Pseudoglandular Stage Of Lung Development Research Articles

Sex-Specific Differences in MicroRNA Expression During Human Fetal Lung Development.

Background: Sex-specific differences in fetal lung maturation have been well described; however, little is known about the sex-specific differences in microRNA (miRNA) expression during human fetal lung development. Interestingly, many adult chronic lung diseases also demonstrate sex-specific differences in prevalence. The developmental origins of health and disease hypothesis suggests that these sex-specific differences in fetal lung development may influence disease susceptibility later in life. In this study, we performed miRNA sequencing on human fetal lung tissue samples to investigate differential expression of miRNAs between males and females in the pseudoglandular stage of lung development. We hypothesized that differences in miRNA expression are present between sexes in early human lung development and may contribute to the sex-specific differences seen in pulmonary diseases later in life. Methods: RNA was isolated from human fetal lung tissue samples for miRNA sequencing. The count of each miRNA was modeled by sex using negative binomial regression models in DESeq2, adjusting for post-conception age, age2, smoke exposure, batch, and RUV factors. We tested for differential expression of miRNAs by sex, and for the presence of sex-by-age interactions to determine if miRNA expression levels by age were distinct between males and females. Results: miRNA expression profiles were generated on 298 samples (166 males and 132 females). Of the 809 miRNAs expressed in human fetal lung tissue during the pseudoglandular stage of lung development, we identified 93 autosomal miRNAs that were significantly differentially expressed by sex and 129 miRNAs with a sex-specific pattern of miRNA expression across the course of the pseudoglandular period. Conclusion: Our study demonstrates differential expression of numerous autosomal miRNAs between the male and female developing human lung. Additionally, the expression of some miRNAs are modified by age across the pseudoglandular stage in a sex-specific way. Some of these differences in miRNA expression may impact susceptibility to pulmonary disease later in life. Our results suggest that sex-specific miRNA expression during human lung development may be a potential mechanism to explain sex-specific differences in lung development and may impact subsequent disease susceptibility.

Pre- and postnatal exposure of mice to concentrated urban PM2.5 decreases the number of alveoli and leads to altered lung function at an early stage of life

Gestational exposure to air pollution is associated with negative outcomes in newborns and children. In a previous study, we demonstrated a synergistic negative effect of pre- and postnatal exposure to PM2.5 on lung development in mice. However, the means by which air pollution affects development of the lung have not yet been identified.In this study, we exposed pregnant BALB/c mice and their offspring to concentrated urban PM2.5 (from São Paulo, Brazil; target dose 600 μg/m3 for 1 h daily). Exposure was started on embryonic day 5.5 (E5.5, time of placental implantation). Lung tissue of fetuses and offspring was submitted to stereological and transcriptomic analyses at E14.5 (pseudoglandular stage of lung development), E18.5 (saccular stage) and P40 (postnatal day 40, alveolarized lung). Additionally, lung function and cellularity of bronchoalveolar lavage (BAL) fluid were studied in offspring animals at P40.Compared to control animals that were exposed to filtered air throughout gestation and postnatal life, PM-exposed mice exhibited higher lung elastance and a lower alveolar number at P40 whilst the total lung volume and cellularity of BAL fluid were not affected. Glandular and saccular structures of fetal lungs were not altered upon gestational exposure; transcriptomic signatures, however, showed changes related to DNA damage and its regulation, inflammation and regulation of cell proliferation. A differential expression was validated at E14.5 for the candidates Sox8, Angptl4 and Gas1.Our data substantiate the in utero biomolecular effect of gestational exposure to air pollution and provide first-time stereological evidence that pre- and early life-postnatal exposure compromise lung development, leading to a reduced number of alveoli and an impairment of lung function in the adult mouse.

