Sphingolipids in thyroid eye disease

Graves’ disease (GD) is caused by an autoimmune formation of autoantibodies and autoreactive T-cells against the thyroid stimulating hormone receptor (TSHR). The autoimmune reaction does not only lead to overstimulation of the thyroid gland, but very often also to an immune reaction against antigens within the orbital tissue leading to thyroid eye disease, which is characterized by activation of orbital fibroblasts, orbital generation of adipocytes and myofibroblasts and increased hyaluronan production in the orbit. Thyroid eye disease is the most common extra-thyroidal manifestation of the autoimmune Graves’ disease. Several studies indicate an important role of sphingolipids, in particular the acid sphingomyelinase/ceramide system and sphingosine 1-phosphate in thyroid eye disease. Here, we discuss how the biophysical properties of sphingolipids contribute to cell signaling, in particular in the context of thyroid eye disease. We further review the role of the acid sphingomyelinase/ceramide system in autoimmune diseases and its function in T lymphocytes to provide some novel hypotheses for the pathogenesis of thyroid eye disease and potentially allowing the development of novel treatments

The Role of Sphingolipids and Ceramide in Pulmonary Inflammation in Cystic Fibrosis

Sphingolipids and in particular ceramide have been shown to be critically involved in the response to many receptor-mediated, but also receptor-independent, mainly stress stimuli. Recent studies demonstrate that ceramide plays an important role in the pathogenesis of cystic fibrosis, a hereditary metabolic disorder caused by mutations of the Cystic Fibrosis Transmembrane Conductance Regulator. Patients with cystic fibrosis suffer from chronic pulmonary inflammation and microbial lung infections, in particular with Pseudomonas aeruginosa. Chronic pulmonary inflammation in these patients seems to be the initial pathophysiological event. Inflammation may finally result in the high infection susceptibility of these patients, fibrosis and loss of lung function. Recent studies demonstrated that ceramide accumulates in lungs of cystic fibrosis mice and causes age-dependent pulmonary inflammation as indicated by accumulation of neutrophils and macrophages in the lung and increased pulmonary concentrations of Interleukins 1 and 8, death of bronchial epithelial cells, deposition of DNA in bronchi and high susceptibility to Pseudomonas aeruginosa infections. Genetic or pharmacological inhibition of the acid sphingomyelinase blocks excessive ceramide production in lungs of cystic fibrosis mice and corrects pathological lung findings. First clinical studies confirm that inhibition of the acid sphingomyelinase with small molecules might be a novel strategy to treat patients with cystic fibrosis

Cell-intrinsic ceramides determine T cell function during melanoma progression

Acid sphingomyelinase (Asm) and acid ceramidase (Ac) are parts of the sphingolipid metabolism. Asm hydrolyzes sphingomyelin to ceramide, which is further metabolized to sphingosine by Ac. Ceramide generates ceramide-enriched platforms that are involved in receptor clustering within cellular membranes. However, the impact of cell-intrinsic ceramide on T cell function is not well characterized. By using T cell-specific Asm- or Ac-deficient mice, with reduced or elevated ceramide levels in T cells, we identified ceramide to play a crucial role in T cell function in vitro and in vivo. T cell-specific ablation of Asm in Smpd1fl/fl/Cd4cre/+ (Asm/CD4cre) mice resulted in enhanced tumor progression associated with impaired T cell responses, whereas Asah1fl/fl/Cd4cre/+ (Ac/CD4cre) mice showed reduced tumor growth rates and elevated T cell activation compared to the respective controls upon tumor transplantation. Further in vitro analysis revealed that decreased ceramide content supports CD4+ regulatory T cell differentiation and interferes with cytotoxic activity of CD8+ T cells. In contrast, elevated ceramide concentration in CD8+ T cells from Ac/CD4cre mice was associated with enhanced cytotoxic activity. Strikingly, ceramide co-localized with the T cell receptor (TCR) and CD3 in the membrane of stimulated T cells and phosphorylation of TCR signaling molecules was elevated in Ac-deficient T cells. Hence, our results indicate that modulation of ceramide levels, by interfering with the Asm or Ac activity has an effect on T cell differentiation and function and might therefore represent a novel therapeutic strategy for the treatment of T cell-dependent diseases such as tumorigenesis

