Browsing by Subject "Lysosomes"

Now showing 1 - 7 of 7
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    Studies on the breakdown of glycogen in the lysosomes: The effects of hydrocortisone
    (Murcia : F. Hernández, 2000) Kalamidas, Stefanos; Kotoulas, Othon B.
    The effects of hydrocortisone on newborn rat liver were studied by using biochemical assays, electron microscopy and quantitative morphometry. Hydrocortisone increased the number of lysosomes in the hepatocytes. Most of the lysosomes represented glycogen-containing autophagic vacuoles. The glucocorticoid also increased the activity of the liver glycogen-hydrolyzing acid glucosidase and the breakdown of glycogen inside lysosomes. The activity of the liver acid mannose 6-phosphatase was decreased. This may be related to the stimulation of autophagic mechanisms in the newborn rat hepatocytes
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    Targeting exogenous β-Defensin to the endolysosomal compartment via a vehicle guided system
    (Universidad de Murcia. Departamento de Biología Celular e Histología, 2017) Carvelli, Lorena; Libin, Yuan; Esfandnia, Sherry; Zhang, Yan; Presley, John F.; Morales, Carlos R.
    A number of pathogens for which there are no effective treatments infect the cells via endocytosis. Once in the endosomes, the pathogens complete their life cycle by overriding normal lysosomal functions. Recently, our laboratory identified the lysosomal targeting signal of prosaposin, which is recognized by the sorting receptor “sortilin”. Based on this evidence, we tested whether the antimicrobial peptide β-Defensin linked to the targeting sequence of prosaposin (βDPSAP) could be redirected from its secretory pathway to the endolysosomal compartment. To this effect, βDPSAP was transfected into COS-7 cells. The sub-cellular distribution of βD-PSAP was analyzed by confocal microscopy and differential centrifugation. Confocal microscopy demonstrated that βD-PSAP overlaid with the lysosomal marker LAMP1, indicating that the construct reached endosomes and lysosomes. Differential centrifugation also showed that βD-PSAP was in the lysosomal fractions. In addition, our binding inhibition assay demonstrated that βD-PSAP bound specifically to sortilin. Similarly, the delivery of βDPSAP was abolished after overexpressing a truncated sortilin. These results indicate that the prosaposin Cterminus and D/C-domain (prosaposin targeting sequence) was an effective “guidance system” to redirect βD-PSAP to the endolysosomal compartment. In the future, this and other fusion proteins with antimicrobial properties will be assembled to our “biotic vehicle” to target pathogens growing within these compartments.
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    The degradation of glycogen in the lysosomes of newborn rat hepatocytes: glycogen-, maltose- and isomaltose-hydrolyzing acid alpha glucosidase activities in liver
    (Murcia : F. Hernández, 1999) Kalamidas, Stefanos; Kotoulas, Othon B.
    The lysosomal glucosidase activities and glycogen degradation in newborn rat liver were studied by using biochemical assays, electron microscopy and quantitative morphometry. Glycogen-hydrolyzing, maltose-hydrolyzing and isomaltose-hydrolyzing activities were low at birth but increased afterwards. At the age of 6 hours they were markedly elevated. Actinomycin prevented the development of glucosidase activities indicating that these depend on protein synthesis. Parenteral glucose inhibited all three activities. This was apparently due to the abolition of normal postnatal hypoglycemia and the need for blood glucose. Cyclic AMP increased the glycogen-hydrolyzing but not the maltose-hydrolyzing activity. Propranolol inhibited the glycogen-hydrolyzing but not the maltose-hydrolyzing activity. The observations of this study provide further support for the hypothesis made by previous investigators that these activities are due to different enzymes.
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    The interactomics of sortilin: an ancient lysosomal receptor evolving new functions
    (Murcia : F. Hernández, 2009) Canuel, Maryssa; Libin, Yuan; Morales, Carlos R.
