Protective effect of paeoniflorin against optic nerve crush.

In order to evaluate the efficacy of traditional paeonia extract paeoniflorin against optic nerve crush, 16 Brown Norway rats were divided into two groups at random, with 8 rats in each group. In paeoniaflorin-treated group, 2 mg paeoniaflorin (total volume: 1 mL) was injected into rat's peritoneum one time a day for a period of 8 days. Rats in untreated group were given a single dose of vehicle. The optic nerve was crushed by a special forceps for 30 s in the left eye and a sham procedure was performed in the right eye on the 2nd day after the first injection. The retrograde fluorogold labeling of ganglion cells was conducted 5 days after optic nerve crush. The whole retina was flat-mounted thereafter. The ganglion cells that survived the crush were counted under fluorescent microscope by using an automatic counting software. As compared with the contralateral eye, the survival rate of ganglion cells in the left eye increased from 40.22% to 64.53% with a significant difference found between them (t=2.55, P=0.023). The results suggested that the paeonia extract paeoniflorin possessed a protective effect against optic nerve crush.

Inhibitory effect of dexamethasone on TGF-beta1 expression of rabbit ciliary pigment epithelia cultured in vitro.

In order to elucidate the effect of dexamethasone on the expression of transforming growth factor-beta1 (TGF-beta1) in ciliary pigment epithelial (CPE) cells cultured in vitro, rabbit CPE cells were cultured in vitro, treated with DMEM medium containing 0, 1 x 10(-8), 5 x 10(-8), 10 x 10(-8) and 50 x 10(-8) mol/L dexamethasone respectively for 5 days. The TGF-beta1 expression was detected by immunohistochemistry Supervision methods and analyzed semi-quantitatively by HMIAS-2000 image system. As opposed to in vivo, rabbit CPE cells expressed TGF-beta1 under cultured circumstance in vitro. The gray scales of the positive yellow staining in the groups of 1 x 10(-8), 5 x 10(-8), 10 x 10(-8) and 50 x 10(-8) mol/L dexamethasone were 136.57 +/- 4.43, 140.20 +/- 6.10, 142.98 +/- 2.99, 146.80 +/- 1.68 and 150.05 +/- 1.94 respectively. When the concentrations of dexamethasone were equal to or higher than 5 x 10(-8) mol/L and, the expression of TGF-beta1 was inhibited. 10(-7) mol/L dexamethasone showed a significant inhibition. It was suggested that CPE cells possess the potential ability of synthesizing and expressing TGF-beta1. The inhibition of TGF-beta1 expression by dexamethasone may be beneficial to the treatment of proliferative vitroretinopathy, also exert some influence on the secretion of aqueous humor and ciliary inflammation.

Inhibitory effect of tissue transglutaminase (tTG) antisense oligodeoxynucleotides on tTG expression in cultured bovine trabecular meshwork cells.

To study the effect of tTG fully phosphorothioated antisense oligodeoxynucleotides (tTG-ASDON) on tTG expression in cultured bovine trabecular meshwork cells (BTMCs) in vitro and explore a new treatment alternative for primary open angle glaucoma (POAG), the ASDON1 and ASDON2 complementary to the protein codogram region of tTG were designed, synthesized and phosphorothioated according to the secondary structure of tTG. The ASDON1 and ASDON2 were embedded in Lipofectamine and transfected into BTMCs. The untreated group served as negative controls. The expression of tTG in the mRNA and protein level were measured by semi-quantitative RT-PCR and immunohistochemical technique-Supervision method respectively. Our results showed that both the mRNA and the protein of tTG with tTG-ASDON and tTG-ASDON2 were significantly decreased as compared with that of the controls (P < 0.05). On the other hand, no significant difference was found between the ASDON1 group and the ASDON2 group. It is concluded that the expression of tTG mRNA and protein in cultured BTMC are down-regulated by tTG- ASDON. As a result, tTG-ASDON may be used for the treatment of POAG through the inhibitory effect on the expression of tTG.

Regulatory effect of dexamethasone on aquaporin-1 expression in cultured bovine trabecular meshwork cells.

To evaluate the effect of dexamethasone on the expression of aquaporin-1 (AQP-1) in cultured bovine trabecular meshwork cells, bovine trabecular meshwork cells were cultured in vitro and reproduced to the third and the fourth generation, then treated with dexamethasone at the concentrations of 5, 25, 50, 250 microg/L respectively for 7 days. Immunohistochemical technique-supervision method was employed to measure, and image analysis system to analyze the expression of AQP-1 in normal cultured bovine trabecular meshwork cells and those treated with dexamethasone. In normal bovine trabecular meshwork cells, the grayscale of AQP-1 positive staining was 167.94 +/- 1.18, while it was 168.92 +/- 0.91, 176.72 +/- 1.80, 180.64 +/- 1.31, 185.64 +/- 1.58 in cells treated with 5, 25, 50, 250 microg/L concentrations of dexamethasone. When the concentration of dexamethasone was higher than 25 microg/L, the expression of AQP-1 was significantly inhibited (P < 0.05). The regulation of AQP-1 expression by dexamethasone in cultured bovine trabecular meshwork cells in vitro may be one of causes that retard the aqueous outflow in glucocorticoid- induced glaucoma.

Inhibitory mechanism of inteferon-gamma on human fibroblasts from Tenon's capsule.

The inhibitory mechanism of interferon-gamma (IFN-gamma) on the fibroblasts from Tenon's capsule was studied. By using immunohistochemical SP method and pathological image system, the inhibitory effects of IFN-gamma on the expression of transforming growth factor beta receptor I in the in vitro cultured fibroblasts from Tenon's capsule were quantitatively analyzed. The results showed that IFN-gamma could reduce the expression of transforming growth factor beta receptor I in the fibroblasts with the following dose-effect relationship: Y = 1937.5-134.2 Igx (r=-0.971, P<0.01). It was concluded that IFN-gamma could inhibit the expression of transforming growth factor beta receptor I in the fibroblasts from Tenon's capsule. The modulation of the transforming growth factor beta receptor I expression by IFN-gamma may be beneficial to the alleviation of the hyperplasia of scar after trabeculectomy.

Expression of tissue transglutaminase in cultured bovine trabecluar meshwork cells.

To study whether cultured bovine trabecluar meshwork cells (BTMC) are capable of expressing tTG in protein and at mRNA level, BTMC were cultured in vitro and passaged three times, then the cells were transferred onto or cultured on sterile cover or submitted to isolation of RNA with Trizol, and the expression of tTG was detected by immunohistochemical technique and reverse transcription polymerase chain reaction (RT-PCR) respectively. Our results showed that tTG immunostaining was positive in the cytoplasm and rarely in the nucleus of cultured BTMC. No immunostaining was seen in the negative control. Moreover, a single RT-PCR amplified product whose sequence and size were in accordance with our known parameters was obtained. The expression of tTG in cultured BTMC was confirmed in protein and at mRNA level. BMTC is available more readily for the investigation of the relationship between tTG and primary open-angle glaucoma.