Caractérisation du phénotype hépatosplénique d'un nouveau modèle de rats acéruloplasminémiques pour une meilleure compréhension des mécanismes physiopathologiques de l'acéruloplasminémie héréditaire

Hereditary aceruloplasminemia is a rare genetic disease, resulting in a lack of functional ceruloplasmin in plasma. This pathology leads to iron overload, especially in hepatocytes. However, liver and spleen macrophages are not iron overloaded, despite the reported important role of ceruloplasmin in iron egress from macrophages. Our hypothesis was that ceruloplasmin is not essential for iron egress from macrophages, and that some mechanisms, not in direct link with the ceruloplasmin reported role, are the cause of the hepatosplenic phenotype observed in the disease. We performed a synthesis of aceruloplasminemia clinical cases reported in the literature, confirming by this way the presence of an hépatosplenic phenotype that suggest an imperfect undersanding of ceruloplasmin role. We then performed and characterized a new rat model of aceruloplasminemia, reproducing the human hepatosplenic phenotype. The data obtained in this model suggested that ceruloplasmin is not essential for macrophage iron egress. Otherwise, some other results highlighted abnormalities regarding to non-iron metals metabolism, in link with aceruloplasminemia.L’acéruloplasminémie héréditaire est une pathologie génétique rare, résultant en une absence de céruloplasmine fonctionnelle dans le plasma. La pathologie entraîne des surcharges en fer, notamment au niveau hépatocytaire. Cependant, les macrophages hépatiques et spléniques des patients ne semblent pas impactés par la surcharge en fer, alors même que la céruloplasmine est décrite comme nécessaire à la sortie du fer macrophagique. Notre hypothèse était que la céruloplasmine n’est pas indispensable à la sortie du fer macrophagique, et que des mécanismes sans lien direct avec le rôle rapporté de la céruloplasmine sont à l’origine du phénotype hépatosplénique rencontré dans la pathologie. Nous avons réalisé une synthèse des cas cliniques d’acéruloplasminémie rapportés dans la littérature, confirmant la présence d’un phénotype hépatosplénique suggérant que le rôle de la céruloplasmine est incomplètement compris. Nous avons par la suite développé et caractérisé un modèle original de rats acéruloplasminémiques, reproduisant le phénotype hépatosplénique humain. Les données récoltées dans ce modèle suggéraient que la céruloplasmine n’est pas nécessaire à la sortie du fer macrophagique. D’autres résultats ont par ailleurs mis en évidence des anomalies du métabolisme des métaux non-ferreux, en lien avec la pathologie

Characterization of the hepatosplenic phenotype from a new aceruloplasminemic rat model for a better understanding of hereditary aceruloplasminemia pathophysiological mechanisms

L’acéruloplasminémie héréditaire est une pathologie génétique rare, résultant en une absence de céruloplasmine fonctionnelle dans le plasma. La pathologie entraîne des surcharges en fer, notamment au niveau hépatocytaire. Cependant, les macrophages hépatiques et spléniques des patients ne semblent pas impactés par la surcharge en fer, alors même que la céruloplasmine est décrite comme nécessaire à la sortie du fer macrophagique. Notre hypothèse était que la céruloplasmine n’est pas indispensable à la sortie du fer macrophagique, et que des mécanismes sans lien direct avec le rôle rapporté de la céruloplasmine sont à l’origine du phénotype hépatosplénique rencontré dans la pathologie. Nous avons réalisé une synthèse des cas cliniques d’acéruloplasminémie rapportés dans la littérature, confirmant la présence d’un phénotype hépatosplénique suggérant que le rôle de la céruloplasmine est incomplètement compris. Nous avons par la suite développé et caractérisé un modèle original de rats acéruloplasminémiques, reproduisant le phénotype hépatosplénique humain. Les données récoltées dans ce modèle suggéraient que la céruloplasmine n’est pas nécessaire à la sortie du fer macrophagique. D’autres résultats ont par ailleurs mis en évidence des anomalies du métabolisme des métaux non-ferreux, en lien avec la pathologie.Hereditary aceruloplasminemia is a rare genetic disease, resulting in a lack of functional ceruloplasmin in plasma. This pathology leads to iron overload, especially in hepatocytes. However, liver and spleen macrophages are not iron overloaded, despite the reported important role of ceruloplasmin in iron egress from macrophages. Our hypothesis was that ceruloplasmin is not essential for iron egress from macrophages, and that some mechanisms, not in direct link with the ceruloplasmin reported role, are the cause of the hepatosplenic phenotype observed in the disease. We performed a synthesis of aceruloplasminemia clinical cases reported in the literature, confirming by this way the presence of an hépatosplenic phenotype that suggest an imperfect undersanding of ceruloplasmin role. We then performed and characterized a new rat model of aceruloplasminemia, reproducing the human hepatosplenic phenotype. The data obtained in this model suggested that ceruloplasmin is not essential for macrophage iron egress. Otherwise, some other results highlighted abnormalities regarding to non-iron metals metabolism, in link with aceruloplasminemia

