Diet-Induced Circadian Enhancer Remodeling Synchronizes Opposing Hepatic Lipid Metabolic Processes.

Overnutrition disrupts circadian metabolic rhythms by mechanisms that are not well understood. Here, we show that diet-induced obesity (DIO) causes massive remodeling of circadian enhancer activity in mouse liver, triggering synchronous high-amplitude circadian rhythms of both fatty acid (FA) synthesis and oxidation. SREBP expression was rhythmically induced by DIO, leading to circadian FA synthesis and, surprisingly, FA oxidation (FAO). DIO similarly caused a high-amplitude circadian rhythm of PPARα, which was also required for FAO. Provision of a pharmacological activator of PPARα abrogated the requirement of SREBP for FAO (but not FA synthesis), suggesting that SREBP indirectly controls FAO via production of endogenous PPARα ligands. The high-amplitude rhythm of PPARα imparted time-of-day-dependent responsiveness to lipid-lowering drugs. Thus, acquisition of rhythmicity for non-core clock components PPARα and SREBP1 remodels metabolic gene transcription in response to overnutrition and enables a chronopharmacological approach to metabolic disorders.

An intrinsically disordered region controlling condensation of a circadian clock component and rhythmic transcription in the liver.

Circadian gene transcription is fundamental to metabolic physiology. Here we report that the nuclear receptor REV-ERBα, a repressive component of the molecular clock, forms circadian condensates in the nuclei of mouse liver. These condensates are dictated by an intrinsically disordered region (IDR) located in the protein's hinge region which specifically concentrates nuclear receptor corepressor 1 (NCOR1) at the genome. IDR deletion diminishes the recruitment of NCOR1 and disrupts rhythmic gene transcription in vivo. REV-ERBα condensates are located at high-order transcriptional repressive hubs in the liver genome that are highly correlated with circadian gene repression. Deletion of the IDR disrupts transcriptional repressive hubs and diminishes silencing of target genes by REV-ERBα. This work demonstrates physiological circadian protein condensates containing REV-ERBα whose IDR is required for hub formation and the control of rhythmic gene expression.

Isoform-specific functions of PPARγ in gene regulation and metabolism.

Peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor that is a vital regulator of adipogenesis, insulin sensitivity, and lipid metabolism. Activation of PPARγ by antidiabetic thiazolidinediones (TZD) reverses insulin resistance but also leads to weight gain that limits the use of these drugs. There are two main PPARγ isoforms, but the specific functions of each are not established. Here we generated mouse lines in which endogenous PPARγ1 and PPARγ2 were epitope-tagged to interrogate isoform-specific genomic binding, and mice deficient in either PPARγ1 or PPARγ2 to assess isoform-specific gene regulation. Strikingly, although PPARγ1 and PPARγ2 contain identical DNA binding domains, we uncovered isoform-specific genomic binding sites in addition to shared sites. Moreover, PPARγ1 and PPARγ2 regulated a different set of genes in adipose tissue depots, suggesting distinct roles in adipocyte biology. Indeed, mice with selective deficiency of PPARγ1 maintained body temperature better than wild-type or PPARγ2-deficient mice. Most remarkably, although TZD treatment improved glucose tolerance in mice lacking either PPARγ1 or PPARγ2, the PPARγ1-deficient mice were protected from TZD-induced body weight gain compared with PPARγ2-deficient mice. Thus, PPARγ isoforms have specific and separable metabolic functions that may be targeted to improve therapy for insulin resistance and diabetes.

A coregulator shift, rather than the canonical switch, underlies thyroid hormone action in the liver.

