Keratin K has been identified

Keratin 8 (K8) has been identified as the first intermediate filament expressed during development in amphibians and mammals. In zebrafish, a type II cytokeratin 8 (zf-K8) cDNA has been cloned. zf-K8 gene was maternally inherited and expressed in all surface cells throughout the embryonic stages older than blastula. In these embryonic stages, zf-K8 was also expressed in presumptive intestinal cells and the pectoral fin, and in the adult, its expression was observed in the colorectal luminal cells and superficial cell layers of skin and fins, and scales (Imboden et al., 1997). Cyt-1 was reported as a novel type I cytokeratin, which was expressed in the outer enveloping layer in zebrafish embryos (Sagerstrom et al., 1996). We report the identification of a novel type I cytokeratin gene, dorsal aorta and pronephric duct keratin type-1 gene (DAPK-1) from zebrafish cDNA, and the expression in early dorsal aorta and pronephric duct. DAPK-1 cDNA (1.3 kb) contains an open reading frame for 431 amino acids. Comparative studies of amino IAA-94 receptor sequence revealed 87% identity between the goldfish keratin (GK) and the zebrafish DAPK-1. Application of DAPK-1 as a marker gene for dorsal aorta and pronephric duct also was discussed.
Materials and methods
Results and discussion
Acknowledgements
Introduction Stroke is a leading cause of mortality worldwide and is the leading cause of disability in the United States, representing a high-cost burden [1], [2]. Several experimental animal models of stroke are available in which reproducible injuries can be obtained through a combination of ischemia [3] and hypoxia [4], effectively mimicking the human condition. Ischemic stroke is caused by a disruption of blood flow that deprives the afflicted brain region of oxygen and nutrients, resulting in membrane depolarization and glutamate release [5]. Exposure to glutamate has been associated with a form of calcium-dependent delayed cell death linked to neurotoxicity [6], [7]. A mechanism of calcium-mediated neuronal death has been hypothesized in ischemia [8], central to which is the observation that hypoxia leads to the translocation of calcium from extracellular to intracellular spaces [9]. Death-associated protein kinase (DAPK) is an intracellular calcium/calmodulin (CaM)-regulated protein kinase (CaMK) whose mRNA levels increase in regions of tissue damage in an animal model of cerebral ischemia [10]. DAPK mRNA levels are also increased in the developing hippocampus [10], reaching maximal levels between embryonic day 22 and postnatal day 7. This represents the period of brain development characterized by apoptotic cell death [11], [12], [13], suggesting a role for DAPK in the regulation of apoptotic cell death during development. Consistent with this, cell culture models have demonstrated the involvement of DAPK in neuronal apoptosis that requires its intact catalytic domain [14], [15]. These findings raise the possibility that inhibition of the DAPK catalytic activity might forestall apoptotic neuronal death following ischemia. The targeting of DAPK in stroke is attractive, particularly because DAPK appears to function early in pathways of apoptosis [16], [17], potentially prior to the commitment of a cell to death. Changes in DAPK concentration or catalytic activity in response to cerebral ischemia have not been reported. This knowledge is required for a complete understanding of the role of DAPK in ischemia, and for future investigations into the potential of DAPK as a therapeutic target for the treatment of stroke. To examine the effect of hypoxic-ischemic injury on temporal patterns of DAPK activity in the brain, we used a well-characterized postnatal day 7 (P7) rodent model of perinatal cerebral hypoxia-ischemia (HI) [4], [18], an age approximately equivalent to the 34-week gestation human newborn [19]. We report here an increased DAPK specific activity in the hippocampus from the ischemic right cerebral hemisphere, compared to the uninjured left hemisphere, at a late time point following HI. This corresponds to a timeframe in which repair processes have already been initiated [20]. Based on these animal model results, we examined changes in DAPK activity during nerve growth factor (NGF)-induced differentiation of the PC12 cell line. We found an increase in DAPK catalytic activity and protein levels upon treatment with NGF, consistent with a potential role in neuronal differentiation or survival.