Following hepatic injury or stress, gluconeogenic and acute-phase response genes are rapidly upregulated to restore metabolic homeostasis and limit tissue damage. Regulation of the liver-restricted insulin-like growth factor binding protein 1 (IGFBP-1) gene is dramatically altered by changes in the metabolic state and hepatectomy, and thus it provided an appropriate reporter to assess the transcriptional milieu in the liver during repair and regeneration. The cytokine interleukin-6 (IL-6) is required for liver regeneration and repair, and it transcriptionally upregulates a vast array of genes during liver growth by unknown mechanisms. The liver, which plays an important role in maintaining metabolic and synthetic homeostasis, constitutes a conditional renewal system in which parenchymal cells normally in G 0 may be induced to proliferate following toxic damage, hepatitis, and surgical resection that culminates in the rapid restoration of hepatic parenchyma (35). To maintain glucose balance following the acute loss of liver mass posthepatectomy, phosphoenolpyruvate carboxykinase (PEPCK), glucose-6-phosphatase (G6Pase), and other genes involved in gluconeogenesis are rapidly upregulated in the regenerating liver (14, 37, 54, 55). However, the molecular mechanism by which the liver maintains metabolic homeostasis despite the acute loss of twothirds of hepatic tissue or after injury is not known.Liver regeneration, a hyperplastic response, involves the proliferation of the mature functioning cells composing the intact organ (35,54,55). Of the known cytokines released after liver injury or hepatectomy, interleukin-6 (IL-6) has been shown to be required for normal liver regeneration and repair (8,24). In IL6 Ϫ/Ϫ mice, a highly significant reduction in hepatocyte DNA synthesis, increased liver necrosis, discrete G 1 -phase abnormalities including absence of STAT3 activation, reduction in AP-1 activation, and selective abnormalities in gene expression are observed posthepatectomy and after carbon tetrachloride injury, all of which are corrected by injection with IL-6.Among those genes whose expression is abnormal in IL-6 Ϫ/Ϫ livers after partial hepatectomy are those encoding proteins involved in cell cycle progression such as AP-1 factors, c-Myc, and cyclin D1. However, a number of other genes with less clear connection to cell growth show blunted induction in the absence of IL-6, including the insulin-like growth factor binding protein 1 (IGFBP-1) gene. The mechanism by which IL-6 activates such a vast array of genes is unknown. We chose to study IGFBP-1 because of its proposed role in both hepatic growth and metabolism.IGFBP-1 is an immediate-early gene induced at the transcriptional level in the remnant liver following partial hepatectomy (26,38,50). It is distinct in that its plasma level is dynamically regulated by changes in the metabolic state and after hepatic injury. Of the known upregulators of IGFBP-1 transcription, only IL-6 and phorbol esters have been demonstrated to overcome the strong inhibition of IGFB...

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Transforming growth factor-␤ (TGF-␤) functions as an antiproliferative factor for hepatocytes. However, for unexplained reasons, hepatocytes become resistant to TGF-␤ signals and can proliferate despite the presence of TGF-␤ during liver regeneration. TGF-␤ is up-regulated during liver regeneration, although it is not known whether it is active or latent. TGF-␤ activity may be examined by assessing Smad activation, a downstream signaling pathway. Smad pathway activation during liver regeneration induced by partial hepatectomy or CCl 4 injury was examined by assessing the levels of phospho-Smad2 and Smad2-Smad4 complexes. We found that Smad proteins were slightly activated in quiescent liver, but that their activation was further en-

