The scale of zebrafish pectoral fin buds is determined by intercellular K+ levels and consequent Ca2+-mediated signaling via retinoic acid regulation of Rcan2 and Kcnk5b.

K+ channels regulate morphogens to scale adult fins, but little is known about what regulates the channels and how they control morphogen expression. Using the zebrafish pectoral fin bud as a model for early vertebrate fin/limb development, we found that K+ channels also scale this anatomical structure, and we determined how one K+-leak channel, Kcnk5b, integrates into its developmental program. From FLIM measurements of a Förster Resonance Energy Transfer (FRET)-based K+ sensor, we observed coordinated decreases in intracellular K+ levels during bud growth, and overexpression of K+-leak channels in vivo coordinately increased bud proportions. Retinoic acid, which can enhance fin/limb bud growth, decreased K+ in bud tissues and up-regulated regulator of calcineurin (rcan2). rcan2 overexpression increased bud growth and decreased K+, while CRISPR-Cas9 targeting of rcan2 decreased growth and increased K+. We observed similar results in the adult caudal fins. Moreover, CRISPR targeting of Kcnk5b revealed that Rcan2-mediated growth was dependent on the Kcnk5b. We also found that Kcnk5b enhanced depolarization in fin bud cells via Na+ channels and that this enhanced depolarization was required for Kcnk5b-enhanced growth. Lastly, Kcnk5b-induced shha transcription and bud growth required IP3R-mediated Ca2+ release and CaMKK activity. Thus, we provide a mechanism for how retinoic acid via rcan2 can regulate K+-channel activity to scale a vertebrate appendage via intercellular Ca2+ signaling.

WSB1, a Hypoxia-Inducible E3 Ligase, Promotes Myofibroblast Accumulation and Attenuates Alveolar Epithelial Regeneration in Mouse Lung Fibrosis.

Idiopathic pulmonary fibrosis is a progressive interstitial lung disease for which there is no curative therapy available. Repetitive alveolar epithelial injury repair, myofibroblast accumulation, and excessive collagen deposition are key pathologic features of idiopathic pulmonary fibrosis, eventually leading to cellular hypoxia and respiratory failure. The precise mechanism driving this complex maladaptive process remains inadequately understood. WD repeat and suppressor of cytokine signaling box containing 1 (WSB1) is an E3 ubiquitin ligase, the expression of which is associated strongly with hypoxia, and forms a positive feedback loop with hypoxia-inducible factor 1α (HIF-1α) under anoxic condition. This study explored the expression, cellular distribution, and function of WSB1 in bleomycin (BLM)-induced mouse lung injury and fibrosis. WSB1 expression was highly induced by BLM injury and correlated with the progression of lung fibrosis. Significantly, conditional deletion of Wsb1 in adult mice ameliorated BLM-induced pulmonary fibrosis. Phenotypically, Wsb1-deficient mice showed reduced lipofibroblast to myofibroblast transition, but enhanced alveolar type 2 proliferation and differentiation into alveolar type 1 after BLM injury. Proteomic analysis of mouse lung tissues identified caveolin 2 as a potential downstream target of WSB1, contributing to BLM-induced epithelial injury repair and fibrosis. These findings unravel a vital role for WSB1 induction in lung injury repair, thus highlighting it as a potential therapeutic target for pulmonary fibrosis.

U2AF2-SNORA68 promotes triple-negative breast cancer stemness through the translocation of RPL23 from nucleoplasm to nucleolus and c-Myc expression.

BACKGROUND: Small nucleolar RNAs (snoRNAs) play key roles in ribosome biosynthesis. However, the mechanism by which snoRNAs regulate cancer stemness remains to be fully elucidated. METHODS: SNORA68 expression was evaluated in breast cancer tissues by in situ hybridization and qRT‒PCR. Proliferation, migration, apoptosis and stemness analyses were used to determine the role of SNORA68 in carcinogenesis and stemness maintenance. Mechanistically, RNA pull-down, RNA immunoprecipitation (RIP), cell fractionation and coimmunoprecipitation assays were conducted. RESULTS: SNORA68 exhibited high expression in triple-negative breast cancer (TNBC) and was significantly correlated with tumor size (P = 0.048), ki-67 level (P = 0.037), and TNM stage (P = 0.015). The plasma SNORA68 concentration was significantly lower in patients who achieved clinical benefit. The SNORA68-high patients had significantly shorter disease-free survival (DFS) (P = 0.036). Functionally, SNORA68 was found to promote the cell stemness and carcinogenesis of TNBC in vitro and in vivo. Furthermore, elevated SNORA68 expression led to increased nucleolar RPL23 expression and retained RPL23 in the nucleolus by binding U2AF2. RPL23 in the nucleolus subsequently upregulated c-Myc expression. This pathway was validated using a xenograft model. CONCLUSION: U2AF2-SNORA68 promotes TNBC stemness by retaining RPL23 in the nucleolus and increasing c-Myc expression, which provides new insight into the regulatory mechanism of stemness.

