Examination and classification of human olfactory mucosa in patients with clinical olfactory disturbances

Summary To determine objectively the degree of olfactory disturbance, we biopsied the olfactory mucosa from patients who complained of anosmia. The olfactory disturbances in this study were caused by choanal atresia, chronic sinusitis, viral inflammation, and head trauma, as well as by congenital and idiopathic anosmia. The biopsy specimens were examined by light microscopy and the degree of mucosal degeneration present was classified according to five grades. The clinical courses of the patients studied paralleled the changes found in the olfactory mucosa.     

The human olfactory mucosa

Studies of the tissues of the human olfactory mucosa have been performed to investigate olfactory dysfunction and, more recently, olfactory mucosa has attracted a novel interest of investigators because it can be used as an early marker of neurodegenerative conditions of the brain and as a source of multipotent neural stem cells, with applications in regenerative medicine. The olfactory mucosa is readily available to the otolaryngologist, but the harvesting of this tissue must be safe, effective, and reliable, obtaining as little tissue as necessary, while avoiding unnecessary harm to the remaining olfactory tissue and function. The purpose of this review is to summarize the results of the most important studies and knowledge with regard to the human olfactory mucosa and its applications, emphasizing the issue of the distribution of the olfactory mucosa in the nasal cavities.     

Carnosine-like immunoreactivity in human olfactory mucosa

By means of immunohistochemical techniques it is demonstrated for the first time ever that carnosine (beta-alanyl-L-histidine)-like immunoreactivity (LI) is localized in the olfactory receptor neuron in human olfactory mucosa. Carnosine-LI was found in all regions of the receptor cells, i.e. in dendrites, olfactory vesicles, soma and axons. Supporting cells and basal cells were not immunoreactive. The present results suggest that carnosine play a major role in the human olfactory neuron and supports the concept that this peptide is a putative neurotransmitter in vertebrate olfactory systems.     

A re-evaluation of the classification of olfactory epithelia in patients with olfactory disorders

Summary We have previously demonstrated that human olfactory epithelia can be classified into five grades according to the degree of degeneration present in patients with various kinds of olfactory disorders. In practice, however, the occurrence of additional types of cell changes in other kinds of olfactory disorders and findings with immunohistochemical techniques have led us to re-evaluate our previous classification. In the present study, changes in olfactory epithelia from ten patients with various kinds of olfactory disorders are discussed and a revised classification is proposed. Microvillar and differentiating cells were also evaluated in the epithelium studied. Correspondence to: M. Yamagishi     

Abnormalities of axon growth in human olfactory mucosa

OBJECTIVES/HYPOTHESIS: Random biopsies of the human adult olfactory mucosa often demonstrate degenerative changes in the olfactory epithelium (OE) in both dysosmic and normosmic patients and, consequently, have limited diagnostic usefulness. However, detailed analysis of the subepithelial tissue with specific attention to the fascicles of the olfactory nerve and abnormalities of axonal growth may improve the correlation of histopathology with sensory function. STUDY DESIGN: Retrospective review of human OE biopsies. METHODS: Mucosal biopsies from the olfactory area obtained from 27 subjects were examined by light and electron microscopy, with particular attention to the olfactory nerve fascicles; results were correlated with clinical status. Immunohistochemical analysis was used to characterize the extent of axonal depletion, relative maturity of the parent population, and aberrant axonal growth. RESULTS: As expected, there are areas of respiratory metaplasia and neuronal depletion in normosmic as well as dysosmic patients. The degree of axon degeneration within the fascicles correlates better with individual olfactory status. Immature neurons predominate, and re-entrant neuromas develop in patients with olfactory loss caused by disconnection from the olfactory bulb. Individuals with olfactory loss caused by epithelial damage as with chronic rhinosinusitis display evidence of nerve fascicle degeneration and intraepithelial neuromas. CONCLUSION: The status of olfactory axons provides useful information on the overall condition of the olfactory periphery and improves the diagnostic usefulness of mucosal biopsies. In addition to an assessment of the epithelium per se, the fascicles of the olfactory nerve need to be characterized for a complete analysis of the olfactory mucosa.     

