NASA has reported pulmonary abnormalities in astronauts on space missions, but the molecular changes in lung tissue remain unknown. The goal of the present study was to explore the effects of spaceflight on expression of extracellular matrix (ECM), cell adhesion and pro-fibrotic molecules in lungs of mice flown on Space Shuttle Endeavour (STS-118). C57BL/6Ntac mice housed in animal enclosure modules during a 13-day mission in space (FLT) were euthanized within hours after return; ground controls were treated similarly for comparison (GRD). Analysis of genes associated with ECM and adhesion molecules was performed according to quantitative RT-PCR. The data revealed that FLT lung samples had statistically significant transcriptional changes, i.e., at least 1.5-fold, in 25 out of 84 examined genes (P < 0.05); 15 genes were up-regulated and 10 were down-regulated. The genes that were up-regulated by more than 2-fold were Ctgf, Mmp2, Ncam1, Sparc, Spock1, and Timp3, whereas the most down-regulated genes were Lama1, Mmp3, Mmp7, vcam-1, and Sele. Histology showed profibrosis-like changes occurred in FLT mice, more abundant collagen accumulation around blood vessels, and thicker walls compared with lung samples form GRD mice. Immunohistochemistry was used to compare expression of six selected proteins associated with fibrosis. Immunoreactivity of four proteins (MMP-2, CTGF, TGF-beta1, and NCAM) was enhanced by spaceflight, whereas, no difference was detected in expression of MMP-7 and MMP-9 proteins between the FLT and GRD groups. Taken together, the data demonstrate that significant changes can be readily detected shortly after return from spaceflight in the expression of factors that can adversely influence lung function.