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It is currently accepted that inflammation plays an important role in the pathogenesis of connective tissue disease-associated pulmonary arterial hypertension (CTD-PAH). However, the efficacy of immunosuppressive therapy remains anecdotal. The objective of this systematic review was to evaluate the efficacy of immunosuppressive therapy in patients with CTD-PAH and to further assess whether response differs between CTD subtypes and clinical features.We systematically searched studies reporting the treatment response of immunosuppressants and biological agents in CTD-PAH from PUBMED, EMBASE, the Cochrane Library, and Scopus. Studies had to report treatment regime and response criteria. The risk of bias was assessed using the Newcastle-Ottawa scale.Seven independent cohorts, 1 trial, and 1 case-series encompassing 439 patients with CTD-PAH were included. Patients were divided into 2 groups according to the therapeutic regimen. There were 146 patients in the immunosuppressants group with better heart function at baseline and 52.1% (76/146) of them were responders. There were 236 patients treated with immunosuppressants combined with PAH-specific therapy who showed more severity at baseline and 41.1% (97/236) of them were responders. Among different CTD subtypes, patients with systemic lupus erythematosus-associated PAH (SLE-PAH) showed a better response to immunosuppressants (response rate 48.1%). What is more, 1 randomized controlled trial showed the potential therapeutic value of rituximab (n = 57) in CTD-PAH patients.Current studies support the use of immunosuppressive therapy in CTD-PAH, especially in SLE-PAH. Further studies on biological agents and the therapeutic effect of different immunosuppressants are still needed.

Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease in Western countries. There is growing evidence that dysbiosis of the intestinal microbiota and disruption of microbiota-host interactions contribute to the pathology of NAFLD. We previously demonstrated that gut microbiota derived tryptophan metabolite indole-3-acetate (I3A) was decreased in both cecum and liver of high-fat diet-fed mice and attenuated the expression of inflammatory cytokines in macrophages and TNF-a and fatty acid induced inflammatory responses in an aryl-hydrocarbon receptor (AhR) dependent manner in hepatocytes. In this study, we investigated the effect of orally administered I3A in a mouse model of diet induced NAFLD. Western diet (WD)-fed mice given sugar water (SW) with I3A showed dramatically decreased serum ALT, hepatic TG, liver steatosis, hepatocyte ballooning, lobular inflammation, and hepatic production of inflammatory cytokines, compared to WD-fed mice given only SW. Metagenomic analysis show that I3A administration did not significantly modify the intestinal microbiome, suggesting that I3A’s beneficial effects likely reflect the metabolite’s direct actions on the liver. Administration of I3A partially reversed WD induced alterations of liver metabolome and proteome, notably, decreasing expression of several enzymes in hepatic lipogenesis and β-oxidation. Mechanistically, we also show that AMP-activated protein kinase (AMPK) mediates the anti-inflammatory effects of I3A in macrophages. The potency of I3A in alleviating liver steatosis and inflammation clearly demonstrates its potential as a therapeutic modality for preventing the progression of steatosis to NASH.

1,3,5-Tri-O-caffeoyl quinic acid is a caffeoylquinic acid derivative isolated from the roots of Arctium lappa L. Our previous studies have revealed that the ethyl acetate extract of the roots of A. lappa L. and the caffeoylquinic acids contained in it possess antioxidant properties, especially 1,3,5-tri-O-caffeoyl quinic acid. The present study aimed to investigate the protective effects of 1,3,5-tri-O-caffeoyl quinic acid against hydrogen peroxide-induced oxidative stress and explore the underlying mechanism. We found that 1,3,5-tri-O-caffeoyl quinic acid prevented the decline of cell viability and excessive release of lactate dehydrogenase induced by hydrogen peroxide. In addition, Hoechst 33 342 staining and Annexin V-PI double staining showed that 1,3,5-tri-O-caffeoyl quinic acid inhibited hydrogen peroxide-induced neuronal cell apoptosis. 1,3,5-Tri-O-caffeoyl quinic acid reduced the excessive production of intracellular reactive oxygen species, decreased the malondialdehyde content, and improved the activity of superoxide dismutase. Furthermore, 1,3,5-tri-O-caffeoyl quinic acid restored the loss of mitochondrial membrane potential in SH-SY5Y cells induced by hydrogen peroxide. 1,3,5-Tri-O-caffeoyl quinic acid downregulated the overexpression of proapoptotic proteins, including Bax, cytochrome c, cleaved caspase-9, and cleaved caspase-3 as well as promoted the expression of the antiapoptotic protein Bcl-2. Moreover, the phosphorylation of mitogen-activated protein kinases induced by hydrogen peroxide was inhibited by 1,3,5-tri-O-caffeoyl quinic acid. Pretreatment with 1,3,5-tri-O-caffeoyl quinic acid also promoted the activation of phosphorylated Akt. Taken together, these findings suggest that 1,3,5-tri-O-caffeoyl quinic acid exerts protective effects against hydrogen peroxide-induced neuronal apoptosis. In addition, inhibition of the mitogen-activated protein kinase signaling pathway and the activation of Akt are implicated in the antioxidant activity of 1,3,5-tri-O-caffeoyl quinic acid, giving new insight in searching for a compound with antioxidant activity for the treatment of oxidative stress-associated neurological diseases.

