Okajimas Folia Anat. Jpn., 58(4-6) : 583-594, March 1982 Some Observations on the Fine Structure of the Goblet Cells in the Nasal Respiratory Epithelium of the Rat, with Special Reference to the Well-Developed Agranular Endoplasmic Reticulum By TORAO YAMAMOTO and HARUHIKO MASUDA Department of Anatomy, Faculty of Medicine, Kyushu University, Fukuoka 812, Japan -Received for Publication, November 16, 1981- Key words : Fine structure, Goblet cells, Nasal respiratory epithelium, Agranular endoplasmic reticulum, Rat Summary : The fine structure of the goblet cells in the nasal respiratory epithelium of the rat was examined, in comparison with that of the intestinal goblet cells, by electron microscopy. The nasal goblet cells are characterized by the remarkable development of agranular endoplasmic reticulum in the apical cytoplasm ; on the other hand, the intestinal goblet cells are completely devoid of this type of endoplasmic reticulum. In the nasal goblet cells mucus secretory granules are embedded in the networks of agranular endoplasmic reticulum, when accumulating in the apical cytoplasm. The basal half of the cells is abundantly provided with parallel arrayed cisterns of granular endoplasmic reticulum. The distribution pattern of endoplasmic reticulum as mentioned above is quite similar to that in the non-ciliated cells (Clara cells) of the terminal bronchiole. The study suggests that the nasal goblet cells of the rat are not only involved in the mucus secretion, but also play at least in part the same functional role as do the Clara cells. However, the functional implication of the agranular endoplasmic reticulum in the nasal goblet cells is in reality not known at present. It is known that the goblet cells responsible for mucus secretion occur in the mucosa of the intestinal tracts and the air passages, in which their population is variable in place to place, of mammals. Although the fine structure of the intestinal goblet cells has been well documented so far (Palay, 1958 ; Freeman, 1962, 1966 ; Hollmann, 1963 ; Neutra and Leblond, 1966 ; Kurosumi, Shibuichi and Tosaka, 1981; and others), very little is known about the fine morphology of the goblet cells in the air passages (Matulionis and Parks, 1973 ; Okano and Sugawa, 1965 ; Hansell and Moretti, 1969). The present paper describes some ultrastructural aspects, particularly well-developed agranular endoplasmic reticulum, characteristic of the This study was supported by Grant-in-Aid for scientific research from the Ministry of Education of Japan. 583
584 T. Yamamoto and H. Masuda goblet cells of the nasal respiratory epithelium of the rat, in comparison with those of the intestine. Materials and Methods White rats were anesthetized with nembutal injected intraperitoneally, and per fused with half strength Karnovsky fixative buffered with 0.1 M cacodylate through the left ventricule. Then, a small piece of the nasal mucosa and the jejunum was removed form the animals respectively. Immediately after removal, all these materials were cut into small bits and then transfered into and fixed in the same fixative as above for 2 hr. Subsequently, materials were fixed in an ice cold 2% osmic acid buffered with 0.1 M cacodylate for 2 hr. Following dehydration in graded ethanols, specimens were embedded in epoxy resin. Thin sections were made with a diamond knife on a Porter-Blum ultramicrotome, stained with uranyl acetate and lead tartrate solution and then examined in a Hitachi H-300 electron microscope. Observation The goblet cells in the nasal respiratory epithelium occur relatively abundantly among ciliated cells (Fig. 1). These cells do not show such a typical goblet shape as usually seen in the intestine (Fig. 5), but exhibit a slender, tall columnar profile with more or less undulated outline. When the apical cytoplasm is packed with large amounts of secretory granules, the supranuclear half of the cells extends slightly laterally (Fig. 1). The lumina' surface provided with sparse, short microvilli shows an irregular contour which seems due to the accumulation of secretory granules, and often here cytoplasmic protrusions containing secretory granules can be observed, suggesting the occurrence of apocrine secretion (Fig. 1). The lateral surf aces are attached to those of adjacent cells by a tight junction just beneath the microvilli. Apart from the tight junction, there can be observed sporadic desmosomes along the length of the intercellular clefts. The interdigitation of the lateral membranes between adjoining cells which is consistently encountered in the intestinal goblet cells (Fig. 5) is very scanty or absent in the nasal goblet cells (Fig. 1). Secretory granules which are bounded by a single membrane and more or less uniform in size can be usually observed accumulated in the apical