Regulation of the flow of fatty acids (FA) by futile cycle (TAG/FA cycle) in white adipose tissue (WAT) is an important mechanism of controlling metabolism of FA and therefore its regulation is in the interest of research as a possible therapeutic target in the treatment of obesity and insulin resistance. The study of the murine model suggests that the treatment of the n-3 PUFA with mild caloric restriction (CR) probably induces the TAG/FA cycle mainly in epididymal depot. It also reduces inflammation in WAT and the risk of cardiovascular diseases. We focused on monitoring the gene expression encoding key enzymes of the TAG/FA cycle (PEPCK, ATGL, HSL, DGAT1 and DGAT2) in dorzolumbar (DL), gonadal (GON) and mesenteric (MEZ) depot in a murine model C57BL/6 using qPCR. We were interested in the influence of the composition of fats in the diet, the influence of administration of n-3 PUFA with combination of 10% CR and the difference of gene expression among depots of WAT. The results indicate that the supplementation of high-fat diet with n-3 PUFA contributes mainly to reduction of gene expression for DGAT2 and the combination of n-3 PUFA and CR increases expression of genes influencing the TAG/FA cycle. In MEZ unlike GON and DL fat depot were no significant differences in gene expression, while the different...

Acclimation to cold or hardening is known for many decades through its beneficial effects on human health. In contrast, sudden exposure to cold, cold shock, is a great risk of cerebral and cardiac injury, especially in the elderly. There is very little published data on the cellular and molecular mechanisms induced by cold adaptation in heart and brain. The aim of this work was to describe and compare different properties heart, liver, brain and brown adipose tissue mitochondria of rats housed at 25 ± 1 řC and at mild cold (9 ± 1 řC, 5 weeks). The high-resolution oxygraphy, spectrophotometry and Western blotting analyses were used. We found differences in the respiratory control between the heart and liver. Cold acclimation decreased activity of the Krebs cycle enzymes. Fatty acid contribution to the respiration reached the maximum in brown fat and the minimum in the hippocampus. However, further study is necessary.

The prevalence of obesity has already epidemic dimensions. Recently, the obesogens have been identified as the main cause in addition to excessive food intake, the lack of physical activity and the genetic background. These substances damage the metabolic processes, interfere with the hormonal functions and impair the energy balance in behalf of gaining weight and obesity. The theoretical part of this work deals with obesity, adipose tissue, lipid droplet and obesogens. From the obesogens there is closely specified a group of persistent organic pollutants (POP) from which one representative was used in the practical part of this work. The aim of the practical part was to describe the cellular model of differentiation the mesenchymal stem cell into adipocytes and to investigate the effect of one of the most frequently occurring obesogen on the expression genes of lipid metabolism and insulin signalling pathway. The morphological changes were observed in cells during differentiation (at days 0, 4, 10 and 21). The mesenchymal cells of the elongated spindle shape changed into adipocytes filled with lipid droplets. Oil Red O staining was used for quantification of accumulated lipids. The differentiation to adipocytes was confirmed by fluorescence immunocytochemistry using a specific protein FABP4. The...