CRISPR interference (CRISPRi) enables programmable, reversible, and titratable repression of gene expression (knockdown) in mammalian cells. Initial CRISPRi-mediated genetic screens have showcased the potential to address basic questions in cell biology, genetics, and biotechnology, but wider deployment of CRISPRi screening has been constrained by the large size of single guide RNA (sgRNA) libraries and challenges in generating cell models with consistent CRISPRi-mediated knockdown. Here, we present next-generation CRISPRi sgRNA libraries and effector expression constructs that enable strong and consistent knockdown across mammalian cell models. First, we combine empirical sgRNA selection with a dual-sgRNA library design to generate an ultra-compact (1-3 elements per gene), highly active CRISPRi sgRNA library. Next, we rigorously compare CRISPRi effectors to show that the recently published Zim3-dCas9 provides the best balance between strong on-target knockdown and minimal nonspecific effects on cell growth or the transcriptome. Finally, we engineer a suite of cell lines with stable expression of Zim3-dCas9 and robust on-target knockdown. Our results and publicly available reagents establish best practices for CRISPRi genetic screening.

We numerically demonstrate highly efficient mid-infrared quasi-parametric chirped-pulse amplification (QPCPA) based on a recently developed Sm3+-doped La3Ga5.5Nb0.5O14 (Sm:LGN) crystal. At pump wavelength around 1 µm, the broadband absorption of Sm3+ on idler pulses can enable QPCPA for femtosecond signal pulses centered at 3.5 or 5 µm, with a conversion efficiency approaching the quantum limit. Due to suppression of back conversion, such mid-infrared QPCPA exhibits robustness against phase-mismatch and pump-intensity variation. The Sm:LGN-based QPCPA will provide an efficient approach for converting currently well-developed intense laser pulses at 1 µm to mid-infrared ultrashort pulses.

Rubidium (Rb) is an important microelement for the human body, which can kill and inhibit bacteria. Rubidium-containing polyhydrated ionogens have been applied to treat refractory wounds. In this work, for the first time, Rb was added into calcium alginate hydrogel (Rb-CA gel) and then made into a dressing by freeze-drying. The addition of Rb to CA maintained a well-ordered porous structure with only slightly reduced swelling and storage modulus and increased roughness and in vitro degradation rate. Experiments in vitro demonstrated that Rb-CA gels could not only inhibit the growth of Staphylococcus aureus and Pseudomonas aeruginosa ubiquitous but were nontoxic and even conducive to human umbilical vein endothelial cells (HUVECs), fibroblasts, and keratinocytes. HUVECs exposed to Rb-CA gels exhibited enhanced migration and tubule formation ability, increased vascular endothelial growth factor secretion, and improved activation of the nuclear factor (erythroid-derived 2)-like 2 (NRF2)/heme-oxygenase-1 (HO-1) signaling pathway. Rb-CA gels also promoted the migration of fibroblasts and keratinocytes. In vivo, in a type II diabetic male Sprague-Dawley rat wound model, neovascularization, re-epithelialization, and collagen deposition were all improved greatly by the Rb-CA gel, with the upregulation of NRF2 and HO-1 protein secretion that inhibited the oxidative stress reaction in the wound sites. Moreover, the Rb-CA gel exhibited a strong anti-inflammatory effect on the wound. In summary, the application of Rb-CA gel dressings to diabetic wounds could produce various synergetic enhancements in terms of angiogenesis, re-epithelialization, and collagen deposition. Hence, Rb-CA gels can be used as a novel therapeutic strategy to promote the healing of diabetic wounds.

