Evaluation of plant extracts : artemisia afra and annona muricata for inhibitory activities against mycobacterium tuberculosis and human immunodeficiency virus

Mycobacterium tuberculosis and Human Immuno-Deficiency Virus (HIV) have a high prevalence in South Africa. The development and spread of drug resistant tuberculosis is a serious problem which is exacerbated by tuberculosis (TB) co-infection in HIV patients. Traditional medicinal plants like Annona muricata and Artemisia afra are used for respiratory ailments and antiviral therapies respectively. The aim of this study was to evaluate Annona muricata (ethanolic extract) and Artemisia afra (ethanolic and aqueous extracts) for inhibitory activities against M. tuberculosis and HIV. In vitro bioassays for anti-TB activity included: microplate alamar blue assay (MABA), flow cytometry and ρ-iodonitrotetrazolium chloride assays while anti-HIV activity was determined using an HIV-1 reverse transcriptase colorimetric ELISA kit and an HIV-1 integrase colorimetric immunoassay. Cytotoxicity of plant extracts were assessed by the MTT assay on Chang Liver and HepG2 cells. Potential synergistic effects were determined using the basis of Combination Index. Potential interactions of plant extracts with drug metabolic pathways were evaluated with the Glutathione-S-Transferase assay kit as well as the CYP3A4 assay kit. A. muricata ethanolic extract exhibited anti-TB activity with MIC 125 μg/mL. MABA was shown to be the most sensitive and effective method for the detection of anti-TB activity. Artemisia afra aqueous extract showed HIV-1 reverse transcriptase inhibition exhibiting ˃85 percent inhibition at 1 mg/mL while the ethanolic extracts of A. afra and A. muricata showed inhibition of HIV-1 integrase activity at ˃86.8 percent and ˃88.54 percent respectively at concentrations >0.5 - 4 mg/mL. The aqueous extract of A. afra displayed inhibition of HIV-1 integrase ˃52.16 percent at 0.5 mg/mL increasing to 72.89 percent at 4 mg/ml of the extract. A. muricata was cytotoxic at an IC50 of 30 μg/mL and 77 μg/mL on Chang Liver and HepG2 cells respectively, whilst A. afra aqueous and ethanol extracts were not cytotoxic to both cell lines. The ethanolic extract of A. muricata showed both antagonistic and synergistic properties at various IC values, when used in conjunction with rifampicin. A. afra ethanolic extract interrupted GST activity while aqueous extracts of A. afra and A. muricata had a slight effect. All extracts interrupted CYP3A4 activity, however the ethanolic extracts of A. muricata and A. afra showed greater inhibition than the aqueous extract of A. afra. These extracts should be investigated further as they could be an important source of compounds for treatment of M. tuberculosis and HIV respectively

Carotid Artery Intima-Media Thickness, Carotid Plaque and Coronary Heart Disease and Stroke in Chinese

Background: Our aim was to prospectively investigate the association between carotid artery intima-media thickness (IMT) as well as carotid plaque and incidence of coronary heart disease (CHD) and stroke in Chinese, among whom data are limited. Methods and Findings: We conducted a community-based cohort study composed of 2190 participants free of cardiovascular disease at baseline in one community. During a median 10.5-year follow up, we documented 68 new cases of coronary heart disease and 94 cases of stroke. The multivariate relative risks (RRs) associated with a change of 1 standard deviation of maximal common carotid IMT were 1.38 (95% confidence interval [CI], 1.12–1.70) for CHD and 1.47 (95% CI, 1.28–1.69) for stroke. The corresponding RRs with internal carotid IMT were 1.47 (95% CI, 1.21–1.79) for CHD and 1.52 (95% CI, 1.31–1.76) for stroke. Carotid plaque measured by the degree of diameter stenosis was also significantly associated with increased risk of CHD (p for trend<0.0001) and stroke (p for trend<0.0001). However, these associations were largely attenuated when adjusting for IMT measurements. Conclusions: This prospective study indicates a significant association between carotid IMT and incidence of CHD and stroke in Chinese adults. These measurements may be useful for cardiovascular risk assessment and stratification in Chinese

Prothrombotic gene variation and cerebral ischaemia of arterial origin

Stroke is the second most common cause of death and a leading cause of adult disability worldwide. Ischaemia is the cause in about 80% of all strokes. Atherosclerotic lesions in the cerebropetal and intracranial arteries lead to cerebral ischaemia of arterial origin. Cerebral ischaemia and atherosclerosis have a genetic basis. Genetic factors may help to identify stroke patients at high risk for new events. We studied associations between prothrombotic gene variation and cerebral ischaemia of arterial origin. Chapter 3 is a cohort study among 887 patients with cerebral ischaemia. During a 4.6-year mean follow-up period new vascular events occurred in 135 patients. None of 22 selected prothrombotic variants was associated with the occurrence of new vascular events. This study does not support the use of prothrombotic gene variants to identify stroke patients at increased risk for new vascular events. Chapter 4 is a case-control study in 316 long-term survivors (cases) and 887 patients with recent cerebral ischaemia (controls). Only two of 23 prothrombotic variants were associated with reduced survival after stroke. These variants do not appear to play an important role in survival after cerebral ischemia. In Chapter 5 we studied the effect of prothrombotic gene variation on atherosclerosis in 689 patients with nondisabling cerebral ischaemia. None of 22 prothrombotic variants was associated with carotid intima-media thickness or age. Only one variant was associated with symptomatic carotid stenosis. Prothrombotic gene variation does not seem to affect the pathogenesis of atherosclerosis. In Chapter 6 we compared genotype frequencies between 621 stroke patients with large vessel disease (LVD) and 266 patients with small vessel disease (SVD). Only one of 22 variants was not equally prevalent among LVD and SVD patients. Prothrombotic gene variants may not be specific for one stroke subtype. Claims about genetic associations with specific subtypes should be interpreted with caution. In Chapter 7 we found the carotid intima-media thickness to be larger in patients with LVD than in those with SVD. Carotid intima-media thickness is an established marker of artherosclerosis. This supports the hypothesis that LVD and SVD have a different pathogenesis. Chapter 8 is a case-control study among 190 young women with ischaemic stroke and 767 women without cardiovascular disease. The Phenylalanine allele of the Tyr204Phe variant conferred a 9-fold increased risk of stroke. The combination of oral contraceptive use and carriership of the 204Phe allele led to a 20-fold increase in stroke risk. TheTyr204Phe variant of the coagulation factor XIII subunit A gene constitutes a strong and common genetic factor for ischaemic stroke in young women. Genetic screening preceding oral contraceptive use is not warranted because of the low incidence of ischaemic stroke among young women. Chapter 9 is a nested case-control study in patients with cerebral ischaemia on oral anticoagulant treatment. A variant in the alpha fibrinogen gene was associated with a decreased anticoagulant-related bleeding risk and factor V Leiden conferred an increased bleeding risk. If replicated, these findings may have implications for the future selection of patients for oral anticoagulant treatment