Isolation of mutant renal (LLC-PK1) epithelia defective in basolateral, Na(+)-independent glucose transport.
To obtain mutant renal epithelia defective in the uptake of 2-deoxyglucose (Na(+)-independent glucose transport), LLC-PK1 renal epithelial cells were subjected to a combination of DNA alkylation, tritium-suicide with 2-[3H]deoxyglucose, and replica plating. One of the mutant sublines obtained, LLC-PK1M-7A, possesses only 40% of the 2-deoxyglucose uptake rate of the parent line, LLC-PK1M, and this defect is stable over at least 30 population doublings. Initial rate of 3-O-methylglucose transport into these mutant cells is only 20% of the parental rate, and efflux of 3-O-methylglucose from the mutant cells is correspondingly low. Glucose metabolism in the mutant cells does not appear to be altered, nor is free glucose accumulated in the cells against a concentration gradient. The uptake rate of L-leucine is the same in both mutant and parent, whereas the (Na(+)-dependent) uptake of alpha-methyl-D-glucoside and methylaminoisobutyric acid is greater in the mutant than the parent. The population doubling times of LLC-PK1M and LLC-PK1M-7A cultures are similar. LLC-PK1M-7A cell morphology is similar to LLC-PK1M when cultures are subconfluent, but on reaching confluence, LLC-PK1M-7A appear larger than LLC-PK1M cells. Their measured cell volume is then 150% of the volume of the parent cells. Future studies of the LLC-PK1M-7A mutant, and acquisition of additional mutant sublines, should elucidate the roles of transepithelial glucose transport in proximal tubular cell physiology.
Mullin
J M
JM
Lankenau Medical Research Center, Philadelphia, Pennsylvania 19151.
Snock
K V
KV
McGinn
M T
MT
Kofeldt
L M
LM
D019325:3-O-Methylglucose; D000818:Animals; D001693:Biological Transport, Active / Q000187:drug effects; D002460:Cell Line; D002999:Clone Cells; D003571:Cytochalasin B / Q000494:pharmacology; D003837:Deoxy Sugars / Q000378:metabolism*; D003847:Deoxyglucose / Q000378:metabolism*; D004847:Epithelial Cells; D004848:Epithelium / Q000187:drug effects / Q000378:metabolism; D005947:Glucose / Q000378:metabolism*; D007668:Kidney; D007700:Kinetics; D008757:Methylglucosides / Q000378:metabolism; D009154:Mutation; D010693:Phloretin / Q000494:pharmacology; D012964:Sodium / Q000494:pharmacology
D003837:Deoxy Sugars; D008757:Methylglucosides; D019325:3-O-Methylglucose; D003571:Cytochalasin B; D003847:Deoxyglucose; D012964:Sodium; D005947:Glucose; D010693:Phloretin