Cerebral aneurysm formation and rupture and TNF-α expression. Representative gross images of normal cerebral blood vessels in (A) sham operated mice, (B) unruptured aneurysms, and (C) ruptured aneurysms. H & E staining of normal cerebral blood vessels in sham operated mice demonstrates two to three layers of smooth muscle cells and a single, thin, continuous layer of endothelial cells (A2 to 3). In both unruptured cerebral aneurysms (B2 to 3) and ruptured aneurysms (C2 to 3) there are thin and thick areas of vascular wall. In unruptured aneurysms in the areas of thin vascular wall, there is intact endothelial and smooth muscle cell layers, but this is discontinuous in the areas of thick vascular wall. In ruptured aneurysms these areas are sometimes discontinuous in both thick and thin areas of the vascular walls. Trichrome staining of normal cerebral blood vessels in sham operated mice (A4 to 5) demonstrates one layer of elastic lamina. This is disorganized in unruptured aneurysms (B4 to 5) and ruptured aneurysms (C2 to 3). TNF-α (green) expression is nearly non-existent in sham operated mice (A6 to 7). The expression of TNF-α (green) is decreased in sham mice (A6 to 7) as compared to unruptured (B6 to 7) and furthermore ruptured aneurysms (C6 to 7). SMC-22α (red) is observed in sham operated mice (A6) and co-localizes with TNF-α (green) to appear yellow in unruptured (B6) and ruptured aneurysms (B7). Normal sized cerebral blood vessels also stain only red for SM-22α in vascular segments adjacent to cerebral aneurysms (B6). Macrophages are nearly non-existent in sham operated mice (A7). Macrophages (red) co-localize with TNF-α (green) and appear yellow in unruptured aneurysms (B7) and more so in ruptured aneurysms (C7). Nuclei are stained blue with DAPI. DAPI, 4',6-diamidino-2-phenylindole; H & E, hematoxylin and eosin; SM-22α, smooth muscle cell 22 alpha.