Mobilization of Mg2+ from rat heart and liver mitochondria following the interaction of thyroid hormone with the adenine nucleotide translocase.

The in vitro addition of thyroid hormone to isolated rat heart or liver mitochondria induces the extrusion of approximately 2-4 nmol Mg2+/mg protein from both mitochondria preparations. The mobilization of Mg2+ is not accompanied by extrusion of matrix ATP or K+, or by mitochondria swelling, thus excluding that the phenomenon occurs through the nonspecific opening of the mitochondrial permeability transition pore. Moreover, the Mg2+ extrusion is completely prevented by bongkrekic acid, a membrane-permeant inhibitor of the adenine nucleotide translocase (AdNT), and by cyclosporine, which has also been reported to inhibit AdNT in a bongkrekate-like manner, operating at the matrix site of the translocase. By contrast, atractyloside, another specific inhibitor of AdNT that operates at the cytosolic site of the AdNT, only partially affects the Mg2+ mobilization (< 30% inhibition). These findings and the binding of 125I-labeled thyroid hormone to both the dimeric and monomeric moiety of AdNT support the hypothesis that AdNT can operate as a specific receptor for thyroid hormone in the mitochondria, and suggest that thyroid hormone operates at the matrix site of the translocase. In addition, these observations may imply that some of the so called "nongenomic effects" exerted by thyroid hormone on mitochondrial metabolism could occur through changes in the matrix content of Mg2+.

Profiles of immunoregulatory cytokine production in vitro in patients with IgA nephropathy and their kindred.

We hypothesized that the altered immunoglobulin synthesis and/or lymphocyte function apparent in patients with IgA nephropathy (IgAN) is due to a primary defect in lymphokine regulation. In addition, we reasoned that such changes in lymphokine production might be, at least partially, genetically determined. To assess the extent of lymphocyte abnormalities, we investigated the profile of cytokine production from peripheral blood mononuclear cells (PBMC) in 34 IgAN patients and 44 of their first degree relatives, 10 of whom had persistent microhaematuria. Compared with healthy volunteers (n = 34), PBMC from patients showed increased IL-2 production both spontaneously or after phytohaemagglutinin (PHA) (20 micrograms/ml) stimulation, whereas IL-4 and interferon-gamma (IFN-gamma) production were significantly higher only after stimulation. Microhaematuric relatives had a similar pattern of cytokine production, whereas non-microhaematuric relatives showed no significant difference versus normals. The altered pattern of cytokine production appeared to be quite specific to IgAN patients and their microhaematuric relatives, because patients with other forms of primary glomerulonephritis (n = 17) did not differ from normal individuals. Patients and relatives that hyperproduced IL-4 were also hyperproducers of IL-2. No such congruence was seen in any other group or with any other pairing of cytokines. We propose that a subpopulation of IgAN patients bear lymphocytes intrinsically hyperresponsive. Among those individuals such hyperresponsiveness may be causally related to the pathogenesis and/or character of IgAN.

Hormonal stimulation of Mg2+ uptake in hepatocytes. Regulation by plasma membrane and intracellular organelles.

Collagenase dispersed rat liver hepatocytes release Mg2+ when stimulated with norepinephrine or accumulate Mg2+ when stimulated with vasopressin, respectively. Mg2+ fluxes in either direction account for a net loss or gain of approximately 10% of total cell magnesium and are rapidly reversible. Both stimulated Mg2+ efflux and Mg2+ influx require physiological concentration of extracellular NaCl and Ca2+. In the absence of extracellular Na+, Mg2+ efflux, but not influx, can be observed in the presence of extracellular Cl-. Under these conditions, the efflux is inhibited by the Cl-/HCO3- exchanger inhibitor 4,4'-dinitrostilbene-2,2'-disulfonic acid. In hepatocytes, Mg2+ influx, but not efflux, is completely inhibited by thapsigargin, a specific inhibitor of the endoplasmic reticulum Ca2+ ATPase. Several lines of evidence, such as measurements of cytosolic Ca2+ or of cytosolic Ca2+ buffering, indicate that the effect of thapsigargin in inhibiting Mg2+ influx could not be explained by an increase in cytosolic Ca2+. Instead, the inhibition of hepatocyte Mg2+ influx was found to be the result of the depletion of the Ca2+ stored within the endoplasmic reticulum.

Regulation of magnesium uptake and release in the heart and in isolated ventricular myocytes.

