Browsing by Subject "CARS"
Now showing items 1-3 of 3
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Continuous Grading of Early Fibrosis in NAFLD Using Label-Free Imaging : A Proof-of- Concept Study (Helsingin yliopisto, 2016)Maksan rasvoittumisen ajatellaan joskus olevan vain metabolisen oireyhtymän hyvälaatuinen maksamanifestaatio, vaikka se johtaa, erityisesti maksan fibrotisoituessa, kasvaneeseen kardiovaskulaarikuolleisuuteen ja pahimmillaan maksasyöpään. Tästä kansanterveydellisesti merkittävästä roolista huolimatta rasvamaksa on alidiagnosoitu ja sen patogeneettiset mekanismit ovat huonosti ymmärrettyjä, molemmat osin hyvien diagnostisten työkalujen puutteesta johtuen. Päätimme selvittää uusien non-lineaaristen kuvantamismodaliteettien soveltuvuutta rasvamaksan varhaisessa diagnostiikassa. Selvitimme 32 lihavuusleikkauspotilaan maksakudoskoepalojen kudosarkkitehtuurin perinteisin kudosvärjäysmenetelmin ja vertasimme tuloksia samojen potilaiden non-lineaarisesti kuvannettuihin näytteisiin. Kuvansimme minimaalisesti käsitellyistä jääleikkeistä coherent anti-Stokes Raman scattering (CARS) ja second harmonic generation (SHG) signaalit, joita hyödyntäen arvioimme näytteiden rasva- ja kollageenipitoisuuden. Kehitimme myös kuva-analytiikka-algoritmin, joka mahdollistaa kollageenisignaalin automatisoidun ja objektiivisen arvioimisen. Näytteistä mittaamamme SHG- ja CARS-signaalit korreloivat hyvin patologien arvioihin näytteiden fibrotisoitumis- ja rasvoittumisasteista, minkä lisäksi automatisoitu algoritmimme erotteli luotettavasti jopa varhaisen fibroosin normaalista kudoksesta. Myös terveiksi luokiteltujen näytteiden maksakudoksen keskellä esiintyi hienoa fibrillaarista SHG-signaalia. Tunnistamamme SHG-signaali oli spesifiä kolokalisoituen hyvin kollageeni-I- ja III-vasta-ainevärjäyksien kanssa. Tuloksemme osoittavat, että multimodaalisten kuvantamismenetelmien avulla voidaan tunnistaa maksan fibrillaarista kollageenia perinteisiä kudostutkimusmenetelmiä herkemmin. Kehittämämme kuva-analyysi-algoritmi puolestaan arvioi varhaisen fibroosin määrän nopeasti, objektiivisesti ja jatkuvalla skaalalla. Yhdessä nämä löydökset luovat edellytykset tutkia esimerkiksi maksan vähäisen fibroosin kliinistä merkitystä ja paranemistaipumusta isommissa potilasaineistoissa.
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(Helsingfors universitet, 2014)The aim of this thesis was to investigate the potential of label-free CARS microscopy as a new method for chemically-specific imaging of live cells and particle-cell interactions in a drug delivery context. Cells used to mimic the intestinal epithelium, Caco-2 cells and HT-29 cells and nano-/ microcrystal particle interactions with macrophages were studied. More information about drug absorption from intestinal and particle cell interactions are needed, since many novel drugs lack properties needed for good bioavailability. It would be beneficial if these events could be visualized without labels. CARS microscopy was found to be well suited to imaging live Caco-2 and HT-29 cells that were grown on PTFE Transwell inserts. CARS microscopy revealed lipid droplets inside these cells. The size of lipid droplets increased in Caco-2 cells a lot during a three week period so that at the end a large part of the inner part of the cell was filled with lipid droplets. It was also observed that Caco-2 cells and HT-29 cells can grow on top of each other on Transwell inserts and not just as a monolayer. These two facts could cause variations in drug absorption studies based on Caco-2 cell monolayers. CARS microscopy was able to detect nanocrystals as small as about 500 nm with label-free, molecular-specific CARS microscope inside RAW 264.7 macrophages after incubation of 120 min. This observation was important, since nanocrystal drug formulations are gaining interest in the field of pharmacy. Nanocrystals can be used in parenteral drug formulations as well as in oral dosage forms. In suspensions, nanocrystals can be used to cause long lasting drug release. Nanocrystals can be also used to enhance poor bioavailability of drugs. Whether these nanocrystals are used in parenteral formulations or in oral drug formulations it is evident that imaging techniques are needed to image interactions between these nanocrystals and cells. CARS microscopy could be one of those techniques, since it is suitable for live cell imaging and it can be used to image nanocrystals that are not labeled. The results in this thesis suggest that CARS microscopy could be used as fast imaging technique for nanocrystal particle cell interactions. Overall, CARS microscopy is a relatively new imaging method that shows much promise as a label-free chemically specific imaging technique for imaging cells and cell-particle interactions in a drug delivery context. As the technique becomes more widely available and undergoes some technical developments, it will become much more widespread imaging method in the future.
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(2018)Co-amorphous mixtures have rarely been formulated as oral dosage forms, even though they have been shown to stabilize amorphous drugs in the solid state and enhance the dissolution properties of poorly soluble drugs. In the present study we formulated tablets consisting of either spray dried co-amorphous ibuprofen-arginine or indomethacin-arginine, mannitol or xylitol and polyvinylpyrrolidone K30 (PVP). Experimental design was used for the selection of tablet compositions, and the effect of tablet composition on tablet characteristics was modelled. Multimodal non-linear imaging, including coherent anti-Stokes Raman scattering (CARS) and sum frequency/second harmonic generation (SFG/SHG) microscopies, as well as scanning electron microscopy, X-ray diffractometry and Fourier-transform infrared spectroscopy were utilized to characterize the tablets. The tablets possessed sufficient strength, but modelling produced no clear evidence about the compaction characteristics of co-amorphous salts. However, co-amorphous drug-arginine mixtures resulted in enhanced dissolution behaviour, and the PVP in the tableting mixture stabilized the supersaturation. The co-amorphous mixtures were physically stable during compaction, but the excipient selection affected the long term stability of the ibuprofen-arginine mixture. CARS and SFG/SHG proved feasible techniques in imaging the component distribution on the tablet surfaces, but possibly due to the limited imaging area, recrystallization detected with xray diffraction was not detected.
