Identification and characterization of lipids from endosomes purified by electromagnetic chromatography

Traditional separation techniques do not yield endolysosomes of sufficient purity to permit detailed biochemical characterization of this important class of intracellular vesicles. Here, we have used a magnetic chromatography technique to isolate the endosomes from rat peritoneal macrophages and studied their lipid composition. Electromagnetic isolation works by retention of colloidal iron containing vesicles on magnetic column. The data suggested that both early and late endosomes were rich in cholesterol, whereas sphingomyelin (SM) and specific phospholipids like phosphatidylcholine. phosphatidylethanolamine, phosphatidylglycerol and phosphatidylserine are enriched in the late compartments. Our results also indicated that the purified fractions are enriched in raft lipids like SM, but not in cholesterol. The endosomal purification method described here yields pure endosomes with little or no contamination from mitochondria and hence could be used for further biochemical and marker analysis, giving insight into mechanisms of endocytic traffic.     

Prognostic significance of CHAC1 expression in breast cancer

Molecular Biology Reports - An emerging component of Unfolded Protein Response (UPR) pathway, cation transport regulator homolog 1 (CHAC1) has been conferred with the ability to degrade...     

Pulmonary Artery Pseudoaneurysm in COVID-19-Associated Pulmonary Mucormycosis: Case Series and Systematic Review of the Literature

Mycopathologia - Literature on COVID-19-associated pulmonary mucormycosis (CAPM) is sparse. Pulmonary artery pseudoaneurysm (PAP) is an uncommon complication of pulmonary mucormycosis (PM), and...     

Descriptions of Deladenus albizicus n. sp. and D. processus n. sp. (Nematoda: Hexatylina) from Haryana,India

Two different nematodes were isolated from the bark of Albizia lebbeck trees; one from insect infested and another from noninfested, healthy tree. Based on the biological, morphological, and molecular evidences, the nematodes are described as Deladenus albizicus n. sp. and D. processus n. sp. (Nematoda: Hexatylina). Deladenus albizicus n. sp., isolated from insect-infested tree, multiplied on the fungus Nigrospora oryzae. Myceliophagous females of this nematode reproduced by parthenogenesis and spermathecae were indistinct. Infective females, readily produced in the cultures, are dorsally curved. Only one type of males containing small-sized sperms in their genital tracts were produced in the culture. Myceliophagous females: L = 0.75 to 1.71 mm, a = 32.3 to 50.8, b = 9.3 to 11.2, b’ = 5.2 to 7.3, c = 27.2 to 35.6, V = 91.0 to 93.3, c’ = 2.0 to 2.9, stylet = 11 to 12 µm, excretory pore in the region of median pharyngeal bulb, 43 to 47 µm anterior to hemizonid. Deladenus processus n. sp., isolated from bark of healthy A. lebbeck tree, was cultured on Alternaria alternata. Myceliophagous females reproduced by amphimixis and their spermathecae contained rounded sperms. Infective females were never produced, even in old cultures. Myceliophagous females: L = 0.76 to 0.99 mm, a = 34 to 49, b = 13.3 to 17.7, b’ = 3.8 to 5.8, c = 19.6 to 22.8, V = 92.2 to 93.5, c’ = 2.7 to 3.5, stylet = 6 to 7 µm, excretory pore in the proximity of hemizonid, tail conoid, tapering from both sides to a long pointed central process. It is proposed to classify Deladenus species in three groups: durus, siricidicola, and laricis groups based on female and spermatogonia dimorphism, mode of reproduction, and insect parasitism.     

Chloroplast and nuclear microsatellite analysis of Aegilops cylindrica

Aegilops cylindrica Host (2n=4x=28, genome CCDD) is an allotetraploid formed by hybridization between the diploid species Ae. tauschii Coss. (2n=2x=14, genome DD) and Ae. markgrafii (Greuter) Hammer (2n=2x=14, genome CC). Previous research has shown that Ae. tauschii contributed its cytoplasm to Ae. cylindrica. However, our analysis with chloroplast microsatellite markers showed that 1 of the 36 Ae. cylindrica accessions studied, TK 116 (PI 486249), had a plastome derived from Ae. markgrafii rather than Ae. tauschii. Thus, Ae. markgrafii has also contributed its cytoplasm to Ae. cylindrica. Our analysis of chloroplast and nuclear microsatellite markers also suggests that D-type plastome and the D genome in Ae. cylindrica were closely related to, and were probably derived from, the tauschii gene pool of Ae. tauschii. A determination of the likely source of the C genome and the C-type plastome in Ae. cylindrica was not possible.     

alpha-Lipoic acid ameliorates altered colonic contractility and intestinal transit in STZ-diabetic rats

alpha-Lipoic acid treatment (100 mg/kg/day for 2 weeks after 6 weeks of untreated diabetes) of streptozotocin diabetic rats partially but significantly reversed both reduced contractile response of distal colon to acetylcholine and delayed transit of charcoal meal in small intestine compared to diabetic control. These effects of alpha-Lipoic acid were associated with complete reversal of diabetes induced increased plasma lipid peroxidation level. alpha-Lipoic acid had no effect on any of the parameters measured in non-diabetic rats. These findings demonstrate contribution of oxidative stress in the development of physiological changes of gut in diabetes.     