The acid ceramidase/ceramide axis controls parasitemia in Plasmodium yoelii-infected mice by regulating erythropoiesis

Acid ceramidase (Ac) is part of the sphingolipid metabolism and responsible for the degradation of ceramide. As bioactive molecule, ceramide is involved in the regulation of many cellular processes. However, the impact of cell-intrinsic Ac activity and ceramide on the course of Plasmodium infection remains elusive. Here, we use Ac-deficient mice with ubiquitously increased ceramide levels to elucidate the role of endogenous Ac activity in a murine malaria model. Interestingly, ablation of Ac leads to alleviated parasitemia associated with decreased T cell responses in the early phase of Plasmodium yoelii infection. Mechanistically, we identified dysregulated erythropoiesis with reduced numbers of reticulocytes, the preferred host cells of P. yoelii, in Ac-deficient mice. Furthermore, we demonstrate that administration of the Ac inhibitor carmofur to wildtype mice has similar effects on P. yoelii infection and erythropoiesis. Notably, therapeutic carmofur treatment after manifestation of P. yoelii infection is efficient in reducing parasitemia. Hence, our results provide evidence for the involvement of Ac and ceramide in controlling P. yoelii infection by regulating red blood cell development

Acid Sphingomyelinase Deficiency Ameliorates Farber Disease

Farber disease is a rare lysosomal storage disorder resulting from acid ceramidase deficiency and subsequent ceramide accumulation. No treatments for Farber disease are clinically available, and affected patients have a severely shortened lifespan. We have recently reported a novel acid ceramidase deficiency model that mirrors the human disease closely. Acid sphingomyelinase is the enzyme that generates ceramide upstream of acid ceramidase in the lysosomes. Using our acid ceramidase deficiency model, we tested if acid sphingomyelinase could be a potential novel therapeutic target for the treatment of Farber disease. A number of functional acid sphingomyelinase inhibitors are clinically available and have been used for decades to treat major depression. Using these as a therapeutic for Farber disease, thus, has the potential to improve central nervous symptoms of the disease as well, something all other treatment options for Farber disease can’t achieve so far. As a proof-of-concept study, we first cross-bred acid ceramidase deficient mice with acid sphingomyelinase deficient mice in order to prevent ceramide accumulation. Double-deficient mice had reduced ceramide accumulation, fewer disease manifestations, and prolonged survival. We next targeted acid sphingomyelinase pharmacologically, to test if these findings would translate to a setting with clinical applicability. Surprisingly, the treatment of acid ceramidase deficient mice with the acid sphingomyelinase inhibitor amitriptyline was toxic to acid ceramidase deficient mice and killed them within a few days of treatment. In conclusion, our study provides the first proof-of-concept that acid sphingomyelinase could be a potential new therapeutic target for Farber disease to reduce disease manifestations and prolong survival. However, we also identified previously unknown toxicity of the functional acid sphingomyelinase inhibitor amitriptyline in the context of Farber disease, strongly cautioning against the use of this substance class for Farber disease patients

Stress induces major depressive disorder by a neutral sphingomyelinase 2-mediated accumulation of ceramide-enriched exosomes in the blood plasma

Major depressive disorder (MDD) is a very common, severe disease with a lifetime prevalence of ~ 10%. The pathogenesis of MDD is unknown and, unfortunately, therapy is often insufficient. We have previously reported that ceramide levels are increased in the blood plasma of patients with MDD and in mice with experimental MDD. Here, we demonstrate that ceramide-enriched exosomes in the blood plasma are increased in mice with stress-induced MDD. Genetic studies reveal that neutral sphingomyelinase 2 is required for the formation of ceramide-enriched exosomes in the blood plasma. Accordingly, induced deficiency of neutral sphingomyelinase 2 prevented mice from the development of stress-induced MDD. Intravenous injection of microparticles from mice with MDD or injection of ceramide-loaded exosomes induced MDD-like behavior in untreated mice, which was abrogated by ex vivo pre-incubation of purified exosomes with anti-ceramide antibodies or ceramidase. Mechanistically, injection of exosomes from mice with MDD or injection of ex vivo ceramide-loaded microparticles inhibited phospholipase D (PLD) in endothelial cells in vitro and in the hippocampus in vivo and thereby decreased phosphatidic acid in the hippocampus, which has been previously shown to mediate MDD by plasma ceramide. In summary, our data indicate that ceramide-enriched exosomes are released by neutral sphingomyelinase 2 into the blood plasma upon stress and mediate stress-induced MDD

