Key hepatic signatures of human and mouse nonalcoholic steatohepatitis: A transcriptome-proteome data meta-analysis.

Background: Despite the global prevalence of nonalcoholic fatty liver disease (NAFLD), its pathophysiology remains unclear. In this study, we established highly confident nonalcoholic steatohepatitis (NASH) gene signatures and evaluated the pathological mechanisms underlying NASH through a systematic meta-analysis of transcriptome and proteome datasets obtained from NASH patients and mouse models. Methods: We analyzed NASH transcriptome datasets from 539 patients and 99 mice. A whole-liver tissue proteome dataset was used to confirm the protein level dysregulation of NASH signatures significant in both humans and mice. Results: In total, 254 human and 1,917 mouse NASH gene signatures were established. Up-regulated genes of 254 human signatures were associated with inflammation, steatosis, apoptosis, and extracellular matrix organization, whereas down-regulated genes were associated with response to metal ions and lipid and amino acid metabolism. When different mouse models were compared against humans, models with high fat and high fructose diet most closely resembled the genetic features of human NAFLD. Cross-species analysis revealed 66 genes that were concordantly dysregulated between human and mouse NASH. Among these, 14 genes were further validated to be dysregulated at the protein level. The resulting 14 genes included some of the well-established NASH associated genes and a promising NASH drug target. Functional enrichment analysis revealed that dysregulation of amino acid metabolism was the most significant hepatic perturbation in both human and mouse NASH. Conclusions: We established the most comprehensive hepatic gene signatures for NASH in humans and mice to date. To the best of our knowledge, this is the first study to collectively analyze the common signatures between human and mouse NASH on a transcriptome-proteome scale.

A Novel Competitive Binding Screening Assay Reveals Sennoside B as a Potent Natural Product Inhibitor of TNF-α.

Natural products (NPs) have played a significant role in drug discovery for diverse diseases, and numerous attempts have been made to discover promising NP inhibitors of tumor necrosis factor α (TNF-α), a major therapeutic target in autoimmune diseases. However, NP inhibitors of TNF-α, which have the potential to be developed as new drugs, have not been reported for over a decade. To facilitate the search for new promising inhibitors of TNF-α, we developed an efficient competitive binding screening assay based on analytical size exclusion chromatography coupled with liquid chromatography-tandem mass spectrometry. Application of this screening method to the NP library led to the discovery of a potent inhibitor of TNF-α, sennoside B, with an IC50 value of 0.32 µM in TNF-α induced HeLa cell toxicity assays. Surprisingly, the potency of sennoside B was 5.7-fold higher than that of the synthetic TNF-α inhibitor SPD304. Molecular docking was performed to determine the binding mode of sennoside B to TNF-α. In conclusion, we successfully developed a novel competition binding screening method to discover small molecule TNF-α inhibitors and identified the natural compound sennoside B as having exceptional potency.

Selenium bioisosteric replacement of adenosine derivatives promoting adiponectin secretion increases the binding affinity to peroxisome proliferator-activated receptor δ.

N6-(3-Iodobenzyl)adenosine-5'-N-methyluronamide (1a, IB-MECA) exhibited polypharmacological characteristics targeting A3 adenosine receptor (AR), peroxisome proliferator-activated receptor (PPAR) γ, and PPARδ, simultaneously. The bioisosteric replacement of oxygen in 4'-oxoadenosines with selenium significantly increased the PPARδ-binding activity. 2-Chloro-N6-(3-iodobenzyl)-4'-selenoadenosine-5'-N-methyluronamide (3e) and related 4'-selenoadenosine derivatives significantly enhanced adiponectin biosynthesis during adipogenesis in human bone marrow mesenchymal stem cells (hBM-MSCs). The PPARδ-binding affinity, but not the A3 AR binding affinity, of 4'-selenoadenosine derivatives correlated with their adiponectin secretion stimulation. Compared with the sugar ring of 4'-oxoadenosine, that of 4'-selenoadenosine was more favorable in forming the South sugar conformation. In the molecular docking simulation, the South sugar conformation of compound 3e formed additional hydrogen bonds inside the PPARδ ligand-binding pocket compared with the North conformation. Therefore, the sugar conformation of 4'-selenoadenosine PPAR modulators affects the ligand binding affinity against PPARδ.