cytoplasm (Figs. 1, 2). Individual granules remain in discrete form, and do not fuse to each other to develop into large droplets as seen in the intestinal goblet cells (Figs. 1, 2, and 5). The agranular endoplasmic reticulum is abundant in the apical cytoplasm and particularly concentrated in the supranuclear region above the Golgi apparatus (Figs. 3, 4). Here agranular endoplasmic reticulum is tubular in profile and constitutes complex networks of tubules (Fig. 4). Even in the apical cytoplasm occupied with secretory granules, the agranular endoplasmic reticulum fills the narrow cytoplasm between secretory granules (Figs. 2, 3). Although the agranular endoplasmic reticulum appears to contain some flocculent materials in its cavity and occasionally a few secretory granules are seen embedded among networks of the reticulum, secretory granules show neither continuity with nor development from the agranular endoplasmic reticulum (Figs. 2, 3 and 4). The Golgi apparatus consisting of stacked lamellae, vesicles and vacuoles is located above the nucleus, but appears to be less developed than that in the intestinal goblet cells (Figs. 1, 3 and 5). A variety of condensing vacuoles are also seen
Fine Structure of the Nasal Goblet Cells 585 situated in the cytoplasm, being embraced by the Golgi lamellae (Fig. 1). The granular endoplasmic reticulum which is arranged parallel in stacks, is well developed mainly in the basal half of the cells and often shows continuities with agranular endoplasmic reticulum (Figs. 1, 4). No dilatations suggesting direct formation of secretory granules can be observed in this reticulum. Elongated mitochondria with relatively numerous cristae are seen distributed throughout the cythplasm. They are moderate in number, but more or less numerous as compared with those of the intestinal goblet cells. Lysosomes are occasionally found in the transition area from agranular to granular endoplasmic reticulum, though very infrequent. The goblet cells of the intestial mucosa show a typical goblet shape, when the apical cytoplasm is filled with mucus secretory granules (Fig. 5). Secretory granules tend to produce varying sizes of granules by fusion (Fig. 5). The cytoplasm between granules is extremely scanty. The nucles with irregular contour is usually located in the basal half of the cells. In maturing goblet cells, the Golgi apparatus is well developed and occupies most of the central column of the supranuclear cytoplasm (Fig. 5). This organelle is closely associated with different condensing vacuoles which contain some flocculent materials similar to those of secretory granules. The remaining cytoplasm is evenly distributed with parallel arrayed granular endoplasmic reticulum and free ribosomes being devoid of agranular endoplasmic reticulum. Discussion The present study has demonstrated the prominent occurrence of agranular endoplasmic reticulum in the apical cytoplasm of the goblet cells in the nasal respiratory epithelium of the rat. This feature is quite different from that of the intestinal goblet cells, in which agranular endoplasmic reticulum is entirely absent in the apical cytoplasm. Similar occurrence of agranular endoplasmic reticulum has been reported in the goblet-type cells of the nasal respiratory epithelium of the mouse (Matulionis and Parks, 1973). In these cells, the agranular endoplasmic reticulum is distributed densely in the apical cyto plasm of the cells containing few or no secretion droplets ; however, it is relatively reduced in amount following droplet accumulation. In the nasal goblet cells of the rat, on the other hand, the agranular endoplasmic reticulum seems to remain unchanged in amount, even after granule accumulation, forming a dense mass of agranular endoplasmic reticulum in the cytoplasm beneath granule accumulation. Thus, there is a minor difference in the distribution pattern of agranular endoplasmic reticulum be tween the nasal goblet cells of the rat and the nasal goblet-type cells of the mouse ; however, these two cells are probably the same in nature. Therefore, the goblet cells with well-developed agranular endoplasmic reticulum mignt be specific for the nasal respiratory mucosa of mammals. Since the development of agranular endoplasmic reticulum and the presence of secretory granules in the apical cytoplasm, and the abundance of granular endoplasmic reticulum in the basal cytoplasm are characteristic of the non-ciliated. cells (Clara cells) in the terminal bronchiole (Cutz and Conen, 1971), it may be reasonable to presume that the nasal goblet cells of the rat are a kind of modified Clara cells which differentiated to possess an additional function of mucus secretion, although the Clara cells have not yet been well established to be mucus secreting cells