Barrett's esophagus (BE) is an intestinal metaplasia that occurs in the setting of chronic acid and bile reflux and is associated with a risk for adenocarcinoma. Expression of intestine-specific transcription factors in the esophagus likely contributes to metaplasia development. Our objective was to explore the effects of an intestine-specific transcription factor when expressed in the mouse esophageal epithelium. Transgenic mice were derived in which the transcription factor Cdx2 is expressed in squamous epithelium using the murine Keratin-14 gene promoter. Effects of the transgene upon cell proliferation and differentiation, gene expression, and barrier integrity were explored. K14-Cdx2 mice express the Cdx2 transgene in esophageal squamous tissues. Cdx2 expression was associated with reduced basal epithelial cell proliferation and altered cell morphology. Ultrastructurally two changes were noted. Cdx2 expression was associated with dilated space between the basal cells and diminished cell-cell adhesion caused by reduced Desmocollin-3 mRNA and protein expression. This compromised epithelial barrier function, as the measured trans-epithelial electrical resistance (TEER) of the K14-Cdx2 epithelium was significantly reduced compared to controls (1189 Ohm*cm2 ±343.5 to 508 Ohm*cm2±92.48, p = 0.0532). Secondly, basal cells with features of a transitional cell type, intermediate between keratinocytes and columnar Barrett's epithelial cells, were observed. These cells had reduced keratin bundles and increased endoplasmic reticulum levels, suggesting the adoption of secretory-cell features. Moreover, at the ultrastructural level they resembled “Distinctive” cells associated with multilayered epithelium. Treatment of the K14-Cdx2 mice with 5′-Azacytidine elicited expression of BE-associated genes including Cdx1, Krt18, and Slc26a3/Dra, suggesting the phenotype could be advanced under certain conditions. We conclude that ectopic Cdx2 expression in keratinocytes alters cell proliferation, barrier function, and differentiation. These altered cells represent a transitional cell type between normal squamous and columnar BE cells. The K14-Cdx2 mice represent a useful model to study progression from squamous epithelium to BE.

Acute liver failure caused by viral hepatitis or toxic damage involves both apoptotic and necrotic pathways. IGF binding protein-1 (IGFBP-1), a hepatocyte-derived secreted protein, is required for normal liver regeneration. To determine whether IGFBP-1 could prevent liver injury that entails direct stimulation of hepatocyte apoptosis, IGFBP-1 -/-mice, IGFBP-1 +/+ mice, and mice pretreated with Ab's against IGFBP-1 were treated with a normally sublethal dose of Fas agonist. IGFBP-1 deficiency was associated with massive hepatocyte apoptosis and caspase activation within 3 hours of Fas agonist treatment, which could be corrected by pretreatment with IGFBP-1. IGFBP-1-deficient livers had enhanced signaling via the integrin receptor at early times (0.5 to 1 hour) after Fas agonist treatment accompanied by elevated activated matrix metalloproteinase-9 (MMP-9), a known target of fibronectin signaling and activator of TGF-β. Within 3 hours of Fas agonist treatment, elevated expression of active TGF-β1, a hepatocyte apoptogen, was observed in IGFBP-1-deficient livers that correlated with the appearance of the apoptotic process. Both MMP-9 and TGF-β1 expression were suppressed by IGFBP-1 treatment, supporting their role in the apoptotic process. IGFBP-1 -/-mice also displayed increased injury in a toxic hepatic injury model caused by CCl 4 . These findings indicate that IGFBP-1 functions as a critical hepatic survival factor in the liver by reducing the level of proapoptotic signals. transcription factors (20, 21) that is also required for liver regeneration. C/EBPβ deficiency in the liver confers resistance to Fas-mediated apoptosis in the hepatocytes, as shown by reduced activation of caspase-3 and increased expression of antiapoptotic protein Bcl-x L in Fas-treated C/EBPβ -/-livers (22).We wondered if IGFBP-1 deficiency would result in an apoptotic defect that was similar to that observed with C/EBPβ deficiency. On the contrary, we found that abnormalities in C/EBPβ expression did not play a prominent role in Fas-mediated hepatic injury sustained in IGFBP-1 -/-livers. Instead, after Fas stimulation, IGFBP-1 deficiency was associated with rapid onset of massive hepatocyte apoptosis that could be corrected by pretreatment with IGFBP-1. Fibronectin signaling was elevated early in IGFBP-1-deficient livers and was associated with proteolytic activation of matrix metalloproteinase-9 (MMP-9), enhanced activation of the proapoptotic TGF-β1, caspase-3 and caspase-8 activation, and ultimately breakdown of fibronectin. Treatment of IGFBP-1 -/-livers with IGFBP-1 corrected these abnormalities and the associated morbidity and hepatic defects, establishing IGFBP-1 as a critical hepatic survival factor. MethodsAnimal studies and generation of mutant mice. Generation of IGFBP-1 -/-animals was described previously (2). Studies were performed on IGFBP-1 -/-and IGFBP-1 +/+ mice 12-16 weeks of age in a B6.129 hybrid background. Littermates and backcrossing were used to provide a uniform background. IGFBP-1 -/-phenotype was confirmed by ...