Immunohistopathology of variations of human olfactory mucosa

Summary The characteristics of the human olfactory mucosa were studied immunohistologically. Regular, tonal distribution of the supporting cells, multilayered olfactory receptor cells and basal cells was commonly found in the olfactory mucosa of the human fetus. In contrast, most of the olfactory mucosa in the adult varied to some extent. In the relatively thick, slightly degenerated olfactory mucosa, olfactory marker protein positive receptor cells were arranged irregularly. The most common evidence for variation was the decrease or disappearance of the olfactory receptor cells. Serous-type lactorferrin-containing glandular acini were characteristically found beneath degenerated epithelium. Islands of respiratory epithelium were also seen. The ductules of the Bowman's glands were distended and the openings of these ductules were wide. There was invagination or epithelial cell processes into the glandular lumina. These findings suggest that the epithelial cells of Bowman's glands play an important role in the regeneration of the human olfactory mucosa. Offprint requests to: T. Nakashima     

放疗后嗅粘膜超微结构观察

嗅区粘膜放疗后可引起迟发性嗅觉障碍，为了解其机制，对放疗后嗅粘膜用透射电镜观察其超微结构改变。发现其上皮层变性嗅神经原增多；固有层内纤维母细胞增多，胞浆内含有大量粗面内质网和线粒体，在纤维母细胞周围有大量整齐排列的胶原纤维。放疗后嗅粘膜的纤维化是嗅觉障碍的原因之一。     

Odor stick identification test for Japanese patients with olfactory disturbances

In recent years, a new stick-type odor identification test, the odor-stick identification test for Japanese (OSIT-J) has been developed in Japan. Thirteen odors familiar to Japanese people are used in this test. The OSIT-J is an olfactory discrimination test and is significantly correlated with the average recognition threshold of T & T olfactometry, which is the standard olfactory acuity test used in Japan. In this study, we evaluated the accuracy of the OSIT-J in patients with olfactory disturbances. We compared the OSIT-J and T & T olfactometry results and examined the sensitivity and specificity of the OSIT-J. Using the OSIT-J, olfactory disturbances were diagnosed in more than 70% based on the average recognition threshold determined by T & T olfactometry. OSIT-J is a simple test and is recommended for use in clinical practice for evaluating olfactory disturbances.     

Main causes and diagnostic evaluation in patients with primary complaint of olfactory disturbances

IntroductionEstablishing a diagnosis in patients with olfactory disturbances has always been challenging for physicians.One reason for this is the rarity of some of the diseases that affect this sense, such as Kallmann's syndrome and post-viral olfactory loss.ObjectiveTo identify the major causes of olfactory disturbances and to describe the diagnostic evaluation in outpatients attended to at an ambulatory clinic specialized in olfaction disorders.MethodsA retrospective analysis was performed in outpatients with primary olfactory complaint attended to between June 1, 2011 and September 30, 2013 in a center specialized in olfactory disorders. Patient history, nasofibroscopy, and the University of Pennsylvania Smell Identification Test (UPSIT) comprised the examination.ResultsSixty-two patients were evaluated. The major causes were chronic rhinosinusitis (31%); rhinitis, primarily the allergic type (19%); post-viral olfactory loss (13%); and post-traumatic loss (8%). UPSIT scores were statistically different among different etiologies (p = 0.01).ConclusionsThe major diagnoses that should be part of the physician assessment when a patient complains of olfactory disturbance are chronic rhinosinusitis with and without polyps, allergic rhinitis, post-viral olfactory loss, and post-traumatic loss.     

变应性鼻炎伴嗅觉障碍小鼠嗅黏膜的观察

目的观察变应性鼻炎伴嗅觉障碍小鼠嗅黏膜形态结构的改变及相关免疫组织化学变化,探讨变应性鼻炎对嗅觉功能的影响。方法将40只BALB/C小鼠随机分为实验组(30只)和对照组(10只)。实验组应用卵清蛋白致敏后,利用埋藏食物小球实验(buried food pellet test,BFT)评估小鼠嗅觉功能,ELISA法测定并比较血清中IgE水平。建立变应性鼻炎伴嗅觉障碍小鼠动物模型。取小鼠鼻腔嗅黏膜,观察形态结构的改变,检测各组嗅黏膜中嗅觉标记蛋白(olfactory marker protein,OMP)和神经元特异性烯醇化酶(neuron- specific enolase,NSE)的表达。结果变应性鼻炎伴嗅觉障碍组嗅黏膜OMP阳性的成熟嗅感觉神经元(olfctory receptor neurons,ORNs)减少,与变应性鼻炎不伴嗅觉障碍组及对照组相比差异存在统计学意义(P=0.046<0.05,P=0.039<0.05)。变应性鼻炎伴嗅觉障碍组嗅黏膜NSE阳性的成熟ORNs减少,与变应性鼻炎不伴嗅觉障碍组及对照组相比差异存在统计学意义(P=0.041<0.05,P=0.033<0.05)。结论通过卵清蛋白致敏激发可以成功建立变应性鼻炎伴嗅觉障碍小鼠的动物模型。变应性鼻炎的炎症反应导致嗅黏膜的改变,可能是变应性鼻炎嗅觉障碍的发病机制之一。     

