DNA Microarray Reveals Novel Genes Induced by Mechanical Forces in Fetal Lung Type II Epithelial Cells

Abstract

Mechanical forces are essential for normal fetal lung development. However, the cellular and molecular mechanisms regulating this process are still poorly defined. In this study, we used oligonucleotide microarrays to investigate gene expression in cultured embryonic d 19 rat fetal lung type II epithelial cells exposed to a level of mechanical strain similar to the developing lung. Significance Analysis of Microarrays (SAM) identified 92 genes differentially expressed by strain. Interestingly, several members of the solute carrier family of amino acid transporter (Slc7a1, Slc7a3, Slc6a9, and tumor-associated protein 1) genes involved in amino acid synthesis (Phgdh, Psat1, Psph, Cars, and Asns), as well as the amiloride-sensitive epithelial sodium channel gene (Scnn1a) were up-regulated by the application of force. These results were confirmed by quantitative real-time PCR (qRT-PCR). Thus, this study identifies genes induced by strain that may be important for amino acid signaling pathways and protein synthesis in fetal type II cells. In addition, these data suggest that mechanical forces may contribute to facilitate lung fluid reabsorption in preparation for birth. Taken together, the present investigation provides further insights into how mechanical forces may modulate fetal lung development. N ormal lung growth and development during fetal life are critical for extrauterine survival. Premature infants are often born before sufficient lung maturation has occurred, and, as a result, they experience a high rate of long-term pulmonary complications such as bronchopulmonary dysplasia. Understanding the mechanisms of prenatal lung growth and development is a crucial step for the design of preventive and therapeutic strategies.Mechanical forces generated in utero by repetitive breathing movements and by fluid distension are essential to mammalian lung development (1-3). Previous in vitro experiments have demonstrated that application of force to cultured type II epithelial cells induces cell proliferation (4) and differentiation (5). Other studies have begun to identify mechanoreceptors and cell signaling pathways mediating fetal lung growth and maturation (4 -8). However, the precise molecular and cellular mechanisms by which lung cells sense mechanical stimuli to influence lung development are still poorly defined.DNA microarray technology provides a powerful tool for rapid, comprehensive, and quantitative analysis of gene expression profile. This technique has been previously used to assess changes in gene expression in endothelial cells (9), bladder smooth muscle cells (10), and human pulmonary A549 (11) and H441 (12) epithelial cells exposed to mechanical strain.Therefore, the goal of the present study was to identify genes differentially expressed by mechanical stress of fetal type II epithelial cells. We used an in vitro model system in which embryonic d (E) 19 type II cells are exposed to mechanical forces similar to that observed in the developing lung. These studies revealed that mechanical force...

Microarray-Based Gene Expression Profiling Suggests adaptation of Lung Epithelial Cells Subjected to Chronic Cyclic Strain

“…Detailed comparisons with another gene microarray study, which revealed a highly altered expression of 18 genes in primary rat AT I-like cells [24], did not identify any similar changes. However, comparisons with the gene microarray study of Wang et al ., which revealed 57 differentially expressed genes in primary fetal AT II cells [11], confirmed similar changes for at least four up-regulated genes (SLC7A3, SLC6A9 and SLC29A3 coding for solute carrier (SLC) group of membrane transport proteins, and TRIB3 (Tribbles homolog 3)) in A549 cells (data not shown).…”

“…In contrast to expression studies investigating individual strain-regulated genes in lung epithelial cells, our and other gene microarray studies investigating a large panel of genes were not performed in a time-dependent manner [11,22,23]. Merely one study analyzed the entire gene expression profile at two time points, but the maximum time was already at 6 hours of cyclic distension [24].…”

“…These values might vary in old or injured lungs because the distension of cells depends on many factors, including the functional residual capacity [8]. Since it is currently not possible to determine exactly the cellular distension in vivo , most research groups studying the effect of mechanical strain on lung cells in vitro perform their experiments with a cell surface elongation between 10 and 20 % to mimic physiological conditions [1,2,9,10,11,12]. Depending on the cell type higher magnitudes result in a pro-inflammatory cell phenotype and cell death [13,14].…”

“…Gene microarray studies have been performed to identify genes, which are up- or down-regulated in alveolar epithelial cells subjected to cyclic mechanical strain in vitro [11,22,23,24]. However, a comparison of these studies revealed only few parallels probably because of differences in the cell type used and strain conditions applied.…”

“…(15 min -1 ) [24]. Also, the time of analysis varied strongly (1 to 24 h), and only one study analyzed long-term effects of cyclic strain on the gene expression [11]. Therefore, our study aimed at the identification of genes, which are differentially expressed in various lung epithelial cells each subjected to long-term cyclic distension at adult frequency conditions.…”