CancerMHL: the database of integrating key DNA methylation, histone modifications and lncRNAs in cancer.

The discovery of key epigenetic modifications in cancer is of great significance for the study of disease biomarkers. Through the mining of epigenetic modification data relevant to cancer, some researches on epigenetic modifications are accumulating. In order to make it easier to integrate the effects of key epigenetic modifications on the related cancers, we established CancerMHL (http://www.positionprediction.cn/), which provide key DNA methylation, histone modifications and lncRNAs as well as the effect of these key epigenetic modifications on gene expression in several cancers. To facilitate data retrieval, CancerMHL offers flexible query options and filters, allowing users to access specific key epigenetic modifications according to their own needs. In addition, based on the epigenetic modification data, three online prediction tools had been offered in CancerMHL for users. CancerMHL will be a useful resource platform for further exploring novel and potential biomarkers and therapeutic targets in cancer. Database URL: http://www.positionprediction.cn/.

Naringin Protects against Non-Alcoholic Fatty Liver Disease by Promoting Autophagic Flux and Lipophagy.

The autophagic degradation of lipid droplets, termed lipophagy, is the main mechanism contributing to lipid consumption in hepatocytes. Identifying effective and safe natural compounds that target lipophagy to eliminate excess lipids may be a potential therapeutic strategy for non-alcoholic fatty liver disease (NAFLD). Here the effects of naringin on NAFLD and the underlying mechanisms involved are investigated. Naringin treatment effectively relieves HFD-induced hepatic steatosis in mice and inhibits PA-induced lipid accumulation in hepatocytes. Increased p62 and LC3-II levels are observed with excess lipid support autophagosome accumulation and impaired autophagic flux. Treatment with naringin restores TFEB-mediated lysosomal biogenesis, thereby promoting the fusion of autophagosomes and lysosomes, restoring impaired autophagic flux and further inducing lipophagy. However, the knockdown of TFEB in hepatocytes or the hepatocyte-specific knockout of TFEB in mice abrogates naringin-induced lipophagy, eliminating its therapeutic effect on hepatic steatosis. These results demonstrate that TFEB-mediated lysosomal biogenesis and subsequent lipophagy play essential roles in the ability of naringin to mitigate hepatic steatosis and suggest that naringin is a promising drug for treating NAFLD.

Maternal COVID-19 Distress and Chinese Preschool Children's Problematic Media Use: A Moderated Serial Mediation Model.

Introduction: Maternal distress increased during the COVID-19 pandemic, significantly impacting children's media use. The purpose of this study was to explore the influence mechanism of maternal COVID-19 distress on preschoolers' problematic media us through a moderated mediation model; specifically, we examined the possible mediating roles of parenting stress and negative instrumental use of media in parenting and the moderating role of supportive co-parenting. Methods: An online survey was conducted in a sample of 1357 children (Mage = 4.01, SD = 1.06; 47.4% boys) and their parents from six public kindergartens in Shanghai, China. The mothers provided information by completing measures on their levels of distress related to COVID-19, parenting stress levels, digital parenting practices, and perception of supportive co-parenting from their partners. Additionally, both parents rated their children's problematic media use. Results: (1) maternal COVID-19 distress was significantly and positively related to children's problematic media use; (2) this relationship was sequentially mediated by parenting stress and parents' negative instrumental use of media in parenting; and (3) supportive co-parenting moderated the serial mediation path by reducing the effect of maternal COVID-19 distress on parenting stress. Conclusion: The findings provide some support and guidance for preventing children's problematic media use and enhancing parental adaptation during the COVID-19 pandemic or in potentially adverse situations.

Water-responsive gel extends drug retention and facilitates skin penetration for curcumin topical delivery against psoriasis.

