Fibrinogen synthesis in serum-free hepatocyte cultures: Stimulation by glucocorticoids

Grieninger, Gerd; Hertzberg, Kathe M.; Pindyck, Johanna

doi:10.1073/pnas.75.11.5506

Cited by 62 publications

(37 citation statements)

References 24 publications

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“…This behavior of albumin synthesis in our cultures is unchanged by inclusion of serum in the medium (15) and appears to be related to a decline of albumin messenger RNA (unpublished data). Although a number ofhormones have been shown to stimulate synthesis of specific plasma proteins in this culture system (8,9), none can replace the requirement of the cells for insulin to restore albumin synthesis. Insulin is also required in primary frog hepatocyte cultures for continued albumin synthesis (25).…”

Section: Methodsmentioning

confidence: 99%

“…Culture medium was replaced with an equal volume of fresh medium every 24 hr unless otherwise indicated. Heparin sodium salt was included, at a concentration of 15 ,ug/ml, in medium intended for fibrinogen assay (8). Insulin (bovine pancreas, crystalline, Sigma; 24.3 units/mg) was routinely used.…”

Section: Methodsmentioning

confidence: 99%

“…The preparation of hepatocyte suspensions from 16-day-old chicken embryos by perfusion of the liver, mechanical disruption, and treatment with purified dissociating enzymes was performed essentially as described (8,12). Modified Ham's F-12 medium (13) was used without insulin or serum supplementation so that the cells were washed, plated, and maintained in chemically defined hormone-free medium.…”

Section: Methodsmentioning

confidence: 99%

“…Clear interpretation of the effect of insulin was further complicated by the simultaneous presence of serum (6,7) or mixtures ofhormones (5,6). Some ofthese agents have been shown individually to stimulate the synthesis of specific plasma proteins in culture (8)(9)(10).…”

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Direct effect of insulin on the synthesis of specific plasma proteins: biphasic response of hepatocytes cultured in serum- and hormone-free medium.

Liang¹,

Grieninger²

1981

Proc. Natl. Acad. Sci. U.S.A.

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Monolayers of chicken embryo hepatocytes, cultured in chemically defined medium, retain the ability to synthesize a wide spectrum of plasma proteins for several days in the absence of added hormones. Addition of insulin to the medium elicited a biphasic stimulation ofplasma protein synthesis: a rapid response of the synthesis of a limited number of plasma proteins (e.g., albumin and a,-globulin "M"), then, after prolonged exposure to the hormone, the involvement ofadditional plasma proteins (e.g., fibrinogen and lipoproteins). Synthesis of transferrin and a few other plasma proteins was not affected by the presence of insulin. The degree of stimulation for the most responsive plasma proteins ranged between 2-to 4-fold during the early phase and 10-and even 30-fold during the late phase ofthe cells' response to insulin. Stimulated synthesis in the early phase was detected within 1 hr and was rapidly reversible. Plasma protein synthesis in culture was sensitive to concentrations ofinsulin below0.35 nM, well within the physiological range. The delayed response was elicited only at higher hormone levels. Parallels between the control ofsynthesis ofplasma proteins in this system and that observed in diabetic animals suggest that the embryonic chicken hepatocytes may be a useful model for studying liver function in diabetes as well as insulin action in general.The anabolic effect of insulin on carbohydrate metabolism in the liver is well documented (1). However, the role of insulin in regulating the hepatic function of plasma protein synthesis has not been clearly defined. In the early 1960s, experiments with perfused livers of diabetic rats showed a decrease in total plasma protein synthesis (2) as well as reduced synthesis of albumin (3) relative to that in normal rat liver. More recently, it has been reported that the lowered synthesis of hepatic secretory proteins by the diabetic rat can be restored to normal by injection of insulin (4).To investigate the effect of insulin on plasma protein synthesis without the influence of other body organs, a number of rat liver-derived systems have been used, including isolated perfused liver (5) and dispersed cell suspensions (6, 7). These studies reported modest stimulation of albumin synthesis (less than 2-fold) with little, if any, effect on fibrinogen synthesis. Long-term responses to the hormone were not rigorously examined, due to the limited viability of the systems employed. Clear interpretation of the effect of insulin was further complicated by the simultaneous presence of serum (6, 7) or mixtures ofhormones (5, 6). Some ofthese agents have been shown individually to stimulate the synthesis of specific plasma proteins in culture (8-10).Our study was undertaken to examine the direct action of insulin alone on the hepatocellular synthesis of a wide range of plasma proteins. It has been made possible by development of a hepatocyte monolayer culture derived from chicken embryos, which can sustain synthesis of a broad spectrum ofplasma proteins in medium free of...

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

See 2 more Smart Citations

Direct effect of insulin on the synthesis of specific plasma proteins: biphasic response of hepatocytes cultured in serum- and hormone-free medium.

Liang¹,

Grieninger²

1981

Proc. Natl. Acad. Sci. U.S.A.

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“…However, the mechanisms by which acute-phase reactants are induced are poorly understood. Macrophage factors, in particular interleukin I (2), and glucocorticoids (3,4) have both been implicated in induction of some acute-phase reactant proteins. We (5), Baumann et al (6), and Feinberg et al (7) have recently shown that AGP and its mRNA are induced several hundred-fold by glucocorticoids in the rat hepatoma cell line HTC.…”

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confidence: 99%

Glucocorticoid-mediated induction of alpha 1-acid glycoprotein: evidence for hormone-regulated RNA processing.