Malformations adénomatoïdes kystiques du poumon : diagnostic, prise en charge, hypothèses physiopathologiques

Congenital cystic adenomatoid malformations (CCAM) of the lung are the most frequent congenital lung malformations. Their diagnosis is based on histological features. CCAM consist of bronchopulmonary cystic lesions which are classified according to the presence and cysts size. Type I CCAM are composed of large cysts (>2 cm) lined by a columnar pseudostratified epithelium. Type II CCAM contain multiple small cystic lesions (<1 cm) lined by a flattened cuboidal epithelium. Type III CCAM are more solid and contain immature structures resembling the pseudoglandular stage of lung development. Ultrasonography (US) allows early detection during the second trimester of pregnancy as cystic, and/or hyperechoic fetal lung lesions. Although most CCAM remain asymptomatic, CCAM can cause polyhydramnios or fetal hydrops, respiratory distress at birth, infections and pneumothoraces during infancy, and may give rise to malignancies. Serial US allow detection of complications, and planification of delivery. Complicated forms require an urgent treatment. In fetuses with a macrocystic life-threatening lesion, a thoraco-amniotic shunt can be placed. Microcystic compressive forms may respond to prenatal steroids. Post-natal symptomatic lesions require early surgery. The treatment of asymptomatic forms remains controversial. Some recommend a non-operative approach with a long-term clinical and radiological following, whereas other favour a preventive surgical excision. The origin of CCAM remains unknown. Recent advances suggest a transient and focal abnormality in lung development which may result from an airway obstruction. This article reviews the diagnosis, treatment, and pathophysiology of CCAM.

Expression of A-, B-, and ΓENAC in Human Fetal and Newborn Lung

Background: Airway epithelial sodium channel ENaC is critical to perinatal lung fluid clearance. Data on ENaC protein expression in developing human lung are limited and absent for infants with RDS or BPD. Objective: To characterize the expression of α-, β-, and γENaC expression in lung tissue from human fetuses and newborns with and without RDS or BPD. Methods: Lung tissue was obtained from autopsies (n = 23) from fetuses (GA 12 - 21 wks), preterm infants with (GA 25 - 28 wks) or without RDS (GA 23 - 25 wks), and infants with BPD (GA 24 - 30 wks at birth). The primary antibodies were rabbit polyclonal αENaC (1:60 Sigma-Aldrich Inc, MO, US), βENaC (1:2250 Aviva Systems Biology, CA, US) or γENaC (1:200 Lifespan Biosciences Inc, WA, US). Results: In fetal lung developing respiratory, glandular epithelium stained with α-, β-, and γENaC from 12+3 wks GA. In preterm infants without RDS expression of α-, β-, and γENaC was apparent in single type II pneumocytes, while type I pneumocytes remained unstained. In RDS and BPD the expression of ENaC subunits in alveolar epithelium was largely absent but reactive, hyperplastic type II pneumocytes stained strongly for α- and βENaC. Throughout development, and in infants with RDS or BPD α-, β-, and γENaC stained the bronchiolar epithelium. Conclusions: In human fetal lung all three ENaC subunits were expressed during the pseudoglandular stage of lung development. Expression of ENaC subunits in bronchioles may indicate a role for conductive airways in perinatal lung liquid clearance.

RUNX transcription factors: association with pediatric asthma and modulated by maternal smoking.

Intrauterine smoke exposure (IUS) is a strong risk factor for development of airways responsiveness and asthma in childhood. Runt-related transcription factors (RUNX1-3) have critical roles in immune system development and function. We hypothesized that genetic variations in RUNX1 would be associated with airway responsiveness in asthmatic children and that this association would be modified by IUS. Family-based association testing analysis in the Childhood Asthma Management Program genome-wide genotype data showed that 17 of 100 RUNX1 single-nucleotide polymorphisms (SNPs) were significantly (P < 0.03-0.04) associated with methacholine responsiveness. The association between methacholine responsiveness and one of the SNPs was significantly modified by a history of IUS exposure. Quantitative PCR analysis of immature human lung tissue with and without IUS suggested that IUS increased RUNX1 expression at the pseudoglandular stage of lung development. We examined these associations by subjecting murine neonatal lung tissue with and without IUS to quantitative PCR (N = 4-14 per group). Our murine model showed that IUS decreased RUNX expression at postnatal days (P)3 and P5 (P < 0.05). We conclude that 1) SNPs in RUNX1 are associated with airway responsiveness in asthmatic children and these associations are modified by IUS exposure, 2) IUS tended to increase the expression of RUNX1 in early human development, and 3) a murine IUS model showed that the effects of developmental cigarette smoke exposure persisted for at least 2 wk after birth. We speculate that IUS exposure-altered expression of RUNX transcription factors increases the risk of asthma in children with IUS exposure.

