Mechanosensory hair cells and supporting cells develop from common precursors located in the prosensory domain of the developing cochlear epithelium. Prosensory celldifferentiation into hair cells or supporting cells proceeds from the basal to the apical region of the cochleae, but the mechanism and significance of this basal-to-apical wave of differentiation remain to be elucidated. Here, we investigated the role of Hedgehog (Hh) signaling in cochlear development by examining the effects of up- and downregulation of Hh signaling in vivo . The Hh effector smoothened (Smo) was genetically activated or inactivated specifically in the developing cochlear epithelium after prosensory domain formation. Cochleae expressing a constitutively active allele of Smo showed only one row of inner hair cells with no outer hair cells (OHCs); abnormal undifferentiated prosensory-like cells were present in the lateral compartment instead of OHCs and their adjacent supporting cells. This suggests that Hh signaling inhibits prosensory celldifferentiation into hair cells or supporting cells and maintains their properties as prosensory cells. Conversely, in cochlea with the Smo conditional knockout ( Smo CKO), haircelldifferentiation was preferentially accelerated in the apical region. Smo CKO mice survived after birth, and exhibited haircell disarrangement in the apical region, a decrease in haircell number, and hearing impairment. These results indicate that Hh signaling delays haircell and supporting celldifferentiation in the apical region, which forms the basal-to-apical wave of development, and is required for the proper differentiation, arrangement and survival of hair cells and for hearing ability.
Fig. 1. Schematic diagram of development of the chick inner ear and its sensory patches. (A) The inner ear of a bird originates by invagination of the otic placode and remodelling of the resulting otic vesicle to form the labyrinth. The mature structure contains seven vestibular sensory patches, involved in perception of gravity and acceleration: three cristae (in the ampullae of the semicircular canals) and four maculae, those of the utricle, the saccule, the lagena, and the macula neglecta (which we neglect). The basilar papilla, an elongated sensory region extending along the cochlear duct, serves auditory function. (B) The patterns of Notch ligand expression in a sensory patch before, during and after hair-celldifferentiation. One of the earliest genes to be expressed in prospective sensory patches codes for the Notch ligand Ser1. Within the Ser1 domains, nascent hair cells expressing Delta1 can be detected from E3.5 in vestibular regions, and from E5 in the cochlear duct. Delta1 expression foreshadows the differentiation of hair cells, which become identifiable by morphological and other molecular criteria about 24 hours later. Notch1 itself is expressed in sensory as well as non- sensory regions of the developing otocyst.
now show that combining Neurog1 in one allele with removal of floxed Atoh1 in a self-terminating conditional mutant (Atoh1-Cre; Atoh1 f/kiNeurog1 ) mouse results in significantly more differentiated inner HCs and outer HCs that have a prolonged longevity of 9 months compared with Atoh1 self-terminating littermates. Stereocilia bundles are partially disorganized, disoriented and not HC type specific. Replacement of Atoh1 with Neurog1 maintains limited expression of Pou4f3 and Barhl1 and rescues HCs quantitatively, but not qualitatively. OC patterning and supporting celldifferentiation are also partially disrupted. Diffusible factors involved in patterning are reduced (Fgf8) and factors involved in cell-cell interactions are affected (Jag1, Hes5). Despite the presence of many HCs with stereocilia these mice are deaf, possibly owing to HC and OC patterning defects. This study provides a novel approach to disrupt OC development through modulating the HC-specific intracellular TF network. The resulting disorganized OC indicates that normally differentiated HCs act as ‘ self-organizers ’ for OC development and that Atoh1 plays a crucial role to initiate HC stereocilia differentiation independently of HC viability.
