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    Methyl pyruvate protects a normal lung fibroblast cell line from irinotecan-induced cell death: Potential use as adjunctive to chemotherapy
    (Public Library of Science, 2017-08) Monchusi, B.; Ntwasa, M.
    The Warburg Effect, characterized by increased rate of glycolysis even under normoxic conditions, is one of the hallmarks of cancer. Relatively lower oxidative phosphorylation (OXPHOS) is also a characteristic feature in cancer cells. We hypothesized that interference with this phenomenon, by introducing exogenous pyruvate, would upset this cancer phenotype and boost the energy requirements of normal cells. We find that methyl pyruvate protects irinotecan-treated normal lung fibroblast cell line (MRC-5) probably by turning off the p53/p21 axis of the apoptotic pathways. When the MRC-5 fibroblasts recover in drug-free medium, the intrinsic apoptotic pathway is also turned off and the cells survive with no discernible exponential growth during the observation period. In contrast, the mere introduction of exogenous pyruvate kills the lung cancer cell line (A549). Although, functional p53 is important in the drug-induced cancer cell death, it is probably not essential because cancer cell lines with mutated p53 also die albeit less efficiently. We conclude that methyl pyruvate may preferentially kill cancer cells and protect normal cells during chemotherapy.
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    In Vitro Inhibition of Angiogenesis by Antibodies Directed against the 37kDa/67kDa Laminin Receptor
    (Public Library of Science, 2013-03-12) Khusal, R.; Da Costa Dias, B; Moodley, K.; Penny, C.; Reusch, U.; Knackmuss, S.; Little, M.; Weiss, S.F.T.
    The 37kDa/67kDa laminin receptor (LRP/LR) is a central receptor mediating interactions between tumour cells and the basement membrane and is thereby a key player in adhesion and invasion, essential processes in metastatic cancer. To affect continued tumour growth, tumours induce angiogenesis for the constant delivery of nutrients and oxygen. This study aims to determine the blocking effect of the anti-LRP/LR specific antibody, W3 on the angiogenic potential of HUVE (human umbilical vein endothelial) cells. Flow cytometric analysis revealed that 97% of HUVE cells display cell surface LRP/LR. An angiogenesis assay was conducted employing HUVE cells seeded on the basement membrane reconstituent Matrigel™ supplemented with the pro-angiogenic factor vascular endothelial growth factor (VEGF). Post 18h incubation at 37°C tubular structures, namely tube lengths were assessed. Treatment of established tubular structures with 100 μg/ml anti-LRP/LR specific antibody completely blocked angiogenesis. Our findings suggest a central role of the 37kDa/67kDa LRP/LR in tube formation and recommends anti-LRP/LR specific antibodies as potential therapeutic tools for treatment of tumour angiogenesis.
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    Downregulation of the Non-Integrin Laminin Receptor Reduces Cellular Viability by Inducing Apoptosis in Lung and Cervical Cancer Cells.
    (Public Library of Science., 2013-03-05) Moodley, K.; Weiss, S.F.T.
    The non-integrin laminin receptor, here designated the 37-kDa/67-kDa laminin receptor (LRP/LR), is involved in many physiologically relevant processes, as well as numerous pathological conditions. The overexpression of LRP/LR on various cancerous cell lines plays critical roles in tumour metastasis and angiogenesis. This study investigated whether LRP/LR is implicated in the maintenance of cellular viability in lung and cervical cancer cell lines. Here we show a significant reduction in cellular viability in the aforementioned cell lines as a result of the siRNA-mediated downregulation of LRP. This reduction in cellular viability is due to increased apoptotic processes, reflected by the loss of nuclear integrity and the significant increase in the activity of caspase-3. These results indicate that LRP/LR is involved in the maintenance of cellular viability in tumorigenic lung and cervix uteri cells through the blockage of apoptosis. Knockdown of LRP/LR by siRNA might represent an alternative therapeutic strategy for the treatment of lung and cervical cancer.
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    Assessing Global Transcriptome Changes in Response to South African Cassava Mosaic Virus [ZA-99] Infection in Susceptible Arabidopsis thaliana.
    (Public Library of Science., 2013-06-27) Pierce, E.J.; Rey, M.E.C.
    In susceptible plant hosts, co-evolution has favoured viral strategies to evade host defenses and utilize resources to their own benefit. The degree of manipulation of host gene expression is dependent on host-virus specificity and certain abiotic factors. In order to gain insight into global transcriptome changes for a geminivirus pathosystem, South African cassava mosaic virus [ZA:99] and Arabidopsis thaliana, 4×44K Agilent microarrays were adopted. After normalization, a log2 fold change filtering of data (p<0.05) identified 1,743 differentially expressed genes in apical leaf tissue. A significant increase in differential gene expression over time correlated with an increase in SACMV accumulation, as virus copies were 5-fold higher at 24 dpi and 6-fold higher at 36 dpi than at 14 dpi. Many altered transcripts were primarily involved in stress and defense responses, phytohormone signalling pathways, cellular transport, cell-cycle regulation, transcription, oxidation-reduction, and other metabolic processes. Only forty-one genes (2.3%) were shown to be continuously expressed across the infection period, indicating that the majority of genes were transient and unique to a particular time point during infection. A significant number of pathogen-responsive genes were suppressed during the late stages of pathogenesis, while during active systemic infection (14 to 24 dpi), there was an increase in up-regulated genes in several GO functional categories. An adaptive response was initiated to divert energy from growth-related processes to defense, leading to disruption of normal biological host processes. Similarities in cell-cycle regulation correlated between SACMV and Cabbage leaf curl virus (CaLCuV), but differences were also evident. Differences in gene expression between the two geminiviruses clearly demonstrated that, while some global transcriptome responses are generally common in plant virus infections, temporal host-specific interactions are required for successful geminivirus infection. To our knowledge this is the first geminivirus microarray study identifying global differentially expressed transcripts at 3 time points.
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    High resolution imaging study of interactions between the 37 kDa/67 kDa laminin receptor and APP, beta-secretase and gamma-secretase in Alzheimer's disease.
    (Public Library of Science., 2014-06-27) Jovanovic, K.; Loos, B.; Da Costa Dias, B.; Penny, C.; Weiss, S.F.T.
    Alzheimer's disease (AD) is the most prevalent form of dementia affecting the elderly. Neurodegeneration is caused by the amyloid beta (Aβ) peptide which is generated from the sequential proteolytic cleavage of the Amyloid Precursor Protein (APP) by the β- and γ- secretases. Previous reports revealed that the 37 kDa/67 kDa laminin receptor (LRP/LR) is involved in APP processing, however, the exact mechanism by which this occurs remains largely unclear. This study sought to assess whether LRP/LR interacted with APP, β- or γ-secretase. Detailed confocal microscopy revealed that LRP/LR showed a strong co-localisation with APP, β- and γ-secretase, respectively, at various sub-cellular locations. Superresolution Structured Illumination Microscopy (SR-SIM) showed that interactions were unlikely between LRP/LR and APP and β-secretase, respectively, while there was strong co-localisation between LRP/LR and γ-secretase at this 80 nm resolution. FRET was further employed to assess the possibility of protein-protein interactions and only an interaction between LRP/LR and γ-secretase was found. FLAG co-immunoprecipitation confirmed these findings as LRP/LR co-immunoprecipitated with γ-secretase, but failed to do so with APP. These findings indicate that LRP/LR exerts its influence on Aβ shedding via a direct interaction with the γ-secretase and possibly an indirect interaction with the β-secretase.