S32 Cyclical mechanical stretch enhances the pro-fibrotic responses of primary embryonic foetal fibroblasts, but not ADAM33 expression

RationaleA Disintegrin And Metalloprotease (ADAM)33 is a susceptibility gene associated with asthma, bronchial hyperresponsiveness (BHR) and reduced lung function in young children. It is selectively expressed in mesenchymal cells, including...

Selective Toll-Like Receptor Expression in Human Fetal Lung

Toll-like receptors (TLRs) are critical components of the innate immune system, acting as pattern recognition molecules and triggering an inflammatory response. TLR associated gene products are of interest in modulating inflammatory-related pulmonary diseases of the neonate. The ontogeny of TLR-related genes in human fetal lung has not been previously described and could elucidate additional functions and identify strategies for attenuating the effects of fetal inflammation. We examined the expression of 84 TLR-related genes on 23 human fetal lung samples from three groups with estimated ages of 60 (57-59 d), 90 (89-91 d), and 130 (117-154 d) d. By using a false detection rate algorithm, we identified 32 genes displaying developmental regulation with TLR2 having the greatest up-regulation of TLR genes (9.2-fold increase) and TLR4 unchanged. We confirmed the TLR2 up-regulation by examining an additional 133 fetal lung tissue samples with a fluorogenic polymerase chain reaction assay (TaqMan) and found an exponential best-fit curve during the study time. The best-fit curve predicts a 6.1-fold increase from 60 to 130 d. We conclude that TLR2 is developmentally expressed from the early pseudoglandular stage of lung development to the canalicular stage.

An FGF–WNT gene regulatory network controls lung mesenchyme development

Lung mesenchyme is a critical determinant of the shape and size of the lung, the extent and patterning of epithelial branching, and the formation of the pulmonary vasculature and interstitial mesenchymal components of the adult lung. Fibroblast growth factor 9 (FGF9) is a critical regulator of lung mesenchymal growth; however, upstream mechanisms that modulate the FGF mesenchymal signal and the downstream targets of mesenchymal FGF signaling are poorly understood. Here we have identified a robust regulatory network in which mesenchymal FGF signaling regulates β-Catenin mediated WNT signaling in lung mesenchyme. By conditionally inactivating β-Catenin in lung mesenchyme, we show that mesenchymal WNT-β-Catenin signaling is essential for lung development and acts to regulate the cell cycle G1 to S transition and the FGF responsiveness of mesenchyme. Together, both FGF and WNT signaling pathways function to sustain mesenchymal growth and coordinate epithelial morphogenesis during the pseudoglandular stage of lung development.

QS144. Endotoxin Alters Branching Morphogenesis and Matrix Deposition of The Fetal Lung

