Pediatric Pulmonology 8:40-44 (1990)
Brush Cells in the Human Fetal Trachea Mary F. DiMaio, MD,' Meyer Kattan, MD,' Doina Ciurea, DVM, PhD,2 Joan Gil, MD,* and Renate Dische, M D ~ Summary. We performed a morphologic examination of human fetal lung tissue, using scanning and transmission electron microscopy, in order to establish the presence of brush cells in extrapulmonary and intrapulmonary airways, and developing acinar epithelium. Brush cells, characterized by a border of regular straight microvilli containing a filamentous core, were observed within the tracheal epithelium of a 19-20 week gestational age fetus. These cells constituted 0.5% of the total epithelial cell population. Brush cells were not seen within the bronchial, bronchiolar or developing acinar epithelium. Our study shows that brush cells occur infrequently but normally in the developing tracheal epithelium of the second trimester fetus. Pediatr Pulmonol 1990; 8:40-44. Key words: Scanning and transmission electron microscopy; fetus, 19-20 weeks GA; 0.5% cell frequency.
INTRODUCTION
existence and distribution of this cell in the human fetal airway.
Respiratory brush cells have been described in the trachea and bronchi of man and other mammals. '-11 These cells are distinguished by a border of regular, straight MATERIALS AND METHODS microvilli that protrude into the respiratory lumen and Lung Preparation contain a filamentous core. 1237 Similar cells have been Our material consisted of surgically obtained fetal described in the alveoli of adult rats but are not normally fragments from 10 electively terminated normal pregnanfound in the human alveolus. l2-I6 The initial time of cies ranging in gestational age from 14 to 26 weeks. appearance of the brush cell in mammals in unknown. Gestational age was determined by fetal footlength acLeeson' noted these cells in the trachea of fetal rabbits, cording to published standards. l 9 All tissue was procured and Jeffery and Reid' demonstrated possible precursors for study within 10 minutes of fetal demise. Sections of of this cell in the rat trachea at 21 days gestation. trachea and mainstem bronchus were taken from 3 feAlthough it is assumed that brush cells appear postna- tuses ranging in gestation from 14 to 22 weeks. A rantally in man, there are no studies citing their presence or dom thin section of lung tissue was taken from 9 fetuses absence in the human fetus or infant.8 We recently re- ranging in gestation from 19 to 26 weeks; identification ported the presence of alveolar brush cells in a 4 month as to its precise anatomic origin was not possible. This old infant with desquamative interstitial pneumonitis and lung parenchyma provided developing acinar epithelium suggested that their presence was a regenerative response for study. In 5 fetuses, ranging in gestation from 22 to 24 to injury.17 However, we could not preclude that they weeks, the parenchyma provided intrapulmonary bronwere congenital in origin. The purpose of this study was to examine the fetal airway epithelium for the presence of brush cells in fetuses ranging in gestational age from From the Jack and Lucy Clark Department of Pediatrics' and Lillian 14 to 26 weeks. Using scanning and transmission elec- and Henry M. Stratton-Hans Popper Department of Pathology', tron microscopy we studied 1) the extrapulmonary air- Mount Sinai School of Medicine, New York, New York. ways (trachea and mainstream bronchus); 2) the intraReceived July 11, 1989; (revision) accepted for publication September pulmonary airways (bronchus and bronchioles); and 3 ) 19, 1989. developing acinar epithelium for the presence of brush cells. This work was supported in part by the MCH Training Grant The identification of brush cells in the human lung has MCJ001069, by the U.S. Department of Health and Human Services, led to some controversy because of morphologic differ- and by National Heart, Lung, and Blood Institute grant HL34196. ences between the human and other s p e ~ i e s . ~ ~ ~ ,De~ , ' , ' ' Address correspondence and reprint requests to Dr. M. DiMaio, Mt. spite the controversy, our findings of this unusual cell in Sinai Medical Center, Division of Pediatric Pulmonology, Box 1202, humans""' led us to perform a study to determine the One Gustave L. Levy Place, New York, NY 10029. 0 1990 Wiley-Liss, Inc.
