Tracheal Secretory Cells Research Articles

Regulation of PACAP receptors in respiratory distress syndrome (RDS) of preterms and newborns

s of the 51st Workshop for Pediatric Research 51st Workshop for Pediatric Research Gottingen, Germany 16-17 April 2015 This supplement has not been sponsored. Meeting abstracts Background Prevalence of respiratory distress syndrome (RDS) caused by primary surfactant deficiency reaches 50% in infants below 32 weeks of gestational age. Increased alveolar surface tension leads to atelectatic and/or dystelectatic lung areas followed by inflammation resulting in development of a bronchopulmonary dysplasia (BPD) in 8% of these children. Standard RDS treatment consists of administration of exogenous surfactant. With the introduction of less invasive application (LISA) of exogenous surfactant, the use of Surfactant has significantly increased. Pituitary adenylate cyclase-activating polypeptide (PACAP) and its receptors (VPAC1, VPAC2, PAC1) trigger inflammatory reactions as well as anti-inflammatory processes. Materials and methods Using cell cultures, FACS and ELISA methods we analyzed surfactant effects on inflammation and PACAP-regulation and of its receptors in vivo and ex vivo. Results We were able to show that lipopolysaccharide (LPS) in prestimulated cells (PBMC) after surfactant treatment resulted in a significant up-regulation of pro-inflammatory interleukin (IL)-8. In tracheal secretory cells of surfactant-treated neonates we found an up-regulation of pro-inflammatory receptor VPAC1. Furthermore, in cell cultures we identified changes in the proliferation rates. Amniotic infection syndrome (AIS) is one of the major causes for premature birth resulting in RDS requiring exogenous surfactant. After pre-stimulation of PBMCs and subsequent addition of surfactant, pro-inflammatory signals were fortified. These results were confirmed by analysis of tracheal secretions of preterms and newborns. Conclusions Current findings may imply that safety and efficacy of routine surfactant administration should be reassessed as surfactant potentially triggers pulmonary inflammation and thus may contribute to BPD development.

Vitamin A deficiency and inflammation: the pivotal role of secretory cells in the development of atrophic, hyperplastic and metaplastic change in the tracheal epithelium in vivo.

We showed previously that the proliferation of hamster airway secretory cells decreases during vitamin A deficiency (VAD) but later increases when submucosal inflammation develops (Virchows Arch [B] 59:231-242, 1990). This observation has important biological implications since two morphological extremes (atrophy and quiescence versus hyperplasia and hyperproliferation) are reported in the literature for VAD tracheal epithelium in vivo. In the present study, histological slides of tracheal rings from 35-day-old control and VAD hamsters (Virchows Arch [B] 45:197-219, 1984) were reviewed again. Rings from VAD hamsters were selected based on the absence or presence of a florid submucosal inflammation. Quantitative analyses were made on the cartilaginous part of rings from the anterior third of the trachea. When inflammation was absent, a mucociliary pseudostratified epithelium was, for the most part, maintained. The mitotic rate (MR, 6 h colchicine blockade) of secretory cells was markedly reduced (29-fold) but that of basal cells was not changed significantly. Moreover, cell density was not changed by VAD but ciliated cells and secretory cells were decreased and basal cells were increased, proportionally. We call this "minimal morphological change." Thinning (atrophy) of the minimally changed epithelium was associated with focal cell sloughing. Small scattered foci of epidermoid metaplasia (multiple layers of highly keratinized cells which were extremely flat, so that the epithelium was thin and attenuated) were also seen. We call this "atrophic epidermoid metaplasia." When inflammation was present, hyperplastic changes (stratification and epidermoid metaplasia) predominated and cells were in mitosis at all epithelial levels (low, middle, superficial) except in the most superficial (terminally differentiated) squames. The tracheal epithelium was thickened and hypercellular. The cells were piled up at the stratified lesions, and epithelial height, cell density and epithelial MR were significantly increased compared with the non-inflamed VAD epithelium. The effects of VAD and inflammation on cell proliferation were analyzed further by studying 7 h bromodeoxyuridine (BrdU) labelling patterns of cells in VAD tracheal epithelium, with and without submucosal inflammation. In addition, inflammation was induced in "minimally changed epithelium" by mild mechanical injury. The BrdU labelling patterns confirmed that DNA synthesis by secretory cells is reduced markedly by VAD. However, this suppression is overidden by the influx of inflammatory cells (the nature of the stimulus is unknown). The results indicate that the morphological contrasts (atrophy and hyperplasia) seen in the trachea during VAD in vivo are related to extremes in proliferation rates of tracheal secretory cells, regulated by VAD alone (minimal replication) and by inflammation (maximal replication).

