Browsing by Author "Williams, Ricquelle Daphne"

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    The antimycobacterial activity of phytocannabinoids
    (Stellenbosch : Stellenbosch University, 2023-02) Williams, Ricquelle Daphne; Mavumengwana, Vuyo; Loxton, Andre; Smith, Liezel; Allie, Nasiema; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. Molecular Biology and Human Genetics.
    ENGLISH ABSTRACT: Tuberculosis (TB) is a deadly and communicable disease that is caused by the bacterium, Mycobacterium tuberculosis (M.tb). M.tb is skilled at manipulating and evading hostdefense mechanisms by alveolar macrophages, allowing its survival and replication intracellularly. Since the current treatment regimen makes use of antibiotics, the everincreasing number of multi-drug resistant (MDR) M.tb strains has resulted in a need for research into alternative options. Although most anti-TB drug discovery strategies target the actual pathogen, slow pace in discovering new antimycobacterials is testament to the need to change strategies. Host directed therapy (HDT) is an intervention where the eradication of the intracellular pathogen is mediated by the host immune response modulated by small molecules. Cannabis sativa L. (C. sativa) is valued for its psychoactive potentials and varied ethnobotanical medicinal properties due to a plethora of bioactive constituents. In addition to the plant’s utility as a treasure trove for medicinal applications, this research study aims to present a case for the use of small molecules derived from C. sativa as an alternative HDT against TB. We aimed to evaluate the antimycobacterial effect of crude extracts of two C. sativa plants, one grown outdoor (C. sativa plant 1 or P1) and one grown under controlled indoor conditions (C. sativa plant 2 or P2) and evaluated their bioactive organic extracts activity in THP-1 macrophages infected with mycobacteria. In addition, we isolated endophytic fungi from C. sativa and evaluated their antimycobacterial activity and whether they produce similar compounds to the host plant. Herein, it was demonstrated that the dichloromethane (DCM) extract of C. sativa plant 1 (DP1) and the methanol and ethyl acetate extracts of C. sativa plant 2 (MP2 and EP2) stimulated THP-1 macrophages in the killing of Mycobacterium smegmatis (M. smegmatis) mc2155 compared to untreated macrophages. In addition, the methanol extract of C. sativa plant 2 (MP2) displayed the best activity with a percentage survival of 14.31% (p = 0.000009) at 6-hours post treatment; 2.63% at 12-hours post treatment (p = 0.0001) and 0% survival at 24-hours post treatment (p = 0.0005). The metabolite profile showed that these three extracts (DP1, MP2 and EP2) share three compounds, cannabinol (CBN), cannabigerol (CBG), and cannabielsoin (CBE), which could be the cause of macrophage stimulation. However, MP2, which showed the best activity, contains cannabidiol (CBD), which could be the cannabinoid causing the increased activity. Although the actual bioactivities of CBN, CBG, CBE and CBD remain speculative until further purification, characterization and reevaluations are carried out, a cautious judgement can be made that these compounds likely play a beneficial role. Fungal endophytes Alternaria alternata (A. alternata), Alternaria infectoria (A. infectoria), Fusarium incarnatum (F. incarnatum), and Fusarium chlamydosporum (F. chlamydosporum) were isolated from surface sterilized buds of C. sativa and identified using molecular and phylogenetic methods. A. alternata showed some compounds (via LC-QTOF-MS) which were previously isolated in C. sativa, with its extracts exhibiting immunomodulatory activity against THP-1 macrophages infected with M. smegmatis mc2155. The percentage survival for treated THP-1 cells was found to be 51.81% at 6 hours (p = 0.0003) compared to 81.91% untreated. Overall, these results establish that cannabinoids are able to influence THP-1 infected cells’ ability to clear mycobacterial infection albeit a low percentage cell survival. To develop specialized HDT strategies targeting macrophages, a deeper comprehension of the mutual interaction between cannabis and immunity is required. Future studies to isolate and characterize compound(s) from the endophytic fungi and plant extracts could result in lead development of naturally sourced drugs for host-directed TB treatment.