Brush Cells in the Human Fetal Trachea
41
Fig. 1. a: Electron micrograph taken from the trachea of a 19 week gestation fetus showing a columnar brush cell adjacent to a ciliated cell. Numerous blunt microvilli protrude into the tracheal lumen. b: Electron micrographtaken from the same specimen showing a globular-shaped brush cell adjacent to a cell
bearing cilia and microvilli. Compared to the ciliated cell, the brush cell contributes a small surface area to the epithelial airway. Uranyl acetate and lead citrate. Original magnification x 6,600, a and b.
chial or bronchiolar epithelium for examination. The tissues were immediately fixed for transmission electron microscopy by immersion in 1.5% glutaraldehyde in 0.1 M cacodylate buffer (pH = 7.3) for 24 hours. The tissue was then diced into numerous blocks of 1 mm length, placed in 1.5% osmium tetroxide with S-collidine buffer (pH = 7.4) for 1 hour, and post-fixed in uranyl acetate in maleate buffer (pH = 6) for 45 minutes. The osmolarity of these solutions had been adjusted to 330 mOsm. After dehydration in graded ethanol, the tissue samples were embedded in Epon 812. For scanning electron microscopy, one thin slice of lung parenchyma was taken from 4 fetuses, ranging in gestation from 21 to 24 weeks, in order to study bronchiolar and developing acinar epithelium. The tissue was fixed in 1.5% glutaraldehyde for 24 hours, diced into small blocks, and placed in 1.5% osmium tetroxide with S-collidine buffer for 1 hour. After dehydration in graded ethanol, critical point drying was accomplished with the use of a Bomar instrument.
Sampling Procedure
For transmission electron microscopical examination four blocks of lung parenchyma were selected randomly. Two blocks from trachea and one block from bronchus and intrapulmonary bronchus were also randomly chosen. Ultrathin section (60-90 mm) were cut with a diamond knife on an LKB ultramicrotome. The sections were mounted on copper grids and double stained with uranyl acetate and lead Using a JEM CXlOO transmission electron microscope a total of 4 2.5 2 grids (mean t SE) from trachea, mainstem bronchus, and intrapulmonary bronchus were examined for brush cells. Five grids from each block of lung parenchyma were also examined for a total of 180 grids. Brush cells, when identified, were photographed. A nuclear cell count was performed by counting the nuclei of all the respiratory epithelial cells present in the specimen in which brush cells were seen. A total of 385 cells were counted. For scanning electron microscopy, one randomly se-
42
DiMaio et al.
Fig. 2. a: Higher magnification of Figure l a showing fine filaments within the microvilli. The filaments extend into the apical cytoplasm (arrow). A desmosome is present (arrowhead). b: Higher magnificationof Figure 1b showing the microvilli in lon-
gitudinal and cross section. Filaments within the microvilli end in a terminal web (arrowheads). Uranyl acetate and lead citrate. Original magnification ~ 2 0 , 0 0 0a, and b.
lected block of tissue was attached to a specimen stub and coated with gold-palladium by the use of a Hummcr sputtering device. The specimens were viewed in their entirety under a HlTACHl SEM 5300-KEVEX 2000 scanning microscope for the presence of brush cclls.