The small nucleolar RNA SNORA51 enhances breast cancer stem cell-like properties via the RPL3/NPM1/c-MYC pathway.

Breast cancer stem cells (BCSCs) are key players in carcinogenesis and development. Small nucleolar RNAs (snoRNAs) seem to have a crucial influence on regulating stem cell-like properties in various cancers, but the underlying mechanism in breast cancer has not been determined. In this study, we first found that the expression of SNORA51 might be strongly and positively related to BCSCs-like properties. SNORA51 expression was assessed in breast cancer tissues (n = 158 patients) by in situ hybridization. Colony formation, cell counting kit-8, and sphere formation assays were used to detect cell proliferation and self-renewal, respectively. Wound healing and transwell assays were used to detect cell migration. Coimmunoprecipitation and molecular docking were used to determine the underlying mechanism through which SNORA51 regulates BCSCs-like properties. High SNORA51 expression was associated with a worse prognosis, overall survival, and disease-free survival, in 158 breast cancer patients and was also closely related to lymph node status, ER status, the Ki-67 index, histological grade, and TNM stage. Further analysis proved that SNORA51 could enhance and maintain stem cell-like properties, including cell proliferation, self-renewal, and migration, in breast cancer. Moreover, high SNORA51 expression could reduce nucleolar RPL3 expression, induce changes in the expression of NPM1 in the nucleolus and nucleoplasm, and ultimately increase c-MYC expression. Taken together, our findings demonstrated that SNORA51 could enhance BCSCs-like properties via the RPL3/NPM1/c-MYC pathway both in vitro and in vivo. Therefore, SNORA51 might be a significant biomarker and potential therapeutic target and might even provide a new viewpoint on the regulatory mechanism of snoRNAs in breast cancer or other malignant tumors.

Construction of an Endogenously Activated Signal Amplification Assay for In Situ Imaging of MicroRNA and Guided Precise Photodynamic Therapy for Cancer Cells.

The construction of a sensitive strategy for in situ visualizing and dynamic tracing intracellular microRNA is of great importance. Via the toehold-mediated strand displacement process, the catalytic hairpin assembly (CHA) could offer several hundreds-fold signal amplifications. However, the CHA may produce certain background interferences since microRNA may exist in normal cells. In this work, we constructed an endogenously and sequentially activated signal amplification strategy to provide the amplified dual-color fluorescence imaging of microRNA in living cancer cells, which was comprised of two successive reaction processes: the activation of the preprotective catalytic probe by the endogenous glutathione (GSH) and the subsequent catalytic hairpin assembly on the surface of the upconversion nanoprobe triggered by the specific microRNA. Since the concentration of GSH in cancer cells was much higher than that in normal cells and the extracellular environment, the activation of the designed nanoprobe could be controlled at the desirable site. With the merits of the endogenous initiation and selective activation, the designed nanoprobe could achieve the bioimaging of microRNA in living cancer cells with high precision and reliability. Furthermore, via the introduction of a photosensitizer molecule into the DNA strand, the designed nanoplatform could achieve the precise photodynamic therapy (PDT) for cancer cells and malignant tumors under the irradiation of the NIR laser. This work provided a new avenue to achieve the accurate imaging of intracellular microRNA and guided precise PDT, which would offer powerful hints to the early diagnosis and therapy of malignant tumors.

Pan-cancer analysis of NLRP3 inflammasome with potential implications in prognosis and immunotherapy in human cancer.