Interleukin-6, C/EBP-beta and PPAR-gamma expression correlates with intramuscular liposarcoma growth in mice: The impact of voluntary physical activity levels

International audienceIL-6 is an axial cytokine overexpressed in cancer to promote growth and increase resistance to anticancer therapies. As the application of IL-6-targeting therapies are still limited, alternative non aggressive and adjuvant approaches, like physical activity (PA) could be useful to reverse IL-6 effects. To get more insights into liposarcoma (LS) pathophysiology, we investigated potential molecular links between IL-6 and LS growth and we tested the impact of PA on such mechanism in an orthotopic model of intramuscular LS. Initially active nude mice have received an intramuscular injection of either human SW872 cells or vehicle, then were respectively randomized into voluntary-active or inactive mice with open or restricted access to activity-wheels. We found that LS-bearing mice exhibited-6 fold increase in circulating IL-6 comparing to controls, with a concomitant decrease in hepatic drug-metabolizing enzymes expression. Circulating IL-6 levels were positively correlated with intra-tumor IL-6 expression (r = 0.85, P < 0.01). Interestingly, intra-tumor IL-6, C/EBP-4 and PPAR-gamma expression were correlated together and with greater tumor mass and autophagy markers, notably, GABARAPL-1. Intriguingly, we found that maintaining a spontaneous PA after tumor injection did not reduce the levels of IL-6, but even enhanced tumor growth, induced body weight loss and increased the risk of developing lung metastasis. Our findings suggest that (1) IL-6, C/EBP-a and PPAR-gamma exert a potential role in promoting growth of dedifferentiated LS and (2) that PA failed to mechanistically interfere with these factors, but enhanced LS growth via other independent-mechanisms. The preclinical data reported here could be helpful in the sub-molecular classification of LS patients to improve diagnosis and design a low-risk treatment. Circulating IL-6 could serve as an indicator for treatment follow-up and, perhaps, for infra-radiologic LS relapses

Iron as a Therapeutic Target in <i>HFE</i>-Related Hemochromatosis: Usual and Novel Aspects

Genetic hemochromatosis is an iron overload disease that is mainly related to the C282Y mutation in the HFE gene. This gene controls the expression of hepcidin, a peptide secreted in plasma by the liver and regulates systemic iron distribution. Homozygous C282Y mutation induces hepcidin deficiency, leading to increased circulating transferrin saturation, and ultimately, iron accumulation in organs such as the liver, pancreas, heart, and bone. Iron in excess may induce or favor the development of complications such as cirrhosis, liver cancer, diabetes, heart failure, hypogonadism, but also complaints such as asthenia and disabling arthritis. Iron depletive treatment mainly consists of venesections that permit the removal of iron contained in red blood cells and the subsequent mobilization of stored iron in order to synthesize hemoglobin for new erythrocytes. It is highly efficient in removing excess iron and preventing most of the complications associated with excess iron in the body. However, this treatment does not target the biological mechanisms involved in the iron metabolism disturbance. New treatments based on the increase of hepcidin levels, by using hepcidin mimetics or inducers, or inhibitors of the iron export activity of ferroportin protein that is the target of hepcidin, if devoid of significant secondary effects, should be useful to better control iron parameters and symptoms, such as arthritis