Thyroid hormones (THs) are powerful regulators of metabolism with major effects on body weight, cholesterol, and liver fat that have been exploited pharmacologically for many years. Activation of gene expression by TH action is canonically ascribed to a hormone-dependent "switch" from corepressor to activator binding to thyroid hormone receptors (TRs), while the mechanism of TH-dependent repression is controversial. To address this, we generated a mouse line in which endogenous TRß1 was epitope-tagged to allow precise chromatin immunoprecipitation at the low physiological levels of TR and defined high-confidence binding sites where TRs functioned at enhancers regulated in the same direction as the nearest gene in a TRß-dependent manner. Remarkably, although positive and negative regulation by THs have been ascribed to different mechanisms, TR binding was highly enriched at canonical DR4 motifs irrespective of the transcriptional direction of the enhancer. The canonical NCoR1/HDAC3 corepressor complex was reduced but not completely dismissed by TH and, surprisingly, similar effects were seen at enhancers associated with negatively as well as positively regulated genes. Conversely, coactivator CBP was found at all TH-regulated enhancers, with transcriptional activity correlating with the ratio of CBP to NCoR rather than their presence or absence. These results demonstrate that, in contrast to the canonical "all or none" coregulator switch model, THs regulate gene expression by orchestrating a shift in the relative binding of corepressors and coactivators.

Assessment and comparison of viability assays for cellular products.

BACKGROUND AIMS: Accurate assessment of cell viability is crucial in cellular product manufacturing, yet selecting the appropriate viability assay presents challenges due to various factors. This study compares and evaluates different viability assays on fresh and cryopreserved cellular products, including peripheral blood stem cell (PBSC) and peripheral blood mononuclear cell (PBMC) apheresis products, purified PBMCs and cultured chimeric antigen receptor and T-cell receptor-engineered T-cell products. METHODS: Viability assays, including manual Trypan Blue exclusion, flow cytometry-based assays using 7-aminoactinomycin D (7-AAD) or propidium iodide (PI) direct staining or cell surface marker staining in conjunction with 7-AAD, Cellometer (Nexcelom Bioscience LLC, Lawrence, MA, USA) Acridine Orange/PI staining and Vi-CELL BLU Cell Viability Analyzer (Beckman Coulter, Inc, Brea, CA, USA), were evaluated. A viability standard was established using live and dead cell mixtures to assess the accuracy of these assays. Furthermore, precision assessment was conducted to determine the reproducibility of the viability assays. Additionally, the viability of individual cell populations from cryopreserved PBSC and PBMC apheresis products was examined. RESULTS: All methods provided accurate viability measurements and generated consistent and reproducible viability data. The assessed viability assays were demonstrated to be reliable alternatives when evaluating the viability of fresh cellular products. However, cryopreserved products exhibited variability among the tested assays. Additionally, analyzing the viability of each subset of the cryopreserved PBSC and PBMC apheresis products revealed that T cells and granulocytes were more susceptible to the freeze-thaw process, showing decreased viability. CONCLUSIONS: The study demonstrates the importance of careful assay selection, validation and standardization, particularly for assessing the viability of cryopreserved products. Given the complexity of cellular products, choosing a fit-for-purpose viability assay is essential.

Effectiveness of Global Leadership Initiative on Malnutrition and Subjective Global Assessment for diagnosing malnutrition and predicting wound healing in patients with diabetic foot ulcers.

Malnutrition significantly hampers wound healing processes. This study aimed to compare the effectiveness of the Global Leadership Initiative on Malnutrition (GLIM) and Subjective Global Assessment (SGA) in diagnosing malnutrition and predicting wound healing in patients with diabetic foot ulcers (DFU). GLIM criteria were evaluated for sensitivity (SE), specificity (SP), positive predictive value, negative predictive value and kappa (κ) against SGA as the reference. Modified Poisson regression model and the DeLong test investigated the association between malnutrition and non-healing ulcers over 6 months. This retrospective cohort study included 398 patients with DFU, with a mean age of 66·3 ± 11·9 years. According to SGA and GLIM criteria, malnutrition rates were 50·8 % and 42·7 %, respectively. GLIM criteria showed a SE of 67·3 % (95 % CI 60·4 %, 73·7 %) and SP of 82·7 % (95 % CI 76·6 %, 87·7 %) in identifying malnutrition, with a positive predictive value of 80·0 % and a negative predictive value of 71·1 % (κ = 0·50) compared with SGA. Multivariate analysis demonstrated that malnutrition, as assessed by SGA, was an independent risk factor for non-healing (relative risk (RR) 1·84, 95 % CI 1·45, 2·34), whereas GLIM criteria were associated with poorer ulcer healing in patients with estimated glomerular filtration rate ≥ 60 ml/min/1·73m2 (RR: 1·46, 95 % CI 1·10, 1·94). SGA demonstrated a superior area under the receiver's operating characteristic curve for predicting non-healing compared with GLIM criteria (0·70 (0·65-0·75) v. 0·63 (0·58-0·65), P < 0·01). These findings suggest that both nutritional assessment tools effectively identify patients with DFU at increased risk, with SGA showing superior performance in predicting non-healing ulcers.

