Phenotypic characterization of rat hepatoma cell lines and lineage-specific regulation of gene expression by differentiation agents.

Hepatoma cell lines can be characterized by their expression of hepatocyte- and biliary-specific genes and by their response to differentiating agents in a lineage-dependent manner. These characteristics can be used to map the maturational lineage position of the cell lines. Tissue-specific gene expression and regulation by heparin, dimethylsulfoxide (DMSO), and sodium butyrate (SB) were examined in three rat hepatoma cell lines and two rat liver epithelial cell lines. Based on antigenic profiles and gene expression in serum-supplemented medium, the hepatoma cell lines could be organized in distinct categories of hepatic differentiation. All three hepatomas expressed the following five genes: gamma-glutamyl transpeptidase (GGT), glutathione-S-transferase pi (Yp), glutamine synthetase, and alpha 5 and beta 1 integrin. Cell line H4AzC2 also expressed alpha-fetoprotein (AFP), albumin. IGF II receptor, and the biliary/oval cell antigens OC.2 and OC.3, a phenotype characteristic of fetal hepatocytes. FTO-2B cells lacked AFP, OC.2, and OC.3 but expressed albumin and IGF II receptor in addition to the five commonly expressed genes, consistent with a more hepatocyte-like phenotype. Cell line H5D-7 expressed neither albumin nor the IGF II receptor, but did express OC.2, OC.3, and alpha 3 integrin in addition to the five commonly expressed genes, characteristic of biliary epithelial cells. Regulation of gene expression by heparin, DMSO, and SB was examined in cells cultured in hormonally defined medium. The patterns of regulation of AFP, albumin, GGT, and Yp were dependent upon the state of differentiation of the cell. FTO-2B cells regulated genes in a manner similar to that of E16 fetal hepatocytes, H4AzC2 regulated genes characteristic of both hepatocytic and biliary lineages, and H5D.7 regulated only biliary genes. Suppression of GGT by DMSO was uniformly observed. The three cell lines expressed equal amounts of HNF-4, but FTO-2B cells expressed more HNF-3 beta and less HNF-3 alpha, while the reverse was true of H4AzC2 and H5D.7 cells.

Electron-beam computed tomography, coronary artery calcium, and evaluation of patients with coronary artery disease.

PURPOSE: To briefly review the role of calcium in the pathophysiology of atherosclerosis and to comprehensively review and analyze studies of coronary artery calcium detected by electron-beam computed tomography (CT). DATA SOURCES: The English-language literature located through MEDLINE and Current Contents. STUDY SELECTION: All studies of electron-beam CT in symptomatic and asymptomatic patients with and without known coronary artery disease were selected. DATA EXTRACTION: Significant findings on the association of cardiac risk factors and angiographically evident coronary artery disease with coronary artery calcium detected on electron-beam CT were compared. Prospective data on clinical outcomes in patients with coronary artery calcium were assessed. DATA SYNTHESIS: Coronary artery calcium is common in patients with known coronary artery disease or risk factors for coronary artery disease, and it becomes more common with increasing age. Coronary artery calcium detected by electron-beam CT is a sensitive but not a specific indicator of angiographically evident atherosclerosis; sensitivity is increased and specificity is decreased for angiographically significant disease. Test characteristics can be adjusted to improve specificity at the cost of sensitivity. Very limited data suggest that patients with coronary artery calcium are more likely to have cardiac events. CONCLUSIONS: Electron-beam CT is a promising new tool for the evaluation of coronary artery disease because patients who have coronary artery calcium are likely to have angiographically evident atherosclerosis. However, too few data currently exist to support the broad use of this tool in clinical decision making during the evaluation of patients with known or suspected coronary artery disease.

Maturation-dependent gene expression in a conditionally transformed liver progenitor cell line.