NLRP3 inflammasome was introduced as a double-edged sword in tumorigenesis and influenced immunotherapy response by modulating host immunity. However, a systematic assessment of the NLRP3-inflammasome-related genes across human cancers is lacking, and the predictive role of NLRP3 inflammasome in cancer immunotherapy (CIT) response remains unexplored. Thus, in this study, we performed a pan-cancer analysis of NLRP3-inflammasome-related genes across 24 human cancers. Out of these 24 cancers, 15 cancers had significantly different expression of NLRP3-inflammasome-related genes between normal and tumor samples. Meanwhile, Cox regression analysis showed that the NLRP3 inflammasome score could be served as an independent prognostic factor in skin cutaneous melanoma. Further analysis indicated that NLRP3 inflammasome may influence tumor immunity mainly by mediating tumor-infiltrating lymphocytes and macrophages, and the effect of NLRP3 inflammasome on immunity is diverse across tumor types in tumor microenvironment. We also found that the NLRP3 inflammasome score could be a stronger predictor for immune signatures compared with tumor mutation burden (TMB) and glycolytic activity, which have been reported as immune predictors. Furthermore, analysis of the association between NLRP3 inflammasome and CIT response using six CIT response datasets revealed the predictive value of NLRP3 inflammasome for immunotherapy response of patients in diverse cancers. Our study illustrates the characterization of NLRP3 inflammasome in multiple cancer types and highlights its potential value as a predictive biomarker of CIT response, which can pave the way for further investigation of the prognostic and therapeutic potentials of NLRP3 inflammasome.

Efficacy and safety of transurethral resection of bladder tumour combined with chemotherapy and immunotherapy in bladder-sparing therapy in patients with T1 high-grade or T2 bladder cancer: a protocol for a randomized controlled trial.

BACKGROUND: Bladder cancer is the tenth most common cancer worldwide. For patients with T1 high-grade or T2 bladder cancer, radical cystectomy is recommended. However, radical cystectomy is associated with various complications and has a detrimental impact on the quality of life. Bladder-sparing therapy has been widely explored in patients with muscle-invasive bladder cancer, and whether a combination of transurethral resection of bladder tumour (TURBT) with chemotherapy and immunotherapy shows definite superiority over TURBT plus chemotherapy is still a matter of debate. The aim of this study is to investigate the efficacy and safety of TURBT combined with chemotherapy and immunotherapy in bladder-sparing therapy in patients with T1 high-grade or T2 bladder cancer who are unwilling or unsuitable to undergo radical cystectomy. METHODS: An open-label, multi-institutional, two-armed randomized controlled study will be performed with 86 patients with T1 high-grade or T2 bladder cancer meeting the eligibility criteria. Participants in the experimental group (n = 43) will receive TURBT combined with chemotherapy (GC: gemcitabine 1000 mg/m2 on the 1st day and the 8th day, cisplatin 70 mg/m2 on the 2nd day, repeated every 21 days) and immunotherapy (toripalimab 240 mg on the 5th day, repeated every 21 days), and those in the control group (n = 43) will receive TURBT plus chemotherapy (GC). The primary outcome is pathological response, and the secondary outcomes include progression-free survival, overall survival, toxicities, and quality of life. DISCUSSION: To the best of our knowledge, this is the first study to evaluate the efficacy and safety of TURBT combined with GC regimen and toripalimab in bladder-sparing therapy in patients with T1 high-grade or T2 bladder cancer. The expected benefit is that the combination of TURBT with chemotherapy and immunotherapy would be more effective than TURBT plus chemotherapy without compromising the quality of life and increasing the toxicity. TRIAL REGISTRATION: ChiCTR2200060546, chictr.org.cn, registered on June 14, 2022.

Hyperactive DNA Cutting for Unbiased UHPLC-MS/MS Quantification of Epigenetic DNA Marks by Engineering DNase I Mutants.

Epigenetic DNA modifications, such as 5-methylcytosine, 5-hydroxymethylcytosine, and 5-formylcytosine, are associated with a variety of diseases and potential biomarkers for cancer diagnosis and therapy. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) assays are considered to be the gold standard for qualitative and quantitative detection of DNA modifications. DNA digestion for converting long DNA polymer into 2'-deoxynucleosides is an important preprocessing step to achieve sensitive and accurate LC-MS/MS quantification. Here, we showed that, as stimulated by divalent metal ions, Mg2+ and Mn2+, the engineered human DNase I Q9R:E13R:N74K mutant can efficiently digest DNA in the presence of monovalent metal ions at a high concentration (e.g., 1 M NaCl), showing hyperactivity on DNA cutting. We also found that the engineered DNase I mutants display exceptional DNA-cutting activity over a wider pH range (5.5-9.5). Due to their hyperactivity and high salt tolerance, the engineered DNase I mutants cut DNA 5mC and dC efficiently. Benefitting from this DNA-cutting hyperactivity, we demonstrated an LC-MS/MS assay for unbiased and accurate quantification of DNA 5mC.