Objective This study is intended to figure out the function of microRNA-218 (miR-218) together with microphthalmia-associated transcription factor (MITF) on the cardiac fibrosis and cardiac function impairment in rat models of myocardial infarction (MI). Results The rats with MI exhibited cardiac function impairment, cardiac fibrosis, oxidative stress, cardiomyocyte apoptosis, as well as inflammatory injury. Additionally, upregulated miR-218 and downregulated MITF were detected in cardiac tissues of MI rats. MI rats injected with miR-218 inhibitors or overexpressed MITF exhibited elevated MITF expression, improved cardiac function, and diminished pathological damages, infarct size, cardiomyocyte apoptosis, cardiac fibrosis, oxidative stress as well as inflammatory injury in cardiac tissues. Furthermore, downregulated miR-218 and MITF aggravated the conditions than downregulation of miR-218 alone in MI rats. Methods MI models were performed in rats, and then the rats were injected with miR-218 inhibitors and/or MITF overexpression plasmid to elucidate the role of miR-218 and/or MITF on the cardiac function, pathological damage, cardiac fibrosis, angiogenesis, oxidative stress and inflammatory injury of cardiac tissues in MI rats by performing a series of assays. Conclusion Collectively, we found that the suppression of miR-218 alleviates cardiac fibrosis and cardiac function impairment, and stimulates angiogenesis in MI rats through inhibiting MITF.

There is increasing evidence that the intestinal microbiota plays a mechanistic role in the etiology of non-alcoholic fatty liver disease (NAFLD). Animal and human studies have linked small molecule metabolites produced by commensal bacteria in the gut contribute to not only intestinal inflammation, but also to hepatic inflammation. These immunomodulatory metabolites are capable of engaging host cellular receptors, and may mediate the observed association between gut dysbiosis and NAFLD. This review focuses on the effects and potential mechanisms of three specific classes of metabolites that synthesized or modified by gut bacteria: short chain fatty acids, amino acid catabolites, and bile acids. In particular, we discuss their role as ligands for cell surface and nuclear receptors regulating metabolic and inflammatory pathways in the intestine and liver. Studies reveal that the metabolites can both agonize and antagonize their cognate receptors to reduce or exacerbate liver steatosis and inflammation, and that the effects are metabolite- and context-specific. Further studies are warranted to more comprehensively understand bacterial metabolite-mediated gut-liver in NAFLD. This understanding could help identify novel therapeutics and therapeutic targets to intervene in the disease through the gut microbiota.

A magnetic covalent organic framework nanocomposite (Fe3O4@COF(Tp-NDA)) was synthesized via a solvothermal method, used as a magnetic adsorbent for the extraction of polycyclic aromatic hydrocarbons (PAHs) from lake water, tea, coffee, and fried chicken, and detected using a high performance liquid chromatography-ultraviolet detector. The synthesized magnetic adsorbent was characterized via transmission electron microscopy, X-ray diffraction, Fourier transform infrared spectroscopy, N2 adsorption-desorption isotherm analysis and vibrating sample magnetometry. Parameters that affected the extraction conditions and desorption conditions were optimized. Adsorption equilibrium could be attained within 3 min. The prepared magnetic material could be reused 10 times. The limits of detection and quantification were 0.05-0.25 μg L-1 and 0.17-0.83 μg L-1, respectively. The recovery was 74.6-101.8% with a relative standard deviation of below 4.2%. The method was successfully used to detect PAHs in various samples.

The human gastrointestinal (GI) tract is colonized by a highly diverse and complex microbial community (i.e., microbiota). The microbiota plays an important role in the development of the immune system, specifically mediating inflammatory responses, however the exact mechanisms are poorly understood. We have developed a mathematical model describing the effect of indole on host inflammatory signaling in HCT-8 human intestinal epithelial cells. In this model, indole modulates transcription factor nuclear factor κ B (NF-κB) and produces the chemokine interleukin-8 (IL-8) through the activation of the aryl hydrocarbon receptor (AhR). Phosphorylated NF-κB exhibits dose and time-dependent responses to indole concentrations and IL-8 production shows a significant down-regulation for 0.1 ng/mL TNF-α stimulation. The model shows agreeable simulation results with the experimental data for IL-8 secretion and normalized NF-κB values. Our results suggest that microbial metabolites such as indole can modulate inflammatory signaling in HTC-8 cells through receptor-mediated processes.

