No description available.

Background: A major problem in the treatment of human hypertension is the late diagnosis of hypertension and, hence, the delayed start of treatment. Very often, hypertension has existed for a long time and cardiac damage has already developed. Therefore, we tested whether lateonset antihypertensive treatment is effective in lowering blood pressure (BP) and in reducing or even preventing left ventricular hypertrophy and fibrosis. Methods: Twenty-one male 60-week-old spontaneously hypertensive rats (SHR) were included. Fourteen rats received oral treatment with captopril (CAP) either as monotherapy or combined with nifedipine (CAP + NIF) over 22 weeks. Seven untreated SHR served as controls. We examined the therapeutic effects on BP, heart weight and histological and biochemical markers of left ventricular remodeling and fibrosis. Results: At 82 weeks of age, BP was reduced in the CAP and CAP + NIF groups by 44 and 51 mmHg, respectively (p < 0.001), but not in untreated controls. Despite the late therapy start, cardiac hypertrophy and fibrosis were attenuated compared to controls. Both treatments reduced heart weight by 1.2 mg/g (25%, p = 0.001) and collagens I and III by 66% and 60%, respectively (p < 0.001), thus proving nearly equivalent cardioprotective efficacy. Conclusion: These data clearly emphasize the benefit of antihypertensive treatment in reducing BP and mitigating the development of cardiac damage even when treatment is started late in life.

TGF-β isoforms are important signalling molecules in wound repair in the skin. Transforming growth factor β3 (TGF-β3) has been implicated in scarless healing. In both animal and human models the application of exogenous TGF-β3 causes a reduction in the inflammatory response and improves the architecture of the neodermis. Research into the influence of TGF-β on scarring has tended to focus on fibroblasts. However, keratinocytes play a major role in scarring both indirectly, as a result of their influence over the behaviour of fibroblasts and also by directly influencing wound contraction. Thus, experiments were carried out to investigate the influence of TGF-β3 on the behaviours of a keratinocyte cell line (HaCaT). Incubation with TGF-β3 increased cell spreading and appeared to reduce cell-surface contacts indicated by both SPR imaging and a detachment assay. TGF-β3 also caused a decreased cell alignment response to microcontact printed protein patterns, in part due to the deposition of laminin which is associated with the TGF-β induced cell migration. There is evidence that TGF-β isoforms differentially influence the outcome of wound healing. Similar to the results produce following addition of exogenous TGF-β3, the neutralisation of TGF-β1 and 2 has been shown to reduce scar formation in the adult wounds. During reepithelialisation keratinocytes experience a dynamic environment. Both extracellular matrix proteins and growth factors influence the progression of wound repair which includes both cell migration and proliferation. Few studies have examined collective cell behaviour in response to TGF-β isoforms and ECM coated substrates. Thus both wound closure and cell proliferation assays were conducted for different ECM proteins fibronectin, laminin and collagen type I and for TGF-β1, 2 and 3. Rates of wound closure were significantly reduced on laminin coated substrates while cell proliferation rates were increased. TGF-β2 and 3 induced significant increases in wound closure rates. This appeared to correspond with an increase in the number of cells independently migrating out from the wound margins. Only TGF-β3 caused a significant decrease in cell proliferation over a 4 day period. Laminin332 deposition is central to the reepithelialisation process and is known to be induced in response to TGF-β. Thus experiments were carried out to investigate HaCaT cell laminin332 deposition in response to TGF-β1, 2 and 3. Both an immunofluorescence staining technique and an ELISA based semi-quantification method was used. Following 4 day incubation all TGF-β isoforms significantly increased laminin332 deposition; however TGF-β2 and 3 caused the most significant increases. Integrin receptors enable cell-matrix interactions during wound repair. TGF-β is known to influence the expression of integrin subunits. Thus, experiments were carried out to compare the influence of each TGF-β isoform on the expression of subunits α3, α2, α5, β1 and β4. All TGF-β isoforms significantly increased all subunit expression. TGF-β3 caused the most significant increase in β4 and both TGF-β2 and 3 caused the most significant increase in α2. While there were differences in cell responses to each isoforms, TGF-β3 did not stand out from the other two isoforms. Interestingly, TGF-β2 shared more similarities with TGF-β3 than it did with TGF-β1, in its role in enhancing wound closure and LN332 deposition. These comparative studies have shown that differences exist in the way TGF-β isoforms influence HaCaT cell behaviour, namely migration, laminin deposition and integrin expression. / EPSRC and DTA grant

