Receptors for advanced glycation end-products (RAGE) are multi-ligand cell surface receptors highly expressed in the lung that modulate pulmonary inflammation during disease. However, the contributions of RAGE signaling are unknown during pulmonary organogenesis. In order to test the hypothesis that RAGE misexpression adversely affects lung morphogenesis, conditional transgenic mice were generated that over-express RAGE in alveolar type II cells of the lung. When RAGE is over-expressed throughout embryogenesis, severe lung hypoplasia ensues, culminating in perinatal lethality. Flow cytometry and immunohistochemistry employing cell-specific markers for various distal cell types demonstrated anomalies in key epithelial cell populations resulting from RAGE up-regulation through embryonic (E) 18.5. Electron microscopy also identified significant morphological disturbances to distal cell types including separation from the basement membrane. Possible mechanisms leading to the disappearance of pulmonary tissue by increased RAGE expression were then evaluated. A time course of lung organogenesis commencing at E12.5 demonstrated that increased RAGE expression primarily alters lung morphogenesis beginning at E16.5. TUNEL immunohistochemistry and immunoblotting for active caspase-3 confirm a shift toward apoptosis in lungs from RAGE over-expressing mice when compared to wild type controls. Assaying for NF-κB also revealed elevated nuclear translocation in lungs from transgenic mice compared to controls. An RT-PCR assessment of genes regulated by NF-κB demonstrated elevated expression of Fas ligand, suggesting increased activity of the Fas-mediated signal transduction pathway in which ligand-receptor interaction triggers cell death. These data provide evidence that RAGE expression must be tightly regulated during organogenesis. Furthermore, additional elucidation of RAGE signaling potentially involved in branching morphogenesis and cell cycle abnormalities may provide insight into the progression of RAGE-mediated lung diseases.

The objective of this study was to examine the influence of age on food intake, tissue and organ mass and NF-ĸB and SIRT6 levels in various tissues. The transcription factor, Nuclear Factor Kappa-B (NF-ĸB), is associated with both catabolic and anabolic pathways of muscle metabolism and may be involved in age-related muscle loss. SIRT6 is a member of the sirtuin family of proteins that function as protein lysine deacetylases and are associated with longevity in a number of organisms. Male Sprague-Dawley rats, aged 6 months (Adult) and 21 months (Old) were fed a commercially available diet for 10-17 days. Old rats consumed less food per body weight (BW) each day than Adult rats (1.45% g diet/g BW vs. 2.4% g diet/g BW). However, when intake data were expressed as g/diet per day there was no significant difference between groups. For skeletal muscle tissue, the average mass of gastrocnemius and soleus (g muscle/g BW) was significantly lower in Old rats. Levels of NF-ĸB (p65/RelA) and SIRT6 were measured by Western blot analysis in gastrocnemius, tibialis anterior, quadriceps, soleus, lung, heart, kidney and liver. NF-ĸB levels were higher in gastrocnemius of Old rats compared to Adult rats. No significant age-specific differences in SIRT6 protein levels were noted in the tissues examined. Interestingly, when examined independent of age, levels of SIRT6 were significantly different between certain tissues. Data from this study suggest that age affects muscle loss and NF-ĸB in a tissue-specific manner. Furthermore, these findings indicate tissue-specific but not age-specific differences in SIRT6 protein levels.