Association and Interaction between Heavy Metals and Hyperuricemia in a Taiwanese Population.

The prevalence of hyperuricemia in Taiwan is high, and hyperuricemia has been associated with a risk of developing several diseases. Although the traditional risk factors for hyperuricemia are well known, the relationship between heavy metals and hyperuricemia is still undefined. Therefore, the aim of this study was to investigate the relationship between hyperuricemia and heavy metals. A total of 2447 participants (977 males and 1470 females) residing in southern Taiwan were enrolled, and levels of the following heavy metals were measured: lead in blood, and nickel, chromium, manganese, arsenic (As), copper, and cadmium in urine. Hyperuricemia was defined as a serum uric acid level greater than 7.0 mg/dL (416.5 µmol/L) in men and 6.0 mg/dL (357 µmol/L) in women. The participants were divided into two groups: those without hyperuricemia (n = 1821; 74.4%) and those with hyperuricemia (n = 626; 25.6%). Multivariate analysis showed that only high urine As (log per 1 µg/g creatinine; odds ratio, 1.965; 95% confidence interval, 1.449 to 2.664; p < 0.001), young age, male sex, high body mass index, high hemoglobin, high triglycerides, and low estimated glomerular filtration rate were significantly associated with hyperuricemia. In addition, the interactions between Pb × Cd (p = 0.010), Ni × Cu (p = 0.002), and Cr × Cd (p = 0.001) on hyperuricemia were statistically significant. Increasing levels of Pb and Cr yielded an increased prevalence of hyperuricemia, and the effect was progressively greater for increasing Cd. Moreover, increasing levels of Ni yielded an increased prevalence of hyperuricemia, and the effect was progressively greater for increasing Cu. In conclusion, our results show that high urine As is associated with hyperuricemia, and some interactions of heavy metals on hyperuricemia are noted. We also found that young age, male sex, high BMI, high hemoglobin, high triglycerides, and low eGFR were significantly associated with hyperuricemia.

IKZF3 amplification frequently occurs in HER2-positive breast cancer and is a potential therapeutic target.

Breast cancer is one of the leading causes of cancer death in women, and although treatment outcome has substantially improved in the past decades, advanced or metastatic breast cancers still carry a poor prognosis. Gene amplification is one of the frequent genetic alterations in cancer, and oncogene amplification may be associated with cancer aggressiveness and oncogenicity. Targeting amplified genes such as HER2 has vastly improved disease outcome and survival, and anti-HER2 therapeutics have revolutionized the standard of care in HER2 breast cancer. Besides currently known druggable gene amplifications including ERBB2 and FGFR2, other frequently amplified genes are relatively less well known for function and clinical significance. By querying four large databases from TCGA and AACR-Genie, from a total of 11,890 patients with invasive ductal breast carcinoma, we discover IKZF3, CCND1, ERBB2 to be consistently amplified across different cohorts. We further identify IKZF3 as a frequently amplified gene in breast cancer with a prevalence of 12-15% amplification rate. Interestingly, IKZF3 amplification is frequently co-amplified with ERBB2/HER2, and is also associated with worse prognosis compared to IKZF3 non-amplified cancers. Analysis of HER2 breast cancer patients treated with trastuzumab revealed decrease in both ERBB2/HER2 and IKZF3 expression. Further investigation using the DepMap for gene dependency by genome-wide CRISPR screening revealed dependence on IKZF3 in HER2 breast cancer cell lines. Our study utilized an integrative analysis of large-scale patient genomics, transcriptomics and clinical data to reveal IKZF3 as a frequently amplified gene, and suggest a potential role of IKZF3 as a druggable target for HER2 breast cancer.

CDK4/6 inhibitors downregulate the ubiquitin-conjugating enzymes UBE2C/S/T involved in the ubiquitin-proteasome pathway in ER + breast cancer.

