Studies of ionic mechanisms in model cell lines of prostate cancer

Metastasis is the major problem in clinical management of cancer, including prostate cancer (PCa). Biochemical conditions in tumour microenvironment (especially O2 and Ca2+ levels) are important for modulating metastatic cell behaviours (MCBs). Ionic mechanisms, in particular voltage-gated sodium channel (VGSC) and intracellular Ca2+ activities, were shown previously to be involved in MCBs in PCa. This PhD tested the hypotheses (1) that MCBs would be sensitive to hypoxia and be inhibited by VGSC blockers including those selective for the persistent current and (2) that VGSC activity would control intracellular Ca2+ signalling. Experiments were carried on a variety of model cell lines in a comparative approach: strongly metastatic Mat-LyLu and weakly metastatic AT-2 rat PCa cells, and analogous human PCa cell lines (PC-3M and LnCaP, respectively). Experiments on Mat-LyLu cells showed that hypoxia (2 % O2, 24 h) increased mRNA expression of Nav1.7, the predominant VGSC α-subunit expressed in PCa; however, both plasma membrane and intracellular VGSC protein levels were reduced. There was no change in cellular proliferation, Matrigel invasion or lateral motility, whilst transverse migration increased significantly. Treatment (24 h) with the VGSC blockers ranolazine, riluzole, lidocaine and procaine (micromolar concentrations) decreased Nav1.7 mRNA and total VGSC protein levels, and suppressed Matrigel invasion in both normoxia (~21 % O2) and hypoxia. PC-3M (but not LNCaP or Mat-LyLu) cells showed spontaneous, transient elevations of intracellular Ca2+ (“Ca2+ oscillations”). Ionic substitution and pharmacological experiments suggested that Ca2+ influx and release from intracellular stores both contributed to the oscillations. Importantly, acute treatment with tetrodotoxin and ranolazine decreased the oscillation amplitude and frequency. Thus, both hypotheses were confirmed, i.e. (1) that hypoxia increases PCa (Mat-LyLu) cell migration and VGSC expression and (2) that intracellular Ca2+ oscillations are downstream to VGSC activity. Overall, the thesis concludes that VGSC blockers could serve clinically as anti-metastatic agents

Studies of ionic mechanisms in model cell lines of prostate cancer

Metastasis is the major problem in clinical management of cancer, including prostate cancer (PCa). Biochemical conditions in tumour microenvironment (especially O2 and Ca2+ levels) are important for modulating metastatic cell behaviours (MCBs). Ionic mechanisms, in particular voltage-gated sodium channel (VGSC) and intracellular Ca2+ activities, were shown previously to be involved in MCBs in PCa. This PhD tested the hypotheses (1) that MCBs would be sensitive to hypoxia and be inhibited by VGSC blockers including those selective for the persistent current and (2) that VGSC activity would control intracellular Ca2+ signalling. Experiments were carried on a variety of model cell lines in a comparative approach: strongly metastatic Mat-LyLu and weakly metastatic AT-2 rat PCa cells, and analogous human PCa cell lines (PC-3M and LnCaP, respectively). Experiments on Mat-LyLu cells showed that hypoxia (2 % O2, 24 h) increased mRNA expression of Nav1.7, the predominant VGSC α-subunit expressed in PCa; however, both plasma membrane and intracellular VGSC protein levels were reduced. There was no change in cellular proliferation, Matrigel invasion or lateral motility, whilst transverse migration increased significantly. Treatment (24 h) with the VGSC blockers ranolazine, riluzole, lidocaine and procaine (micromolar concentrations) decreased Nav1.7 mRNA and total VGSC protein levels, and suppressed Matrigel invasion in both normoxia (~21 % O2) and hypoxia. PC-3M (but not LNCaP or Mat-LyLu) cells showed spontaneous, transient elevations of intracellular Ca2+ (“Ca2+ oscillations”). Ionic substitution and pharmacological experiments suggested that Ca2+ influx and release from intracellular stores both contributed to the oscillations. Importantly, acute treatment with tetrodotoxin and ranolazine decreased the oscillation amplitude and frequency. Thus, both hypotheses were confirmed, i.e. (1) that hypoxia increases PCa (Mat-LyLu) cell migration and VGSC expression and (2) that intracellular Ca2+ oscillations are downstream to VGSC activity. Overall, the thesis concludes that VGSC blockers could serve clinically as anti-metastatic agents.EThOS - Electronic Theses Online ServicePro Cancer Research Fund (PCRF)GBUnited Kingdo

Anti-proliferative and cytotoxic activities of Allium autumnale P. H. Davis (Amaryllidaceae) on human breast cancer cell lines MCF-7 and MDA-MB-231

Abstract Background Natural products obtained from plants can be potent sources for developing a variety of pharmaceutical products. Allium species have been widely studied for their anti-cancer effects and presented promising results as potential anti-cancer agents. Breast cancer (BCa) is one of the most commonly diagnosed types of cancer in women. In this study, we aimed to investigate the anti-proliferative, cytotoxic and anti-metastatic effects of bulb and stem extracts from Allium autumnale P. H. Davis (Amaryllidaceae), an endemic Allium species to the island of Cyprus, in a comparative approach to weakly metastatic MCF-7 and strongly metastatic MDA-MB-231 breast cancer (BCa) cell lines. Methods Possible cytotoxic, anti-proliferative and anti-metastatic effects of the Allium extracts on MCF-7 and MDA-MB-231 cells were tested using trypan blue exclusion, MTT and wound heal assays, respectively. Gas Chromatography Mass Spectroscopy (GC-MS) analysis was performed to determine the prominent medically important compounds in Allium autumnale bulb (AAB) and Allium autumnale stem (AAS) extracts. Student unpaired t-test or ANOVA followed by Newman-Keuls post hoc analysis (INSTAT Software) was used where appropriate. Results Our results demonstrate that AAB extract (24, 48 and 72 h) exerts significant anti-proliferative effect on both MCF-7 and MDA-MB-231 cells where this effect for AAS extract was observed only at high (5000 and 10,000 μg/mL) concentrations. Cell viability experiments revealed that AAB extract incubations caused more cytotoxicity on both BCa cell lines compared to the AAS. In contrast, there was no effect on lateral motilities of either cell line. Conclusion Overall, our studies demonstrated the anti-cancer activities associated with Allium autumnale, revealing it’s cytotoxic and anti-proliferative potential to be further utilized in in vivo studies