Excess iron accumulation mediated senescence in diabetic kidney injury.

Cellular senescence and iron accumulation were separately observed in diabetic nephropathy (DN). Limited evidence supports that iron was significantly accumulated in senescent cells. We aimed to explore whether iron is involved in the pathogenesis role of senescence in DN. Renal cells were treated with high glucose (HG, 35 mM) for 10 or 15 days, and DN mice were induced by high-fat diet and streptozotocin. Gene ontology enrichment, gene set enrichment analysis analysis, ß-galactosidase staining, 5-ethynyl-2-deoxyuridine staining, and western blot depicted the upregulated senescence pathway in vitro and in vivo of DN. Lactate dehydrogenase (LDH) release was increased by HG and reversed by p16/p21 knockdown, and the supernatant of HG-treated cells caused increased LDH release from normal cells. Iron metabolism-related protein expression was disordered after HG exposure concomitant with senescence. Ferric ammonium citrate (50 µM) upregulated gamma-H2A.X variant histone and increased the senescence markers in HG-treated cells. The treatment of deferoxamine (0.5 µM) had the opposite effect. Compared to the non-DN individual, increased ferritin and senescence markers were verified in DN mice and patients, and the co-localization of ferritin and senescence markers was observed by immunofluorescence. These results suggested that accumulated iron was correlated with aggravated DNA damage and accelerated senescence, and revealed the role of iron in the cellular senescence of diseases.

Indirubin-3'-oxime promotes the efficacy of GnRHa in obese-induced central precocious puberty and maintains normal bone growth and body weight via the ERK-Sp1-KISS-1/GPR54 axis.

Central precocious puberty (CPP) is a widespread developmental abnormality. The application of gonadotrophin-releasing hormone agonist (GnRHa) is widely useful for the medical therapy of CPP. This study aimed to investigate the combination effect and mechanism of indirubin-3'-oxime (I3O), an active ingredient analogue of traditional Chinese medicine, and GnRHa treatment on the progression of CPP. First, female C57BL/6 mice were fed with a high-fat diet (HFD) for the induction of precocious puberty and treated with GnRHa and I3O alone or in combination. Development of sexual maturation, bone growth and obesity were determined by vaginal opening detection, H&E staining and ELISA. The protein and mRNA expression levels of related genes were evaluated via western blotting, immunohistochemical method and RT-qPCR. Subsequently, tBHQ, an inhibitor of ERK, was applied to verify whether the mechanism of I3O was associated with this signaling. The results showed that the treatment of I3O alone or in combination with GnRHa could alleviate the HFD-induced earlier vaginal opening and serum levels of the gonadal hormone in mice. And, I3O could significantly eliminate the role of growth deceleration of GnRHa in bone development and reversed the side effect of GnRHa on body weight. More importantly, we found that I3O decreased the expression of KISS-1 and GPR54 by suppressing the phosphorylation of ERK1/2 and Sp1 in the hypothalamus in mice. In summary, these data indicated that I3O could promote the efficacy of GnRHa in HFD-induced precocious puberty, and maintain bone growth and body weight in mice via the ERK-Sp1-KISS-1/GPR54 axis.

Lower serum insulin-like growth factor-1 levels are independently associated with anemia in patients undergoing maintenance hemodialysis.