Bone repair is modulated by different stimuli. There is evidence that the Transforming Growth Factor-beta (TGF-¿) super-family of cytokines have significant effects on bone structure by regulating the replication and differentiation of chondrocytes, osteoblasts and osteoclasts. There is also significant evidence that interactions with extracellular matrix molecules also influence cell behaviour. This study aimed at determining the role of the TGF-¿s, Collagen type I, Fibronectin and Laminin in bone cell behaviour. To do this MG63 bone cells were used to examine cell adhesion and alignment to different micro-contact printed ECM protein patterns of different widths. The study also aimed at examining how TGF-¿1, 2 and 3 and their solvent and carrier (HCl and BSA, respectively) effected cell surface interactions, cell morphology, cell proliferation and integrin expression. Finally, this study also aimed at examining how the TGF-¿s and their solvent and carrier influenced wound closure in an in vitro wound closure model and how TGF-¿s influence ECM secretion and integrin expression. 5, 10, 25, 50 and 100¿m wide repeat gratings of Collagen type I, Fibronectin and Laminin patterns were stamp patterned onto glass slides and plated with MG63 cells at 50,000 cells per coverslip. Cells on the fibronectin pattern attached and elongated soon after seeding, but did not adhere readily to collagen and laminin and appeared more rounded until 18hrs after seeding. Cells aligned significantly well on the 50¿m and 100¿m wide fibronectin patterned coverslips with mean angles of alignment ~7.87¿ ¿ 3.06SD and 6.45¿ ¿ 5.08SD, respectively, compared to those with smaller width (p<0.001). In comparison, cells aligned less readily to the other two ECM proteins, showing optimal alignments of 9.66¿ ¿ 4.18SD and 14.36¿ ¿ 1.57SD to the 50¿m wide collagen and laminin patterns, respectively. Differences in cell length mirrored those of alignment, with cells acquiring the greatest length when showing the greatest degree of alignment. The results indicate that MG63 cells responded significantly better to 50 and 100¿m wide fibronectin patterns compared to those with smaller width (p<0.001) indicating that the cells may attach mostly via fibronectin specific integrins. Cell surface attachment was examined via a trypsinisation assay in which the time taken to trypsinise cells from the surface provided a means of assessing the strength of attachment. The results indicated that treatment with the solvent (HCl), TGF-¿1, 2 and 3 all decreased cell attachment, but this effect was significantly greater in the case of HCl and TGF-¿3 (p<0.001). However, there were significant differences in trypsinisation rates between HCl and TGF-¿3 (p<0.001). The wound healing response to the TGF-¿s and their solvent/carrier was also investigated in 300¿m ± 10-30¿m SD wide model wounds induced in fully confluent monolayers of MG63 bone cells. The results indicated that TGF-¿3 and HCl significantly enhance wound closure when compared against negative controls, TGF-¿1 and TGF-¿2 treatment (p<0.001). It was also found that TGF-¿1 and TGF-¿2 treatment significantly improved wound closure rate in comparison to the controls (p<0.001). Experiments were performed to determine if the HCl effects on wound closure were dose dependent. Cells were incubated with 20¿M, 40¿M, 80¿M and 160¿M concentrations of HCl prior to wounding and wound closure rates were recorded. Wound closure was dependent on HCl dose with the 80¿M and 160¿M concentrations inducing increases in wound closure rates that were both significantly greater than those induced by 20¿M, 40¿M and control treatments (p<0.001). However, there were significant differences in wound closure between the 80¿M and 160¿M treatment groups after 30hrs of treatment (p<0.001). The effect of different TGF-¿ isomers and their combinations on proliferation rate and cell length of human bone cells were also assessed. The results suggest that cell morphology changes were observed significantly more in cells treated with TGF-¿(2+3) and TGF-¿(1+3) (p<0.001). Any cell treated with TGF-¿1, TGF-¿(1+2) and TGF-¿(1+2+3) showed significantly less elongation compared to the control and other TGF-¿ isomers. In terms of proliferation rate, TGF-¿3 and TGF-¿(2+3) increased cell numbers more than TGF-¿1, TGF-¿2 and other combinations. TGF-¿1 and its combinations did not show significant proliferation and attachment compared to the control due to perhaps its inhibitory effect in contact with human bone cells. Immunostaining indicated that treatment with TGF-¿3 significantly promoted the secretion of collagen type I and anti-human fibronectin in addition to integrin (¿3 and ¿1) expression. Statistically TGF-¿3 and their combinations showed significant differences in number of cells stained for collagen type I, anti-human fibronectin, ¿3 and ¿1integrin. Any cell treated with TGF-¿1 or any combination with TGF-¿1 showed significantly lower cell number stained with the same proteins and integrins (p<0.001). Imaging with WSPR allowed observation of the focal contacts without the need for immunostaining. WSPR images revealed guided cells with high contrast band like structures at the border of cells distal to the edge of guidance cue to which they aligned and with less concentrically formed band like features across the cell body. It is believed that the high contrast features are associated with the formation of focal contacts on the edge of the cells distal to the edge of fibronectin patterns, which suggests that cell guidance is aided by a decrease in cell attachment along a guidance feature. The WSPR experiments also indicated that TGF-¿s influenced the distribution of focal contacts. In the case of TGF-¿1 treated cells the bright high contrast regions were intense but only arranged around the periphery of the cell. In TGF-¿2 and TGF-¿3 cells the bright contrast regions were weaker but again mostly localised around the periphery. These findings supported the earlier trypsinisation results.

