Human Keratinocyte Responses to Woodsmoke Chemicals.

Air pollution consists of complex mixtures of chemicals with serious deleterious health effects from acute and chronic exposure. To help understand the mechanisms by which adverse effects occur, the present work examines the responses of cultured human epidermal keratinocytes to specific chemicals commonly found in woodsmoke. Our earlier findings with liquid smoke flavoring (aqueous extract of charred wood) revealed that such extracts stimulated the expression of genes associated with oxidative stress and proinflammatory response, activated the aryl hydrocarbon receptor, thereby inducing cytochrome P4501A1 activity, and induced cross-linked envelope formation, a lethal event ordinarily occurring during terminal differentiation. The present results showed that furfural produced transcriptional responses resembling those of liquid smoke, cyclohexanedione activated the aryl hydrocarbon receptor, and several chemicals induced envelope formation. Of these, syringol permeabilized the cells to the egress of lactate dehydrogenase at a concentration close to that yielding envelope formation, while furfural induced envelope formation without permeabilization detectable in this way. Furfural (but not syringol) stimulated the incorporation of amines into cell proteins in extracts in the absence of transglutaminase activity. Nevertheless, both chemicals substantially increased the amount of cellular protein incorporated into envelopes and greatly altered the envelope protein profile. Moreover, the proportion of keratin in the envelopes was dramatically increased. These findings are consistent with the chemically induced protein cross-linking in the cells. Elucidating mechanisms by which this phenomenon occurs may help understand how smoke chemicals interact with proteins to elicit cellular responses, interpret bioassays of complex pollutant mixtures, and suggest additional sensitive ways to monitor exposures.

Integrative transcriptomic and metabolomic analyses reveal the phenylpropanoid and flavonoid biosynthesis of Prunus mume.

Prunus mume is an important medicinal plant with ornamental and edible value. Its flowers contain phenylpropanoids, flavonoids and other active components, that have important medicinal and edible value, yet their molecular regulatory mechanisms in P. mume remain unclear. In this study, the content of total flavonoid and total phenylpropanoid of P. mume at different developmental periods was measured first, and the results showed that the content of total flavonoid and total phenylpropanoid gradually decreased in three developmental periods. Then, an integrated analysis of transcriptome and metabolome was conducted on three developmental periods of P. mume to investigate the law of synthetic accumulation for P. mume metabolites, and the key enzyme genes for the biosynthesis of phenylpropanoids and flavonoids were screened out according to the differentially expressed genes (DEGs). A total of 14,332 DEGs and 38 differentially accumulate metabolites (DAMs) were obtained by transcriptomics and metabolomics analysis. The key enzyme genes and metabolites in the bud (HL) were significantly different from those in the half-opening (BK) and full-opening (QK) periods. In the phenylpropanoid and flavonoid biosynthesis pathway, the ion abundance of chlorogenic acid, naringenin, kaempferol, isoquercitrin, rutin and other metabolites decreased with the development of flowers, while the ion abundance of cinnamic acid increased. Key enzyme genes such as HCT, CCR, COMT, CHS, F3H, and FLS positively regulate the downstream metabolites, while PAL, C4H, and 4CL negatively regulate the downstream metabolites. Moreover, the key genes FLS (CL4312-2, CL4312-3, CL4312-4, CL4312-5, CL4312-6) regulating the synthesis of flavonols are highly expressed in bud samples. The dynamic changes of these metabolites were validated by determining the content of 14 phenylpropanoids and flavonoids in P. mume at different developmental periods, and the transcription expression levels of these genes were validated by real-time PCR. Our study provides new insights into the molecular mechanism of phenylpropanoid and flavonoid accumulation in P. mume.

Natural compounds ursolic acid and digoxin exhibit inhibitory activities to cancer cells in RORγ-dependent and -independent manner.