within the microvilli. The filaments extended into the apical cytoplasm and in one cell appeared to end in a terminal web (Fig. 2a,b). The mitochondria of the brush cells were small and less plentiful in number when compared to the ciliated cells. The Golgi complexes were well developed, and several lysosomes were seen in one cell. Rough endoplasmic reticulum was sparse, but numerous free ribosomes were scattered throughout the cytoplasm. Smooth endoplasmic reticulum and pinocytotic vesicles were found in the apical cytoplasm of the cell. Tight junctions and desmosomes joined the brush cells to the adjacent ciliated cells. Brush cells were not found in sections of mainstem bronchus, intrapulmonary bronchus, bronchioles, or developing acinar epithelium by either scanning or transmission electron microscopy. We observed other cells, resembling brush cells, within the tracheal, bronchial, and bronchiolar epithelium. These cells were rare and were covered by numerous microvilli measuring 0.25-0.3 pm in thickness (Fig. 3 ) . The microvilli frequently branched and did not form a very regular border. The cells contained apical
RESULTS
We examined the tracheal epithelium of two fetuses of 14 and 19-20 weeks gestational age. Transmission electron microscopy revealed the presence of brush cells within the epithelium of the 19-20 week old fetus. These cells occurred with a frequency of 0.5% and were always found between ciliated cells. The brush cells varied in shape from columnar to globular. Compared to the neighboring ciliated cells, they presented a small surface area to the luminal surface and were wider at the base than at the apex (Fig. la,b). The brush cells were devoid of cilia and basal bodies and exhibited numerous, blunt, straight microvilli of regular length protruding into the tracheal lumen. The microvilli measured 0.25-0.3 pm in thickness. Bundles of fine filaments were present
Brush Cells in the Human Fetal Trachea
43
Fig. 3. Electron micrograph taken from a 20 week gestation fetus showing a bronchiolarcell covered with microvilli some of which branch (arrowheads). The microvilli appear to have a
fuzzy coat consistent with glycocalyx. Two basal bodies (arrows) are present within the apical cytoplasm. Uranyl acetate and lead citrate. Original magnification x 16,000.
smooth endoplasmic reticulum, pinocytotic vesicles, and a few basal bodies.
appear as stubby. The authors state that the rat brush cell does not have an “adult” appearance until 19 days postnatally.8 In light of our findings and those of previous investigators, it is probable that interspecies variation and developmental stage account for differences in the morphological appearance of brush cells, Based on our ultrastructural observation of cells bearing a regular microvillus border, we conclude that brush cells are present in the trachea of human second trimester fetuses. This ultrastructural criterion is used for all rnamalian species in order to classify a cell as a brush cell. 1.6,7,18 Cells meeting this criterion have been described in the trachea, bronchi, and bronchioles of children and a d ~ l t s . ~ - ~ However, ,~,I’ other investigators refute the existence of brush cells in man and believe that some workers are merely describing precursors of the ciliated cell.” The brush cells that we describe do not demonstrate basal bodies or cilia. We did observe cells within the the fetal airway bearing numerous microvilli and containing basal bodies. These cells resemble indifferent cells undergoing differentiation to ciliated cells, as described by McDowell et a1.,I8 and are probably not brush cells. The observation of a cell bearing microvilli and containing basal bodies indicated to several investigators the role of the “brush” cell in c i l i ~ g e n e s i s .Other ~ ~ ~ , pos~ sible functions of the brush cell have been explored. The presence of a microvillous border and apical vesicles are features of cells with an absorptive function. I,‘,’ Since
DISCUSSION
In this study we performed a rnorphologic examination of human fetal lung tissue in order to determine if brush cells are present prenatally in the extrapulmonary and intrapulmonary epithelium and developing acinar epithelium. We found brush cells in the developing tracheal epithelium of one human second trimester fetus. In mammals, brush cells are distinguished by a border of regular, straight microvilli protruding into the respiratory lumen and containing a filamentous core. 1 3 6 , 7 3 1 9 The human fetal tracheal brush cells that we describe are morphologically similar to but not identical with the brush cells of the adult rat trachea and calf bronchus.1363Z1 In the adult rat and calf the microvilli appear more stubby when compared to the human fetus. Whereas the rat microvilli measure 1.5 to 1.8 p m in width, the calf microvilli measure 0.25 p m in width. Thus, the human fetal microvilli have a width identical with that of the calf. Although the microfilaments contained within the human fetal microvilli occasionally end in a terminal web, this has not been observed in other mammals. A probable precursor of the brush cell has been identified in fetal rats.8 Based on the published electron micrograph, the microvillus border is less orderly than that of the adult rat, and the microvilli do not
44
DiMaio et al.
the Golgi apparatus appears prominent, we suggest that these cells might be involved in a secretory function rather than an absorptive process. Luciano et al. observed associations between rat tracheal brush cells and nerve fibers and speculated that brush cells may be chemoreceptors.2’ We did not observe nerve fibers adjacent to fetal brush cells. Hijiya showed that bleomycin-induced injury in rat lung causes the appearance of increased numbers of alveolar brush cells. 12,13 He suggests that the prevalence of brush cells may be the result of injury and hypoxia. We recently reported the presence of alveolar brush cells in a 4 month old infant with desquamative interstitial pneumonitis and suggested that their presence may represent a regenerative response to injury. l 7 However, we could not preclude that cells at that location were congenital in origin. Our present study has shown that brush cells do not occur normally in the developing acinar epithelium. Whether in this infant they were a response to injury or an abnormality in cellular development in unknown. In conc1usion, we have demonstrated the prenatal existence of brush cells in the human trachea. Further work is warranted in order to elucidate their function.
