Monoacylglycerol lipase reprograms hepatocytes and macrophages to promote liver regeneration.

Background & Aims: Liver regeneration is a repair process in which metabolic reprogramming of parenchymal and inflammatory cells plays a major role. Monoacylglycerol lipase (MAGL) is an ubiquitous enzyme at the crossroad between lipid metabolism and inflammation. It converts monoacylglycerols into free fatty acids and metabolises 2-arachidonoylglycerol into arachidonic acid, being thus the major source of pro-inflammatory prostaglandins in the liver. In this study, we investigated the role of MAGL in liver regeneration. Methods: Hepatocyte proliferation was studied in vitro in hepatoma cell lines and ex vivo in precision-cut human liver slices. Liver regeneration was investigated in mice treated with a pharmacological MAGL inhibitor, MJN110, as well as in animals globally invalidated for MAGL (MAGL-/-) and specifically invalidated in hepatocytes (MAGLHep-/-) or myeloid cells (MAGLMye-/-). Two models of liver regeneration were used: acute toxic carbon tetrachloride injection and two-thirds partial hepatectomy. MAGLMye-/- liver macrophages profiling was analysed by RNA sequencing. A rescue experiment was performed by in vivo administration of interferon receptor antibody in MAGLMye-/- mice. Results: Precision-cut human liver slices from patients with chronic liver disease and human hepatocyte cell lines exposed to MJN110 showed reduced hepatocyte proliferation. Mice with global invalidation or mice treated with MJN110 showed blunted liver regeneration. Moreover, mice with specific deletion of MAGL in either hepatocytes or myeloid cells displayed delayed liver regeneration. Mechanistically, MAGLHep-/- mice showed reduced liver eicosanoid production, in particular prostaglandin E2 that negatively impacts on hepatocyte proliferation. MAGL inhibition in macrophages resulted in the induction of the type I interferon pathway. Importantly, neutralising the type I interferon pathway restored liver regeneration of MAGLMye-/- mice. Conclusions: Our data demonstrate that MAGL promotes liver regeneration by hepatocyte and macrophage reprogramming. Impact and Implications: By using human liver samples and mouse models of global or specific cell type invalidation, we show that the monoacylglycerol pathway plays an essential role in liver regeneration. We unveil the mechanisms by which MAGL expressed in both hepatocytes and macrophages impacts the liver regeneration process, via eicosanoid production by hepatocytes and the modulation of the macrophage interferon pathway profile that restrains hepatocyte proliferation.

MAIT cell inhibition promotes liver fibrosis regression via macrophage phenotype reprogramming.

Recent data have shown that liver fibrosis can regress even at later stages of cirrhosis and shifting the immune response from pro-inflammatory towards a resolutive profile is considered as a promising option. The immune regulatory networks that govern the shift of the inflammatory phenotype and thus potential reversal of liver fibrosis are lesser known. Here we show that in precision-cut human liver slices obtained from patients with end-stage fibrosis and in mouse models, inhibiting Mucosal-Associated Invariant T (MAIT) cells using pharmacological or antibody-driven approaches, limits fibrosis progression and even regresses fibrosis, following chronic toxic- or non-alcoholic steatohepatitis (NASH)-induced liver injury. Mechanistic studies, combining RNA sequencing, in vivo functional studies (performed in male mice) and co-culture experiments indicate that disruption of the MAIT cell-monocyte/macrophage interaction results in resolution of fibrosis both by increasing the frequency of restorative Ly6Clo at the expenses of pro-fibrogenic Ly6Chi monocyte-derived macrophages and promoting an autophagic phenotype in both subsets. Thus, our data show that MAIT cell activation and the consequential phenotype shift of liver macrophages are important pathogenic features of liver fibrosis and could be targeted by anti-fibrogenic therapy.

Isobaric labeling-based quantitative proteomics of FACS-purified immune cells and epithelial cells from the intestine of Crohn's disease patients reveals proteome changes of potential importance in disease pathogenesis.

