Distribution And Expression Of Pulmonary Ionocyte-related Factors CFTR,ATP6V0D2,and ATP6V1C2 In The Lungs Of Yaks At Different Ages

Bos grunnien,as a kind of bovine species endemic to the plateau,is mainly distributed in the Qinghai-Tibet Plateau and its adjacent alpine and sub-alpine regions in China.It has strong adaptability to the alpine low oxygen ecological environment above 3000 meters,and has formed a series of unique ecological and physiological adaptive characteristics.Pulmonary Ionocyte is a new type of cell found in human and mouse lungs that regulates the physiological environment of the airway surface and plays a key role in the pathogenesis of pulmonary cystic fibrosis.It plays a key role in the damage and tissue repair of many airway diseases,including asthma,chronic obstructive pulmonary disease and bronchitis.Low oxygen environment at high altitude and many lung diseases will lead to pulmonary fibrosis,causing a series of pathological changes of the body.However,yaks can adapt to low oxygen environment at high altitude well,and their lungs will not suffer pathological manifestations such as pulmonary fibrosis caused by low oxygen.The aim of this study was to study the distribution and expression characteristics of pulmonary ionic cell-related factors CFTR,ATP6V0D2 and ATP6V1C2 in four age groups of yak lungs,and to explore the possible distribution locations of pulmonary ionic cells in different ages of yak lungs and the regulatory role of these related factors in the structure formation of yak lungs in adapting to the hypoxia environment at high altitude.The lung tissue of healthy yak in four age groups of primary(1-6 days of age),juvenile(1-2 years of age),adult(3-6 years of age)and old(7-10 years of age)was taken as the research object.Firstly,the expressions of CFTR,ATP6V0D2 and ATP6V1C2 in lung of yaks of different age groups were detected by qRT-PCR and Western-blot.Then,the exact distribution of CFTR,ATP6V0D2 and ATP6V1C2 in the lungs of yak at different age groups was studied by immunohistochemical technique,and the comparison was made in the lungs of adult cattle and mice.The results showed that:1.The qRT-PCR results showed that CFTR,ATP6V0D2 and ATP6V1C2 m RNA were expressed in different age groups in yak lung.The expression level of CFTR was the highest in the adult group,followed by the elderly group,and the lowest in the young group,with significant differences among different age groups(P < 0.05).The expression trend of ATP6V1C2 was the same as that of CFTR.The expression level of ATP6V0D2 in primary group was the highest,and the difference was significant(P < 0.05);Compared with the other three groups,the difference was significant(P < 0.05);There was no significant difference in expression levels between the adult group and the elderly group(P > 0.05).2.Western-blot results showed that the expression level of CFTR protein in the primary group was significantly higher than that in the other three groups(P < 0.05),there was no significant difference among other groups(P > 0.05).The expression level of ATP6V0D2 in adult group was significantly higher than that in the other three groups(P < 0.05),and there were significant differences among other groups(P < 0.05).ATP6V1C2 was the highest expression in the juvenile group(P < 0.05),followed by the adult group,and the newborn group had the lowest expression level(P < 0.05).3.Immunohistochemical results showed that CFTR was mainly distributed in ciliated cells and Clara cells in the epithelial mucosa layer of bronchioles and branches of yak,smooth muscle cells and glands in the wall of the vessel,and smooth muscle cells in the middle membrane of the accompanying pulmonary artery.It was strongly positive in neonatal group and positive in other groups.ATP6V0D2 was mainly distributed in ciliated cells and Clara cells of the bronchiole and its branches.It was strongly positive in the adult group and the elderly group.ATP6V1C2 was mainly distributed in ciliary cells and Clara cells,smooth muscle cells and fibroblasts in the mucosal layer of the bronchus and its branches in the lung,accompanied by fibroblasts in the pulmonary artery wall.The expression of ATP6V1C2 was strongly positive in primary group.By comparison,CFTR,ATP6V0D2 and ATP6V1C2 were also distributed in the ciliated cells and Clara cells of the bronchiole and its branches in adult cattle and mice,but not in the airway wall or the accompanying pulmonary artery wall.In conclusion,yak ciliary cells and Clara cells share common characteristics with mouse and human pulmonary ionocytes,but they are widely distributed and abundant.These cells may have similar functions to pulmonary ionocytes and can play a similar role in preventing pulmonary fibrosis as pulmonary ionocytes.CFTR,ATP6V0D2 and ATP6V1C2 were all distributed in lung tissues of yaks of different ages.Except that ATP6V0D2 was only distributed in ciliated cells and Clara cells in the epithelial mucosa layer of bronchioles and its branches,and was strongly expressed in the adult group and the elderly group.CFTR and ATP6V1C2 were distributed in large amount in ciliary cells and Clara cells of epithelial mucosa,smooth muscle cells of duct wall,smooth muscle cells of accompanying pulmonary artery wall and fibroblasts of each age group,and all of them were strongly expressed in primary group.CFTR,ATP6V0D2 and ATP6V1C2 were strongly expressed in neonatal and adult lung,and their protein expression reached the highest level.These results are directly related to the formation of adaptive structure of yak lung to hypoxic environment,and may play an important physiological function.However,the exact relationship between them and lung ion cells needs further study.