PURPOSE: Despite significant improvement in therapeutic development in the past decades, breast cancer remains a formidable cause of death for women worldwide. The hormone positive subtype (HR(+)) (also known as luminal type) is the most prevalant category of breast cancer, comprising ~70% of patients. The clinical success of the three CDK4/6 inhibitors palbociclib, ribociclib, and abemaciclib has revolutionized the treatment of choice for metastatic HR(+) breast cancer. Accumulating evidence demonstrate that the properties of CDK4/6 inhibitors extend beyond inhibition of the cell cycle, including modulation of immune function, sensitizing PI3K inhibitors, metabolism reprogramming, kinome rewiring, modulation of the proteosome, and many others. The ubiquitin-proteasome pathway (UPP) is a crucial cellular proteolytic system that maintains the homeostasis and turnover of proteins. METHODS: We performed transcriptional profiling of the HR(+) breast cancer cell lines MCF7 and T47D treated with Palbociclib. Differential expressed genes were analyzed for novel pathways enriched. The results were further validated with biochemical assays and with real world clinical database cohorts. RESULTS: We uncovered a novel mechanism that demonstrate the CDK4/6 inhibitors suppress the expression of three ubiquitin conjugating enzymes UBE2C, UBE2S, UBE2T. Further validation in the HR(+) cell lines show that Palbociclib and ribociclib decrease UBE2C at both the mRNA and protein level, but this phenomenon was not shared with abemaciclib. These three E2 enzymes modulate several E3 ubiquitin ligases, including the APC/C complex which plays a role in G1/S progression. We further demonstrate the UBE2C/UBE2T expression levels are associated with breast cancer survival, and HR(+) breast cancer cells demonstrate dependence on the UBE2C. CONCLUSIONS: Our study suggests a novel link between CDK4/6 inhibitor and UPP pathway, adding to the potential mechanisms of their clinical efficacy in cancer.

Sorafenib is an antagonist of the aryl hydrocarbon receptor.

Sorafenib is an orally administered inhibitor of several tyrosine protein kinases. Treatment with sorafenib induces autophagy, which may suppress the growth of hepatocellular carcinoma (HCC) and other cancers. Aryl hydrocarbon receptor (AhR) is activated by xenbiotics and is involved in detoxification, but also plays other physiological roles. The following results were obtained. ITE and ß-NF are endogenous and synthetic AhR ligands, respectively. One µM sorafenib can strongly suppress baseline as well as 0.5 µM ITE- and 1 µM ß-NF-induced transcriptional activity of the aryl hydrocarbon response element (AHRE) in both human and mouse cells. Cytochrome p450 (CYP) 1A1 is mainly transcribed by activated AhR. Sorafenib (2-15 µM) strongly and dose-dependently suppressed baseline as well as 2 µM ITE- and 10 µM ß-NF-induced CYP1A1 mRNA and protein expression. Ligand-activated AhR translocates from the cytoplasm to the nucleus. While sorafenib was found to suppress AhR activity, the drug alone was able to induce AhR translocation into the nucleus. Sorafenib's antagonistic action on AhR was comparable to that of the known AhR antagonist CH-223191 in human liver and ovarian cell lines. In summary, we demonstrate that sorafenib is a potent AhR antagonist and likely endocrine disruptor of the AhR. Moreover, sorafenib offers potential benefit for diseases treatable through AhR suppression strategies. Further investigation is warranted into sorafenib's AhR antagonistic behavior.

The human telomeric proteome during telomere replication.

Telomere shortening can cause detrimental diseases and contribute to aging. It occurs due to the end replication problem in cells lacking telomerase. Furthermore, recent studies revealed that telomere shortening can be attributed to difficulties of the semi-conservative DNA replication machinery to replicate the bulk of telomeric DNA repeats. To investigate telomere replication in a comprehensive manner, we develop QTIP-iPOND - Quantitative Telomeric chromatin Isolation Protocol followed by isolation of Proteins On Nascent DNA - which enables purification of proteins that associate with telomeres specifically during replication. In addition to the core replisome, we identify a large number of proteins that specifically associate with telomere replication forks. Depletion of several of these proteins induces telomere fragility validating their importance for telomere replication. We also find that at telomere replication forks the single strand telomere binding protein POT1 is depleted, whereas histone H1 is enriched. Our work reveals the dynamic changes of the telomeric proteome during replication, providing a valuable resource of telomere replication proteins. To our knowledge, this is the first study that examines the replisome at a specific region of the genome.

Cyproterone acetate acts as a disruptor of the aryl hydrocarbon receptor.

Prostate cancer is a major cause of death in males. Cyproterone acetate (CPA), the steroidal anti-androgen for part of androgen deprivation therapy, may block the androgen-receptor interaction and then reduce serum testosterone through its weak anti-gonadotropic action. In addition to CPA inducing hepatitis, CPA is known to cause liver tumors in rats also. Aryl hydrocarbon receptor (AhR) is a cytoplasmic receptor and regulates multiple physiological functions. CYP1A1 is an AhR-targeted gene. We found that CPA induced CYP1A1 expression, transcriptional activity of the aryl hydrocarbon response element (AHRE), and the nuclear localization of AhR in mouse Hepa-1c1c7 cells. However, CPA suppressed CYP1A1 mRNA expression and the transcriptional activity of AHRE in human HepG2 and MCF7 cells, and also decreased AhR ligand-induced CYP1A1 protein expression and transcriptional activity of AHRE in HepG2 cells. In summary, CPA is an AhR agonist in mouse cells, but an AhR antagonist in human cells. Accordingly, CPA potentially plays a role as an endocrine disruptor of the AhR. This study helps us to understand why CPA induces acute hepatitis, gene mutation, and many other side effects. In addition, it may trigger further studies investigating the relationships between CPA, glucocorticoid receptor and castration-resistant prostate cancer in the future.