CXCL14 downregulation in human keratinocytes is a potential biomarker for a novel in vitro skin sensitization test.

To elucidate the roles of epidermal keratinocytes in the toxicological outcomes of chemically induced contact dermatitis, genome-scale transcriptional analyses were performed using normal human keratinocytes (NHKCs) treated with 10 µM sodium lauryl sulfate (SLS) or 5 µM urushiol. In Gene Ontology (GO) enrichment analyses, SLS- and urushiol-induced upregulated DEGs are commonly associated with the regulation of pro-inflammatory responses and epidermal differentiation processes in NHKCs whereas cellular protein metabolic process was also identified as a commonly downregulated DEG signature. Among the downregulated DEGs, CXCL14 was investigated as a potential biomarker for a new in vitro skin sensitization test using OECD TG429 reference chemicals. CXCL14 was significantly downregulated in NHKCs in response to 62.5% of the OECD TG429 sensitizers in a concentration-dependent manner. When the sensitizer-induced upregulation of chemokine CXCL8 was included in the analysis, 87.5% of the OECD TG429 reference sensitizing chemicals significantly induced either CXCL8 upregulation or CXCL14 downregulation in NHKCs. Only one OECD TG429 reference non-sensitizer changed the constitutive CXCL14 expression in NHKCs whereas five out of six non-sensitizers altered CXCL8 production. The reference irritating non-sensitizer SLS caused a false-positive outcome. The downregulation of constitutively expressed CXCL14 was regulated by both the MAPK/ERK and JAK3/STAT6 pathways in NHKCs. CXCL14 can be used as a mechanism-based biomarker in the development of in vitro skin sensitization tests and may help improve the distinction between allergenic sensitizers and non-sensitizers.

Keratinocyte-derived IL-36γ plays a role in hydroquinone-induced chemical leukoderma through inhibition of melanogenesis in human epidermal melanocytes.

Chemical leukoderma is an acquired type of vitiligo that can be initiated by various exogenous chemicals such as hydroquinone (HQ), rhododendrol (RD), or 4-tertiary butyl phenol (4-TBP). Despite the importance of epidermal keratinocytes in diverse dermatological conditions, their toxicological role in chemical leukoderma is poorly understood. To elucidate their role in the pathogenesis of chemical leukoderma, genome-scale transcriptional analysis was performed in human epidermal keratinocytes (HEKs) treated with a sub-cytotoxic HQ concentration (10 µM). The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway-based functional enrichment analysis of HQ-induced differentially expressed genes (DEGs) revealed that HQ significantly upregulated DEGs related to the IL-17 signaling pathway and significantly downregulated DEGs associated with melanogenesis in HEKs. The meta-analysis between the HQ-induced and cytokine-induced transcriptional data (GSE53751) showed that 58 DEGs were commonly upregulated between HQ- and IL-17A-treated HEKs. Notably, the expression of IL36G was significantly increased in HEKs in response to both HQ and IL-17A. IL-36γ (2 µg/ml) directly inhibits melanin biosynthesis in cultured human epidermal melanocytes (HEMs) and downregulates the gene transcription of key enzymes in the melanogenesis pathway including TYR, DCT, and TYRP1. Moreover, IL-36γ autocrinally regulated keratinocyte function to produce the proinflammatory cytokines IL-36γ, IL-6, and CXCL8/IL-8 in a concentration-dependent manner, suggesting that IL-36γ may stimulate the amplification cycle of cutaneous inflammation. In this regard, hydroquinone-induced IL-36γ from human keratinocytes plays a pivotal role in the development of chemical leukoderma by autocrinally or paracrinally modulating the crosstalk between keratinocytes and melanocytes.

2-Phenyl-8-(1-phenylallyl)-chromenone compounds have a pan-PPAR modulator pharmacophore.