Crohn's disease (CD) is a chronic condition characterized by recurrent flares of inflammation in the gastrointestinal tract. Disease etiology is poorly understood and is characterized by dysregulated immune activation that progressively destroys intestinal tissue. Key cellular compartments in disease pathogenesis are the intestinal epithelial layer and its underlying lamina propria. While the epithelium contains predominantly epithelial cells, the lamina propria is enriched in immune cells. Deciphering proteome changes in different cell populations is important to understand CD pathogenesis. Here, using isobaric labeling-based quantitative proteomics, we perform an exploratory study to analyze in-depth proteome changes in epithelial cells, immune cells and stromal cells in CD patients compared to controls using cells purified by FACS. Our study revealed increased proteins associated with neutrophil degranulation and mitochondrial metabolism in immune cells of CD intestinal mucosa. We also found upregulation of proteins involved in glycosylation and secretory pathways in epithelial cells of CD patients, while proteins involved in mitochondrial metabolism were reduced. The distinct alterations in protein levels in immune- versus epithelial cells underscores the utility of proteome analysis of defined cell types. Moreover, our workflow allowing concomitant assessment of cell-type specific changes on an individual basis enables deeper insight into disease pathogenesis.

MEFV and NLRP3 Inflammasome Expression Is Attributed to Immature Macrophages and Correlates with Serum Inflammatory Proteins in Crohn´s Disease Patients.

Inflammasomes are intracellular protein complexes whose activation results in proinflammatory cytokines. Inflammasomes are implicated in Crohn´s disease (CD) pathogenesis, yet the contribution of inflammasomes in intestinal epithelial cells (IECs) versus lamina propria (LP) macrophages is poorly understood. Whether inflammasome expression in intestinal tissue reflects the serum inflammatory protein profile of patients is also not known. We aimed to determine the intestinal cell types where inflammasome expression is increased in CD and if they correlate with the serum protein profile. RT-PCR and NanoString nCounter technology were used to characterize inflammasome gene expression in CD patients and controls. The mucosa, LP and IEC cell fractions and FACS-sorted cells were analyzed. Proximity extension assay with a 92-protein panel was used to determine the serum inflammatory protein profile. Compositional analysis was used to correlate ileum inflammasome gene expression with intestinal mononuclear phagocyte populations. We show that NLRP3 and MEFV inflammasome sensors and downstream effector expression including IL-1ß are increased in inflamed mucosa of IBD patients and correlate with disease activity. Inflammasome gene expression increased with the abundance of immature intestinal macrophages, and increased IL-1ß released by CD LP cells correlated with immature macrophage frequency. Inflammasome gene expression was also increased in circulating monocytes, the precursors of immature intestinal macrophages. Finally, the serum inflammatory profile of CD patients correlates with ileal expression of genes related to NLRP3 and MEFV inflammasomes. Overall, we show that MEFV and NLRP3 inflammasome expression in CD intestine is attributed to the accumulation of immature macrophages and correlates with serum inflammatory proteins.

TREM-1+ Macrophages Define a Pathogenic Cell Subset in the Intestine of Crohn's Disease Patients.

BACKGROUND AND AIMS: Uncontrolled activation of intestinal mononuclear phagocytes [MNPs] drives chronic inflammation in inflammatory bowel disease [IBD]. Triggering receptor expressed on myeloid cells 1 [TREM-1] has been implicated in the pathogenesis of IBD. However, the role of TREM-1+ cell subsets in driving IBD pathology and the link with clinical parameters are not understood. We investigated TREM-1 expression in human intestinal MNP subsets and examined blocking TREM-1 as a potential IBD therapy. METHODS: TREM-1 gene expression was analysed in intestinal mucosa, enriched epithelial and lamina propria [LP] layers, and purified cells from controls and IBD patients. TREM-1 protein on immune cells was assessed by flow cytometry and immunofluorescence microscopy. Blood monocyte activation was examined by large-scale gene expression using a TREM-1 agonist or LP conditioned media [LP-CM] from patients in the presence or absence of TREM-1 and tumour necrosis factor [TNF] antagonist antibodies. RESULTS: TREM-1 gene expression increases in intestinal mucosa from IBD patients and correlates with disease score. TREM-1+ cells, which are mainly immature macrophages and CD11b+ granulocytes, increase among LP cells from Crohn's disease patients and their frequency correlates with inflammatory molecules in LP-CM. LP-CM from Crohn's disease patients induces an inflammatory transcriptome in blood monocytes, including increased IL-6 expression, which is reduced by simultaneous blocking of TREM-1 and TNF. CONCLUSIONS: High intestinal TREM-1 expression, reflecting a high frequency of TREM-1+ immature macrophages and TREM-1+CD11b+ granulocytes, is linked to the deleterious inflammatory microenvironment in IBD patients. Therefore, blocking the TREM-1 pathway, especially simultaneously with anti-TNF therapy, has potential as a new IBD therapy.