MONIKULTTUURINEN KASVATUSKUMPPANUUS VARHAISKASVATUKSESSA : Monikulttuuristen perheiden vanhempien näkemyksiä

Opinnäytetyön tarkoituksena on kuvailla monikulttuuristen perheiden vanhempien kokemuksia ja toiveita kasvatuskumppanuudesta varhaiskasvatuksessa. Opinnäytetyö on kvalitatiivinen ja se on toteutettu teemahaastatteluiden avulla. Haastattelut on tehty yksilöhaastatteluina. Opinnäytetyön toimeksiantajana toimi oululainen päiväkoti. Teoreettinen viitekehys opinnäytetyössä pohjautuu vahvasti kasvatuskumppanuuden periaatteisiin, joita ovat kuuleminen, kunnioitus, luottamus ja dialogi. Lisäksi opinnäytetyössä on teoriaa kulttuurista ja monikulttuurisuudesta. Haastattelurunko sisälsi lähinnä avoimia kysymyksiä, jotka pohjautuivat tietoperustaan. Haastattelut on analysoitu teemoittelun avulla. Teemat olivat samat kuin tietoperustassa. Opinnäytetyön pääteemoja ovat kasvatuskumppanuus ja monikulttuurisuus, jotka sisältävät vielä alateemoja. Tuloksissa nousee esiin pääosin positiiviset kokemukset kasvatuskumppanuudesta varhaiskasvatuksessa. Kasvatuskumppanuus koetaan tärkeänä ja kasvatusvastuuta jaetaan mielellään. Kulttuurieroja ei arjessa huomata juurikaan, mutta perheen kulttuuritausta toivotaan huomioitavan hienovaraisesti, liikaa korostamatta. Johtopäätöksenä voidaan sanoa, että varhaiskasvatukseen ja kasvatuskumppanuuden toteutumiseen ollaan tyytyväisiä.The purpose of this thesis is to describe experiences and wishes of parents of multicultural families about educational partnership in early childhood education. This thesis is qualitative and the material for it was collected with individual theme interviews. The thesis was commissioned by a certain kindergarten from Oulu. The theoretical frame of this thesis includes principles of educational partnership such as hearing, respect, trust and dialog. There is also theory of culture and multiculturalism in this thesis. Open questions of the interview were based on the theoretical frame. The interviews were analyzed by thematic analyzing. Themes were same as in theoretical frame. The main themes of this thesis are educational partnership and multiculturalism which includes subthemes. The results of our thesis show positive experiences in educational partnership and early childhood education. The parents feel the importance of educational partnership and responsibility of raising a child is shared gladly. The parents do not feel cultural differences in everyday life, but there is a wish for sensitive consideration of culture which does not overemphasize the culture. In summary it can be said, that the parents are satisfied with educational partnership and early childhood education

Engineered liposomes sequester bacterial exotoxins and protect from severe invasive infections in mice

Gram-positive bacterial pathogens that secrete cytotoxic pore-forming toxins, such as Staphylococcus aureus and Streptococcus pneumoniae, cause a substantial burden of disease. Inspired by the principles that govern natural toxin-host interactions, we have engineered artificial liposomes that are tailored to effectively compete with host cells for toxin binding. Liposome-bound toxins are unable to lyse mammalian cells in vitro. We use these artificial liposomes as decoy targets to sequester bacterial toxins that are produced during active infection in vivo. Administration of artificial liposomes within 10 h after infection rescues mice from septicemia caused by S. aureus and S. pneumoniae, whereas untreated mice die within 24-33 h. Furthermore, liposomes protect mice against invasive pneumococcal pneumonia. Composed exclusively of naturally occurring lipids, tailored liposomes are not bactericidal and could be used therapeutically either alone or in conjunction with antibiotics to combat bacterial infections and to minimize toxin-induced tissue damage that occurs during bacterial clearance

Antiviral and Anti-Inflammatory Activities of Fluoxetine in a SARS-CoV-2 Infection Mouse Model