Psoriasis is a chronic inflammatory skin disease characterized by erythema, scaling, and skin thickening. Topical drug application is recommended as the first-line treatment. Many formulation strategies have been developed and explored for enhanced topical psoriasis treatment. However, these preparations usually have low viscosity and limited retention on the skin surface, resulting in low drug delivery efficiency and poor patient satisfaction. In this study, we developed the first water-responsive gel (WRG), which has a distinct water-triggered liquid-to-gel phase transition property. Specifically, WRG was kept in a solution state in the absence of water, and the addition of water induced an immediate phase transition and resulted in a high viscosity gel. Curcumin was used as a model drug to investigate the potential of WRG in topical drug delivery against psoriasis. In vitro and in vivo data showed that WRG formulation could not only extend skin retention but also facilitate the drug permeating across the skin. In a mouse model of psoriasis, curcumin loaded WRG (CUR-WRG) effectively ameliorated the symptoms of psoriasis and exerted a potent anti-psoriasis effect by extending drug retention and facilitating drug penetration. Further mechanism study demonstrated that the anti-hyperplasia, anti-inflammation, anti-angiogenesis, anti-oxidation, and immunomodulation properties of curcumin were amplified by enhanced topical drug delivery efficiency. Notably, neglectable local or systemic toxicity was observed for CUR-WRG application. This study suggests that WRG is a promising formulation for topically psoriasis treatment.

The structural discrepancy between the small and large gut microbiota of Asiatic toad (Bufo gargarizans) during hibernation.

Hibernating amphibians are suitable for the research on the adaptation of gut microbiota to long-term fasting and cold stresses. However, the previous studies mainly focus on the large or whole gut microbiota but not the small gut microbiota. To test the structural discrepancy between the small and large gut microbiota during hibernation, we performed two independent batches of 16S rRNA gene amplicon sequencing to profile the small and large gut microbiota of hibernating Asiatic toad (Bufo gargarizans) from two wild populations. Both batches of data revealed that Proteobacteria, Bacteroidetes, and Firmicutes were the three most dominant phyla in the small and large gut microbiota. Three core OTUs with 100% occurrence in all gut microbiotas were annotated as Pseudomonas. A significant structural discrepancy was detected between the small and large gut microbiota. For instance, Proteobacteria assembled in the small intestine with a higher proportion than it did in the large intestine, but Bacteroidetes and Firmicutes assembled in the large intestine with a higher proportion than they did in the small intestine. The large gut microbiota exhibited higher diversity than the small gut microbiota. Nevertheless, a severe batch effect existed in the structural analysis of the gut microbiotas. The large gut microbiota showed a better resistance to the batch effect than the small gut microbiota did. This study provides preliminary evidence that microbes assemble in the small and large intestines of amphibians with discrepant patterns during hibernation.

Astragaloside IV ameliorates diet-induced hepatic steatosis in obese mice by inhibiting intestinal FXR via intestinal flora remodeling.

BACKGROUND: Nonalcoholic fatty liver disease (NAFLD) is a major clinical and public health burden worldwide with no established pharmacological therapy. Changes in the intestinal flora and associated metabolite bile acids (BAs) have been described in NAFLD. Astragaloside IV (AS-IV) is a low drug permeability saponin with protective effects against multiple diseases. However, the specific mechanism underlying the involvement of AS-IV in the regulation of NAFLD is yet to be clarified. PURPOSE: This study aimed to investigate the effect of AS-IV on NAFLD and explore whether intestinal flora was involved. METHODS: The effect of AS-IV was evaluated on high-fat diet-fed mice. Real-time PCR, immunohistochemistry, immunofluorescence, and biochemical analyses were performed. 16S rRNA gene sequencing and UPLC-TQMS were used to determine the alterations in the intestinal flora and concentration of BAs. Fecal microbiota transplantation (FMT) and intestine-specific farnesoid X receptor (FXR) knockout were also performed. RESULTS: AS-IV treatment alleviated diet-induced metabolic impairments, particularly hepatic steatosis. These changes occurred in the setting of decreased intestinal bile salt hydrolase (BSH)-expressing flora. Further analysis showed that the reduced BSH activity increased intestinal tauro-ß-muricholic acid levels, an inhibitor of intestinal FXR. Inhibition of intestinal FXR signaling by AS-IV was accompanied by decreased expression of intestinal fibroblast growth factor 15 and subsequent hepatic FXR activation as well as increased glucagon-like peptide-1 and decreased ceramide production, all of which contribute to the inhibition of sterol regulatory element-binding protein-1c-mediated hepatic steatosis. Furthermore, intestine-specific Fxr knockout and FMT further demonstrated an FXR- and intestinal flora-dependent preventive effect of AS-IV on hepatic steatosis. CONCLUSION: These results show that the changes in intestinal flora and BAs serve an essential role in the remission of hepatic steatosis by AS-IV, thereby suggesting that AS-IV may be used as a prebiotic agent to provide viable treatment for NAFLD.