Suppressing the activating cleavage of Notch with a ␥ -secretase inhibitor modulates Notch signaling and induces haircelldifferentiation. DAPT has been shown to regulate ␥ -secretase in a dose-dependent manner, and hence Notch signaling in mouse kidney (Cheng et al., 2003) and thymus cultures (Doerfler et al., 2001; Hadland et al., 2001), and in whole animals (Dovey et al., 2001; Geling et al., 2002). Although ␥ -secretase modulates the activation of several proteins (Kimberly and Wolfe, 2003), inhibition of ␥ - secretase produces phenotypes similar to those of Notch mutations in zebrafish (Geling et al., 2002) and Drosophila (Micchelli et al., 2003), strongly suggesting that the disruption of Notch signaling is the dominant effect of DAPT treatment. Using increasing concentrations of DAPT in cochleae explants, we have discovered that Notch regulates haircelldifferentiation in a dose-dependent manner in the mouse cochlea. The concentrations of DAPT used were not toxic and did not seem to modify levels of genes non- specifically, as the expression levels of ␥ -tubulin, COUP-TFI and COUP-TFII were unaltered. Pharmacological suppression of Notch activation with a different ␥ -secretase inhibitor demonstrated a Notch-dependent regulation of excess haircell development, primarily in the middle to basal turns (Yamamoto et al., 2006). It is unclear whether this difference of where in the duct excess hair cells differentiate is due to the different ␥ -secretase inhibitor used, the stage of culture, or both. Importantly, high doses of DAPT further attenuated Notch signaling to produce a greater number of hair cells in the apex in COUP-TFI –/– than in wild-type cochleae. Consistent with our results, genetic redundancy and gene-dose effects of Notch signaling components have been documented for several Notch signaling mutants that produce excess hair cells in the organ of Corti
Since the VDR is widely expressed and is missing from all cells of the VDR-null mice, the organ and cell type in which VDR acts to maintain normal hair cycling is uncertain. To identify the cell population responsible for the defect and rule out a systemic cause for the alopecia, hair-reconstitution assays were performed in nude mice. Implantation of a mixture of keratinocytes and activat- ed dermal papilla cells into a nude mouse host recapitu- lates the process of hair follicle morphogenesis (17), lead- ing to a functional hair follicle. The use of activated (early anagen) dermal papilla cells is critical, since these cells do not retain their ability to induce the formation of functional hair follicles during all stages of the hair cycle. By implanting wild-type or VDR-null activated dermal papilla cells with wild-type or VDR-null ker- atinocytes, we were able to obtain hair follicles with epi- dermal and mesodermal components differing in VDR status (VDR +/+ versus VDR –/– ) and analyze which cell
be responsible for the increased timing to the basilar membrane. Thus, the medial OC efferents would exert their effect mechanically. Sridhar et al (1995) reported an additional slower response o f sound-evoked nerve activity that is also efferent- modulated. They propose that the slow effect is attributable to release o f calcium from the subsurface cistemae o f the OHC, causing calcium-gated potassium channels to open and leading to hyperpolarisation o f the haircell on a much larger time-scale (Sridhar et al, 1997). Reiter and Liberman (1995) believe that protection against temporary threshold shifts caused by high-level tones is correlated with the OC slow effect. Efferent stimulation also reduces the level o f otoacoustic distortion products (Mountain, 1980; Siegel and Kim, 1982). Otoacoustic emissions (OAEs) are low- intensity sounds, produced specifically by the cochlea and, most probably, by the cochlear outer hair cells as they undergo fast, reversible length changes. Changes in their amplitude as a result o f acoustic overstimulation reflect structural and fimctional damage to the hearing organ.
Using auditory evoked potential audiometry (AEP), we examined the effects of sex and reproductive condition on hearing ability in the round goby. In addition, we related female hearing measurements to plasma 17β-estradiol (E2) – a crucial hormone for reproductive development in female fishes (Kime, 1993) – and tested whether variation in hearing ability in both sexes was associated with haircell density in the saccule, the primary auditory end organ in fishes (Popper and Fay, 1999) [but see Lu et al. for roles of utricle and lagena (Lu et al., 2003; Lu et al., 2004)]. Auditory threshold, suprathreshold response amplitude and suprathreshold latency were measured in response to tones and a single-pulse round goby vocalization. We predicted that females would have superior hearing ability relative to males (lower thresholds, higher response amplitudes, shorter latencies), and that plasma E2 level would correlate positively with enhanced auditory phenotypes in females. No specific predictions were made about the effect of reproductive condition on hearing ability in males because while heightened sensitivity to neighboring male vocalizations could be advantageous for assessment purposes, it could also conceivably increase the probability of engaging in costly agonistic behaviors.
