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Betaine, taurine, and inositol participate as osmolytes in liver cell volume homeostasis and interfere with cell function. In this study we investigated whether osmolytes are also released from the intact liver independent of osmolarity changes. In the perfused rat liver, phagocytosis of carbon particles led to a four- to fivefold stimulation of taurine efflux into the effluent perfusate above basal release rates. This taurine release was inhibited by 70-80% by the anion exchange inhibitor DIDS or by pretreatment of the rats with gadolinium chloride. Administration of vasopressin, cAMP, extracellular ATP, and glucagon also increased release of betaine and/or taurine, whereas insulin, extracellular UTP, and adenosine were without effect. In isolated liver cells, it was shown that parenchymal cells and sinusoidal endothelial cells, but not Kupffer cells and hepatic stellate cells, release osmolytes upon hormone stimulation. This may be caused by a lack of hormone receptor expression in these cells, because single-cell fluorescence measurements revealed an increase of intracellular calcium concentration in response to vasopressin and glucagon in parenchymal cells and sinusoidal endothelial cells but not in Kupffer cells and hepatic stellate cells. The data show that Kupffer cells release osmolytes during phagocytosis via DIDS-sensitive anion channels. This mechanism may be used to compensate for the increase in cell volume induced by the ingestion of phagocytosable material. The physiological significance of hormone-induced osmolyte release remains to be evaluated.

Background & Aims: CD95 (Apo-1/Fas) ligand suppresses inflammatory responses in immune-privileged organs. In this study, modulation of the hepatic CD95 receptor/ligand system by interferon gamma and cyclosporin A was investigated., Methods: CD95 receptor and ligand expression were measured at the messenger RNA level by using quantitative reverse-transcription polymerase chain reaction and immunocytochemistry in primary cultures of rat Kupffer cells, hepatocytes, and T lymphocytes. Soluble CD95 in culture supernatants was detected by enzyme-linked immunosorbent assay and apoptosis by the TUNEL method., Results: Interferon gamma treatment led to an increase in CD95 ligand messenger RNA levels in Kupffer cells followed by an overexpression of the soluble CD95 receptor. Supernatants derived from 24-hour but not from 48-hour interferon gamma-treated Kupffer cells killed lymphocytes by a CD95-dependent mechanism. Cyclosporin A inhibited CD95 ligand expression in Kupffer cells and lymphocyte killing. In liver parenchymal cells, interferon gamma increased messenger RNA levels of the transmembrane CD95 isoform and sensitivity of these cells toward CD95-mediated apoptosis., Conclusions: The expression pattern of CD95 receptor and ligand in response to interferon gamma points to a coordinated interplay between Kupffer cells, hepatocytes, and T lymphocytes in which Kupffer cells may regulate programmed cell death of T lymphocytes and hepatocytes.

The expression of glutamine synthetase (GS) was studied in cultured quiescent hepatic stellate cells (HSC) and during their transformation into myofibroblast-like cells. GS mRNA was detectable in quiescent HSC (1-day culture); however, the enzyme protein was not expressed, as assessed by Western blot analysis, immunocytochemistry and the absence of detectable enzyme activity. Similar findings were obtained after 2 days of culture; in addition, the mRNA levels had dropped by about 70%, but they increased again thereafter during the process of HSC transformation in culture, as indicated by the expression of alpha-smooth-muscle actin. In parallel with the accumulation of alpha-smooth-muscle actin, GS was expressed, as shown by Western blot analysis and immunocytochemistry, and enzyme activity increased from undetectable levels in quiescent cells to 0.13+/-0.01 micromol/h per mg of cell protein within 7-14 days. This value compares with GS activity in liver parenchymal cells of 0.57+/-0.03 micromol/h per mg of cell protein. The findings suggest that activation of HSC results in the de novo expression of GS protein and activity, and this may serve as another marker of HSC transformation.

Betaine, also known as trimethylglycine, is an important human nutrient obtained from a variety of foods and also can be synthesized from choline. Betaine is much more abundant in kidney and liver compared to other mammalian organs. The principal role of betaine in the kidney is osmoprotection in cells of the medulla and it enters these cells via the betaine/γ-aminobutyric acid (GABA) transporter protein (BGT1), which is upregulated by hyperosmotic stress. This process has been studied in great detail. In liver, the main role of betaine is a methyl donor in the methionine cycle. However, recent studies showed that BGT1 is much more abundant in liver compared to kidney medulla. Despite this, the role of BGT1 in liver has received little attention. Entry of betaine into liver cells is a necessary first step for its action at the cellular level. Increased interest in betaine has developed because of a number of therapeutic uses. These include treatment of nonalcoholic fatty liver and hyperhomocysteinemia, a risk factor for atherosclerotic disease. Several important questions need to be addressed to better understand the potential of betaine as a therapeutic agent for other liver diseases, such as alcohol-induced injury. Heavy alcohol consumption is the most common cause for liver-related deaths and altered liver metabolism may contribute to hepatic, vascular, coronary, and cerebral diseases. © 2014 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