The prediction of tumor and normal tissues based on the DNA methylation values of ten key sites.

Abnormal DNA methylation can alter the gene expression to promote or inhibit tumorigenesis in colon adenocarcinoma (COAD). However, the finding important genes and key sites of abnormal DNA methylation which result in the occurrence of COAD is still an eventful task. Here, we studied the effects of DNA methylation in the 12 types of genomic features on the changes of gene expression in COAD, the 10 important COAD-related genes and the key abnormal DNA methylation sites were identified. The effects of important genes on the prognosis were verified by survival analysis. Moreover, it was shown that the important genes were participated in cancer pathways and were hub genes in a co-expression network. Based on the DNA methylation levels in the ten sites, the least diversity increment algorithm for predicting tumor tissues and normal tissues in seventeen cancer types are proposed. The better results are obtained in jackknife test. For example, the predictive accuracies are 94.17 %, 91.28 %, 89.04 % and 88.89 %, respectively, for COAD, rectum adenocarcinoma, pancreatic adenocarcinoma and cholangiocarcinoma. Finally, by computing enrichment score of infiltrating immunocytes and the activity of immune pathways, we found that the genes are highly correlated with immune microenvironment.

Phillygenin ameliorates nonalcoholic fatty liver disease via TFEB-mediated lysosome biogenesis and lipophagy.

BACKGROUND: Lipophagy is an autophagic process, which delivers the intracellular lipid droplets to the lysosomes for degradation. Recent studies revealed that the impairment of lysosomal biogenesis and autophagic flux led to dysregulation of lipophagy in hepatocytes, which exacerbated the development of nonalcoholic fatty liver disease (NAFLD). Therefore, agents restoring autophagic flux and lipophagy in hepatocytes may have therapeutic potential against this increasingly prevalent disease. Phillygenin (PHI), a lignin extracted from Forsythia suspense, exerts hepatoprotective and anti-inflammatory effects. However, the effect of PHI on NAFLD remains unknown. PURPOSE: This study aimed to investigate the protective effect of PHI on NAFLD and elucidate the underlying mechanism. METHODS: The effects of PHI were examined in palmitate (PA)-stimulated AML12 cells and primary hepatocytes, as well as in NAFLD mice induced by a high-fat diet (HFD). We also used transcription factor EB (TFEB) knockdown hepatocytes and hepatocyte-specific TFEB knockout (TFEBΔhep) mice for mechanistic studies. In vivo and in vitro studies were performed using western blots, immunofluorescence techniques, and transmission electron microscopy. RESULTS: Our results indicated that autophagic flux and lysosome biogenesis in PA-stimulated hepatocytes were impaired. PHI alleviated lipid deposition by increasing lysosomal biogenesis and autophagic flux. It also stimulated the release of endoplasmic reticulum Ca2+ to activate calcineurin, which regulated TFEB dephosphorylation and nuclear translocation, and promoted lysosomal biogenesis. In addition, PHI blocked the NLRP3 inflammasome pathway and improved hepatocyte inflammation in an autophagy-dependent manner. Consistent with the in vitro results, PHI improved hepatic steatosis and inflammation in HFD mice, but these beneficial effects were eliminated in hepatocyte-specific TFEB knockout mice. CONCLUSION: Despite PHI has been reported to have anti-hepatic fibrosis effects, whether it has a hepatoprotective effects against NAFLD and the underlying molecular mechanism remain unclear. Herein, we found that PHI restored lipophagy and suppressed lipid accumulation and inflammation by regulating the Ca2+-calcineurin-TFEB axis in hepatocytes. Thus, PHI represents a therapeutic candidate for the treatment of NAFLD.