Cytoskeleton stiffness regulates cellular senescence and innate immune response in Hutchinson-Gilford Progeria Syndrome.

Hutchinson-Gilford progeria syndrome (HGPS) is caused by the accumulation of mutant prelamin A (progerin) in the nuclear lamina, resulting in increased nuclear stiffness and abnormal nuclear architecture. Nuclear mechanics are tightly coupled to cytoskeletal mechanics via lamin A/C. However, the role of cytoskeletal/nuclear mechanical properties in mediating cellular senescence and the relationship between cytoskeletal stiffness, nuclear abnormalities, and senescent phenotypes remain largely unknown. Here, using muscle-derived mesenchymal stromal/stem cells (MSCs) from the Zmpste24-/- (Z24-/- ) mouse (a model for HGPS) and human HGPS fibroblasts, we investigated the mechanical mechanism of progerin-induced cellular senescence, involving the role and interaction of mechanical sensors RhoA and Sun1/2 in regulating F-actin cytoskeleton stiffness, nuclear blebbing, micronuclei formation, and the innate immune response. We observed that increased cytoskeletal stiffness and RhoA activation in progeria cells were directly coupled with increased nuclear blebbing, Sun2 expression, and micronuclei-induced cGAS-Sting activation, part of the innate immune response. Expression of constitutively active RhoA promoted, while the inhibition of RhoA/ROCK reduced cytoskeletal stiffness, Sun2 expression, the innate immune response, and cellular senescence. Silencing of Sun2 expression by siRNA also repressed RhoA activation, cytoskeletal stiffness and cellular senescence. Treatment of Zmpste24-/- mice with a RhoA inhibitor repressed cellular senescence and improved muscle regeneration. These results reveal novel mechanical roles and correlation of cytoskeletal/nuclear stiffness, RhoA, Sun2, and the innate immune response in promoting aging and cellular senescence in HGPS progeria.

Setting boundaries for genome-wide heterochromatic DNA deletions through flanking inverted repeats in Tetrahymena thermophila.

Eukaryotic cells pack their genomic DNA into euchromatin and heterochromatin. Boundaries between these domains have been shown to be set by boundary elements. In Tetrahymena, heterochromatin domains are targeted for deletion from the somatic nuclei through a sophisticated programmed DNA rearrangement mechanism, resulting in the elimination of 34% of the germline genome in ∼10,000 dispersed segments. Here we showed that most of these deletions occur consistently with very limited variations in their boundaries among inbred lines. We identified several potential flanking regulatory sequences, each associated with a subset of deletions, using a genome-wide motif finding approach. These flanking sequences are inverted repeats with the copies located at nearly identical distances from the opposite ends of the deleted regions, suggesting potential roles in boundary determination. By removing and testing two such inverted repeats in vivo, we found that the ability for boundary maintenance of the associated deletion were lost. Furthermore, we analyzed the deletion boundaries in mutants of a known boundary-determining protein, Lia3p and found that the subset of deletions that are affected by LIA3 knockout contained common features of flanking regulatory sequences. This study suggests a common mechanism for setting deletion boundaries by flanking inverted repeats in Tetrahymena thermophila.

Markers of Accelerated Skeletal Muscle Regenerative Response in Murphy Roths Large Mice: Characteristics of Muscle Progenitor Cells and Circulating Factors.

The "super-healing" Murphy Roths Large (MRL/MpJ) mouse possesses a superior regenerative capacity for repair of many tissues, which makes it an excellent animal model for studying molecular and cellular mechanisms during tissue regeneration. As the role of muscle progenitor cells (MPCs) in muscle-healing capacity of MRL/MpJ mice has not been previously studied, we investigated the muscle regenerative capacity of MRL/MpJ mice following muscle injury, and the results were compared to results from C57BL/6J (B6) age-matched control mice. Our results show that muscle healing upon cardiotoxin injury was accelerated in MRL/MpJ mice and characterized by reduced necrotic muscle area, reduced macrophage infiltration, and more regenerated myofibers (embryonic myosin heavy chain+/centronucleated fibers) at 3, 5, and 12 days postinjury, when compared to B6 age-matched control mice. These observations were associated with enhanced function of MPCs, including improved cell proliferation, differentiation, and resistance to stress, as well as increased muscle regenerative potential when compared to B6 MPCs. Mass spectrometry of serum proteins revealed higher levels of circulating antioxidants in MRL/MpJ mice when compared to B6 mice. Indeed, we found relatively higher gene expression of superoxide dismutase 1 (Sod1) and catalase (Cat) in MRL/MpJ MPCs. Depletion of Sod1 or Cat by small interfering RNA impaired myogenic potential of MRL/MpJ MPCs, indicating a role for these antioxidants in muscle repair. Taken together, these findings provide evidence that improved function of MPCs and higher levels of circulating antioxidants play important roles in accelerating muscle-healing capacity of MRL/MpJ mice. Stem Cells 2019;37:357-367.

