Hepatocarcinoma-Intestine-Pancreas/Pancreatic Associated Protein (HIP/PAP) Is Expressed and Secreted by Proliferating Ductules as well as by Hepatocarcinoma and Cholangiocarcinoma Cells

Hepatocarcinoma-intestine-pancreas/pancreatic associated protein (HIP/PAP) gene was identified because of its increased expression in 25% of human hepatocellular carcinoma. HIP/PAP protein, a C-type lectin, binds laminin, acts as an adhesion molecule for hepatocytes, and has also been described as an acute phase secretory protein during acute pancreatitis in humans and rats. We investigated HIP/PAP protein expression in patients with various liver diseases associated with ductular reaction. At the same time, we analyzed patients with hepatocellular carcinoma and cholangiocarcinoma, and tested HIP/PAP protein levels in sera to establish the pattern of secretion. Our data show that HIP/PAP expression was not restricted to hepatocellular carcinoma, but was also detected in cholangiocarcinoma cells as well as in reactive non-malignant bile ductules. In contrast, HIP/PAP protein expression was undetectable in normal mature hepatocytes, but some ductular cells localized at the interface of portal tracts with parenchyma were HIP/PAP immunoreactive in normal liver. Finally, we present evidence that HIP/PAP serum levels were increased in 21/28 (75%) patients with hepatocellular carcinoma, and in 25/51 (49%) patients with nonmalignant cirrhosis. Altogether, these results suggest that HIP/PAP protein may be implicated in hepatocytic and cholangiolar differentiation and proliferation.     

HIP/PAP, a member of the reg family, is expressed in glucagon-producing enteropancreatic endocrine cells and tumors

Hepatocarcinoma-intestine-pancreas/pancreatitis-associated protein (HIP/PAP) protein, a member of the reg family, is constitutively expressed by some specialized epithelial cell subsets in the digestive tract and the pancreas. We performed a detailed analysis of the expression of HIP/PAP protein in normal digestive endocrine cells according to their localization, lineage, and differentiation stage, and in digestive endocrine tumors according to their site of origin and hormonal profile. In both adult and fetal normal tissues, HIP/PAP expression was detected only in endocrine cells of the small intestine, ascending colon, and pancreas. Two different expression patterns were identified: (a) a strong cytoplasmic labeling observed in the endocrine cells of the digestive mucosa and the outer rim of Langerhans islets specialized in the synthesis of glucagon and glucagon-like peptides; (b) a weak cytoplasmic immunoreactivity observed in the other pancreatic endocrine cell populations. HIP/PAP expression was detected in 36 of the 184 cases of digestive endocrine tumors examined; 32 of these cases (89%) were pancreatic. The 2 patterns observed in the normal state were retained: (a) a strong labeling was observed in 5% to 100% of tumor cells in 26 tumors, all expressing glucagon or glucagon-like peptides; (b) a weak labeling was present in 10 tumors, presenting various hormonal profiles. In conclusion, a strong expression of HIP/PAP is characteristic of glucagon-producing normal and neoplastic enteropancreatic endocrine cells. Our results lend further support to the concept that members of the reg family play regulatory roles in various endocrine cell populations and that their expression in endocrine cells is lineage-specific.     

Expression of hypoxia-inducible, membrane-bound carbonic anhydrase isozyme XII in mouse tissues

Carbonic anhydrase (CA) XII is a membrane-associated enzyme that has been demonstrated to be normally expressed in some human tissues, to be upregulated in some cancers, and to be a hypoxia-inducible gene product. In mouse, CA XII has been recently localized in the kidney. In the present study, we investigated CA XII gene and protein expression in other mouse tissues, with the kidney serving as a positive control for the reagents. The expression of CA XII mRNA was examined using polymerase chain reaction (PCR) amplification of commercial cDNAs produced from selected mouse tissues. A strong positive signal for CA XII mRNA was detected in the kidney, and weak signals were obtained in the testis and lung. Heart, spleen, liver, and skeletal muscle were negative. Immunohistochemical staining was performed using a mouse CA XII-specific antibody and biotin-streptavidin complex method. The results showed high expression of CA XII in the kidney, as expected. It was also highly expressed in the surface epithelial cells of the colon, whereas it was absent in the stomach, proximal small intestine, pancreas, liver, heart, and skeletal muscle. The maturing sperm cells showed a weak staining in a pattern that most probably indicates expression in the developing acrosomal membrane. The high expression in the kidney and colon suggests a role for CA XII in the maintenance of body ion and pH homeostasis in the mouse. However, the present findings demonstrated that CA XII has a very limited distribution in mouse tissues outside these two organs.     

