Serpinb3 is overexpressed in the liver in presence of iron overload.

Iron overload results in cellular toxicity, tissue injury, organ fibrosis and increased risk of neoplastic transformation. SerpinB3 is a serine protease inhibitor overexpressed in the liver in oxidative stress conditions, able to induce fibrosis and increased risk of malignant transformation. Aim of the present study was to assess the effect of iron overload on SerpinB3 expression in the liver using in vivo and in vitro models.The expression of Serpinb3 was assessed in the liver of hemojuvelin knockout mice (Hjv-/-), an established model of hereditary hemochromatosis, and of wild type control mice, following dietary or pharmacological iron manipulation. To assess the direct effect of iron in vitro, cell lines were treated with different concentration of hemin or with an iron chelator.Hepatic Serpinb3 mRNA and protein were highly expressed in Hjv-/- mice, but not in wild type controls fed with a standard diet. Serpinb3 became detectable in wild type mice fed with a high iron diet or injected with iron dextran; these treatments further induced Serpinb3 expression in Hjv-/- mice. Livers expressing Serpinb3 showed a positive staining also for HIF-2α in the same areas. Hemin promoted induction of SerpinB3 mRNA in HeLa and HA22T/VGH cells, but a mild stimulation of SerpinB3 promoter activity in HeLa and Huh7 cells. In conclusion, Serpinb3 is strongly induced by iron in the mouse liver. The molecular link between iron, ROS and SerpinB3 seems to be HIF-2α, which is induced by iron overload and was previously found capable of up-regulating SerpinB3 at the transcriptional level.

Hfe and Hjv exhibit overlapping functions for iron signaling to hepcidin.

UNLABELLED: Functional inactivation of HFE or hemojuvelin (HJV) is causatively linked to adult or juvenile hereditary hemochromatosis, respectively. Systemic iron overload results from inadequate expression of hepcidin, the iron regulatory hormone. While HJV regulates hepcidin by amplifying bone morphogenetic protein (BMP) signaling, the role of HFE in the hepcidin pathway remains incompletely understood. We investigated the pathophysiological implications of combined Hfe and Hjv ablation in mice. Isogenic Hfe (-)/(-) and Hjv (-)/(-) mice were crossed to generate double Hfe (-)/(-) Hjv (-)/(-) progeny. Wild-type control and mutant mice of all genotypes were analyzed for serum, hepatic, and splenic iron content, expression of iron metabolism proteins, and expression of hepcidin and Smad signaling in the liver, in response to a standard or an iron-enriched diet. As expected, Hfe (-)/(-) and Hjv (-)/(-) mice developed relatively mild or severe iron overload, respectively, which corresponded to the degree of hepcidin inhibition. The double Hfe (-)/(-) Hjv (-)/(-) mice exhibited an indistinguishable phenotype to single Hjv (-)/(-) counterparts with regard to suppression of hepcidin, serum and hepatic iron overload, splenic iron deficiency, tissue iron metabolism, and Smad signaling, under both dietary regimens. We conclude that the hemochromatotic phenotype caused by disruption of Hjv is not further aggravated by combined Hfe/Hjv deficiency. Our results provide genetic evidence that Hfe and Hjv operate in the same pathway for the regulation of hepcidin expression and iron metabolism. KEY MESSAGES: Combined disruption of Hfe and Hjv phenocopies single Hjv deficiency. Single Hjv(-)/(-) and double Hfe(-)/(-)Hjv(-)/(-) mice exhibit comparable iron overload. Hfe and Hjv regulate hepcidin via the same pathway.

Better documentation improves patient care.

This article is the sixth in a series of seven describing the journey within NHS Lanarkshire in partnership with the University of the West of Scotland to support nursing and midwifery leadership roles through Scotland's Leading Better Care programme. Preceding articles have provided an overview of the programme and discussed a range of staff development work programmes. This article describes work carried out on clinical documentation to promote delivery of the three quality ambitions of safe, effective and person-centred care.

The role of calcium influx pathways in phospholipase D activation in bovine adrenal glomerulosa cells.

