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Alveolar epithelium, besides exerting a key role in gas exchange and surfactant production, plays important functions in host defense and inflammation. Pathological conditions associated to alveolar dysfunction include Acute Respiratory Distress Syndrome (ARDS), asthma, chronic obstructive pulmonary disease (COPD) and idiopathic pulmonary fibrosis (IPF). The use of predictive in vitro models of human alveolar epithelium is nowadays required for the study of disease mechanisms, as well as of pharmacokinetic parameters of pulmonary drugs delivery. Here, we employed a novel 3D model of human alveoli, namely EpiAlveolar™, consisting of primary alveolar epithelial cells, pulmonary endothelial cells and fibroblasts, that reflects properly the in vivo -like conditions. In EpiAlveolar™ we performed a characterization of Organic Cation Transporters (OCTs and OCTNs) expression and activity and we found that OCTN2, OCT1 and OCT3 are expressed on the basolateral membrane; instead, ATB0,+ transporter for cationic and neutral amino acids, which shares with OCTN2 the affinity for carnitine as substrate, is readily detectable and functional at the apical side. We also show that these transporters differentially interact with anticholinergic drugs. Overall, our findings reveal close similarities of EpiAlveolar™ with the tracheal/bronchial epithelium (EpiAirway™ model) and entrust this alveolar tissue as a potential tool for the screening of biopharmaceuticals molecules. • SLC6A14/ATB0,+ is strongly expressed at the apical side of alveolar epithelium. • OCT1, OCT3, and OCTN2 are active at the basolateral side. • All these transporters are involved in the interaction with respiratory drugs. [ABSTRACT FROM AUTHOR]

Abstract: Arginine is a semi-essential amino acid that plays an important role in the regulation of metabolic processes associated with several pathological/physiological conditions. In the vasculature, it mainly exerts its biological functions as a substrate of two alternative pathways: the conversion to nitric oxide (NO) by nitric oxide synthase (NOS) and the breakdown to urea and ornithine by arginase. To determine arginine metabolism, in the current study we propose an original radiochemical technique that allows the simultaneous monitoring of NOS and arginase activation within intact cells. Taking advantage of this method, we show here the consequences of different experimental conditions known to modulate endothelial homeostasis on arginine metabolism. [Copyright &y& Elsevier]

Abstract: Endothelial metabolism of arginine plays a key role in vascular homeostasis. While it is documented that the availability of extracellular arginine is critical for nitric oxide synthesis by eNOS, little is known about the relationships existing between arginine transport and the activity of arginase, the enzyme responsible for the production of ornithine and urea. The present study aims to characterize the role of PKC in the regulation of arginine transport and metabolism by human umbilical vein (HUVEC) and aortic (HAEC) endothelial cells. The results obtained demonstrate that the activation of PKCα by phorbol esters or thymeleatoxin causes a transient increase of arginine transport through system y+, referable to the induction of SLC7A2 mRNAs and to the increased expression of CAT2 transporters. PKCα-dependent stimulation of arginine transport requires the activation of MEK/ERK1/2 cascade, which leads to the stimulation of AP-1 and to the consequent induction of CAT2 expression. In parallel, PKCα activation also increases arginase expression and activity and promotes eNOS phosphorylation, resulting in decreased NO production. It is concluded that the activation of PKCα stimulates arginine entry in human endothelial cells and shifts the metabolism of the cationic amino acid from NO synthesis to arginase-dependent production of ornithine and urea. This metabolic deviation may contribute to the endothelial dysfunction associated with conditions of PKC overactivity. [Copyright &y& Elsevier]

Abstract: Lysinuric Protein Intolerance (LPI, MIM 222700) is a recessive aminoaciduria caused by defective cationic amino acid transport in epithelial cells of intestine and kidney. SLC7A7, the gene mutated in LPI, codifies for the y+LAT1 subunit of system y+L amino acid transporter. LPI patients frequently display severe complications, such as pulmonary disease, haematological abnormalities and disorders of the immune response. The transport defect may explain only a part of the clinical aspects of the disease, while the mechanisms linking the genetic defect to the clinical features of the patients remain thus far obscure. The aim of the study is to investigate the consequences of SLC7A7 mutations on specific macrophage functions, so as to evaluate if a macrophage dysfunction may have a role in the development of pulmonary and immunological complications of LPI. The results presented 1) confirm previous data obtained in one LPI patient, demonstrating that arginine influx through system y+L is markedly compromised in LPI macrophages; 2) demonstrate that also system y+L-mediated arginine efflux is significantly lower in LPI macrophages than in normal cells and 3) demonstrate that the phagocytic activity of LPI macrophages is severely impaired. In conclusion, SLC7A7/y+LAT1 mutations lead to a defective phenotype of macrophages, supporting the pathogenetic role of these cells in the development of LPI-associated complications. [Copyright &y& Elsevier]