Teaching life cycle assessment in higher education

The International Journal of Life Cycle Assessment - Scientific Life Cycle Assessment (LCA) literature provides some examples of LCA teaching in higher education, but not a structured overview of...     

Serum Metabolite Biomarkers Discriminate Healthy Smokers from COPD Smokers

COPD (chronic obstructive pulmonary disease) is defined by a fixed expiratory airflow obstruction associated with disordered airways and alveolar destruction. COPD is caused by cigarette smoking and is the third greatest cause of mortality in the US. Forced expiratory volume in 1 second (FEV1) is the only validated clinical marker of COPD, but it correlates poorly with clinical features and is not sensitive enough to predict the early onset of disease. Using LC/MS global untargeted metabolite profiling of serum samples from a well-defined cohort of healthy smokers (n = 37), COPD smokers (n = 41) and non-smokers (n = 37), we sought to discover serum metabolic markers with known and/or unknown molecular identities that are associated with early-onset COPD. A total of 1,181 distinct molecular ions were detected in 95% of sera from all study subjects and 23 were found to be differentially-expressed in COPD-smokers vs. healthy-smokers. These 23 putative biomarkers were differentially-correlated with lung function parameters and used to generate a COPD prediction model possessing 87.8% sensitivity and 86.5% specificity. In an independent validation set, this model correctly predicted COPD in 8/10 individuals. These serum biomarkers included myoinositol, glycerophopshoinositol, fumarate, cysteinesulfonic acid, a modified version of fibrinogen peptide B (mFBP), and three doubly-charged peptides with undefined sequence that significantly and positively correlate with mFBP levels. Together, elevated levels of serum mFBP and additional disease-associated biomarkers point to a role for chronic inflammation, thrombosis, and oxidative stress in remodeling of the COPD airways. Serum metabolite biomarkers offer a promising and accessible window for recognition of early-stage COPD.     

Myristate exposure in the human immunodeficiency virus type 1 matrix protein is modulated by pH

Human immunodeficiency virus type 1 (HIV-1) encodes a polypeptide called Gag that is capable of forming virus-like particles (VLPs) in vitro in the absence of other cellular or viral constituents. During the late phase of HIV-1 infection, Gag polyproteins are transported to the plasma membrane (PM) for assembly. A combination of in vivo, in vitro, and structural studies have shown that Gag targeting and assembly on the PM are mediated by specific interactions between the myristoylated matrix [myr(+)MA] domain of Gag and phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2]. Exposure of the MA myristyl (myr) group is triggered by PI(4,5)P2 binding and is enhanced by factors that promote protein self-association. In the studies reported here, we demonstrate that myr exposure in MA is modulated by pH. Our data show that deprotonation of the His89 imidazole ring in myr(+)MA destabilizes the salt bridge formed between His89(Hδ2) and Glu12(COO-), leading to tight sequestration of the myr group and a shift in the equilibrium from trimer to monomer. Furthermore, we show that oligomerization of a Gag-like construct containing matrix-capsid is also pH-dependent. Disruption of the His?Glu salt bridge by single-amino acid substitutions greatly altered the myr-sequestered?myr-exposed equilibrium. In vivo intracellular localization data revealed that the H89G mutation retargets Gag to intracellular compartments and severely inhibits virus production. Our findings reveal that the MA domain acts as a “pH sensor” in vitro, suggesting that the effect of pH on HIV-1 Gag targeting and binding to the PM warrants investigation.     

Oxidative alterations of cyclooxygenase during atherogenesis

Nitric oxide (*NO) and eicosanoids are critical mediators of physiological and pathophysiological processes. They include inflammation and atherosclerosis. *NO production and eicosanoid synthesis become disrupted during atherosclerosis and thus, it is important to understand the mechanisms that may contribute to this outcome. We, and others, have shown that nitrogen oxide (NO(x)) species modulate cyclooxygenase (COX; also known as prostaglandin H(2) synthase) activity and alter eicosanoid production. We have determined that peroxynitrite (ONOO(-)) has multiple effects on COX activity. ONOO(-) can provide the peroxide tone necessary for COX activation, such that simultaneous exposure of COX to its arachidonic acid substrate and ONOO(-) results in increased eicosanoid production. Alternatively, in the absence of arachidonic acid, ONOO(-) can modify COX through nitration of an essential tyrosine residue (Tyr385) such that it is incapable of catalysis. In this regard, we have shown that COX nitration occurs in human atherosclerotic tissue and in aortic lesions from ApoE(-/-) mice kept on a high fat diet. Additionally, we have demonstrated that Tyr nitration in ApoE(-/-) mice is dependent on the inducible form of NO synthase (iNOS). Under conditions where ONOO(-) persists and arachidonic acid is not immediately available, the cell may try to correct the situation by responding to ONOO(-) and releasing arachidonic acid via a signaling pathway to favor COX activation. Other post-translational modifications of COX by NO(x) species include S-nitrosation of cysteine (Cys) residues (which may have an activating effect) and Cys oxidation. The central focus of this review will include a discussion of how NO(x) species alter COX activity at the molecular level and how these modifications may contribute to altered eicosanoid output during atherosclerosis and lesion development.     

