PURPOSE: 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.
PURPOSE: 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.