The long noncoding RNA H19 was reported to associate with melanogenesis. However, it remains unknown whether H19 expression will be changed by UVB irradiation and whether H19 will regulate melanocytes melanogenesis by paracrine effects. Here, we analysed the expression changes of H19 irradiated by UVB in keratinocytes and explored the mechanism of melanogenesis stimulated by H19 through paracrine effects. First, after keratinocytes were exposed to UVB irradiation, expression of H19 and pro-opiomelanocortin (POMC) was measured by qRT-PCR. Also, α-melanocyte-stimulating hormone (α-MSH) contents in cells supernatant were measured by ELISA. Then, H19 siRNAs were designed and transfected into keratinocytes by liposome. The expression changes of H19, POMC and α-MSH were detected. Besides, expression of p53 was detected by Western blot. After that, supernatant of keratinocytes with H19 siRNAs or negative control siRNA was cocultured with immortalized melanocyte line PIG1. Expression levels of MiTF, TYR, Rab27A, TYRP2, FSCN1 and MYO5A in PIG1 cells were detected by Western blot and qRT-PCR. We found that H19 expression of keratinocytes cells decreased after UVB irradiation. However, the levels of POMC, α-MSH and p53 were upregulated in UVB-irradiated cells. Compared with the negative control, H19 siRNAs could significantly increase the expression of POMC, α-MSH and p53. After supernatant of keratinocytes transfected with H19 siRNAs was cocultured with PIG1 cells, the levels of MiTF, TYR and Rab27A were upregulated in PIG1 cells. In conclusion, UVB-inhibited H19 may promote α-MSH secretion by p53 in keratinocytes and then regulate melanocytes melanogenesis through paracrine effects.

Arthritis is an autoimmune disease that adversely affects the quality of life of patients. Treatment options for autoimmunity include the use of chemical and biologic agents that suppress inflammation. However, these can result in patients being vulnerable to opportunistic infections. Adoptive T regulatory (Treg) cell transfer can specifically block auto-inflammatory immune cells in an antigen-specific manner while maintaining an intact immune response to pathogens. However, isolating sufficient Treg cells for adoptive cell transfer therapy is challenging; therefore, improving Treg cell survival and function could have important impacts on adoptive Treg transfer therapy. To explore the use of bacterial vaccines as autoimmune therapies, we engineered an attenuated brucellosis vaccine (Brucella melitensis ΔvjbR Tna) to produce indole, a bacterial metabolite that induces Treg expansion. In the collagen induced arthritis (CIA) murine model, Treg transfer following BmΔvjbR Tna treatment decreased arthritis incidence and score. CD4+FoxP3+ T regulatory cells and CD8+ T cells in spleen and lymph nodes were increased in BmΔvjbR Tna treated CIA mice. BmΔvjbR Tna injection also promoted CD4+ cell proliferation, and increased IL-2 and IFN-g expression. The levels of some pro-inflammatory cytokines (IL-1β and IL-6) were also dramatically reduced in BmΔvjbR Tna infected bone marrow derived macrophages (BMDMs). Taken together, these findings indicate that the indole-producing BmΔvjbR Tna strain increases Treg cell populations and proliferation, suppresses proinflammatory cytokine secretion, and reduces autoimmune responses.

The occurrence of hyperpigmentation or hypopigmentation after inflammation is a common condition in dermatology and cosmetology. Since the exact mechanism of its occurrence is not yet known, prevention and treatment are troublesome. Previous studies have confirmed that α‑melanocyte‑stimulating hormone, stem cell factor and other factors can promote melanogenesis‑related gene expression through the activation of signaling pathways. Recent studies have revealed that a variety of inflammatory mediators can also participate in the regulation of melanogenesis in melanocytes. In this review, we summarized that interleukin‑18, interleukin‑33, granulocyte‑macrophage colony stimulating factor, interferon‑γ, prostaglandin E2 have the effect of promoting melanogenesis, while interleukin‑1, interleukin‑4, interleukin‑6, interleukin‑17 and tumor necrosis factor can inhibit melanogenesis. Further studies have found that these inflammatory factors may activate or inhibit melanogenesis‑related signaling pathways (such as protein kinase A and mitogen activated protein kinase) by binding to corresponding receptors, thereby promoting or inhibiting the expression of melanogenesis‑related genes and regulating skin pigmentation processes. This suggests that the development of drugs or treatment methods from the perspective of regulating inflammation can provide new ideas and new targets for the treatment of pigmented dermatosis. This review outlines the current understanding of the inflammation factors' roles in melanogenesis.

