Suitability of LLC-PK1 pig kidney cells for the study of drug action on renal cell cholesterol uptake: identification and characterization of low-density lipoprotein receptors.
BACKGROUND
The purpose of this study was to identify and characterize the presence of low-density lipoprotein receptors (LDLr) in LLC-PK(1) cells.


METHODS
LLC-PK(1) cells were assessed for the presence of LDLr by conducting dose-response, LDL specific binding and competitive studies with DiI-LDL, and Western blot and RT-polymerase chain reaction (PCR) analyses. Assay conditions with IgG-C7, a monoclonal antibody (mAb) to the LDLr, were optimized, including temperature, preincubation time, and concentration in LLC-PK(1) cells.


RESULTS
LLC-PK(1) cells express LDL receptors as determined by LDL specific and competitive binding studies and Western blot and RT-PCR analysis (specific binding 0.5 ng DiI-LDL/mug of cellular protein).


CONCLUSIONS
Taken together, these findings confirm the presence of LDL receptors on LLC-PK1 cells and support the appropriateness of using these cells in studies involving renal cell cholesterol uptake and metabolism.
Chung
Nancy S
NS
Division of Pharmaceutics and Biopharmaceutics, Faculty of Pharmaceutical Sciences, The University of British Columbia, Vancouver, British Columbia, Canada.
Sachs-Barrable
Kristina
K
Lee
Stephen D
SD
Wasan
Kishor M
KM
D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive / Q000187:drug effects; D001681:Biological Assay; D015153:Blotting, Western; D002232:Carbocyanines / Q000494:pharmacology*; D018929:Cell Culture Techniques; D002784:Cholesterol / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D005240:Feasibility Studies; D005453:Fluorescence; D018374:LLC-PK1 Cells; D008077:Lipoproteins, LDL / Q000494:pharmacology*; D018613:Microscopy, Confocal; D011485:Protein Binding; D011973:Receptors, LDL / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013552:Swine
D002232:Carbocyanines; D008077:Lipoproteins, LDL; D011973:Receptors, LDL; C024286:3,3'-dioctadecylindocarbocyanine; D002784:Cholesterol