Isolation of Two Plasticizers,Bis(2-ethylhexyl) Terephthalate and Bis(2-ethylhexyl) Phthalate,from Capparis spinosa L. Leaves

Many plants have been known to be contaminated and accumulate plasticizers from the environment, including water sources, soil, and atmosphere. Plasticizers are used to confer elasticity and flexibility to various fiber and plastic products. Consumption of plasticizers can lead to many adverse effects on human health, including reproductive and developmental toxicity, endocrine disruption, and cancer. Herein, we report for the first time that two plasticizers, bis(2-ethylhexyl) terephthalate (DEHT) and bis(2-ethylhexyl) phthalate (DEHP), have been isolated from the leaves of Capparis spinosa L. (the caper bush), a plant that is widely used in food seasonings and traditional medicine. 297 mg/kg of DEHT and 48 mg/kg of DEHP were isolated from dried and grounded C. spinosa L. leaves using column chromatography and semi-preparative high-performance liquid chromatography. Our study adds to the increase in the detection of plasticizers in our food and medicinal plants and to the alarming concern about their potential adverse effects on human health.     

Meta-Modeling of Methylprednisolone Effects on Glucose Regulation in Rats

A retrospective meta-modeling analysis was performed to integrate previously reported data of glucocorticoid (GC) effects on glucose regulation following a single intramuscular dose (50 mg/kg), single intravenous doses (10, 50 mg/kg), and intravenous infusions (0.1, 0.2, 0.3 and 0.4 mg/kg/h) of methylprednisolone (MPL) in normal and adrenalectomized (ADX) male Wistar rats. A mechanistic pharmacodynamic (PD) model was developed based on the receptor/gene/protein-mediated GC effects on glucose regulation. Three major target organs (liver, white adipose tissue and skeletal muscle) together with some selected intermediate controlling factors were designated as important regulators involved in the pathogenesis of GC-induced glucose dysregulation. Assessed were dynamic changes of food intake and systemic factors (plasma glucose, insulin, free fatty acids (FFA) and leptin) and tissue-specific biomarkers (cAMP, phosphoenolpyruvate carboxykinase (PEPCK) mRNA and enzyme activity, leptin mRNA, interleukin 6 receptor type 1 (IL6R1) mRNA and Insulin receptor substrate-1 (IRS-1) mRNA) after acute and chronic dosing with MPL along with the GC receptor (GR) dynamics in each target organ. Upon binding to GR in liver, MPL dosing caused increased glucose production by stimulating hepatic cAMP and PEPCK activity. In adipose tissue, the rise in leptin mRNA and plasma leptin caused reduction of food intake, the exogenous source of glucose input. Down-regulation of IRS-1 mRNA expression in skeletal muscle inhibited the stimulatory effect of insulin on glucose utilization further contributing to hyperglycemia. The nuclear drug-receptor complex served as the driving force for stimulation or inhibition of downstream target gene expression within different tissues. Incorporating information such as receptor dynamics, as well as the gene and protein induction, allowed us to describe the receptor-mediated effects of MPL on glucose regulation in each important tissue. This advanced mechanistic model provides unique insights into the contributions of major tissues and quantitative hypotheses for the multi-factor control of a complex metabolic system.     

Anti-CD70 Immunocytokines for Exploitation of Interferon-γ-Induced RIP1-Dependent Necrosis in Renal Cell Carcinoma

Metastatic renal cell carcinoma (RCC) is an incurable disease in clear need of new therapeutic interventions. In early-phase clinical trials, the cytokine IFN-γ showed promise as a biotherapeutic for advanced RCC, but subsequent trials were less promising. These trials, however, focused on the indirect immunomodulatory properties of IFN-γ, and its direct anti-tumor effects, including its ability to kill tumor cells, remains mostly unexploited. We have previously shown that IFN-γ induces RIP1 kinase-dependent necrosis in cells lacking NF-κB survival signaling. RCC cells display basally-elevated NF-κB activity, and inhibiting NF-κB in these cells, for example by using the small-molecule proteasome blocker bortezomib, sensitizes them to RIP1-dependent necrotic death following exposure to IFN-γ. While these observations suggest that IFN-γ-mediated direct tumoricidal activity will have therapeutic benefit in RCC, they cannot be effectively exploited unless IFN-γ is targeted to tumor cells in vivo. Here, we describe the generation and characterization of two novel ‘immunocytokine’ chimeric proteins, in which either human or murine IFN-γ is fused to an antibody targeting the putative metastatic RCC biomarker CD70. These immunocytokines display high levels of species-specific IFN-γ activity and selective binding to CD70 on human RCC cells. Importantly, the IFN-γ immunocytokines function as well as native IFN-γ in inducing RIP1-dependent necrosis in RCC cells, when deployed in the presence of bortezomib. These results provide a foundation for the in vivo exploitation of IFN-γ-driven tumoricidal activity in RCC.     