Our previous study found that Ganoderma lucidum polysaccharide (GLP), bioactive ingredients from Ganoderma lucidum, protected fibroblasts from photoaging. However, whether GLP can affect melanogenesis in melanocytes through regulating paracrine mediators that secreted by keratinocytes and fibroblasts is unclear. We aimed to investigate the efficacy and mechanisms of action of GLP in melanogenesis by regulating paracrine effects of keratinocytes and fibroblasts. The effect of GLP on cell viability affected by GLP was measured by the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. After an immortal keratinocyte line (HaCaT) and primary fibroblasts (FB) were treated with GLP, the supernatants of HaCaT and FB cells were collected and cocultured with an immortalized melanocyte line (PIG1). The expression levels of melanogenesis-associated genes in PIG1 cells were measured by quantitative real-time polymerase chain reaction (qRT-PCR) and western blot analysis. Furthermore, FRS-2, ERK, JNK, and p38 phosphorylation levels were measured. Then, major melanogenic paracrine mediators in HaCaT and FB cells treated with GLP were evaluated by qRT-PCR and enzyme-linked immunosorbent assay (ELISA). In addition, the expression of IL-6 and STAT3 was examined in HaCaT and FB cells. GLP was not cytotoxic to HaCaT and FB cells. The supernatants of GLP-treated HaCaT and FB cells downregulated the expression levels of MITF, TYR, TYRP1, TYRP2, RAB27A, and FSCN1 genes and inhibited the phosphorylation of FRS-2, ERK, JNK, and p38 in PIG1 cells. GLP also decreased FGF2 secretion in HaCaT and FB cells. Moreover, GLP reduced IL-6 expression and STAT3 phosphorylation in HaCaT and FB cells. GLP reduced melanogenesis in melanocytes by inhibiting the paracrine effects of keratinocytes and fibroblasts via IL-6/STAT3/FGF2 pathway.

It is well established that the dynamics of signaling and gene expression in a clonal population of cells vary both in the absence and presence of stimuli. In order to elucidate the stochasticity in signaling and downstream cellular responses, we have formulated a mathematical model and utilized it along with single-cell experimental measurements to investigate the cell-to-cell variability in the response of mouse macrophages to lipopolysaccharide (LPS). Since the single-cell mathematical model is computationally expensive to simulate, a semi-stochastic model that combines a deterministic model with parameters that have distributions at the single-cell level is more viable than a stochastic model. This modeling approach requires an accurate deterministic model a priori. Motivated by above considerations, this work aimed to quantitatively calibrate the deterministic model that can simulate the average single-cell dynamics via a systematic approach, which combines single-cell experimental measurements and a parameter selection method, with the LPS-induced $\text{NF}\kappa\mathrm{B}$ signal pathway as an example case.

Ultraviolet (UV)-induced pigmentation is very common in clinical practice, but the current treatments are rarely effective, accompanied by some side effects. Ganoderma lucidum polysaccharide (GLP) is a natural antioxidant with no toxic side effects, which can antagonize UVB-induced fibroblast photo aging. The study aims to explore the role of GLP in inhibiting UVB-induced melanogenesis and its possible mechanism. The expression of melanogenesis genes such as microphthalmia-associated transcription factor (MITF), tyrosine (TYR), tyrosinase related protein 1 (TYRP1), tyrosinase related protein 2 (TYRP2), ras-related protein Rab-27A (Rab27A), and Myosin shows an upward trend after exposure of B16F10 and PIG1 cells to UVB irradiation, but GLP can downregulate the expression of genes related to UVB-induced melanogenesis. GLP can inhibit UVB-activated protein kinase A (PKA) and mitogen-activated protein kinase (MAPK) signaling pathways. Besides, GLP protects mitochondria from UVB damage and inhibits reactive oxygen species (ROS) production. Also, UVB-induced cyclic adenosine monophosphate (cAMP) can be inhibited. It has been found in the experiments of UVB-induced skin pigmentation in zebrafish that GLP is capable of inhibiting UVB-induced skin pigmentation. Meanwhile, it can greatly relieve erythema reaction in guinea pig skin caused by high-dosage UVB irradiation. In conclusion, this study shows that GLP can inhibit UVB-induced melanogenesis by antagonizing cAMP/PKA and ROS/MAPK signaling pathways and is a potential natural safe whitening sunscreen additive.