Cardioprotection effect of Yiqi-Huoxue-Jiangzhuo formula in a chronic kidney disease mouse model associated with gut microbiota modulation and NLRP3 inflammasome inhibition.

BACKGROUND: The pathogenesis and treatment of cardiovascular disease mediated by chronic kidney disease (CKD) are key research questions. Specifically, the mechanisms underlying the cardiorenal protective effect of Yiqi-Huoxue-Jiangzhuo formula (YHJF), a traditional Chinese herbal medicine, have not yet been clarified. METHODS: A classical CKD mouse model was constructed by 5/6 nephrectomy (Nx) to study the effects of YHJF intervention on 5/6 Nx mice cardiorenal function, gut microbial composition, gut-derived metabolites, and NLRP3 inflammasome pathways. RESULTS: YHJF improved cardiac dysfunction and reversed left ventricular hypertrophy, myocardial hypertrophy, and interstitial fibrosis in 5/6 Nx mice. In addition, YHJF inhibited activation of the NLRP3 inflammasome and downregulated the expression of TNF-α and IL-1ß both in the heart and serum; reconstitution of the intestinal flora imbalance was also found in 5/6 Nx mice treated with YHJF. Spearman's correlation and redundancy analyses showed that changes in the intestinal flora of 5/6 Nx mice were related to clinical phenotype and serum inflammatory levels. CONCLUSIONS: Treatment with YHJF effectively protected the heart function of 5/6 Nx mice; this effect was attributed to inhibition of NLRP3 inflammasome activation and regulation of intestinal microbial composition and derived metabolites. YHJF has potential for improving intestinal flora imbalance and gut-derived toxin accumulation in patients with CKD, thereby preventing cardiovascular complications.

Quercetin protects against palmitate-induced pancreatic ß-cell apoptosis by restoring lysosomal function and autophagic flux.

Quercetin, a natural flavonoid, has been reported to prevent pancreatic ß-cell apoptosis in animal models of diabetes. However, the underlying mechanism remains unclear. We investigated the mechanisms through which quercetin protects ß cells from palmitate-induced apoptosis and determined whether autophagy is involved in this process. We found that quercetin treatment partially reduced palmitate-induced ß-cell apoptosis. This protective effect was abolished by pharmacologic inhibition of autophagy and by silencing a key autophagy gene. Further analysis revealed that palmitate treatment promoted the expression of LC3 II, a marker of autophagosomes, but resulted in the blockade of autophagic flux due to lysosome dysfunction. Defective lysosome accumulation can cause lysosomal membrane permeabilization and the release of cathepsins from lysosome into the cytosol that triggers apoptosis. Treatment with quercetin reversed lysosomal dysfunction and promoted autophagosome-lysosome fusion, which restored defective autophagic flux and provoked autophagy. Overall, our results indicate that lysosomal dysfunction is a major factor that contributes to ß-cell apoptosis and demonstrates that quercetin improves cell survival by restoring lysosomal function and autophagic flux. This study provides new evidence regarding the anti-apoptotic mechanism of quercetin in the treatment of type 2 diabetes.

Intestinal Flora Mediates Antiobesity Effect of Rutin in High-Fat-Diet Mice.