REFERENCES 1. Jeffery PK, Reid L. New observations of rat airway epithelium: A
2.
3. 4.
5.
quantitative and electron microscopic study. J Anat. 1975; 120: 2% -320. Reid L, Jones R. Bronchial mucosal cells. Fed Proc. 1979; 38: 191-196. Rhodin JAG. Ultrastructure and function of the human tracheal mucosa. The ciliated cell. Am Rev Respir Dis. 1966; 93:l-15. Basset F, Poirier J, LeCrom M, Turiaf J. Etude ultrastructure de l’epithelium bronchiolaire humain. Z Zellforsch Microsk Anat. 1971; 116:425-442. Watson JHL, Bnnkman GL. Electron microscopy of the epithelial
cells of normal and bronchitic human bronchus. Am Rev Respir Dis. 1964; 90:851-866. 6. Allan EM. The ultrastructure of the brush cell in bovine lung. Res Vet Sci. 1978; 25:314-317. 7. Rhodin J. Ultrastructure of the tracheal ciliated mucosa in rat and man. Ann Otol Rhino1 Laryngol. 1959; 68964-974. 8. Jeffery PK, Reid L. Ultrastructure of airway epithelium and submucosal gland during development. In: Hodson WA, ed. Development of the Lung. New York: Marcel Dekker Inc., 1977; 87134. 9. Leeson TS. The development of the trachea in the rabbit with particular reference to its fine structure. Anat Anz. 1961; 110: 214-223. 10. Christensen TG, Breuer R, Hornstra LJ, Lucey EC, Snider GL. The ultrastructure of hamster bronchial epithelium. Exp Lung Res. 1987; 13:253-277. I I . Gordon RE, Kattan M. Absence of cilia and basal bodies with a predominance of brush cells in the respiratory mucosa from a patient with immotile cilia syndrome. Ultrastruct Pathol. 1984; 6:4S-49. 12. Hijiya K, Okada Y, Tankdwa H. Ultrastructural study of the alveolar brush ccll. J Electron Microsc (Tokyo). 1977; 26321329. 13. Hijiya K. Electron microscope study of the alveolar brush cell. J Electron Microsc (Tokyo). 1978; 27:223-227. 14. Filippenke LN. Light and electron microscopic study of the alveolar brush cells of the rat lung. Biull Eksp Biol Med. 1978; 86592-596. 15. Meyrick B, Reid L. The alveolar brush cell in rat lung-a third pneumonocyte. J Ultrastruct Res. 1968; 23:7 1-80. 16. Foliguet B, Grignon G. Le pneumocyte de type 111. La cellule alveolaire en brosse du poumon du rat. Etude en microscopie electronique transmission. Poumon-Coeur. 1980; 36: 149-1 53. 17. DiMaio MF, Dische R, Gordon RE, Kattan M. Alveolar brush cells in an infant with desquamative interstitial pneumonitis. Pediatr Pulmonol. 1988; 4:185-191. 18. McDowell EM, Barett LA, Glavin F, Harris CC, Trump BF. The respiratory epithelium, I . Human bronchus. JNC. 61539-545, 1978. 19. Scammon RR. Two simple nomographs for estimating age and some of the major external dimensions of the human fetus. Anat Rec. h8:221-225, 1937. 20. Sato T. A modified method for lead staining of thin sections. J Electron Microsc (Tokyo). 16: 133, 1967. 21. Luciano L, Reale E, Ruska H. Uber eine chemorezeptive sinneszelle in der trachea der ratte. Z Zellforsch. 85:350-375, 1968.
