The Influence Of Ammonia Stimulation On Airway Mucus Secretion In Piglets

Airway mucus traps inhaled harmful gases,dust,pathogens and transports them out of the lungs by ciliary beating to maintain the health of airway.Mucus hypersecretion is a feature of many airway diseases.The high concentration of ammonia in the pig house in winter not only affects the growth performance of pigs,but also induces airway diseases.This study first collected the trachea and the upper right lung of the healthy pigs,and then observed the distribution of the main epithelial cell types(Goblet cells,Basal cells and Clara cells)and mucus in the trachea and bronchus by histological staining.Then,ammonia stimulation experiment was carried out,twelve 7-week-old piglets were randomly divided into four groups(NC group,D4 group,D8 group,D12 group),with three piglets in each group.The group NC was used as the control,and the experimental groups were exposed in a chamber with an ammonia concentration of 56 mg/m3 for 4 days(D4 group),8 days(D8 group)and 12 days(D12 group),respectively(8 hours for each day;from 8:00 am to 16:00 pm).At the end of the stimulation,the right upper lobes of the piglets were collected to make paraffin sections,the effects of ammonia stimulation on the secretion of airway mucus of piglets were observed.RNA was extracted from the lower lobe of the left lung,and the expression of genes related to inflammation and mucus were detected.The results are as follows:1.Distribution of main cells in the respiratory epithelium of pigsK14 is marker of Basal cells,and SCGB1A1 is marker of Clara cells.Immunohistochemistry showed that the differentiation potential of Basal cells and Clara cells were distributed in the respiratory epithelium of the pigs,and Basal cells and Clara cells were abundant in tracheal epithelium.MUC5 AC is marker of Goblet cells.Immunohistochemistry of MUC5 AC showed that Goblet cells were widely distributed in the respiratory epithelium of pigs.2.Effects of ammonia stimulation on airway mucus secretion in pigletsAB-PAS staining showed that with the respiratory tract diameter decrease,the content of mucus gradually reduced to disappear.AB-PAS staining of respiratory tract with same diameter in piglets,showed that the content of mucus in the airway epithelium increased with the extension of the ammonia stimulation time,and was significantly higher in D12 group than that in NC group(P<0.05).Immunohistochemistry showed that the content of MUC5 AC in the airway epithelium was also increased after ammonia stimulation.3.Changes of main epithelial cells and related factors in airway after ammonia stimulation(1)K5,K14,TP63 are markers of Basal cells,and immunofluorescence showed that the content of K14 in the airway epithelium of piglets was increased after ammonia stimulation.The results of Q-PCR showed that the expression of K5,K14 and TP63 in the lung tissue of the piglets was increased after ammonia stimulation.(2)SCGB1A1 is marker of Clara cells.Immunohistochemistry of SCGB1A1 showed that there was a small amount of Clara cells in the airway epithelium of piglets before and after ammonia stimulation.The results of Q-PCR showed that the expression of SCGB1A1 was significantly lower in D12 group than that of the NC group(P<0.05).(3)ABCA3 is marker of type Ⅱ alveolar cells.Q-PCR results showed that the expression of ABCA3 in lung tissue was significantly higher after ammonia stimulation(P<0.05).The S100A9,AQP5 and EGFR genes are genes associated with airway mucus secretion,and the Q-PCR results showed that the expression of S100A9 in D12 group was significantly higher than that in NC group(P<0.05).Compared with the NC group,the expression of AQP5 in lung tissue decreased in D4 group,and the expression level of EGFR increased in D8 group.Conclusion: Goblet cells,Basal cells and Clara cells are widely distributed in the proximal airway of pigs.With the decrease of the diameter of the respiratory tract,secreted mucus gradually reduced to disappear.Ammonia stimulation resulted in the mucus secretion and the number of Goblet cells,Basal cells and type Ⅱ alveolar cells in piglet respiratory epithelium increased,but the number of Clara cells decreased.The expression of genes related to inflammation and mucus secretion also changed.