Structure of the myristylated human immunodeficiency virus type 2 matrix protein and the role of phosphatidylinositol-(4,5)-bisphosphate in membrane targeting

During the late phase of retroviral replication, newly synthesized Gag proteins are targeted to the plasma membrane (PM), where they assemble and bud to form immature virus particles. Membrane targeting by human immunodeficiency virus type 1 (HIV-1) Gag is mediated by the PM marker molecule phosphatidylinositol-(4,5)-bisphosphate [PI(4,5)P₂], which is capable of binding to the matrix (MA) domain of Gag in an extended lipid conformation and of triggering myristate exposure. Here, we show that, as observed previously for HIV-1 MA, the myristyl group of HIV-2 MA is partially sequestered within a narrow hydrophobic tunnel formed by side chains of helices 1, 2, 3, and 5. However, the myristate of HIV-2 MA is more tightly sequestered than that of the HIV-1 protein and does not exhibit concentration-dependent exposure. Soluble PI(4,5)P₂ analogs containing truncated acyl chains bind HIV-2 MA and induce minor long-range structural changes but do not trigger myristate exposure. Despite these differences, the site of HIV-2 assembly in vivo can be manipulated by enzymes that regulate PI(4,5)P₂ localization. Our findings indicate that HIV-1 and HIV-2 are both targeted to the PM for assembly via a PI(4,5)P₂-dependent mechanism, despite differences in the sensitivity of the MA myristyl switch, and suggest a potential mechanism that may contribute to the poor replication kinetics of HIV-2.     

Tyrosine nitration in prostaglandin H(2) synthase

In this study, we investigated the effects of various nitrogen oxide (NO(x)) species on the extent of prostaglandin H(2) synthase-1 (PGHS-1) nitration in purified protein and in vascular smooth muscle cells. We also examined PGHS-1 activity under these conditions and found the degree of nitration to correlate inversely with enzyme activity. In addition, since NO(x) species are thought to invoke damage during the pathogenesis of atherosclerosis, we examined human atheromatous tissue for PGHS-1 nitration. Both peroxynitrite and tetranitromethane induced Tyr nitration of purified PGHS-1, whereas 1-hydroxy-2-oxo-3-(N-methyl-aminopropyl)-3-methyl-1-triazene (NOC-7; a nitric oxide-releasing compound) did not. Smooth muscle cells treated with peroxynitrite showed PGHS-1 nitration. The extent of nitration by specific NO(x) species was determined by electrospray ionization mass spectrometry. Tetranitromethane was more effective than peroxynitrite, NOC-7, and nitrogen dioxide at nitrating a tyrosine-containing peptide (12%, 5%, 1%, and <1% nitration, respectively). Nitrogen dioxide and, to a lesser extent, peroxynitrite, induced dityrosine formation. Using UV/Vis spectroscopy, it was estimated that the reaction of PGHS-1 with excess peroxynitrite yielded two nitrated tyrosines/PGHS-1 subunit. Finally, atherosclerotic tissue obtained from endarterectomy patients was shown to contain nitrated PGHS-1. Thus, prolonged exposure to elevated levels of peroxynitrite may cause oxidative damage through tyrosine nitration.     

Castration induced changes in dog prostate gland associated with diminished activin and activin receptor expression

This study was conducted to evaluate the effect of androgen ablation on dog prostate gland structure and the proliferation capacity of the prostatic cells and their association with the expression of Activin A and Activin RIIA receptor. The effect of androgen on the prostate gland was compared in intact and castrated dogs after one and two weeks. Specific primary antibodies were used to immunolocalize activin-A, activin receptor type II A and the proliferation marker (PCNA). The results showed that the glandular acini of the prostate gland of intact dogs are lined by tall columnar secretory cells and less abundant flattened basal cells and surrounded by a thin fibromuscular tissue. The cytoplasm of the glandular cells exhibited an intense immunoreaction for activin A and activin RIIA receptor while basal cells expressed PCNA. Castration induced a remarkable atrophy of the prostatic acini associated with a progressive loss of secretory epithelial cells, which showed a dramatic decrease to complete disappearance of Activin A and Activin RIIA receptor immunoreactions. The remaining cells of the atrophied acini continue to express PCNA and the inter-acinar fibromuscular tissue showed a remarkable increase in its mass and are induced to express PCNA. These results indicated that androgen is required for the survival of epithelial cells and to maintain growth-quiescent fibromuscular cells, while basal cell proliferation is androgen independent. The changes in the Activin A and Activin RIIA receptor localization and their association with the dynamic pattern of prostate gland regression after castration suggested that Activin A and Activin RIIA receptor expression are androgen dependent.