The GA-DELLA-OsMS188 module controls male reproductive development in rice.

Pollen protects male sperm and allows flowering plants to adapt to diverse terrestrial environments, thereby leading to the rapid expansion of plants into new regions. The process of anther/pollen development is coordinately regulated by internal and external factors including hormones. Currently, the molecular mechanisms underlying gibberellin (GA)-mediated male reproductive development in plants remain unknown. We show here that rice DELLA/SLR1, which encodes the central negative regulator of GA signaling, is essential for rice anther development. The slr1-5 mutant exhibits premature programmed cell death of the tapetum, lacks Ubisch bodies, and has no exine and no mature pollen. SLR1 is mainly expressed in tapetal cells and tetrads, and is required for the appropriate expression of genes encoding key factors of pollen development, which are suggested to be OsMS188-targeted genes. OsMS188 is the main component in the essential genetic program of tapetum and pollen development. Further, we demonstrate that SLR1 interacts with OsMS188 to cooperatively activate the expression of the sporopollenin biosynthesis and transport-related genes CYP703A3, DPW, ABCG15 and PKS1 for rapid formation of pollen walls. Overall, the results of this study suggest that the GA hormonal signal is integrated into the anther genetic program and regulates rice anther development through the GA-DELLA-OsMS188 module.

DEAD-box helicase 27 enhances stem cell-like properties with poor prognosis in breast cancer.

BACKGROUND: Although the rapid development of diagnosis and treatment has improved prognosis in early breast cancer, challenges from different therapeutic response remain due to breast cancer heterogeneity. DEAD-box helicase 27 (DDX27) had been proved to influence ribosome biogenesis and identified as a promoter in gastric and colorectal cancer associated with stem cell-like properties, while the impact of DDX27 on breast cancer prognosis and biological functions is unclear. We aimed to explore the influence of DDX27 on stem cell-like properties and prognosis in breast cancer. METHODS: The expression of DDX27 was evaluated in 24 pairs of fresh breast cancer and normal tissue by western blot. We conducted Immunohistochemical (IHC) staining in paraffin sections of 165 breast cancer patients to analyze the expression of DDX27 and its correlation to stemness biomarker. The Cancer Genome Atlas-Breast Cancer (TCGA-BRCA) database and the Clinical Proteomic Tumor Analysis Consortium (CPTAC) database were used to analyze the expression of DDX27 in breast cancer. Kaplan-Meier survival analysis were used to investigate the implication of DDX27 on breast cancer prognosis. Western blot, CCK-8 assay, Transwell assay and wound-healing assay were carried out to clarify the regulation of DDX27 on stem cell-like properties in breast cancer cells. Gene Set Enrichment Analysis (GSEA) was performed to analyze the potential molecular mechanisms of DDX27 in breast cancer. RESULTS: DDX27 was significantly high expressed in breast cancer compared with normal tissue. High expression of DDX27 was related to larger tumor size (p = 0.0005), positive lymph nodes (p = 0.0008), higher histological grade (p = 0.0040), higher ki-67 (p = 0.0063) and later TNM stage (p < 0.0001). Patients with high DDX27 expression turned out a worse prognosis on overall survival (OS, p = 0.0087) and disease-free survival (DFS, p = 0.0235). Overexpression of DDX27 could enhance the expression of biomarkers related to stemness and promote stem cell-like activities such as proliferation and migration in breast cancer cells. CONCLUSION: DDX27 can enhance stem cell-like properties and cause poor prognosis in breast cancer, also may be expected to become a potential biomarker for breast cancer therapy.

Identification of the prognostic value of ferroptosis-related gene signature in breast cancer patients.

