Badanie zmian chorobowych w modelu mysim cukrzycy z użyciem obrazowania ramanowskiego

Raman imaging was applied to identify biochemical changes associated with the tissue alterations in the type 2 diabetes mellitus (T2DM) murine model compared with the control. The main focus was on the cell nuclei of the inner-most layer of the vessel wall, i.e. endothelium that is considerably affected in the presence of the cardiovascular risk factors. To achieve the aim of the thesis it was necessary to create an appropriate research methodology that was done based on imaging of endothelial cell nuclei (ECN) in the statistically numerous group of hypertensive NO-deficient, hypertensive nitrate-treated and control mice. Finally, the developed approach was applied to identify the biochemical alterations occurring in the endothelial cell nuclei (ECN) of diabetic (db/db) and control (db/+) mice.The Raman chemical images were obtained by integrating bands related to various compounds of the tissue, enabling visualization of the heterogeneity of the studied system. K-means Cluster Analysis of the obtained Raman images enabled the identification of the ECN in the tissue. The analysis of the ECN in the group of hypertension suggests that the biochemical alterations occurring in control, hypertensive NO-deficient and hypertensive nitrate-treated mice are attributed mostly to changes in the conformation of the secondary structure of proteins in ECN. In the diabetes model, the most striking difference between ECN of the diseased and control mice is the ratio between DNA and proteins that is higher for control ECN relatively to the ECN in diabetic animals. Finally, Hierarchical Cluster Analysis (HCA) served to classify data according to their similarities in both studied models, i. e. diabetes and hypertension.Results obtained in this work demonstrate the potential of Raman imaging to study biochemical changes occurring in the endothelial cell nuclei upon vascular risk factors such as hypertension and diabetes.

Celem pracy było wyznaczenie zmian w składzie biochemicznym jąder komórek śródbłonka w tkankach aorty pochodzących od myszy cukrzycowych (db/db) oraz kontrolnych (db+). Pierwszym etapem pracy było stworzenie metodologii do analizy jąder komórek śródbłonka z zastosowaniem spektroskopii ramanowskiej i technik chemometrycznych. Opracowano metodologię na podstawie eksperymentu uwzględniającego trzy grupy myszy: kontrolne, z nadciśnieniem wywołanym brakiem NO oraz myszy leczone azotanem(V) sodu. Następnie zastosowano tą metodologię do indentyfikacji zmian w składzie jąder komórek śródbłonka myszy cukrzycowych i kontrolnych.