Hiro-oki Okamura

Six1 controls patterning of the mouse otic vesicle

Six1 is a member of the Six family homeobox genes, which function as components of the Pax-Six-Eya-Dach gene network to control organ development. Six1 is expressed in otic vesicles, nasal epithelia, branchial arches/pouches, nephrogenic cords, somites and a limited set of ganglia. In this study, we established Six1-deficient mice and found that development of the inner ear, nose, thymus, kidney and skeletal muscle was severely affected. Six1-deficient embryos were devoid of inner ear structures, including cochlea and vestibule, while their endolymphatic sac was enlarged. The inner ear anomaly began at around E10.5 and Six1 was expressed in the ventral region of the otic vesicle in the wild-type embryos at this stage. In the otic vesicle of Six1-deficient embryos, expressions of Otx1, Otx2, Lfng and Fgf3, which were expressed ventrally in the wild-type otic vesicles, were abolished, while the expression domains of Dlx5, Hmx3, Dach1 and Dach2, which were expressed dorsally in the wild-type otic vesicles, expanded ventrally. Our results indicate that Six1 functions as a key regulator of otic vesicle patterning at early embryogenesis and controls the expression domains of downstream otic genes responsible for respective inner ear structures. In addition, cell proliferation was reduced and apoptotic cell death was enhanced in the ventral region of the otic vesicle, suggesting the involvement of Six1 in cell proliferation and survival. In spite of the similarity of otic phenotypes of Six1- and Shh-deficient mice, expressions of Six1 and Shh were mutually independent.

Human CLC-KB Gene Promoter Drives the EGFP Expression in the Specific Distal Nephron Segments and Inner Ear

Human CLC-KB has been identified as a kidney-specific member of the CLC chloride channel family, and mutations of the human CLC-KB gene are known to cause Bartter syndrome type III. A precise understanding of the localization of this channel in the human kidney is imperative to our understanding of the pathophysiology, but this has remained unclear due to the high homology between human CLC-KB and CLC-KA, another kidney-specific member of the same family. The high intraspecies homology also rules out an exact correlation of the human isoforms (CLC-KA and CLC-KB) to the mouse and rat isoforms (CLC-K1 and CLC-K2, respectively). This study created transgenic mice harboring the enhanced green fluorescence protein (EGFP) gene driven by an 11-kbp human CLC-KB gene promoter. Three transgenic lines were generated, and all of them showed EGFP fluorescence in the kidney, with an identical pattern of localization to the thick ascending limb of Henles loop, distal tubules, connecting tubules, and intercalated cells of the collecting duct. This localization is exactly the same as that of mouse CLC-K2 identified in a previous report (Kobayashi et al. J Am Soc Neph 12: 1327-1334, 2001). EGFP fluorescence was also detected in the inner ear, more specifically in marginal cells of the stria vascularis and dark cells of the vestibular labyrinth, suggesting that human CLC-KB could play an important role in the fluid transport mechanism of the inner ear. The results (1) confirmed that CLC-KB is the true human homologue of rat and mouse CLC-K2 and (2) established that the 11-kbp human CLC-KB gene promoter is sufficient to elicit the precise expression in specific cell types of the kidney and inner ear.

Histopathological Examination of Ruptured Carotid Artery after Irradiation

We report 2 cases of carotid artery rupture after irradiation that was performed 1 year and 17 years before the ruptures. When irradiation-induced arterial rupture occurs, it usually does so within a few months following irradiation. However, the histopathological sections obtained in the present cases revealed carotid artery necrosis that was presumably induced by irradiation. Carotid artery rupture is sudden, massive hemorrhage that ranks among the most dreaded complications in the head and neck. However, several patients have been saved by hospital personnel who discovered the rupture in time to take appropriate measures such as cleaning of the wound and protection with myocutaneous or myofascial flaps. Therefore, it is important to be aware of the possibility of rupture or perforation of major vessels after irradiation, even when the radiation therapy was performed a long time ago.

Late laryngo-tracheal cartilage necrosis with external fistula 44 years after radiotherapy

Major late complications, following radiotherapy of head and neck carcinomas, such as laryngeal oedema, perichondritis and chondronecrosis usually occur between three and 12 months after treatment. However, the present case displayed necrosis of the laryngo-tracheal cartilage and ulceration of anterior neck skin with a tracheal fistula 44 years after irradiation. The reasons for the long interval between irradiation and late complications may be explained by long-standing hypovascularity and/or infection of the irradiated area. Histological study revealed chondronecrosis without inflammatory cells in the laryngo-tracheal cartilage and bacterial colonization of subcutaneous tissue. Necrotic tissue was removed and tracheostomy was performed. The fistula was almost completely closed using a delto-pectoral cutaneous flap and the clinical course of patient has been good. This paper demonstrates the possibility of laryngo-tracheal necrosis in cases that had received radiation as long ago as 44 years.