SCOPE: Intestinal flora plays a critical role in the development of . Rutin is a natural flavonoid with potential prebiotic effects on regulating the intestinal flora composition that is beneficial for host health. Therefore, this study hypothesizes that rutin supplementation has beneficial effects on high-fat-diet (HFD)-induced obesity and metabolic disorder through the modulation of intestinal flora in mice. METHODS AND RESULTS: The obesity-alleviating property of rutin using 6-week-old C57BL/6J male mice fed on HFD with or without rutin supplementation for 16 weeks is investigated. Rutin supplementation effectively reduces body-weight gain, insulin resistance, and acted favorably on the intestinal barrier, thereby reducing endotoxemia and systemic inflammation. Sequencing of 16S rRNA genes from fecal samples indicate that rutin exerted modulatory effects on HFD-induced intestinal flora disorders (e.g., rutin decreased Firmicutes abundance and increased Bacteroidetes and Verrucomicrobia abundance). Antibiotic treatment and fecal microbiota transplantation further demonstrate that the salutary effects of rutin on obesity control are strongly dependent on the intestinal flora. CONCLUSION: Rutin can be considered as a prebiotic agent for improving intestinal flora disorders and obesity-associated metabolic perturbations in obese individuals.

Alantolactone-loaded chitosan/hyaluronic acid nanoparticles suppress psoriasis by deactivating STAT3 pathway and restricting immune cell recruitment.

Psoriasis is a common chronic immune-mediated skin disease characterized by hyperproliferation and aberrant differentiation of keratinocytes and massive infiltration of inflammatory immune cells. Recent studies showed that Signal Transducer and Activator of Transcription 3 (STAT3), which plays an important role in cell survival, proliferation, differentiation, angiogenesis, and immune responses, is constitutively activated in epidermal keratinocytes of human psoriatic skin lesions. In addition, STAT3 promotes the differentiation and expansion of T cells and facilitates cytokine production, thereby exacerbating the condition of psoriasis. Alantolactone (ALT) is a sesquiterpene lactone compound that could selectively suppress STAT3 activation, but its effectiveness and application in psoriasis treatment have not been determined. In this study, we developed ALT loaded chitosan/hyaluronic acid nanoparticles (CHALT), and investigated its therapeutic potential for psoriasis therapy. CHALT effectively abrogated the hyperproliferation by inducing ROS-mediated apoptosis with loss of mitochondrial membrane potential, and also inhibited IL-6-induced STAT3 signaling activation and inflammatory reaction in HaCaT cell line. In an Imiquimod (IMQ)-induced psoriasis model, the topical treatment of psoriasis lesions with CHALT effectively attenuated the STAT3 hyperactivation within keratinocytes and ameliorated the symptoms of psoriasis. In addition, it was found that CHALT restricted the recruitment of immune cells. These results indicated that ALT-based nanoformulation CHALT holds great potential for psoriasis therapy.

MicroRNA-100-5p and microRNA-298-5p released from apoptotic cortical neurons are endogenous Toll-like receptor 7/8 ligands that contribute to neurodegeneration.

BACKGROUND: MicroRNA (miRNA) expression in the brain is altered in neurodegenerative diseases. Recent studies demonstrated that selected miRNAs conventionally regulating gene expression at the post-transcriptional level can act extracellularly as signaling molecules. The identity of miRNA species serving as membrane receptor ligands involved in neuronal apoptosis in the central nervous system (CNS), as well as the miRNAs' sequence and structure required for this mode of action remained largely unresolved. METHODS: Using a microarray-based screening approach we analyzed apoptotic cortical neurons of C56BL/6 mice and their supernatant with respect to alterations in miRNA expression/presence. HEK-Blue Toll-like receptor (TLR) 7/8 reporter cells, primary microglia and macrophages derived from human and mouse were employed to test the potential of the identified miRNAs released from apoptotic neurons to serve as signaling molecules for the RNA-sensing receptors. Biophysical and bioinformatical approaches, as well as immunoassays and sequential microscopy were used to analyze the interaction between candidate miRNA and TLR. Immunocytochemical and -histochemical analyses of murine CNS cultures and adult mice intrathecally injected with miRNAs, respectively, were performed to evaluate the impact of miRNA-induced TLR activation on neuronal survival and microglial activation. RESULTS: We identified a specific pattern of miRNAs released from apoptotic cortical neurons that activate TLR7 and/or TLR8, depending on sequence and species. Exposure of microglia and macrophages to certain miRNA classes released from apoptotic neurons resulted in the sequence-specific production of distinct cytokines/chemokines and increased phagocytic activity. Out of those miRNAs miR-100-5p and miR-298-5p, which have consistently been linked to neurodegenerative diseases, entered microglia, located to their endosomes, and directly bound to human TLR8. The miRNA-TLR interaction required novel sequence features, but no specific structure formation of mature miRNA. As a consequence of miR-100-5p- and miR-298-5p-induced TLR activation, cortical neurons underwent cell-autonomous apoptosis. Presence of miR-100-5p and miR-298-5p in cerebrospinal fluid led to neurodegeneration and microglial accumulation in the murine cerebral cortex through TLR7 signaling. CONCLUSION: Our data demonstrate that specific miRNAs are released from apoptotic cortical neurons, serve as endogenous TLR7/8 ligands, and thereby trigger further neuronal apoptosis in the CNS. Our findings underline the recently discovered role of miRNAs as extracellular signaling molecules, particularly in the context of neurodegeneration.