It has been proposed that key enzymes of ureagenesis and the alanine aminotransferase activity predominate in periportal hepatocytes. However, ureagenesis from alanine, when measured in the perfused liver, did not show periportal predominance and even the release of the direct products of alanine transformation, lactate and pyruvate, was higher in perivenous cells. An alternative way of analyzing the functional distributions of alanine aminotransferase and the urea cycle along the hepatic acini would be to measure alanine and urea production from precursors such as lactate or pyruvate plus ammonia. In the present work these aspects were investigated in the bivascularly perfused rat liver. The results of the present study confirm that gluconeogenesis and the associated oxygen uptake tend to predominate in the periportal region. Alanine synthesis from lactate and pyruvate plus ammonia, however, predominated in the perivenous region. Furthermore, no predominance of ureagenesis in the periportal region was found, except for conditions of high ammonia concentrations plus oxidizing conditions induced by pyruvate. These observations corroborate the view that data on enzyme activity or expression alone cannot be extrapolated unconditionally to the living cell. The current view of the hepatic ammonia-detoxifying system proposes that the small perivenous fraction of glutamine synthesizing perivenous cells removes a minor fraction of ammonia that escapes from ureagenesis in periportal cells. However, since urea synthesis occurs at high rates in all hepatocytes with the possible exclusion of those cells not possessing carbamoyl-phosphate synthase, it is probable that ureagenesis is equally important as an ammonia-detoxifying mechanism in the perivenous region. [ABSTRACT FROM AUTHOR]

Summary.  Using microdialysis, we have monitored extracellular levels of amino acids and related amines in the human liver at three stages of the transplantation procedure: donor retrieval, back table preparation and during 48 h post-implantation. By comparing the ratio of mean levels at the donor and back table stages, with the ratio between early (2–6 h) and late (43–48 h) post-reperfusion, these amines were classified into one of three groups. In one group, back table levels were markedly higher than during the donor stage, with levels declining over time post-reperfusion. A second group had much lower levels in the back table than during the donor phase, and post-reperfusion levels were either stable or increased over time. Concentrations of amino acids in the final group remained relatively constant at all stages. This study illustrates the value of microdialysis in providing organ-specific metabolic data that may indicate specific mechanisms of poor graft function. [ABSTRACT FROM AUTHOR]

Cell hydration changes critically affect liver metabolism and gene expression. In the course of gene expression studies using nylon cDNA-arrays we found that hyperosmolarity (405 mosmol/l) suppressed the betaine-homocysteine methyltransferase (Bhmt) mRNA expression in H4IIE rat hepatoma cells. This was confirmed by Northern blot and real-time quantitative RT-PCR analysis, which in addition unraveled a pronounced induction of Bhmt mRNA expression by hypoosmotic (205 mosmol/l) swelling. Osmotic regulation of Bhmt mRNA expression was largely paralleled at the levels of Bhmt protein and enzymatic activity. Like hyperosmotic NaCl, hyperosmotic raffinose but not hyperosmotic urea suppressed Bhmt mRNA expression, suggesting that cell shrinkage rather than increased ionic strength or hyperosmolarity per se is the trigger. Hypoosmolarity increased the expression of a reporter gene driven by the entire human BHMT promoter, whereas destabilization of BHMT mRNA was observed under hyperosmotic conditions. Osmosensitivity of Bhmt mRNA expression was impaired by inhibitors of tyrosine kinases and cyclic nucleotide-dependent kinases. The osmotic regulation of BHMT may be part of a cell volume-regulatory response and additionally lead to metabolic alterations that depend on the availability of betaine-derived methyl groups. [ABSTRACT FROM AUTHOR]

Betaine-homocysteine S-methyltransferase (BHMT) is the only enzyme known to catabolize betaine. In addition to being a substrate for BHMT, betaine also functions as an osmoprotectant that accumulates in the kidney medulla under conditions of high extracellular osmolarity. The mechanisms that regulate the partitioning of betaine between its use as a methyl donor and its accumulation as an osmoprotectant are not completely understood. The aim of this study was to determine whether BHMT expression is regulated by salt intake. This report shows that guinea pigs express BHMT in the liver, kidney, and pancreas and that the steady-state levels of BHMT mRNA in kidney and liver decrease 68% and 93% in guinea pigs consuming tap water containing high levels of salt compared with animals provided untreated tap water. The animals consuming the salt water also had ∼50% less BHMT activity in the liver and kidney, and steady-state protein levels decreased ∼30% in both organs. Pancreatic BHMT activity and protein levels were unaffected by the high salt treatment. The complex mechanisms involved in the downregulation of hepatic and renal BHMT expression in guinea pigs drinking salt water remain to be clarified, but the physiological significance of this downregulation may be to expedite the transport and accumulation of betaine into the kidney medulla under conditions of high extracellular osmolarity. [ABSTRACT FROM AUTHOR]