No description available.

No description available.

Thesis (M.Sc. Agriculture (Agricultural Economics)) -- University of Limpopo, 2022 / The agricultural sector is dominated by broiler production and it is the key source of animal proteins followed by beef. South Africa is described as a net importer of chicken meat, given that broiler consumption of broiler is greater than supply. Additionally, the South African Poultry industry has raised concerns regarding the high influx of cheap broiler imports into the domestic market. This led to an increase of the ad valorem tariff charged on poultry imports in April 2020 from a previous adjustment in September 2013. It is vital to understand how import tariff changes affect the broiler sector since little is known about how domestic broiler production is responding to an increased tariff. The study, therefore, attempted to determine the relationship between domestic broiler production, import tariff, domestic prices, and import volume. The study aimed to investigate the long-run relationship between import tariff, import quantity, domestic production, and prices (retail and producer) in the broiler sector of South Africa for the period (April 2010 – June 2020). Brazilian frozen chicken imports were selected for study given their relatively high domestic demand in South Africa. In addition, Brazil primarily faces the import tariffs charged by South Africa as the main country of origin for South African imports. The Augmented Dickey-Fuller test, Johansen Cointegration tests, and the Error Correction Model were used as analytical tools in achieving the study objectives. The results for the Augmented Dickey-Fuller test and the Johansen cointegration test showed that all variables were stationary at first difference and cointegrated. The ECM results concluded the existence of a long-run relationship between domestic production, ad valorem tariff, and import volume. As the tariff charged changes, the domestic production increased by 4% in the long run which might be deemed a small advantage in terms of the production scale and therefore not sufficient. The study recommended that strategies that reduce barriers of entry for small-scale farmers such as reduced production costs be implemented to boost domestic production. Lastly, since domestic production is responding positively to tariff adjustment, a thorough investigation is necessary to prove dumping allegations against broiler importers as this will enable the country to impose anti-dumping duties on all countries. / Agri SETA

