Pulmonary Alveolar Type II Cells Isolated from Rats

Dobbs, Leland G.; Mason, Robert J.

doi:10.1172/jci109313

Cited by 329 publications

(44 citation statements)

References 46 publications

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“…Rat alveolar type II cells were isolated and cultured as described (15,27). A type I-like cell phenotype was generated by culturing type II cells on plastic dishes in DMEM plus 10% FBS without KGF (16).…”

Section: Methodsmentioning

confidence: 99%

Identification and characterization of a lysophosphatidylcholine acyltransferase in alveolar type II cells

Chen

Hyatt

Mucenski

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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187

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Pulmonary surfactant is a complex of lipids and proteins produced and secreted by alveolar type II cells that provides the low surface tension at the air-liquid interface. The phospholipid most responsible for providing the low surface tension in the lung is dipalmitoylphosphatidylcholine. Dipalmitoylphosphatidylcholine is synthesized in large part by phosphatidylcholine (PC) remodeling, and a lysophosphatidylcholine (lysoPC) acyltransferase is thought to play a critical role in its synthesis. However, this acyltransferase has not yet been identified. We have cloned full-length rat and mouse cDNAs coding for a lysoPC acyltransferase (LPCAT). LPCAT encodes a 535-aa protein of Ϸ59 kDa that contains a transmembrane domain and a putative acyltransferase domain. When transfected into COS-7 cells and HEK293 cells, LPCAT significantly increased lysoPC acyltransferase activity. LPCAT preferred lysoPC as a substrate over lysoPA, lysoPI, lysoPS, lysoPE, or lysoPG and prefers palmitoyl-CoA to oleoyl-CoA as the acyl donor. This LPCAT was preferentially expressed in the lung, specifically within alveolar type II cells. Expression in the fetal lung and in rat type II cells correlated with the expression of the surfactant proteins. LPCAT expression in fetal lung explants was sensitive to dexamethasone and FGFs. KGF was a potent stimulator of LPCAT expression in cultured adult type II cells. We hypothesize that LPCAT plays a critical role in regulating surfactant phospholipid biosynthesis and suggest that understanding the regulation of LPCAT will offer important insight into surfactant phospholipid biosynthesis.biochemistry ͉ lung ͉ surfactant ͉ phosphatidylcholine

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Section: Methodsmentioning

confidence: 99%

Identification and characterization of a lysophosphatidylcholine acyltransferase in alveolar type II cells

Chen

Hyatt

Mucenski

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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175

187

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“…Conversely, low temperatures decrease metabolic rate, and may therefore lower the rate of surfactant secretion. In ATII cells isolated from homeothermic rats, a decrease in incubation temperature to 5°C virtually abolishes surfactant secretion (Dobbs and Mason, 1979). Basal secretion is also significantly lower in type II cells isolated from warm-active dunnarts when incubated at 15°C compared to 37°C (Ormond et al, 2001).…”

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confidence: 96%

“…In direct contrast, type II cells isolated from homeothermic mammals such as humans and rats do not respond to cholinergic agonists (Dobbs and Mason, 1979). This suggests that heterothermic mammals may have a direct role for the parasympathetic nervous system in regulating surfactant secretion at low body temperatures.…”

mentioning

confidence: 99%

Thermal acclimation of surfactant secretion and its regulation by adrenergic and cholinergic agonists in type II cells isolated from warm-active and torpid golden-mantled ground squirrels,Spermophilus lateralis

Ormond

Orgeig

Daniels

et al. 2003

Journal of Experimental Biology

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SUMMARYHomeothermic mammals experience pulmonary surfactant dysfunction with relatively small fluctuations in body temperature. However, ground squirrels survive dramatic changes in body temperature during hibernation, when body temperature drops from 37°C to 0-5°C during prolonged torpor bouts. Using type II cells isolated from both warm-active and torpid squirrels, we determined the effect of assay temperature, autonomic agonists and torpor on surfactant secretion. Basal secretion was significantly higher in type II cells isolated from torpid squirrels compared with warm-active squirrels when assayed at the body temperature of the animal from which they were isolated(4°C and 37°C, respectively). A change in assay temperature significantly decreased surfactant secretion. However, the change in secretory rate between 37°C and 4°C was less than expected if due to temperature alone (Q10 range=0.8-1.2). Therefore, the surfactant secretory pathway in squirrel type II cells demonstrates some temperature insensitivity. When incubated at the body temperature of the animal from which the cells were isolated, the adrenergic agonist, isoproterenol, significantly increased surfactant secretion in both warm-active and torpid squirrel type II cells. However, the cholinergic agonist, carbamylcholine chloride, only increased secretion in torpid squirrel type II cells when incubated at 4°C. Torpor did not affect basal cAMP production from isolated type II cells. However, the production of cAMP appears to be upregulated in response to isoproterenol in torpid squirrel type II cells. Thus, at the cellular level, both the secretory and regulatory pathways involved in surfactant secretion are thermally insensitive. Upregulating basal secretion and increasing the sensitivity of type II cells to cholinergic stimulation may be adaptative characteristics of torpor that enable type II cells to function effectively at 0-5°C.