Natural compounds ursolic acid (UA) and digoxin isolated from fruits and other plants display potent anti-cancer effects in preclinical studies. UA and digoxin have been at clinical trials for treatment of different cancers including prostate cancer, pancreatic cancer and breast cancer. However, they displayed limited benefit to patients. Currently, a poor understanding of their direct targets and mechanisms of action (MOA) severely hinders their further development. We previously identified nuclear receptor RORγ as a novel therapeutic target for castration-resistant prostate cancer (CRPC) and triple-negative breast cancer (TNBC) and demonstrated that tumor cell RORγ directly activates gene programs such as androgen receptor (AR) signaling and cholesterol metabolism. Previous studies also demonstrated that UA and digoxin are potential RORγt antagonists in modulating the functions of immune cells such as Th17 cells. Here we showed that UA displays a strong activity in inhibition of RORγ-dependent transactivation function in cancer cells, while digoxin exhibits no effect at clinically relevant concentrations. In prostate cancer cells, UA downregulates RORγ-stimulated AR expression and AR signaling, whereas digoxin upregulates AR signaling pathway. In TNBC cells, UA but not digoxin alters RORγ-controlled gene programs of cell proliferation, apoptosis and cholesterol biosynthesis. Together, our study reveals for the first-time that UA, but not digoxin, acts as a natural antagonist of RORγ in the cancer cells. Our finding that RORγ is a direct target of UA in cancer cells will help select patients with tumors that likely respond to UA treatment.

Exploring the mechanism of the Fructus Mume and Rhizoma Coptidis herb pair intervention in Ulcerative Colitis from the perspective of inflammation and immunity based on systemic pharmacology.

PURPOSE: Ulcerative Colitis (UC) is a chronic nonspecific inflammatory disease of the colon and rectum. Fructus Mume (FM) and Rhizoma Coptidis (RC) exert effects on inflammatory and immune diseases. We evaluated the hypothesis of the FM and RC (FM-RC) herb pair remedy in alleviating dextran sulfate sodium (DSS)-induced colitis, through network pharmacology-based analyses, molecular docking, and experimental validation. METHODS: The Traditional Chinese medicine systematic pharmacology analysis platform(TCMSP) and Swiss database were used to predict potential targets of FM-RC and the GeneCards database was utilized to collect UC genes. Cytoscape software was used to construct and analyze the networks, and DAVID was utilized to perform enrichment analysis. AutoDock software was used to dock the core chemical components of the FM-RC herb pair with key UC targets. Animal experiments were performed to validate the prediction results and general conditions and body weight were observed. Pathological changes in colon tissue were observed by staining with hematoxylin and eosin. The levels of TNF-α, IL-8, IL-17, and IL-4 in serum and colon tissue were detected by ELISA. RESULTS: Eighteen effective components of the herb couple were screened, and their potential therapeutic targets in the treatment of UC were acquired from 110 overlapped targets. GO and KEGG analyses revealed that these targets were highly correlated with protein autophosphorylation, plasma membrane, ATP binding, cancer pathways, the PI3K-AKt signaling pathway, and the Rap1 signaling pathway. Molecular docking established the core protein interactions with compounds having a docking energy < 0 kJ·mol-1, indicating the core active components had strong binding activities with the core targets. FM-RC herb pair relieved pathological indicators and reduced the concentration of TNF-α, IL-8, and IL-17 and increased IL-4 levels in the serum and colon tissues of UC rats. CONCLUSION: Collectively, FM-RC herb pair administration alleviated UC. These beneficial effects targeted MAPK1 signaling related to inflammation and immunity, which provided a basis for a better understanding of FM-RC in the treatment of UC.

Deregulation of Cholesterol Homeostasis by a Nuclear Hormone Receptor Crosstalk in Advanced Prostate Cancer.

