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Decreased pulmonary expression of Forkhead Box f1 (Foxf1) transcription factor was associated with lethal alveolar hemorrhage in 55% of the Foxf1 +/- newborn mice. The severity of the pulmonary abnormalities correlates with the levels of Foxf1 mRNA. Defects in alveolarization and vasculogenesis were observed in subsets of the Foxf1 +/- mice with relatively low levels of expression from the normal Foxf1 allele. Lung hemorrhage was coincident with disruption of the mesenchymal-epithelial cell interfaces in the alveolar and bronchiolar regions of the lung parenchyma and was associated with increased apoptosis and reduced surfactant protein B (SP-B) expression. Finally, the lung defect associated with the Foxf1 +/- mutation was accompanied by reduced expression of vascular endothelial growth factor (VEGF), the VEGF receptor 2 (Flk-1), bone morphogenetic protein 4 (Bmp-4), and the transcription factors of the Brachyury T-Box family (Tbx2-Tbx5) and Lung Kruppel-like Factor. Reduction in the level of Foxf1 caused neonatal pulmonary hemorrhage and abnormalities in alveologenesis, implicating this transcription factor in the regulation of mesenchyme-epithelial interaction critical for lung morphogenesis.

Previous studies have shown that haploinsufficiency of the splanchnic and septum transversum mesoderm Forkhead Box (Fox) f1 transcriptional factor caused defects in lung and gallbladder development and that Foxf1 heterozygous (؉/؊) mice exhibited defective lung repair in response to injury. In this study, we show that Foxf1 is expressed in hepatic stellate cells in developing and adult liver, suggesting that a subset of stellate cells originates from septum transversum mesenchyme during mouse embryonic development. Because liver regeneration requires a transient differentiation of stellate cells into myofibroblasts, which secrete type I collagen into the extracellular matrix, we examined Foxf1 ؉/؊ liver repair following carbon tetrachloride injury, a known model for stellate cell activation. We found that regenerating Foxf1 ؉/؊ liver exhibited defective stellate cell activation following CCl 4 liver injury, which was associated with diminished induction of type I collagen, ␣-smooth muscle actin, and Notch-2 protein and resulted in severe hepatic apoptosis despite normal cellular proliferation rates. Furthermore, regenerating Foxf1 ؉/؊ livers exhibited decreased levels of interferon-inducible protein 10 (IP-10), delayed induction of monocyte chemoattractant protein 1 (MCP-1) levels, and aberrantly elevated expression of transforming growth factor ␤1. T he Forkhead Box (Fox) family of transcription factors shares homology in the winged helix DNA binding domain, 1 and its members play important roles in cellular proliferation, differentiation, and metabolic homeostasis. [2][3][4][5][6] Haploinsufficiency of the Foxf1 gene (previously known as HFH-8 or Freac-1) in heterozygous (ϩ/Ϫ) mice causes perinatal lethality from pulmonary hemorrhage and severe defects in alveolarization, vascularization, and fusion of lung lobes. 7-9 Lung hemorrhage was observed in one half of newborn Foxf1 ϩ/Ϫ mice that had an 80% reduction in pulmonary Foxf1 levels (low Foxf1 ϩ/Ϫ) and reduced expression of genes involved in lung morphogenesis. 7 Interestingly, expression of these lung developmental genes was unchanged in 40% of the newborn Foxf1 ϩ/Ϫ mice that had near wildtype (WT) pulmonary levels of Foxf1 messenger RNA (mRNA) (high Foxf1 ϩ/Ϫ mice) without pulmonary hemorrhage, but they exhibited diminished alveolar septation. 7 Moreover, the high Foxf1 ϩ/Ϫ mice had normal life spans and adult lung morphology, suggesting that these mice compensated for the alveolar septation defect but exhibited defective lung repair in response to injury. 10 Liver development initiates at 9 days postcoitum (dpc) of mouse embryogenesis, when the cardiac mesenchyme induces the hepatic primordium to emerge from the foregut endoderm that invades the septum transversum mesenchyme. 6 Previous expression studies have shown

