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Isolation and characterisation of alveolar type II pneumocytes from adult bovine lung.

ABSTRACT: Alveolar type II (ATII) cells play a key role as part of the distal lung epithelium, including roles in the innate immune response and as self-renewing progenitors to replace alveolar type I (ATI) cells during regeneration of the alveolar epithelium. Their secretion of surfactant protein helps to maintain homeostasis in the distal lung and exert protective, antimicrobial properties. Despite the cell's crucial roles, they remain difficult to study, in part due to inefficient and expensive isolation methods, a propensity to differentiate into alveolar type I cells in culture and susceptibility to fibroblast overgrowth from primary isolations. Published methods of isolation often require specialist technology, negatively impacting the development of in vitro models of disease, including bovine tuberculosis (BTB), a serious re-emerging disease in both animals and humans worldwide. We present here a simple and cost-effective method that may be utilised in the generation of bovine primary ATII cells. These exhibit an ATII phenotype in 2D and 3D culture in our studies and are conducive to further study of the role of ATII cells in bovine respiratory diseases.

SUBMITTER: Lee DF

PROVIDER: S-EPMC6085293 | biostudies-literature | 2018 Aug

REPOSITORIES: biostudies-literature

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Isolation and characterisation of alveolar type II pneumocytes from adult bovine lung.

Lee Diane Frances DF Salguero Francisco Javier FJ Grainger Duncan D Francis Robert James RJ MacLellan-Gibson Kirsty K Chambers Mark Andrew MA

Scientific reports 20180809 1

Alveolar type II (ATII) cells play a key role as part of the distal lung epithelium, including roles in the innate immune response and as self-renewing progenitors to replace alveolar type I (ATI) cells during regeneration of the alveolar epithelium. Their secretion of surfactant protein helps to maintain homeostasis in the distal lung and exert protective, antimicrobial properties. Despite the cell's crucial roles, they remain difficult to study, in part due to inefficient and expensive isolati ...[more]

PMID: 30093682

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Project description:The regulation by cyclic AMP and fatty acids of phosphatidylcholine (PC) synthesis in rat alveolar type II cells was studied. In contrast with results with hepatocytes, cyclic AMP and its potent chlorophenylthio analogue had no inhibitory effect on [Me-14C]choline incorporation into PC in pulse-chase studies with alveolar type II cells. The inclusion of the fatty acids palmitate, oleate or linoleate in the chase incubation medium stimulated the incorporation of [Me-14C]choline into PC by type II cells. The effect of palmitate, which was more pronounced than that of the other fatty acids, appeared to be concentration-dependent. Increased [Me-14C]choline incorporation into PC was paralleled by an accelerated disappearance of the radiolabel from choline phosphate, which is consistent with an activation of CTP:choline-phosphate cytidylyltransferase. This enzyme is considered to be rate-limiting in the synthesis of PC de novo by type II cells. As fatty acids are also substrate for PC synthesis, their effect could also be due to compensation for a fatty acid deficiency. To test this possibility, fatty acid synthesis in the type II cells was stimulated by addition of lactate. Even then, an additional stimulation of PC synthesis by palmitate was observed, which supports the regulatory influence of exogenous fatty acids. Incubation of type II cells in the presence of 0.2 mM-palmitate resulted in a 45% increase in the membrane-bound CTP:choline-phosphate cytidylyltransferase activity, whereas the soluble activity remained unchanged. Choline kinase activity in the soluble fraction increased by 48%. However, the increase in choline kinase is unlikely to be responsible for the increased metabolic flux through the choline phosphate pathway, because there is a relatively large pool of choline phosphate in type II cells. Therefore it is suggested that the microsomal CTP:choline-phosphate cytidylyltransferase is the form of this enzyme which is active in surfactant PC synthesis, and possibly has a regulatory role in this pathway.

Dataset Information

Isolation and characterisation of alveolar type II pneumocytes from adult bovine lung.

Publications

Isolation and characterisation of alveolar type II pneumocytes from adult bovine lung.

Similar Datasets

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