ABSTRACT
Title
Genistein protection in LHON disease: study of molecular mechanism in cybrid cell models
Authors
D. Catanzaro
Doctorate school in Pharmacological Science, course of Molecular and Cellular Pharmacology. Dept. of Pharmacology and Anaesthesiology – University of Padova, Italy
Doctorate school in Pharmacological Science, course of Molecular and Cellular Pharmacology. Dept. of Pharmacology and Anaesthesiology – University of Padova, Italy
Abstract
Leber's hereditary optic neuropathy (LHON), is a maternally inherited disease characterized by loss of central vision associated with mitochondrial DNA point mutations affecting complex I subunits of the respiratory chain. Over 95% of LHON cases are connected with one of three most frequent mtDNA mutations (G11778A, T14484C and G3460A) [1-2], but only 50% of men and 10% of women having one of this mutations, develop optic neuropathy. The incomplete penetrance and the male prevalence suggest that additional genetic (nuclear or mitochondrial) and/or environmental factors must modulate phenotype expression in LHON disease [3-4].
Cybrids obtained by fusion of donor enucleated fibroblasts and mtDNA-depleted osteosarcoma cells (rho0 cells), represent an useful experimental cell model to study LHON pathology and pharmacology excluding the influence of donor nuclear genome. Previous experiments have shown that estrogens might positively prevent oxidative stress damage in LHON cybrids [5] and preliminary data indicate that similar protection might be obtained by genistein, one of the soy isoflavones with estrogen-like activity.
In this study we investigated the molecular mechanism of genistein (0.1 µM), in cybrids incubated in basal condition and in galactose (glucose-free) medium, an experimental condition forcing the cells to rely mainly on the mitochondrial respiratory chain to produce ATP. Results show that in basal condition genistein induced the increase of p-ERK, of Mn-SOD expression and antioxidant enzymatic activity and also of mtDNA copies. Galactose incubation caused a decreased rate of the oxygen consumption, decreased total ATP content, increased apoptotic rate in LHON cybrids compared with control, and genistein at least in part prevented all these alterations. These results support our preliminary data and suggest that genistein by different molecular mechanism at mitochondrial level might be involved in the protection of LHON cell damage from oxidative stress.
Cybrids obtained by fusion of donor enucleated fibroblasts and mtDNA-depleted osteosarcoma cells (rho0 cells), represent an useful experimental cell model to study LHON pathology and pharmacology excluding the influence of donor nuclear genome. Previous experiments have shown that estrogens might positively prevent oxidative stress damage in LHON cybrids [5] and preliminary data indicate that similar protection might be obtained by genistein, one of the soy isoflavones with estrogen-like activity.
In this study we investigated the molecular mechanism of genistein (0.1 µM), in cybrids incubated in basal condition and in galactose (glucose-free) medium, an experimental condition forcing the cells to rely mainly on the mitochondrial respiratory chain to produce ATP. Results show that in basal condition genistein induced the increase of p-ERK, of Mn-SOD expression and antioxidant enzymatic activity and also of mtDNA copies. Galactose incubation caused a decreased rate of the oxygen consumption, decreased total ATP content, increased apoptotic rate in LHON cybrids compared with control, and genistein at least in part prevented all these alterations. These results support our preliminary data and suggest that genistein by different molecular mechanism at mitochondrial level might be involved in the protection of LHON cell damage from oxidative stress.
- Carelli Vet al.(2004) - Prog Retin Eye Res, 23: 53–89.
- Man PYet al.(2009) - J Med Genet, 46: 145–58.
- Phasukkijwatana Net al.(2010) - Hum Genet 2010, 128: 39–49.
- Kirkman MAet al.(2009) - Brain, 132: 2317–26.
- Giordano C, Montopoli M et al. (2011) – Brain, 134:220-34.