Qing-Re-Xiao-Zheng Formula Modulates Gut Microbiota and Inhibits Inflammation in Mice With Diabetic Kidney Disease.

Evidence indicates that the metabolic inflammation induced by gut microbiota dysbiosis contributes to diabetic kidney disease. Prebiotic supplementations to prevent gut microbiota dysbiosis, inhibit inflammatory responses, and protect the renal function in DKD. Qing-Re-Xiao-Zheng formula (QRXZF) is a Traditional Chinese Medicine (TCM) formula that has been used for DKD treatment in China. Recently, there are growing studies show that regulation of gut microbiota is a potential therapeutic strategy for DKD as it is able to reduce metabolic inflammation associated with DKD. However, it is unknown whether QRXZF is effective for DKD by regulating of gut microbiota. In this study, we investigated the reno-protective effect of QRXZF by exploring its potential mechanism between gut microbiota and downstream inflammatory pathways mediated by gut-derived lipopolysaccharide (LPS) in the kidney. High-fat diet (HFD) and streptozotocin injection-induced DKD mice model was established to assess the QRXZF effect in vivo. Mice treated with QRXZF for 8 weeks had significantly lower levels of urinary albumin, serum cholesterol and triglycerides. The renal injuries observed through histological analysis were attenuated as well. Also, mice in the QRXZF group had higher levels of Zonula occludens protein-1 (ZO-1) expression, lower levels of serum fluorescein-isothiocyanate (FITC)-dextran and less-damaged colonic mucosa as compared to the DKD group, implying the benefit role for the gut barrier integrity. QRXZF treatment also reversed gut dysbiosis and reduced levels of gut-derived LPS. Notably, the expression of toll-like receptor 4 (TLR4) and nuclear factor-κB (NF-κB), which are important inflammation pathways in DKD, were suppressed in the QRXZF groups. In conclusion, our results indicated that the reno-protective effects of QRXZF was probably associated with modulating gut microbiota and inhibiting inflammatory responses in the kidney.

Decisive Effects of Life Stage on the Gut Microbiota Discrepancy Between Two Wild Populations of Hibernating Asiatic Toads (Bufo gargarizans).

Until now, the effects of driving factors on the gut microbiota of amphibians are still mostly confounded. Due to a long-term fasting, hibernating amphibians are ideal experimental materials to explore this question. In this study, we characterized the small intestine microbiota of adult hibernating Asiatic toads (Bufo gargarizans) collected from two geographical populations using 16S rRNA amplicon sequencing technique and evaluated the effects of non-dietary factors (e.g., sex and host genetic background). Proteobacteria (0.9196 ± 0.0892) was characterized as the most dominant phylum in the small gut microbiota of hibernating Asiatic toads, among which five core OTUs were identified and three were classified into Pseudomonas. In view of the coincidence between the dominant KEGG pathways (such as the two-component system) and Pseudomonas, Pseudomonas appeared to be a key adaptor for small gut microbiota during hibernation. Furthermore, we detected a greater discrepancy of gut microbiota between geographical populations than between sexes. Both sex and host genetic background showed a minor effect on the gut microbiota variation. Finally, life stage was determined to be the decisive factor driving the gut microbiota discrepancy between populations. However, a large proportion of the gut microbiota variation (∼70%) could not be explained by the measured deterministic factors (i.e., sex, location, body length, and routine blood indices). Therefore, other factors and/or stochastic processes may play key roles in shaping gut bacterial community of hibernating amphibians.

