ALBERT

Description: Background: Macrophage activation syndrome (MAS) in adults, also called reactive haemophagocytic syndrome or haemophagocytic lymphohistiocytosis, is the result of inappropriate proliferation and activation of the mononuclear phagocytic system secondary to an underlying disease. Persistent high levels of inflammatory cytokines with impaired cytotoxic capacity of T-lymphocytes and natural killer cells seem to be the main trigger of the syndrome. There are a variety of diseases that can lead to fulminant uncontrolled MAS, mainly viral infections and conditions of immunologic dysregulation such as Systemic Lupus Erythematodes, Morbus Still and neoplastic disorders. Despite the variety of associated diseases, the clinical presentation is uniform with high fever, splenomegaly, lymphadenopathy, hepatomegaly, rash and neurologic abnormalities. MAS is associated with substantial morbidity and mortality. Beside the clinical presentation, extreme elevation of plasma markers for macrophage hyperactivation are mandatory in diagnosing and monitoring MAS (hyperferritinemia, and extreme elevation of neopterin and sIL2R), whereas the phenomenon of hemophagocytosis (±cytopenias) is more variable. Different therapeutic strategies including high-dose steroids, intravenous immunoglobulin (IVIG) [Emmenegger U, et al. American Journal of Haematology2001;68:4–10] and intravenous etoposide (VP16)[Imashuku S, et al. Journal of Clinical Oncology2001;19:2665–267] have been reported to induce remissions, but tend to be incomplete and of short duration. These treatments are mainly based on studies in children with familial haemophagocytic lymphohistiocytosis and have considerable side-effects. The optimal treatment-approach in adults with MAS remains unclear. Faced with high mortality of MAS at our institution, we accepted VP16 as a paticularly important component of therapy but were struck by fast and live threatening relapses during neutropenic episodes following classical pulse-application of VP16 (2x150mg/m²/d x 2/week). We initiated oral low dose VP16 in refractory RHS and observed substantial effects. Methods and Patients: Our treatment concept was that of uninterrupted macrophage mitigation by continuous low-dose oral VP16 after a first un- or only partially successful episode of high-dose methylprednisolone or IVIG. In the present study we describe the effect of low dose oral VP16 after a first episode treated with high-dose methylprednisolone or IVIG in 3 patients with relapsing or resistant RHS. Close monitoring of cytokine activation was performed by serial measurements of ferritin, sIL2R, transcobalamin II and neopterin (at least twice weekly). 2 of our patients suffered from recurrent MAS in the context of chronic active Epstein-Barr virus infection, 1 patient showed 2 episodes of MAS secondary to Morbus Still. VP16-treatment was dosed at diurnal 50mg for 3 days followed by 15–20mg daily avoiding neutropenic episodes. Results: During the time of observation (4 years) we have ascertained a total of 9 episodes of MAS, 3 of them classified as severe (admission to intensive care unit). 4 episodes of MAS have been managed by oral VP16 exclusively. Decrease of cytokine activation and clinical outcome in episodes treated with oral VP16 proofed to be superior to the other treatment-strategies. Conclusion: Compared to steroids and IVIG, strictly monitored long-term low-dose oral VP16 (for several weeks to months) seems to be a safe and most effective treatment strategy in patients with life-threatening reactive MAS.

Description: The ability of red blood cells (RBCs) to transport gases, their lifespan as well as their rheological properties invariably depend on the deformability, hydration, and membrane stability of these cells, which can be measured by Laser optical rotational red cell analyser (Lorrca® Maxsis, RR Mechatronics). The osmoscan mode of Lorrca is currently used in diagnosis of rare anemias in clinical laboratories. However, a broad range of normal values for healthy subjects reduces the sensitivity of this method for diagnosis of mild disease phenotype. In this pilot study, we explored the impact of age and gender of 45 healthy donors, as well as RBC age on the Lorrca indices. Whereas gender did not affect the Lorrca indices in our study, the age donors had a profound effect on the O_hyper parameter. To study the impact of RBC age on the osmoscan parameters, we have isolated low (L)-, medium (M)-, or high (H)- density fractions enriched with young, mature, and senescent RBCs, respectively, and evaluated the influence of RBC age-related properties, such as density, morphology, and redox state, on the osmoscan indices. As before, O_hyper was the most sensitive parameter, dropping markedly with an increase in RBC density and age. Senescence was associated with a decrease in deformability (EI_max) and tolerability to low and high osmolatites (Area). L-fraction was enriched with reticulocytes and cells with high projected area and EMA staining, but also contained a small number of cells small in projected area and most likely, terminally senescent. L-fraction was on average slightly less deformable than mature cells. The cells from the L-fraction produced more oxidants and NO than all other fractions. However, RBCs from the L-fraction contained maximal levels of reduced thiols compared to other fractions. Our study suggests that reference values for O_hyper should be age-stratified, and, most probably, corrected for the average RBC age. Further multi-center study is required to validate these suggestions before implementing them into clinical practice.

Description: (1) Background: It is known that sickle cells contain a higher amount of Ca2+ compared to healthy red blood cells (RBCs). The increased Ca2+ is associated with the most severe symptom of sickle cell disease (SCD), the vaso-occlusive crisis (VOC). The Ca2+ entry pathway received the name of Psickle but its molecular identity remains only partly resolved. We aimed to map the involved Ca2+ signaling to provide putative pharmacological targets for treatment. (2) Methods: The main technique applied was Ca2+ imaging of RBCs from healthy donors, SCD patients and a number of transgenic mouse models in comparison to wild-type mice. Life-cell Ca2+ imaging was applied to monitor responses to pharmacological targeting of the elements of signaling cascades. Infection as a trigger of VOC was imitated by stimulation of RBCs with lysophosphatidic acid (LPA). These measurements were complemented with biochemical assays. (3) Results: Ca2+ entry into SCD RBCs in response to LPA stimulation exceeded that of healthy donors. LPA receptor 4 levels were increased in SCD RBCs. Their activation was followed by the activation of Gi protein, which in turn triggered opening of TRPC6 and CaV2.1 channels via a protein kinase Cα and a MAP kinase pathway, respectively. (4) Conclusions: We found a new Ca2+ signaling cascade that is increased in SCD patients and identified new pharmacological targets that might be promising in addressing the most severe symptom of SCD, the VOC.