Background: The effects of immaturity and hypoplasia of the premature lung can be affected by proinflammatory stimuli in late gestation or the postnatal period from acute lung injury secondary to intensive ventilatory management or the metabolic consequences of surgery. These stimuli alter alveolarization and contribute to bronchopulmonary dysplasia. While prior research has focused primarily on late gestational effects of inflammation on alveolar development, we sought to study whether early gestational exposure to endotoxin affects branching morphogenesis, during the critical pseudoglandular stage of lung development. Methods: Gestational day 15 (E15) fetal rat lung explants (term=22 d) at the pseudoglandular stage were incubated at 37°C in 95% air/5% CO2 as submersion cultures on inserts in serum-free BGJb media supplemented with antibiotics and ascorbic acid. Explants were treated with either 200 ng/mL or 2 μg/mL LPS with controls and examined daily by phase microscopy. After 5 days, explants were fixed in 4% formaldehyde, paraffin embedded and sectioned at 5 μm in the coronal plane. Immunohistochemical analysis was performed with PECAM to define endothelial cells, VEGF to examine endothelial mitogenesis and COX-2 antibodies as a marker for prostaglandin synthesis. Air space fraction was analyzed with Image J software. Results: Phase contrast microscopy and hematoxylin-eosin histology revealed progressive, dose-related changes in air sac contraction and interstitial thickening. Compared with control E15 explants, day 5 explants incubated with high dose LPS demonstrated thickened and shrunken airway sacs with stunted branching and increased matrix deposition in interstitial areas (graph). By immunohistochemical staining, COX-2 was quantitatively increased after high dose LPS exposure, while PECAM was reduced. VGEF expression was unaltered. Conclusions: This is the first evidence of an inflammatory effect of LPS on the early phase of lung development in the fetal rat, affecting branching morphogenesis during the pseudoglandular phase. Fetal endothelial cells are clearly affected, while COX-2 elevation suggests activation of an as yet undefined fetal pulmonary inflammatory cascade. We speculate that the proinflammatory stimuli that may ultimately lead to abnormal pulmonary development affect epithelial-mesenchymal interaction and differentiation at a much earlier gestational age than was previously recognized.

APOPTOSIS OF MESENCHYMAL CELLS DURING THE PSEUDOGLANDULAR STAGE OF LUNG DEVELOPMENT AFFECTS BRANCHING MORPHOGENESIS

Apoptosis has been implicated as an important process in the development of several organ systems, including limbs, kidneys, and the heart. In developing murine lungs, the authors found that apoptosis was more predominant during the pseudoglandular stage of lung development than during the saccular stage, with 93% of the apoptotic structures in the mesenchyme. Murine lung explants cultured in the presence of zinc chloride and aurintricarboxylic acid, 2 blockers of endonuclease function, showed decreased branching. These observations suggest that apoptosis predominates in mesenchymal cells during the pseudoglandular stage of lung development, and may be important for normal progression of lung branching morphogenesis.

Effects of acute alcohol intoxication in the second trimester of pregnancy on development of the murine fetal lung

We hypothesized that administration of alcohol during the second trimester of gestation at the pseudoglandular phase of lung development might lead to aberrant differentiation and growth, similar to that seen in congenital cystic adenomatoid malformation in human. We further hypothesized that these effects would be apparent morphologically and by altered Hoxb5 expression. C57BL/6J mice, exposed to ethanol at embryonic day (E) 11.5 to E13.5, which corresponds to the pseudoglandular stage of lung development, were examined at E18.5. The lungs were analyzed histologically by immunostaining. The average body and lung weight of alcohol-exposed (AE) fetuses were lower than those of control fetuses, the reduction in lung mass being more than the body weight. Histology showed that AE lungs were less developed and exhibited higher expression of Hoxb5 in AE lungs than controls. Alcohol exposure at E13.5 affected fetal lung development, with delayed differentiation and increased Hoxb5.

Embryonic Essential Myosin Light Chain Regulates Fetal Lung Development in Rats

Congenital diaphragmatic hernia (CDH) is currently the most life-threatening congenital anomaly the major finding of which is lung hypoplasia. Lung hypoplasia pathophysiology involves early developmental molecular insult in branching morphogenesis and a late mechanical insult by abdominal herniation in maturation and differentiation processes. Since early determinants of lung hypoplasia might appear as promising targets for prenatal therapy, proteomics analysis of normal and nitrofen-induced hypoplastic lungs was performed at 17.5 days after conception. The major differentially expressed protein was identified by mass spectrometry as myosin light chain 1a (MLC1a). Embryonic essential MLC1a and regulatory myosin light chain 2 (MLC2) were characterized throughout normal and abnormal lung development by immunohistochemistry and Western blot. Disruption of MLC1a expression was assessed in normal lung explant cultures by antisense oligodeoxynucleotides. Since early stages of normal lung development, MLC1a was expressed in vascular smooth muscle (VSM) cells of pulmonary artery, and MLC2 was present in parabronchial smooth muscle and VSM cells of pulmonary vessels. In addition, early smooth muscle differentiation delay was observed by immunohistochemistry of alpha-smooth muscle actin and transforming growth factor-beta1. Disruption of MLC1a expression during normal pulmonary development led to significant growth and branching impairment, suggesting a role in branching morphogenesis. Both MLC1a and MLC2 were absent from hypoplastic fetal lungs during pseudoglandular stage of lung development, whereas their expression partially recovered by prenatal treatment with vitamin A. Thus, a deficiency in contractile proteins MLC1a and MLC2 might have a role among the early molecular determinants of lung hypoplasia in the rat model of nitrofen-induced CDH.

