Lena Erlandsson

Interferon-β is required for interferon-α production in mouse fibroblasts

The type I interferons — interferon-a (IFN-a) and interferon-b (IFN-b) — are critical for protection against viruses during the acute stage of viral infection [1,2]. Furthermore, type I interferons have been implicated as important mediators in the regulation of lymphocyte development [3], immune responses [4,5] and the maintenance of immunological memory of cytotoxic T cells [6,7]. The different IFN-a subtypes are encoded by 12 genes in the mouse [8] whereas IFN-b is encoded for by only one gene [9]. IFN-a and IFN-b have a high degree of sequence homology and are thought to interact with the same surface receptor on target cells [10,11]. As an approach to analysing the different biological functions of IFN-a and IFN-b, we have generated a mouse strain with an inactivated IFN-b gene. We report here that embryonic fibroblasts from such mice produce neither IFN-b nor IFN-a upon Sendai virus infection, whereas the production of IFN-a by leukocytes from the same strain of mice is intact. IFN-a production in embryonic fibroblasts from IFN-b–/– mice could be rescued by ‘priming’ the cells using exogenous IFN-b. These results imply a unique role for IFN-b in the induction of type I interferons in peripheral tissues.

Oxidative stress in preeclampsia and the role of free fetal hemoglobin.

Preeclampsia is a leading cause of pregnancy complications and affects 3–7% of pregnant women. This review summarizes the current knowledge of a new potential etiology of the disease, with a special focus on hemoglobin-induced oxidative stress. Furthermore, we also suggest hemoglobin as a potential target for therapy. Gene and protein profiling studies have shown increased expression and accumulation of free fetal hemoglobin in the preeclamptic placenta. Predominantly due to oxidative damage to the placental barrier, fetal hemoglobin leaks over to the maternal circulation. Free hemoglobin and its metabolites are toxic in several ways; (a) ferrous hemoglobin (Fe2+) binds strongly to the vasodilator nitric oxide (NO) and reduces the availability of free NO, which results in vasoconstriction, (b) hemoglobin (Fe2+) with bound oxygen spontaneously generates free oxygen radicals, and (c) the heme groups create an inflammatory response by inducing activation of neutrophils and cytokine production. The endogenous protein α1-microglobulin, with radical and heme binding properties, has shown both ex vivo and in vivo to have the ability to counteract free hemoglobin-induced placental and kidney damage. Oxidative stress in general, and more specifically fetal hemoglobin-induced oxidative stress, could play a key role in the pathology of preeclampsia seen both in the placenta and ultimately in the maternal endothelium.

Mice with an inactivated joining chain locus have perturbed IgM secretion.

A mouse with an inactivated joining chain locus was produced by gene targeting in embryonic stem cells by deleting the first exon. Heterozygote (J+ / − ) and homozygote (J− / − ) offspring from these mice showed normal total serum immunoglobulin levels and a normal peripheral B cell compartment when compared to wild‐type littermates. The distribution of serum immunoglobulin isotypes in serum was different; IgA levels were elevated while IgM levels were reduced in J− / − mice as compared to wild‐type mice. High molecular weight serum IgM was reduced in J+ / − and J− / − mice and instead found in oligomeric form of undefined structure. Furthermore, serum IgM from J+ / − and J− / − mice showed a reduced ability to activate complement. The number of splenic and bone marrow IgM plaque‐forming cells were reduced in unimmunized J+ / − as well as in J− / − mice. Furthermore, the number of plaque‐forming cells was reduced in B cells from both J+ / − and J− / − mice after stimulation with lipopolysaccharide in vitro. The perturbation of IgM production in J− / − mice appears to affect a late stage of differentiation, because cells with intracellular IgM were readily detected both in vivo and in vitro. Finally, after immunization with T‐dependent or T‐independent antigens the IgM component of the immune response was reduced in J− / − mice while only a marginal reduction of the IgG response was detected.

A1M Ameliorates Preeclampsia-Like Symptoms in Placenta and Kidney Induced by Cell-Free Fetal Hemoglobin in Rabbit

Preeclampsia is one of the most serious pregnancy-related diseases and clinically manifests as hypertension and proteinuria after 20 gestational weeks. The worldwide prevalence is 3-8% of pregnancies, making it the most common cause of maternal and fetal morbidity and mortality. Preeclampsia lacks an effective therapy, and the only “cure” is delivery. We have previously shown that increased synthesis and accumulation of cell-free fetal hemoglobin (HbF) in the placenta is important in the pathophysiology of preeclampsia. Extracellular hemoglobin (Hb) and its metabolites induce oxidative stress, which may lead to acute renal failure and vascular dysfunction seen in preeclampsia. The human endogenous protein, α1-microglobulin (A1M), removes cell-free heme-groups and induces natural tissue repair mechanisms. Exogenously administered A1M has been shown to alleviate the effects of Hb-induced oxidative stress in rat kidneys. Here we attempted to establish an animal model mimicking the human symptoms at stage two of preeclampsia by administering species-specific cell-free HbF starting mid-gestation until term, and evaluated the therapeutic effect of A1M on the induced symptoms. Female pregnant rabbits received HbF infusions i.v. with or without A1M every second day from gestational day 20. The HbF-infused animals developed proteinuria and a significantly increased glomerular sieving coefficient in kidney that was ameliorated by co-administration of A1M. Transmission electron microscopy analysis of kidney and placenta showed both intracellular and extracellular tissue damages after HbF-treatment, while A1M co-administration resulted in a significant reduction of the structural and cellular changes. Neither of the HbF-treated animals displayed any changes in blood pressure during pregnancy. In conclusion, infusion of cell-free HbF in the pregnant rabbits induced tissue damage and organ failure similar to those seen in preeclampsia, and was restored by co-administration of A1M. This study provides preclinical evidence supporting further examination of A1M as a potential new therapy for preeclampsia.