Structure-aware machine learning identifies microRNAs operating as Toll-like receptor 7/8 ligands.

MicroRNAs (miRNAs) can serve as activation signals for membrane receptors, a recently discovered function that is independent of the miRNAs' conventional role in post-transcriptional gene regulation. Here, we introduce a machine learning approach, BrainDead, to identify oligonucleotides that act as ligands for single-stranded RNA-detecting Toll-like receptors (TLR)7/8, thereby triggering an immune response. BrainDead was trained on activation data obtained from in vitro experiments on murine microglia, incorporating sequence and intra-molecular structure, as well as inter-molecular homo-dimerization potential of candidate RNAs. The method was applied to analyse all known human miRNAs regarding their potential to induce TLR7/8 signalling and microglia activation. We validated the predicted functional activity of subsets of high- and low-scoring miRNAs experimentally, of which a selection has been linked to Alzheimer's disease. High agreement between predictions and experiments confirms the robustness and power of BrainDead. The results provide new insight into the mechanisms of how miRNAs act as TLR ligands. Eventually, BrainDead implements a generic machine learning methodology for learning and predicting the functions of short RNAs in any context.

A novel strategy based on targeted cellular metabolomics for quantitatively evaluating anti-aging effect and screening effective extracts of Erzhi Wan.

The complexity of ingredients in traditional Chinese medicine (TCM) makes it challenging to clarify its efficacy in an acceptable and scientific approach. The present study was aimed to use quantification results from targeted cellular metabolomics to evaluate anti-aging efficacy of a famous Chinese medicine formula, Erzhi Wan (EZW), and screen possible effective extracts, depending on the developed strategy integrating multivariate receiver operating characteristic (ROC) curve and analytic hierarchy process (AHP). In this study, senescent NRK cells induced by D-galactose were treated with drug-containing serum of EZW and four kinds of extracts (petroleum ether, ethyl acetate, butanol and water). Intermediates of two major metabolic pathways for energy synthesis, tricarboxylic acid (TCA) cycle and glycolysis, were accurately quantified by GC-MS/MS to identify discriminate metabolites for clarifying therapeutic mechanism of EZW based on multivariate statistical analysis. Senescent and non-senescent cells were successfully distinguished using these metabolites by ROC curve analysis. Next, these metabolites were used as evaluation indexes to quantitatively reflect different effect of EZW and its extracts, according to the role of them in distinguishing groups and in conjunction with AHP. In vitro detection of senescence-associated ß-galactosidase (SA-ß-gal) activity was used to verify the reliability of evaluation results. The reversal after treatment of drug-containing serum of EZW and extracts was observed, and the petroleum ether extract might be the potential active extract responsible for the major anti-aging effect of EZW, which was in agreement with in vitro experiments. Altogether, metabolomics was a powerful approach for evaluation efficacy and elucidation action mechanisms of TCM. The integrated evaluation strategy in this paper with properties of high practicality, feasibility and effectivity was expected to provide a new insight into comprehensive and quantitative efficacy evaluation.

Extracellular Matrix Stiffness Regulates DNA Methylation by PKCα-Dependent Nuclear Transport of DNMT3L.

Extracellular matrix (ECM) stiffness has profound effects on the regulation of cell functions. DNA methylation is an important epigenetic modification governing gene expression. However, the effects of ECM stiffness on DNA methylation remain elusive. Here, it is reported that DNA methylation is sensitive to ECM stiffness, with a global hypermethylation under stiff ECM condition in mouse embryonic stem cells (mESCs) and embryonic fibroblasts compared with soft ECM. Stiff ECM enhances DNA methylation of both promoters and gene bodies, especially the 5' promoter regions of pluripotent genes. The enhanced DNA methylation is functionally required for the loss of pluripotent gene expression in mESCs grown on stiff ECM. Further experiments reveal that the nuclear transport of DNA methyltransferase 3-like (DNMT3L) is promoted by stiff ECM in a protein kinase C α (PKCα)-dependent manner and DNMT3L can be binding to Nanog promoter regions during cell-ECM interactions. These findings unveil DNA methylation as a novel target for the mechanical sensing mechanism of ECM stiffness, which provides a conserved mechanism for gene expression regulation during cell-ECM interactions.