Brush Cells in the Human Fetal Trachea Mary F. DiMaio, MD,' Meyer Kattan, MD,' Doina Ciurea, DVM, PhD,2 Joan Gil, MD,* and Renate Dische, M D ~ Summary. We performed a morphologic examination of human fetal lung tissue, using scanning and transmission electron microscopy, in order to establish the presence of brush cells in extrapulmonary and intrapulmonary airways, and developing acinar epithelium. Brush cells, characterized by a border of regular straight microvilli containing a filamentous core, were observed within the tracheal epithelium of a 19-20 week gestational age fetus. These cells constituted 0.5% of the total epithelial cell population. Brush cells were not seen within the bronchial, bronchiolar or developing acinar epithelium. Our study shows that brush cells occur infrequently but normally in the developing tracheal epithelium of the second trimester fetus. Pediatr Pulmonol 1990; 8:40-44. Key words: Scanning and transmission electron microscopy; fetus, 19-20 weeks GA; 0.5% cell frequency.
INTRODUCTION
existence and distribution of this cell in the human fetal airway.
Respiratory brush cells have been described in the trachea and bronchi of man and other mammals. '-11 These cells are distinguished by a border of regular, straight MATERIALS AND METHODS microvilli that protrude into the respiratory lumen and Lung Preparation contain a filamentous core. 1237 Similar cells have been Our material consisted of surgically obtained fetal described in the alveoli of adult rats but are not normally fragments from 10 electively terminated normal pregnanfound in the human alveolus. l2-I6 The initial time of cies ranging in gestational age from 14 to 26 weeks. appearance of the brush cell in mammals in unknown. Gestational age was determined by fetal footlength acLeeson' noted these cells in the trachea of fetal rabbits, cording to published standards. l 9 All tissue was procured and Jeffery and Reid' demonstrated possible precursors for study within 10 minutes of fetal demise. Sections of of this cell in the rat trachea at 21 days gestation. trachea and mainstem bronchus were taken from 3 feAlthough it is assumed that brush cells appear postna- tuses ranging in gestation from 14 to 22 weeks. A rantally in man, there are no studies citing their presence or dom thin section of lung tissue was taken from 9 fetuses absence in the human fetus or infant.8 We recently re- ranging in gestation from 19 to 26 weeks; identification ported the presence of alveolar brush cells in a 4 month as to its precise anatomic origin was not possible. This old infant with desquamative interstitial pneumonitis and lung parenchyma provided developing acinar epithelium suggested that their presence was a regenerative response for study. In 5 fetuses, ranging in gestation from 22 to 24 to injury.17 However, we could not preclude that they weeks, the parenchyma provided intrapulmonary bronwere congenital in origin. The purpose of this study was to examine the fetal airway epithelium for the presence of brush cells in fetuses ranging in gestational age from From the Jack and Lucy Clark Department of Pediatrics' and Lillian 14 to 26 weeks. Using scanning and transmission elec- and Henry M. Stratton-Hans Popper Department of Pathology', tron microscopy we studied 1) the extrapulmonary air- Mount Sinai School of Medicine, New York, New York. ways (trachea and mainstream bronchus); 2) the intraReceived July 11, 1989; (revision) accepted for publication September pulmonary airways (bronchus and bronchioles); and 3 ) 19, 1989. developing acinar epithelium for the presence of brush cells. This work was supported in part by the MCH Training Grant The identification of brush cells in the human lung has MCJ001069, by the U.S. Department of Health and Human Services, led to some controversy because of morphologic differ- and by National Heart, Lung, and Blood Institute grant HL34196. ences between the human and other s p e ~ i e s . ~ ~ ~ ,De~ , ' , ' ' Address correspondence and reprint requests to Dr. M. DiMaio, Mt. spite the controversy, our findings of this unusual cell in Sinai Medical Center, Division of Pediatric Pulmonology, Box 1202, humans""' led us to perform a study to determine the One Gustave L. Levy Place, New York, NY 10029. 0 1990 Wiley-Liss, Inc.