586 T. Yamamoto and H. Masuda (Cutz and Conen, 1971 ; Kuhn, Callaway and Askin, 1974 ; Petrik and Collet, 1974). It was reported that in the tracheal mucosa of the mouse, in which goblet cells are actually absent, the non-ciliated cells showed a positive reaction to PAS test (Hansell and Moretti, 1969). Therefore, the explanation is acceptable that the nonciliated cells develop freely into goblet cells in the upper respiratory tract but fail to do so in the lower tract (Matulionis and Parks, 1973). In the goblet-type cells of the nasal respiratory epithelium of the mouse, the possibility was suggested that secretory granules are formed directly in the granular endoplasmic reticulum without passing through the Golgi apparatus (Matulionis and Parks, 1973). However, the present study does not provide any evidences to support such possibility in the nasal goblet cells of the rat. In the Clara cells of the rat, Kuhn et al (1974) described that the agranular endoplasmic reticulum which is well developed in the apical cytoplasm may be a major site of the production of secretory granules. In the present study, however, the secretory granules are not confirmed to develop directly in the agranular endoplasmic reticulum, but appear to mature in the Golgi apparatus in the same fashion as do in the intestinal goblet cells. The present study suggests that the nasal goblet cells of the respiratory mucosa of the rat are primarily similar, if not identical, in function to the Clara cells, but have differentiated, in addition, to produce mucus in adjustment to the environmental condition of the nasal cavity. References Am. J. Path., 62: 127-142, 1971. 2) Freeman, J. A. : Fine structure of the goblet cell mucous secretory process. Anat. Rec., 144: 341-358, 1962. 3) Freeman, J. A. Goblet cell fine structure. Anat. Rec., 154: 121-148, 1966. 4) Hansell, M. M. and R. L. Moretti: Ultrastructure of the mouse tracheal epithelium. J. Morph., 128: 159-170, 1969. 5) Hollman, K.: The fine structure of the goblet cells in the rat intestine. Ann. New York Acad. Sci., 106: 545-554, 1963. 6) Kuhn, C., L. A. Callaway and F. B. Askin : The formation of granules in the bronchiolar Clara cells of the rat. 1. Electron microscopy. J. Ultrastruct. Res., 49: 387-400, 1974. 7) Kurosumi, K., I. Shibuichi and H. Tosaka : Ultrastructural studies on the secretory mechanism of goblet cells in the rat jejunal epithelium. Arch. histol. jap., 44: 263-284, 1981. 8) Matulionis, D. H. and H. F. Parks : Ultrastructural morphology of the normal nasal respiratory epithelium of the mouse. Anat. Rec., 176: 65-84, 1973. 9) Neutra, M. R. and C. P. Leblond : Synthesis of the carbohydrate of mucus in the Golgi complex as shown by electron microscopic radioautography of goblet cells from rats injected with glucose-3h. J. Cell Biol., 30: 119-136, 1966. 10) Okano, M. and Y. Sugawa : Ultrastructure of the respiratory mucous epithelium of the canine nasal cavity. Arch. histol. jap., 26: 1-21, 1965. 11) Palay, S. L.: The morphology of secretion. In : Palay, S. L. ed. "Frontiers in cytology". 305-342, Yale Univ. Press. New Haven, 1958. 12) Petrik, P. and A. J. Collet : Quantitative electron microscopic autoradiography of in vivo incorporation of 3H-choline, 31-1- leucine, 3H-acetate and 3H-galactose in nonciliated bronchiolar (Clara) cells of mice. Am. J. Anat., 139: 519-534, 1974. 1) Cutz, E. and P. E. Conen : Ultrastructure and cytochemistry of Clara cells.
Fine Structure of the Nasal Goblet Cells 587 PLATES
588 T. Yamamoto and H. Masuda Explanation Plate of Figures I Fig. 1. An electron micrograph of the nasal respiratory epithelium of the rat at low magnification. Four goblet cells, in which the apical cytoplasm is packed with secretory granules, are seen together with ciliated cells. The apical cytoplasmic protrusion containing secretory granules is observed in the goblet cell at the right. In these goblet cells, the Golgi apparatus is seen prominently above the nucleus. x 7,000.
Plate 589 I T. Yamamoto and H. Masuda
590 T. Yamamoto and H. Masuda Plate II Fig. 2. The apical portion of the ciliated cell (C) and goblet cells. Secretory granules of the goblet dells are seen in discrete form. The cytoplasm between granules is occupied with agranular endoplasmic reticulum. x 20,000. Fig. 3. The cytoplasm above the Golgi apparatus (G) is provided with numerous agranular endoplasmic reticulum, among which secretory granules are present. x 20,000.
T. Yamamoto and H. Masuda 591 Plate II
592 T. Yamamoto and H. Masuda Plate III Fig. 4. A transition area between agranular and granular endoplasmic reticulum. Here can be observed the continuity between two types of endoplasmic reticulum. x 25,000. Fig. 5. A low magnification of electron micrograph showing a typical goblet cell in the jejunum of the rat. x 6,000.
Plate 593 III T. Yamamoto and H. Masuda