Estradiol biosynthesis in canine lens epithelial cells.

Purpose: To confirm that lens epithelial cells (LEC) synthesize 17β-estradiol, active estrogen, and to identify the pathway(s) by which normal and cataractous LEC synthesize 17β-estradiol.

Methods: ELISA was used to measure estradiol in aqueous humor; immunohistochemical staining was used to localize estradiol, testosterone and sulfatase; tritiated water release assay was used to measure aromatase activity; and qRT-PCR was used to quantify expression of aromatase and sulfatase in normal and cataractous canine and human LEC.

Results: Canine eyes with and without cataracts had no differences in aqueous humor estradiol levels; however, cataractous LEC had more intense immunoreactivity for estradiol than normal LEC.

The effect of phosphorylated Akt inhibition on posterior capsule opacification in an ex vivo canine model.

Purpose: To evaluate whether inhibition of phosphorylated Akt (pAkt) would reduce or prevent posterior capsule opacification (PCO) in an ex vivo canine lens capsule model.

Methods: Normal and cataractous lenses (n=6) were evaluated for pAkt via immunohistochemistry and immunoblotting. Primary cultures of lens epithelial cells (LEC) were exposed to ultraviolet light (UV) to induce pAkt.

ERalpha increases expression and interacts with TERT in cataractous canine lens epithelial cells.

Purpose: Estrogen receptor alpha (ERalpha) expression has previously been evaluated in lens epithelial cells (LEC). However, its function in the lens has not been determined. One potential function may be its interaction with the catalytic subunit of telomerase (TERT), which is present in normal LEC and higher in LEC that have undergone epithelial to mesenchymal transition (EMT).

Role of bacteria in the pathogenesis of recurrent uveitis in horses from the southeastern United States.

Objective: To determine the role of intraocular bacteria in the pathogenesis of equine recurrent uveitis (ERU) in horses from the southeastern United States by evaluating affected eyes of horses with ERU for bacterial DNA and intraocular production of antibodies against Leptospira spp.

Sample Population: Aqueous humor, vitreous humor, and serum samples of 24 clinically normal horses, 52 horses with ERU, and 17 horses with ocular inflammation not associated with ERU (ie, non-ERU inflammation).

Procedures: Ribosomal RNA quantitative PCR (real-time PCR) assay was used to detect bacterial DNA in aqueous humor and vitreous humor from clinically normal horses (n = 12) and horses with chronic (> 3-month) ERU (28).

Effect of grape polyphenols on oxidative stress in canine lens epithelial cells.

Objective: To evaluate whether the effects of oxidative stress could be attenuated in cultures of canine lens epithelial cells (LECs) by incubation with grape seed proanthocyanidin extract (GSE), resveratrol (RES), or a combination of both (GSE+RES).

Sample Population: Primary cultures of canine LECs.

Procedures: LECs were exposed to 100MM tertiary butyl-hydroperoxide (TBHP) with or without GSE, RES, or GSE+RES.

Prevention of posterior capsular opacification through cyclooxygenase-2 inhibition.

Purpose: To determine if cyclooxygenase-2 (COX-2) is upregulated when lens epithelial cells (LEC) in clinical samples of cataracts and posterior capsule opacification (PCO) undergo epithelial-mesenchymal transition (EMT)-like changes. We also wanted to learn if inhibition of the enzymatic activity of COX-2 could prevent PCO formation.

Methods: To ensure that EMT-like changes were occurring in LEC, real-time RT-PCR was used to examine expression of EMT markers.

Expression and characterization of the catalytic subunit of telomerase in normal and cataractous canine lens epithelial cells.

Purpose: To determine whether the catalytic subunit of telomerase reverse transcriptase (TERT) is regionally distributed in canine lens epithelial cells (LEC), compare TERT and the RNA subunit of telomerase (TR) mRNA expression and TERT protein expression in normal and cataractous LEC, and to evaluate whether telomerase activity is present in the cytoplasm and nucleus from normal LEC. Finally, the expression of p23 and heat shock protein 90 (hsp90), coactivators of TERT in neoplastic cells, were evaluated in normal and cataractous LEC.

Methods: TERT protein was detected by imunohistochemical staining and western immunoblotting in normal and cataractous LEC.

Ultraviolet irradiation up-regulates telomerase transcription and activity in lens epithelial cells.

Purpose: Ultraviolet irradiation (UVR) increases telomerase activity in various cell types including skin, a sun-exposed organ. The lens is also continually exposed to UVR and we hypothesized that lenses exposed to UVR would have increased telomerase activity, with up-regulated TERT and TR, the two main components of the telomerase holoenzyme. To evaluate whether the cornea would protect lenses from such changes, whole globes, as well as isolated lenses, were exposed to UVR, and lenses were evaluated for changes in telomerase activity.