Antiproliferative activity of aqueous leaf extract of annona muricata (linn.) on rat prostate, BPH-1 cells and some target genes.

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Annona muricata (Linn.) is an evergreen tropical tree of the Annonaceae family that posseses phytotherapeutic bioactive compounds known as annonaceous acetogenins effective for the treatment of cancers and other conditions. This study aimed at investigating the effect of the aqueous leaf extract of A. muricata on human BPH-1 cells, the prostate organ as well as certain target cellular genes of proliferative activity. Dried A. muricata leaves were pulverized, and the aqueous crude extract obtained. HPLC was used for monitoring various batches of the A. muricata leaf extract (AMLE). The MTT assay was performed on BPH-1 cells (1 x 105 per well) using AMLE concentrations of 0.5, 1.0 and 1.5 mg/mL for 24, 48 and 72 hours. Microscopic examination of proliferation as well as morphology of the cells was carried out. RT-PCR was used to examine possible target genes, Bax and Bcl-2, using mRNA extracted from cells. Fifteen (15) F344 male rats (150-200 g) were placed in three groups of five (5). The low dose (LD) and high dose (HD) groups were gavaged 30 mg/mL and 300 mg/mL respectively and fed ad libitum alongside five (5) control (ctrl) group rats. Rats were sacrificed after 60 days. Whole blood was sampled by cardiac puncture and processed for biochemical assay. Prostate, seminal vesicles and testes were harvested, weighed and stored for histological examination. HPLC chromatographic fingerprint monitoring of different batches of AMLE yielded eight peaks. In vitro cell viability analysis showed a dose dependent growth inhibitory effect of AMLE on BPH-1 cells. The IC50 obtained for AMLE was 1.36 mg/ml. Increasing doses of AMLE directly up-regulated the levels of the pro-apototic protein Bax, whiles down-regulating the levels of the anti-apoptotic protein Bcl-2. In vivo studies showed statistically significant seminal vesicle indices recorded for both (LD) and (HD) groups (p=0.004 and p=0.009 respectively) compared with control (ctrl) group. There was no statistically significant difference between PSA levels of test groups compared with control groups. Histological examinations of the prostatic and seminal vesicle tissues however showed dose dependent morphological changes with AMLE treatment. The acinii were empty of secretions and there was marked atrophy with increased cellularity observed. AMLE exerts decreased secretory activity on prostate with flattening of acinar epithelial linings being demonstrated. The prostate epithelial and stromal cells were flattened with scanty prostatic secretions in the lumen. Thus, AMLE has antiproliferative activity against BPH-1 cells.
A thesis submitted in partial fulfilment of the requirements for the degree of Master of Science (Chemical Patholgy) in the Department of Molecular Medicine, School of Medical Sciences,