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Kerstin Birke, Matthias Zenkel, Friedrich E. Kruse, Ulrich-Christoph Welge-Luessen, Anselm G. Juenemann; The Migratory Effect Of Homocysteine On Primary Human Astrocytes. Invest. Ophthalmol. Vis. Sci. 2011;52(14):2455.
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An increased homocysteine level in the blood plasm is a risk factor for primary open angle glaucoma (POAG). Moreover, in POAG patients the homocysteine levels are increased in the aqueous humor. In POAG degenerations of the optic nerve axons are a severe implication. The degenerative processes are often associated with astrocytosis or increased migration of astrocytes. At present it is not clarified which process is cause or effect. Therefore, the direct effect of homocysteine on primary human astrocytes was investigated in vitro.
Primary astrocytes were isolated from 5 donors. Cultures of passages 3 to 5 were treated with 0, 2 and 3mM homocysteine in DMEM/F12 supplemented with 0.1% FCS for 0-114h. The proliferation rate of astrocytes was analyzed with a BrdU-kit. Metabolic activity was analyzed photometrical by use of a CCK-8-kit. The migration was observed by time-lapse video microscopy over 18h. Expression levels of MMP2, MMP9, TIMP1-3, Elastin and Hsp27 mRNA were analyzed and quantified by real-time PCR after 18h treatment. Changes on the protein expression level, of the described factors were analyzed by immunofluorescence microscopy.
Treatment with homocysteine had no influence on the proliferation rate of astrocytes when compared with controls. After 18h the metabolic activity was increased 5-6 fold as compared with untreated astrocytes. By time-lapse video microscopy an increased migration of treated astrocytes was observed. The mRNA expression of MMP2, MMP9 and Hsp27 was increased and the mRNA expression of TIMP1-3 and Elastin was decreased after 18h treatment when compared with controls. By immunofluorescence an augmented Hsp27 labeling was observed after 18h treatment when compared with controls.
Due to its migration-stimulating capacity demonstrated in primary human astrocytes, homocysteine might be involved in increasing migration of astrocytes also in POAG.
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