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Lens epithelial cell apoptosis and intracellular Ca2+ increase in the presence of xanthurenic acid.

Malina H, Richter C, Frueh B, Hess OM - BMC Ophthalmol (2002)

Bottom Line: In the control cells gelsolin stained the perinuclear region, whereas in the presence of 10 microM xanthurenic acid gelsolin is translocated to the cytoskeleton, but does not lead to cytoskeleton breakdown.At low (5 to 10 microM) of xanthurenic acid concentration, the elongation of the cytoskeleton was associated with migration of mitochondria and cytochrome c release.We observed an induction of calpain Lp 82 and an increase of free Ca2+ in the cells in a xanthurenic acid concentration-dependent manner.

View Article: PubMed Central - HTML - PubMed

Affiliation: Swiss Cardiovascular Research Center, Inselspital, CH-3010 Bern, Switzerland. halina.malina@dkf2.unibe.ch

ABSTRACT

Background: Xanthurenic acid is an endogenous product of tryptophan degradation by indoleamine 2,3-dioxygenase (IDO). We have previously reported that IDO is present in mammalian lenses, and xanthurenic acid is accumulated in the lenses with aging. Here, we studied the involvement of xanthurenic acid in the human lens epithelial cell physiology.

Methods: Human lens epithelial cells primary cultures were used. Control cells, and cells in the presence of xanthurenic acid grow in the dark. Western blot analysis and immunofluorescence studies were performed.

Results: In the presence of xanthurenic acid human lens epithelial cells undergo apoptosis-like cell death. In the control cells gelsolin stained the perinuclear region, whereas in the presence of 10 microM xanthurenic acid gelsolin is translocated to the cytoskeleton, but does not lead to cytoskeleton breakdown. In the same condition caspase-3 activation, and DNA fragmentation was observed. At low (5 to 10 microM) of xanthurenic acid concentration, the elongation of the cytoskeleton was associated with migration of mitochondria and cytochrome c release. At higher concentrations xanthurenic acid (20 microM and 40 microM) damaged mitochondria were observed in the perinuclear region, and nuclear DNA cleavage was observed. We observed an induction of calpain Lp 82 and an increase of free Ca2+ in the cells in a xanthurenic acid concentration-dependent manner.

Conclusions: The results show that xanthurenic acid accumulation in human lens epithelial cells disturbs the normal cell physiology and leads to a cascade of pathological events. Xanthurenic acid induces calpain Lp82 and caspases in the cells growing in the dark and can be involved in senile cataract development.

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Staining of the mitochondria (Mito-Tracker Red CMXRos), nucleus (Hoechst) and cell membranes (DiOC18) in HuLEC grown in the presence of xanthurenic acid for 96 hours: (A-C), control cells: (A) mitochondria, (B) nucleus, (C) membranes; (D-F) with 10 μM xanthurenic acid: (D) mitochondria, (E) membranes, (F) merge of D, E, and nucleus stained by Hoechst; (G-I) with 20 μM xanthurenic acid: (G) mitochondria, (H) merge of (G) and Hoechst staining, (I) membranes
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Figure 6: Staining of the mitochondria (Mito-Tracker Red CMXRos), nucleus (Hoechst) and cell membranes (DiOC18) in HuLEC grown in the presence of xanthurenic acid for 96 hours: (A-C), control cells: (A) mitochondria, (B) nucleus, (C) membranes; (D-F) with 10 μM xanthurenic acid: (D) mitochondria, (E) membranes, (F) merge of D, E, and nucleus stained by Hoechst; (G-I) with 20 μM xanthurenic acid: (G) mitochondria, (H) merge of (G) and Hoechst staining, (I) membranes

Mentions: The cell membranes were stained with DiOC18 and co-stained with Mito-Tracker Red CMXRos, and the nucleus was stained with Hoechst 33342 (Fig. 6). In the control cell mitochondria were in the perinuclear region and the cell membranes were uniformly stained with DiOC18 (Fig. 6A,6B,6C) In the presence of 10 μM xanthurenic acid the mitochondria migrated to the cell periphery and the cell membranes were not uniformly stained (Fig. 6D,6E,6F). In the presence of 20 μM of xanthurenic acid the mitochondrial structure was destroyed, nuclear DNA was degraded and membranes were not stained with DiOC18 (Fig. 6G,6H,6I).