BACKGROUND: Breast cancer (BRCA) is a malignant tumor with high morbidity and mortality, which is a threat to women's health worldwide. Ferroptosis is closely related to the occurrence and development of breast cancer. Here, we aimed to establish a ferroptosis-related prognostic gene signature for predicting patients' survival. METHODS: Gene expression profile and corresponding clinical information of patients from The Cancer Genome Atlas (TCGA) database and Gene Expression Omnibus (GEO) database. The Least absolute shrinkage and selection operator (LASSO)-penalized Cox regression analysis model was utilized to construct a multigene signature. The Kaplan-Meier (K-M) and Receiver Operating Characteristic (ROC) curves were plotted to validate the predictive effect of the prognostic signature. Gene Ontology (GO) and Kyoto Encyclopedia of Genes, Genomes (KEGG) pathway and single-sample gene set enrichment analysis (ssGSEA) were performed for patients between the high-risk and low-risk groups divided by the median value of risk score. RESULTS: We constructed a prognostic signature consisted of nine ferroptosis-related genes (ALOX15, CISD1, CS, GCLC, GPX4, SLC7A11, EMC2, G6PD and ACSF2). The Kaplan-Meier curves validated the fine predictive accuracy of the prognostic signature (p < 0.001). The area under the curve (AUC) of the ROC curves manifested that the ferroptosis-related signature had moderate predictive power. GO and KEGG functional analysis revealed that immune-related responses were largely enriched, and immune cells, including activated dendritic cells (aDCs), dendritic cells (DCs), T-helper 1 (Th1), were higher in high-risk groups (p < 0.001). Oppositely, type I IFN response and type II IFN response were lower in high-risk groups (p < 0.001). CONCLUSION: Our study indicated that the ferroptosis-related prognostic signature gene could serve as a novel biomarker for predicting breast cancer patients' prognosis. Furthermore, we found that immunotherapy might play a vital role in therapeutic schedule based on the level and difference of immune-related cells and pathways in different risk groups for breast cancer patients.

LncRNA SPRY4-IT1 regulates breast cancer cell stemness through competitively binding miR-6882-3p with TCF7L2.

SPRY4-intronic transcript 1 has been found in several kinds of cancers, but the role of SPRY4-IT1 in breast cancer stem cells has not been studied. We investigated whether SPRY4-IT1 is involved in the promotion of breast cancer stem cells (BCSCs). We used qRT-PCR to detect the expression of SPRY4-IT1 in MCF-7 cells and MCF-7 cancer stem cells (MCF-7 CSCs). The effects of SPRY4-IT1 on the proliferation and renewal ability of breast cancer cells were investigated by in vitro and in vivo assays (ie in situ hybridization, colony formation assay, sphere formation assay, flow cytometry assay, western blotting, xenograft model and immunohistochemistry). The mechanism of SPPRY4-IT1 as a ceRNA was studied by a dual-luciferase reporter assay and bioinformatic analysis. In our study, SPRY4-IT1 was up-regulated in MCF-7 CSCs compared with MCF-7 cells, and high SPRY4-IT1 expression was related to reduced breast cancer patient survival. Furthermore, SPRY4-IT1 overexpression promoted breast cancer cell proliferation and stemness in vitro and in vivo. In addition, SPRY4-IT1 knockdown suppressed BCSC renewal ability and stemness maintenance in vivo and in vitro. The dual-luciferase reporter assays indicated that SPRY4-IT1 as a sponge for miR-6882-3p repressed transcription factor 7-like 2 (TCF7L2) expression. Taken together, these findings demonstrated that SPRY4-IT1 promotes proliferation and stemness of breast cancer cells as well as renewal ability and stemness maintenance of BCSCs by increasing the expression of TCF7L2 through targeting miR-6882-3p.

Prognostic relevance of miR-137 and its liver microenvironment regulatory target gene AFM in hepatocellular carcinoma.

MiR-137 has been identified as potential hepatocellular carcinoma (HCC) prognostic biomarkers. Highly relevant HCC prognostic biomarkers may be derived from combinations of miR-137 with its target genes involved in the regulation of liver microenvironment. This study aimed at the discovery of such a combination with improved HCC prognosis performance than miR-137 or its target gene alone in a significantly higher number of HCC patients than previous studies. Analysis of the differentially expressed micro RNAs (miRNAs) between cancer and noncancer tissues reconfirmed miR-137 to be among the most relevant prognostic miRNAs and the data of 375 HCC patients and 50 normal cases were from the Cancer Genome Atlas (TCGA) data sets. Target genes were identified by the established search methods and Kaplan-Meier survival analysis of HCC patients was used to evaluate the overall survival (OS) and recurrence-free survival (RFS). Cox proportional hazards regression indicated that the miR-137 and its target gene AFM combination is an independent prognostic factor for the OS and RFS in HCC. In vitro experiments validated that miR-137 could bind to 3'-untranslated region of the AFM and promote the invasion and metastasis of HCC cell lines. The expressions of miR-137 and its liver microenvironment regulatory target gene AFM in combination significantly correlated with HCC progression in a higher number of patients than in previous studies, which suggested their potential as prognostic biomarkers for HCC.