Lectin Cytochemistry Reveals Differences between Hamster Trachea and Bronchus in the Composition of Epithelial Surface Glycoconjugates and in the Response of Secretory Cells to Neutrophil Elastase

Hamsters exposed to an intratracheal instillation of human neutrophil elastase (HNE) accumulate an abnormally high number of secretory granules in bronchial but not tracheal epithelial cells. We employed lectin cytochemistry to investigate possible differences in the epithelial cell surface glycoconjugate layer in trachea compared to bronchus which might explain the regional dissimilarity in response to HNE. Portions of glutaraldehyde-fixed trachea and bronchi were incubated in one of several ferritin-labeled lectins prior to embedding for transmission electron microscopy. Lectins from Ricinus communis, Helix pomatia, and Triticum vulgaris bound to the surface of tracheal secretory cells in moderate to profuse amounts, while most bronchial secretory cells showed little or no label with these lectins. Gold-labeled Helix pomatia agglutinin (HPA), a lectin specific for secretory cells, showed a decrease in surface binding to all tracheal secretory cell types within 2 h of HNE instillation, compared to saline controls. In contrast, the majority of bronchial secretory cells showed an HNE-induced increase in surface label from extremely low levels in saline controls. The low levels of lectin binding to bronchial cells, in contrast to the trachea, may indicate the lack of a protective surface glycoconjugate coat, thus explaining the vulnerability of these cells to HNE. The rise in number of accessible HPA binding sites on the surface of bronchial secretory cells exposed to HNE may represent an important event in the pathologic accumulation of secretory granules by these cells.

An immunohistochemical characterization of rhesus monkey respiratory secretions using monoclonal antibodies.

A series of 12 monoclonal antibodies was made against mucous and serous components of rhesus monkey trachea. These antibodies were used to localize secretory products in the respiratory epithelium by immunohistochemical methods. Mucous cells of the tracheal surface epithelium and submucosal glands were shown by histochemical methods to be a homogeneous population containing periodate-reactive sulfated glycoconjugates. Immunohistochemical staining using the monoclonal antibodies on glycol methacrylate sections serial to those stained histochemically revealed a more antigenically heterogeneous population of secretory cells. Four distinct staining patterns were observed with the 12 monoclonal antibodies. Eight antibodies reacted with most, but not all, mucous and serous cells. Two antibodies recognized subpopulations of secretory cells. One antibody stained tracheal mucous cells with a concentration of reaction product on the luminal border, and one antibody stained only serous cells within the glands. Ten of the 12 antibodies were shown to react in an ELISA with the high molecular weight void volume containing mucous glycoproteins separated on a Sepharose CL-4B gel filtration column. None of the antibodies recognized blood group antigens. We conclude: (1) that production of a large panel of monoclonal antibodies to respiratory tract mucins from primates is feasible, (2) that the monoclonal antibodies will recognize epitopes on biochemically identifiable glycoproteins, and (3) that the intracellular mucous products of tracheal secretory cells exhibit greater heterogeneity than is detectable by conventional histochemistry.

Effect of BR-227, a new bromhexine derivative, on secretory activities of tracheal secretory cells.

The effects of BR-227 on secretory activities of canine tracheal secretory cells including behavior of mucus glycoproteins were investigated histologically and histochemically. Following BR-227 treatment at a concentration of alcian blue at pH 2.5 and periodic acid-Schiff (AB (pH 2.5)/PAS) decreased in a concentration-dependent manner. Furthermore, a decrease in the thickness of the acini of submucosal glands and in marked increase in the ratio of acinar inner diameter of the gland to tracheal wall were induced after application of BR-227 at a concentration range of 10(-7) to 10(-4) M. The numbers of goblet and submucosal glandular cells which stained blue and purple with AB (pH 2.5)/PAS were decreased by BR-227 treatment in a concentrations-dependent way, whereas the cells which stained red were markedly increased. In the experiment using a combination of AB at pH 1.0 and PAS, a decrease in sulfated glycoproteins in those secretory cells was observed. Total saccharide and protein concentrations in the incubation fluid increased with BR-227 treatment, while N-acetylhexomsamine concentration tended to decrease. These findings suggest that BR-227 stimulates secretory activities of both goblet cells and submucosal glands, and lowers mucus viscosity in the secretory cells.