We have isolated a conditionally transformed liver progenitor cell line with phenotypic similarities to both hepatoblasts (bipotent embryonic liver cells that give rise to hepatocytes and intrahepatic biliary epithelial cells) and liver epithelial cells (primitive hepatic cells isolated from adult livers capable of generating both hepatocytic and biliary lineages). Cell line L2039 was derived from E14 fetal mouse liver after transformation with temperature-sensitive SV-40 large T antigen. At 33 degrees C, these cells have an epithelial morphology with a high nucleocytoplasmic ratio and express both hepatocytic and biliary genes, including albumin, alpha-fetoprotein, glutamine synthetase, insulinlike growth factor II receptor, fibronectin and laminin, and cytokeratins 8 and 19, a set of markers characteristic for hepatoblasts. The presence of cytokeratin 14, vimentin, and several oval-cell antigens link cell line L2039 to nonparenchymal liver epithelial cell populations thought to contain progenitor cells. Serum-free, hormonally defined media conditions and extracellular matrix requirements were determined for growth and differentiation of this cell line. During culture on type IV collagen at 39 degrees C, L2039 cells cease dividing and demonstrate hepatocytic differentiation with the assumption of a hepatocytelike morphology and glucocorticoid-dependent regulation of liver-specific genes, including albumin, alpha-fetoprotein, phosphoenolpyruvate carboxykinase, and liver-enriched transcription factors. The number of albumin-positive cells increases during culture at 39 degrees C, indicating that L2039 cells convert from a prehepatocytic to a hepatocytic phenotype. Under conditions specific for hepatocytic differentiation, C/EBPs were expressed and differentially regulated, with C/EBPbeta and C/EBPdelta upregulated early and C/EBPalpha only slightly expressed after 7 d, indicating that C/EBPalpha may not be a crucial factor in commitment to the hepatocytic phenotype.

Paraproteinemia, plasmacytoma, myeloma and HIV infection.

We have identified by MEDLINE search the cases of gammaglobinopathy and plasma cell malignancy in HIV-positive patients reported in the English language literature. The average age at presentation among HIV-positive patients with plasma cell disorders is 33 years, far younger than the average age of presentation in the general population. Some of these patients present with transient paraproteinemias, while others have persistent paraproteins, which may or may not be associated with true plasma cell malignancies. In most cases in which it has been examined, the paraprotein contains high-titer anti-HIV activity. The presence of high-titer anti-HIV activity in the paraproteins of AIDS patients suggests that an antigen-driven process in response to HIV infection may contribute to the early development of plasma cell disorders in these patients. Recent work in plasma cell tumorigenesis has indicated that transformation at a single point in the B lymphocyte lineage can give rise to either lymphoma or myeloma, dependent upon environmental factors such as T cell function, which may be required for directing transformed lymphocytes from lymphoma and towards plasma cell differentiation. This may explain why B lineage oncogenesis in AIDS patients favors the development of lymphoma over that of myeloma.

Hypercalcemia and alkalosis due to the milk-alkali syndrome: a case report and review.

At one time, when antacids were the primary medical means of treating peptic ulcer disease, the milk-alkali syndrome was not an uncommon cause of hypercalcemia. The simultaneous occurrence of hypercalcemia, alkalosis, and renal failure, in conjunction with the appropriate history of ingestion fof antacids, was suggestive of the syndrome. With the advent of antisecretory therapy, however, the milk-alkali syndrome has become an uncommon diagnosis. I report a case of milk-alkali syndrome and review the history of this syndrome as reported in the medical literature. Contemporary reports have focused on understanding the pathophysiology of the syndrome. Recent series have identified a shifting demographic profile, as increasing numbers of elderly women consume calcium carbonate as an anti-osteoporosis measure.

Sleep, genes and death: fatal familial insomnia.

Over the past 30 years, significant progress has been made in understanding the physiologic mechanisms of sleep. Insomnia, a common complaint in general medical practice, and other sleep disorders have become increasingly recognized. In 1986, a heritable total insomnia was described and termed fatal familial insomnia; since then, the pathology of this disease has been shown to involve an accumulation of prion particles in the brains of affected patients. Prions have been more commonly associated with the transmission of spongiform encephalopathies such as scrapie (in sheep), Creutzfeldt-Jakob disease and Kuru. We briefly review the physiological and biochemical characteristics of normal sleep, describe the typical clinical characteristics of fatal familial insomnia and describe the current understanding of how prions cause neurodegenerative diseases, including fatal familial insomnia.

Disruption of cell-cell adhesion in the presence of sodium butyrate activates expression of the 92 kDa type IV collagenase in MDCK cells.