    The delivery of soluble lysosomal proteins to the lysosomes is dependent primarily on the mannose 6- phosphate receptor (MPR). The MPR has been demonstrated to attain the early endosomes via a process that requires the interaction of its cytosolic domain with the GGA and AP-1 adaptor proteins. Additionally, the MPR can be recycled back to the trans-Golgi network (TGN) through its interaction with the retromer complex. Interestingly, in I-cell disease (ICD), in which the MPR pathway is non-functional, many soluble lysosomal proteins continue to traffic to the lysosomes. This observation led to the discovery that sortilin is responsible for the MPR-independent targeting of the sphingolipid activator proteins (SAPs) and acid sphingomyelinase (ASM). More recently, our laboratory has tested the hypothesis that sortilin is also capable of sorting a variety of cathepsins that exhibit varying degrees of MPR-independent transport. We have demonstrated that the transport of cathepsin D is partially dependent upon sortilin, that cathepsin H requires sortilin, and that cathepsins K and L attain the lysosomes in a sortilin-independent fashion. As a type-1 receptor, sortilin also has numerous cytosolic binding partners. It has been observed that like the MPR, the anterograde trafficking of sortilin and its cargo require both GGAs and AP-1. Similarly, the retrograde recycling pathway of sortilin also involves an interaction with retromer through a YXXf site in the cytosolic tail of sortilin. In conclusion, the cytosolic domains of sortilin and MPR possess a high degree of functional homology and both receptors share a conserved trafficking mechanism.
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    The sorting and trafficking of lysosomal proteins
    (Murcia : F. Hernández, 2006) Ni, X.; Canuel, Maryssa; Morales, Carlos R.
    For a long time lysosomes were considered terminal organelles involved in the degradation of different substrates. However, this view is rapidly changing by evidence demonstrating that these organelles and their content display specialized functions in addition to the degradation of substances. Many lysosomal proteins have been implicated in specialized cellular functions and disorders such as antigen processing, targeting of surfactant proteins, and most lysosomal storage disorders. To date, about fifty lysosomal hydrolases have been identified, and the majority of them are targeted to the lysosomes via the mannose-6-phosphate receptor (M6P-Rc). However, recent studies on the intracellular trafficking of the nonenzymic lysosomal proteins prosaposin and GM2 activator (GM2AP) demonstrated that they use an alternative receptor termed “sortilin”. Existing evidence suggests that some hydrolases traffic to the lysosomes in a mannose 6-phophate-indepentend manner. The possibility that sortilin is implicated in the targeting of some soluble hydrolases, as well as the consequences of this process, is addressed in the present review.
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    The value of conjunctival biopsy in childhood cystinosis
    (Murcia : F. Hernández, 1989) Cruz-Sánchez, F. F.; Cervós-Navarro, J.; Rodríguez-Prados, S.; Lennert, T.
    Cystinosis is frequently presented with cystine storage in the cornea and cnnjiinctiva. and the diagnosis can be established by slit-lamp esamination. It can also be confirmed by elcctron microscopy of a conjunctival biopsy . The present paper reports on n 16-nionth-old boy with Fanconi's syndronie. in whoni the slit-lanip examination did not show crystal deposits of cystinc in the conjunctiva. The ~iltrastructurals tudj,o f the conjunctival biopsy demonstruted polygonal crystals within douhle mcmbrane-liniited organelles located in fibrobliists. Similar crystals were subsequently found in a hidney biopsy. We therefore think that conjunctival biopsy is a valuable diagnostic tool prior to pcrforming renal biopsy, even in cases with negativc findings hy ophthalmologic examination.
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    Ultrastructural localisation of acid phosphatase in intestinal eosinophilic granule cells (EGC) of rainbow trout
    (Murcia : F. Hernández, 1992) Powell, Mark D.; Briand, Heather A.; Wright, Glenda M.; Burka, John F.
    Enzyme cytochemistry was used to investigate possible lysosome involvement in capsaicin induced degranulation of the eosinophilic granule cell (EGC) of the rainbow trout intestine. Three adult rainbow trout (Oncorhynchus mykiss) were injected intra 'k eritoneally with capsaicin in a saline vehicle (0.5 pg.g- body weight). Following a 2 hour period of incubation, the fish were killed, and a mid portion of the intestine was dissected and fixed in cold glutaraldehyde buffered with sodium cacodylate. Vibratome sections were incubated in either reaction medium containing B-glycerophosphate and cerium chloride in acetate buffer or substrate (Bglycerophosphate) deficient control medium. Sections were then refixed in osmium tetroxide and processed for electron microscopy. Acid phosphatase was found to be localised within lysosomes. The enzyme was not found in the large cytoplasmic granules under normal or capsaicin-stimulated conditions. EGCs which had migrated to the lamina propria in response to the capsaicin stimulation had a distinct multivesicular granule morphology. These multivesicular granules did not contain acid phosphatase suggesting that this form of EGC degranulation is not a lysosomally mediated event.