Das globale Phänomen Übergewicht erhöht das Risiko für die Entwicklung von Diabetes, Atherosklerose und Herz-Kreislauf-Erkrankungen. Diese sind assoziiert mit der Expression des Transmembranproteins Glycoprotein nonmetastatic melanoma protein b (Gpnmb), das von Makrophagen und dendritischen Zellen exprimiert wird. Wir haben die Rolle von Gpnmb in genetisch- und diät-induzierter Atherosklerose sowie in diät-induzierter Adipositas in Gpnmb-Knockout- und Wildtyp-Mäusen untersucht. Körpergewicht und Blutfette wurden in beiden Erkrankungen nicht von Gpnmb beeinflusst. Gpnmb wurde in Makrophagen von atherosklerotischen Läsionen stark exprimiert, jedoch hatte das Fehlen von Gpnmb keinen Einfluss auf die Größe der Aortenläsion. In Übergewicht konnten wir dagegen einen größeren Effekt von Gpnmb detektieren. Gpnmb hatte einen positiven Einfluss auf den Insulin- und Glukoseplasmaspiegel sowie auf die Leberfibrose bei adipösen Mäusen. Gpnmb-Knockout-Tiere besaßen mehr Makrophagen im epididymalen Fettgewebe. Da Gpnmb in den entsprechenden Makrophagen von Wildtyp-Mäusen stark exprimiert wurde, könnte Gpnmb eine abschwächende Rolle auf Entzündungen des Fettgewebes haben. Dies wird durch in-vitro-Daten bestätigt, wo Gpnmb hauptsächlich in reparativen, TGFβ-stimulierten Makrophagen exprimiert wurde. Diese Expression führte jedoch nur zu einer leichten entzündungshemmenden Wirkung. Eine weitere Aufgabe von Makrophagen, die Autophagie, wurde durch Gpnmb nicht beeinflusst. Dies ist überraschend, da die Expression, die Freisetzung und der Abbau von Gpnmb durch Bafilomycin, einem Inhibitor des letzten Schritts der Autophagie, auf einzigartige Weise erhöht wurde. Zusammenfassend ist die Gpnmb-Expression in voll ausgereiften Makrophagen stark induziert und kann durch lysosomale Hemmung weiter gesteigert werden. Bei Adipositas verhindert Gpnmb die Entwicklung einer Insulinresistenz, möglicherweise durch Dämpfung der Entzündung des Fettgewebes. / Obesity, an emerging global phenomenon, increases the risk for the development of diabetes, atherosclerosis and cardiovascular diseases. Those metabolic diseases have been associated with the expression of glycoprotein nonmetastatic melanoma protein b (Gpnmb), a transmembrane protein that is expressed by macrophages and dendritic cells. We studied the role of Gpnmb in genetically- and diet-induced atherosclerosis as well as diet-induced obesity in Gpnmb-knockout and respective wildtype control mice. The absence of Gpnmb did not affect body weight and blood lipid parameters in both diseases. Whereas Gpnmb was strongly expressed in atherosclerotic lesion-associated macrophages, the absence of Gpnmb did not influence the development of aortic lesion size. On the other hand, the absence of Gpnmb elicited stronger effects in obesity. We observed a positive influence of Gpnmb on insulin and glucose plasma levels as well as liver fibrosis in obese mice. Moreover, Gpnmb-knockout animals contained more macrophages in epididymal adipose tissue. Gpnmb was strongly expressed in adipose tissue macrophages in wildtype mice, suggesting an alleviating role of Gpnmb on adipose tissue inflammation. This was corroborated by in vitro data where Gpnmb was mostly expressed in reparative macrophages stimulated with transforming growth factor β (TGFβ). However, this expression resulted only in a mild anti-inflammatory effect. Another important macrophage feature, autophagy, was not influenced by Gpnmb. This is surprising as Gpnmb expression, shedding and degradation was uniquely increased by bafilomycin, an inhibitor of the last step of autophagy. Taken together, Gpnmb expression is strongly induced in fully mature macrophages and can be further increased due to lysosomal inhibition. In obesity, Gpnmb prevents the development of insulin resistance possibly by dampening adipose tissue inflammation.

The diploma thesis deals the problematics of weight reduction with the using of bariatric- metabolic surgery and focuses on the changing risk components of the metabolic syndrome during one year long observation of 45 probands who have undergone Partial Jejuno-Ileal Diversion, Laparoscopic Sleeve Gastrectomy or Laparoscopic Gastric Plication. The main aim of the diploma thesis is to evaluate the changing risk components of the metabolic syndrome during weight reduction after undergoing bariatric surgery. The thesis shows that in the studied sample of bariatric patients it is an effective method of weight reduction (in PJID the success rate was 48 % EWL, in LGCP 51 % EWL and the most successful was LSG with 76 % EWL) with metabolic effect such as for example observed positive changes in risk components of the metabolic syndrome - reduction of morning glucose levels, increase of HDL cholesterol and decrease of triacylglycerols in the blood, decrease of waist circumferences and decrease of blood pressure or elimination of metabolic syndrome. Up to 68.9 % of the monitored probands showed signs of metabolic syndrome when evaluating the initial measurement before bariatric surgery, the remaining 22.2 % of the probands showed the signs after the year's observation. As part of the risk assessment for the...

