Hypoxic repression of lactate dehydrogenase-B in retina.
In order to elucidate mechanisms controlling hypoxia induced gene repression in the retina, we studied expression of the lactate dehydrogenase gene LDH-B. RT/PCR was used to isolate cDNA fragments of the LDH-A and LDH-B genes from both rat and chick. Northern analysis was used to measure LDH-B mRNA expression and in situ hybridization to localize LDH-B mRNA during development of the avascular chick retina. Primary retinal cultures of rat and chick were subjected to hypoxia and reperfusion paradigms, and levels of LDH-A and LDH-B mRNA expression were measured by Northern and semi-quantitative RT/PCR analyses. Northern analysis demonstrated that LDH-B mRNA is developmentally regulated in the chick retina according to a time course that correlates with oxygen availability. In developing chick retina in situ hybridization localized LDH-B mRNA to aerobic regions of the retina, coincident with the distribution of active mitochondria. Northern and semi-quantitative RT/PCR analyses demonstrated that LDH-B mRNA levels decrease in primary rat and chick retinal cells in response to hypoxia. LDH-B mRNA increased to control levels in retinal cells exposed to hypoxia then reperfused with oxygen, while the opposite was true for LDH-A, an hypoxia inducible gene. These data demonstrate that LDH-B gene repression after hypoxia and reactivation after oxygen reperfusion occurs in both vascular and avascular retinal cells. Oxygen regulated expression of LDH-B is opposite and complementary to that of LDH-A and provides a system to elucidate the mechanisms underlying hypoxic gene repression and reactivation after reperfusion. Mechanisms controlling gene repression and reactivation based on oxygen availability are likely to be critically involved in ischemic damage and neovascular retinopathy.
Buono
R J
RJ
Department of Pathology, Anatomy, and Cell Biology, Jefferson Medical College, 1020 Locust St., Philadelphia, PA, 19107, USA.
Lang
R K
RK
D000818:Animals; D015152:Blotting, Northern; D002478:Cells, Cultured; D002642:Chick Embryo; D005786:Gene Expression Regulation; D005865:Gestational Age; D000860:Hypoxia / Q000201:enzymology*; D017403:In Situ Hybridization; D007527:Isoenzymes; D007770:L-Lactate Dehydrogenase / Q000235:genetics*; D010477:Perfusion; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D012160:Retina / Q000196:embryology / Q000201:enzymology*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D007527:Isoenzymes; D012333:RNA, Messenger; D007770:L-Lactate Dehydrogenase