Comparison of the in Vitro effect of two-dimensional and three-dimensional polycaprolactone polymers on cell morphology, viability and cytotoxicity

Please read the abstract in the dissertation CopyrightDissertation (MSc)--University of Pretoria, 2010.Physiologyunrestricte

Qualitative assessment of smooth muscle cells propagated on 2D-and 3D-polycaprolactone polymers via scanning electron microscope

Polycaprolactone (PCL) polymers which illustrate both biocompatibility and resorbability for replacement or bulking of damaged or diseased tissue are important in tissue engineering. Cytocompatibilty of these polymers was assessed on two-dimensional PCL disks and threedimensional PCL solid and PCL hollow microspheres using human uterine mixed leiomyosarcoma (SKUT-1) and hamster ductus deferens leiomyosarcoma (CRL-1701) cell lines. Possible PCL cytotoxicity and morphology were investigated in SKUT- and CRL-1701 cells. SKUT cells cultured in disk and microsphere extracts between 24 h and 5 day time periods displayed statistically increased metabolic activity, though activity decreased significantly on 1 month and 1 year extracts. However, the metabolic activity of CRL-1701 cells was similar to controls. Activity increased significantly on the 1 month extracts and decreased significantly on the 1 year extracts. Scanning electron microscopy illustrated increased cell density of cells attached to pre-conditioned disks. After 5 days, cells were spindle-shaped, following microspheres contours indicating high focal adhesion. Both cell lines migrated inside the hollow microspheres, indicating that they benefit from the sheltered environment. This in vitro study suggests that hollow microspheres allow for further cell expansion with a sheltered environment to protect cells from sheer stress experienced in vivo.The Biomaterials and Polymer Division from the Council of Scientific and Industrial Research (Pretoria, South Africa) and the Department of Physiology, University of Pretoria (Pretoria, South Africa).http://www.jstage.jst.go.jp/browse/biomedre

The effects of various n-3 and n-6 polyunsaturated fatty acids on the secretion of insulin like growth factor-I (IGF-I) by MC3T3-E1 osteoblast-like cells

Previous research has shown that some PUFAs have a positive effect on bone, although the cellular mechanism for this is unclear. One such hypothesis is that PUFAs may increase the secretion of IGF-I by osteoblasts. IGFs are autocrine and paracrine stimulators of osteoblasts resulting in increased gene transcription and proliferation. The purpose of this pilot study was to test this hypothesis by treating the cells with various representatives of the PUFA families and examining the effects thereof on secretion of IGF-I. MC3T3-E1 murine osteoblast-like cells were exposed to the n-6 PUFAs arachidonic acid (AA) and gamma-linolenic acid, the n-3 PUFAs eicosapentaenoic acid and docosahexaenoic acid at concentrations of 20ug/ml and a vehicle (ethanol, 0.1%). Prostaglandin E2 (1uM), a product of AA metabolism, as well as parathyroid hormone (0.01uM) were included in the study as positive controls. In some cases indomethacin (1uM) a cyclo-oxygenase blocker was added 45 minutes prior to addition of AA. After 24 hours the conditioned media were harvested and stored at -70 degrees celsius until IGF-I concentrations were determined by means of ELISA. Cell numbers were obtained by means of a standardised crystal violet staining method. Results from this experiment showed that parathyroid hormone increased IGF-I secretion as expected. The PUFAs all stimulated IGF-I secretion with the n-3 PUFAs having the largest effect. PGE2 did not increase IGF-I secretion as expected, this could be due to the fact that the concentration used might not have been optimal for this model. The results obtained support the claims that the n-3 PUFAs have a beneficial effect on osteoblastic IGF-I secretion, however, more experiments need to be conducted in order to verify these results. This work was supported by grants from the National Research Foundation (M.Coetzee) and the Research Development Fund (M.Coetzee).Poster presented at the University of Pretoria Health Sciences Faculty Day, 20 August 2008, Pretoria, South Africa. This poster was also presented at the Physiology Society of Southern Africa (PSSA) Conference (16-19 September 2008) as well as the Suid-Afrikaanse Wetenskap en Kuns Kongres (SAWA) on the 2nd October 2008.Financial support: National Research Foundation and the Researchers Development Progra

Influence of Sutherlandia frutescens extracts on cell numbers, morphology and gene expression in MCF-7 cells

Sutherlandia frutescens is a well-known South African herbal remedy traditionally used for stomach problems, internal cancers, diabetes, various inflammatory conditions and recently to improve the overall health in cancer and HIV/AIDS patients. The influence of crude Sutherlandia frutescens extracts (prepared with 70% ethanol) was investigated on cell numbers, morphology, and gene expression profiles in a MCF-7 human breast adenocarcinoma cell line. Time-dependent (24, 34, 48 and 72 h) and dose-dependent (0.5–2.5 mg/ml) studies were conducted utilizing spectrophotometrical analysis with crystal violet as DNA stain. A statistically significant decrease to 50% of malignant cell numbers was observed after 24 h of exposure to 1.5 mg/ml Sutherlandia frutescens extract when compared to vehicle-treated controls. Morphological characteristics of apoptosis including cytoplasmic shrinking, membrane blebbing and apoptotic bodies were observed after 24 h of exposure. A preliminary global gene expression profile was obtained by means of microarray analysis and revealed valuable information about the molecular mechanisms and signal transduction associated with 70% ethanolic Sutherlandia frutescens extracts.This study was supported by grants from the Struwig-Germishuysen Trust