ATPase Copper Transporting Beta (ATP7B) Is a Novel Target for Improving the Therapeutic Efficacy of Docetaxel by Disulfiram/Copper in Human Prostate Cancer.

Docetaxel has been the standard first-line chemotherapy for lethal metastatic prostate cancer (mPCa) since 2004, but resistance to docetaxel treatment is common. The molecular mechanisms of docetaxel resistance remain largely unknown and could be amenable to interventions that mitigate resistance. We have recently discovered that several docetaxel-resistant mPCa cell lines exhibit lower uptake of cellular copper and uniquely express higher levels of a copper exporter protein ATP7B. Knockdown of ATP7B by silencing RNAs (siRNA) sensitized docetaxel-resistant mPCa cells to the growth-inhibitory and apoptotic effects of docetaxel. Importantly, deletions of ATP7B in human mPCa tissues predict significantly better survival of patients after their first chemotherapy than those with wild-type ATP7B (P = 0.0006). In addition, disulfiram (DSF), an FDA-approved drug for the treatment of alcohol dependence, in combination with copper, significantly enhanced the in vivo antitumor effects of docetaxel in a docetaxel-resistant xenograft tumor model. Our analyses also revealed that DSF and copper engaged with ATP7B to decrease protein levels of COMM domain-containing protein 1 (COMMD1), S-phase kinase-associated protein 2 (Skp2), and clusterin and markedly increase protein expression of cyclin-dependent kinase inhibitor 1 (p21/WAF1). Taken together, our results indicate a copper-dependent nutrient vulnerability through ATP7B exporter in docetaxel-resistant prostate cancer for improving the therapeutic efficacy of docetaxel.

Water-soluble chromenylium dyes for shortwave infrared imaging in mice.

In vivo imaging using shortwave infrared light (SWIR, 1000-2000 nm) benefits from deeper penetration and higher resolution compared to using visible and near-infrared wavelengths. However, the development of biocompatible SWIR contrast agents remains challenging. Despite recent advancements, small molecule SWIR fluorophores are often hindered by their significant hydrophobicity. We report a platform for generating a panel of soluble and functional dyes for SWIR imaging by late-stage functionalization of a versatile fluorophore intermediate, affording water-soluble dyes with bright SWIR fluorescence in serum. Specifically, a tetra-sulfonate derivative enables clear video-rate imaging of vasculature with only 0.05 nmol dye, and a tetra-ammonium dye shows strong cellular retention for tracking of tumor growth. Additionally, incorporation of phosphonate functionality enables imaging of bone in awake mice. This modular design provides insights for facile derivatization of existing SWIR fluorophores to introduce both solubility and bioactivity towards in vivo bioimaging.

The ionophore thiomaltol induces rapid lysosomal accumulation of copper and apoptosis in melanoma.

In this report, we investigate the toxicity of the ionophore thiomaltol (Htma) and Cu salts to melanoma. Divalent metal complexes of thiomaltol display toxicity against A375 melanoma cell culture resulting in a distinct apoptotic response at submicromolar concentrations, with toxicity of Cu(tma)2 > Zn(tma)2 >> Ni(tma)2. In metal-chelated media, Htma treatment shows little toxicity, but the combination with supplemental CuCl2, termed Cu/Htma treatment, results in toxicity that increases with suprastoichiometric concentrations of CuCl2 and correlates with the accumulation of intracellular copper. Electron microscopy and confocal laser scanning microscopy of Cu/Htma treated cells shows a rapid accumulation of copper within lysosomes over the course of hours, concurrent with the onset of apoptosis. A buildup of ubiquitinated proteins due to proteasome inhibition is seen on the same timescale and correlates with increases of copper without additional Htma.