Perfused rat hearts release or accumulate approximately 10% of total Mg2+ content when stimulated with norepinephrine (NE) or carbachol, respectively. Collagenase-dispersed rat ventricular myocytes increase or decrease total cell Mg2+ by 1 mM within 5 minutes when stimulated with these same transmitters. Measurements of Mg2+ transport using 28Mg or atomic absorbance spectrophotometry indicate that the rate and the extent of both stimulated Mg2+ efflux and influx are independent of the concentration of extracellular Mg2+ (0 to 1.2 mM). Mg2+ release induced by NE is rapidly reversed by the addition of carbachol, and Mg2+ uptake induced by carbachol is reversed by NE. Decreasing extracellular Na+ or Ca2+ decreases or abolishes Mg2+ efflux from myocytes. Cd2+ or other Ca2+ channel blockers also inhibit Mg2+ efflux in the presence of a physiological concentration of extracellular Ca2+. Replacement of extracellular Ca2+ with Sr2+ or with Mn2+ decreases or abolishes both stimulated efflux and influx of Mg2+. Redistribution of 85Sr in myocytes and in the supernatant indicates that under those conditions Sr2+ is released or accumulated by NE or carbachol in a manner similar to that of Mg2+. Hence, at least in the case of Sr2+, the inhibition of Mg2+ fluxes can be explained by the transport of Sr2+ rather than Mg2+ through the transport(s) systems. By contrast, replacement of extracellular Ca2+ with Ba2+ inhibits stimulated Mg2+ uptake but not Mg2+ release. These results indicate that cardiac myocytes have a major pool of Mg2+ that can be rapidly mobilized upon hormonal stimulation. The net uptake and release of Mg2+ are quantitatively similar and appear to be independent of the extracellular Mg2+ concentrations but are affected, to various degrees, by the presence of other cellular or extracellular cations.

Cell magnesium transport and homeostasis: role of intracellular compartments.

Magnesium transport across the plasma membrane of cardiac and liver cells appears to be under hormonal control. The increase in cytosolic cAMP, following the adrenergic stimulation of both cell types, results in a major Mg2+ efflux from perfused rat hearts or livers and from collagenase-dispersed ventricular myocytes or hepatocytes. By contrast, the activation of protein kinase C by carbachol, vasopressin, phorbol-myristate acetate or diacylglycerol analogs induces Mg2+ accumulation in either of the experimental models. As for the role of intracellular compartments on Mg2+ homeostasis, the cAMP-mediated Mg2+ efflux largely depends on the mobilization of Mg2+ from mitochondria via the mitochondrial adenine nucleotide translocase. By contrast, Mg2+ influx appears to be related to the endo(sarco)plasmic reticulum and its dynamic handling of cytosolic Ca2+.

Regulation of Mg2+ uptake in isolated rat myocytes and hepatocytes by protein kinase C.

A large Mg2+ cell uptake against concentration gradients is stimulated in collagenase-dispersed rat myocytes by carbachol and in hepatocytes by carbachol or vasopressin. The signalling pathway(s) responsible for this stimulation of Mg2+ uptake was investigated by using various activators or inhibitors of protein kinase C (PKC) and by correlating Mg2+ uptake with cell PKC activity and cAMP content. In both cell preparations, the direct stimulation of PKC by diacylglycerol analogs or phorbol esters reproduce the same pattern of Mg2+ uptake as that induced by carbachol or vasopressin. These data indicate that the activation of PKC is responsible for a stimulation of Mg2+ uptake by myocytes or hepatocytes, whereas increase in cAMP in these cells stimulates Mg2+ release.

Involvement of peripheral blood monocytes in haemodialysis: in vivo induction of tumour necrosis factor alpha, interleukin 6 and beta 2-microglobulin.

Our aim was to evaluate the role of three different variables in the activation of the monocyte system: dialysis membrane (cuprophane or polyacrylonitrile), dialysate (acetate or bicarbonate), and procedure (standard or high-flux haemodialysis). By ELISA test we measured the 'in vivo' intracellular (monocyte-associated) production and extracellular release of tumour necrosis factor alpha (TNF alpha), interleukin-6 (Il-6) and beta 2-microglobulin (beta 2-M) by monocytes from 20 uraemic patients before and after the dialysis session. At the beginning of the dialysis session, uraemic patients' cultured monocytes spontaneously released a greater amount of TNF alpha, Il-6 and beta 2-M compared to normal controls. However, no differences in cytokine and beta 2-M were observed in monocyte lysate between the two groups. At the end of the dialysis session, cultured monocytes from patients treated with cellulosic membranes and acetate dialysate showed greater TNF alpha values than normal controls (P less than 0.001 and P less than 0.05 respectively). Moreover, TNF alpha and Il-6 values were strictly correlated (P less than 0.05). These results clearly show an activation of the monocyte system in uraemic patients undergoing periodic haemodialysis. The implicated factors may be multiple, such as complement activating cellulosic membranes or acetate dialysate. The production of TNF alpha, Il-6 and beta 2-M may explain some of the pathological findings observed in long-term haemodialysis patients.