Sandwich-Structured Upconversion Nanoprobes Coated with a Thin Silica Layer for Mitochondria-Targeted Cooperative Photodynamic Therapy for Solid Malignant Tumors.

Upconversion nanoparticles-based photodynamic nanotheranostic agents (UCNPs-PDT) have received great interest due to improved tissue penetration, weak autofluorescence, and low biotoxicity. However, conventional UCNPs-PDT are often limited by low energy transfer efficiency from UCNPs to photosensitizer (PS) molecules and insufficient generation and limited diffusion distance of reactive oxygen species (ROSs). Herein, an "all in one" nanotheranostic agent has been developed which has multicolor sandwich-structured UCNPs (SWUCNPs) as the core, a thin silica layer with a mitochondria-targeted group for loading dual PS as the medium layer, and polyethylene glycol-folic acid (PEG-FA) chains as the outer layer. Multicolor SWUCNPs simultaneously achieve two-photon fluorescence imaging and serve as energy donor for dual PS molecules. The thin luminescence layer and silica layer control most UCNPs activators and PS molecules in the effective energy transfer distance to guarantee a high energy transfer efficiency. Via FA-mediated endocytosis, the nanotheranostic agent is selectively endocytosed by cancer cells, is released from the endosome/lysosome, targets the mitochondria, and in situ produces ROSs under excitation from NIR, leading to significant mitochondria-mediated cell apoptosis. Furthermore, the established nanotheranostic agent shows tumor targetability, increased generation of ROSs, high PDT efficacy, significant cell apoptosis, minimal systemic cytotoxicity, and efficacious in vivo tumor inhibition.

Clinical implications of a novel prognostic factor AIFM3 in breast cancer patients.

BACKGROUND: In a time of increasing concerns over personalized and precision treatment in breast cancer (BC), filtering prognostic factors attracts more attention. Apoptosis-Inducing Factor Mitochondrion-associated 3 (AIFM3) is widely expressed in various tissues and aberrantly expressed in several cancers. However, clinical implication of AIFM3 has not been reported in BC. The aim of the study is to investigate the crystal structure, clinical and prognostic implications of AIFM3 in BC. METHODS: AIFM3 expression in 151 BC samples were assessed by immunohistochemistry (IHC). The Cancer Genome Atlas (TCGA) and Kaplan-Meier survival analysis were used to demonstrate expression and survival of AIFM3 signature. Gene Set Enrichment Analysis (GSEA) was performed to investigate the mechanisms related to AIFM3 expression in BC. RESULTS: AIFM3 was significantly more expressed in breast cancer tissues than in normal tissues. AIFM3 expression had a significant association with tumor size, lymph node metastasis, TNM stage and molecular typing. Higher AIFM3 expression was related to a shorter overall survival (OS) and disease-free survival (DFS). Lymph node metastasis and TNM stage were independent factors of AIFM3 expression. The study presented the crystal structure of AIFM3 successfully and predicted several binding sites when AIFM3 bonded to PTPN12 by Molecular Operating Environment software (MOE). CONCLUSIONS: AIFM3 might be a potential biomarker for predicting prognosis in BC, adding to growing evidence that AIFM3 might interact with PTPN12.

The effect of di-2-ethylhexyl phthalate on inflammation and lipid metabolic disorder in rats.

