| The degeneration and loss of auditory hair cells is the major cause of deafness in humans. In mammals including humans, auditory hair cells are terminally differentiated cells and incapable of regeneration after loss caused by genetic or environmental factors. Some exciting progress has been made recently in the research on hair cell regeneration. Supernumerary outer and/or inner hair cells can be produced in embryonic cochlear explant cultures either following retinoic acid treatment or in normal serum-free medium, or in mice in which a specific gene is deleted. Such studies may have a significant therapeutic value. By applying regenerated hair cells to replace the lost or injured hair cells, hearing disorders may be eventually treated.Recently, the greater epithelial ridge (GER) of mammalian cochlea has attracted researchers' attention. Studies show that when forced to overexpress Math1, a positive regulator gene for hair cell differentiation, the GER cells of the postnatal rat cochlear turned into robust hair cells. Studies also show that some important genes and growth factor receptors were involved in the differentiation and regeneration of hair cells, in addition, these genes and receptors such as Hes1, Hes5, FGFR1 express in the GER. Thus, the GER is an important cell population and may be one of the progenitor cells pools of hair cells. So far we know little about the GER. In the first part of the experiment, we observe morphological development of the GER of rat cochleae at various stages.Hair cell regeneration is likely to repeat the differentiating process of hair cell in normal development. Thus, understanding the mechanisms of hair cell differentiation would be helpful to stimulate hair cell regeneration. Some specific genes can regulate the differentiating process of hair cells. Recent studies have suggested that p27Kipl, a cyclin-dependent kinase inhibitor (CDKI) that functions as an inhibitor of cell cycle progression, plays an essential role in development of the organ of Corti. In embryonic development, the cells expressed p27Kipl stop mitosis and exit cell cycle. p27Kipl are thought to be responsible for preventing auditory hair cells and supporting cells of mammals from re-entering the cell cycle. In the second part of the experiment, we observe p27Kipl expression in the GER of rat cochleae at various stages and investigate the relative mechanism of hair cell differentiation.Part one: Morphological change in the greater epithelial ridge of rat cochleae at various stagesIn order to observe morphological development of the greater epithelial ridge (GER), we selected the rat cochleae in the embryonic day 14, 16, 18, 20 and postnatal day 0, 2, 5, 7, 10, 14 (E14, E16, E18, E20, P0, P2, P5, P7, P1, P14). After fixation and embedment, the cochlea samples were cut into 7 m sections with a cryostat. The sections were processed for Hematoxylin and Eosin staining. We compare the morphology of basal, middle and apical turn of the same one rat cochlea. We also compare the morphology of the same turns of rat cochleae at different stages.At E14, according to the spiral vessel, a landmark for thelocation of the primordial OC, we can found the thickening ventromedial region of the otocyst, in which the primordial OC is located. In apical turn, the thickening region has no change. In middle turn, the region adjacent to the GER becomes concave and forms the primordial OC. In basal turn, the primordial OC develop further: auditory hair cells and several supporting cells can just be recognizable. So we can conclude that differentiating and maturing of hair cells were developed from basal turn to apical turn and the latter had a delay of 1-2 days.Comparing the rat cochleae at E16, E20, PO, P14, we find that the cytoarchitecture rearrangement and cells reduction take place in the GER. We also find that these changes start from the GER' s luminal surface. As hair cells matured at P14, the GER disappears and forms the inner spiral sulcus. We can see the GER as a morphological symbol of the cochlea being im... |
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