Ectopic expression of C/EBPα in the lung epithelium disrupts late lung development

The lung develops from the endoderm through a process of branching morphogenesis. This process is highly active during the pseudoglandular stage of lung development and continues into the canalicular stage, resulting in the formation of terminal sacs. CCAAT/enhancer binding proteins (C/EBPs) are transcription factors regulating central aspects of differentiation and proliferation. We report here the developmental expression of C/EBPalpha, -beta, and -delta in the lung. C/EBPalpha exhibits a dynamic expression pattern and is first detected during the late pseudoglandular stage. At this stage, expression is observed in a subset of epithelial cells in the distal parts of the branching tubules. The expression of C/EBPalpha is confined to nonproliferating cells. To examine the role of C/EBPalpha in lung development, we generated transgenic mice ectopically expressing C/EBPalpha in the lung epithelium using the human surfactant protein C promoter. Lungs from these mice were of normal size but exhibited a phenotype characterized by fewer and larger developing epithelial tubules, indicating that the branching process was affected. No effects on overall proliferation or cellular differentiation were observed. When this phenotype was compared with that of mice carrying a targeted mutation of the Cebpa gene, the Cebpa-/- mice exhibited a similar developmental phenotype. In conclusion, our results show a role for C/EBPalpha in lung development and suggest a function in the later stages of lung branching morphogenesis.

Lung regeneration: Implantation of fetal rat lung fragments into adult rat lung parenchyma

The capability of regeneration of lung tissues in adults is limited after chronic destruction. Bone marrow-derived stem cells, retinoic acid, growth factors, and other approaches have been attempted to promote or facilitate this process. We hypothesized that fetal lung tissues, with great potential for growth and differentiation, could be used for lung regeneration. Day 17 fetal lung tissue fragments at the pseudoglandular stage of lung development from Lewis rats were implanted into adult Lewis rat lungs. For group 1, fetal lung fragments were injected into the adult left lung parenchyma; for group 2, fetal fragments were injected with the left lung partially resected; for group 3, adult fragments were injected; and for groups 4 and 5, fetal fragments were implanted into the omentum and subcutaneous tissue, respectively. The grafts implanted into pulmonary parenchyma were differentiated with opening of the alveolar space after 4 weeks and were advanced further with morphologic features similar to those of neonatal lungs after 8 and 12 weeks. The implants were connected with pulmonary circulation determined by means of perfusion with India ink. These changes appeared to be further enhanced in animals with partial lung resection that might have facilitated the maturation of implanted fetal lung tissues through mechanical factors, soluble factors, or both. Fetal lung tissues did not mature when implanted into the omentum or subcutaneous tissue. Adult lung fragments did not expand after being reimplanted back into the same animal. Fetal lung tissue might be an option for further investigation into lung regeneration.

Angiogenesis and morphogenesis of murine fetal distal lung in an allograft model.

