Cellular origin of regenerating parenchyma in a mouse model of severe hepatic injury.

Several treatments in rodents, including administration of the alkylating agent dipin, followed by two-thirds partial hepatectomy in mice combine destruction of liver parenchyma with hepatocyte mitoinhibition. These treatments induce proliferation of bile epithelial-like cells (termed oval cells), development of foci composed of small hepatocytes, and eventual replacement of damaged parenchyma by healthy hepatocytes. It has been proposed that these oval cells represent transitional cells in a nonhepatocytic liver facultative stem cell lineage that can give rise to the small hepatocyte foci, and that these foci eventually become confluent and replace liver parenchyma. In this study, we used in vivo cell lineage marking in genetically chimeric livers to test the hypothesis that hepatocytes can serve as the precursor cell type to the small hepatocyte foci that develop in mouse liver after treatment with dipin plus partial hepatectomy. Although we do not exclude the possibility that some small hepatocyte foci may be stem cell-derived, we demonstrate that hepatocyte-derived foci are present after dipin-induced liver damage in mice.

Hepatocyte transplantation in a model of toxin-induced liver disease: variable therapeutic effect during replacement of damaged parenchyma by donor cells.

To provide long-term therapy in patients with severe toxin-induced hepatic parenchymal damage, donor hepatocytes would need to replicate and replace a large portion of the damaged parenchyma. Using a mouse model developed to reproduce this type of hepatic injury, we found that hepatocyte transplantation only slightly improved survival after transplantation despite the fact that many non-survivors showed moderate liver repopulation by donor cells. Perhaps accounting for this outcome, donor parenchyma in non-survivors did not have typical lobular organization. These results indicate that the re-creation of functional parenchyma by transplanted hepatocytes requires time, during which donor cells proliferate and then establish normal parenchymal architecture.

Mainstream and sidestream cigarette smoke condensates suppress macrophage responsiveness to interferon gamma.

Sidestream smoke evolves from the smoldering end of a cigarette while the smoker is not puffing, and contributes substantially to environmental tobacco smoke (ETS). In contrast, main stream smoke emerges from the butt end of the cigarette and is mainly inhaled by the smoker. This study was performed to compare the effects of short-term exposure to cigarette smoke condensates prepared from sidestream (CSC-SS) and mainstream cigarette smoke (CSC-MS) on macrophage basal metabolism and responsiveness to two different stimuli, bacterial lipopolysaccharide (LPS) and interferon gamma (IFNgamma). Despite their generation at different temperatures and their different chemical composition, CSC - SS and CSC - MS had similar effects on macrophages. Both enhanced macrophage basal metabolism and responsiveness to LPS. Macrophage responsiveness to IFNgamma, assessed by their expression of four functional capacities, was suppressed by both CSC-SS and CSC-MS. The four assessed IFNgamma-inducible functional capacities were: enhanced phagocytosis of immuoglobulin-opsonized sheep red blood cells, TPA-induced peroxide production, class II major histocompatibility complex expression, and nitric oxide synthesis with LPS co-stimulation. The effects of CSC - SS and CSC - MS were similar qualitatively; they differ quantitatively in some cases, with CSC-MS generally effective at lower concentrations (expressed as cigarette-equivalents) than CSC-SS. Considering dilution of sidestream smoke in room air and loss during passage through the respiratory system, we expect to deliver the maximal dose to lung macrophages in situ only in rooms dense with smokers. However, only a fraction of the maximal dose can partially suppress induction of some functions, such as nitric oxide production and MHC expression. Macrophages play critical roles in tissue modeling during development. Of particular concern are neonates, whose organs are still undergoing growth and development, and are therefore susceptible to impaired development. If involuntary exposure to ETS hinders induction of macrophage functional capacities by cytokines, then development of the lungs and perhaps other organs would be impaired. In general, since macrophages are potent effectors and regulators of immunity, impairment of their responsiveness to cytokine must disrupt the proper functioning of the immune system.

Liver disease and compensatory growth: unexpected lessons from genetically altered mice.