Metastatic castration-resistant prostate cancer (mCRPC) features high intratumoral cholesterol levels, due to aberrant regulation of cholesterol homeostasis. However, the underlying mechanisms are still poorly understood. The retinoid acid receptor-related orphan receptor gamma (RORγ), an attractive therapeutic target for cancer and autoimmune diseases, is strongly implicated in prostate cancer progression. We demonstrate in this study that in mCRPC cells and tumors, RORγ plays a crucial role in deregulation of cholesterol homeostasis. First, we found that RORγ activates the expression of key cholesterol biosynthesis proteins, including HMGCS1, HMGCR, and SQLE. Interestingly, we also found that RORγ inhibition induces cholesterol efflux gene program including ABCA1, ABCG1 and ApoA1. Our further studies revealed that liver X receptors (LXRα and LXRß), the master regulators of cholesterol efflux pathway, mediate the function of RORγ in repression of cholesterol efflux. Finally, we demonstrated that RORγ antagonist in combination with statins has synergistic effect in killing mCRPC cells through blocking statin-induced feedback induction of cholesterol biosynthesis program and that the combination treatment also elicits stronger anti-tumor effects than either alone. Altogether, our work revealed that in mCRPC, RORγ contributes to aberrant cholesterol homeostasis by induction of cholesterol biosynthesis program and suppression of cholesterol efflux genes. Our findings support a therapeutic strategy of targeting RORγ alone or in combination with statin for effective treatment of mCRPC.

Nuclear receptor RORγ inverse agonists/antagonists display tissue- and gene-context selectivity through distinct activities in altering chromatin accessibility and master regulator SREBP2 occupancy.

The nuclear receptor RORγ is a major driver of autoimmune diseases and certain types of cancer due to its aberrant function in T helper 17 (Th17) cell differentiation and tumor cholesterol metabolism, respectively. Compound screening using the classic receptor-coactivator interaction perturbation scheme led to identification of many small-molecule modulators of RORγ(t). We report here that inverse agonists/antagonists of RORγ such as VTP-43742 derivative VTP-23 and TAK828F, which can potently inhibit the inflammatory gene program in Th17 cells, unexpectedly lack high potency in inhibiting the growth of TNBC tumor cells. In contrast, antagonists such as XY018 and GSK805 that strongly suppress tumor cell growth and survival display only modest activities in reducing Th17-related cytokine expression. Unexpectedly, we found that VTP-23 significantly induces the cholesterol biosynthesis program in TNBC cells. Our further mechanistic analyses revealed that VTP-23 enhances the local chromatin accessibility, H3K27ac mark and the cholesterol master regulator SREBP2 recruitment at the RORγ binding sites, whereas XY018 exerts the opposite activities. Yet, they display similar inhibitory effects on circadian rhythm program. Similar distinctions and contrasting activities between TAK828F and SR2211 in their effects on local chromatin structure at Il17 genes were also observed. Together, our study shows for the first-time that structurally distinct RORγ antagonists possess different or even contrasting activities in tissue/cell-specific manner. Our findings also highlight that the activities at natural chromatin are key determinants of RORγ modulators' tissue selectivity.

Ultra high performance liquid chromatography with quadrupole time-of-flight tandem mass spectrometry and liquid chromatography with tandem mass spectrometry combined with chemometric analysis as an approach for the quality evaluation of Mume Fructus.

Mume Fructus is an important traditional Chinese medicine that has been widely used in the treatment of intestinal diseases and asthma for thousands of years. In order to evaluate the quality of Mume Fructus in different processing methods, the main chemical components in Mume Fructus were investigated and a method was established for simultaneous quantification of organic acids of Mume Fructus. First, an optimized ultra-performance liquid chromatography-quadrupole-time of flight tandem-mass spectrometry method was used to identify the structures of main components in Mume Fructus. A total of 41 chemical compounds were identified, including 11 organic acids, 13 flavonoids, and three fatty acids. The contents of 11 organic acids in 18 batches of Mume Fructus from different processing methods were simultaneously determined by a liquid chromatography with tandem mass spectrometry method. The results of quantitative and hierarchical cluster analysis indicated that Mume Fructus under different processing methods were rich in the above 11 organic acids and the contents were obviously different. Taken together, the proposed quality evaluation method was fast and comprehensively reflects the content of the main chemical components in Mume Fructus under different processing methods, and provides a useful reference for the quality control and evaluation of Mume Fructus.

Dietary Conjugated Linoleic Acid Modulates the Hepatic Circadian Clock Program via PPARα/REV-ERBα-Mediated Chromatin Modification in Mice.

