In order to investigate whether cultured normal human lens epithelial cells (LEC) express transforming growth factor beta (TGF-beta), reverse transcriptase polymerase chain reaction (RT-PCR) and immunohistochemical methods were used for detection of TGF-beta mRNA and protein in cultured normal human LEC. The results showed that a single RT-PCR amplified product about 310bp was obtained, and the sequence was homologous to the known sequence. TGF-beta immunostain was positive in the plasma of LEC. It was suggested that normal human LEC could produce TGF-beta, and LEC could be affected by TGF-beta through autocrine action.
Cells, Cultured ; Epithelial Cells/cytology ; Epithelial Cells/*metabolism ; Lens, Crystalline/cytology ; Lens, Crystalline/*metabolism ; RNA, Messenger/biosynthesis ; RNA, Messenger/genetics ; Transforming Growth Factor beta/*biosynthesis ; Transforming Growth Factor beta/genetics

To investigate the role of interleukin-l beta(IL-1β)and interleukin-8(IL-8) in the pathogenesis of vernal keratoconjunctivitis.MethodsThe levels of two cytokines of peripheral blood serum were determined with radioimmunoassay in 13 volunteers,13 patients with quiet vernal keratoconjunctivitis and 13 patients with active vernal keratoconjunctivitis.ResultsIL-1βserum levels in active and quiet vernal keratoconjunctivitis were(0.134 6±0.022 9)ng/ml,(0.213 1±0.036 0)ng/ml,IL-8 serum levels in active and quiet vernal keratoconjunctivitis were(0.230 4±0.014 5)ng/ml,(0.323 8±0.036 9)ng/ml,which were higher than(0.075 0±0.010 8)ng/ml,(0.67 2±0.023 1)ng/ml of normal controls,and the differences among the three groups were statistically significant(P<0.001).ConclusionIL-1β and IL-8 might play a significant role in the pathogenesis of vernal keratoconjunctivitis,and their possible roles in estimating and treating the diesease are worthy of further investigation.

Whether transforming growth factor-beta2 (TGF-beta2) induces apoptosis of human trabecular meshwork cells was investigated in vitro. Cultured 3-5 passage human trabecular meshwork cells were treated with 0 (control), 0.32, 1, 3.2 ng/ml TGF-beta2 for 48 h and divided into control group and experimental group. The apoptosis of human trabecular meshwork cells was examined by transmission electron microscopy, TUNEL technique and flow cytometry. The results showed characteristic morphologic changes of apoptotic cells were observed under transmission electron microscopy. DNA fragmentation of human trabecular meshwork cells was found by TUNEL technique. Quantitative analysis of flow cytometry showed that percentages of apoptotic human trabecular meshwork cells were (2.79 +/- 0.44)%, (4.43 +/- 1.17)% and (9.60 +/- 2.05)% respectively with different concentrations [1 ng/ml (P<0.05), 3.2 ng/ml (P<0.01)] of TGF-beta2 with the difference being significant between experimental group and control group [(1.41 +/- 0.34)%]. It was concluded that TGF-beta2 can induce apoptosis of human trabecular meshwork cells in vitro and may be involved in the decrease of trabecular meshwork cells in the patients with primary open angle glaucoma and aging of normal people.
Apoptosis/*drug effects ; Cells, Cultured ; Trabecular Meshwork/*cytology ; Transforming Growth Factor beta/*pharmacology ; Transforming Growth Factor beta2

To explore the biological effects of light on trabecular cells, cultured bovine trabecular cells were exposed to visible light of different wavelength with different energy. Cellular morphology, structure, proliferation, and phagocytosis were observed. The cells showed no remarkable changes when the energy was low. When the exposure energy reached 1.12 mW/cm2, the cytoplasm showed a rough appearance, and cell proliferation and phagocytosis decreased. This phototoxicity was strong with white light (compound chromatic light), moderate with violet light or yellow light, and mild with red light.
Cell Proliferation/radiation effects ; Cells, Cultured ; *Light ; Phagocytosis/radiation effects ; Trabecular Meshwork/cytology ; Trabecular Meshwork/*radiation effects