example, two recent studies (Hawkins et al., 2003; Hawkins et al., 2007) used genomic profiling to identify a large number of genes that are altered in the avian auditory and/or vestibular epithelium after HC damage. This characterization was accomplished with gene microchips, which define relative levels of tens of thousands of transcripts in a given tissue and allow one to compare levels of expression across tissue samples. Microchip analyses enable the identification of molecular markers and signaling pathways here- tofore unknown to be relevant for HC regeneration. These studies have confirmed differential expression levels of some candidate genes using quantitative polymerase chain reaction and in situ hybridization, which provides additional assurance of the validity of the approach. Follow-up technologies must now be developed to enable investigators to accept or reject hypotheses regarding the role of specific candidate genes in HC regeneration. Two examples of these are gene knock-down and gene misexpression. Inhibition of gene function through knockdown can be accom- plished with RNA interference or RNA antisense approaches. Gene misexpression can be accomplished through delivery of conditionally or constitutively active genes into cells of interest. To our knowledge, no studies involving genetic perturbation in the post-embryonic amphibian or avian inner ear have been per- formed. However, delivery of modified nucleic acids into the developing chicken otocyst has been accomplished using retrivirus (e.g., Fekete et al., 1998), electroporation (e.g., Daudet and Lewis, 2005) and antisense morpholinos (Gerlach-Bank et al., 2004). The challenge is now to bring these methods for gene transfection and transduction into tissues of the mature inner ear. Another new area of study in the genetics of HC loss and regeneration has recently been developed in zebrafish, a classic model for genetics experiments. Hair cells of the lateral line neuromast are positioned
Trichoepithelioma is the most common benign tumor of the hair follicle origin in the present study with microscopic features showing basaloid cells (like cylindroma that form primi- tive hair follicle germ structures with fibromyxoid stroma. Cells are often in fronds, may have papillary mesenchymal bodies (Figure 1). Pilomatrixoma on microscopy showing biphasic pattern of keratinized ghost cells and basaloid cells. Case also showed giant cell reaction (Figure 2). Trichofolliculoma on microscopy revealed dilated hair follicle arising from surface epithelium with numerous secondary hair follicles arising from it (Figure 3). Sebaceous carcinoma on microscopy showing large pleomorphiccells with multivacuolated clear cytoplasm and at a foci showing squamoid differentiation is seen (Figure 4).
split hair bundles, suggesting that hair cells employ redundant mechanisms for maintaining proper hair bundle morphology. During early postnatal development, the apical region of the haircell undergoes a shape change driven by actomyosin forces (Etournay et al., 2010). Also during this time period, densely bundled rootlet structures form at the base of the stereocilia to anchor them into the cuticular plate, an actin meshwork that provides rigid support (DeRosier and Tilney, 1989; Kitajiri et al., 2010). We suggest that the rootlets of the stereocilia and the cuticular plate serve as additional physical constraints to maintain the position of the basal body and the V-shape of the hair bundle in conjunction with Lis1-dynein. Lis1 is also required for positioning of the Golgi and mitochondria, as well as for haircell survival. Importantly, cell death is not limited to cells with abnormal hair bundles, suggesting that it is not merely a consequence of hair bundle defects. Further study is needed to understand these additional functions of Lis1-dynein.
