PURPOSE: The purpose of this study was to examine the molecular basis for steroid-induced cataract and the specific aim was to investigate the potential effects of glucocorticoids on modulation of alpha-crystallin gene expression in lens epithelial cells. METHOD: Bovine lens epithelial explants, B-3 human lens epithelial cells, and alphaTN4 mouse lens epithelial cells were incubated in the absence or presence of dexamethasone. At the indicated time point, total cellular RNA was isolated and subjected to RT-PCR and Northern blot analysis to assess mRNA expression of alphaA- and alphaB-crystallin. RESULTS: The mRNA expression of alphaA-crystallin was drastically downregulated by treatment with dexamethasone in bovine, human, and mouse lens epithelial cells. The mRNA level of alphaA-crystallin was also decreased in response to hydrocortisone but not to estrogen. The stability of the alphaA-crystallin mRNA was decreased after dexamethasone treatment. CONCLUSIONS: These results suggest that the alteration of alphaA-crystallin gene expression by glucocorticoids may result in abnomal structure and differentiation of lens epithelial cells on steroid-induced cataract formation.
alpha-Crystallins* ; Animals ; Blotting, Northern ; Cataract ; Dexamethasone ; Epithelial Cells* ; Estrogens ; Gene Expression* ; Glucocorticoids* ; Humans ; Hydrocortisone ; Mice ; RNA ; RNA, Messenger

The purpose of this study was to find out effects of treatment of ginsenoside Re, Rc and EGCG on mRNA expressions of leptin, hormone sensitive lipase (HSL) and resistin in 3T3-L1 adipocytes. The concentrations of EGCG were treated with 0.01 x 10(-7), 0.1 x 10(-7), 1 x 10(-7) and 1 x 10(-6) M or 100 microgram/ml ginsenoside Re, Rc in culture cell for 13 days. mRNA expression of leptin wasn't expressed in preadipocyte but according to differentiation of adipocyte, the that of mRNA expression was decreased at gensenosids or EGCG treated cells compared with non treated adipocyte. Expression of HSL mRNA was increased in G-Re, G-Rc and EGCG treated cells compared with non treated cells. The resistin level was significantly decreased in adipocytes treated with G-Re, G-Rc and EGCG. These pattern was similar to leptin expression.These results support that treatment of gensenosides or EGCG in 3T3-L1 adipocyte resulted to affect of leptin and resistin as well as HSL mRNA levels, accordingly, levels of leptin and HSL will be acted by signalling body fat stores to the hypothalamus which in turn regulates food intake and energy expenditure to maintain body weight homeostasis. And also regulation of resistin mRNA will prevent to diabetics attacked with obesity. In conclusion, we suggest that consumption of ginseng saponine or EGCG might prevent human diabetics or/and obesity.
3T3-L1 Cells ; Adipocytes* ; Adipose Tissue ; Body Weight ; Catechin* ; Eating ; Energy Metabolism ; Homeostasis ; Humans ; Hypothalamus ; Leptin* ; Obesity ; Panax* ; Resistin* ; RNA, Messenger* ; Saponins ; Sterol Esterase* ; Tea*

An after-cataract is caused by the proliferation of residual cells over the equator of the lens. These cells subsequently migrate to the posterior lens capsule, where they undergo aberrant differentiation into fiber-like cells or transdifferentiation into fibroblast-like cells. To study the precise molecular mechanisms of transdifferentiation, an attempt was made to establish an in vitro system, in which the lens epithelial cells (LECs) of the pre-equatorial zone could be transdifferentiated into fibroblast-like cells. The required conditions for culturing the LECs were identified as consisting of four phases; intact bovine explants, explant-cultured, serum-modulated and additionally modulated LECs. The LECs of each phase were compared by examining changes in the expression of the epithelial-mesenchymal transition (EMT) -related genes and changes in cellular morphology and adhesion. The explants that were cultured in a medium containing 10% fetal bovine serum (FBS) for 2 weeks, showed changes in the expression of the EMT-related genes, although the other explant-cultured cells maintained an epithelial morphology. To introduce a transition into mesenchymal cells, the explant cultures were subcultured in a medium containing 20% FBS for six passages. These cells displayed an elongated morphology and were able to grow and migrate in a similar way to fibroblast cells. The expression of the EMT-related genes, such as, extracellular matrix proteins and integrins, was altered. This was similar to the alteration of the 3-dimensional collagen gels model previously reported. During a further process of EMT by additional serum modulation, the inhibitory effect of disintegrin on cell adhesion was gradually decreased, integrin expression was differentially regulated and alpha-smooth muscle actin was post-translationally modified from the point of passage number six. Overall, it can be concluded that terminal transdifferentiation accompanies changes in the cytoskeletal proteins and cell surface molecules. These are modulated in systematic patterns of post-transcriptional and post-translational regulation and patterns of gene regulation, by the synergic effects of several transforming factors contained in serum. Therefore, posterior capsular opacification may also be accompanied by this molecular mechanism.
Animals ; Blood Proteins/*pharmacology ; Cattle ; Cell Adhesion/drug effects ; Cell Differentiation/drug effects/physiology ; Cells, Cultured ; Epithelial Cells/*cytology/physiology ; Fibroblasts/*cytology/physiology ; Gene Expression/drug effects ; Lens, Crystalline/*cytology ; Support, Non-U.S. Gov't ; alpha-Crystallin A Chain/genetics ; alpha-Crystallin B Chain/genetics

Crystallins are the major proteins found in the lens, and the localization of specific crystallins is well known. Overexpression and accumulation of alphaB-crystallin has been observed in response to stress conditions or in certain diseases, such as brain tumors and neurodegenerative diseases. The purpose of this study was to examine whether alpha-crystallins are modified during pathological myofibroblastic changes in lens epithelial cells. Lens epithelial cells attached to the anterior capsules of patients with nuclear or anterior polar cataracts were analyzed quantitatively for alpha-crystallin proteins and mRNAs using Western blot and RT-PCR analysis., respectively. The degree of modification of alpha-crystallins was determined by 2-dimensional gel electrophoresis followed by Western blotting. Higher molecular weight protein bands that were immunoreactive to anti-alphaA- and anti-alphaB-crystallin antibodies around 45 kDa accumulated more in the anterior polar cataract samples than in those with the nuclear type of cataracts. Also monomeric alphaB-crystallins accumulated more in lens epithelial cells of patients with anterior polar cataracts. By comparison, no significant changes were found in the levels of the mRNAs encoding alphaA- and alphaB-crystallins in the different types of cataracts. Both alphaA- and alphaB-crystallin proteins seemed to undergo more extensive modification in anterior polar cataracts. Conclusion. In addition to fibrotic changes, which accompany increased levels of extracellular matrix molecules, accumulation and abnormal modification of alpha-crystallins might be implicated in the pathogenic mechanism of this type of cataract.
Adult ; Cataract/*genetics/metabolism ; Epithelial Cells/metabolism ; Female ; Human ; Lens, Crystalline/metabolism ; Male ; Middle Aged ; RNA, Messenger/analysis ; Reverse Transcriptase Polymerase Chain Reaction ; Support, Non-U.S. Gov't ; alpha-Crystallin A Chain/*genetics/metabolism ; alpha-Crystallin B Chain/*genetics/metabolism