Whether cultured bovine trabecular meshwork cells and trabecular tissue ex vivo express insulin-like growth factor-I (IGF-I) messenger RNA (mRNA) and protein was investigated. Total RNA of cultured bovine trabecular meshwork cells as well as trabecular meshwork tissue freshly excised from bovine eyes was extracted, and reverse transcriptase-polymerase chain reaction (RT-PCR) was used to detect IGF-I mRNA. RT-PCR product was verified by sequencing. Immunohistochemical stain was used to detect IGF-I protein. The results showed that a single PCR amplified product was obtained, and the sequence was homologous to the known sequence.. IGF-I immunostain was positive in the cytoplasm of trabecular meshwork cells. It was concluded that trabecular meshwork cells produce IGF-I and contribute to the presence of IGF-I in trabecular meshwork microenvironment as well as aqueous humor. Trabecular meshwork cells were affected by IGF-I not only through paracrine, but also autocrine action. Whether abnormal down-regulations in IGF-I production may contribute to the pathogenesis of primary open-angle glaucoma and the possibility of promoting the autocrine action of IGF-I by trabecular meshwork cells to treat the disease is worth further investigation.
Cells, Cultured ; Cloning, Molecular ; Glaucoma, Open-Angle/etiology ; Glaucoma, Open-Angle/metabolism ; Insulin-Like Growth Factor I/biosynthesis ; Insulin-Like Growth Factor I/*genetics ; RNA, Messenger/biosynthesis ; RNA, Messenger/genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Sequence Analysis, DNA ; Trabecular Meshwork/cytology ; Trabecular Meshwork/*metabolism

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.
Cells, Cultured ; Glaucoma, Open-Angle/metabolism ; RNA, Messenger/biosynthesis ; RNA, Messenger/genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Trabecular Meshwork/*metabolism ; Transglutaminases/*biosynthesis ; Transglutaminases/genetics

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.

In order to explore whether the conventional use of 5-fluorouracil (5-Fu) had any toxic effects on trabecular meshwork cells, bovine trabecular meshwork cells were cultured in vitro and exposed to 5-Fu at different concentrations. The cellular morphology, ultrastructure, mortality and phagocytosis were studied under light microscopy, transmission electron microscopy and methods of Wright's stain. It was found that the toxic effects of 5-Fu on the cells were in a dose-dependent mode. 1×10-1 mg/ml of 5-Fu caused a large part of cells rounded up, while 1×10-3 mg/ml of the drug only a rough appearance of the cell surface. Exposure to 1×10-2 mg/ml of 5-Fu made mitochrone swollen and rough endoplasmic reticulum enlarged, with the cell mortality being 50.5 %. The latex microspheres engulfed in cytoplasm in cells receiving 1×10-1 and 1×10-2 mg/ml of 5-Fu were significantly decreased as compared with those in the control group (P<0.01). It was concluded that the safe concentration of 5-Fu on bovine trabecular meshwork cells was 1×10-3 mg/ml and the conventional dosage of 5-Fu in clinical practice would not cause injury to trabecular meshwork cells.

In order to explore whether the conventional use of 5-fluorouracil (5-Fu) had any toxic effects on trabecular meshwork cells, bovine trabecular meshwork cells were cultured in vitro and exposed to 5-Fu at different concentrations. The cellular morphology, ultrastructure, mortality and phagocytosis were studied under light microscopy, transmission electron microscopy and methods of Wright's stain. It was found that the toxic effects of 5-Fu on the cells were in a dose-dependent mode. 1×10-1 mg/ml of 5-Fu caused a large part of cells rounded up, while 1×10-3 mg/ml of the drug only a rough appearance of the cell surface. Exposure to 1×10-2 mg/ml of 5-Fu made mitochrone swollen and rough endoplasmic reticulum enlarged, with the cell mortality being 50.5 %. The latex microspheres engulfed in cytoplasm in cells receiving 1×10-1 and 1×10-2 mg/ml of 5-Fu were significantly decreased as compared with those in the control group (P<0.01). It was concluded that the safe concentration of 5-Fu on bovine trabecular meshwork cells was 1×10-3 mg/ml and the conventional dosage of 5-Fu in clinical practice would not cause injury to trabecular meshwork cells.

In order to investigate whether cultured normal human lens epithelial cells (LEC) express transforming growth factor beta (TGF-beta), reverse transcriptase polymerase chain reaction (RT-PCR) and immunohistochemical methods were used for detection of TGF-beta mRNA and protein in cultured normal human LEC. The results showed that a single RT-PCR amplified product about 310bp was obtained, and the sequence was homologous to the known sequence. TGF-beta immunostain was positive in the plasma of LEC. It was suggested that normal human LEC could produce TGF-beta, and LEC could be affected by TGF-beta through autocrine action.
Cells, Cultured ; Epithelial Cells ; cytology ; metabolism ; Humans ; Lens, Crystalline ; cytology ; metabolism ; RNA, Messenger ; biosynthesis ; genetics ; Transforming Growth Factor beta ; biosynthesis ; genetics