Gian Gazzola

Publications (2)4.46 Total impact

• Article: In Lysinuric Protein Intolerance system y⁺L activity is defective in monocytes and in GM-CSF-differentiated macrophages

  Amelia Barilli, Bianca Rotoli, Rossana Visigalli, Ovidio Bussolati, Gian Gazzola, Zamir Kadija, Giuseppe Rodi, Francesca Mariani, Maria Ruzza, Maurizio Luisetti, Valeria Dall&apos, Asta
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  ABSTRACT: Abstract Background In the recessive aminoaciduria Lysinuric Protein Intolerance (LPI), mutations of SLC7A7/y+LAT1 impair system y+L transport activity for cationic amino acids. A severe complication of LPI is a form of Pulmonary Alveolar Proteinosis (PAP), in which alveolar spaces are filled with lipoproteinaceous material because of the impaired surfactant clearance by resident macrophages. The pathogenesis of LPI-associated PAP remains still obscure. The present study investigates for the first time the expression and function of y+LAT1 in monocytes and macrophages isolated from a patient affected by LPI-associated PAP. A comparison with mesenchymal cells from the same subject has been also performed. Methods Monocytes from peripheral blood were isolated from a 21-year-old patient with LPI. Alveolar macrophages and fibroblastic-like mesenchymal cells were obtained from a whole lung lavage (WLL) performed on the same patient. System y+L activity was determined measuring the 1-min uptake of [3H]-arginine under discriminating conditions. Gene expression was evaluated through qRT-PCR. Results We have found that: 1) system y+L activity is markedly lowered in monocytes and alveolar macrophages from the LPI patient, because of the prevailing expression of SLC7A7/y+LAT1 in these cells; 2) on the contrary, fibroblasts isolated from the same patient do not display the transport defect due to compensation by the SLC7A6/y+LAT2 isoform; 3) in both normal and LPI monocytes, GM-CSF induces the expression of SLC7A7, suggesting that the gene is a target of the cytokine; 4) GM-CSF-induced differentiation of LPI monocytes is comparable to that of normal cells, demonstrating that GM-CSF signalling is unaltered; 5) general and respiratory conditions of the patient, along with PAP-associated parameters, markedly improved after GM-CSF therapy through aerosolization. Conclusions Monocytes and macrophages, but not fibroblasts, derived from a LPI patient clearly display the defect in system y+L-mediated arginine transport. The different transport phenotypes are referable to the relative levels of expression of SLC7A7 and SLC7A6. Moreover, the expression of SLC7A7 is regulated by GM-CSF in monocytes, pointing to a role of y+LAT1 in the pathogenesis of LPI associated PAP.
  Orphanet Journal of Rare Diseases. 01/2010;
• Article: Arginine transport in human erythroid cells: discrimination of CAT1 and 4F2hc/y+LAT2 roles.

  Bianca Maria Rotoli, Ellen I Closs, Amelia Barilli, Rossana Visigalli, Alexandra Simon, Alice Habermeier, Nicoletta Bianchi, Roberto Gambari, Gian C Gazzola, Ovidio Bussolati, Valeria Dall'Asta
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  ABSTRACT: Since arginine metabolites, such as nitric oxide and polyamines, influence the expression of genes involved in erythroid differentiation, the transport of the cationic amino acid may play an important role in erythroid cells. However, available data only concern the presence in these cells of CAT1 transporter (system y(+)), while no information exists on the role of the heterodimeric transporters of system y(+)L (4F2hc/y(+)LAT1 and 4F2hc/y(+)LAT2) which operates transmembrane arginine fluxes cis-inhibited by neutral amino acids in the presence of sodium. Using erythroleukemia K562 cells and normal erythroid precursors, we demonstrate here that arginine transport in human erythroid cells is due to the additive contributions of a leucine-sensitive and leucine-insensitive component. In both cell types, leucine inhibition of arginine influx is much less evident in the absence of sodium, a hallmark of system y(+)L. In K562 cells, N-ethylmaleimide, a known inhibitor of CAT transporters (system y(+)), suppresses only a fraction of arginine influx corresponding to leucine-insensitive uptake. Moreover, in Xenopus oocytes coexpressing 4F2hc and y(+)LAT2, leucine exerts a marked inhibition of arginine transport, partially dependent on sodium, while no inhibition is seen in oocytes expressing CAT1. Lastly, silencing of SLC7A6, the gene for y(+)LAT2, lowers arginine transport and doubles the intracellular content of the cationic amino acid in K562 cells. We conclude that arginine transport in human erythroid cells is due to both system y(+) (CAT1 transporter) and system y(+)L (4F2hc/y(+)LAT2 isoform), which mainly contribute, respectively, to the influx and to the efflux of the cationic amino acid.
  Pflügers Archiv - European Journal of Physiology 07/2009; 458(6):1163-73. · 4.46 Impact Factor