Vannice¹,

Taylor²,

Ringold³

1984

Proc. Natl. Acad. Sci. U.S.A.

196

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We have studied the glucocorticoid-mediated accumulation of a,-acid glycoprotein (AGP) knRNA in HTC rat hepatoma cells. In contrast to the well-characterized primary response of mouse mammary tumor vills, in viro transcription assays in isolated nuclei show that the rate of transcription of the AGP gene is high even in the absence of hormone. Despite the constitutive transcription of the AGP gene, no detectable AGP RNA can be found in either the cytoplasm or the nuclei of untreated cells. Previous experiments have shown that the glucocorticoid induction of AGP RNA requires ongoing protein synthesis. In conjunction with the present study, our data suggest that glucocot-ticoids stimulate accumulation of AGP RNA by inducing an RNA processing factor that allows production of stable transcripts.The acute phase reactant a,-acid glycoprotein (AGP)K also called orosomucoid, is one of several plasma proteins synthesized by the liver in response to various stressful stimuli. Physical trauma such as surgery or wounding, bacterial infection, or nonspecific inflammatory stimuli such as subcutaneous injection of turpentine elicit the so-called acutephase response (for review, see ref. 1). In addition to AGP, whose physiological function remains obscure, a variety of protease inhibitors (e.g., a1-anti-trypsin) and iron scavengers (e.g., haptoglobin and hemopexin) are also induced. However, the mechanisms by which acute-phase reactants are induced are poorly understood. Macrophage factors, in particular interleukin I (2), and glucocorticoids (3, 4) have both been implicated in induction of some acute-phase reactant proteins. We (5), Baumann et al. (6), and Feinberg et al. (7) have recently shown that AGP and its mRNA are induced several hundred-fold by glucocorticoids in the rat hepatoma cell line HTC.Glucocorticoids, and steroid hormones in general, act by altering the rates of transcription of specific genes. The model for steroid hormone action, originally proposed by Jensen et al. (8) The gene encoding AGP, however, appears to be regulated in quite a different manner. The several hundred-fold increase in AGP RNA observed in glucocorticoid-treated HTC cells is dependent on ongoing protein synthesis (5-7); this is in contrast to the induction of MMTV RNA, which proceeds normally in the absence of protein synthesis (16,17). Thus, the induction of AGP RNA by glucocorticoids is a secondary action of the hormone, presumably mediated by a protein that is induced by direct action of the glucocorticoid-receptor complex. In this manuscript, we describe experiments showing that the induction of AGP RNA occurs at a post-transcriptional level. Our data implicate a glucocorticoid-regulated RNA processing system in the production of AGP RNA. MATERIALS AND METHODSCells and Cell Culture. The cell lines used in these studies were derived from the rat hepatoma cell line HTC (18). JZ.1 and MSC1 are HTC-derived cell lines containing 1 and -10 MMTV proviral copies, respectively. They are described in more detail elsewhere (19). These ce...

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Liver Nonparenchymal Cells Are Stimulated to Provide Interleukin 6 for Induction of the Hepatic Acute-Phase Response in Endotoxemia but Not in Remote Localized Inflammation

Billiar

Curran²,

Williams³

et al. 1992

Arch Surg

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It has been postulated that Kupffer cells provide signals that regulate hepatocyte responses in sepsis and inflammation. Although in vitro data support such a hypothesis, to our knowledge, no in vivo evidence has been reported. We injected rats with lipopolysaccharide intraperitoneally to simulate sepsis or turpentine intramuscularly to mimic localized inflammation. Both treatments are known to induce the hepatic acute-phase response. Liver nonparenchymal cells and hepatocytes were isolated and placed in culture. Hepatocyte fibrinogen synthesis was measured as an indication of interleukin 6 exposure, while nonparenchymal interleukin 6 production was measured directly. Both lipopolysaccharide and turpentine stimulated a sharp increase in hepatocyte fibrinogen synthesis (turpentine greater than lipopolysaccharide). However, only lipopolysaccharide injection was associated with increased nonparenchymal cell interleukin 6 synthesis. Increased circulating levels of interleukin 6 could be found only after lipopolysaccharide injection. In addition, tumor necrosis factor synthesis was enhanced by lipopolysaccharide but not turpentine. Our data show that nonparenchymal cells are stimulated to provide the interleukin 6 signal to hepatocytes in endotoxemia but not in remote localized inflammation, even though both treatments stimulate the hepatic acute-phase response. Our findings support paracrine functions for liver sinusoidal cells in certain septic states.

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Fibrinogen synthesis in serum-free hepatocyte cultures: Stimulation by glucocorticoids

Abstract: Fibrinogen synthesis was investi ated in cultures of chicken embryo hepatocytes initiated and maintained in chemically defined serum-free medium. lI-Hydroxy glucocorticoids caused a 3-0old stimulation of fibrinogen synthesis.

Cited by 62 publications

References 24 publications

Direct effect of insulin on the synthesis of specific plasma proteins: biphasic response of hepatocytes cultured in serum- and hormone-free medium.

Direct effect of insulin on the synthesis of specific plasma proteins: biphasic response of hepatocytes cultured in serum- and hormone-free medium.

Glucocorticoid-mediated induction of alpha 1-acid glycoprotein: evidence for hormone-regulated RNA processing.

Liver Nonparenchymal Cells Are Stimulated to Provide Interleukin 6 for Induction of the Hepatic Acute-Phase Response in Endotoxemia but Not in Remote Localized Inflammation

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