Neovascularization is crucial to lung morphogenesis; however, factors determining vessel growth and formation are poorly understood. The goal of our study was to develop an allograft model that would include maturation of the distal lung, thereby ultimately allowing us to study alveolar development, including microvascular formation. We transplanted 14-day gestational age embryonic mouse lung primordia subcutaneously into the back of nude mice for 3.5-14 days. Lung morphogenesis and neovascularization were evaluated by light microscopy, in situ hybridization, and immunohistochemical techniques. Embryonic 14-day gestational age control lungs had immature structural features consistent with pseudoglandular stage of lung development. In contrast, 14 days after subcutaneous transplantation of a 14-day gestational age lung, the allograft underwent significant structural morphogenesis and neovascularization. This was demonstrated by continued neovascularization and cellular differentiation, resulting in mature alveoli similar to those noted in the 2-day postnatal neonatal lung. Confirmation of maturation of the allograft was provided by progressive type II epithelial cell differentiation as evidenced by enhanced local expression of mRNA for surfactant protein C and a threefold (P < 0.008) increase in vessel formation as determined by immunocytochemical detection of platelet endothelial cell adhesion molecule-1 expression. Using the tyrosine kinase Flk-1 receptor (flk-1) LacZ transgene embryos, we determined that the neovascularization within the allograft was from the committed embryonic lung endothelium. Therefore, we have developed a defined murine allograft model that can be used to study distal lung development, including neovascularization. The model may be useful when used in conjunction with an altered genetic background (knockout or knock in) of the allograft and has the further decided advantage of bypassing placental barriers for introduction of pharmacological agents or DNA directly into the lung itself.

Heparin inhibits lung branching morphogenesis: potential role of smooth muscle cells in cleft formation.

Lung branching morphogenesis is the process by which the embryonic lung undergoes repetitive branching to form the bronchial tree. This process occurs during the pseudoglandular stage of lung development and requires epithelial-mesenchymal interactions. Coinciding with lung branching morphogenesis is the appearance of parabronchial smooth muscle cells (PSMCs) and the accumulation of extracellular matrices (ECMs) around the developing airways. The authors previously reported in preliminary form that heparin prevents the branching of murine lung explants (Roman et al., Am Rev Respir Dis. 1991; 143:A401); this article corroborates those early observations and expands them by demonstrating that heparin results in disruption of PSMC distribution and abnormal organization of ECMs around the developing airways. These changes were associated with inhibition of lung branching morphogenesis in the absence of effects on cell proliferation. The data provide further support for the role of ECMs in lung branching morphogenesis, and points to PSMCs as potential players in this process.

Heparin Inhibits Lung Branching Morphogenesis: Potential Role of Smooth Muscle Cells in Cleft Formation

Lung branching morphogenesis is the process by which the embryonic lung undergoes repetitive branching to form the bronchial tree. This process occurs during the pseudoglandular stage of lung development and requires epithelial-mesenchymal interactions. Coinciding with lung branching morphogenesis is the appearance of parabronchial smooth muscle cells (PSMCs) and the accumulation of extracellular matrices (ECMs) around the developing airways. The authors previously reported in preliminary form that heparin prevents the branching of murine lung explants (Roman et al., Am Rev Respir Dis. 1991; 143:A401); this article corroborates those early observations and expands them by demonstrating that heparin results in disruption of PSMC distribution and abnormal organization of ECMs around the developing airways. These changes were associated with inhibition of lung branching morphogenesis in the absence of effects on cell proliferation. The data provide further support for the role of ECMs in lung branching morphogenesis, and points to PSMCs as potential players in this process.

Ontogeny of neuroepithelial bodies: Correlations with mitogenesis and innervation

