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Additional file 1: Appendix S1. Supplementary methods. Table S1. Metabolic and anthropometric parameters of the randomized participants before and after weight loss. Table S2. BMI [mean and 95% CI] and estimated mean differences within intention to treat analysis. Table S3. Frequency of impaired glucose metabolism before (T-3) and after (T0) weight loss. Table S4. Change of HOMA-IR and glucoseAUC [mean and 95% CI] compared to baseline within per-protocol analysis. Table S5. HRQoL of the participants and effects of weight loss. Table S6. Physical component summary [PCS] score [mean and 95% CI] and estimated mean differences within per-protocol analysis. Table S7. Mental component summary [MCS] score [mean and 95% CI] and estimated mean differences within per-protocol analysis. Table S8. Independent association of NPR-C after weight loss with weight regain at month 48 (ΔBMIT0T48). Table S9. Independent association of NPR-C after weight loss with FM regain at month 48 (ΔFMT0T48). Figure S1. Flow chart of the randomized controlled trial. f/m indicates female/male. BMI during the randomized controlled trial.

Additional file 1. Electronic supplementary material.

Additional file 2. PRISMA checklist.

T cells represent a central component of the human adaptive immune system. Among them, CD4+ T helper cells and CD8+ cytotoxic T cells can be distinguished which either mediate the actions of other immune cells or kill affected cells, respectively. Their proper function is dependent on an appropriate surrounding micromilieu, which classically includes other cell types as well as chemokines and cytokines. Interestingly, recent findings suggest that also the osmolarity of the microenvironment may have an influence. Since osmolarity is substantially higher in lymphatic tissue and other immunologically active compartments than in plasma, I investigated what kind of alterations this hypertonicity can evoke in different effector T cell species. Due to their relevance in autoimmune and tumor diseases, Th17 and CD8+ T cells have been in the focus of my studies. To investigate phenotypical and metabolic changes, Th17 cells and CD8+ T cells have been cultured and stimulated in hypertonic medium and subsequently analyzed. Additionally, shRNA and molecular inhibitors were utilized to uncover possible underlying molecular pathways. Consequently, to analyze functional alterations in hypertonicity, CD8+ T cells specific for MART-1 were purified and their cytolytic capability towards target cells presenting MART-1 was evaluated in a hypertonic environment. Upon activation under hypertonic conditions, the Th17 phenotype was significantly augmented within the Th17 cell population, though this was going hand in hand with induction of anti-inflammatory markers such as FOXP3 and TGF-β expression. I was able to illustrate that both effects were mediated via the p38 MAP kinase pathway. Moreover, cytotoxic T cells showed significant increase of their cytolytic capability in hypertonic medium. It correlated with enhanced expression of granzyme B, perforin and TNF-α as well as alternative cytolytic molecules such as TRAIL. Similarly, the cytotoxic T cells demonstrated augmented cell activation and cell metabolism in a hyperosmotic microenvironment. Concurrence of the upregulation of pro- and anti-inflammatory features in Th17 cell subsets in a hypertonic microenvironment indicates a relevant role in Th17-mediated autoimmune diseases. Moreover, enhanced cytotoxic activity of CD8+ T cells in higher osmolarity may represent a vital part of the defense system protecting the human body from pathogens and preventing tumor growth. These findings may bring about therapeutic innovations in the highly investigated field of adoptive T cell therapies, making the deployed CD8+ T cells even more effective. Lastly, considering osmolarity when investigating T cell functionality will become essential in future to obtain reliable, relevant and translational findings., T-Zellen stellen einen zentralen Bestandteil des Immunsystems des menschlichen Körpers dar. Man unterscheidet CD4+ T-Helferzellen und CD8+ zytotoxische T-Zellen, deren jeweilige Funktionen vom jeweils umgebenden Mikromilieu beeinflusst werden, das sich klassischerweise aus anderen Zellen, Pathogenen und Zytokinen zusammensetzt. Neue Erkenntnisse zeigen jedoch, dass auch die Osmolarität der Umgebung einen Einfluss auf die Zellen haben kann. Die Osmolarität in lymphatischem Gewebe und anderen immunologisch aktiven Kompartimenten ist signifikant höher als im Blutplasma. Daher habe ich die Veränderungen untersucht, die diese Hyperosmolarität in verschiedenen Effektor-T-Zellen hervorrufen. Im Fokus meiner Untersuchungen standen Th17- und zytotoxische CD8+ T-Zellen, da sie im Rahmen von Autoimmun- und Tumorerkrankungen eine klinisch relevante Rolle spielen. Zur Untersuchung phänotypischer und metabolischer Veränderungen wurden Th17- und CD8+ T-Zellen in hypertonischem Medium kultiviert, stimuliert und anschließend analysiert. Der zugrundeliegende Mechanismus wurde mithilfe von shRNA sowie molekularen Inhibitoren untersucht. Zur Analyse funktioneller Veränderungen wurden MART-1-spezifische CD8+ T-Zellen isoliert, deren zytolytisches Potenzial anschließend in Zytotoxizitäts¬tests untersucht wurde. Th17-Zellen zeigten einen verstärkten Th17-Phänotyp sowie antiinflammatorische Eigenschaften wie FOXP3- und TGF-β-Expression, wenn sie unter hypertonischen Bedingungen aktiviert wurden. Beide Veränderungen wurden durch den p38-MAP-Kinase-Signalweg einschließlich NFAT5 und SGK1 reguliert. Hingegen zeigten zytotoxische T-Zellen in hypertonischem Medium signifikant verbesserte zytolytische Fähigkeiten, die durch verstärkte Expression von Granzyme B, Perforin, und TNF-α sowie weiterer zytolytischer Moleküle wie TRAIL vermittelt wurden. Ebenso wiesen zytotoxische T-Zellen auch eine gesteigerte Aktivierung und erhöhte Stoffwechselleistung unter hypertonischen Bedingungen auf. Die Verstärkung sowohl von pro- als auch antiinflammatorischen Charakteristiken in Th17-Zellen in hypertonischer Umgebung deutet auf eine mögliche Relevanz dieser Ergebnisse in der Pathogenese von Th17-vermittelten Autoimmunerkrankungen hin. Die gesteigerte Aktivität von CD8+ T-Zellen in hypertonischer Umgebung könnte Teil der gesunden Immunabwehr sein, die den menschlichen Körper vor Pathogenen oder bösartigem Tumorwachstum schützt. Darüber hinaus ergeben sich aus diesen Ergebnissen möglicherweise praktische Anwendungen auf dem Gebiet der adoptiven T-Zelltherapie. Letztlich zeigen meine Ergebnisse, dass die Berücksichtigung von osmotischen Faktoren bei Untersuchungen zu T-Zellfunktionen relevant ist.