Analysis of KRAP expression and localization,and genes regulated by KRAP in a human colon cancer cell line

We previously identified the human KRAP (Ki-ras-induced actin-interacting protein) gene from the cDNA library of human colon cancer HCT116 cells as one of the genes whose expression levels were up-regulated by activated Ki-ras. Although the KRAP gene is structurally conserved from fish to mammalian species, the expression pattern and function of KRAP still remain to be elucidated. Here, we have generated a specific polyclonal antibody for KRAP and characterized the histological expression of KRAP in mouse tissues. KRAP was ubiquitously expressed in mouse tissues, with high levels in pancreas, liver, and brown adipose tissues, and KRAP was co-localized with filamentous actin along the apical membranes in both pancreas and liver tissues. A subfractionation study revealed that KRAP is a cytoplasmic protein and that the majority is associated with the cytoskeleton. Furthermore, microarray gene expression profile by inhibiting KRAP expression in HCT116 cells showed that several receptors and signal molecules frequently deregulated in cancers were differentially expressed in the KRAP-knockdown cells. All of these results suggested that KRAP might be a cytoskeleton-associated protein involving the structural integrity and/or signal transductions in human cancers. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Timing and expression pattern of carbonic anhydrase II in pancreas.

In the search for genetic markers for assessing the role of duct cells in pancreas growth, we examined whether carbonic anhydrase II (CAII) has ductal cell specificity. We determined the distribution and timing of CAII expression in mouse pancreas from embryonic stage to adult. The pancreatic ducts only start expressing CAII at embryonic day (E) 18.5, with increases after birth. Around E15.5, glucagon-positive cells, but not insulin-positive cells, also express CAII, with further increases by adult. CAII expression was restricted to cells within ductal structures and glucagon-positive cells with no colocalization with any insulin-positive cells at any time. In the human pancreas, CAII expression is restricted to the ducts. Furthermore, the activity of a 1.6-kb fragment of the human promoter with Luciferase assays was moderately strong compared with the cytomegalovirus promoter in human pancreatic duct cell line (PANC-1). Thus, we believe that the CAII gene could serve as a useful pancreatic duct cell marker.     

Tissue-specific expression, developmental regulation, and genetic mapping of the gene encoding CCAAT/enhancer binding protein

This paper presents the results of experiments that determine the chromosomal location of the mouse gene encoding CCAAT/enhancer binding protein (C/EBP) and measure its expression as a function of tissue type and temporal period of development in mice and rats. Three alleles of the C/EBP gene were identified according to restriction fragment length polymorphisms. The strain distribution pattern of the three alleles was determined in recombinant inbred mouse strains and compared to that of other mouse genes. These results mapped the gene to a position within 2.5 centimorgans (cM) of the structural gene encoding glucose phosphate isomerase on chromosome 7 of the mouse. The expression pattern of the C/EBP gene was studied by a combination of nucleic acid hybridization and antibody staining assays. High levels of C/EBP mRNA were observed in tissues known to metabolize lipid and cholesterol-related compounds at uncommonly high rates. These included liver, fat, intestine, lung, adrenal gland, and placenta. More detailed analysis of two of these tissues, liver and fat, showed that C/EBP expression was limited to fully differentiated cells. Moreover, analysis of the temporal pattern of expression of C/EBP mRNA in two tissues, liver and intestine, revealed a coordinated induction just prior to birth. These observations raise the possibility that the synthesis of C/EBP may be responsive to humoral factors and that modulation in C/EBP expression might mediate coordinated changes in gene expression that facilitate adaptive challenges met during development or during the fluctuating physiological states of adult life.     

Fetal antigen 1 (FA1) in the human pancreas: cell type expression, topological and quantitative variations during development

Monospecific rabbit anti-human fetal antigen 1 (FA1), was used to examine the distribution of FA1 during the development of the human fetal pancreas and liver using an indirect immunoperoxidase technique. FA1 was expressed by 94% of the glandular epithelial cells of the branching ducts in the pancreatic anlage at week 7 of gestation. This pattern changed during the development of the human pancreas, 64% of the glandular cells being FA1 positive at week 17 of gestation, decreasing to 11% in the infant (4 months after birth). In the infant and adults the FA1 expression was restricted to a subpopulation of -cells within the islets of Langerhans. Insulin immunoreactive cells were scattered throughout the epithelium of primitive branching pancreatic ducts at week 7 of gestation, well before the formation of islets. From the 7th through to the 17th week of gestation, FA1 was found in the cytoplasm of fetal hepatocytes, whereas no staining was observed in the liver from a 4-month-old infant. No FA1 expression was found in the epithelium of the developing gut. The present findings indicate that the glandular epithelial cells in the developing pancreas may serve as stem cells, which, if appropriately induced, may differentiate into endocrine cells. Fetal antigen 1 (FA1) may take part in or be a result of this differentiation.