This paper summarizes current knowledge and advances speculation about the formation of the neuroendocrine system of mammalian lungs (comprising uninnervated solitary and clustered small-granule cells and innervated neuroepithelial bodies). It relates the initial appearance of neuroendocrine cells to regulation of mitotic activity in the epithelium during the development of the lung and pays special attention to the later ingrowth of nerves that converts some of them into neuroepithelial bodies, structures considered ideally adapted to function as chemoreceptors. A few original observations from ongoing immunohistochemical, electron microscopic, and analytical studies have been included here and there to point the discussion. The neuroendocrine cells are derived from undifferentiated precursors present in the endodermal pulmonary epithelium. At an early pseudoglandular stage of lung development these precursors begin to differentiate into neuroendocrine small-granule cells, commencing in the larynx and upper trachea, and expanding centrifugally into pulmonary airways almost as rapidly as these are laid down. Subsequently many of the intrapulmonary small-granule cell clusters become innervated. This event, the delayed appearance of small-granule cells synthesizing other than the dominant peptides and amines (calcitonin gene-related peptide and serotonin in rodents, gastrin-releasing peptide and serotonin in human beings), and other regional adjustments yield the population distribution present in the lungs of adults. Neuroendocrine cell precursors normally differentiate into typical serotonin- or peptide-synthesizing small-granule cells without requiring direct contact by nerves, and dissociated cells from a previously innervated population continue to exhibit physiological characteristics of oxygen sensors despite the loss of contact with nerves. Development of the innervation occurs in stages. Small-granule cell clusters are reached first by ganglion cells derived from pulmonary neuroblasts and later on by processes of extrinsic sensory nerves. The latter not only convey information to the central nervous system but also serve in a variety of ways to extend the neuroepithelial bodies' sphere of influence within the lung itself. Microsc. Res. Tech. 37:43–61, 1997. © 1997 Wiley-Liss, Inc.

Ontogeny of neuroepithelial bodies: correlations with mitogenesis and innervation.

This paper summarizes current knowledge and advances speculation about the formation of the neuroendocrine system of mammalian lungs (comprising uninnervated solitary and clustered small-granule cells and innervated neuroepithelial bodies). It relates the initial appearance of neuroendocrine cells to regulation of mitotic activity in the epithelium during the development of the lung and pays special attention to the later in growth of nerves that converts some of them into neuroepithelial bodies, structures considered ideally adapted to function as chemoreceptors. A few original observations from ongoing immunohistochemical, electron microscopic, and analytical studies have been included here and there to point the discussion. The neuroendocrine cells are derived from undifferentiated precursors present in the endodermal pulmonary epithelium. At an early pseudoglandular stage of lung development these precursors begin to differentiate into neuroendocrine small-granule cells, commencing in the larynx and upper trachea, and expanding centrifugally into pulmonary airways almost as rapidly as these are laid down. Subsequently many of the intrapulmonary small-granule cell clusters become innervated. This event, the delayed appearance of small-granule cells synthesizing other than the dominant peptides and amines (calcitonin gene-related peptide and serotonin in rodents, gastrin-releasing peptide and serotonin in human beings), and other regional adjustments yield the population distribution present in the lungs of adults. Neuroendocrine cell precursors normally differentiate into typical serotonin- or peptide-synthesizing small-granule cells without requiring direct contact by nerves, and dissociated cells from a previously innervated population continue to exhibit physiological characteristics of oxygen sensors despite the loss of contact with nerves. Development of the innervation occurs in stages. Small-granule cell clusters are reached first by ganglion cells derived from pulmonary neuroblasts and later on by processes of extrinsic sensory nerves. The latter not only convey information to the central nervous system but also serve in a variety of ways to extend the neuroepithelial bodies' sphere of influence within the lung itself.

Cell-Matrix Interactions in Gland Development in the Lung

The tubulo-acinar glands of the conducting airways participate in lung host defense and the pathogenesis of chronic inflammatory airway disease. Models of gland development may provide clues to mechanisms that promote hypersecretion in diseases such as chronic bronchitis and cystic fibrosis. The developmental anatomy of airway glands was described 30 years ago, but the regulation of gland formation remains a mystery. During the pseudoglandular stage of lung development, poorly differentiated surface epithelial cells grow radially from the lumen and invade the airway submucosa. The basal lamina is remodeled during this process and animal studies indicate a correlation between the expression of the 72-kD gelatinase (MMP-2) and gland morphogenesis. Cell culture models with lung epithelial and mesenchymal elements mimic early gland morphogenesis. Studies using this model have corroborated the involvement of 72-kD gelatinase in the formation of gland-like structures. While the secretion and activation of 72-kD gelatinase is correlated with epithelial matrix invasion, the presence of activated 72-kD gelatinase is not by itself sufficient to permit epithelial matrix invasion. Whether the 72-kD gelatinase participates in a more complex proteolytic cascade or whether appropriate localization of the enzyme is critical to airway epithelial morphogenesis is the focus of ongoing investigation.