Expression of neuron-specific enolase and olfactory marker protein in the developing olfactory mucosa of human fetuses]

OBJECTIVE: To study the expression of neuron-specific enolase (NSE) and olfactory marker protein (OMP) in the developing olfactory mucosa of human fetuses. METHOD: The expression of NSE and OMP in the olfactory mucosa of 6 human fetuses (12, 16, 20, 24, 28 and 34 weeks) was studied using the technique of immunohistochemistry. RESULTS: NSE immunological positive reactions were seen in all 6 fetal mucosa from gestational 12 (G12) to G34, with plenty of positive-stained dual-pole neuron cells. At G12, the positive cells aligned tightly, the cell bodies were localized in the lower portion of olfactory epithelium and the positive-stained area occupied upper 2/3 of fetal nasal mucosa. With the development, the positive cells gradually became multilayer, but the density and the relative area of positive-cells reduced. At G34, the positive cells were located only in upper 1/3 of nasal mucosa. OMP-positive reactions were localized in a few dual-pole neurons at G12, the number was much less than NSE-positive cells in the same fetus. With the development, the OMP-positive cells gradually increased with most of the cell bodies located in the upper portion of epithelium, but number still relatively less than the NSE-positive cells at the same age. CONCLUSION: At G12, there were lots of olfactory neuron in the olfactory mucosa and only a few olfactory neurons had became mature. With the development, the olfactory epithelial area reduced but the number of mature olfactory neurons increased. At the last trimester, fetal olfactory sensor was almost matured.     

人嗅黏膜细胞色素P4502A表达的免疫印迹和免疫组化分析

目的　研究人类细胞色素P4 5 0 (cytochromeP 4 5 0 ,CYP 4 5 0 ) 2A(CYP2A)在鼻黏膜中的表达特点和意义。方法　应用免疫印迹方法分别检测 36例鼻腔鼻窦疾病患者和孕 96、119、16 0、182、185d的 5例胎儿不同部位鼻组织中CYP2A的表达水平 ,并用免疫组化方法检测CYP2A在嗅区黏膜中分布特点。结果　成人标本中 10块嗅区黏膜标本中 8例免疫印迹阳性 ,而 37块呼吸区黏膜标本均为阴性。胎儿嗅区黏膜标本免疫印迹均为阳性 ,其表达水平随胎龄有增加的趋势 ,仅 1例呼吸区黏膜检测到CYP2A。免疫印迹定量分析显示 ,成人嗅区鼻黏膜微粒体CYP2A蛋白质水平在0 1～ 1 1pmol/mg之间 ,胎儿的鼻微粒体蛋白质含量在 0 8～ 5 3pmol/mg之间 ,胎儿一般高于成人。免疫组化结果显示 ,CYP2A免疫活性存在于嗅黏膜的支持细胞和Bowman腺。结论　CYP2A存在于嗅黏膜提示 :嗅黏膜是吸入性外源性物质生物转化的靶位 ,CYP2A还与嗅觉感受有关 ,胎儿期CYP2A嗅区鼻黏膜的表达也可能使母体衍生毒素对胎儿成长发育有潜在威胁     

Immunoreactive somatostatin in the peripheral olfactory system and nasal mucosa of the human

Immunohistochemically, the cyclic retradecapeptide somatostatin was demonstrated in periglomerular cells and fibres of the glomerular layer (Fig. 3) of the human olfactory bulb as well as in the superficial granule layer and fibres (Fig. 2) of the olfactory tract. Additionally, somatostatin was found in endothelial cells of subepithelial arterioles and capillaries of the regio olfactoria (Fig. 4a, b) and superior turbinate. We assume that somatostatin has a paracrine transmitter function in the peripheral olfactory system and a regulative function on the blood flow of the superior nasal mucosa.     

胚胎型神经细胞黏附分子在人嗅黏膜内的发育表达

目的：探讨胚胎发育期人嗅黏膜内胚胎型神经细胞黏附分子(E-NCAM)的表达变化规律及生理功能。方法：使用免疫组织化学方法对人胚胎发育不同时期嗅黏膜内E-NCAM的表达情况进行检测。结果：E-NCAM的表达开始于胚胎发育的第14周，阳性反应位于嗅神经元细胞胞体、树突、轴突及固有层内的嗅神经纤维束；随胚胎发育阳性细胞数目逐渐增多，嗅神经纤维束染色逐渐增强；第28周达峰值后逐渐下降。在发育不同时期，E-NCAM阳性细胞均位于黏膜层的中下部分，黏膜顶层的支持细胞和嗅神经元胞核不表达E-NCAM。结论：E-NCAM的表达同相应的神经元可塑性变化有一致的时序，在胚胎发育前期E-NCAM的高表达，有助于正在分化的神经元生长；发育后期E-NCAM表达下调，有助于成熟神经元细胞的形态结构的稳定性。     