Due to the intrinsic stochasticity, the signaling dynamics in a clonal population of cells exhibit cell-to-cell variability at the single-cell level, which is distinct from the population-average dynamics. Frequently, flow cytometry is widely used to acquire the single-cell level measurements by blocking cytokine secretion with reagents such as Golgiplug™. However, Golgiplug™ can alter the signaling dynamics, causing measurements to be misleading. Hence, we developed a mathematical model to infer the average single-cell dynamics based on the flow cytometry measurements in the presence of Golgiplug™ with lipopolysaccharide (LPS)-induced NF κ B signaling as an example. First, a mathematical model was developed based on the prior knowledge. Then, average single-cell dynamics of two key molecules (TNF α and I κ B α ) in the NF κ B signaling pathway were measured through flow cytometry in the presence of Golgiplug™ to validate the model and maximize its prediction accuracy. Specifically, a parameter selection and estimation scheme selected key model parameters and estimated their values. Unsatisfactory results from the parameter estimation guided subsequent experiments and appropriate model improvements, and the refined model was calibrated again through the parameter estimation. The inferred model was able to make predictions that were consistent with the experimental measurements, which will be used to construct a semi-stochastic model in the future.

Ethnopharmacological relevance Chloasma, senile plaques, vitiligo and other pigmentary disorders seriously affect patients’ appearance and life quality. Medicinal plant is the product of long-term medical practice worldwide, with the advantages of outstanding curative properties and less side effects. Recently, research were made to explore the value of medicinal plants in the treatment of pigmentary disorders, and remarkable results were achieved. Aim of the review This review outlines the current understanding of the role and potential mechanisms of medicinal plants (including active ingredients, extracts and prescriptions) in pigmentary disorders, especially Chinese medicinal plants, provides the preclinical evidence for the clinical benefits. This study hopes to provide comprehensive information and reliable basis for exploring new therapeutic strategies of plant drugs in the treatment of skin pigmented diseases. Methods The literature information was obtained from the scientific databases (up to Oct, 2017), mainly from the PubMed, Web of Science and CNKI databases, and was to identify the experimental studies on the regulating melanogenesis role of the active agents from herbal medicine and the involved mechanisms. The search keywords for such work included: “pigmentary” or “pigmentation”, “melanogenesis”, and “traditional Chinese medicine” or “Chinese herbal medicine”, “herb”, “medicinal plant”. Results We summarized the function of medicinal plants involved in melanogenesis, especially Chinese medicine. It was reported that the active ingredients, extracts, or prescriptions of medicinal plants can regulate the expression of genes related to melanogenesis by affecting the signaling pathways such as MAPK and PKA, thereby regulating pigment synthesis. Some of them can promote melanogenesis (such as isoliquiritigenin, geniposide; Cornus officinalis Siebold & Zucc., Eclipta prostrata (L.) L.; the Bairesi complex prescription, etc.). While others have the opposite effect (such as biochanin A, Gomisin N; Panax ginseng C.A. Meyer, Nardostachys chinensis Bat.; Sanbaitang, etc.). Conclusion Asian medicinal plants, especially their active ingredients, have multilevel effects on melanogenesis by regulating melanogenesis-related genes or signaling pathways. They are of great clinical value for the treatment of skin pigmentary disorders. However, the experimental effect, safety, and functional mechanism of the medicinal plants require further determination before studying their clinical efficacy.

SUMMARY The gut microbiota plays a significant role in the progression of fatty liver disease; however, the mediators and their mechanisms remain to be elucidated. Comparing metabolite profile differences between germ-free and conventionally raised mice against differences between mice fed a low- and high-fat diet (HFD), we identified tryptamine and indole-3-acetate (I3A) as metabolites that depend on the microbiota and are depleted under a HFD. Both metabolites reduced fatty-acid- and LPS-stimulated production of pro-inflammatory cytokines in macrophages and inhibited the migration of cells toward a chemokine, with I3A exhibiting greater potency. In hepatocytes, I3A attenuated inflammatory responses under lipid loading and reduced the expression of fatty acid synthase and sterol regulatory element-binding protein-1c. These effects were abrogated in the presence of an aryl-hydrocarbon receptor (AhR) antagonist, indicating that the effects are AhR dependent. Our results suggest that gut microbiota could influence inflammatory responses in the liver through metabolites engaging host receptors., In Brief Dysbiosis of the intestinal microbiota is an emerging factor contributing to the progression of fatty liver disease. Krishnan et al. utilize metabolomics and biochemical assays in conjunction with animal and cell culture models to identify microbiota-dependent metabolites that engage a host receptor to affect liver inflammatory responses under lipid loading.

