Dynamic alteration of low-density lipoprotein receptor after exposure to transforming growth factor-beta2 in human Tenon's capsule fibroblasts.
OBJECTIVE
The present study investigated dynamic alteration of low-density lipoprotein receptor and its binding and uptake of low-density lipoprotein (LDL) after exposure to transforming growth factor-beta(2) (TGF-beta(2)) in human Tenon's capsule fibroblasts.


METHODS
Tenon's capsule fibroblasts obtained from elective cataract surgery patients were cultured and stimulated with different concentrations (0.1-10 ng/mL) of TGF-beta(2) for 24, 48, and 72 h. The LDLr mRNA and protein levels were analyzed by relative quantification real-time RT-PCR and Western blot analysis, respectively. The binding and uptake of DiO (3,3'-dioctadecyloxacarbocyanine)-labeled LDL was assessed by confocal microscopy.


RESULTS
Real-time RT-PCR and Western blot analyses showed similar results revealing that after exposure to TGF-beta(2), the expression of protein and mRNA of LDLr occurred in a concentration-dependent and time-dependent manner with a peak at a concentration of 1.0 ng/mL at 72 h in Tenon's capsule fibroblasts. Confocal microscopy showed that DiO-LDL binding and uptake were time-dependent, reaching saturation at approximately 6 h.


CONCLUSIONS
This study shows that LDLrs were overexpressed in the activated Tenon's capsule fibroblasts in a concentration-dependent and time-dependent manner after exposure to TGF-beta(2). The results suggest that LDLr in the activated Tenon's capsule fibroblasts may become a novel focus as a target receptor for controlled drug delivery, particularly in anti-scarring therapy during excessive conjunctival wound healing.
Shao
Tingting
T
State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, People's Republic of China.
Gao
Qianying
Q
Jiang
Ruzhang
R
Duan
Yongheng
Y
Sun
Xuerong
X
Ge
Jian
J
D015153:Blotting, Western; D002232:Carbocyanines / Q000737:chemistry; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D005123:Eye / Q000166:cytology / Q000378:metabolism; D005260:Female; D005347:Fibroblasts / Q000187:drug effects* / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D008077:Lipoproteins, LDL / Q000378:metabolism*; D008297:Male; D018613:Microscopy, Confocal; D008875:Middle Aged; D011485:Protein Binding; D012333:RNA, Messenger / Q000378:metabolism; D011973:Receptors, LDL / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D053781:Transforming Growth Factor beta2 / Q000008:administration & dosage / Q000494:pharmacology*
D002232:Carbocyanines; D008077:Lipoproteins, LDL; D012333:RNA, Messenger; D011973:Receptors, LDL; D053781:Transforming Growth Factor beta2; C098044:3,3'-dioctadecyloxacarbocyanine