Brush Cells in the Human Fetal Trachea
41
Fig. 1. a: Electron micrograph taken from the trachea of a 19 week gestation fetus showing a columnar brush cell adjacent to a ciliated cell. Numerous blunt microvilli protrude into the tracheal lumen. b: Electron micrographtaken from the same specimen showing a globular-shaped brush cell adjacent to a cell
bearing cilia and microvilli. Compared to the ciliated cell, the brush cell contributes a small surface area to the epithelial airway. Uranyl acetate and lead citrate. Original magnification x 6,600, a and b.
chial or bronchiolar epithelium for examination. The tissues were immediately fixed for transmission electron microscopy by immersion in 1.5% glutaraldehyde in 0.1 M cacodylate buffer (pH = 7.3) for 24 hours. The tissue was then diced into numerous blocks of 1 mm length, placed in 1.5% osmium tetroxide with S-collidine buffer (pH = 7.4) for 1 hour, and post-fixed in uranyl acetate in maleate buffer (pH = 6) for 45 minutes. The osmolarity of these solutions had been adjusted to 330 mOsm. After dehydration in graded ethanol, the tissue samples were embedded in Epon 812. For scanning electron microscopy, one thin slice of lung parenchyma was taken from 4 fetuses, ranging in gestation from 21 to 24 weeks, in order to study bronchiolar and developing acinar epithelium. The tissue was fixed in 1.5% glutaraldehyde for 24 hours, diced into small blocks, and placed in 1.5% osmium tetroxide with S-collidine buffer for 1 hour. After dehydration in graded ethanol, critical point drying was accomplished with the use of a Bomar instrument.
Sampling Procedure
For transmission electron microscopical examination four blocks of lung parenchyma were selected randomly. Two blocks from trachea and one block from bronchus and intrapulmonary bronchus were also randomly chosen. Ultrathin section (60-90 mm) were cut with a diamond knife on an LKB ultramicrotome. The sections were mounted on copper grids and double stained with uranyl acetate and lead Using a JEM CXlOO transmission electron microscope a total of 4 2.5 2 grids (mean t SE) from trachea, mainstem bronchus, and intrapulmonary bronchus were examined for brush cells. Five grids from each block of lung parenchyma were also examined for a total of 180 grids. Brush cells, when identified, were photographed. A nuclear cell count was performed by counting the nuclei of all the respiratory epithelial cells present in the specimen in which brush cells were seen. A total of 385 cells were counted. For scanning electron microscopy, one randomly se-
42
DiMaio et al.
Fig. 2. a: Higher magnification of Figure l a showing fine filaments within the microvilli. The filaments extend into the apical cytoplasm (arrow). A desmosome is present (arrowhead). b: Higher magnificationof Figure 1b showing the microvilli in lon-
gitudinal and cross section. Filaments within the microvilli end in a terminal web (arrowheads). Uranyl acetate and lead citrate. Original magnification ~ 2 0 , 0 0 0a, and b.
lected block of tissue was attached to a specimen stub and coated with gold-palladium by the use of a Hummcr sputtering device. The specimens were viewed in their entirety under a HlTACHl SEM 5300-KEVEX 2000 scanning microscope for the presence of brush cclls.