The relationship between serum insulin-like growth factor-1 (IGF-1) levels and anemia in patients undergoing maintenance hemodialysis (MHD) remains unclear. This cross-sectional study included patients who underwent MHD treatment for >3 months at our dialysis center in March 2021. Demographic and clinical data were recorded. Blood samples were collected before the hemodialysis sessions, and general serum biochemical parameters, routine blood markers, and serum IGF-1 levels were measured. Patients were divided into a group without anemia (hemoglobin ≥110 g/L) and a group with anemia (hemoglobin <110 g/L), and multivariable linear and binary logistic regression analyses were performed to study the relationship between the levels of serum IGF-1 and anemia. A total of 165 patients (male/female = 99:66) with MHD were enrolled in the study, with a median age of 66.0 (58.0, 75.0) years and a median dialysis vintage of 27.0 (12.0, 55.0) months. The mean hemoglobin level was 96.38 ± 16.72 g/L, and 126 patients had anemia (76.4%). Compared to patients without anemia, patients with anemia had lower serum IGF-1 and triglyceride levels and higher intravenous iron supplementation on dialysis (all p < 0.05). After adjusting for confounding factors in different models, the nine-model multivariate binary logistic regression analyses also confirmed that lower serum IGF-1 levels and serum IGF-1 < 197.03 ng/ml were both independently associated with anemia in patients undergoing MHD. However, further multicenter studies with larger sample sizes are required to confirm these findings.

SR9009 has REV-ERB-independent effects on cell proliferation and metabolism.

The nuclear receptors REV-ERBα and -ß link circadian rhythms and metabolism. Like other nuclear receptors, REV-ERB activity can be regulated by ligands, including naturally occurring heme. A putative ligand, SR9009, has been reported to elicit a range of beneficial effects in healthy as well as diseased animal models and cell systems. However, the direct involvement of REV-ERBs in these effects of SR9009 has not been thoroughly assessed, as experiments were not performed in the complete absence of both proteins. Here, we report the generation of a mouse model for conditional genetic deletion of REV-ERBα and -ß. We show that SR9009 can decrease cell viability, rewire cellular metabolism, and alter gene transcription in hepatocytes and embryonic stem cells lacking both REV-ERBα and -ß. Thus, the effects of SR9009 cannot be used solely as surrogate for REV-ERB activity.

Purinergic P2X7 Receptor Mediates the Elimination of Trichinella spiralis by Activating NF-κB/NLRP3/IL-1ß Pathway in Macrophages.

Trichinellosis is one of most neglected foodborne zoonoses worldwide. During Trichinella spiralis infection, the intestinal immune response is the first line of defense and plays a vital role in the host's resistance. Previous studies indicate that purinergic P2X7 receptor (P2X7R) and pyrin domain-containing protein 3 (NLRP3) inflammasome are involved in the intestinal immune response in T. spiralis infection. However, the precise role of P2X7R and its effect on NLRP3 remains largely underdetermined. In this study, we aimed to investigate the role of P2X7R in the activation of NLRP3 in macrophages during the intestinal immune response against T. spiralis We found that T. spiralis infection upregulated expression of P2X7R and activation of NLRP3 in macrophages in mice. In vivo, P2X7R deficiency resulted in increased intestinal adult and muscle larval burdens, along with decreased expression of NLRP3/interleukin-1ß (IL-1ß) in macrophages from the infected mice with T. spiralis In In vitro experiments, P2X7R blockade inhibited activation of NLRP3/IL-1ß via NF-κB and thus reduced the capacity of macrophages to kill newborn larvae of T. spiralis These results indicate that P2X7R mediates the elimination of T. spiralis by activating the NF-κB/NLRP3/IL-1ß pathway in macrophages. Our findings contribute to the understanding of the intestinal immune mechanism of T. spiralis infection.

Landscape of the long non-coding RNA transcriptome in human heart.

Long non-coding RNAs (lncRNAs) have been revealed to play essential roles in the human cardiovascular system. However, information about their mechanisms is limited, and a comprehensive view of cardiac lncRNAs is lacking from a multiple tissues perspective to date. Here, the landscape of the lncRNA transcriptome in human heart was summarized. We summarized all lncRNA transcripts from publicly available human transcriptome resources (156 heart samples and 210 samples from 29 other tissues) and systematically analysed all annotated and novel lncRNAs expressed in heart. A total of 7485 lncRNAs whose expression was elevated in heart (HE lncRNAs) and 453 lncRNAs expressed in all 30 analysed tissues (EIA lncRNAs) were extracted. Using various bioinformatics resources, methods and tools, the features of these lncRNAs were discussed from various perspectives, including genomic structure, conservation, dynamic variation during heart development, cis-regulation, differential expression in cardiovascular diseases and cancers as well as regulation at transcriptional and post-transcriptional levels. Afterwards, all the features discussed above were integrated into a user-friendly resource named CARDIO-LNCRNAS (http://bio-bigdata.hrbmu.edu.cn/CARDIO-LNCRNAS/ or http://www.bio-bigdata.net/CARDIO-LNCRNAS/). This study represents the first global view of lncRNAs in the human cardiovascular system based on multiple tissues and sheds light on the role of lncRNAs in developments and heart disorders.