A clinical study of olfactory dysfunction in patients with Mikulicz's disease

Objective

Mikulicz's disease (MD) is differentiated from Sjögren's syndrome as an immunoglobulin G4 (IgG4) systemic disease. MD patients often report olfactory dysfunction (OD). To analyze cases of OD associated with MD, we studied clinicopathological and serological findings of MD patients.

Methods

A total of 44 MD patients (17 males and 27 females) were examined for OD. We evaluated clinicopathological and serological findings of these patients by dividing them into OD (+) and OD (−) groups.

Results

The mean IgG4 concentration (SD) in such cases was 950.5 (797.5) mg/dl. Of the 44 patients, 20 (45%) had OD even though no abnormalities, such as obstructive and inflammatory disease, were detected in their nasal cavities and sinuses. The two groups did not show significant differences in background characteristics, such as age, sex, IgG4 concentration, presence or absence of allergic rhinitis, and presence or absence of extrasalivary gland lesions. We found abundant IgG4-positive plasmacytes in the nasal mucosa specimens of the OD (+) group but not in that of the OD (−) group.

Conclusions

Nasal mucosa in the MD patients with OD was infiltrated with IgG4-positive plasmacytes. We concluded that OD may be associated with infiltration by IgG4-positive cells.     

人嗅黏膜细胞色素P4502A表达的免疫印迹和免疫组化分析

目的 研究人类细胞色素P450(cytochrome P-450，CYP-450)2A(CYP2A)在鼻黏膜中的表达特点和意义。方法 应用免疫印迹方法分别检测36例鼻腔鼻窦疾病患者和孕96、119、160、182、185d的5例胎儿不同部位鼻组织中CYP2A的表达水平，并用免疫组化方法检测CYP2A在嗅区黏膜中分布特点。结果 成人标本中10块嗅区黏膜标本中8例免疫印迹阳性，而37块呼吸区黏膜标本均为阴性。胎儿嗅区黏膜标本免疫印迹均为阳性，其表达水平随胎龄有增加的趋势，仅1例呼吸区黏膜检测到CYP2A。免疫印迹定量分析显示，成人嗅区鼻黏膜微粒体CYP2A蛋白质水平在0．1—1．1pmol／mg之间，胎儿的鼻微粒体蛋白质含量在0．8—5．3pmol／mg之间，胎儿一般高于成人。免疫组化结果显示，CYP2A免疫活性存在于嗅黏膜的支持细胞和Bowman腺。结论 CYP2A存在于嗅黏膜提示：嗅黏膜是吸入性外源性物质生物转化的靶位，CYP2A还与嗅觉感受有关，胎儿期CYP2A嗅区鼻黏膜的表达也可能使母体衍生毒素对胎儿成长发育有潜在威胁。     

A freeze-fracture study of receptor axons and Schwann cells in the human olfactory mucosa

Electron micrographs of freeze-fracture replicas from the human olfactory mucosa were analysed regarding the structure of the axons of the olfactory receptor cells. In the lamina propria, numerous axons were generally invested with one Schwann cell. The ensheathed axons were often found in close contact with one another. Membrane specializations were not found at these sites, nor were tightening membrane junctions observed in the mesaxons. The Schwann cell plasmalemma exhibited caveolae, whose neck was surrounded by uniformly sized intramembranous particles evenly distributed over the axolemmal fracture planes. There was a marked difference in particle density between the P face (about 850/micron 2) and the E face (about 180/micron 2).     

Immunopathology of olfactory mucosa following injury to the olfactory bulb

Removal of the olfactory bulb was performed on rats in an attempt to elucidate the processes of olfactory dysfunction following head injury. Degeneration and regeneration of the olfactory mucosa were examined, histopathologically and immunohistochemically. We used antisera to olfactory marker protein (OMP) and neuron specific enolase (NSE) as a marker of the mature olfactory receptor neurons. Following rapid degeneration after bulbectomy, the olfactory receptor neurons regenerated. OMP and NSE containing cells re-appeared 49 days later. However, the cell population of the neuroepithelium did not revert to the numbers observed in the non-operated neuroepithelium, even three months later. The lack of a connection between regenerated axons and the olfactory bulb may result in immature neuronal replacement and reduce the number of olfactory receptor neurons.