Biosynthesized ZnO Nanoparticles Using Albizia lebbeck Extract Induced Biochemical and Morphological Alterations in Wistar Rats

Nano-based particles synthesized via green routes have a particular structure that is useful in biomedical applications as they provide cheap, eco-friendly, and non-toxic nanoparticles. In the present study, we reported the effect of various concentrations of Zinc oxide nanoparticles synthesized using A. lebbeck stem bark extract (ZnO NPsAL) as stabilizing agent on rat biochemical profiles and tissue morphology. Adult Wistar rats weighing 170 ± 5 g were randomly classified into eight groups of five rats each; Group A served as a control fed with normal diet and water. Groups B1, B2, C1, C2, D1, D2, and E were treated with 40 mg/kg and 80 mg/kg of the 0.01, 0.05, and 0.1 M biosynthesized ZnO NPsAL and zinc nitrate daily by the gavage method, respectively. The rats were anesthetized 24 h after the last treatment, blood samples, kidney, heart, and liver tissues were collected for biochemical and histopathological analysis. The rats mean body weight, serum alkaline phosphatase, alanine aminotransferase, creatinine, urea, bilirubin, protein, albumin, globulin, total cholesterol, triacylglycerol, and high-density lipoprotein were significantly altered with an increased concentration of biosynthesized ZnO NPsAL when compared with the control group (p < 0.05; n ≥ 5). Furthermore, histopathological analysis of treated rats’ kidney, heart, and liver tissue revealed vascular congestion, tubular necrosis, inflammation, and cytoplasmic vacuolation. Biosynthesized ZnO NPsAL showed significant alteration in biochemical parameters and tissue morphology in rats with increasing concentrations of the nanoparticles

Riluzole: Anti-invasive effects on rat prostate cancer cells under normoxic and hypoxic conditions

Anti-invasive effects of riluzole and ranolazine, a neuro-protectant and an anti-anginal drug, respectively, on Mat-LyLu rat prostate cancer (PCa) cells were tested in vitro (a) at non-toxic doses and (b) under both normoxic and hypoxic conditions, the latter common to growing tumours. Tetrodotoxin (TTX) was used as a positive control. Hypoxia had no effect on cell viability but reduced growth at 48 hours. Riluzole (5 mu mol/L) or ranolazine (20 mu mol/L) had no effect on cell viability or growth under normoxia or hypoxia over 24 hours. Matrigel invasion was not affected by hypoxia but inhibited by TTX, ranolazine and riluzole under a range of conditions. The expression of Nav1.7 mRNA, the prevailing, pro-invasive voltage-gated sodium channel alpha-subunit (VGSC alpha), was up-regulated by hypoxia. Riluzole had no effect on Nav1.7 mRNA expression in normoxia but significantly reduced it in hypoxia. VGSC alpha protein expression in plasma membrane was reduced in hypoxia; riluzole increased it but only under hypoxia. It was concluded (a) that riluzole and ranolazine have anti-invasive effects on rat PCa cells and (b) that Nav1.7 mRNA and protein expression can be modulated by riluzole under hypoxia. Overall, therefore, riluzole and ranolazine may ultimately be "repurposed" as anti-metastatic drugs against PCa

Prediction of Cell Migration in MDA-MB 231 and MCF-7 Human Breast Cancer Cells Treated with <i>Albizia Lebbeck</i> Methanolic Extract Using Multilinear Regression and Artificial Intelligence-Based Models

Breast cancer is a common cancer affecting women worldwide, and it progresses from breast tissue to other parts of the body through a process called metastasis. Albizia lebbeck is a valuable plant with medicinal properties due to some active biological macromolecules, and it’s cultivated in subtropical and tropical regions of the world. This study reports the phytochemical compositions, the cytotoxic, anti-proliferative and anti-migratory potential of A. lebbeck methanolic (ALM) extract on strongly and weakly metastatic MDA-MB 231 and MCF-7 human breast cancer cells, respectively. Furthermore, we employed and compared an artificial neural network (ANN), an adaptive neuro-fuzzy inference system (ANFIS), and multilinear regression analysis (MLR) to predict cell migration on the treated cancer cells with various concentrations of the extract using our experimental data. Lower concentrations of the ALM extract (10, 5 & 2.5 μg/mL) showed no significant effect. Higher concentrations (25, 50, 100 & 200 μg/mL) revealed a significant effect on the cytotoxicity and proliferation of the cells when compared with the untreated group (p n ≥ 3). Furthermore, the extract revealed a significant decrease in the motility index of the cells with increased extract concentrations (p n ≥ 3). The comparative study of the models observed that both the classical linear MLR and AI-based models could predict metastasis in MDA-MB 231 and MCF-7 cells. Overall, various ALM extract concentrations showed promising an-metastatic potential in both cells, with increased concentration and incubation period. The outcomes of MLR and AI-based models on our data revealed the best performance. They will provide future development in assessing the anti-migratory efficacies of medicinal plants in breast cancer metastasis