Human Intestinal Mononuclear Phagocytes in Health and Inflammatory Bowel Disease.

Inflammatory bowel disease (IBD), including Crohn's disease and ulcerative colitis, is a complex immune-mediated disease of the gastrointestinal tract that increases morbidity and negatively influences the quality of life. Intestinal mononuclear phagocytes (MNPs) have a crucial role in maintaining epithelial barrier integrity while controlling pathogen invasion by activating an appropriate immune response. However, in genetically predisposed individuals, uncontrolled immune activation to intestinal flora is thought to underlie the chronic mucosal inflammation that can ultimately result in IBD. Thus, MNPs are involved in fine-tuning mucosal immune system responsiveness and have a critical role in maintaining homeostasis or, potentially, the emergence of IBD. MNPs include monocytes, macrophages and dendritic cells, which are functionally diverse but highly complementary. Despite their crucial role in maintaining intestinal homeostasis, specific functions of human MNP subsets are poorly understood, especially during diseases such as IBD. Here we review the current understanding of MNP ontogeny, as well as the recently identified human intestinal MNP subsets, and discuss their role in health and IBD.

Immune cell-derived cytokines contribute to obesity-related inflammation, fibrogenesis and metabolic deregulation in human adipose tissue.

Adipose tissue contains a variety of immune cells, which vary in abundance and phenotype with obesity. The contribution of immune cell-derived factors to inflammatory, fibrotic and metabolic alterations in adipose tissue is not well established in human obesity. Human primary adipose tissue cells, including pre-adipocytes, endothelial cells and mature adipocytes, were used to investigate deregulation of cell- and pathway-specific gene profiles. Among factors known to alter adipose tissue biology, we focus on inflammatory (IL-1ß and IL-17) and pro-fibrotic (TGF-ß1) factors. rIL-1ß and rIL-17 induced concordant pro-inflammatory transcriptional programs in pre-adipocytes and endothelial cells, with a markedly more potent effect of IL-1ß than IL-17. None of these cytokines had significant effect on fibrogenesis-related gene expression, contrasting with rTGF-ß1-induced up-regulation of extracellular matrix components and pro-fibrotic factors. In mature adipocytes, all three factors promoted down-regulation of genes functionally involved in lipid storage and release. IL-1ß and IL-17 impacted adipocyte metabolic genes in relation with their respective pro-inflammatory capacity, while the effect of TGF-ß1 occurred in face of an anti-inflammatory signature. These data revealed that IL-1ß and IL-17 had virtually no effect on pro-fibrotic alterations but promote inflammation and metabolic dysfunction in human adipose tissue, with a prominent role for IL-1ß.

High-finesse Fabry-Perot cavities with bidimensional Si3N4 photonic-crystal slabs.

