Nan Hee Kim

Genistein inhibits aldose reductase activity and high glucose-induced TGF-β2 expression in human lens epithelial cells

Aldose reductase (AR) and TGF-beta have been implicated in the development of diabetic complications, such as cataracts. In an attempt to obtain potential agents for the prevention of diabetic cataracts from natural products, we purified genistein from the roots of Pueraria lobata and investigated its inhibitory effects upon AR activity and its antioxidant effects on rat lenses. The inhibition of AR activity by genistein increased in a dose-dependent manner and the opacities of lenses were significantly improved when treated with genistein. In addition, we determined the effects of genistein on mechanisms induced by exposure to high glucose in human lens epithelial (HLE-B3) cells. We found that genistein was able to reduce the expression of TGF-beta2, alphaB-crystallin, and fibronectin mRNAs in HLE-B3 cells that were cultured in high glucose conditions. In addition, a reduction in glutathione (GSH) levels and thiobarbituratic acid-reactive substances was observed. These results show that genistein is protective against lens opacity and also inhibits high glucose-mediated toxic effects in HLE-B3 cells. These effects are likely achieved by preventing AR and cellular oxidation; therefore, genistein may be a potential therapeutic agent for preventing and treating complications associated with diabetes mellitus, such as diabetic cataracts.

Methylglyoxal induces cellular damage by increasing argpyrimidine accumulation and oxidative DNA damage in human lens epithelial cells

Methylglyoxal (MGO) is a cytotoxic metabolite and modifies tissue proteins through the Maillard reaction, resulting in advanced glycation end products (AGEs), which can alter protein structure and functions. Several MGO-derived AGEs have been described, including argpyrimidine, a fluorescent product of the MGO reaction with arginine residues. Herein, we evaluated the cytotoxic role of MGO in human lens epithelial cell line (HLE-B3). HLE-B3 cells were exposed to 400 microM MGO in the present or absence of pyridoxamine for 24h. We then examined the formation of argpyrimidine, apoptosis and oxidative stress in HLE-B3 cells. In MGO-treated HLE-B3 cells, the accumulation of argpyrimidine was markedly increased, and caspase-3 and 8-hydroxydeoxyguanosine (8-OHdG) were highly expressed, which paralleled apoptotic cell death. However, pyridoxamine (AGEs inhibitor) prevented the argpyrimidine formation and apoptosis of MGO-treated HLE-B3 cells. These results suggested that the accumulation of argpyrimidine and oxidative DNA damage caused by MGO are involved in apoptosis of HLE-B3 cells.

Extract of Cassiae Semen and its major compound inhibit S100b-induced TGF-β1 and fibronectin expression in mouse glomerular mesangial cells

Non-enzymatic glycation reactions between reducing sugar and free reactive amino groups of protein lead to the formation of advanced glycation end products, which increase under conditions of aging or diabetes. A previous study showed that extracts of Cassiae Semen (CS), the seed of Cassia tora, had inhibitory activity on advanced glycation end products formation in vitro. To examine the pharmacological effects of a butanol-soluble extract of CS under conditions of diabetic nephropathy, we evaluated the expression of transforming growth factor-beta1 (TGF-beta1) and fibronectin, key mediators of diabetic nephropathy, in mouse glomerular mesangial cells cultured in the presence of S100b (a specific ligand for receptor of advanced glycation end products). CS inhibited S100b-induced TGF-beta1 and fibronectin expression in mouse mesangial cells by suppressing activation of Smad2/3, extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK), and oxidative stress. Moreover, CS suppressed nuclear factor-kappa B (NF-kappaB) activation in S100b-stimulated mouse mesangial cells. To identify the active compounds of CS, three major compounds, rubrofusarin-6-O-beta-d-gentiobioside (CS-A), toralactone-9-O-beta-d-gentiobioside (CS-B), and cassiaside (CS-C), were tested in cells. Of these compounds, CS-A significantly decreased the expression of TGF-beta1 and fibronectin and NF-kappaB DNA binding activity. These findings suggest that CS, especially CS-A, has potential as a preventive agent for advanced glycation end products-related diabetic complications.

Cytotoxic role of methylglyoxal in rat retinal pericytes: Involvement of a nuclear factor-kappaB and inducible nitric oxide synthase pathway