Microsystems are a broad category of engineered technologies in the micro and nano scale that have a diverse range of applications. They are emerging as a powerful tool in the field of biomedical research, drug discovery, as well as clinical diagnostics and prognostics, especially with regards to cancer. One of the major challenges in precision and personalized medicine in cancer lies in the technical difficulties of ex-vivo cell culture and propagation of the limited number of primary cells derived from patients. Therefore, our aims are to 1. Develop a biologically relevant platform for culturing cancer cells and characterize how it influences the cell growth and phenotype compared to conventional 2-dimensional(2D) cell culturing techniques, 2. Isolate secondary metabolites from endophytic fungi and screen them on the platform for potential anticancer properties in a preliminary drug discovery pipeline, 3. Design and develop biosensors for quantifying cell responses in real-time within these systems. Several biomaterial scaffolds with microscale architectures have been utilized for engineering the tumor extracellular matrix, but very few studies have thoroughly characterized the phenotypic changes in their cell models, which is critical for translational applications of biomaterial systems. The overall objective of these studies is to engineer a biomimetic platform for the culture of breast cancer cells in vitro and to quantify and profile their phenotypic changes. In order to do this, we first evaluated a blank-slate matrix consisting of thiolated collagen, hyaluronic acid and heparin, cross-linked chemically via Michael addition reaction using diacrylate functionalized poly (ethylene glycol). The hydrogel network was used with triple-negative breast cancer cells and showed significant changes in characteristics, with cells self-assembling to form a 3D spheroid morphology, with higher viability, and exhibiting significantly lower cell death upon chemotherapy treatment, as well as had a decrease in proliferation. Furthemore, the transcriptomic changes quantified using RNA-Seq and Next-Gen Sequencing showed the dramatic changes in some of the commonly targeted pathways in cancer therapy. Furthermore, we were able to show the importance of our biomimetic platform in the process of drug discovery using fungal endophytes and their secondary metabolites as the source for potential anticancer molecules. Additionally, we developed gold nanoparticle and antibody-based (ICAM1 and CD11b) sensors to quantify cell responses spatiotemporally on our platform. We were able to show quenching of the green fluorescent fluorophores due to the Förster Resonance Energy Transfer mechanism between the fluorophore and the gold nanometal surface. We also observed antigen-dependent recovery of fluorescence and inhibition of energy transfer upon the antibody binding to the cell-surface receptors. Future efforts are directed towards incorporating the hydrogel system with antigen-dependent sensors in a conceptually-designed microfluidic platform to spatiotemporally quantify the expression of surface proteins in various cells of the tumor stroma. This includes the migration,infiltration, and polarization of specific immune cells. This approach will provide further insight into the heterogeneity of cells at the single-cell resolution in defined spaces within the 3D microfluidic platform. / Doctor of Philosophy / Microsystems are a broad category of engineered technologies in the micro and nano scale that have a diverse range of applications. They are emerging as a powerful tool in the field of biomedical research, drug discovery, as well as clinical diagnostics and prognostics, especially with regards to cancer. However, a major challenge in being able to offer personalized medicine to cancer patients comes from the difficulty of growing cells from the patient's tumor biopsy in a laboratory for further screening and analysis. There are also limited resources available for real-time expression of proteins on cell-surfaces, that could be potential biomarkers and targets for treatment. Various natural and synthetic polymers are biocompatible and have been used widely in engineering the tumor extracellular matrix. However, the effect of hydrogels derived from these polymers on the specific tumor cells are not always well characterized. Our studies explore the influence of a biohybrid hydrogel on breast cancer cells and our results show that the microscale architecture of the hydrogel platform works as a suitable scaffold for recapitulating the 3-dimensional(3D) breast tumor microenvironment, and can also be employed in the drug discovery process. Additionally, we developed a nano-scale biosensor to enable the quantification of specific cell-surface proteins in real-time. Ongoing and future efforts are focused on designing and fabricating a microfluidic device with precise control over the design of space and special chambers for cell culture. These will be used for studying interactions of various cells in the tumor microenvironment that influence cancer progression. Integrating these micro-scale systems, including sensors will allow researchers to quantify cell behavior in response to the variable factors they are exposed to, as well as provide insight to answer fundamental questions about cancer biology that are limited by the conventional 2D cell culture systems.