Light scattering by a two-dimensional photonic-crystal slab (PCS) can result in marked interference effects associated with Fano resonances. Such devices offer appealing alternatives to distributed Bragg reflectors and filters for various applications, such as optical wavelength and polarization filters, reflectors, semiconductor lasers, photodetectors, bio-sensors and non-linear optical components. Suspended PCS also have natural applications in the field of optomechanics, where the mechanical modes of a suspended slab interact via radiation pressure with the optical field of a high-finesse cavity. The reflectivity and transmission properties of a defect-free suspended PCS around normal incidence can be used to couple out-of-plane mechanical modes to an optical field by integrating it in a free-space cavity. Here we demonstrate the successful implementation of a PCS reflector on a high-tensile stress Si3N4 nanomembrane. We illustrate the physical process underlying the high reflectivity by measuring the photonic-crystal band diagram. Moreover, we introduce a clear theoretical description of the membrane scattering properties in the presence of optical losses. By embedding the PCS inside a high-finesse cavity, we fully characterize its optical properties. The spectrally, angular- and polarization-resolved measurements demonstrate the wide tunability of the membrane's reflectivity, from nearly 0 to 99.9470±0.0025%, and show that material absorption is not the main source of optical loss. Moreover, the cavity storage time demonstrated in this work exceeds the mechanical period of low-order mechanical drum modes. This so-called resolved-sideband condition is a prerequisite to achieve quantum control of the mechanical resonator with light.

Experimental GVD engineering in slow light slot photonic crystal waveguides.

The use in silicon photonics of the new optical materials developed in soft matter science (e.g. polymers, liquids) is delicate because their low refractive index weakens the confinement of light and prevents an efficient control of the dispersion properties through the geometry. We experimentally demonstrate that such materials can be incorporated in 700 µm long slot photonic crystal waveguides, and hence can benefit from both slow-light field enhancement effect and slot-induced ultra-small effective areas. Additionally, we show that their dispersion can be engineered from anomalous to normal regions, along with the presence of multiple zero group velocity dispersion (ZGVD) points exhibiting Normalized Delay Bandwidth Product as high as 0.156. The reported results provide experimental evidence for an accurate control of the dispersion properties of fillable periodical slotted structures in silicon photonics, which is of direct interest for on-chip all-optical data treatment using nonlinear optical effects in hybrid-on-silicon technologies.

Enhanced nonlinear interaction in a microcavity under coherent excitation.

The large field enhancement that can be achieved in high quality factor and small mode volume photonic crystal microcavities leads to strengthened nonlinear interactions. However, the frequency shift dynamics of the cavity resonance under a pulsed excitation, which is driven by nonlinear refractive index change, tends to limit the coupling efficiency between the pulse and the cavity. As a consequence, the cavity enhancement effect cannot last for the entire pulse duration, limiting the interaction between the pulse and the intra-cavity material. In order to preserve the benefit of light localization throughout the pulsed excitation, we report the first experimental demonstration of coherent excitation of a nonlinear microcavity, leading to an enhanced intra-cavity nonlinear interaction. We investigate the nonlinear behavior of a Silicon-based microcavity subject to tailored positively chirped pulses, enabling to increase the free carrier density generated by two-photon absorption by up to a factor of 2.5 compared with a Fourier-transform limited pulse excitation of equal energy. It is accompanied by an extended frequency blue-shift of the cavity resonance reaching 19 times the linear cavity bandwidth. This experimental result highlights the interest in using coherent excitation to control intra-cavity light-matter interactions and nonlinear dynamics of microcavity-based optical devices.

Liquid sensor based on high-Q slot photonic crystal cavity in silicon-on-insulator configuration.

We present the realization of an optical sensor based on an infiltrated high-Q slot photonic crystal cavity in a nonfreestanding membrane configuration. Successive infiltrations by liquids with refractive indices ranging from 1.345 to 1.545 yield a sensitivity S of 235 nm/RIU (refractive index unit), while the Q-factor is comprised between 8000 and 25,000, giving a sensor figure of merit up to 3700. This sensor has a detection limit of 1.25×10⁻5. The operation of this device on a silicon-on-insulator (SOI) substrate allows a straightforward integration in the silicon photonics platform, while providing a compliant mechanical stability.

High efficiency asymmetric directional coupler for slow light slot photonic crystal waveguides.