Ceruloplasmin deficiency does not induce macrophagic iron overload: lessons from a new rat model of hereditary aceruloplasminemia

International audienceHereditary aceruloplasminemia (HA), related to mutations in the ceruloplasmin (Cp) gene, leads to iron accumulation. Ceruloplasmin ferroxidase activity being considered essential for macrophage iron release, macrophage iron overload is expected, but it is not found in hepatic and splenic macrophages in humans. Our objective was to get a better understanding of the mechanisms leading to iron excess in HA. A clustered regularly interspaced short palindromic repeats (CRISPR)/ CRISPR associated protein 9 (Cas9) knockout of the Cp gene was performed on Sprague-Dawley rats. We evaluated the iron status in plasma, the expression of iron metabolism genes, and the status of other metals whose interactions with iron are increasingly recognized. In Cp rats, plasma ceruloplasmin and ferroxidase activity were absent, together with decreased iron concentration and transferrin saturation. Similarly as in humans, the hepatocytes were iron overloaded conversely to hepatic and splenic macrophages. Despite a relative hepcidin deficiency in Cp rats and the loss of ferroxidase activity, potentially expected to limit the interaction of iron with transferrin, no increase of plasma non-transferrin-bound iron level was found. Copper was decreased in the spleen, whereas manganese was increased in the plasma. These data suggest that the reported role of ceruloplasmin cannot fully explain the iron hepatosplenic phenotype in HA, encouraging the search for additional mechanisms.-Kenawi, M., Rouger, E., Island, M.-L., Leroyer, P., Robin, F., Remy, S., Tesson, L., Anegon, I., Nay, K., Derbré, F., Brissot, P., Ropert, M., Cavey, T., Loréal, O. Ceruloplasmin deficiency does not induce macrophagic iron overload lessons from a new rat model of hereditary aceruloplasminemia

Intermittent reloading does not prevent iron availability decrease and hepcidin upregulation caused by hindlimb unloading

International audienceNew findings - What is the central question of this study? Could skeletal muscle be involved in microgravity-induced iron misdistribution by modulating expression of hepcidin, the master regulator of iron metabolism? What is the main finding and its importance? We demonstrate, in rats, that hepcidin upregulation is not a transient adaptation associated with early exposure to microgravity and that intermittent reloading does not limit microgravity-induced iron misdistribution despite having a beneficial effect on soleus muscle wasting. Abstract - In humans, exposure to microgravity during spaceflight causes muscle atrophy, changes in iron storage and a reduction in iron availability. We previously observed that during 7 days of simulated microgravity in rats, hepcidin plays a key role in iron misdistribution, and we suggested that a crosstalk between skeletal muscle and liver could regulate hepcidin synthesis in this context. In the present study in rats, we investigated the medium-term effects of simulated microgravity on iron metabolism. We also tested whether intermittent reloading (IR) to target skeletal muscle atrophy limits iron misdistribution efficiently. For this purpose, Wistar rats underwent 14 days of hindlimb unloading (HU) combined or not combined with daily IR. At the end of this period, the serum iron concentration and transferrin saturation were significantly reduced, whereas hepatic hepcidin mRNA was upregulated. However, the main signalling pathways involved in hepcidin synthesis in the liver (BMP-small mothers against decapentaplegic (SMAD), interleukin-6-STAT3 and ERK1/2) were unaffected. Unlike what was observed after 7 days of HU, the iron concentration in the spleen, liver and skeletal muscle was comparable between control animals and those that underwent HU or HU plus IR for 14 days. Despite its beneficial effect on soleus muscle atrophy and slow-to-fast myosin heavy chain distribution, IR did not significantly prevent a reduction in iron availability and hepcidin upregulation. Altogether, these results highlight that iron availability is durably reduced during longer exposure to simulated microgravity and that the related hepcidin upregulation is not a transient adaptation to these conditions. The results also suggest that skeletal muscle does not necessarily play a key role in the iron misdistribution that occurs during simulated microgravity