mechanotransducer (MET) currents recorded from a control (A) and a gelsolin knockout (B) P6 apical-coil OHC. MET currents (bottom panels) were elicited by applying sinusoidal force stimuli to the hair bundles while changing the membrane potential between 2121 mV and +99 mV in 20 mV nominal increments from the holding potential of 281 mV (middle panel). For clarity only responses at 2121 mV are shown. The driver voltage (DV) signal of 640 V at 50 Hz to the fluid jet is shown above the traces (negative deflections of the DV are inhibitory). The arrows indicate the closure of the transducer channels, i.e. disappearance of the resting current, during inhibitory bundle displacements. Dashed lines indicate the holding or resting current. C, Peak-to-peak current-voltage curves were obtained from four control and three knockout OHCs (P6) using 1.3 mM extracellular Ca 2+ . The fits through the data are according to eqn.1 (see Methods) with values: control k = 494651, V r = 2.360.4 mV, V s = 4063 mV, and
Cell divisions and therefore replication errors occur in many tissues, and by analogy to species molecular clocks, a reliable somatic cell molecular clock should allow com- parisons among tissues. CSX tags have been previously sampled from human large and small intestinal crypts [3,5,21], which are also clonal structures . In contrast to hair, intestinal methylation increased throughout life, with average levels exceeding hair later in life (Figure 5). Intestinal methylation is also primarily sampled from dif- ferentiated cells and therefore its genealogy can be divided into three phases (neogenesis, stem cell and differentia- tion). In contrast to years of anagen, differentiated crypt cells only divide a few times and survive about a week. As in hair, average neogenesis and differentiation intervals are similar regardless of age, and therefore the age-related increase in intestinal methylation suggests crypt stem cells divide more often than bulge stem cells. Alternatively, other factors such as differences in environment or subtle differences in low-level gene expression (CSX is normally expressed only in the heart ) may influence how methylation changes with age. However, a recent study of human endometrium was also consistent with replication errors as a source for age-related CSX methylation because percent methylation correlated with likely numbers of menstrual cycles .
cells and mesenchyme with trace amounts present in the adult hair cells. Similarly, the limited expression o f cytokeratin in the developing sehsory epithelia decreases after E l 2.5 (Kuijpers et a l, 1991b). The transient co-expression o f vimentin and cytokeratin within the developing epithelia may play a role in differentiation and can be used as an early marker of differentiation (Viebahn, et a l, 1997). Glial filaments and desmin have not been observed at any point in the development o f hair cells in the guinea pig, rat or human (Raphael, 1987; Kuijpers et a l, 1991b; Oesterle et a l, 1990). More than most other cells, hair cells rely on actin. Actin is present in at least four regions of the haircell (Flock and Cheung, 1977; Slepecky and Chamberlin, 1982; Drenckhahn et a l, 1991). The majority of actin is found in the stereocilia, where it is present as bundles of parallel filaments that extend into the cuticular plate. Apically, it forms a circumferential ring of filaments associated with the adherens junctions at the reticular lamina. It is also a component of the cortical lattice just inside the cell membrane along the lateral wall and it is found in the cytoplasm (Holley et a l, 1992). Each o f these regions has different actin-binding proteins which include profilin (Flock et al., 1982), tropomyosin (Slepecky and Chamberlin, 1985b), spectrin (Holley and Ashmore, 1990; Ylikoski et a l, 1990), a-actinin (Slepecky and Savage, 1994), and fimbrin (Matsudaira et a l, 1983). Co-localisation studies show that fimbrin cross links the highly organised parallel actin filaments in the stereocilia o f both IHC’s and OHC’s (Drenckhahn et a l, 1991). This cross-linking contributes to the stiffness o f the stereocilia. In the rat, fimbrin is first noted at E l 8 in IHCs and by birth it is present in OHCs (Zine et a l, 1995). It is an essential part of the formation and ontogenesis of stereocilia.