Mechanical interactions of cells with their immediately surrounding extracellular matrix (ECM) is now known to be critical in pathophysiology. For example, during cancer progression, while uncontrollable cell division leads to tumor formation, the subsequent metastatic migration of cells from the primary tumor site to distant parts of the body causes most cancer-related deaths. The metastatic journey requires cells to be able to adopt different shapes and move persistently through the highly fibrous native ECM, thereby requiring significant spatiotemporal reorganization of the cell cytoskeleton. While numerous studies performed on flat 2-dimensional culture platforms and physiological 3D gels have elucidated cytoskeletal reorganization, our understanding on how cells adapt to natural fibrous microenvironments and regulate their behavior in response to specific ECM biophysical cues including fiber size, spacing, alignment and stiffness remains in infancy. Here, we utilize the non -electrospinning Spinneret tunable engineered parameters (STEP) technique to manufacture ECM mimicking suspended fibrous matrices with precisely controlled fiber diameters, network architecture, inter-fiber spacing and structural stiffness to advance our fundamental understanding of how external cues affect cytoskeleton-based cellular forces in 3-distinct morphological processes of the cell cycle starting from division to spreading and migration. Mechanobiological insights from these studies are implemented to deliver intracellular cargo inside cells using electrical fields. Holistically, we conclude that fibrous environments elicit multiple new cell behaviors never before reported. Specifically, our new findings include (i) design of fiber networks regulates actin networks and cell forces to sculpt nuclei in varying shapes: compressed ovals, tear drop, and invaginations, and drive the nuclear translocation of transcription factors like YAP/TAZ. In all these shapes, nuclei remain rupture-free, thus demonstrating the unique adaptability of cells to fibers, (ii) dense crosshatch networks are fertile environments for persistent 1D migration in 3D shapes of rounded nuclei and low density of actin networks, while sparse fiber networks induce 2D random migration in flattened shapes and well-defined actin stress fibers, (iii) actin retraction fiber-based stability regulates mitotic errors. Cells undergoing mitosis on single fibers exhibit significant 3D movement, and those attached to two fibers can have rotated mitotic machinery, both conditions contributing to erroneous division, and (iv) a bi-phasic force response to electroporation that coincides with actin cytoskeleton remodeling. Cells on suspended fibers can withstand higher electric field abuse, which opens opportunities to deliver cargo of varying sizes inside the cell. Taken altogether, our findings provide new mechanobiological understanding of cell-fiber interactions at high spatiotemporal resolution impacting cell migration, division and nuclear mechanics-key behaviors in the study of pathophysiology. / Doctor of Philosophy / Cancer, one of the major pathophysiological conditions, progresses within the living body through spreading of malignant cells from the primary tumor to distant secondary sites, ultimately leading to life-ending outcomes. Such spreading of cancer also known as cancer metastasis requires mechanical interactions of cells with their immediately surrounding microenvironment or the extracellular matrix (ECM). Cells utilize their cytoskeleton, a dynamic internal network of filamentous proteins, to adopt various morphologies, exert mechanical forces and physically remodel their local environment as they navigate through the highly fibrous native ECM. While previous research has elucidated how biochemical factors and bulk matrix properties regulate such cytoskeletal organization and single cell behavior, our understanding of how cells adapt to fibrous environments and respond to local biophysical cues like fiber diameter, spacing, alignment and stiffness remains in infancy. Here we use the non -electrospinning Spinneret tunable engineered parameters (STEP) to generate suspended nanofiber networks of tunable geometric and mechanical properties to mimic the native cellular environment. We discover that cells elongated within these ECM-mimicking environments utilize a unique cytoskeletal caging structure to regulate the shape and response of their nuclei in a fiber -diameter and organization-dependent manner. Additionally, we demonstrate that these elongated cell morphologies often observed during metastatic cancer cell movements, is achievable not only in aligned fibers but can also be induced by dense networks of fibers in a crossing organization. Specifically, such dense crosshatch networks allow cells to migrate persistently at high speeds while cells on sparsely spaced networks demonstrate slower and random movements. As cells elongated during interphase rounded up to undergo division, we find that the underlying fiber-geometry modulates mitotic dynamics through differential levels of actin retraction fiber-mediated stability, leading to significant alterations in orientation of mitotic machinery and mitotic spindle defects. Finally, we utilize these mechanobiological insights on cytoskeletal organization and cell shape control to optimize intracellular delivery of cargo using high-voltage electric fields. We demonstrate suspended cells are capable of withstanding higher electric fields and identify multistage cell contractility recovery dynamics, which correlate with cytoskeletal disruption and reassembly. Taken altogether, our findings provide a comprehensive understanding of the fibrous ECM-mediated regulation of the cytoskeletal organization and its impact in cell migration, division and nuclear mechanics. Knowledge obtained from this study will improve our understanding of cancer metastasis and provide predictive data for in vivo cellular response, essential for cytoskeleton-targeting cancer therapies.

Previous research has shown that Taiwan's military spending is affected either by China's military buildup or the US's military pipeline. This study investigates whether it is also true an ongoing US-China relationship has dynamic effects. Three major findings are obtained from the statistical analyses. First and foremost, the level of US-China conflict has a contemporaneous positive effect on Taiwan's military spending. Second, the analyses also indicate that the volatility of US-China relations has negative effects on Taiwan's military spending. This finding suggests that instability in US-China relations will prompt Taiwan to decrease its military spending due to a higher amount of perceived security on the one hand, and Taiwan wants to avoid further provoking China on the other. Third, analyses indicate that an error correction model fares better than a simple budgetary incremental model in explaining the re-equilibrating effects of GNP growth on Taiwan's military spending. Overall, the results demonstrate the interplay of domestic and international constraints and may help to predict what will be the expected military spending when Taiwan's economy changes. I suggest that Taiwan's military spending is likely to be influenced by US-China relations as well as by foreign investment and domestic economic constraints as long as the United States policy toward the Taiwan problem remains unchanged.