An asymmetric directional coupler scheme for the efficient injection of light into slow light slot photonic crystal waveguide modes is proposed and investigated using finite-difference time-domain simulation. Coupling wavelengths can be flexibly controlled by the geometrical parameters of a side-coupled subwavelength corrugated strip waveguide. This approach leads to a ~1dB insertion loss level up to moderately high light group indices (nG≈30) in wavelength ranges of 5-10nm. This work brings new opportunities to inject light into the slow modes of slot photonic crystal waveguides for on-chip communications using hybrid silicon photonics or sensing based on hollow core waveguides.

T cell-derived IL-22 amplifies IL-1ß-driven inflammation in human adipose tissue: relevance to obesity and type 2 diabetes.

Proinflammatory cytokines are critically involved in the alteration of adipose tissue biology leading to deterioration of glucose homeostasis in obesity. Here we show a pronounced proinflammatory signature of adipose tissue macrophages in type 2 diabetic obese patients, mainly driven by increased NLRP3-dependent interleukin (IL)-1ß production. IL-1ß release increased with glycemic deterioration and decreased after gastric bypass surgery. A specific enrichment of IL-17- and IL-22-producing CD4(+) T cells was found in adipose tissue of type 2 diabetic obese patients. Coculture experiments identified the effect of macrophage-derived IL-1ß to promote IL-22 and IL-17 production by human adipose tissue CD4(+) T cells. Reciprocally, adipose tissue macrophages express IL-17 and IL-22 receptors, making them sensitive to IL-17 and IL-22. IL-22 increased IL-1ß release by inducing pro-IL-1ß transcription through activation of C-Jun pathways in macrophages. In sum, these human data identified IL-1ß and the T-cell cytokine IL-22 as key players of a paracrine inflammatory pathway previously unidentified in adipose tissue, with a pathological relevance to obesity-induced type 2 diabetes. These results provide an additional rationale for targeting IL-1ß in obesity-linked type 2 diabetes and may have important implications for the conception of novel combined anti-IL-1ß and anti-IL-22 immunotherapy in human obesity.

Lung tumor microenvironment induces specific gene expression signature in intratumoral NK cells.

Natural killer (NK) cells are able to recognize and kill tumor cells, however whether they contribute to tumor immunosurveillance is still debated. Our previous studies demonstrated the presence of NK cells in human lung tumors. Their comparison with NK cells from non-tumoral lung tissues and with blood NK cells from the same individuals revealed a decreased expression of some NK receptors and impaired ex vivo cytotoxic functions occurring specifically in NK cells isolated from the tumor microenvironment. The aim of the present study was to characterize the transcriptional profile of such intratumoral NK cells, by comparative microarray analysis of sorted NK cells isolated from non-tumoral (Non-Tum-NK) and tumoral (Tum-NK) lung tissues of 12 Non-Small Cell Lung Cancer patients. Our results reveal a specific gene expression signature of Tum-NK cells particularly in activation processes and cytotoxicity, confirming that tumor environment induces modifications in NK cells biology. Indeed, intratumoral NK cells display higher expression levels of NKp44, NKG2A, Granzymes A and K, and Fas mRNA. A particular pattern of receptors involved in chemotaxis was also observed, with an overexpression of CXCR5 and CXCR6, and a lower expression of CX3CR1 and S1PR1 genes in Tum-NK as compared to Non-Tum-NK cells. The precise identification of the molecular pathways modulated in the tumor environment will help to decipher the role of NK cells in tumor immunosurveillance and will open future investigations to manipulate their antitumoral functions.

Enhanced localization of light in slow wave slot photonic crystal waveguides.

A flexible design of slot photonic crystal waveguide with a wide comb is investigated. Introduction of a carefully designed comb within the photonic crystal waveguide allows an accurate dispersion engineering in order to achieve slow light and increase the optical confinement within the comb. The strong light confinement results in an extremely small nonlinear effective area around 0.015 µm2. We report experimental realization of a comb photonic crystal waveguide with measured group indices higher than 100 in a Mach-Zehnder interferometer configuration and extract losses limited to 3.7 dB for a 100 µm device at n(g)=37.