Transport characteristics of HL-1 cells: a new model for the study of adenosine physiology in cardiomyocytes.
Adenosine is a physiologically important nucleoside in the cardiovascular system where it can act as a cardioprotectant and modulator of energy usage. Adenosine transporters (ATs) modulate cellular adenosine levels, which, in turn, can affect a number of processes such as receptor activation and glucose uptake, but their role in cardiac physiology is poorly understood. Therefore, we have developed a new cell model by determining various adenosine-related characteristics of HL-1, an immortalized atrial cardiomyocyte murine cell line. Adenosine uptake in HL-1 cells is sodium independent, saturable, and inhibitable by nucleoside transport inhibitors (nitrobenzylthioinosine (NBTI), dipyridamole, dilazep). Reverse transcription--polymerase chain reaction analysis confirmed that HL-1 cells possess mouse equilibrative nucleoside transporters 1 and 2 (mENT1, mENT2) and kinetic analyses indicate moderate-affinity (Km = 51.3 +/- 12.9 microM), NBTI-sensitive adenosine transport. NBTI binds at a high-affinity single site (B(max) = 520 +/- 10 fmol/mg protein, Kd = 0.11 +/- 0.04 nM, 1.6 x 10(5) NBTI-binding sites/cell). HL-1 cells possess adenosine receptor, metabolic enzyme, protein kinase C isoform, and insulin-stimulated glucose transport profiles that match normal mouse heart. Therefore, HL-1 is an excellent model to study ATs within cardiomyocytes and the first model for evaluating in detail the role of the ATs in modulating effects of adenosine.
Chaudary
Naz
N
Department of Biology, York University, Toronto, ON, Canada.
Shuralyova
Iirina
I
Liron
Tamar
T
Sweeney
Gary
G
Coe
Imogen R
IR
D000241:Adenosine / Q000378:metabolism*; D000243:Adenosine Deaminase / Q000378:metabolism; D000248:Adenosine Kinase / Q000378:metabolism; D000345:Affinity Labels / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001692:Biological Transport; D015152:Blotting, Northern; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D017931:DNA Primers / Q000737:chemistry; D033721:Equilibrative Nucleoside Transporter 1 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D033722:Equilibrative-Nucleoside Transporter 2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D007328:Insulin / Q000494:pharmacology; D026901:Membrane Transport Proteins; D051379:Mice; D008954:Models, Biological; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism*; D033703:Nucleoside Transport Proteins; D011493:Protein Kinase C; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D013868:Thioinosine / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology
D000345:Affinity Labels; D002352:Carrier Proteins; D017931:DNA Primers; D033721:Equilibrative Nucleoside Transporter 1; D033722:Equilibrative-Nucleoside Transporter 2; D007328:Insulin; D026901:Membrane Transport Proteins; D033703:Nucleoside Transport Proteins; C498476:SLC29A1 protein, mouse; C486681:Slc29a2 protein, mouse; C444719:adenosine transporter; D013868:Thioinosine; D000248:Adenosine Kinase; D011493:Protein Kinase C; D000243:Adenosine Deaminase; C001789:4-nitrobenzylthioinosine; D005947:Glucose; D000241:Adenosine