DYRK1A regulates the recruitment of 53BP1 to the sites of DNA damage in part through interaction with RNF169

Human Dual-specificity tyrosine (Y) Regulated Kinase 1A (DYRK1A) is encoded by a dosage dependent gene whereby either trisomy or haploinsufficiency result in developmental abnormalities. However, the function and regulation of this important protein kinase are not fully understood. Here, we report proteomic analysis of DYRK1A in human cells that revealed a novel role of DYRK1A in DNA double-strand breaks (DSBs) repair, mediated in part by its interaction with the ubiquitin-binding protein RNF169 that accumulates at the DSB sites and promotes homologous recombination repair (HRR) by displacing 53BP1, a key mediator of non-homologous end joining (NHEJ). We found that overexpression of active, but not the kinase inactive DYRK1A in U-2 OS cells inhibits accumulation of 53BP1 at the DSB sites in the RNF169-dependent manner. DYRK1A phosphorylates RNF169 at two sites that influence its ability to displace 53BP1 from the DSBs. Although DYRK1A is not required for the recruitment of RNF169 to the DSB sites and 53BP1 displacement, inhibition of DYRK1A or mutation of the DYRK1A phosphorylation sites in RNF169 decreases its ability to block accumulation of 53BP1 at the DSB sites. Interestingly, CRISPR-Cas9 knockout of DYRK1A in human and mouse cells also diminished the 53BP1 DSB recruitment in a manner that did not require RNF169, suggesting that dosage of DYRK1A can influence the DNA repair processes through both RNF169-dependent and independent mechanisms. Human U-2 OS cells devoid of DYRK1A display an increased HRR efficiency and resistance to DNA damage, therefore our findings implicate DYRK1A in the DNA repair processes.     

The RNase PARN Controls the Levels of Specific miRNAs that Contribute to p53 Regulation

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Study of the Geometry and Folding Pattern of Leaves of Mimosa pudica

Many structural and functional properties possessed by plants have great potentials to stimulate new concepts and inno-vative ideas in the field of biomimetic engineering. The key inputs from biology can be used for creation of efficient and op-timized structures. The study of the geometry and folding pattern of leaves of Mimosa pudica,referred as Sensitive Plant,reveals some of the peculiar characteristics during folding and unfolding. When the leaf is touched,it quickly folds its leaflets and pinnae and droops downward at the petiole attachment. With the help of experiments on simulation model,the variations in angle of leaflets and degree of compaction after folding are investigated.     

Thrombospondin-1 expression and modulation of Wnt and hippo signaling pathways during the early phase of Trypanosoma cruzi infection of heart endothelial cells

The protozoan parasite, Trypanosoma cruzi, causes severe morbidity and mortality in afflicted individuals. Approximately 30% of T. cruzi infected individuals present with cardiac pathology. The invasive forms of the parasite are carried in the vascular system to infect other cells of the body. During transportation, the molecular mechanisms by which the parasite signals and interact with host endothelial cells (EC) especially heart endothelium is currently unknown. The parasite increases host thrombospondin-1 (TSP1) expression and activates the Wnt/β-catenin and hippo signaling pathways during the early phase of infection. The links between TSP1 and activation of the signaling pathways and their impact on parasite infectivity during the early phase of infection remain unknown. To elucidate the significance of TSP1 function in YAP/β-catenin colocalization and how they impact parasite infectivity during the early phase of infection, we challenged mouse heart endothelial cells (MHEC) from wild type (WT) and TSP1 knockout mice with T. cruzi and evaluated Wnt signaling, YAP/β-catenin crosstalk, and how they affect parasite infection. We found that in the absence of TSP1, the parasite induced the expression of Wnt-5a to a maximum at 2 h (1.73±0.13), P< 0.001 and enhanced the level of phosphorylated glycogen synthase kinase 3β at the same time point (2.99±0.24), P<0.001. In WT MHEC, the levels of Wnt-5a were toned down and the level of p-GSK-3β was lowest at 2 h (0.47±0.06), P< 0.01 compared to uninfected control. This was accompanied by a continuous significant increase in the nuclear colocalization of β-catenin/YAP in TSP1 KO MHEC with a maximum Pearson correlation coefficient of (0.67±0.02), P< 0.05 at 6 h. In WT MHEC, the nuclear colocalization of β-catenin/YAP remained steady and showed a reduction at 6 h (0.29±0.007), P< 0.05. These results indicate that TSP1 plays an important role in regulating β-catenin/YAP colocalization during the early phase of T. cruzi infection. Importantly, dysregulation of this crosstalk by pre-incubation of WT MHEC with a β-catenin inhibitor, endo-IWR 1, dramatically reduced the level of infection of WT MHEC. Parasite infectivity of inhibitor treated WT MHEC was similar to the level of infection of TSP1 KO MHEC. These results indicate that the β-catenin pathway induced by the parasite and regulated by TSP1 during the early phase of T. cruzi infection is an important potential therapeutic target, which can be explored for the prophylactic prevention of T. cruzi infection.