The steroid hormone aldosterone maintains sodium homeostasis and is therefore important in the control of blood volume and pressure. Angiotensin II (AngII) and elevated extracellular potassium concentrations ([K(+)](e)), the prime physiologic regulators of aldosterone secretion from adrenal glomerulosa cells, activate phospholipase D (PLD) in these cells. The role of Ca(2+) in the activation by these agents is unknown, although nitrendipine, a voltage-dependent Ca(2+) channel antagonist, does not inhibit AngII-elicited PLD activation, despite the fact that this compound blocked elevated [K(+)](e)-stimulated PLD activity. PLD activation triggered by AngII was also unaffected by the T-type calcium channel inhibitor nickel. Nevertheless, Ca(2+) influx was required for AngII-induced PLD activation in both primary cultures of bovine adrenal glomerulosa cells and a glomerulosa cell model, the NCI H295R adrenocortical carcinoma cell line. The involvement of store-operated Ca(2+) (SOC) influx and Ca(2+) release-activated Ca(2+) (CRAC) influx pathways in PLD activation was investigated using thapsigargin, an endoplasmic reticulum Ca(2+) pump inhibitor that empties the store to induce SOC influx, and the SOC inhibitor YM-58483 (BTP2), as well as a CRAC inhibitor, tyrphostin A9. In bovine glomerulosa cells, tyrphostin A9 inhibited AngII-induced PLD activation without affecting elevated [K(+)](e)-stimulated enzyme activity. On the other hand, differences were observed between the bovine adrenal glomerulosa and H295R cells in the involvement of Ca(2+) influx pathways in PLD activation, with the involvement of the SOC pathway suggested in the H295R cells. In summary, our results indicate that Ca(2+) entry only through certain Ca(2+) influx pathways is linked to PLD activation.

Phorbol ester increases mitochondrial cholesterol content in NCI H295R cells.

The first step in steroidogenesis is cholesterol mobilization from cytosolic lipid droplets to the initiating rate-limiting enzyme complex located on the inner mitochondrial membrane. Angiotensin II (AngII), the primary agonist of aldosterone secretion from adrenal glomerulosa cells, is known to induce cholesterol mobilization to mitochondria. However, the role of the protein kinase C (PKC) pathway in mediating cholesterol mobilization is unknown. To determine PKC's involvement, human adrenocortical carcinoma cells were incubated with or without PKC-activating phorbol 12-myristate 13-acetate (PMA) and mitochondrial cholesterol content assayed. Like AngII, PMA significantly elevated mitochondrial cholesterol content as well as aldosterone secretion. Thus, PKC may play a role in cholesterol mobilization to mitochondria and hence steroid production. Atrial natriuretic peptide (ANP) inhibited both AngII- and PMA-stimulated mitochondrial cholesterol content. These findings suggest that the ability of ANP to inhibit steroidogenesis induced by multiple agents may be related to its capacity to reduce cholesterol mobilization.

Characterization and phospholipase D mediation of the angiotensin II priming response in adrenal glomerulosa cells.

Bovine adrenal glomerulosa cells are primed by an initial treatment with angiotensin II (AngII) to respond with enhanced secretion to a second exposure to AngII or agents that increase calcium influx. We hypothesized that the mechanism of priming involves a persistent increase in diacylglycerol (DAG) generated via sustained activity of phospholipase D (PLD). In this report, we sought to define the time frame of this priming response as well as determine its mechanism using assays of aldosterone secretion, PLD activation, and radiolabeled diacylglycerol levels. We found that in primary cultures priming was observed for up to 50 min after AngII washout, suggesting that the priming window is protracted in these cultures relative to freshly isolated cells. The phorbol ester, phorbol 12,13-dibutyrate (PDBu), was used to investigate the role of sustained PLD activation in the persistent DAG and priming responses. PDBu was able to both prime glomerulosa cells to respond with enhanced secretion to AngII and elicit a persistent increase in DAG after PDBu washout. This persistent increase in DAG levels with an initial exposure to PDBu or AngII was not the result of maintained PLD activity after agent removal because PLD activation returned to basal levels by 30 min after washout. Finally, inhibition of PLD signaling during the initial AngII treatment inhibited the subsequent response to AngII or another agent that increases calcium influx. Thus, our results suggest that persistent DAG resulting from PLD signaling mediates the priming response to AngII or PDBu.

A review of the Boston Naming Test and multiple-occasion normative data for older adults on 15-item versions.

The literature reveals numerous versions of the Boston Naming Test (BNT) but normative data for very old people are scarce. Using four shortened versions of the BNT we produced normative data and examined naming ability longitudinally in a population-based sample of community dwelling elderly people. Our initial sample comprised 803 people with an average age of 76 years (range 65-93) who were examined twice, 2 years apart. A sub-sample of 326 people were followed-up 6 years later and re-examined. Results indicated that age and education level, but not gender, affected naming ability.