Abstract
The blood–brain barrier (BBB) is a biological firewall that carefully regulates the cerebral microenvironment by acting as a physical, metabolic and transport barrier. This selectively permeable interface was modelled using the immortalised human cerebral microvascular endothelial cell line (hCMEC/D3) to investigate interactions with the cationic amino acid (CAA) L-arginine, the precursor for nitric oxide (NO), and with asymmetric dimethylarginine (ADMA), an endogenously derived analogue of L-arginine that potently inhibits NO production. The transport mechanisms utilised by L-arginine are known but they are not fully understood for ADMA, particularly at the BBB. This is of clinical significance giving the emerging role of ADMA in many brain and cerebrovascular diseases and its potential as a therapeutic target. We discovered that high concentrations of ADMA could induce endothelial dysfunction in the hCMEC/D3s BBB permeability model, leading to an increase in paracellular permeability to the paracellular marker FITC-dextran (40kDa). We also investigated interactions of ADMA with a variety of transport mechanisms, comparing the data with L-arginine interactions. Both molecules are able to utilise the CAA transport system y+. Furthermore, the expression of CAT-1, the best known protein from this group, was confirmed in the hCMEC/D3s. It is likely that influx systems, such as y+L and b0,+, have an important physiological role in ADMA transport at the BBB. These data are not only important with regards to the brain, but apply to other microvascular endothelia where ADMA is a major area of investigation.
Highlights
asymmetric dimethylarginine (ADMA) is an endogenously derived analogue of the cationic amino acid (CAA), L-arginine, and is produced during routine proteolysis of methylated arginine residues on proteins by a group of enzymes called protein-arginine methyltransferases (PRMTs) (Palm et al, 2007)
Whereas L-arginine is a substrate for nitric oxide synthases (NOS) which catalyse the oxidation of L-arginine to L-citrulline and nitric oxide (NO), ADMA acts as a Abbreviations: ADMA, asymmetric dimethylarginine; BBB, blood-brain barrier; CAA, cationic amino acids; CAT-1, cationic amino acid transporter 1; DDAH, NG, NGdimethylarginine dimethylaminohydrolase; GTPases, guanosine triphosphatases; IFN-γ, interferon gamma; LAT, large neutral amino acid transporters; L-NIO, N
An Octanol-saline partition coefficient (OSPC) was performed on all radiolabelled compounds to assess their relative lipophilicity which would give an indication of their potential to cross cell membranes by passive diffusion and accumulate in the cells
Summary
ADMA is an endogenously derived analogue of the CAA, L-arginine, and is produced during routine proteolysis of methylated arginine residues on proteins by a group of enzymes called protein-arginine methyltransferases (PRMTs) (Palm et al, 2007). Because of its potency as an inhibitor of NO, ADMA has been implicated in a wide variety of pathophysiological conditions throughout the body including hypercholesterolemia, hyperhomocysteinemia and hypertriglyceridemia (Boger et al, 1998; Lundman et al, 2001; Sydow et al, 2003); a host of cardiovascular conditions (Boger, 2003, 2004; Perticone et al, 2005); neuroinflammatory and neurodegenerative diseases including Alzheimer's disease (Selley, 2003; Arlt et al, 2008); cerebrovascular diseases such as stroke (Yoo and Lee, 2001; Brouns et al, 2009); microangiopathy-related cerebral damage (Notsu et al, 2009), and recently, intrauterine foetal growth restriction (Laskowska et al, 2014).
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