Inventory of Novel Animal Models Addressing Etiology of Preeclampsia in the Development of New Therapeutic/Intervention Opportunities.

Preeclampsia is a pregnancy‐related disease afflicting 3–7% of pregnancies worldwide and leads to maternal and infant morbidity and mortality. The disease is of placental origin and is commonly described as a disease of two stages. A variety of preeclampsia animal models have been proposed, but all of them have limitations in fully recapitulating the human disease. Based on the research question at hand, different or multiple models might be suitable. Multiple animal models in combination with in vitro or ex vivo studies on human placenta together offer a synergistic platform to further our understanding of the etiology of preeclampsia and potential therapeutic interventions. The described animal models of preeclampsia divide into four categories (i) spontaneous, (ii) surgically induced, (iii) pharmacologically/substance induced, and (iv) transgenic. This review aims at providing an inventory of novel models addressing etiology of the disease and or therapeutic/intervention opportunities.

Urinary Extracellular Vesicles of Podocyte Origin and Renal Injury in Preeclampsia

Renal histologic expression of the podocyte-specific protein, nephrin, but not podocin, is reduced in preeclamptic compared with normotensive pregnancies. We hypothesized that renal expression of podocyte-specific proteins would be reflected in urinary extracellular vesicles (EVs) of podocyte origin and accompanied by increased urinary soluble nephrin levels (nephrinuria) in preeclampsia. We further postulated that podocyte injury and attendant formation of EVs are related mechanistically to cellfree fetal hemoglobin (HbF) in maternal plasma. Our study population included preeclamptic (n=49) and normotensive (n=42) pregnant women recruited at delivery. Plasma measurements included HbF concentrations and concentrations of the endogenous chelators haptoglobin, hemopexin, and α1- microglobulin. We assessed concentrations of urinary EVs containing immunologically detectable podocyte-specific proteins by digital flow cytometry and measured nephrinuria by ELISA. The mechanistic role of HbF in podocyte injury was studied in pregnant rabbits. Compared with urine from women with normotensive pregnancies, urine from women with preeclamptic pregnancies contained a high ratio of podocin-positive to nephrin-positive urinary EVs (podocin+ EVs-to-nephrin+ EVs ratio) and increased nephrinuria, both of which correlated with proteinuria. Plasma levels of hemopexin, which were decreased in women with preeclampsia, negatively correlated with proteinuria, urinary podocin+ EVs-to-nephrin+ EVs ratio, and nephrinuria. Administration of HbF to pregnant rabbits increased the number of urinary EVs of podocyte origin. These findings provide evidence that urinary EVs are reflective of preeclampsia-related altered podocyte protein expression. Furthermore, renal injury in preeclampsia associated with an elevated urinary podocin+ EVs-to-nephrin+ EVs ratio and may be mediated by prolonged exposure to cellfree HbF.

Pathophysiology of extracellular haemoglobin : use of animal models to translate molecular mechanisms into clinical significance

The bloods major gas exchange is carried out by haemoglobin, a haeme protein that binds iron and oxygen and can have potentially dangerous side‐effects due to redox reactions. Haemoglobin is a very abundant molecule with a concentration of 150 g/l in whole blood, resulting in almost one kg haemoglobin in an adult human body. Normal turnover of red blood cells results in significant haemoglobin release, and pathological conditions that involve haemolysis can lead to massive haemoglobin levels. To control for the potential threat of extracellular haemoglobin, several protective defence systems have evolved. Many pathological conditions, diseases as well as iatrogenic conditions, such as infusion of haemoglobin‐based oxygen carriers, cerebral intraventricular haemorrhage, extracorporeal circulation and the pregnancy complication pre‐eclampsia, involve abnormal levels of haemolysis and extracellular haemoglobin. Although quite different aetiology, the haemoglobin‐induced damage often causes similar clinical sequelae and symptoms. Here, we will give an overview of the pathophysiological mechanisms of extracellular haemoglobin and its metabolites. Furthermore, we will highlight the use of animal models in advancing the understanding of these mechanisms and discuss how to utilize the knowledge in the development of new and better pharmaceutical therapies.