Methylglyoxal (MGO), a cytotoxic metabolite, is produced from glycolysis. Elevated levels of MGO are observed in a number of diabetic complications, including retinopathy, nephropathy and cardiomyopathy. Loss of retinal pericyte, a hallmark of early diabetic retinal changes, leads to the development of formation of microaneurysms, retinal hemorrhages and neovasculization. Herein, we evaluated the cytotoxic role of MGO in retinal pericytes and further investigated the signaling pathway leading to cell death. Rat primary retinal pericytes were exposed to 400muM MGO for 6h. Retinal vessels were prepared from intravitreally MGO-injected rat eyes. We demonstrated apoptosis, nuclear factor-kappaB (NF-kappaB) activation and inducible nitric oxide synthase (iNOS) induction in cultured pericytes treated with MGO and MGO-injected retinal vessels. In MGO-treated pericytes, TUNEL-positive nuclei were markedly increased, and NF-kappaB was translocalized into the nuclei of pericytes, which paralleled the expression of iNOS. The treatment of pyrrolidine dithiocarbamate (an NF-kappaB inhibitor) or l-N6-(1-iminoethyl)-lysine (an iNOS inhibitor) prevented apoptosis of MGO-treated pericytes. In addition, in intravitreally MGO-injected rat eyes, TUNEL and caspase-3-positive pericytes were significantly increased, and activated NF-kappaB and iNOS were highly expressed. These results suggest that the increased expression of NF-kappaB and iNOS caused by MGO is involved in rat retinal pericyte apoptosis.

Scopoletin from the flower buds of Magnolia fargesii inhibits protein glycation, aldose reductase, and cataractogenesis Ex Vivo

Five compounds previously known structures, scopoletin (1), northalifoline (2), stigmast-4-en-3-one (3), tiliroside (4), and oplopanone (5) were obtained from the flower buds of Magnolia fargesii using chromatographic separation methods. The structures of 1–5 were identified by the interpretation of their spectroscopic data including 1D- and 2D-NMR as well as by comparison with reported values. Three compounds 1–3 were found from M. fargesii for the first time in this study. All the isolates (1–5) were subjected to in vitro bioassays to evaluate the inhibitory activity on advanced glycation end products formation and rat lens aldose reductase (RLAR). Compound 1 showed a remarkable inhibitory activity on advanced glycation end products formation with IC50 value of 2.93 μM (aminoguanidine: 961 μM), and showed a significant RLAR inhibitory activity with IC50 value of 22.5 μM (3.3-tetramethyleneglutaric acid: 28.7 μM). Compound 4 exhibited potent inhibitory activity against RLAR (IC50 = 14.9 μM). In the further experiment ex vivo, cataractogenesis of rat lenses induced with xylose was significantly inhibited by compound 1 treatment.

KIOM-79 prevents methyglyoxal-induced retinal pericyte apoptosis in vitro and in vivo.

AIMS OF STUDY The purpose of this study was to investigate the protective effects of KIOM-79, a combination of four plant extracts, on retinal pericytes loss, which is one of the histopathological hallmarks of early diabetic retinopathy. MATERIALS AND METHODS To investigate the protective effect of KIOM-79 on pericyte apoptosis, we have used methylglyoxal (MGO)-treated primary rat retinal pericytes and retinal vessels from intravitreally MGO-injected eyes. Primary rat retinal pericytes were exposed to 400 microM MGO for 6h with or without KIOM-79. 400 microM final intravitreal concentration of MGO with or without KIOM-79 (10 microg/ml final intravitreal concentration) were intravitreally injected into the right eyes of rats for 2 days. The left eyes were received 3mul of saline only. Retinal vessels were then isolated. We examined apoptosis, ROS productions, 8-OHdG in cultured rat retinal pericytes and retinal vessels. RESULTS In CCK-8 assay and TUNEL staining, MGO-induced apoptosis of rat retinal pericytes was markedly inhibited by KIOM-79. KIOM-79 significantly reduced intracellular ROS productions and oxidative damage induced by MGO. In addition, the intravitreal injection of KIOM-79 prevented apoptosis of retinal microvascular cells, MGO accumulation and oxidative DNA damage in intravitreally MGO-injected eye. CONCLUSION These observations suggest that KIOM-79 acts through an antioxidant mechanism to protect against oxidative stress-induced apoptosis in retinal pericytes.

KIOM-79 prevents xylose-induced lens opacity and inhibits TGF-beta2 in human lens epithelial cells cultured under high glucose

AIM OF THE STUDY To investigate the effects of KIOM-79 in preventing the development of diabetic complications, such as cataracts. MATERIALS AND METHODS The inhibitory effects of KIOM-79 were assessed in a model of xylose-induced lens opacity and on changes mediated by high levels of glucose in human lens epithelial (HLE-B3) cells. RESULTS In lenses treated with KIOM-79, opacity was significantly improved and glutathione (GSH) was increased compared to controls. In HLE-B3 cells treated with KIOM-79, high glucose-mediated increases in TGF-beta2, alphaB-crystallin, and fibronectin were significantly inhibited in a dose-dependent manner. KIOM-79 decreased the phosphorylation of p-Smad2/3, pp38MAPK, pp44/42, and NF-kappaB signaling in cells grown under high glucose conditions. CONCLUSION KIOM-79 is protective against lens opacity and protects HLE-B3 cells from the toxic effects of high glucose. Therefore, KIOM-79 may provide a potential therapeutic approach for preventing diabetic complications, such as cataracts.