Scope: Disruptions of circadian rhythm cause metabolic disorders and are closely related to dietary factors. In this study, we investigated the interplays between the dietary conjugated linoleic acid (CLA)-induced hepatic steatosis and the circadian clock regulation, in association with lipid homeostasis. Methods and Results: Exposure of mice to 1.5% dietary CLA for 28 days caused insulin resistance, enlarged livers, caused hepatic steatosis, and increased triglyceride levels. Transcriptional profiling showed that hepatic circadian clock genes were significantly downregulated with increased expression of the negative transcription factor, REV-ERBα. We uncovered that the nuclear receptor (NR) PPARα, as a major target of dietary CLA, drives REV-ERBα expression via its binding to key genes of the circadian clock, including Cry1 and Clock, and the recruitment of histone marks and cofactors. The PPARα or REV-ERBα inhibition blocked the physical connection of this NR pair, reduced the cobinding of PPARα and REV-ERBα to the genomic DNA response element, and abolished histone modifications in the CLA-hepatocytes. In addition, we demonstrated that CLA promotes PPARα driving REV-ERBα transcriptional activity by directly binding to the PPAR response element (PPRE) at the Nr1d1 gene. Conclusions: Our results add a layer to the understanding of the peripheral clock feedback loop, which involves the PPARα-REV-ERBα, and provide guidance for nutrients optimization in circadian physiology.

Freezing-Tolerant, Highly Sensitive Strain and Pressure Sensors Assembled from Ionic Conductive Hydrogels with Dynamic Cross-Links.

Conductive hydrogels have attracted intensive attention for versatile functions in flexible electronics because of their unique combination of mechanical flexibility and conductivity. However, hydrogels containing plenty of water inevitably freeze at subzero temperature, leading to invalid electronics with failed mechanical advantages and negligible conductivity. Moreover, the inferior elasticity and fatigue resistance of hydrogels result in unstable sensing performance and poor reusability of hydrogel-based electronics. Herein, a freezing-tolerant, high-sensitive, durable strain and pressure sensor was constructed from an ionic conductive chitosan-poly(acrylamide-co-acrylic acid) double-network [CS-P(AM-co-AA) DN] hydrogel with dual-dynamic cross-links (chitosan physical network and ionic coordination [CO2LFeIII]), which was feasibly fabricated by soaking the CS-P(AM-co-AA) composite hydrogel in FeCl3 solution. The ions immobilized in dynamic cross-links exerted crucial effects on improving mechanics [prominent tensile performance, supercompressibility, extraordinary elasticity, fast self-recovery capacity, and remarkable fatigue resistance (1000 cycles)]; meanwhile, the free ions in the hydrogel rendered the hydrogel excellent conductivity and strong freezing tolerance concurrently. The sensor assembled from the DN hydrogel exhibited cycling stability and good durability in detecting pressure, various deformations (elongation, compression, and bend), and human motions even at a low temperature (-20 °C). Notably, the sensitivity on detecting strain and pressure at both room and subzero temperature was superior than most of the reported organohydrogel and hydrogel sensors. Thus, we believe that this work will provide a platform for construction and application of high-sensitive strain and pressure hydrogel sensors with cycling stability and good durability in a wide temperature range.

Phytoavailability of bound residue of Carbendazim to Chinese cabbage (Brassica campestris ssp.chinensis) coexisted with Superabsorbent polymers.

Understanding the bioavailability and phytotoxicity of Carbendazim (MBC) bound residues (BR) in soils incubated with different Superabsorbent polymer (SAP) amendment on succeeding crops is essential to assess their environmental fate and risks. In our research, we studied the morphological characteristics and 14C-accumulation of Chinese cabbage and released BR in three typical cultivated soils. The plant dry weight was in order of superabsorbent-hydrogels formulations (HMBC) > MBC > MBC and SAP (MBC-SAP) at 35 d in basic soil 3 (S3), with 675.40 ± 29.07 mg/plant.d.w, 575.93 ± 25.35 mg/plant.d.w and 427.86 ± 18.79 mg/plant.d.w. The whole plant accumulated 2-fold more BR when grew in neutral soil 2 (S2) treated with SAP than MBC at 7 d. The root accumulated a greater proportion of 14C-MBC residue than shoot, with order of MBC-SAP > MBC > HMBC at 21d. The results indicate MBC-BR could be released and accumulated in plant. HMBC promoted the Chinese cabbage growth with lowest 14C accumulation, while MBC-SAP inhibited plant growth with the highest 14C uptake. The released BR rate was 61.43 ± 3.75% of initial BR in MBC-SAP, with 2-fold higher than MBC and HMBC. It is assumed HMBC could be a potential environmentally friendly measure for rational use of pesticides in future.