The forkhead box f1 (Foxf1) transcription factor is expressed in the visceral (splanchnic) mesoderm, which is involved in mesenchymal-epithelial signaling required for development of organs derived from foregut endoderm such as lung, liver, gall bladder, and pancreas. Our previous studies demonstrated that haploinsufficiency of the Foxf1 gene caused pulmonary abnormalities with perinatal lethality from lung hemorrhage in a subset of Foxf1؉/؊ newborn mice. During mouse embryonic development, the liver and biliary primordium emerges from the foregut endoderm, invades the septum transversum mesenchyme, and receives inductive signaling originating from both the septum transversum and cardiac mesenchyme. In this study, we show that Foxf1 is expressed in embryonic septum transversum and gall bladder mesenchyme. Foxf1؉/؊ gall bladders were significantly smaller and had severe structural abnormalities characterized by a deficient external smooth muscle cell layer, reduction in mesenchymal cell number, and in some cases, lack of a discernible biliary epithelial cell layer. This Foxf1؉/؊ phenotype correlates with decreased expression of vascular cell adhesion molecule-1 (VCAM-1), ␣ 5 integrin, plateletderived growth factor receptor ␣ (PDGFR␣) and hepatocyte growth factor (HGF) genes, all of which are critical for cell adhesion, migration, and mesenchymal cell differentiation.At 9-days postcoitum (dpc) 1 during mouse embryogenesis, the liver primordium emerges from the foregut endoderm, invades the septum transversum mesenchyme, and receives bone morphogenetic protein 4 (Bmp-4) and fibroblast growth factor 2 (Fgf2) signaling originating from the septum transversum and cardiac mesenchyme, respectively (1, 2). This hepatic specification is associated with expression of the Foxa2 (HNF-3␤) and Gata4 transcription factors (3). Liver morphogenesis involves a proliferative expansion period and development of the bipotential hepatoblasts that begin to differentiate into hepatocytes and bile duct epithelial cells of the intrahepatic bile ducts (IHBD) at 13.5 dpc (3,4). Interestingly, targeted disruption of either the homeodomain Hex gene or the Hgf gene allows normal development of the mouse hepatic diverticulum, but these cells fail to migrate into the septum transversum and undergo liver morphogenesis (5-7).In the adult liver, bile is synthesized in hepatocytes from cholesterol, secreted into the bile canaliculi and transported through the intrahepatic and extrahepatic biliary system to the gall bladder, where it is stored for secretion into the digestive tract to emulsify lipids (8). The gall bladder and extrahepatic bile ducts (EHBD) develop from the caudal portion of the liver primordium at 10 dpc of mouse embryogenesis (9). However, little is known regarding visceral (splanchnic) mesoderm transcription factors that regulate expression of genes involved in mesenchymal-epithelial induction of gall bladder development from the liver primordium. Visceral mesenchymal expression of the homeodomain Hlx gene is required for the proli...

Most susceptibility to colorectal cancer (CRC) is not accounted for by known risk factors. Because MLH1, MSH2 and MSH6 mutations underlie high-penetrance CRC susceptibility in hereditary nonpolyposis colon cancer (HNPCC), we hypothesized that attenuated alleles might also underlie susceptibility to sporadic CRC. We looked for gene variants associated with HNPCC in Israeli probands with familial CRC unstratified with respect to the microsatellite instability (MSI) phenotype. Association studies identified a new MLH1 variant (415G→C, resulting in the amino acid substitution D132H) in ∼1.3% of Israeli individuals with CRC self-described as Jewish, Christian and Muslim. MLH1 415C confers clinically significant susceptibility to CRC. In contrast to classic HNPCC, CRCs associated with MLH1 415C usually do not have the MSI defect, which is important for clinical mutation screening. Structural and functional analyses showed that the normal ATPase function of MLH1 is attenuated, but not eliminated, by the MLH1 415G→C mutation. The new MLH1 variant confers a high risk of CRC and identifies a previously unrecognized mechanism in microsatellite-stable tumors. These studies suggest that variants of mismatch repair proteins with attenuated function may account for a higher proportion of susceptibility to sporadic microsatellite-stable CRC than previously assumed.To identify variant alleles in MLH1, MSH2 or MSH6 with high sensitivity and specificity, we designed a new high-density oligonucleotide array (HNPCC Chip) that uses improved bioinformatic