Adiponectin is an adipocytokine with insulin-sensitizing, anti-atherogenic, and anti-inflammatory properties. Adiponectin secretion-inducing compounds have therapeutic potential in a variety of metabolic diseases. Phenotypic screening led to the discovery that 5,7-dihydroxy-8-(1-(4-hydroxy-3-methoxyphenyl)allyl)-2-phenyl-4H-chromen-4-one (compound 1) had adiponectin secretion-inducing activity during adipogenesis in human bone marrow mesenchymal stem cells (hBM-MSCs). Compound 1 was originally reported to be an anti-cancer chemical isolated from natural honeybee propolis, and its adiponectin secretion-inducing activity was found in non-cytotoxic concentrations. In a target identification study, compound 1 and its potent synthetic derivative compound 5 were shown to be novel pan-peroxisome proliferator-activator receptor (PPAR) modulators. Molecular docking models with PPARs have indicated that the binding modes of chromenone compounds preferentially interacted with the hydrophobic ligand binding pocket of PPARs. In addition, chromenone compounds have been shown to result in different phenotypic outcomes in the transcriptional regulation of lipid metabolic enzymes than those of selective PPAR mono-agonists for PPARα, PPARγ, and PPARδ. In line with the pharmacology of adiponectin and PPAR pan-modulators, compounds 1 and 5 may have diverse therapeutic potentials to treat cancer and metabolic diseases.

A long-wave UVA filter avobenzone induces obesogenic phenotypes in normal human epidermal keratinocytes and mesenchymal stem cells.

Avobenzone is the most commonly used ultraviolet (UV) A filter ingredient in sunscreen. To investigate the biological activity of avobenzone in normal human epidermal keratinocytes (NHEKs), the genome-scale transcriptional profile of NHEKs was performed. In this microarray study, we found 273 up-regulated and 274 down-regulated differentially expressed genes (DEGs) in NHEKs treated with avobenzone (10 µM). Gene Ontology (GO) enrichment analysis showed that avobenzone significantly increased the DEGs associated with lipid metabolism in NHEKs. In addition, avobenzone increased the gene transcription of peroxisome proliferator-activated receptor γ (PPARγ) and fatty acid binding protein 4 in NHEKs, implicating that avobenzone may be one of the metabolic disrupting obesogens. To confirm the obesogenic potential, we examined the effect of avobenzone on adipogenesis in human bone marrow mesenchymal stem cells (hBM-MSCs). Avobenzone (EC50, 14.1 µM) significantly promoted adipogenesis in hBM-MSCs as its positive control obesogenic chemicals. Avobenzone (10 µM) significantly up-regulated mRNA levels of PPARγ during adipogenesis in hBM-MSCs. However, avobenzone did not directly bind to PPARγ and the avobenzone-induced adipogenesis-promoting activity was not affected by PPARγ antagonists T0070907 and GW9662. Therefore, avobenzone promoted adipogenesis in hBM-MSCs through a PPARγ-independent mechanism. This study suggests that avobenzone functions as a metabolic disrupting obesogen.

Adiponectin-Secretion-Promoting Phenylethylchromones from the Agarwood of Aquilaria malaccensis.

The therapeutic potential of adiponectin regulation has received interest because of its association with diverse human disease conditions, such as diabetes, obesity, atherosclerosis, and cancer. Phenylethylchromone derivatives from Aquilaria malaccensis-derived agarwood promoted adiponectin secretion during adipogenesis in human bone marrow mesenchymal stem cells, and 5,6-dihydroxy-2-(2-phenylethyl)chromone (1) was identified as a new chromone derivative. A target identification study with the most potent adiponectin-secretion-promoting phenylethylchromones, 6-methoxy-2-(2-phenylethyl)chromone (3) and 7-methoxy-2-(2-phenylethyl)chromone (4), showed that they are PPARγ partial agonists. Therefore, the diverse therapeutic effects of agarwood are associated with a PPARγ-mediated adiponectin-secretion-promoting mechanism.