Lymphotoxin-ß Interacts with Methylated EGFR to Mediate Acquired Resistance to Cetuximab in Head and Neck Cancer.

Purpose: In head and neck squamous cell carcinoma (HNSCC), the incidence of RAS mutation, which is the major cause of cetuximab resistance, is relatively rare compared with the other types of cancers, and the mechanism mediating acquired resistance is unclear compared with the driver gene mutation-mediated de novo resistance. Here, we investigated the driver gene mutation-independent mechanism for cetuximab resistance in HNSCC.Experimental Design: We used the in vitro-selected and in vivo-selected cetuximab-resistant sublines of HNSCC cell lines for investigating the mechanism of acquired resistance to cetuximab. Zebrafish model was applied for evaluating the synergistic effect of combinatory drugs for overcoming cetuximab resistance.Results: The cetuximab-resistant HNSCC cells undergo a Snail-induced epithelial-mesenchymal transition. Mechanistically, Snail induces the expression of lymphotoxin-ß (LTß), a TNF superfamily protein that activates NF-κB, and protein arginine methyltransferase 1 (PRMT1), an arginine methyltransferase that methylates EGFR. LTß interacts with methylated EGFR to promote its ligand-binding ability and dimerization. Furthermore, LTß activates the NF-κB pathway through a LTß receptor-independent mechanism. Combination of an EGFR tyrosine kinase inhibitor and a NF-κB inhibitor effectively suppressed cetuximab-resistant HNSCC and interfering with the EGFR-LTß interaction reverses resistance.Conclusions: Our findings elucidate the mechanism of driver gene mutations-independent mechanism of acquired resistance to cetuximab in HNSCC and also provide potential strategies for combating cetuximab resistance. Clin Cancer Res; 23(15); 4388-401. ©2017 AACR.

Programmed Minichromosome Elimination as a Mechanism for Somatic Genome Reduction in Tetrahymena thermophila.

The maintenance of chromosome integrity is crucial for genetic stability. However, programmed chromosome fragmentations are known to occur in many organisms, and in the ciliate Tetrahymena the five germline chromosomes are fragmented into hundreds of minichromosomes during somatic nuclear differentiation. Here, we showed that there are different fates of these minichromosomes after chromosome breakage. Among the 326 somatic minichromosomes identified using genomic data, 50 are selectively eliminated from the mature somatic genome. Interestingly, many and probably most of these minichromosomes are eliminated during the growth period between 6 and 20 doublings right after conjugation. Genes with potential conjugation-specific functions are found in these minichromosomes. This study revealed a new mode of programmed DNA elimination in ciliates similar to those observed in parasitic nematodes, which could play a role in developmental gene regulation.

Activation of aryl hydrocarbon receptor reduces carbendazim-induced cell death.

Carbendazim inhibits microtubule assembly, thus blocking mitosis and inhibiting cancer cell proliferation. Accordingly, carbendazim is being explored as an anticancer drug. Data show that carbendazim increased mRNA and protein expressions and promoter activity of CYP1A1. In addition, carbendazim activated transcriptional activity of the aryl hydrocarbon response element, and induced nuclear translocation of the aryl hydrocarbon receptor (AhR), a sign the AhR is activated. Carbendazim-induced CYP1A1 expression was blocked by AhR antagonists, and was abolished in AhR signal-deficient cells. Results demonstrated that carbendazim activated the AhR, thereby stimulating CYP1A1 expression. In order to understand whether AhR-induced metabolic enzymes turn carbendazim into less-toxic metabolites, Hoechst 33342 staining to reveal carbendazim-induced nuclear changes and flow cytometry to reveal the subG0/G1 population were applied to monitor carbendazim-induced cell apoptosis. Carbendazim induced less apoptosis in Hepa-1c1c7 cells than in AhR signal-deficient Hepa-1c1c7 mutant cells. Pretreatment with ß-NF, an AhR agonist that highly induces CYP1A1 expression, decreased carbendazim-induced cell death. In addition, the lower the level of AhR was, the lower the vitality present in carbendazim-treated cells, including hepatoma cells and their derivatives with AhR RNA interference, also embryonic kidney cells, bladder carcinoma cells, and AhR signal-deficient Hepa-1c1c7 cells. In summary, carbendazim is an AhR agonist. The toxicity of carbendazim was lower in cells with the AhR signal. This report provides clues indicating that carbendazim is more potent at inducing cell death in tissues without than in those with the AhR signal, an important reference for applying carbendazim in cancer chemotherapy.