within the microvilli. The filaments extended into the apical cytoplasm and in one cell appeared to end in a terminal web (Fig. 2a,b). The mitochondria of the brush cells were small and less plentiful in number when compared to the ciliated cells. The Golgi complexes were well developed, and several lysosomes were seen in one cell. Rough endoplasmic reticulum was sparse, but numerous free ribosomes were scattered throughout the cytoplasm. Smooth endoplasmic reticulum and pinocytotic vesicles were found in the apical cytoplasm of the cell. Tight junctions and desmosomes joined the brush cells to the adjacent ciliated cells. Brush cells were not found in sections of mainstem bronchus, intrapulmonary bronchus, bronchioles, or developing acinar epithelium by either scanning or transmission electron microscopy. We observed other cells, resembling brush cells, within the tracheal, bronchial, and bronchiolar epithelium. These cells were rare and were covered by numerous microvilli measuring 0.25-0.3 pm in thickness (Fig. 3 ) . The microvilli frequently branched and did not form a very regular border. The cells contained apical
RESULTS
We examined the tracheal epithelium of two fetuses of 14 and 19-20 weeks gestational age. Transmission electron microscopy revealed the presence of brush cells within the epithelium of the 19-20 week old fetus. These cells occurred with a frequency of 0.5% and were always found between ciliated cells. The brush cells varied in shape from columnar to globular. Compared to the neighboring ciliated cells, they presented a small surface area to the luminal surface and were wider at the base than at the apex (Fig. la,b). The brush cells were devoid of cilia and basal bodies and exhibited numerous, blunt, straight microvilli of regular length protruding into the tracheal lumen. The microvilli measured 0.25-0.3 pm in thickness. Bundles of fine filaments were present
Brush Cells in the Human Fetal Trachea
43
Fig. 3. Electron micrograph taken from a 20 week gestation fetus showing a bronchiolarcell covered with microvilli some of which branch (arrowheads). The microvilli appear to have a
fuzzy coat consistent with glycocalyx. Two basal bodies (arrows) are present within the apical cytoplasm. Uranyl acetate and lead citrate. Original magnification x 16,000.
smooth endoplasmic reticulum, pinocytotic vesicles, and a few basal bodies.
appear as stubby. The authors state that the rat brush cell does not have an “adult” appearance until 19 days postnatally.8 In light of our findings and those of previous investigators, it is probable that interspecies variation and developmental stage account for differences in the morphological appearance of brush cells, Based on our ultrastructural observation of cells bearing a regular microvillus border, we conclude that brush cells are present in the trachea of human second trimester fetuses. This ultrastructural criterion is used for all rnamalian species in order to classify a cell as a brush cell. 1.6,7,18 Cells meeting this criterion have been described in the trachea, bronchi, and bronchioles of children and a d ~ l t s . ~ - ~ However, ,~,I’ other investigators refute the existence of brush cells in man and believe that some workers are merely describing precursors of the ciliated cell.” The brush cells that we describe do not demonstrate basal bodies or cilia. We did observe cells within the the fetal airway bearing numerous microvilli and containing basal bodies. These cells resemble indifferent cells undergoing differentiation to ciliated cells, as described by McDowell et a1.,I8 and are probably not brush cells. The observation of a cell bearing microvilli and containing basal bodies indicated to several investigators the role of the “brush” cell in c i l i ~ g e n e s i s .Other ~ ~ ~ , pos~ sible functions of the brush cell have been explored. The presence of a microvillous border and apical vesicles are features of cells with an absorptive function. I,‘,’ Since
DISCUSSION
In this study we performed a rnorphologic examination of human fetal lung tissue in order to determine if brush cells are present prenatally in the extrapulmonary and intrapulmonary epithelium and developing acinar epithelium. We found brush cells in the developing tracheal epithelium of one human second trimester fetus. In mammals, brush cells are distinguished by a border of regular, straight microvilli protruding into the respiratory lumen and containing a filamentous core. 1 3 6 , 7 3 1 9 The human fetal tracheal brush cells that we describe are morphologically similar to but not identical with the brush cells of the adult rat trachea and calf bronchus.1363Z1 In the adult rat and calf the microvilli appear more stubby when compared to the human fetus. Whereas the rat microvilli measure 1.5 to 1.8 p m in width, the calf microvilli measure 0.25 p m in width. Thus, the human fetal microvilli have a width identical with that of the calf. Although the microfilaments contained within the human fetal microvilli occasionally end in a terminal web, this has not been observed in other mammals. A probable precursor of the brush cell has been identified in fetal rats.8 Based on the published electron micrograph, the microvillus border is less orderly than that of the adult rat, and the microvilli do not
44
DiMaio et al.
the Golgi apparatus appears prominent, we suggest that these cells might be involved in a secretory function rather than an absorptive process. Luciano et al. observed associations between rat tracheal brush cells and nerve fibers and speculated that brush cells may be chemoreceptors.2’ We did not observe nerve fibers adjacent to fetal brush cells. Hijiya showed that bleomycin-induced injury in rat lung causes the appearance of increased numbers of alveolar brush cells. 12,13 He suggests that the prevalence of brush cells may be the result of injury and hypoxia. We recently reported the presence of alveolar brush cells in a 4 month old infant with desquamative interstitial pneumonitis and suggested that their presence may represent a regenerative response to injury. l 7 However, we could not preclude that cells at that location were congenital in origin. Our present study has shown that brush cells do not occur normally in the developing acinar epithelium. Whether in this infant they were a response to injury or an abnormality in cellular development in unknown. In conc1usion, we have demonstrated the prenatal existence of brush cells in the human trachea. Further work is warranted in order to elucidate their function.
