David W. Kastner and Steven L. Castle
Yaku’amide A was isolated in late 2010 from the deep-sea sponge Ceratopsion sp. and exhibits potent inhibitory activity against 39 human cancer cell lines via a potentially novel mode of action. However, the total synthesis requires many steps and produces low yields. In this study, we determined two simpler YA analogues that may retain its anticancer properties and can be synthesized in larger quantities. The analogues replace dehydroisoleucine with either dehydroethylnorvaline or dehydrovaline.
Juan Mejia, Parker Hall, Kelsey Hirschi, Paul R. Reynolds, and Juan Arroyo
The Pyruvate kinase isozymes M2 (PKM2) protein is a metabolic enzyme that regulates the final step of glycolysis. This enzyme is present in highly proliferating cells such as those present in embryonic tissues, cancer and in the placental trophoblast. We recently studied PKM2 protein in the human placenta at term. Our results demonstrated upregulated placental PKM2 during preeclampsia (PE) where trophoblast invasion is decreased and hypoxia is present. Our objective was to determine the cytosolic and nuclear PKM2 expression pattern and determine PKM2 regulation of trophoblast cell invasion. Trophoblast cells were cultured in normoxic and hypoxic conditions and real time cell invasion was determined during activation (Fructose-6-bisphosphate) or inhibition (Shikonin) of PKM2. Treated cells were lysed for western blot analysis of PKM2. During normoxia, we observed: 1) 2-fold increase (p<0.002) in trophoblast invasion when PKM2 was activated; 2) a 68-fold reduction (p<0.002) in trophoblast invasion when PKM2 activation was inhibited; 3) a 1.4-fold induction (p<0.03) of nuclear PKM2 in the trophoblast when PKM2 was activated; 4) a 1.6-fold induction (p<0.04) of nuclear PKM2 after PKM2 inactivation; and 5) a 1.4-fold decrease (p<0.03) in cytosolic PKM2 after PKM2 inactivation. During hypoxia we observed: 1) decreased cell invasion (~3.0-fold; p<0.005) in treated cells; 2) decreased cytosolic PKM2 expression (~1.6-fold; p<0.03) in treated cells; and 3) no significant differences in the expression of cytosolic PKM2. We conclude that PKM2 regulates trophoblast cell invasion depending of its cellular localization. Furthermore, our results suggest that PKM2 regulation is affected by hypoxia. These results suggest that PKM2 could be a mediator of trophoblast cell invasion and its abundance influences the development of complicated pregnancies like PE.
Benjamin Ogilvie, Jon Mitton, Jordan Tucker, Dennis L. Eggett, and Richard A. Robison
Povidone-iodine is an antiseptic that is frequently used to clean skin prior to surgery. Current FDA regulations require that hospitals dispense povidone-iodine from single-use bottles, rather than large, multi-use containers, to prevent microbial contamination. This results in hospitals generating lots of product waste. However, if povidone-iodine vapor can kill microbes, then multi-use containers may be safe to use, since any bacteria that contaminated the inside of the container would soon die. The purpose of this research was to determine whether vapor from povidone-iodine could kill methicillin-resistant Staphylococcus aureus (MRSA), a common cause of hospital-associated infections.
In summary, this experiment demonstrated that the povidone-iodine vapor killed MRSA in a time-dependent manner; most samples demonstrated a 5-log reduction after 80 minutes of povidone-iodine vapor exposure. This bacterial death is likely caused by volatilized iodine, which is surprising, as a typical povidone-iodine solution only has approximately 1% available iodine in solution. Iodine is moderately volatile, having roughly the same heat of vaporization as water (41.57 kJ/mol). A typical multi-use povidone-iodine container probably has a significant amount of iodine vapor inside, making it unlikely to support microbial contamination for very long. It may be possible to safely re-use povidone-iodine containers in a controlled manner.
Katie Pennington, Eranga Roshan, Joshua Andersen, and Katherine K. McCormack
The regulatory protein 14-3-3z promotes cellular growth and survival, and high expression levels of 14-3-3z are associated with mortality in multiple cancer types. 14-3-3z binds cellular proteins in a phosphorylation-dependent manner to modulate their functions. We have been characterizing the interactome of 14-3-3z which will likely lead to candidates for novel therapies for chemoresistant cancers. Tumor cells must be able to survive in a variety of stresses within the tumor microenvironment, including hypoxia, which occurs when tumors lack adequate blood supply. Previous data from our lab suggested that 14-3-3z promotes the adaptation of tumor cells to hypoxic stress. Our hypothesis is that hypoxic stress rearranges the interactome of 14-3-3z to focus on interactions that promote hypoxic stress. We have utilized protein mass spectrometry to find likely interactors of 14-3-3z with an emphasis on interactors that are found in high concentrations in hypoxia. We then validated these interactors using Co-IP and Western blot. Using these techniques, we have found several proteins of interest that warrant further exploration, including TAK1 and the TAK1 binding protein, TAB2, both of which are regulators of apoptosis. We are currently mutating potential phosphorylation sites in these proteins to test 14-3-3z binding and regulatory function. We hypothesize that phosphorylated TAK1 and TAB2 bind to 14-3-3z promoting progrowth pathways.
Differential Expression of mTOR Related Molecules in the Placenta of Patients with Gestational Diabetes Mellitus, Intrauterine Growth Restriction, or Preeclampsia
Katherine Price, Brent Kimbler, Nekel Knowlton, Kelsey Hirschi, Paul Reynolds, and Juan Arroyo
The mechanistic target of rapamycin (mTOR) pathway is involved in placental growth and function during pregnancy. The mTOR pathway responds to nutrient availability and growth factors regulating protein transcription and cell growth. mTOR disruptions are associated with the development of obstetric complications which may result in adverse health outcomes for the mother and/or fetus. The purpose of this study is to identify the differing placental expression of various mTOR-associated proteins during normal gestation (Control), gestational diabetes mellitus (GDM), intrauterine growth restriction (IUGR) and preeclampsia (PE). Immunohistochemistry was used to stain human placenta for activated proteins (phospho; p)AKT, (p)ERK, (p)mTOR, (p)p70 and (p)4EBP1. Real-time PCR array was completed to show differing placental expression of additional mTOR-associated genes during these conditions. We observed: 1) increased (p)AKT during GDM, 2) increased (p)ERK during IUGR, 3) increased (p)mTOR during GDM, while decreased during IUGR and PE, 4) increased (p)p70 during IUGR while decreased during GDM and PE, 5) increased (p)4EBP1 during GDM, IUGR, and PE, and 6) differential placental expression of mTOR pathway associated genes. We conclude that diverse regulation of the mTOR pathway is uniquely involved in the development of the obstetric complications studied. These results may provide insights into the physiological relevance of these pathways, and if so, their modification during gestation may help alleviate these diseases.
Influenza virus is a contagious respiratory pathogen that infects hundreds of thousands of people a year, making it a serious global health concern. The virus has a rapid mutation rate and has developed resistance to current antiviral agents, making it a difficult target for effective treatment. There is an increasing need to identify new treatments for Influenza. Recently, we have tested nutraceuticals as an effective alternative for blocking Influenza. We have tested Jamaican Sorrel, Black Currant Berries, and Manuka Honey paired with Bee Pollen for antiviral activity. We have demonstrated that these nutraceuticals block the 2009 Pandemic California Influenza strain between a concentration range of 1:8-1:16 dilution. Stock solutions of these nutraceuticals have been extracted with water to create a roughly 300 milliosmole solution. They are then neutralized with HEPES buffer, 1N HCl and1N NaOH.
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