The piggyBac transposon-derived genes TPB1 and TPB6 mediate essential transposon-like excision during the developmental rearrangement of key genes in Tetrahymena thermophila.

Ciliated protozoans perform extreme forms of programmed somatic DNA rearrangement during development. The model ciliate Tetrahymena thermophila removes 34% of its germline micronuclear genome from somatic macronuclei by excising thousands of internal eliminated sequences (IESs), a process that shares features with transposon excision. Indeed, piggyBac transposon-derived genes are necessary for genome-wide IES excision in both Tetrahymena (TPB2 [Tetrahymena piggyBac-like 2] and LIA5) and Paramecium tetraurelia (PiggyMac). T. thermophila has at least three other piggyBac-derived genes: TPB1, TPB6, and TPB7 Here, we show that TPB1 and TPB6 excise a small, distinct set of 12 unusual IESs that disrupt exons. TPB1-deficient cells complete mating, but their progeny exhibit slow growth, giant vacuoles, and osmotic shock sensitivity due to retention of an IES in the vacuolar gene DOP1 (Dopey domain-containing protein). Unlike most IESs, TPB1-dependent IESs have piggyBac-like terminal inverted motifs that are necessary for excision. Transposon-like excision mediated by TPB1 and TPB6 provides direct evidence for a transposon origin of not only IES excision machinery but also IESs themselves. Our study highlights a division of labor among ciliate piggyBac-derived genes, which carry out mutually exclusive categories of excision events mediated by either transposon-like features or RNA-directed heterochromatin.

The Copper Balance of Cities: Exploratory Insights into a European and an Asian City.

Material management faces a dual challenge: on the one hand satisfying large and increasing demands for goods and on the other hand accommodating wastes and emissions in sinks. Hence, the characterization of material flows and stocks is relevant for both improving resource efficiency and environmental protection. This article focuses on the urban scale, a dimension rarely investigated in past metal flow studies. We compare the copper (Cu) metabolism of two cities in different economic states, namely, Vienna (Europe) and Taipei (Asia). Substance flow analysis is used to calculate urban Cu balances in a comprehensive and transparent form. The main difference between Cu in the two cities appears to be the stock: Vienna seems close to saturation with 180 kilograms per capita (kg/cap) and a growth rate of 2% per year. In contrast, the Taipei stock of 30 kg/cap grows rapidly by 26% per year. Even though most Cu is recycled in both cities, bottom ash from municipal solid waste incineration represents an unused Cu potential accounting for 1% to 5% of annual demand. Nonpoint emissions are predominant; up to 50% of the loadings into the sewer system are from nonpoint sources. The results of this research are instrumental for the design of the Cu metabolism in each city. The outcomes serve as a base for identification and recovery of recyclables as well as for directing nonrecyclables to appropriate sinks, avoiding sensitive environmental pathways. The methodology applied is well suited for city benchmarking if sufficient data are available.

Electroosmotic flow velocity measurements in a square microchannel.

Experiments were performed using a microparticle image velocimetry (MPIV) for 2D velocity distributions of electroosmotically driven flows in a 40-mm-long microchannel with a square cross section of 200×200 µm. Electroosmotic flow (EOF) bulk fluid velocity measurements were made in a range of streamwise electric field strengths from 5 to 25 kV/m. A series of seed particle calibration tests can be made in a 200×120×24,000-µm untreated polydimethyl siloxane (PDMS channel incorporating MPIV to determine the electrophoretic mobilities in aqueous buffer solutions of 1× TAE, 1× TBE, 10 mM NaCl, and 10 mM borate. A linear/nonlinear (due to Joule heating) flow rate increase with applied field was obtained and compared with those of previous studies. A parametric study, with extensive measurements, was performed with different electric field strength and buffer solution concentration under a constant zeta potential at wall for each buffer. The characteristics of EOF in square microchannels were thus investigated. Finally, a composite correlation of the relevant parameters was developed in the form of [Formula: see text] within ±1% accuracy for 99% of the experimental data.