Establishment and validation of in-house cryopreserved CAR/TCR-T cell flow cytometry quality control.

BACKGROUND: Chimeric antigen receptor (CAR) or T-cell receptor (TCR) engineered T-cell therapy has recently emerged as a promising adoptive immunotherapy approach for the treatment of hematologic malignancies and solid tumors. Multiparametric flow cytometry-based assays play a critical role in monitoring cellular manufacturing steps. Since manufacturing CAR/TCR T-cell products must be in compliance with current good manufacturing practices (cGMP), a standard or quality control for flow cytometry assays should be used to ensure the accuracy of flow cytometry results, but none is currently commercially available. Therefore, we established a procedure to generate an in-house cryopreserved CAR/TCR T-cell products for use as a flow cytometry quality control and validated their use. METHODS: Two CAR T-cell products: CD19/CD22 bispecific CAR T-cells and FGFR4 CAR T-cells and one TCR-engineered T-cell product: KK-LC-1 TCR T-cells were manufactured in Center for Cellular Engineering (CCE), NIH Clinical Center. The products were divided in aliquots, cryopreserved and stored in the liquid nitrogen. The cryopreserved flow cytometry quality controls were tested in flow cytometry assays which measured post-thaw viability, CD3, CD4 and CD8 frequencies as well as the transduction efficiency and vector identity. The long-term stability and shelf-life of cryopreserved quality control cells were evaluated. In addition, the sensitivity as well as the precision assay were also assessed on the cryopreserved quality control cells. RESULTS: After thawing, the viability of the cryopreserved CAR/TCR T-cell controls was found to be greater than 50%. The expression of transduction efficiency and vector identity markers by the cryopreserved control cells were stable for at least 1 year; with post-thaw values falling within ± 20% range of the values measured at time of cryopreservation. After thawing and storage at room temperature, the stability of these cryopreserved cells lasted at least 6 h. In addition, our cryopreserved CAR/TCR-T cell quality controls showed a strong correlation between transduction efficiency expression and dilution factors. Furthermore, the results of flow cytometric analysis of the cryopreserved cells among different laboratory technicians and different flow cytometry instruments were comparable, highlighting the reproducibility and reliability of these quality control cells. CONCLUSION: We developed and validated a feasible and reliable procedure to establish a bank of cryopreserved CAR/TCR T-cells for use as flow cytometry quality controls, which can serve as a quality control standard for in-process and lot-release testing of CAR/TCR T-cell products.

Combinatorial epigenetic regulation of non-coding RNAs has profound effects on oncogenic pathways in breast cancer subtypes.

Although systematic genomic studies have identified a broad spectrum of non-coding RNAs (ncRNAs) that are involved in breast cancer, our understanding of the epigenetic dysregulation of those ncRNAs remains limited. Here, we systematically analysed the epigenetic alterations of microRNAs (miRNAs) and long non-coding RNAs (lncRNAs) in two breast cancer subtypes (luminal and basal). Widespread epigenetic alterations of miRNAs and lncRNAs were observed in both cancer subtypes. In contrast to protein-coding genes, the majority of epigenetically dysregulated ncRNAs were shared between subtypes, but a subset of transcriptomic and corresponding epigenetic changes occurred in a subtype-specific manner. In addition, our findings suggested that various types of epi-modifications might synergistically modulate ncRNA transcription. Our observations further highlighted the complementary dysregulation of epi-modifications, particularly of miRNA members within the same family, which produced the same directed alterations as a result of diverse epi-modifications. Functional enrichment analysis revealed that epigenetically dysregulated ncRNAs were significantly involved in several hallmarks of cancers. Finally, our analysis of epigenetic modification-mediated miRNA regulatory networks revealed that cancer progression was associated with specific miRNA-gene modules in two subtypes. This study enhances understanding of the aberrant epigenetic patterns of ncRNA expression and provides new insights into the functions of ncRNAs in breast cancer subtypes.

