Browsing by Author "Keet, Lana"

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    Development of in vitro models to investigate the anti-inflammatory properties of Cyclopia Maculata and other South African herbal teas : a comparative study
    (Stellenbosch : Stellenbosch University, 2015-04) Keet, Lana; Gelderblom, W. C. A.; Riedel, S.; Swart, Amanda C.; Stellenbosch University. Faculty of Science. Dept. of Biochemistry.
    ENGLISH ABSTRACT: Chronic inflammation is suggested to contribute to cancer development and therefore a potential target for chemoprevention. In the skin, keratinocytes and macrophages play an integral part in acute and chronic inflammation, with interleukin 1-α (IL-1α) and tumor necrosis factor α (TNF-α) as key cytokines governing this process. Green tea (Camellia sinensis) and the South African herbal teas, rooibos (Aspalathus linearis) and honeybush (Cyclopia spp.) displayed antiinflammatory effects in mouse and human skin. To further investigate the antiinflammatory properties of green tea and the herbal teas, rooibos and honeybush (C. subternata and C. maculata) herbal teas, suitable cell culture models were developed and validated utilising human keratinocytes (HaCaT) and monocyte (THP- 1) derived macrophages. Aqueous extracts of the green tea and unfermented herbal teas were prepared and their chemical composition determined by high performance liquid chromatography (HPLC) and the antioxidant activity characterised utilising different antioxidant assays. Green tea and rooibos exhibited similar antioxidant activities while C. maculata displayed the lowest overall antioxidant activity of all the extracts, despite possessing the highest mangiferin level, the major polyphenol in honeybush. The modulation of cytokine release was studied in (i) an UVB-induced pre-exposure HaCaT model monitoring the accumulation of IL-1α and (ii) a LPS stimulated THP-1 macrophage model monitoring the TNF-α release, utilising both a pre-exposure and co-exposure extract regimens. In the pre-exposure HaCaT inflammatory model the UVB-induced IL-1α was decreased by the green tea extract while a far weaker response was obtained with the rooibos extract. Both the honeybush extracts displayed a significant effect in the reduction of IL-1α with C. subternata exhibiting a slight increased protection at a lower extract concentration. In the pre-exposure THP-1 derived macrophage model, green tea and the herbal tea extracts inhibited TNF-α release in a dose dependent manner in the absence of an overt loss in cell viability and apoptosis at lower extract concentrations, suggesting a typical anti-inflammatory effect. In the co-exposure model, the different extracts also exhibited an anti-inflammatory effect at the lowest concentrations in the absence of apoptosis while at higher extract concentrations the effect was masked by a decrease in cell viability and increased apoptosis. C. maculata exhibit differential effects when considering the inhibition of cytokine production and, depending on the cell model, either exhibited a weaker or stronger effect when compared to C. subternata and rooibos. Phenolic diversity of the different teas is likely to explain the differential effects in the antioxidant assays and cell culture models with respect to the regulation of the production of the inflammatory markers. Proposed mechanism for the anti-inflammatory effects include the modulation of oxidative stress via various pathways and the subsequent down regulation of nuclear factor kappa β (NFκB) and activated protein-1 (AP-1) which are key regulators of cytokine production governing the inflammatory response.
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    Modulation of UVB-induced inflammation and oxidative stress by unfermented rooibos and honeybush aqueous extracts: A comparative study
    (Stellenbosch University, 2024-12) Keet, Lana; Lilly, Mariska; Louw, Ann; Abel, Stefan; Stellenbosch University. Faculty of Science. Dept. of Biochemistry.
    Exposure to Ultraviolet B (UVB) radiation induces various biological responses that may contribute to skin cancer development. Natural compounds such as tea polyphenols have demonstrated protective effects against UVB-induced damage. Rooibos (Aspalathus linearis) and honeybush (Cyclopia spp.) are two South African herbal plants that have gained attention for their potential health benefits. Rooibos has recently been registered by the European Commission in its lists of Protected Designations of Origin (PDO) and Protected Geographical Indication (PGI) and therefore has protected status as it is only allowed to be grown in the Cederberg area of the Western Cape, South Africa. The current study explores the preventive effects of unfermented rooibos and honeybush herbal tea aqueous extracts on oxidative stress, inflammation and phospholipid alterations in HaCaT human keratinocytes exposed to UVB. The study utilises a pre-exposure HaCaT model, where cells are treated with tea extracts prior to UVB exposure. An array of assays and techniques were utilized to determine the antioxidant and anti-inflammatory effects on specific cytokines, genes and proteins involved in these processes, as well as ion mobility-mass spectrometry (IM-MS) to investigate effects on the major keratinocyte phospholipids. HPLC analysis showed the presence of the dihydrochalcones, aspalathin and nothofagin, the flavones, isoorientin and orientin, as well as the flavonols, rutin, isoquercitrin, and hyperoside in the rooibos extract. The major compounds found in honeybush extract included the xanthones, mangiferin and iso-mangiferin, and the flavanone, hesperidin. Rooibos demonstrated superior free radical scavenging activity against ABTS and DPPH, antioxidant capacity (ORAC), ferric iron reducing potential (FRAP) and greater inhibition of iron-induced peroxidation (LPO) compared to honeybush. Both teas also exhibited cell-based antioxidant properties by increasing glutathione (GSH) levels, and the increase in GPX1 and CAT expression, while only rooibos upregulated Nrf2 expression. Microarray analysis confirmed the antioxidant activity of both teas in the increased expression of GCLM, GPX and PRDX. The results indicate that honeybush reduces UVB-induced interleukin (IL)-1α accumulation and IL-6 secretion, while both teas decrease IL-8 secretion and NF-κB activation, demonstrating the anti-inflammatory effect of rooibos and honeybush. The treatment of keratinocytes with rooibos and honeybush alone did not affect the levels of any of the lipid species identified in the current study. However, exposure of the cells to UVB significantly decreased the levels of all of evaluated phospholipids. Neither rooibos nor honeybush were able to significantly counteract this decrease in phospholipid levels, with honeybush even further decreasing PC 34:1, PC 36:1, PC 36:4, PE 38:4 and SM 34:1 levels. Rooibos and honeybush contain completely different polyphenol profiles, which is reflected in their activities, with rooibos displaying more antioxidant activity while honeybush has greater anti-inflammatory activity. This research has the potential to inform the production of sunscreen products utilizing natural extracts to protect against the harmful effects of UVB exposure.