The coronavirus disease 2019 (COVID-19) pandemic continues to cause significant morbidity and mortality worldwide. Since a large portion of the world’s population is currently unvaccinated or incompletely vaccinated and has limited access to approved treatments against COVID-19, there is an urgent need to continue research on treatment options, especially those at low cost and which are immediately available to patients, particularly in low- and middle-income countries. Prior in vitro and observational studies have shown that fluoxetine, possibly through its inhibitory effect on the acid sphingomyelinase/ceramide system, could be a promising antiviral and anti-inflammatory treatment against COVID-19. In this report, we evaluated the potential antiviral and anti-inflammatory activities of fluoxetine in a K18-hACE2 mouse model of SARS-CoV-2 infection, and against variants of concern in vitro, i.e., SARS-CoV-2 ancestral strain, Alpha B.1.1.7, Gamma P1, Delta B1.617 and Omicron BA.5. Fluoxetine, administrated after SARS-CoV-2 infection, significantly reduced lung tissue viral titres and expression of several inflammatory markers (i.e., IL-6, TNFα, CCL2 and CXCL10). It also inhibited the replication of all variants of concern in vitro. A modulation of the ceramide system in the lung tissues, as reflected by the increase in the ratio HexCer 16:0/Cer 16:0 in fluoxetine-treated mice, may contribute to explain these effects. Our findings demonstrate the antiviral and anti-inflammatory properties of fluoxetine in a K18-hACE2 mouse model of SARS-CoV-2 infection, and its in vitro antiviral activity against variants of concern, establishing fluoxetine as a very promising candidate for the prevention and treatment of SARS-CoV-2 infection and disease pathogenesis

Potential involvement of the bone marrow in experimental Graves’ disease and thyroid eye disease

IntroductionGraves’ disease is an autoimmune disorder caused by auto-antibodies against the thyroid stimulating hormone receptor (TSHR). Overstimulation of the TSHR induces hyperthyroidism and thyroid eye disease (TED) as the most common extra thyroidal manifestation of Graves’ disease. In TED, the TSHR cross talks with the insulin-like growth factor 1 receptor (IGF-1R) in orbital fibroblasts leading to inflammation, deposition of hyaluronan and adipogenesis. The bone marrow may play an important role in autoimmune diseases, but its role in Graves’ disease and TED is unknown. Here, we investigated whether induction of experimental Graves’ disease and accompanying TED involves bone marrow activation and whether interference with IGF-1R signaling prevents this activation.ResultsImmunization of mice with TSHR resulted in an increase the numbers of CD4-positive T-lymphocytes (p ≤0.0001), which was normalized by linsitinib (p = 0.0029), an increase of CD19-positive B-lymphocytes (p= 0.0018), which was unaffected by linsitinib and a decrease of GR1-positive cells (p= 0.0038), which was prevented by linsitinib (p= 0.0027). In addition, we observed an increase of Sca-1 positive hematopietic stem cells (p= 0.0007) and of stromal cell-derived factor 1 (SDF-1) (p ≤0.0001) after immunization with TSHR which was prevented by linsitinib (Sca-1: p= 0.0008, SDF-1: p ≤0.0001). TSHR-immunization also resulted in upregulation of CCL-5, IL-6 and osteopontin (all p ≤0.0001) and a concomitant decrease of the immune-inhibitory cytokines IL-10 (p= 0.0064) and PGE2 (p ≤0.0001) in the bone marrow (all p≤ 0.0001). Treatment with the IGF-1R antagonist linsitinib blocked these events (all p ≤0.0001). We further demonstrate a down-regulation of arginase-1 expression (p= 0.0005) in the bone marrow in TSHR immunized mice, with a concomitant increase of local arginine (p ≤0.0001). Linsitinib induces an upregulation of arginase-1 resulting in low arginase levels in the bone marrow. Reconstitution of arginine in bone marrow cells in vitro prevented immune-inhibition by linsitinib.ConclusionCollectively, these data indicate that the bone marrow is activated in experimental Graves’ disease and TED, which is prevented by linsitinib. Linsitinib-mediated immune-inhibition is mediated, at least in part, by arginase-1 up-regulation, consumption of arginine and thereby immune inhibition