ApoE deficiency promotes hepatic pathology by aggravating Th17/Treg imbalance in murine schistosomiasis japonica.

AIMS: The Schistosoma japonicum (S japonicum)-infected ApoE gene deficiency (ApoE-/- ) mice were used to determine effect of ApoE on hepatic immunopathology. METHODS: Murine activities and appetite, body weight, and ratio of liver weight to its body weight (Hepatic mass index, HMI) were observed. Worm load and liver egg burden were evaluated as the infection intensity. Number and size of liver egg granulomas and serum levels of alanine aminotransferase (ALT) were investigated. We analysed hepatic fibrosis by markers of fibrosis in tissue, detected hepatic Th17 and Treg frequency by flow cytometry, and measured hepatic expressions of RORγt, Foxp3, IL-17A and TGF-ß1 via qPCR. Lipid metabolism was determined by serum levels of cholesterol (TC) and triglyceride (TG) as well as hepatic Oil red O staining. RESULTS: In the infected ApoE-/- mice, the increased infection intensity aggravated the hepatic immunopathology (evidenced by increased HMI, elevated egg granulomas and increased ALT levels) and fibrosis (increased hepatic collagen deposition). ApoE deficiency resulted in significantly elevated ratio of hepatic Th17/Treg and higher serum levels of TC and TG, along with higher level of hepatic Oil red O staining. CONCLUSIONS: ApoE deficiency promotes hepatic pathology and fibrosis by exacerbating Th17/Treg imbalance and altering lipid metabolism in murine schistosomiasis japonica.

Manganese-doped zinc oxide hollow balls for chemiresistive sensing of acetone vapors.

Both pure and Mn(II)-doped ZnO hollow structures were synthesized by a solvothermal reaction, and their phase structures, morphologies and elemental composition were characterized. SEM and TEM observations show the pure ZnO and the Mn(II)-doped ZnO balls to possess similar hollow structure with a particle size of about 1.5 µm. Their sensing properties were investigated, and the composite containing 1 atom% of Mn(II) (1% Mn-ZnO) is found be display the highest selectivity for acetone. The detection limit is 100 ppm acetone at 234 °C which is 4.6 times lower than that of the pure ZnO. In addition, the response time is shorter. Graphical abstract ZnO and Mn-doped ZnO hollow balls were prepared by a hydrothermal method, and their gas-sensing properties were investigated. Zinc(II) oxide doped with 1 atom% Mn(II) demonstrated an outstanding sensing behavior towards acetone vapors.

Biosynthesis of SnO2 Nanoparticles Based on Response Surface Methodology and the Study of Their Dye Removal.

Litsea cubeba is an evergreen tree from the Lauraceae family, with the common name of mountain pepper in Taiwan. The extracts from different parts such as the bark, leaf, root and fruit have various medicinal properties and have been extensively used in traditional Chinese medicine. SnO2 nanoparticles (NPs) were synthesized using an extract of Litsea cubeba fruit based on response surface methodology (RSM). The phytochemicals present in the extract of Litsea cubeba fruit played a crucial role in the NP formation, which could be confirmed by Fourier-Transform Infrared Spectrometer (FTIR). Analysis of variance (ANOVA) was used to evaluate the validation of the RSM models and indicated that the quadratic model was highly significant and suitable to represent the response of the NP formation yield. The optimum parameters were determined using the Design Expert Program and were as follows: extract volume ratio 30.0%, pH 6.3 and reaction temperature 44.4 °C. Using the optimal reaction conditions from the central composite design (CCD) of RSM, the SnO2 NPs were prepared and characterized by field-emission scanning electron microscopy (FE-SEM), energy dispersive X-ray spectroscopy (EDX), transmission electron microscopy (TEM) and X-ray diffraction (XRD). The FE-SEM results showed that the SnO2 NPs were spherical, and XRD results showed that the NPs had a tetragonal crystal structure. Furthermore, the photodegradation kinetics of malachite green by the SnO2 NPs was represented well with a pseudo first-order model.

Comparative epigenetic analyses reveal distinct patterns of oncogenic pathways activation in breast cancer subtypes.