Localization of Carbonic Anhydrase in Guinea Pig Bowman's Glands

We examined the histochemical localization of carbonic anhydrase (CA) in Bowmans glands by light and electron microscopy. Neither CAI nor CAII was detected immunohistochemically in the duct cells. However, by enzyme histochemistry the duct cells revealed electron-dense precipitates demonstrative of CA in the microvilli and intercellular digitations. The reaction product was also noted in small vesicles in the cytoplasm of duct cells. In cells of the acini, the well-developed short microvilli, basolateral cell membrane, and mitochondria along the basolateral membrane showed strong deposits indicating CA activity. Dense reaction product of CA was also detected in a small core within the electron-lucent granules of the secretory cells, although CAI and CAII were not detected by immunostaining in the secretory granules. Although the functional significance of CA in Bowmans glands is obscure, the enzyme may play a role in regulation of pH and ion balance in the mucous layer covering the olfactory epithelium. The presence of CA activity in the ducts suggests that these structures are not simple tubes serving as a conduit for secretory substances but participate in modifying the luminal content by secreting CA.

Histochemical localization of carbonic anhydrase in the trachea of the guinea pig

Tissue specimens from guinea pigs were examined using an enzyme-histochemical reaction to explore the presence of carbonic anhydrase (CA) activity in the trachea. CA activity was detected in a group of morphologically distinct epithelial cells, in goblet cells, and in glands of the tracheal mucosa. The epithelial cells showing CA activity were distributed singly and sparsely throughout the entire trachea. These cells showed a wide morphological variability and were clearly different from those forming the pseudostratified ciliated epithelium. Their number was higher in sections closer to the tracheal bifurcation than in those near the larynx. Although the nature of these cells is unknown, based on their morphological and histochemical characteristics and their distribution, they may represent a specialized chemoreceptor. To our knowledge, this is the first report of CA localized in tracheal epithelial cells.

New Interdisciplinary Approach for Removing Large Clivus and Upper Cervical Spine Tumors

We have developed a new interdisciplinary approach for removing large clivus and upper cervical spine tumors. This approach is a combination of the Le Fort I osteotomy, midfacial degloving, and median labiomandibular glossotomy. Our approach gives an excellent, wide surgical field from the nasopharynx, including the base of the skull, to the base of the tongue and permits sufficiently safe extirpation of clivus and upper cervical spine tumors that may not be removed by transoral or transcervical approaches. Our approach not only incorporates the merits of each approach but also creates a larger surgical field that may be modified or expanded to accommodate the removal of more bulky tumors in this region. This novel approach will facilitate more successful resection of tumors arising between the nasopharynx, including the skull base, and the retropharyngeal area.

The hearing loss in Wegener's granulomatosis: relationship between hearing loss and serum ANCA.

We observed the change of hearing for 1 year in a patient with Wegeners granulomatosis. We also studied anti-human neutrophil cytoplasm antibody (ANCA), which is believed to be specific for this disease and to be associated with disease activity. From these progresses, changes in hearing were considered to be generally related with the general condition and with the ANCA titer level. Because hearing loss in our case was improved by steroids and immunosuppressants, and because ANCA is the autoantibody observed in angiitis syndrome, hearing loss in this case of Wegeners granulomatosis was suspected of being caused by angiitis of the middle and inner ear induced by autoimmune reaction.

Anomalies of the auditory organ in trisomy 18 syndrome: human temporal bone histopathological study

The purpose of this study was to define the histopathological changes in the temporal bone of a fetus with trisomy 18 syndrome, a stillborn due to perosplanchnia. Several anomalies were found including malformation of the auditory ossicles, residual mesenchyme in the middle ear, aberrant tensor tympani muscle, absence of stapedial tendon, aberrant lateral ampullary nerve and wide endolymphatic sinus. The incus body was deformed and separated from the long process by connective tissue and monocrural stapes was noted in the right ear. Three-dimensional reconstruction images provided a clear view of the auditory ossicle malformation. The abnormal findings in our case indicate that ear anomalies in this syndrome might be derived from the component around the first and second branchial arches.

Rhabdomyoma of the base of the tongue

The histopathological and imaging findings of a rhabdomyoma of the base of the tongue were studied. An immunohistochemical examination of the tumour cells showed positive immunostaining for myoglobin, desmin, and striated muscle actin, but negative immunostaining for smooth muscle actin. Electron microscopy showed many glycogen granules and mitochondria in the tumour cells. The T2-weighted and contrast-enhanced magnetic resonance images (MRI) clearly delineated morphological features of this tumour, but T1-weighted MRI and computed tomography (CT) images showed no important features. These findings are typical for an adult extracardiac rhabdomyoma located in the head and neck region, and they will be useful for diagnosis of this tumour.