiota will be an important step towards the development of novel therapies against microbiome-driven diseases.

Environmental factors dominate over host genetics in shaping human gut microbiota composition

Human gut microbiome composition is shaped by multiple host intrinsic and extrinsic factors, but the relative contribution of host genetic compared to environmental factors remains elusive. Here, we genotyped a cohort of 696 healthy individuals from several distinct ancestral origins and a relatively common environment, and demonstrate that there is no statistically significant association between microbiome composition and ethnicity, single nucleotide polymorphisms (SNPs), or overall genetic similarity, and that only 5 of 211 (2.4%) previously reported microbiome-SNP associations replicate in our cohort. In contrast, we find similarities in the microbiome composition of genetically unrelated individuals who share a household. We define the term biome-explainability as the variance of a host phenotype explained by the microbiome after accounting for the contribution of human genetics. Consistent with our finding that microbiome and host genetics are largely independent, we find significant biome-explainability levels of 16-33% for body mass index (BMI), fasting glucose, high-density lipoprotein (HDL) cholesterol, waist circumference, waist-hip ratio (WHR), and lactose consumption. We further show that several human phenotypes can be predicted substantially more accurately when adding microbiome data to host genetics data, and that the contribution of both data sources to prediction accuracy is largely additive. Overall, our results suggest that human microbiome composition is dominated by environmental factors rather than by host genetics.

Utilization of Murine Laparoscopy for Continuous In-Vivo Assessment of the Liver in Multiple Disease Models

Background Current strategies for follow up of murine models of liver disease are flawed by inability to continuously monitor disease progression in the tissue level, and necessitate sacrifice of animals for tissue sampling. Aims In this study we aimed at developing a safe repetitive tool for sampling livers in vivo, by utilization of a miniaturized endoscopy system for laparoscopic liver biopsies and for injection of tumor cells into livers. Results We report the development of a protocol for murine laparoscopy that allows repeated visualization of murine intra-abdominal organs. The system enables safe and repeated liver biopsies in mice and rats, yielding adequate tissue for histological staining and RNA extraction. In addition, injection of tumor cells into livers facilitates under-vision implantation of hepatic tumors in liver, followed by visualization of tumor growth. Conclusions Murine laparoscopy may be employed as a novel imaging modality for continuous assessment and manipulation of chronic liver disease models.