Breast cancer is a highly heterogeneous disease that is characterized by genetic and epigenetic aberrations; however, our knowledge of epigenetic alterations of breast cancer subtypes remains limited. Here, we portrayed and compared the alterations of six types of histone modifications and DNA methylation between two breast cancer subtypes, luminal and basal. Widespread subtype-specific epigenetic alterations were observed in both subtypes, which preferentially occurred within CpG islands (CGIs) and promoter regions. Specifically, aberrant DNA methylation was mostly located inside CGIs in luminal subtype, whereas in basal subtype it was principally located within CGI shores. Moreover, different types and combinatorial patterns of epigenetic alterations were found to occupy at promoter regions between these two subtypes. And these epigenetic alterations regulated corresponding gene expression in a synergetic way in both subtypes. Functional enrichment analysis highlighted that epigenetically dysregulated genes were significantly involved in the hallmarks of cancers, most of which were subtype specific. Even genes involved in the same hallmarks associated biological processes were affected by various types of epi-modifications in different subtypes. Finally, we revealed distinct patterns of oncogenic pathways activation in different subtypes and provided novel insights into subtype specific therapeutic opportunities. In addition, genes in the key signaling pathways were able to discriminate between disease phenotypes, and subtype-specific progression associated genes were identified. This study presents the aberrant epigenetic patterns of breast cancer subtypes at a genome-wide level, which will be a highly valuable resource for investigations at understanding epigenetic regulation of breast cancer subtypes.

Homoharringtonine production by endophytic fungus isolated from Cephalotaxus hainanensis Li.

Homoharringtonine (HHT), a natural plant alkaloid derived from Cephalotaxus, has demonstrated to have a broad antitumor activity and efficacy in treating human chronic myeloid leukemia. An alternative source is required to substitute for the slow-growing and scarce Cephalotaxus to meet the increasing demand of the drug market. The objective of this study was to screen HHT-producing endophytic fungi from Cephalotaxus hainanensis Li. By screening 213 fungal isolates obtained from the bark parts of Cephalotaxus hainanensis Li, one isolate was found to be capable of biosynthesizing HHT. The fungus was identified as Alternaria tenuissima by morphological characteristics and internal transcribed spacer (ITS) sequence analysis and was named as CH1307. HHT obtained from CH1307 was analyzed through the HPLC and LC-MS/MS and NMR spectroscopy. The extract of the fermentation broth of CH1307 showed antiproliferative activities against K562 (chronic myelocytic leukemia), NB4 (acute promyelocytic leukemia), and HL-60 (promyelocytic leukemia) human cancer cell lines with IC50 values of 67.25 ± 4.26, 65.02 ± 4.75, and 99.23 ± 4.26 µg/mL, respectively. The findings suggest that HHT-producing endophytic fungus, Alternaria tenuissima CH1307 might provide a promising source for the research and application of HHT.

Partial reduction of interleukin-33 signaling improves senescence and renal injury in diabetic nephropathy.

Diabetic nephropathy (DN) is a frequent and costly complication of diabetes with limited understandings of mechanisms and therapies. Emerging evidence points to the important roles of interleukin-33 (IL-33) in acute kidney injury, yet its contribution to DN is still unclear. We here found a ubiquitous increase of IL-33 and its receptor (ST2) in murine models and patients with DN. Surprisingly, both IL-33 and ST2 knockdown aggravated renal lesions in DN, while overexpression of IL-33 also exacerbated the condition. Further population-based analyses revealed a positive correlation of IL-33 expression with renal dysfunction in DN patients. Individuals with high IL-33 expression-related polygenic risk score had a higher DN risk. These findings confirmed the harmful effects of IL-33 on DN. Conversely, endogenous and exogenous partial reduction of IL-33 signaling conferred renoprotective effects in vivo and in vitro. Mechanistically, IL-33 induced senescence by regulating cell cycle factors in HK-2 cells, and accordingly senescence led to renal cell damage through the secretion of senescence-related secretory phenotype (SASP) including IL-33 and prostaglandins. Together, elevated IL-33 accelerates cellular senescence to drive DN possibly by SASP production, while a partial blockage improves renal injury and senescence. Our findings pinpoint a possible and new avenue for DN interventions.