Mulberry leaves ameliorate diabetes via regulating metabolic profiling and AGEs/RAGE and p38 MAPK/NF-κB pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Mulberry leaves have been used as traditional hypoglycemic medicine-food plant for thousand years in China. According to traditional Chinese medicine theory, type 2 diabetes mellitus (T2DM) belongs to the category of XiaoKe. Presently, the research of mulberry leaf hypoglycemic and lipid-lowering direction is mature, but the curative effects of alkaloids, flavonoids, polysaccharides, and other bioactive ingredients and the related mechanism is still unclear. AIM OF THE STUDY: This paper aims to study the efficacy and mechanism of alkaloids, flavonoids, polysaccharides, and other bioactive components in mulberry leaves in the treatment of T2DM individually. MATERIALS AND METHODS: The determination of levels of fasting blood glucose (FBG), triglyceride (TG) and total cholesterol (T-Cho), and pyruvate kinase (PK), hexokinase (HK), and alanine aminotransferase (ALT/GPT) of in plasma of diabetic mice. Urine metabolomics was analyzed by UPLC-QTOF/MS to evaluate differential metabolites from multiple metabolic pathways. The glucose uptake of HepG2 cells and 3T3-L1 cells. Expression of Caspase-3 and caspase-9, inflammatory injury and p38MAPK/NF-κB signaling pathway in GLUTag cells. RESULTS: Our study revealed alkaloids, flavonoids, and polysaccharides in mulberry leaf could increase the levels of PK, HK, and ALT/GPT, and decrease the levels of TG and T-Cho significantly, and regulate glucose, amino acid, and lipid metabolism. Furthermore, 1-deoxynojirimycin (DNJ) and isoquercitrin (QG) both could increase glucose uptake and promote differentiation of HepG2 cells, increase PPARγ, C/EBPα and SREBP-l expression in 3T3-L1 cells, and inhibit AGEs-induced injury and apoptosis in GLUTag cells, reduce the expression of proteins related to AGEs/RAGE and p38MAPK/NF-κB pathway. Notably, isoquercitrin exhibited more pronounced anti-diabetic efficacy. CONCLUSIONS: The alkaloids, flavonoids, and polysaccharides from mulberry leaf exhibited hypoglycemic activity through the regulation of glucose, amino acid, and lipid metabolism. 1-DNJ and QG increased glucose uptake and promoted differentiation of HepG2 cells, increased PPARγ, C/EBPα and SREBP-l expression in 3T3-L1 cells, and inhibited AGEs-induced injury and apoptosis in GLUTag cells via the AGEs/RAGE and p38 MAPK/NF-κB pathway.

Inflammation mobilizes copper metabolism to promote colon tumorigenesis via an IL-17-STEAP4-XIAP axis.

Copper levels are known to be elevated in inflamed and malignant tissues. But the mechanism underlying this selective enrichment has been elusive. In this study, we report a axis by which inflammatory cytokines, such as IL-17, drive cellular copper uptake via the induction of a metalloreductase, STEAP4. IL-17-induced elevated intracellular copper level leads to the activation of an E3-ligase, XIAP, which potentiates IL-17-induced NFκB activation and suppresses the caspase 3 activity. Importantly, this IL-17-induced STEAP4-dependent cellular copper uptake is critical for colon tumor formation in a murine model of colitis-associated tumorigenesis and STEAP4 expression correlates with IL-17 level and XIAP activation in human colon cancer. In summary, this study reveals a IL-17-STEAP4-XIAP axis through which the inflammatory response induces copper uptake, promoting colon tumorigenesis.

Single cell analysis reveals multiple requirements for zinc in the mammalian cell cycle.

Zinc is widely recognized as essential for growth and proliferation, yet the mechanisms of how zinc deficiency arrests these processes remain enigmatic. Here we induce subtle zinc perturbations and track asynchronously cycling cells throughout division using fluorescent reporters, high throughput microscopy, and quantitative analysis. Zinc deficiency induces quiescence and resupply stimulates synchronized cell-cycle reentry. Monitoring cells before and after zinc deprivation we found the position of cells within the cell cycle determined whether they either went quiescent or entered another cell cycle but stalled in S-phase. Stalled cells exhibited prolonged S-phase, were defective in DNA synthesis and had increased DNA damage levels, suggesting a role for zinc in maintaining genome integrity. Finally, we demonstrate zinc deficiency-induced quiescence occurs independently of DNA-damage response pathways, and is distinct from mitogen removal and spontaneous quiescence. This suggests a novel pathway to quiescence and reveals essential micronutrients play a role in cell cycle regulation.

Bioinspired Thiophosphorodichloridate Reagents for Chemoselective Histidine Bioconjugation.

Site-selective bioconjugation to native protein residues is a powerful tool for protein functionalization, with cysteine and lysine side chains being the most common points for attachment owing to their high nucleophilicity. We now report a strategy for histidine modification using thiophosphorodichloridate reagents that mimic post-translational histidine phosphorylation, enabling fast and selective labeling of protein histidines under mild conditions where various payloads can be introduced via copper-assisted alkyne-azide cycloaddition (CuAAC) chemistry. We establish that these reagents are particularly effective at covalent modification of His-tags, which are common motifs to facilitate protein purification, as illustrated by selective attachment of polyarginine cargoes to enhance the uptake of proteins into living cells. This work provides a starting point for probing and enhancing protein function using histidine-directed chemistry.