Superabsorbent hydrogels coating increased degradation and decreased bound residues formation of carbendazim in soil.

The intensive use of pesticides has caused serious environmental pollution and ecological issues. Thus, it is imperative to explore an efficient way to minimize the pesticide residues and pollution. In the present study, we employed the superabsorbent hydrogels (SHs)-coated pesticide 14C-carbendazim (H-14C-MBC) to investigate the fate of MBC in aerobic soils and to assess the soil microbial state during incubation. The results showed that after coating with SHs, MBC dissipation was improved significantly by 34.2-54.1% compared with that in the control (p<0.05), reducing the persistence of MBC in soil matrix. At 100d, the release of 14C-CO2 was enhanced by 68.0% and 46.6% in neutral loamy soil and basic saline soil, respectively, with respect to the control, resulting in more complete degradation and detoxification of MBC. Additionally, the bound residue in soils, which was associated with potential environmental risk and pollution, was reduced by 15.2% and 14.2%, respectively, compared with that in control soils. The microbial diversity of post-H-14C-MBC soil varied, and microbial composition and abundance remained different from the control, even with the refreshment of soil stability and fertility compared with the blank soil. These results demonstrate the environmental behavior of SHs-coated MBC in soils, and illustrate that SHs-encapsulated formulations would be a promising measure for reducing the soil-residue pollution and environmental risk of pesticides.

Effects of superabsorbent polymers on the fate of fungicidal carbendazim in soils.

Superabsorbent polymers (SAPs) have been extensively used as soil amendments to retain water, and they often coexist with pesticides in agricultural fields. However, effects of SAPs on the fate of pesticides in soil remain poorly understood. In this study, a laboratory experiment was conducted to evaluate the effects of SAPs on the transformation of 14C-carbendazim in soils. The results showed that compared to the SAPs-free control, 11.4% relative reduction of 14C-carbendazim extractable residue was observed in red clayey soil with SAPs amendment after 100days of incubation (p<0.05). Carbendazim dissipation was enhanced by 34.7%, while no obvious difference was found in loamy soil and saline soil (p>0.05). SAPs changed the profiles of major metabolites (2-aminobenzimidazole and 2-hydroxybenzimidazole) to some extent. After 100days of SAPs treatment, the mineralization of 14C-carbendazim was significantly reduced by 37.6% and 41.2% in loamy soil and saline soil, respectively, relative to the SAPs-free treatment (p<0.05). SAPs increased the bound residue of carbendazim by 11.1-19.1% in comparison with SAPs-free controls. These findings suggest SAPs amendments significantly affected the fate of carbendazim and attention should be given to the assessment of environmental and ecological safety of pesticides in SAPs-amended soils.

Non-stereoselective transformation of the chiral insecticide cycloxaprid in aerobic soil.

Cycloxaprid (CYC) is one of the most effective neonicotinoid insecticides and is proposed to be a replacement of imidacloprid that has caused concerns over non-targeted resistance and ecological toxicity worldwide. The present study was performed with the 14C-labeled racemic CYC and its two enantiomers in aerobic soil. Racemic CYC and the enantiomers 1S2R-CYC and 1R2S-CYC underwent non-stereoselective degradation in the three soils tested. During the incubation period, CYC was transformed into three achiral degradation products which displayed varying degradation kinetics dependent upon soil properties. The soil properties were found to significantly influence the CYC metabolite profiles. The fastest degradation occurred in loamy soil, whereas the slowest reactions occurred in acidic clay soil. The primary transformation of CYC included cleavage of the oxabridged seven-member ring and CN between chloropyridinylmethyl and imidazalidine ring, carboxylation of the alkene group, and hydroxylation of imidazolidine ring. The results shed light on understanding of CYC degradation and provided useful information for applications and environmental assessments of chiral pesticides.