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“…Primary Culture of Alveolar Type II Cells and Inhibition of Phosphatidylcholine Secretion-Alveolar type II cells were isolated by tissue dissociation with elastase and purification on metrizamide gradients (26). The type II cells were seeded at 2 ϫ 10 6 /35-mm dish in 2 ml of the Dulbecco's modified Eagle's medium (DMEM) containing 10% fetal calf serum.…”

Section: Methodsmentioning

confidence: 99%

Mutational Analysis of Arg197 of Rat Surfactant Protein A

Pattanajitvilai¹,

Kuroki²,

Tsunezawa³

et al. 1998

Journal of Biological Chemistry

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In previous studies, tandem mutagenesis Pulmonary surfactant protein A (SP-A)1 is a hydrophilic C-type mammalian lectin with a subunit molecular mass of 26 -38 kDa that is synthesized primarily by alveolar type II and Clara cells (1, 2). SP-A is composed of four distinct domains (1): 1) an amino-terminal domain that forms interchain disulfide bonds; 2) a collagen-like domain containing multiple Gly-X-Y repeats; 3) a neck region composed of an amphipathic helix and a hydrophobic domain; and 4) a carbohydrate recognition domain (CRD). The mature protein forms an oligomeric bouquet structure composed of six trimeric subunits which are covalently joined via disulfide bonds (1). SP-A binds and aggregates phospholipid vesicles at physiological concentrations of Ca 2ϩ (3, 4). The protein also facilitates uptake of phospholipid by isolated alveolar type II cells (5). These in vitro properties are thought to reflect some of the functions of SP-A in vivo that include structural reorganization and recycling of surfactant lipids. Other important Ca 2ϩ -dependent functions of SP-A include high affinity binding to specific receptors on alveolar type II cells and negative regulation of surfactant secretion (6 -9). SP-A also plays a role in immunoglobulin independent host defense in the lung (10). The protein stimulates macrophage chemotaxis and can enhance phagocytosis of Herpes simplex virus type 1 (11), Staphylococcus aureus (12), and Mycobacterium tuberculosis (13). SP-A also enhances FcR-and CR1-mediated phagocytosis (14). Several proteins that interact with SP-A have been identified on alveolar type II cells and macrophages (15)(16)(17).Mutations in the CRD affect specific type II cell receptor binding, phospholipid binding and aggregation, and inhibition of surfactant secretion as well as augmentation of phospholipid uptake by alveolar type II cells (18,19). The simultaneous substitutions E195Q and R197D of the CRD alter carbohydrate binding specificity, prevent SP-A-mediated phospholipid uptake and aggregation of phospholipid, and destroy the ability of SP-A to compete with its radiolabeled counterpart for surface receptors on alveolar type II cells. These results implicate the Glu 195 and Arg 197 as essential amino acids in SP-A function. This previous conclusion for SP-A is consistent with results first obtained with the closely related MBP-A that defined the active site for carbohydrate binding (20,21). The work with MBP-A followed solution of the crystal structure and unambiguously defined the carbohydrate and Ca 2ϩ -binding sites of the protein. Work with SP-A using site-directed mutagenesis has largely followed from extrapolation of the MBP-A data.In this study, we examined the specific role of amino acid Arg 197 of the SP-A in the interaction of the protein with the type II cell receptor and lipids. Position 197 was replaced with amino acids Ala, Lys, His, Asp, and Asn. These mutants were prepared using the baculovirus expression system in invertebrate (Sf9) cells. Recombinant rat SP-A has been shown to have ...

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Pulmonary Alveolar Type II Cells Isolated from Rats

Abstract: A B S T R A C T

Cited by 329 publications

References 46 publications

Identification and characterization of a lysophosphatidylcholine acyltransferase in alveolar type II cells

Identification and characterization of a lysophosphatidylcholine acyltransferase in alveolar type II cells

Thermal acclimation of surfactant secretion and its regulation by adrenergic and cholinergic agonists in type II cells isolated from warm-active and torpid golden-mantled ground squirrels,Spermophilus lateralis

Mutational Analysis of Arg197 of Rat Surfactant Protein A

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