Hyaluronic acid coated bilirubin nanoparticles attenuate ischemia reperfusion-induced acute kidney injury.

Acute kidney injury (AKI) is a common pathological process that is globally associated with a high morbidity and mortality rate. The underlying AKI mechanisms include over-produced reactive oxygen species (ROS), inflammatory cell infiltration, and high levels of inflammatory mediators. Bilirubin is an endogenous compound with antioxidant, anti-inflammatory and anti-apoptotic properties, and could, therefore, be a promising therapeutic candidate. Nanotechnology-mediated therapy has emerged as a novel drug delivery strategy for AKI treatment. In this study, we report a hyaluronic acid (HA) coated ε-polylysine-bilirubin conjugate (PLBR) nanoparticle (nHA/PLBR) that can selectively accumulate in injured kidneys and alleviate the oxidative/inflammatory-induced damage. The in vitro study revealed that nHA/PLBR has good stability, biocompatibility, and exhibited higher antioxidant as well as anti-apoptotic effects when compared to nPLBR or bilirubin. The in vivo study showed that nHA/PLBR could target and accumulate in the injured kidney, effectively relieve oxidative stress and inflammatory reactions, protect the structure and function of the mitochondria, and more importantly, inhibit the apoptosis of tubular cells in an ischemia/reperfusion-induced AKI rat model. Therefore, nHA/PLBR has the capacity to enhance specific biodistribution and delivery efficiency of bilirubin, thereby providing better treatment for AKI in the future.

Polylysine-bilirubin conjugates maintain functional islets and promote M2 macrophage polarization.

Macrophage polarization is one of the main factors contributing to the proinflammatory milieu of transplanted islets. It causes significant islet loss. Bilirubin exhibits protective effects during the islet transplantation process, but the mode of delivering drugs along with the islet graft has not yet been developed. In addition, it remains unclear whether bilirubin or its derivatives can modulate macrophage polarization during islet transplantation. Therefore, this study aimed to develop an ε-polylysine-bilirubin conjugate (PLL-BR) to encapsulate the islets for protection and to explore its macrophage modulation activities. In in vitro studies, the PLL-BR was shown to tightly adhere to the islet surface. It also exhibited enhanced cytoprotective effects against oxidative and inflammatory conditions by promoting M2-type macrophage polarization. In in vivo studies, the PLL-BR-protected islets successfully prolonged the euglycemia period in diabetic mice and accelerated the blood glucose clearance rate by maintaining the insulin secretion function. Compared to the untreated islets, the PLL-BR-encapsulated islets induced anti-inflammatory responses that were characterized by elevated levels of M2 macrophage markers and local vascularization. In conclusion, PLL-BR can be used as a tool for reprograming macrophage polarization while providing a more efficient immune protection for transplanted islets. STATEMENT OF SIGNIFICANCE: Macrophage polarization is one main factor that caused significant loss of transplanted islets. Bilirubin possesses protective effects toward pancreatic islet, but how to deliver the drug along with the islet graft has not yet been harnessed. More importantly, whether bilirubin or its derivatives could modulate macrophage polarization during the host rejections has also not been answered. In this study, we developed an ε-polylysine-bilirubin conjugate (PLL-BR) to encapsulate the islets and explore its role in macrophage modulation activities. PLL-BR could attach to the surface of islets and exerted high oxidation resistance and anti-inflammatory effect. For the first time, we demonstrate that bilirubin and its derivatives effectively promoted the M2-type macrophage polarization, and optimize the immune microenvironment for islets survival and function.