Additional file 1 Table S1. Search strategies.

We report on a case of very rare autosomal recessive cholesteryl ester storage disease due to lysosomal acid lipase deficiency (LALD). LALD is caused by mutations in the lysosomal acid lipase A (LIPA) gene resulting in cholesteryl ester accumulation in liver, spleen, and macrophages. It can lead to liver failure, accelerated atherosclerosis and premature death. Until recently, treatment options were limited to lipid-lowering medications to control dyslipidemia. Presently, a long-term enzyme replacement therapy with Sebelipase alfa, a recombinant human lysosomal acid lipase, is available for patients with LALD.Our patient's condition became conspicuous at the age of two due to a xanthogranuloma of the chin together with increased lipid levels, elevated liver enzymes and hepatomegaly. It took another five years until our patient was diagnosed with LALD after genetic testing. A bi-weekly therapy with intravenous Sebelipase alfa was started at the age of 26 years. It led to normalization of lipid levels, reduction of liver enzymes and beginning regression of hepatomegaly in the absence of adverse drug reactions after 46 infusions.Since LALD can take a fatal course even in patients with a long-term stable condition, it is essential to identify affected patients early and to treat them appropriately by enzyme replacement therapy. LALD should be suspected in patients with low high-density lipoprotein cholesterol (HDL-C) and high low-density lipoprotein cholesterol (LDL-C) in conjunction with elevated liver enzymes or hepatomegaly.A registry for LALD patients shall help to advance our understanding of the disease as well as improve patient care (NCT01633489). Keywords: Lysosomal acid lipase deficiency (LALD), Cholesteryl ester storage disease, Sebelipase alfa (Kanuma™), LALD registry