Differential effects of Sutherlandia frutescens subs. microphylla on cell numbers, morphology, gene and protein expression in a breast adenocarcinoma and a normal breast epithelial cell line

Show simple item record

dc.contributor.advisor Joubert, Fourie en
dc.contributor.advisor Joubert, Annie M. en
dc.contributor.postgraduate Stander, Barend Andre en
dc.date.accessioned 2013-09-07T09:57:13Z
dc.date.available 2008-08-12 en
dc.date.available 2013-09-07T09:57:13Z
dc.date.created 2008-04-23 en
dc.date.issued 2007 en
dc.date.submitted 2008-08-05 en
dc.description Dissertation (MSc)--University of Pretoria, 2007. en
dc.description.abstract Sutherlandia frutescens is a South African herbal remedy traditionally used for various ailments and lately to improve the overall health in cancer and HIV/AIDS patients. Relatively little is known about the mechanisms of action of the constituents present in S. frutescens. The aim of this project was to examine the in vitro influence of crude ethanolic S. frutescens extracts in human breast adenocarcinoma (MCF-7) and non-tumorigenic breast epithelial (MCF-12A) cells after 48 h of exposure. Dose-dependent studies were conducted on cell numbers and metabolic activity by means of spectrophotometry. Morphological changes were determined with light-, fluorescent- and transmission electron microscopy (TEM). Cell cycle progression and apoptosis were analyzed using flow cytometry. The differential effects of S. frutescens extracts on gene expression levels in both the MCF-7 and MCF-12A cells were conducted utilizing micro array analysis. mTOR kinase activity was measured with an ELISA assay. S. frutescens reduced cell proliferation in both the non-tumorigenic MCF-12A and the tumorigenic MCF-7 cell line in a dose-dependent manner. The tumorigenic MCF-7 cells were more susceptible to S. frutescens treatment compared to the non-tumorigenic MCF-12A cells. Morphological characteristics of apoptosis and autophagy, including cytoplasmic shrinking, membrane blebbing and an increase in autophagic vacuoles were observed in both cell lines with the MCF-7 cells being more susceptible to autophagy and the MCF-12A cells less susceptible to autophagy and apoptotic cell death. TEM confirmed ultrastructural characteristics of autophagy in both cell lines. Flow cytometry revealed a G2/M arrest with no increase in apoptosis in MCF-7 cells and a G2/M arrest with an increase in apoptosis in MCF-12A cells treated with 1.5mg/ml S. frutescens extract. Microarray analyses revealed 325 statistically significantly differentially expressed genes in MCF-7 cells and 1467 genes in MCF-12A cells. The majority of S. frutescens-treated genes were down-regulated when compared to the vehicle-treated control in both cell lines. Several genes involved in DNA replication and repair were differentially expressed in response to S. frutescens exposure. These include Poly (ADP-ribose) polymerase family, member 2 (PARP-2) (down-regulated in both cell lines), PCNA (down-regulated in MCF-7 cells) and growth arrest and DNA-damage-inducible beta (GADD45B) (up¬regulated in MCF-12A cells). This suggests that abrogated expression of genes involved in DNA replication and repair play a role in inducing a G2/M cell cycle arrest in S. frutescens-treated cells. ELISA analysis of the mTOR kinase revealed a decrease in mTOR kinase activity in both cell lines after S. frutescens exposure. Therefore, attenuated mTOR kinase activity as a result of S. frutescens treatment in both cell lines is regarded as a central mediator in inducing autophagy suppressing gene expression and inhibiting ribosome biogenesis. Understanding of in vitro molecular mechanisms of S. frutescens enables researchers to focus on affected cellular mechanisms and identify active compounds with subsequent evaluation as possible candidates for use in anticancer therapy. The current study contributes to the unraveling of the in vitro molecular mechanisms and signal transduction associated with 70% ethanolic S. frutescens extracts, providing a basis for further research on this multi-purpose medicinal plant in Southern Africa. en
dc.description.availability unrestricted en
dc.description.department Physiology en
dc.identifier.citation Stander, BA 2007, Differential effects of Sutherlandia frutescens subs. microphylla on cell numbers, morphology, gene and protein expression in a breast adenocarcinoma and a normal breast epithelial cell line, MSc dissertation, University of Pretoria, Pretoria, viewed yymmdd < http://hdl.handle.net/2263/27037 > en
dc.identifier.other E962/ag en
dc.identifier.upetdurl http://upetd.up.ac.za/thesis/available/etd-08052008-093345/ en
dc.identifier.uri http://hdl.handle.net/2263/27037
dc.language.iso en
dc.publisher University of Pretoria en_ZA
dc.rights © 2007, University of Pretoria. All rights reserved. The copyright in this work vests in the University of Pretoria. No part of this work may be reproduced or transmitted in any form or by any means, without the prior written permission of the University of Pretoria. en
dc.subject Micro array en
dc.subject Mtor kinase en
dc.subject Cell cycle arrest en
dc.subject Autophagy en
dc.subject Sutherlandia frutescens en
dc.subject Anticancer en
dc.subject UCTD en_US
dc.title Differential effects of Sutherlandia frutescens subs. microphylla on cell numbers, morphology, gene and protein expression in a breast adenocarcinoma and a normal breast epithelial cell line en
dc.type Dissertation en


Files in this item

This item appears in the following Collection(s)

Show simple item record