Redox-based reagents for chemoselective methionine bioconjugation.

Cysteine can be specifically functionalized by a myriad of acid-base conjugation strategies for applications ranging from probing protein function to antibody-drug conjugates and proteomics. In contrast, selective ligation to the other sulfur-containing amino acid, methionine, has been precluded by its intrinsically weaker nucleophilicity. Here, we report a strategy for chemoselective methionine bioconjugation through redox reactivity, using oxaziridine-based reagents to achieve highly selective, rapid, and robust methionine labeling under a range of biocompatible reaction conditions. We highlight the broad utility of this conjugation method to enable precise addition of payloads to proteins, synthesis of antibody-drug conjugates, and identification of hyperreactive methionine residues in whole proteomes.

The Intestinal Copper Exporter CUA-1 Is Required for Systemic Copper Homeostasis in Caenorhabditis elegans.

Copper plays key catalytic and regulatory roles in biochemical processes essential for normal growth, development, and health. Defects in copper metabolism cause Menkes and Wilson's disease, myeloneuropathy, and cardiovascular disease and are associated with other pathophysiological states. Consequently, it is critical to understand the mechanisms by which organisms control the acquisition, distribution, and utilization of copper. The intestinal enterocyte is a key regulatory point for copper absorption into the body; however, the mechanisms by which intestinal cells transport copper to maintain organismal copper homeostasis are poorly understood. Here, we identify a mechanism by which organismal copper homeostasis is maintained by intestinal copper exporter trafficking that is coordinated with extraintestinal copper levels in Caenorhabditis elegans Specifically, we show that CUA-1, the C. elegans homolog of ATP7A/B, localizes to lysosome-like organelles (gut granules) in the intestine under copper overload conditions for copper detoxification, whereas copper deficiency results in a redistribution of CUA-1 to basolateral membranes for copper efflux to peripheral tissues. Worms defective in gut granule biogenesis exhibit defects in copper sequestration and increased susceptibility to toxic copper levels. Interestingly, however, a splice isoform CUA-1.2 that lacks a portion of the N-terminal domain is targeted constitutively to the basolateral membrane irrespective of dietary copper concentration. Our studies establish that CUA-1 is a key intestinal copper exporter and that its trafficking is regulated to maintain systemic copper homeostasis. C. elegans could therefore be exploited as a whole-animal model system to study regulation of intra- and intercellular copper trafficking pathways.

Copper regulates cyclic-AMP-dependent lipolysis.

Cell signaling relies extensively on dynamic pools of redox-inactive metal ions such as sodium, potassium, calcium and zinc, but their redox-active transition metal counterparts such as copper and iron have been studied primarily as static enzyme cofactors. Here we report that copper is an endogenous regulator of lipolysis, the breakdown of fat, which is an essential process in maintaining body weight and energy stores. Using a mouse model of genetic copper misregulation, in combination with pharmacological alterations in copper status and imaging studies in a 3T3-L1 white adipocyte model, we found that copper regulates lipolysis at the level of the second messenger, cyclic AMP (cAMP), by altering the activity of the cAMP-degrading phosphodiesterase PDE3B. Biochemical studies of the copper-PDE3B interaction establish copper-dependent inhibition of enzyme activity and identify a key conserved cysteine residue in a PDE3-specific loop that is essential for the observed copper-dependent lipolytic phenotype.

Analysis of Thermal Field Distribution in Winter over Beijing from 1985 to 2015 Using Landsat Thermal Data.

Heat supply, automobile exhaust, industrial production and decrease of thermal inertia in winter caused by the decrease of vegetation coverage leads to an obvious difference in the distribution of the land thermal field in the winter compared with other seasons. The Urban thermal field distribution in the winter directly affects the spread of air pollutants, which has important implications for analyzing the contribution of the thermal field to particulate air pollution. Atmospheric transmissivity and atmospheric upwelling/downwelling radiance in simulations are first calculated using the moderate spectral resolution atmospheric transmittance algorithm and computer model (MODTRAN). Then, we solve the radiative transfer model of the thermal infrared band by constructing a look-up table. In addition, the accuracy estimation is performed using the simulated data, showing that when the error range of emissivity and water vapor content are confined to ±0.005 and ±0.6, respectively, the temperature retrieval error are less than 0.348 and 2.117 K, respectively indicating the high retrieval accuracy of the method. In addition, the long-term sequenced Landsat TM and ETM+ data were selected to retrieve land surface temperature (LST) during 1985-2015. The analysis of the temporal and spatial distribution of thermal fields in Beijing show that the spatial and temporal variations are observable. The spatial variation covers four levels: high temperature is distributed within the second ring, low temperature loops are distributed between the second and the fifth ring, high temperature is distributed in the outer suburb areas and the lowest temperature is distributed in the western mountainous areas. Meanwhile, the temporal variation of thermal field distribution changed a great deal during the rapid development in the past 3 decades: the low temperature loop expanded from the third to the sixth ring; the intensity and scope of the heat island effect within the second ring increased gradually.