Over the last decade, several animal models have been established that permit exploration of liver biology and disease. Although these models have been developed using diverse strategies, including transgene targeting, homozygous gene disruption and administration of hepatotoxic chemicals, each approach creates an animal with hepatocyte damage, resulting in an hepatic microenvironment that supports proliferation of healthy hepatocytes. These models have been used to demonstrate: (1) the remarkable ability of adult hepatocytes to clonally proliferate in response to liver growth signals, (2) the effectiveness of transplanted donor hepatocytes in repopulating damaged liver parenchyma, and (3) the feasibility of reconstituting liver with xenogeneic hepatocytes. This paper reviews the development and use of these models, and outlines their potential future application to the study of hepatic stem cells, therapy of liver disease and hepatic toxicology.

Immunotoxicology of cigarette smoke condensates: suppression of macrophage responsiveness to interferon gamma.

We have investigated systematically the effects of short-term exposure to main stream cigarette smoke condensates (CSC-MS) on basal and inducible functional capacities of murine peritoneal exudate macrophages. Macrophages treated with CSC-MS form granules that fluoresce orange under blue excitation, consistent with the speculation that they are polycyclic aromatic hydrocarbons (PAH). CSC-MS selectively suppressed interferon gamma (IFN gamma) induction of four macrophage functional capacities: enhanced phagocytosis of immunoglobulin-opsonized sheep red blood cells, TPA-induced H2O2 production, class II major histocompatibility complex expression, and nitric oxide synthesis. In contrast, two macrophage functions that are not induced by IFN gamma, basal electron transport and LPS-induced TNF alpha production, were enhanced by treatment with CSC-MS. These results suggest that the suppressive effects of CSC-MS on macrophage responsiveness were selective and were not due to nonspecific inhibition of general functions such as RNA or protein synthesis. Since macrophage responsiveness to IFN gamma can result in induction of functional capacities that are fundamental to immunity, the data suggest that CSC-MS maybe deleterious to the general health of the smoker.

A method for isolating human lung macrophages and observations of fluorescent phagocytes from the lungs of habitual cigarette smokers.

We report the development of a simple, efficient, expedient, and inexpensive procedure for isolating a large and relatively pure population of macrophages (Mphi) from residual (i.e., non-tumor) lung tissue obtained from lung cancer patients undergoing either a lobectomy or pneumonectomy. The proposed technique was founded on observations by fluorescent microscopy of fresh, non-fixed, and non-stained human lung tissue. Examinations of 74 specimens from different patients revealed that most of the Mphi reside as non-adherent cells within the sponge-like lung stroma. Very few Mphi were observed in the lungs of nonsmokers. In contrast, many Mphi were visible in the lungs of habitual smokers. For most specimens from smokers, a few of the Mphi were present as randomly distributed single cells; the majority of the Mphi, however, were in clusters that ranged from a dozen to several hundred cells. The Mphi could be released readily by different mechanical techniques. In the procedure reported herein, pulmonary leukocytes (> 75% Mphi) were dislodged easily from lung tissue with the use of an inexpensive, hand-operated, tissue grinder. The grinder consisted of a glass mortar and Teflon pestle that provided sufficient clearance between the mortar and pestle so as to avert damaging the displaced leukocytes. The leukocytes were then segregated by centrifugation on a density gradient. Further purification was achieved by harvesting Mphi that had been allowed to adhere to serum-coated polystyrene culture dishes (> 90% Mphi). In most experiments, the Mphi yield (approximately 5 x 10(6) Mphi /gr of lung) and Mphi viability (> 85%) were good. A significant advantage of this technique is that it avoids jeopardizing the cells to the hazards associated with enzymes that have been used in techniques employed previously for isolating Mphi from the lung and other organs. Thus, the proposed method provides numerous lung Mphi for detailed studies of their morphology, phenotype, and function. Moreover, lung Mphi were cultured as non-adherent, single cells in a serum- and cytokine-free tissue culture medium for more than 6 weeks. Lung Mphi from habitual smokers displayed a high level of fluorescence that was readily apparent when viewed with a fluorescence microscope that had been configured with either a fluorescein or rhodamine filter. Serial sections of single, living Mphi obtained with the use of a confocal laser scanning microscope revealed that the fluorescence originated from cytoplasmic inclusions. Relative fluorescence intensity was measured by cytometry.(ABSTRACT TRUNCATED AT 400 WORDS)