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Ischemia-reperfusion injury (I/R) is the main cause of primary graft nonfunction. Our aim was to evaluate the effect of excessive versus acute administration of erythropoietin (EPO) in attenuating the hepatic injury induced by I/R in mice. The effect of segmental (70%) hepatic ischemia was evaluated in a transgenic mouse line with constitutive overexpression of human EPO cDNA and in wild-type (WT) mice. Mice were randomly allocated to 5 main experimental groups: (i) WT-sham, (ii) WT ischemia, (iii) WT ischemia + recombinant human erythropoietin (rhEPO), (iv) transgenic-sham, and (v) transgenic ischemia. The EPO-pretreated mice showed a significant reduction in liver enzyme levels and intrahepatic caspase-3 activity and fewer apoptotic hepatocytes (p < 0.05 for all) compared with the WT untreated I/R group. EPO decreased c-Jun N-terminal kinase (JNK) phosphorylation and nuclear factor-κB (NF-κB) expression during I/R. In transgenic I/R livers, baseline histology showed diffused hepatic injury, and no significant beneficial effect was noted between the WT untreated and the transgenic I/R mice. In conclusion, acute pretreatment with EPO in WT mice attenuated in vivo I/R liver injury. However, in excessive EPO overexpression, the initial liver injury abolished the beneficial effect of EPO. These findings have important implications for the potential use of acute EPO in I/R injury during liver transplantation. Key words: ischemia-reperfusion, erythropoietin, liver, transgenic mice, apoptosis, caspase 3, in vivo. La lesion d'ischemie-reperfusion (I/R) est la principale cause de dysfonction primaire du greffon. Nous avons eu pour objectif d'examiner l'effet d'une administration excessive ou aigue d'erythropo'ieetine (EPO) sur l'attenuation de la lesion hepatique induite par l'I/R chez les souris. Nous avons eevalue l'effet d'une ischemie hepatique segmentaire (70 %) dans une ligneee de souris transgeeniques presentant une surexpression constitutive d'ADNc codant pour l'EPO humaine et chez des souris de type sauvage (TS). Nous avons reparti aleeatoirement les souris en 5 groupes experimentaux : (i) non traite; TS ; (ii) ischemie TS; (iii) ischemie TS + EPOrh ; (iv) transgenique non traite; ;et(v) ischemie transgenique. Les souris pretraite;es a l'EPO ont montre une reduction enzymatique significative des taux d'enzymes heipatiques et de l'activite; de la caspase-3 intrahe;patique, et un plus faible taux d'hepatocytes apoptotiques (p < 0,05 pour tous) comparativement au groupe I/R non traite; TS. L'EPO a diminue la phosphorylation de JNK et l'expression de NF-κB durant l'I/R. Dans les foies du groupe I-R transgenique, l'histologie basale a montre une lesion hepatique diffuse, et aucun effet beneefique significatif n'a ete observe chez les souris transgeeniques par comparaison aux souris non trauees TS. En conclusion, un pretraitement aigu a l'EPO a atteenue la leesion heepatique induite par l'I/R in vivo chez les souris TS. Toutefois, dans la surexpression excessive d'EPO, la lesion hepatique initiale a supprime l'effet beneefique de l'EPO. Ces reesultats ont d'importantes implications pour ce qui est de l'utilisation potentielle d'EPO aigue dans la leesion d'I/R durant une transplantation he;patique. Mots-cles: ischemie-reperfusion, eerythropoietine, foie, souris transgeniques, apoptose, caspase 3, in vivo. [Traduit par la Redaction], Introduction Ischemia-reperfusion (I/R) injury is the main cause of both primary graft dysfunction (occurring in 10%-30% of grafts) and primary nonfunction of liver allografts (occurring in 5% of grafts) (Clavien [...]

The active metabolite of vitamin D calcitriol and its analogs are well-known for their anti-inflammatory action in the skin, while their main side effect associated with topical treatment of inflammatory disorders is irritant contact dermatitis. Prostaglandin E2 (PGE2) is proinflammatory at the onset of inflammation and anti-inflammatory at its resolution. We hypothesized that induction of PGE2 synthesis by calcitriol in epidermal keratinocytes may contribute both to its pro-inflammatory and anti-inflammatory effects on the skin. Treatment of human immortalized HaCaT keratinocytes with calcitriol (3-100 nM, 2-24 h) increased PGE2 production due to increased mRNA and protein expression of COX-2, but not to increase of COX-1 or release of arachidonic acid. The effect of calcitriol on COX-2 mRNA was observed also in primary human keratinocytes. The increase in COX-2 mRNA is associated with COX-2 transcript stabilization. Calcitriol exerts this effect by a rapid (2 h) and protein synthesis independent mode of action that is dependent on PKC and Src kinase activities. Treatment with a COX-2 inhibitor partially prevented the attenuation of the keratinocyte inflammatory response by calcitriol. We conclude that upregulation of COX-2 expression with the consequent increase in PGE2 synthesis may be one of the mechanisms explaining the Janus face of calcitriol as both a promoter and attenuator of cutaneous inflammation. [ABSTRACT FROM AUTHOR]

Calcitriol, the hormonal form of vitamin D, inhibited caspase-3-like activation in HaCaT keratinocytes exposed to hyperosmotic and oxidative stresses, heat shock, and the inflammatory cytokine TNF. The hormone also protected the cells from caspase-independent cell death induced by hyperosmotic and oxidative stresses. The protection against hyperosmotic stress is not affected by inhibitors of the EGF receptor, ERK or PI13 kinase pathways, neither is it due to reduced activity of the proapoptotic p38 MAP kinase. These results are in accordance with previous in vivo findings that vitamin D protects epidermal keratinocytes from apoptosis due to UV radiation or chemotherapy. [ABSTRACT FROM AUTHOR]

Agents that increase intracellular cAMP (cAMP elevating agents) and 1,25(OH)2D3 inhibit the proliferation of many cell types. We investigated the combined effect of 1,25(OH)2D3 and cAMP elevating agents on exponentially growing mouse 3T3 fibroblasts. The following cAMP elevating agents were used: theophylline and pentoxyfilline, which inhibit cAMP-dependent phosphodiesterase; prostaglandin E2 which activates adenylate cyclase by a receptor-mediated mechanism; forskolin, which directly stimulates adenylate cyclase; and the cell permeable cAMP analogs 8-bromo cAMP and N6 benzoyl cAMP. 1,25(OH)2D3 and cAMP elevating agents were added to exponentially growing fibroblasts cultured in 96-well microtiter plates and cell number was monitored 3–7 d later. 1,25(OH)2D3 and the cAMP elevating agents as single agents inhibited the growth of the 3T3 cells. The combined treatment of the fibroblasts with 1,25(OH)2D3 and the cAMP elevating agents resulted in an antiproliferative effect that was more than additive. The synergistic interaction depended on the dose of 1,25(OH)2D3 and was apparent already at 10−8 M of the hormone. The specificity of the effect of 1,25(OH)2D3 was demonstrated by the finding that 24,25-dihydroxyvitamin D3, a vitamin D metabolite with low affinity for the vitamin D receptor, did not affect the antiproliferative effect of cAMP elevating agents. From the synergistic interaction between 1,25(OH)2D3 and the cell permeable cAMP analogs, we infer that the site of interaction between the two signaling pathways is distal to the cAMP generating and degrading machinery. [ABSTRACT FROM AUTHOR]