The effects of added glycerol or unprotected free fatty acids or a combination of the two on silage intake, milk production, rumen fermentation and diet digestibility in cows given grass silage based diets

Selostus: Väkirehuun lisätyn glyserolin tai vapaiden rasvahappojen tai näiden yhdistelmän vaikutus maidontuotantoon ja pötsifermentaatioon ruokittaessa lypsylehmiä säilörehuun perustuvalla ruokinnalla

Free PAPP-A: a Novel Marker in Acute Coronary Syndrome Patients

In recent years, one important objective of cardiovascular research has been to find new markers that would improve the risk stratification and diagnosis of patients presenting with symptoms of acute coronary syndrome (ACS). Pregnancy-associated plasma protein A (PAPP-A) is a large metalloproteinase involved in insulin-like growth factor signalling. It is expressed in various tissues and seems to be involved in many physiological and pathological processes, such as folliculogenesis, bone formation, wound healing, pregnancy and atherosclerosis. The aim of this thesis was to investigate PAPP-A in ACS patients. Circulating concentrations of PAPP-A had been previously shown to be elevated in ACS. In this study it was revealed that the form of PAPP-A causing this elevation was the free noncomplexed PAPP-A. Thus, the form of PAPP-A in the circulation of ACS patients differed from the complexed PAPP-A form abundantly present in the circulation during pregnancy. A point-of-care method based on time-resolved immunofluorometric assays was developed, which enabled the rapid detection of free PAPP-A. The method was found to perform well with serum and heparin plasma samples as well as with heparinized whole blood samples. With this method the concentrations of free PAPP-A in healthy individuals were shown to be negligible. When the clinical performance of the method was evaluated with serum samples from ACS patients, it was shown that the free PAPP-A concentration in the admission sample was an independent predictor of myocardial infarction and death. Moreover, as a prognostic marker, free PAPP-A was revealed to be superior to total PAPPA, i.e. the combination of free and complexed PAPP-A, which has been measured by the other groups in this field. As heparin products are widely used as medication in ACS patients, the effect of heparin products on free PAPP-A molecule and circulating concentrations were also investigated in this study. It was shown that intravenous administration of low molecular weight or unfractionated heparin elicits a rapid release of free PAPP-A into the circulation in haemodialysis patients and patients undergoing angiography. Moreover, the interaction between PAPP-A and heparin was confirmed in gel filtration studies. Importantly, the patients included in the clinical evaluation of the free PAPP-A detection method developed had not received any heparin product medication before the admission sample and thus the results were not affected by the heparin effect. In conclusion, free PAPP-A was identified as a novel marker associated with ACS. The point-of-care methods developed enable rapid detection of this molecule which predicts adverse outcome when measured in the admission sample of ACS patients. However, the effect revealed of heparin products on circulating PAPP-A concentrations should be acknowledged when further studies are conducted related to free or total PAPP-A in ACS.

The Influence of Diet and Microbes on Colonic Immune Regulation and their Implications on Type 1 Diabetes

Dietary and microbial factors are thought to contribute to the rapidly increasing prevalence of T1D in many countries worldwide. The impact of these factors on immune regulation and diabetes development in non-obese diabetic (NOD) mice are investigated in this thesis. Diabetes can be prevented in NOD mice through dietary manipulation. Diet affects the composition of intestinal microbiota, which may subsequently influence intestinal immune homeostasis. However, the specific effects of anti-diabetogenic diets on gut immunity and the explicit associations between intestinal immune disruption and type 1 diabetes onset remain unclear. The research presented herein demonstrates that newly weaned NOD mice suffer from a mild level of colitis, which shifts the colonic immune cell balance towards a proinflammatory status. Several aberrations can also be observed in the peritoneal B cells of NOD mice; an increase in activation marker expression, increased trafficking to the pancreatic lymph nodes and significantly higher antigen presenting cell (APC) efficiency towards insulin-specific T cells. A shift towards inflammation is likewise observed in the colon of germ-free NOD mice, but signs of peritoneal B cell activation are lacking in these mice. Remarkably, most of the abnormalities in the colon, peritoneal macrophages and the peritoneal B cell APC activity of NOD mice are abrogated when NOD mice are maintained on a diabetes-preventive, soy-based diet (ProSobee) from the time of weaning. Dietary and microbial factors hence have a significant impact on colonic immune regulation and peritoneal B cell activation and it is suggested that these factors influence diabetes development in NOD mice.

Moving lipids : ceramides, glycosphingolipids and the glycolipid transfer protein

Lipid movement in cells occurs by a variety of methods. Lipids diffuse freely along the lateral plane of a membrane and can translocate between the lipid leaflets, either spontaneously or with the help of enzymes. Lipid translocation between the different cellular compartments predominantly takes place through vesicular transport. Specialized lipid transport proteins (LTPs) have also emerged as important players in lipid movement, as well as other cellular processes. In this thesis we have studied the glycolipid transport protein (GLTP), a protein that transports glycosphingolipids (GSLs). While the in vitro properties of GLTP have been well characterized, its cell biological role remains elusive. By altering GSL and GLTP levels in cells, we have extracted clues towards the protein's function. Based on the results presented in this thesis and in previous works, we hypothesize that GLTP is involved in the GSL homeostasis in cells. GLTP most likely functions as a transporter or sensor of newly synthesized glucosylceramide (GlcCer), at or near the site of GlcCer synthesis. GLTP also seems to be involved in the synthesis of globotriacylceramide, perhaps in a manner that is similar to that of the fourphosphate adaptor protein 2, another GlcCer-transporting LTP. Additionally, we have developed and studied a novel method of introducing ceramides to cells, using a solvent-free approach. Ceramides are important lipids that are implicated in several cellular functions. Their role as proapoptotic molecules is particularly evident. Ceramides form stable bilayer structures when complexed with cholesterol phosphocholine (CholPC), a large-headgroup sterol. By adding ceramide/CholPC complexes to the growth medium, various chain length ceramides were successfully delivered to cells in culture. The uptake rate was dependent on the chain length of the ceramide, where shorter lipids were internalized more quickly. The rate of uptake also determined how the cells metabolised the ceramides. Faster uptake favored conversion of ceramide to GlcCer, whereas slower delivery resulted mainly in breakdown of the lipid.