BACKGROUND: Plasticizer di-2-ethylhexyl phthalate (DEHP) can induce lipid metabolic disorder. There was a chronic low level inflammatory response in adipose tissue of patients with lipid metabolic disorder. But the effect of inflammation on lipid metabolic disorder induced by DEHP is unclear. The present study was undertaken to explore the effect of di-2-ethylhexyl phthalate on inflammation and lipid metabolic disorder in rats. METHODS: Eighty healthy 21-day-old Wistar rats were randomly divided into 4 groups and administered DEHP by gavage at 0, 5, 50, and 500 mg/kg/ d for 8 weeks. Morphological changes of adipose tissue, the levels of IL-1ß, TNF-α, LEP, and ADP in rat serum and adipose tissue, the serum TC, TG, HDL-C and LDL-C, the mRNA and protein expression levels of lipid metabolism-related gene CEBP/ß and inflammation-related gene CD68 were measured. RESULTS: After exposure to DEHP, the weight of rats in the high dose group was significantly higher than that in the control group (p < 0.05). And the number of adipose tissue cells in the medium-dose and high-dose DEHP groups increased, with much more macrophage infiltrated. The levels of LDL-C, HDL-C, TC in serum and LEP in adipose tissue of rats exposed to 500 mg/kg DEHP were significantly higher than those in the control group (p < 0.05); while the level of ADP in adipose tissue in rats exposed to DEHP was significantly lower (p < 0.05). The levels of IL-1ß and TNF-α in surum and adipose tissue of rats exposed to DEHP were significantly higher than those in the control group (p < 0.05). The mRNA and protein expression levels of CEBP/ß and CD68 in adipose tissue of rats exposed to DEHP were significantly higher than those in the control group. The TC, LEP and ADP Levels of rats were significantly different among different subgroup of IL-1ß and TNF-α, and in high level subgroup, the TC, LEP and ADP Levels were increased. The levels of TC and LEP was increased in high level subgroup of CD68. CONCLUSION: DEHP induced more macrophage infiltrated in adipose tissue of rats, promoted the secretion of IL-1ß, TNF-α and the formation of inflammation, and disturbed the normal lipid metabolism and lead to lipid metabolic disorders. What is more, the levels of inflammation were associated with the lipid levels.

Urinary levels of phthalate metabolites and their association with lifestyle behaviors in Chinese adolescents and young adults.

BACKGROUND: Adolescence and young adulthood are critical periods of human growth and development. Phthalates are environmental endocrine disruptors, and their health hazards in adolescents and young adults cannot be ignored. This study was undertaken to assess phthalate exposure and determine the associations between lifestyle behaviors and phthalate metabolite levels in Chinese adolescents and young adults. METHODS: Four hundred and seventy-eight adolescents and young adults aged 16-20 years were included in this study. The levels of mono-ethyl phthalate (MEP), mono-butyl phthalate (MBP), mono-(2-ethylhexyl) phthalate (MEHP), mono-(2-ethyl-5-oxohexyl) phthalate (MEOHP), mono-(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP), mono-(2-ethyl-5-carboxypentyl) phthalate (MECPP) and mono-(2-carboxmethyl)-hexyl phthalate (MCMHP) in the subjects' urine were measured by high-performance liquid chromatography-tandem mass spectrometry. The estimated daily intake (EDI) and hazard index (HI) of phthalates were calculated based on urinary metabolite levels. Relevant information on the subjects was collected via questionnaires. The associations between phthalate metabolite levels and lifestyle behaviors were examined using the independent-sample t-test, Mann-Whitney test and multiple linear regression. RESULTS: In this study, the detection rates of all seven metabolites were >98%. The highest median metabolite concentration was MBP, which was 43.00 µg/L (33.11 µg/g creatinine). The highest median EDI was for di-(2-ethylhexyl) phthalate (DEHP), which was 2.40 µg/kg-bw/day (volume-based) and 1.51 µg/kg-bw/day (creatinine-based). 2.7% (volume-based) and 1.0% (creatinine-based) of the subjects showed excessive HITDI (HI of the tolerable daily intake) values, which indicated the cumulative risk of anti-androgenic effects. Furthermore, factors significantly associated with phthalate metabolite levels included the use of plastic food packages (DEHP metabolites), physical exercise (MEOHP), the frequency of fast food consumption (MBP), and the frequency of skin care cosmetics and color cosmetics use (MEP). CONCLUSION: Our results suggest that Chinese adolescents and young adults are widely exposed to phthalates and their metabolite levels are influenced by lifestyle behaviors.

Multicolor Upconversion Nanoprobes Based on a Dual Luminescence Resonance Energy Transfer Assay for Simultaneous Detection and Bioimaging of [Ca2+ ]i and pHi in Living Cells.