Akt phosphorylates and suppresses the transactivation of retinoic acid receptor alpha

The transactivation of nuclear receptors is regulated by both ligand binding and phosphorylation. We previously showed that RARalpha (retinoic acid receptor alpha) phosphorylation by c-Jun N-terminal kinase contributes to retinoid resistance in a subset of NSCLC cells (non-small cell lung cancer cells), but the aetiology of this resistance in the remainder has not been fully elucidated [Srinivas, Juroske, Kalyankrishna, Cody, Price, Xu, Narayanan, Weigel and Kurie (2005) Mol. Cell. Biol. 25, 1054-1069]. In the present study, we report that Akt, which is constitutively activated in NSCLC cells, phosphorylates RARalpha and inhibits its transactivation. Biochemical and functional analyses showed that Akt interacts with RARalpha and phosphorylates the Ser96 residue of its DNA-binding domain. Mutation of Ser96 to alanine abrogated the suppressive effect of Akt. Overexpression of a dominant-negative form of Akt in an NSCLC cell line decreased RAR phosphorylation, increased RAR transactivation and enhanced the growth-inhibitory effects of an RAR ligand. The findings presented here show that Akt inhibits RAR transactivation and contributes to retinoid resistance in a subset of NSCLC cells.     

Modeling the effects of HER/ErbB1-3 coexpression on receptor dimerization and biological response

The human epidermal growth factor receptor (HER/ErbB) system comprises the epidermal growth factor receptor (EGFR/HER1) and three other homologs, namely HERs 2-4. This receptor system plays a critical role in cell proliferation and differentiation and receptor overexpression has been associated with poor prognosis in cancers of the epithelium. Here, we examine the effect of coexpressing varying levels of HERs 1-3 on the receptor dimerization patterns using a detailed kinetic model for HER/ErbB dimerization and trafficking. Our results indicate that coexpression of EGFR with HER2 or HER3 biases signaling to the cell surface and retards signal downregulation. In addition, simultaneous coexpression of HERs 1-3 leads to an abundance of HER2-HER3 heterodimers, which are known to be potent inducers of cell growth and transformation. Our new approach to use parameter dependence analysis in experimental design reveals that measurements of HER3 phosphorylation and HER2 internalization ratio may prove to be especially useful for the estimation of critical model parameters. Further, we examine the effect of receptor dimerization patterns on biological response using a simple phenomenological model. Results indicate that coexpression of EGFR with HER2 and HER3 at low to moderate levels may enable cells to match the response of a high HER2 expresser.     

Squid (Loligo pealei) giant fiber system: a model for studying neurodegeneration and dementia?

In many neurodegenerative disorders that lead to memory loss and dementia, the brain pathology responsible for neuronal loss is marked by accumulations of proteins in the form of extracellular plaques and intracellular filamentous tangles, containing hyperphosphorylated cytoskeletal proteins. These are assumed to arise as a consequence of deregulation of a normal pattern of topographic phosphorylation-that is, an abnormal shift of cytoskeletal protein phosphorylation from the normal axonal compartment to cell bodies. Although decades of studies have been directed to this problem, biochemical approaches in mammalian systems are limited: neurons are too small to permit separation of cell body and axon compartments. Since the pioneering studies of Hodgkin and Huxley on the giant fiber system of the squid, however, the stellate ganglion and its giant axons have been the focus of a large literature on the physiology and biochemistry of neuron function. This review concentrates on a host of studies in our laboratory and others on the factors regulating compartment-specific patterns of cytoskeletal protein phosphorylation (primarily neurofilaments) in an effort to establish a normal baseline of information for further studies on neurodegeneration. On the basis of these data, a model of topographic regulation is proposed that offers several possibilities for further studies on potential sites of deregulation that may lead to pathologies resembling those seen in mammalian and human brains showing neurodegeneration, dementia, and neuronal cell death.     

Elevated Oxidative Stress and Decreased Antioxidant Function in the Human Hippocampus and Frontal Cortex with Increasing Age: Implications for Neurodegeneration in Alzheimer’s Disease

Oxidative stress and mitochondrial damage are implicated in the evolution of neurodegenerative diseases. Increased oxidative damage in specific brain regions during aging might render the brain susceptible to degeneration. Previously, we demonstrated increased oxidative damage and lowered antioxidant function in substantia nigra during aging making it vulnerable to degeneration associated with Parkinson's disease. To understand whether aging contributes to the vulnerability of brain regions in Alzheimer's disease, we assessed the oxidant and antioxidant markers, glutathione (GSH) metabolic enzymes, glial fibrillary acidic protein (GFAP) expression and mitochondrial complex I (CI) activity in hippocampus (HC) and frontal cortex (FC) compared with cerebellum (CB) in human brains with increasing age (0.01-80 years). We observed significant increase in protein oxidation (HC: p = 0.01; FC: p = 0.0002) and protein nitration (HC: p = 0.001; FC: p = 0.02) and increased GFAP expression (HC: p = 0.03; FC: p = 0.001) with a decreasing trend in CI activity in HC and FC compared to CB with increasing age. These changes were associated with a decrease in antioxidant enzyme activities, such as superoxide dismutase (HC: p = 0.005), catalase (HC: p = 0.02), thioredoxin reductase (FC: p = 0.04), GSH reductase (GR) (HC: p = 0.005), glutathione-s-transferase (HC: p = 0.0001; FC: p = 0.03) and GSH (HC: p = 0.01) with age. However, these parameters were relatively unaltered in CB. We suggest that the regions HC and FC are subjected to widespread oxidative stress, loss of antioxidant function and enhanced GFAP expression during aging which might make them more susceptible to deranged physiology and selective neuronal degeneration.