REFERENCES 1. Jeffery PK, Reid L. New observations of rat airway epithelium: A
2.
3. 4.
5.
quantitative and electron microscopic study. J Anat. 1975; 120: 2% -320. Reid L, Jones R. Bronchial mucosal cells. Fed Proc. 1979; 38: 191-196. Rhodin JAG. Ultrastructure and function of the human tracheal mucosa. The ciliated cell. Am Rev Respir Dis. 1966; 93:l-15. Basset F, Poirier J, LeCrom M, Turiaf J. Etude ultrastructure de l’epithelium bronchiolaire humain. Z Zellforsch Microsk Anat. 1971; 116:425-442. Watson JHL, Bnnkman GL. Electron microscopy of the epithelial
cells of normal and bronchitic human bronchus. Am Rev Respir Dis. 1964; 90:851-866. 6. Allan EM. The ultrastructure of the brush cell in bovine lung. Res Vet Sci. 1978; 25:314-317. 7. Rhodin J. Ultrastructure of the tracheal ciliated mucosa in rat and man. Ann Otol Rhino1 Laryngol. 1959; 68964-974. 8. Jeffery PK, Reid L. Ultrastructure of airway epithelium and submucosal gland during development. In: Hodson WA, ed. Development of the Lung. New York: Marcel Dekker Inc., 1977; 87134. 9. Leeson TS. The development of the trachea in the rabbit with particular reference to its fine structure. Anat Anz. 1961; 110: 214-223. 10. Christensen TG, Breuer R, Hornstra LJ, Lucey EC, Snider GL. The ultrastructure of hamster bronchial epithelium. Exp Lung Res. 1987; 13:253-277. I I . Gordon RE, Kattan M. Absence of cilia and basal bodies with a predominance of brush cells in the respiratory mucosa from a patient with immotile cilia syndrome. Ultrastruct Pathol. 1984; 6:4S-49. 12. Hijiya K, Okada Y, Tankdwa H. Ultrastructural study of the alveolar brush ccll. J Electron Microsc (Tokyo). 1977; 26321329. 13. Hijiya K. Electron microscope study of the alveolar brush cell. J Electron Microsc (Tokyo). 1978; 27:223-227. 14. Filippenke LN. Light and electron microscopic study of the alveolar brush cells of the rat lung. Biull Eksp Biol Med. 1978; 86592-596. 15. Meyrick B, Reid L. The alveolar brush cell in rat lung-a third pneumonocyte. J Ultrastruct Res. 1968; 23:7 1-80. 16. Foliguet B, Grignon G. Le pneumocyte de type 111. La cellule alveolaire en brosse du poumon du rat. Etude en microscopie electronique transmission. Poumon-Coeur. 1980; 36: 149-1 53. 17. DiMaio MF, Dische R, Gordon RE, Kattan M. Alveolar brush cells in an infant with desquamative interstitial pneumonitis. Pediatr Pulmonol. 1988; 4:185-191. 18. McDowell EM, Barett LA, Glavin F, Harris CC, Trump BF. The respiratory epithelium, I . Human bronchus. JNC. 61539-545, 1978. 19. Scammon RR. Two simple nomographs for estimating age and some of the major external dimensions of the human fetus. Anat Rec. h8:221-225, 1937. 20. Sato T. A modified method for lead staining of thin sections. J Electron Microsc (Tokyo). 16: 133, 1967. 21. Luciano L, Reale E, Ruska H. Uber eine chemorezeptive sinneszelle in der trachea der ratte. Z Zellforsch. 85:350-375, 1968.