A significant hallmark of the chronic inflammatory disease, atherosclerosis, is the accumulation of pro-inflammatory M1-like macrophages (macs) and dendritic cells (DCs) in coronary arteries that respond to pro-atherogenic stimuli and fail to digest lipids that contribute to the formation of foam cells in atherosclerotic plaques. Mechanisms that reduce Mac/DC inflammation, increase lipid degradation, and prevent foam cell formation are likely to decrease progression of atherosclerosis. Mechanisms that regulate autophagy in macs and DCs will likely reveal targets for the prevention of atherosclerosis. Mouse models of high-fat diets demonstrate a link between microbiota composition and the development of obesity, diabetes, and atherosclerosis through several mechanisms best demonstrated by the microbiota-dependent production of TMAO that promotes atherosclerosis. We have identified a tryptophan-derived microbiota metabolite, indole, that has novel immunomodulatory properties. This strong association between obesity, inflammation, and the microbiota has led us to the hypothesis that microbiota-derived metabolites in the GI tract might regulate dietary saturated fat-induced inflammation. To test this hypothesis, we generated bone marrow-derived macrophages (BMDMs) and dendritic cells (BMDCs) and tested the effects of indole on these cells in the context of saturated fatty acid-induced inflammation. We investigated pro-inflammatory cytokine production, M1 macrophage development, and metabolic flux. Here, we demonstrate that indole reduces saturated fatty acid-derived inflammation in BMDCs and BMDMs and regulates metabolic programming of phagocytes by inducing autophagy on a level similar to rapamycin.

It is established that the gut microbiota plays an important role in Non-Alcoholic Fatty Liver Disease (NAFLD) and different bacterial taxa have been correlated to disease. However, the specific metabolites through which the microbiota exert their effects and the target pathways in host cells are not fully understood. We used liquid chromatography-mass spectrometry metabolomics to identify two tryptophan-derived metabolites – tryptamine (TA) and indole-3-acetate (I3A) – that require the microbiota and are depleted in high-fat diet mice compared to low fat diet mice. Both I3A and TA reduced palmitate and LPS induced production of pro-inflammatory cytokines (TNFα, IL-1β and MCP-1) in RAW 264.7 macrophages, as well as inhibited macrophage migration toward the chemokine MCP-1. We identified that both palmitate and LPS reduced levels of p-AMPK and the addition of I3A and TA reversed p-AMPK reduction. Since the p-AMPK activator AICAR reduced palmitate and LPS induced inflammatory cytokine production, we propose that I3A and TA modulate inflammation through modulating p-AMPK levels.

Non-alcoholic fatty liver disease (NAFLD) is the most prevalent chronic liver disease in western countries. While it is established that the gut microbiota plays a role in NAFLD, the identities of microbiota metabolites involved and their target pathways in host cells is poorly understood. Based on prior work showing that the microbiota metabolite indole attenuates indicators of inflammation in intestinal epithelial cells, we hypothesized that microbiota metabolites modulate macrophage inflammation in the liver. Mass spectrometry analysis of microbiota metabolites in serum and liver of mice identified tryptamine (TA) and indole-3-acetate (I3A) as depleted in mice on a high-fat diet compared to low-fat diet. Therefore, we investigated the effect of I3A and TA on macrophage inflammation and migration. RAW 264.7 murine macrophages were treated with different concentrations of I3A and TA for 4 hours, stimulated with palmitate for 18 hours followed by LPS for 6 hours in the presence of metabolites. Our results show that treatment with I3A significantly attenuated the induction of the pro-inflammatory cytokines TNFa, IL-1b and MCP-1 by palmitate and LPS in a dose dependent manner at both mRNA and protein levels. Unlike I3A, TA only decreased the gene expression of IL-1b but significantly decreased TNFa and MCP-1 protein levels. Both I3A and TA significantly decreased bone marrow derived macrophage migration towards MCP-1 in a dose dependent manner. Together, our results suggest that microbiota metabolites can modulate NAFLD by directly acting on macrophages. Ongoing work focuses on investigating the mechanisms underlying the anti-inflammatory activity of I3A and TA, which could provide new strategies to treat NAFLD.