We have investigated the effects of altered cell shape on the regulation of the 92 kDa type IV collagenase. In MDCK cells, anti-E-cadherin antibodies alter cell shape by disrupting normal cell-cell contacts, while sodium butyrate causes a marked flattening and spreading of cells. The disruption of cell-cell contacts led to a faint expression of the 92 kDa collagenase. This effect was enhanced by sodium butyrate, which by itself did not induce collagenase expression. In contrast, stromelysin expression was not regulated in these conditions. Although mRNA expression was enhanced, the secreted collagenase activity was not altered in these conditions in either cell line. Examination of cytoskeletal and extracellular matrix proteins and cell-cell and cell-matrix adhesion proteins by immunofluorescence and Western blot revealed a disruption of the actin network, tight junctions, and fibronectin deposition by anti E-cadherin antibodies, and alterations in actin, cytokeratin 8, cytokeratin 14, laminin and beta 1 integrin induced by sodium butyrate. Thus, the induction of collagenase expression in epithelial cells by disrupted cell-cell adhesion and sodium butyrate is associated with changes in cell shape and structure.

Intrauterine contraceptive device-associated actinomycotic abscess and Actinomyces detection on cervical smear.

OBJECTIVE: To summarize clinical features and treatments in reported cases of actinomycotic pelvic abscess occurring in women using intrauterine contraceptive devices (IUD), and to review detection of Actinomyces by cervical smear. DATA SOURCES: The English-language medical literature accessed through MEDLINE. METHODS OF STUDY SELECTION: We identified 92 cases of actinomycotic abscesses associated with IUD use in 63 case reports. In addition, 31 studies of Actinomyces detection were found, 16 of which were studies of Papanicolaou smear-based detection. DATA ABSTRACTION AND SYNTHESIS: Data regarding clinical presentation and treatment were culled from case reports, whereas detection rates of Papanicolaou smear and other methods were obtained from studies of Actinomyces detection. The average patient was 37 years old, had been using an IUD for 8 years, and presented with abdominal pain, weight loss, vaginal discharge, and fever. Laboratory studies commonly revealed anemia, leukocytosis, and an elevated erythrocyte sedimentation rate. Most of these patients underwent operative procedures, usually hysterectomy and salpingoophorectomy. High-dose penicillin was found to be an effective antibiotic. Detection rates of organisms on Papanicolaou smear were somewhat variable; use of other detection methods, including endometrial biopsy, culture, and immunofluorescence, did not improve this variability. CONCLUSIONS: Pelvic actinomycosis associated with the use of IUDs can mimic pelvic malignancy; for that reason, it is often treated surgically. However, if the diagnosis of actinomycosis can be obtained preoperatively, antibiotic treatment may lead to complete resolution. The Papanicolaou smear may be useful in evaluating such patients.

The liver as a stem cell and lineage system.

We propose that the liver is a stem cell and lineage system with many parallels to lineages in the bone marrow, gut, and epidermis, varying from them only in kinetics. All are organized with three compartments: a slow cycling stem cell compartment with cells expressing a fetal phenotype and responding slowly to injury; an amplification compartment with cells of intermediate phenotype rapidly proliferating in response to regenerative stimuli or acute injuries; and a terminal differentiation compartment in which cells increasingly differentiate and gradually lose their ability to divide. In all systems, both those with slow or rapid kinetics, the various compartments are positioned in a polarized organization, are associated with a gradient in the chemistry of the extracellular matrix, and show lineage-position-dependent growth responses, gene expression, pharmacological and toxicological responses, and reaction to viruses and radiation. In general, known oncogens selectively kill cells in the differentiation compartment inducing chronic regenerative responses of the cells in stem cell and/or amplification compartment. Tumors arise by subsequent transformation of the activated stem cells or early precursor cells. The evidence for a lineage model consists of the data implicating gradients in cell size, ploidy, growth potential, and antigenic and gene expression in the liver parenchyma along the sinusoidal plates. The traditional explanation for this heterogeneity is that it represents adaption of cells to a changing sinusoidal microenvironment dictated by the direction of blood flow. However, we review the extant data and suggest that it more readily supports a lineage model involving a maturation process beginning with stem cells and precursors in the periportal zone and ending with sensescing parenchyma near the central vein. Support for this theory is provided by the studies on phenotypic heterogeneity in liver, investigations into the embryology of the liver, and analyses of the responses of liver to chemical and viral oncogens that induce rapid proliferation of small cells with oval-shaped nuclei, "oval cells," now thought to be closely related to liver stem cells. The lineage model provides clarity and insights into many aspects of liver biology and disease including the limited proliferative ability of in vitro parenchymal cultures, liver regeneration, gene expression, viral infection, hepatocellular carcinogenesis, liver cell transplantation, and aging.