The prevention and treatment of obesity is a major problem of health care systems in affluent societies. Metabolism of adipose tissue belongs to the therapeutical targets, since accumulation of adipose tissue is the basis of obesity development. Experiments using transgenic mice with ectopic expression of brown- fat uncoupling protein 1 (UCP1) in white adipose tissue (WAT), verified a concept that obesity could be ameliorated by increasing energy expenditure in WAT. The goal of the experiments of this PhD Thesis was to characterize in detail the phenotype of this unique animal model of obesity resistance. We have shown that mitochondrial uncoupling in WAT resulted in increased oxidation of fatty acids (FA), in face of decreased lipogenesis and induced mitochondrial biogenesis in this tissue. In further studies, we aimed to modulate propensity to obesity be increasing FA oxidation in WAT in response to physiological stimuli. This could be accomplished in response to the combination treatment using n-3 polyunsaturated fatty acids (n-3 PUFA) and mild calorie restriction in mice fed high-fat diet. Synergistic induction of mitochondrial oxidative capacity and lipid catabolism in epididymal WAT was associated with suppression of low-grade inflammation of WAT, which is typical for obesity. The improvement of lipid...

Mesenchymale Stromazellen (MSCs) werden beim Pferd bereits mit vielversprechenden Ergebnissen zur Behandlung von muskuloskelettalen Erkrankungen, insbesondere von Sehnenerkrankungen, eingesetzt. In bisherigen klinischen Studien lag das Hauptaugenmerk auf der Behandlung von Erkrankungen der Oberflächlichen Beugesehne bei Rennpferden, die jedoch in Deutschland nur einen verhältnismäßig kleinen Anteil des Patientenaufkommens darstellen. Die zu erwartenden Ergebnisse nach MSC-Behandlung von Fesselträgererkrankungen sind dagegen noch nicht bekannt. Darüber hinaus sind die grundlegenden Kenntnisse zur Biologie equiner MSCs noch unzureichend, was Verständnis und Optimierung des bestehenden Therapiekonzeptes erschwert. Häufig wird die Verwendung alternativer Gewebequellen für MSCs diskutiert, wobei jedoch nur wenige vergleichende Daten zu den jeweiligen zellulären Eigenschaften vorliegen. Ziel dieser Arbeit war es daher, zum einen mehr Kenntnisse über die zu erwartenden klinischen Ergebnisse nach MSC-Behandlung von Sehnenerkrankungen zu erlangen, einschließlich Erkrankungen des Fesselträgers, zum anderen den Wissensstand hinsichtlich der in-vitro-Charakterisierung equiner MSCs zu erweitern, wobei ein Vergleich klinisch relevanter Charakteristika zwischen MSCs aus verschiedenen Gewebequellen angestrebt wurde. In die klinische Studie wurden 98 Pferde, die aufgrund von Sehnen- und Banderkrankungen mit MSCs behandelt worden waren, einbezogen. Von 58 dieser Tiere konnten Langzeitergebnisse nach einem Beobachtungszeitraum von mindestens einem Jahr erhoben werden. Diese wurden hinsichtlich des Behandlungserfolges sowie möglicher Einflussfaktoren ausgewertet, wobei die Behandlung als erfolgreich bewertet wurde, wenn die Patienten nach dem Beobachtungszeitraum voll trainiert oder im Sport eingesetzt werden konnten und dabei kein Rezidiv aufgetreten war. Die Behandlung mit MSCs wurde bei 84,5 % der Pferde als erfolgreich eingestuft, wobei Erkrankungen der Oberflächlichen Beugesehne mit 84,2 % und Erkrankungen des Fesselträgers mit 83,3 % gleichermaßen gute Ergebnisse zeigten. Tendenziell beeinflussten Nutzungsdisziplin, Erkrankungsstadium und Patientenalter das klinische Ergebnis ebenso wie bei konventioneller Behandlung. Insgesamt war nach MSC-Behandlung das Auftreten von Rezidiven deutlich seltener zu beobachten als in der Literatur für die konventionelle Behandlung beschrieben wird. Für die in-vitro-Studie zur vergleichenden Charakterisierung equiner MSCs aus verschiedenen Quellen wurden Knochenmark, Fett- und Sehnengewebe sowie Nabelschnurblut und -gewebe gewonnen. Aus diesen Proben wurden jeweils die plastikadhärenten MSCs isoliert und hinsichtlich Zellausbeute, Proliferations- und Migrationseigenschaften, tripotentem Differenzierungspotential sowie der Expression der Sehnenmarker Kollagen 1A2 und Skleraxis vergleichend untersucht. Die Ausbeute an MSCs war bei allen soliden Geweben (Fett-, Sehnen-, und Nabelschnurgewebe) hochsignifikant höher (p < 0,001). Ebenso proliferierten MSCs aus Fett- und Sehnengewebe signifi-kant schneller als MSCs aus Knochenmark oder Nabelschnurblut (p < 0,01). Von letzteren wurden darüber hinaus etwa drei viertel aller Zellkulturen vor der achten Passage seneszent. Das höchste Migrationspotential zeigten wiederum MSCs aus Sehnen- und Fettgewebe, wobei hier MSCs aus Nabelschnurgewebe das ungünstigste Ergebnis erzielten (p < 0,01). Die adipogene Differenzierung gelang bei MSCs aus allen Quellen vergleichbar gut. Bei der osteogenen Differenzierung erreichten MSCs aus Knochenmark das beste Ergebnis, während MSCs aus Nabelschnurblut und –gewebe nur schwach osteogen differenzierten (Tag 21: p < 0,01; Tag 35: p < 0,05). Im Gegensatz dazu erreichten MSCs aus Nabelschnurblut bei der chondrogenen Differenzierung die meisten Scorepunkte, MSCs aus Knochenmark dagegen die wenigsten (p < 0,05). Kollagen 1A2 wurde von MSCs aus Fettgewebe am höchsten exprimiert, Skleraxis von MSCs aus Nabelschnurblut. MSCs aus Sehnengewebe exprimierten beide Sehnenmarker auf fast ebenso hohem Level. MSCs aus Knochenmark dagegen zeigten hier jeweils die niedrigste Expression (p < 0,05 für Kollagen 1A2). Basierend auf den Ergebnissen der klinischen Studie ist die MSC-Therapie nach wie vor als vielversprechende Behandlungsoption für Sehnenerkrankungen anzusehen und ist auch für die Behandlung von Fesselträgererkrankungen geeignet. Zukünftige, kontrollierte