Lens epithelial cell apoptosis and intracellular Ca2+ increase in the presence of xanthurenic acid.

Malina H, Richter C, Frueh B, Hess OM - BMC Ophthalmol (2002)

Staining of the mitochondria (Mito-Tracker Red CMXRos), nucleus (Hoechst) and cell membranes (DiOC18) in HuLEC grown in the presence of xanthurenic acid for 96 hours: (A-C), control cells: (A) mitochondria, (B) nucleus, (C) membranes; (D-F) with 10 μM xanthurenic acid: (D) mitochondria, (E) membranes, (F) merge of D, E, and nucleus stained by Hoechst; (G-I) with 20 μM xanthurenic acid: (G) mitochondria, (H) merge of (G) and Hoechst staining, (I) membranes
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC103667&req=5

Figure 6: Staining of the mitochondria (Mito-Tracker Red CMXRos), nucleus (Hoechst) and cell membranes (DiOC18) in HuLEC grown in the presence of xanthurenic acid for 96 hours: (A-C), control cells: (A) mitochondria, (B) nucleus, (C) membranes; (D-F) with 10 μM xanthurenic acid: (D) mitochondria, (E) membranes, (F) merge of D, E, and nucleus stained by Hoechst; (G-I) with 20 μM xanthurenic acid: (G) mitochondria, (H) merge of (G) and Hoechst staining, (I) membranes
Mentions: The cell membranes were stained with DiOC18 and co-stained with Mito-Tracker Red CMXRos, and the nucleus was stained with Hoechst 33342 (Fig. 6). In the control cell mitochondria were in the perinuclear region and the cell membranes were uniformly stained with DiOC18 (Fig. 6A,6B,6C) In the presence of 10 μM xanthurenic acid the mitochondria migrated to the cell periphery and the cell membranes were not uniformly stained (Fig. 6D,6E,6F). In the presence of 20 μM of xanthurenic acid the mitochondrial structure was destroyed, nuclear DNA was degraded and membranes were not stained with DiOC18 (Fig. 6G,6H,6I).

Bottom Line: In the control cells gelsolin stained the perinuclear region, whereas in the presence of 10 microM xanthurenic acid gelsolin is translocated to the cytoskeleton, but does not lead to cytoskeleton breakdown.At low (5 to 10 microM) of xanthurenic acid concentration, the elongation of the cytoskeleton was associated with migration of mitochondria and cytochrome c release.We observed an induction of calpain Lp 82 and an increase of free Ca2+ in the cells in a xanthurenic acid concentration-dependent manner.

View Article: PubMed Central - HTML - PubMed

Affiliation: Swiss Cardiovascular Research Center, Inselspital, CH-3010 Bern, Switzerland. halina.malina@dkf2.unibe.ch

ABSTRACT

Background: Xanthurenic acid is an endogenous product of tryptophan degradation by indoleamine 2,3-dioxygenase (IDO). We have previously reported that IDO is present in mammalian lenses, and xanthurenic acid is accumulated in the lenses with aging. Here, we studied the involvement of xanthurenic acid in the human lens epithelial cell physiology.

Methods: Human lens epithelial cells primary cultures were used. Control cells, and cells in the presence of xanthurenic acid grow in the dark. Western blot analysis and immunofluorescence studies were performed.

Results: In the presence of xanthurenic acid human lens epithelial cells undergo apoptosis-like cell death. In the control cells gelsolin stained the perinuclear region, whereas in the presence of 10 microM xanthurenic acid gelsolin is translocated to the cytoskeleton, but does not lead to cytoskeleton breakdown. In the same condition caspase-3 activation, and DNA fragmentation was observed. At low (5 to 10 microM) of xanthurenic acid concentration, the elongation of the cytoskeleton was associated with migration of mitochondria and cytochrome c release. At higher concentrations xanthurenic acid (20 microM and 40 microM) damaged mitochondria were observed in the perinuclear region, and nuclear DNA cleavage was observed. We observed an induction of calpain Lp 82 and an increase of free Ca2+ in the cells in a xanthurenic acid concentration-dependent manner.

Conclusions: The results show that xanthurenic acid accumulation in human lens epithelial cells disturbs the normal cell physiology and leads to a cascade of pathological events. Xanthurenic acid induces calpain Lp82 and caspases in the cells growing in the dark and can be involved in senile cataract development.

Show MeSH
Related in: MedlinePlus