Iron metabolism in infants: influence of bovine lactoferrin from iron-fortified formula.

OBJECTIVE: The aim of this study was to evaluate whether an iron-fortified formula with a concentration of lactoferrin would significantly improve the hematologic indexes and iron status in term infants compared with those same values in infants fed an iron-fortified formula without lactoferrin. METHOD: In this prospective, multicenter, controlled intervention study, 260 infants ages 4 to 6 mo were selected from six maternal and children's health care hospitals in the area. All infants were divided into two groups with the sequence of outpatient: lactoferrin-fortified formula milk group (fortified group, FG, containing lactoferrin 38 mg/100 g milk and iron element 4 mg/100 g milk) and no lactoferrin fortified milk (control group, CG, containing lactoferrin 0 mg/100 g milk and iron element 4 mg/100 g milk) for 3 mo. The levels of weight, height, and head circumference and the concentration of hemoglobin (Hb), serum ferritin (SF), and serum transferring receptor (sTfR) were measured and sTfR-SF index (TFR-F index), total body iron content (TBIC) and low height for age (HAZ), low weight for age (WAZ), and low weight for height (WHZ) were computed before and after the intervention, respectively. RESULTS: In all, 213 (115 in FG and 98 in CG) infants completed the intervention trial and all measurements of biochemical indicators. There were no significant differences in the average amount of daily intake of formula milk (94.3 ± 9.8 g versus 88.2 ± 8.7 g for FG and CG; P > 0.05) and iron element (3.8 ± 0.4 mg versus 3.7 ± 0.6 mg for FG and CG; P > 0.05). The average amount of daily intake of lactoferrin for infants in FG group was 35.8 ± 3.7 mg. The levels of weight, WAZ, WHZ, Hb, SF, TFR-F index, and TBIC after intervention of infants in FG were all significantly higher than those of infants in CG weight, 8723 ± 245 g versus 8558 ± 214g; WAZ, 1.02 ± 0.31 versus 0.44 ± 0.18; WHZ, 0.98 ± 0.31 versus 0.41 ± 0.12; Hb, 125.5 ± 15.4 g/L versus 116.9 ± 13.1 g/L; SF, 44.7 ± 17.2 µg/L versus 31.6 ± 18.4 µg/L; TFR-F index, 1.88 ± 0.41 versus 1.26 ± 0.39; TBIC, 6.12 ± 0.78 mg/kg versus 5.26 ± 0.55 mg/kg for FG and CG; P < 0.05), but significantly lower (P < 0.05) for the prevalence of anemia (4.1% versus 7.5%), iron deficiency (13.9% versus 24.4%), and iron-deficient anemia (1.7% versus 6.1%). CONCLUSION: When infants who were exclusively breastfed were supplemented with lactoferrin-fortified milk, significant increases in TBIC and iron absorption in the intestine were seen.

Efficacy and feasibility of radiofrequency ablation for hepatocellular carcinoma patients.

BACKGROUND/AIMS: Radiofrequency ablation (RFA) is a new treatment which is used to treat hepatocellular carcinoma (HCC). We performed this clinical trial to investigate whether it could reduce the damage of residual liver function. METHODOLOGY: We studied 40 hepatitis-related chronic patients who underwent RFA for hepatocellular carcinoma. Indocyanine green (ICG) test was performed pre and postoperatively. RESULTS: There were 32 males and 8 females with an average age of 53.98+12.59 years who underwent RFA for HCC. The mean preoperative ICGR15 value of 40 of the patients was (10.17+9.54) lower than the postoperative ICG retention rate at 15 min (ICGR15) value (14.95+12.71).Differences between the preoperative ICGR15 and the postoperative ICGR15 values were not significantly different (p=0.074). The 1-, 2- and 3-year survival rates were 98.7%, 88.8% and 76.4%, respectively. CONCLUSIONS: The results indicate that RFA is a minimally invasive treatment which provides a possible treatment modality for HCC patients with poor liver function and the efficacy is as well as the surgical treatment for HCC patients within the Milan criteria.