Other transcription factors can be integrated into the hemangioblast GRN. Runx1 which is regulated by BMP signalling is essential for definitive hematopoiesis (Lacaud et al., 2002; Pimanda et al., 2007a). It starts to be expressed in the blast colony after 24 hours of colony growth; over the next 24 hours of blast colony development there is a great increase in the appearance of definitive progenitors (Cheng et al., 2008). The homeobox transcription factor Hex is expressed in BL-CFCs and acts to negatively regulate the levels of this progenitor (Kubo et al., 2005). In addition, it appears dispensable for primitive erythroid formation but required for differentiation of definitive hematopoi- etic and endothelial cells (Guo et al., 2003; Kubo et al., 2005). The Cdx genes, Cdx1 and Cdx4, are critical to the development of blood in the zebrafish system through their regulation of Hox genes that promote posterior/hematopoietic fate (Davidson et al., 2003; Davidson and Zon, 2006). The mouse Cdx genes, Cdx1 and Cdx4, show redundant functions in promoting posterior patterning through the regulation of posterior Hox genes (Lohnes, 2003; Subramanian et al., 1995; van den Akker et al., 2002; van Nes et al., 2006). They are expressed during EB differentiation in overlapping time windows which include the period of BL-CFC generation (Lengerke et al., 2008; McKinney-Freeman et al., 2008). When Cdx4 is overexpressed from day 2-4 of serum differentiation and during the blast colony assay it leads to an increase in BL-CFCs (Wang et al., 2005b). While its effect on BL- CFC numbers was not reported, overexpression of Cdx1 during the same time window leads to an increase in hematopoietic progenitors (McKinney-Freeman et al., 2008). Cdx1 or Cdx4 homozygous null mice do not appear to have altered hematopoi- etic development, however, ESCs deficient for either of these genes show reduced hematopoietic potential that can be further abrogated when the remaining Cdx genes including family mem- ber Cdx2 are reduced (Wang et al., 2008). These results point to the importance of the Cdx-hox pathway in regulating commitment and proliferation of the hematopoietic lineage.
which fails to bind estrogen but instead responds to the synthetic ligand 4-hydroxytamoxifen (Littlewood et al., 1995). In the absence of the ligand, the Cre-ER fusion protein is inactive. Upon tamoxifen binding, the receptor is released from the inhibited state, allowing the translocation of the activated Cre enzyme to the nucleus. Therefore, the marking of cells, which is controlled by the activation of the Cre enzyme, can be tightly regulated by the administration of the ligand tamoxifen. Temporal control of Cre alleles can also be achieved by using STOP signals in conjunction with constitutively active regulatory sequences upstream of a reporter gene (such as lacZ or GFP). In this approach, the reporter gene lies downstream of two Cre recombinase recognition (loxP) sites that are separated by a STOP codon. In a cell that has not experienced Cre activation, the STOP codon prohibits the production of the reporter protein (Fig. 3A). Upon Cre importation into the nucleus in the presence of tamoxifen, the STOP codon is excised and the reporter gene is expressed (Fig. 3B) (Danielian et al., 1998). A cell that undergoes such a Cre-mediated recombination event during its developmental history will remain permanently marked by the expression of the activated reporter gene. Importantly, this mark can be stably passed onto all of its progeny. Therefore, this method enables one to specifically mark the cells of interest at defined time-points to test the multipotency and longevity of a putative stem cell population, given that promoters specifically expressed in these populations are known (Fig. 4; Box 2).
A possible association between low levels of photo- exposure and a loss of immune tolerance ﬁ rst appeared when the association between vitamin D and multiple sclerosis was documented. 38 Although vitamin D does have immunoregulatory properties, 39 other immunoregula- tory molecules are generated on light exposure. In this hypothesis, we propose and review the evidence that FFA arises due to the collapse of immune privilege at the hair bulb because photo-protection provided by many facial care products reduces the synthesis of the tryptophan photopro- duct FICZ. Of relevance, FICZ is generated most ef ﬁ ciently by UVB but is also capable of being generated by UVA and visible light. 5 Low levels of FICZ are immunostimulatory with higher levels being immunosuppressive 6 which would support the proposition that UV protective facial products which allow the transmission of visible light but block UV light may have immunostimulatory properties.
Automatic Hair segmentation with better performance and higher accuracy is one of the important task. An algorithm for identity verification using only information from the hair. Face recognition in the wild (i.e., unconstrained settings) is highly useful in a variety of applications, but performance suffers due to many factors, e.g., obscured face, lighting variation, extreme pose angle, and expression. The proposed algorithm is a scholarly hair matcher utilizing shape, shading and surface highlights got from restricted fixes through an Ada-Boost method with avoiding frail classifiers when highlights are absent in the given area. A technique for double order utilizing NN(neural organize) which perform preparing a grouping on same information by utilizing HNN(heuristically prepared neural system).By adding simple parallel processing techniques the performance of HNN increased by 3.4%.