Fig. 4. Ultrastructural and biochemical analysis of lungs from Klf5^/ mice. (A-D) Lungs from control (A,C) and Klf5^/ (B,D) mice (approximately E18) were fixed and processed for electron microscopy as described in the Materials and methods. (B) Immature Type 2 cells (T2) with no lamellar bodies and abundant glycogen particles (g), distributed diffusely throughout the cytoplasm, were observed in the lungs of the Klf5^/ mice. Lipofibroblasts (LF) were found ensheathing the undilated acinar tubules and buds in the periphery of the lung. (A) By comparison, T2 cells (T2) from control mice were more mature, exhibiting multiple lamellar bodies (arrows), smaller patches of glycogen (g), and increased amounts of rough endoplasmic reticulum. (D) In the Klf5^/ mice, immature T2 cells (T2) with no lamellar bodies were observed lining the more dilated acinar tubules found in the central regions of the Klf5^/ lungs. Note the prominent lipofibroblasts (LF) directly adjacent to these epithelial cells. (C) By contrast, differentiating T1 cells (T1) with thin cytoplasmic extensions (arrows) and centrally located glycogen patches (g) were found in the dilated alveolar saccules of control mice. Pulmonary capillaries filled with red blood cells (^*) were found in close apposition to squamous type I cells (arrows). Fibroblasts (F) without lipid droplets were observed in the interior of the alveolar septa. (E) Whole lung homogenates were prepared at E18.5 and 100 µg protein used for western blot using antiserum against the mature SP-B peptide. Mature SP-B was decreased in the lungs of Klf5^/ mice at E18.5, as assessed in four individual animals. Scale bars: 4 µm.