klinische Studien müssen jedoch die Wirksamkeit der MSC-Therapie noch weitergehend bestätigen. Die in-vitro-Studie zeigte signifikante Unterschiede zwischen equinen MSCs aus verschiedenen Quellen auf, die bei der Auswahl einer Gewebequelle für die MSC-Isolierung für klinische Anwendungen berücksichtigt werden sollten. MSCs aus Fettgewebe erscheinen aufgrund ihrer sehr guten Proliferations- und zuverlässigen Differenzierungseigenschaften als eine gute Alternative zu MSCs aus Knochenmark für autologe Therapien. MSCs aus Sehnengewebe sind den hier vorliegenden Ergebnissen zufolge besonders gut für die Behandlung von Sehnenerkrankungen geeignet; vor einer routinemäßigen Anwendung dieser MSCs sollten jedoch ihre Eigenschaften weiterführend untersucht werden. / In horses, mesenchymal stromal cells (MSCs) are used for the treatment of musculoskeletal diseases, especially tendon injuries, with promising results. Previous clinical studies mainly focused on the treatment of superficial digital flexor tendon injuries in racehorses, which, however, represent only a relatively small percentage of the overall equine case load in Germany. Average outcome to be expected following MSC treatment of suspensory ligament injuries was not yet determined. Moreover, basic knowledge on equine MSC biology is still deficient, hampering the understanding and thus the optimisation of the existing treatment regime. The use of alternative MSC sources is frequently discussed, yet to date, only few data comparing the cellular properties of equine MSCs from different sources have been published. The aim of this study was, on the one hand, to gain more knowledge concerning the expected outcome after MSC treatment of tendon injuries, including injuries to the suspensory ligament. On the other hand, it was aimed at expanding the knowledge on equine MSC characterisation in vitro, thereby focusing on the comparison of clinically relevant properties of MSCs derived from different sources. In the clinical study, 98 horses were included, all of which had received MSC treatment for tendon or ligament injuries. In 58 of these horses, long term results after a follow-up period of at least one year could be collected. These data were analysed with respect to treatment outcome and potential influencing factors. Treatment was considered successful when horses were back to full training or competition after the follow-up period, without having suffered a re-injury. The overall success rate was 84.5 %. Success rates in horses suffering from superficial digital flexor tendon injuries and in horses suffering from suspensory ligament injuries were comparably good (84.2 % and 83.3 %, respectively). Similar to conventional therapies, the sports discipline in which the horses performed, age and disease stage tended to influence the outcome. Overall, re-injury rates after MSC treatment were considerably lower than those described in the literature following conventional treatment. For the comparative characterisation of MSCs from different sources in vitro, samples of bone marrow, adipose and tendon tissue, as well as umbilical cord blood and –tissue were collected. Plastic-adherent MSCs were isolated out of these samples and comparatively characterised focusing on cell yields, proliferation and migration properties, trilineage differentiation potential and the expression of the tendon markers collagen 1A2 and scleraxis. MSC yields were significantly higher in all solid tissues (adipose, tendon and umbilical cord tissue) (p < 0.001). Further, MSCs from adipose and tendon tissue proliferated significantly faster than MSCs from bone marrow or umbilical cord blood (p < 0.01). Moreover, approximately three quarters of the samples derived from the latter sources underwent senescence before reaching passage eight. The highest migration potential was found in MSCs derived from tendon and adipose tissue again, while MSCs from umbilical cord tissue showed the least (p < 0.01). The adipogenic differentiation potential was comparably good in MSCs from all different sources. The osteogenic differentiation was most distinct in MSCs from bone marrow, while MSCs from umbilical cord blood and tissue showed only weak evidence of differentiation (day 21: p < 0.01; day 35: p < 0.05). In contrast, following chondrogenic differentiation, MSCs from umbilical cord blood scored highest and MSCs from bone marrow scored lowest (p < 0.05). Collagen 1A2 was most highly expressed in MSCs from adipose tissue, highest scleraxis expression levels were found in MSCs from umbilical cord blood. MSCs from tendon tissue, however, expressed both markers at almost evenly high levels. Contrastingly, lowest expression levels of both markers were found in MSCs derived from bone marrow (p < 0.05 for collagen 1A2). Based on the results of the clinical study, MSC therapy can still be considered a very promising treatment option for tendon diseases and is also a suitable treatment for suspensory ligament injuries. In the future, controlled clinical studies will have to further confirm the efficacy of this treatment regime. The in-vitro-study showed significant differences between equine MSCs derived from different sources, which should be considered when choosing a MSC source for clinical applications. For autologous therapies, MSCs derived from adipose tissue appear to be a good alternative to MSCs derived from bone marrow, due to their remarkable proliferation and reliable differentiation capacities. Furthermore, according to this study, MSCs derived from tendon tissue are especially suitable for treating tendon injuries. Prior to routine clinical applicability of these MSCs, however, their properties should be further investigated.