Intracellular [Ca2+ ]i and pHi have a close relationship, and their abnormal levels can result in cell dysfunction and accompanying diseases. Thus, simultaneous determination of [Ca2+ ]i and pHi can more accurately investigate complex biological processes in an integrated platform. Herein, multicolor upconversion nanoparticles (UCNPs) were prepared with the advantages of no spectral overlapping, single NIR excitation wavelengths, and greater tissue penetration depth. The upconversion nanoprobes were easily prepared by the attachment of two fluorescent dyes, Fluo-4 and SNARF-4F. Based on the dual luminescence resonance energy transfer (LRET) process, the blue and green fluorescence of the UCNPs were specially quenched and selectively recovered after the detachment and/or absorbance change of the attached fluorescent dyes, enabling dual detection. Importantly, the developed nanoprobe could successfully be applied for the detection of [Ca2+ ]i and pHi change in adenosine triphosphate (ATP) and ethylene glycol tetraacetic acid (EGTA) stimulation in living cells.

Dual-Activator Codoped Upconversion Nanoprobe with Core-Multishell Structure for in Vitro and in Vivo Detection of Hydroxyl Radical.

Monitoring the fluctuation of hydroxyl radical (·OH) in the body can serve as an effective tool for the prediction of relative diseases; however, it is highly challenging due to the radical's short lifetime, high reactivity, and extremely low concentration. Sandwich structured lanthanide-doped upconversion nanoparticles (UCNPs) exhibit unique luminescence properties and great prospects in bioimaging. Nonetheless, their rather low luminescence efficiency and intensity are serious limitations for their application. Herein, we report on dual-activator codoped UCNPs with a core-multishell structure that greatly improve the luminescence intensity and lifetime by 46-fold and 2.6-fold, respectively, compared to those of the monoactivator doped sandwich structured UCNPs. Moreover, emitting ions in the designed core-multishell (CMS)-UCNPs were confined in a homogeneous and thin shell layer (∼2 nm); thus, the luminescence resonance energy transfer (LRET)-based CMS-UCNPs@azo dye nanoprobe exhibited a largely shortened energy transfer distance and a pronounced luminescence quenching yield (97%), affording the nearly zero background signal and achieving an ultrahigh sensitivity for the detection of ·OH (with limit of quantitation (LOQ) of 0.10 fM). With good biocompatibility, low biotoxicity, and enhanced luminescence intensity and lifetime, the developed nanoprobe was competent in monitoring the subtle fluctuation of ·OH concentration both in vitro and in vivo.

Prevalence and molecular characterization of Staphylococcus aureus isolated from goats in Chongqing, China.

BACKGROUND: Staphylococcus aureus is an important zoonotic pathogen which not only causes significant economic loss in livestock production but also poses a potential threat to public health. Compared with bovine and swine, the information on the colonization of S. aureus in goats is very limited. To understand the prevalence and characteristics of S. aureus in goats, we used the nasal swabs collected from apparently healthy goats to isolate S. aureus, and tested their antimicrobial susceptibility, virulence gene carrying levels, and multilocus sequence typing (MLST). RESULTS: In 74 nasal swabs of apparently healthy goats, 32 (43.24%) S. aureus strains were isolated and identified, most of which were susceptible to many antibiotics, except for trimethoprim, furazolidone, amoxicillin, lincomycin and roxithromycin, and the resistance incidence of which were 50%, 40.63%, 37.5%, 28.13%, and 21.88% respectively. All the isolates were methicillin-susceptible S. aureus (MSSA) and mecA-negative. Enterotoxin genes were found in 53.13% of the strains. Of which, sej was the most prevalent (21.88%), followed by seb, sec, and see with the same level (18.75%). The most prevalent combination were seb + see and seb + tst. None of the S. aureus isolates harbored sea, sed, seh, eta and etb. Multilocus sequence typing (MLST) revealed 6 new alleles (aroe-552, aroe-553, glpf-500, pta-440, yqil-482 and yqil-496) and 5 new sequence types (STs) (3431,3440,3444,3445 and 3461). Using eBURST, the 5 STs were assigned to clonal complex 522 (CC522) and a further CC with no predicted ancestor. Phylogenetic analysis of seven concatenated MLST alleles revealed that the 5 STs were grouped into cluster I composed of S. aureus mainly from goats and sheep. CONCLUSION: We provide the data for prevalence of S. aureus in goats in Chongqing municipality and their characterization which will help in tracking evolution of epidemic strains and their control methods.