Pharmacological studies on the respiratory tract (Rept. 71): Effects of amhroxol and fominohen on activities of tracheal secretory cells and alveolar type II cells

The effects of amhroxol(AX) and fominoben(FN) (bromhexine derivatives) on activities of tracheal secretory cells and on secretion of pulmonary surfactant were investigated histologically, histochemically or biochemically. Following treatment :f the isolated canine trachea with AX(10−7-10−4M), the number of total glycoprotein-containing goblet cells(GC) was unaltered, while the ratio of acinar inner diameter of submucosal gland(SG) to wall(AWR) increased. AX decreased the i cid glycoprotein(AGP) content in GC and SG and increased. the neutral glycoprotein content in these cells. FN(10−7-10−4) induced, however, no significant change in GC and SG. After AX treatment, phosphatidylcholine (PC), disaturated PC (3PC) and phosphatidylglycerol contents in the pulmonary lavage fluid of rats increased. On the other hand, FN had no such effect, except that the ratio of DSPC content to PC content was increased. These findings suggest that AX has coth secretagogic action on SG and mucolytic action toward AGP in GC and SG and, furthermore, a stimulating action on alveolar type II cells. FN was found to have no effective action on the airway and pulmonary secretory cells.

Effect of pilocarpine on behavior of mucus glycoproteins of canine tracheal secretory cells.

Behavior of mucus glycoproteins in tracheal secretory cells after treatment with pilocarpine was investigated histologically and histochemically using the isolated canine trachea. Following pilocarpine treatment, the number of total and neutral glycoprotein-containing goblet cells was reduced concentration-dependently. The number of acid glycoprotein (AGP)-containing goblet cells was not altered with 10(-7) and 10(-6) M pilocarpine, but significantly decreased with 10(-5) and 10(-4) M pilocarpine. The thickness of the acini of submucosal glands significantly decreased, and the ratio of acinar inner diameter to tracheal wall thickness increased in 10(-5) and 10(-4) M pilocarpine. AGP content in glandular cells increased in 10(-7) and 10(-6) M pilocarpine, but markedly decreased at concentrations of 10(-5) and 10(-4) M. Pilocarpine treatment caused an increase in N-acetylhexosamine concentration in the incubation fluid. Total saccharide concentration in the incubation fluid decreased in 10(-7) and 10(-6) M pilocarpine, but was not apparently altered at concentrations of 10(-5) and 10(-4) M. These findings suggest that lower concentrations of pilocarpine stimulate synthesis of AGP in goblet and glandular cells much more preferentially than it stimulates discharge of AGP from the cells. While at higher concentrations, the AGP-discharge effect overcomes the stimulation in synthesis.

Behavior of mucus glycoproteins of tracheal secretory cells following L-cysteine methyl ester treatment.

The effects of L-cysteine methyl ester on behavior of mucus glycoproteins in canine tracheal secretory cells were investigated histologically and histochemically. Low concentrations (10(-7) and 10(-6)M) of L-cysteine methyl ester produced, as histological changes, decreases in the total goblet cell number and the thickness of the acini of submucosal glands, and an increase in the acinar inner diameter. On the other hand, L-cysteine methyl ester at 10(-5) and 10(-4)M produced an increase in the thickness of the acini and a slight decrease in the acinar inner diameter. Both the total acid glycoprotein and sulphated glycoprotein contents in glandular cells markedly decreased, whereas the neutral glycoprotein content in the cells increased following L-cysteine methyl ester treatment in a concentration-dependent way. Total saccharide and protein concentrations in the incubation fluid increased with 10(-7) and 10(-6)M, but decreased with 10(-5) and 10(-4)M L-cysteine methyl ester. N-Acetylhexosamine concentration in the bathing fluid increased at 10(-7) and 10(-6)M of the agent. These findings suggest that low concentrations of L-cysteine methyl ester stimulate secretory activities of tracheal secretory cells, while at high concentrations the agent stimulates synthesis of mucus glycoproteins in the submucosal glands. Additionally, it is suggested that L-cysteine methyl ester has a marked viscosity lowering effect on the mucus in the secretory cells.