Cellular cardiomyoplasty is a cell therapy using stem cells or progenitor cells for myocardial regeneration to improve cardiac function and mitigate heart failure. Since we first published cellular cardiomyoplasty in 1989, this procedure became the innovative method to treat damaged myocardium other than heart transplantation. A significant improvement in cardiac function, metabolism, and perfusion is generally observed in experimental and clinical studies, but the improvement is mild and incomplete. Although safety, feasibility, and efficacy have been well documented for the procedure, the beneficial mechanisms remain unclear and optimization of the procedure requires further study. This chapter briefly reviews the stem cells used for cellular cardiomyoplasty and their clinical outcomes with possible improvements in future studies.

Cellular cardiomyoplasty is a cell therapy using stem cells or progenitor cells for myocardial regeneration to improve cardiac function and mitigate heart failure. Since we first published cellular cardiomyoplasty in 1989, this procedure became the innovative method to treat damaged myocardium other than heart transplantation. A significant improvement in cardiac function, metabolism, and perfusion is generally observed in experimental and clinical studies, but the improvement is mild and incomplete. Although safety, feasibility, and efficacy have been well documented for the procedure, the beneficial mechanisms remain unclear and optimization of the procedure requires further study. This chapter briefly reviews the stem cells used for cellular cardiomyoplasty and their clinical outcomes with possible improvements in future studies.

Cardiovascular disease is the main cause of death in the United States. Although it is recognized that moderate intensity long-term exercise can decrease the chances of dying from cardiovascular disease by favorably modifying risk factors such as hypertension, obesity, hyperlipidemia, and insulin resistance, physical activity also enhances longevity by mechanisms independent of these risk factors. This review briefly summarizes what is known about the inflammatory nature of atherosclerosis and how long-term aerobic exercise can reduce the atherogenic activity of endothelial cells, blood mononuclear cells, and adipose tissue.