Stem cells in natural product and medicinal plant drug discovery-An overview of new screening approaches

Natural products remain a rich source of new drugs, and the search for bioactive molecules from nature continues to play an important role in the development of new medicines. Also, there is increasing use of herbal medicines for the treatment of a plethora of diseases, and demands for more scientific evidence for their efficacy and safety remains a huge challenge. The propensity of stem cells to differentiate into almost every cell type not only holds promise for the delivery of cell-based therapies for currently incurable diseases or a useful tool in studying cell physiology and pathophysiology. Increasingly, stem cells are becoming an important tool in preclinical drug screening and toxicity testing. In this review, we examine the scientific advances made towards the use of pluripotent stem cells as a model for the screening of plant-based medicines. The combination of well-established in vitro electrophysiological and a plethora of toxicogenomic technologies, together with the optimisation of culture methods of herbal plants and pluripotent stem cells can be explored to establish the basis for efficacy, and tissue/organ-based toxicities of many currently used medicinal plants whose efficacies and toxicities remain unknown

QuinoMit Q10-Fluid attenuates hydrogen peroxide-induced irregular beating in mouse pluripotent stem cell-derived cardiomyocytes

Background: Coenzyme Q10 (CoQ10) is a crucial component of the mitochondrial structure which is involved in producing more than 90% of cellular ATP. This study aimed to investigate the protective effects and underlying mechanisms of QuinoMit Q10-Fluid against hydrogen peroxide (H2O2)-induced arrhythmias on cardiomyocytes (CMs). Methods: Undifferentiated stem cell-derived CMs were cultured in the presence of different concentrations of QuinoMit Q10-Fluid. To investigate if CoQ10 has anti-apoptotic activity, CMs were exposed to H2O2 for up to 100 h with or without CoQ10. The expression levels of cardiac reference genes were determined by RT-PCR. The structural and functional properties of CMs were examined by immunofluorescence and the xCELLigence system. Caspase 3/7 assay was also performed for cell apoptosis study. Results: The study showed that QuinoMit Q10-Fluid inhibits the proliferation of pluripotent stem cells at high concentrations and had less effect on cardiomyogenesis. However, the beating rate of clusters containing CMs generated under QuinoMit Q10-Fluid (1:100) was significantly increased. This increase was accompanied by the up-regulated expression level of some important cardiac markers during differentiation. Treatment of CMs with H2O2 notably induced irregular beating and decreased the amplitude of the beating signal of CMs, concomitantly with increased caspase-3/7 activity. However, CMs pretreated with QuinoMit exhibited a protective effect against H2O2 -induced arrhythmia. Conclusion: Our results reveal that QuinoMit Q10-Fluid attenuates H2O2-induced irregular beating in mouse pluripotent stem cell-derived CMs, at least partly by reducing the generation of ROS, suggesting a protective effect against CM dysfunctions

The proliferative and chronotropic effects of Brillantaisia nitens Lindau (Acanthaceae) extracts on pluripotent stem cells and their cardiomyocytes derivatives

Ethnopharmacological relevance: Brillantaisia nitens Lindau (Acanthaceae) leaves are commonly used in traditional medicine in Africa for the treatment of many disorders including heart diseases and malaria. In this study, we therefore evaluated the effect of the methylene chloride/methanol leaf extract of Brillantaisia nitens on the proliferation of mouse pluripotent stem cells and their cardiomyocyte derivatives. Materials and methods: In this study, we combined two emerging technologies, pluripotent stem cell-derived cardiomyocytes and modern electrophysiology systems (impedance-based real-time) to assess the cytotoxicity of Brillantaisia nitens extract (BNE). Undifferentiated pluripotent cells and cardiomyocytes were exposed to different concentrations of BNE. Cell viability and contraction were monitored by impedance using the xCELLigence system for short-and long-term treatment whereas the excitability of single cardiomyocytes was captured by patch clamp technique after BNE acute exposure. Results: Brillantaisia nitens extract inhibited the proliferation and increased cytotoxicity of embryonic stem cells in a concentration-dependent manner. With the increase in concentration of BNE, beating rate and the contractile amplitude of cardiomyocytes changed significantly. Spontaneous rhythmic activity of cardiomyocytes was completely suppressed after 48 and 24 h exposures to relatively low (4.16 mg/ml) and high (8.32 mg/ml) concentrations of BNE, respectively. Moreover, acute application of 4.16 mg/ml of BNE led to a significant alteration of action potential (AP) parameters such as beating frequency, amplitude and AP duration at 90% of repolarization. Conclusion: Brillantaisia nitens extract inhibits the proliferative capacity of pluripotent stem cells and reduces electrical activity of cardiomyocytes, confirming its depressant action on the heart. (C) 2014 Elsevier Ireland Ltd. All rights reserved

Altered Functional Expression of beta-Adrenergic Receptors in Rhesus Monkey Embryonic Stem Cell-Derived Cardiomyocytes

Pluripotent stem cells have demonstrated the potential to generate large numbers of functional cardiomyocytes (CMs) from different cell sources. Besides Wnt signaling, additional pathways are involved in early cardiac development and function. To date however, no study exists showing the effects of perturbing the canonical Wnt pathway using nonhuman primate embryonic stem (ES) cells. In this study, we investigated the effect of canonical Wnt inhibition during differentiation of nonhuman primate ES cell-derived CMs under defined, growth factor conditions. Rhesus monkey ES (rES) cells were differentiated into spontaneously beating CMs in the absence (control) or presence (treated) of Wnt inhibitor Dickkopf1 (DKK1), vascular endothelial growth factor, and basic fibroblast growth factor combined or added in a sequential manner during differentiation. Quantification and functional characterization of CMs were assessed by molecular and electrophysiological techniques. Analysis revealed no difference in average ratio of spontaneously beating clusters in both control and treated groups. However, the percentage of CMs was significantly reduced and the expressions of specific cardiac markers tested were also decreased in the treated group. Interestingly, we found that in CMs obtained from treated group, beta-adrenergic receptors (beta-ARs) were less expressed, their function was altered and electrophysiological studies revealed differences in action potential responsiveness to beta-AR stimulation. We demonstrated that the Wnt/beta-catenin pathway inhibitor, DKK1 associated with other growth factors repressed functional expression of beta-ARs in rES cell-derived CMs. Thus, control of this pathway in each cell line and source is important for proper basic research and further cell therapy applications

Effects of Synthetic Neural Adhesion Molecule Mimetic Peptides and Related Proteins on the Cardiomyogenic Differentiation of Mouse Embryonic Stem Cells

Background/Aims: Pluripotent stem cells differentiating into cardiomyocyte-like cells in an appropriate cellular environment have attracted significant attention, given the potential use of such cells for regenerative medicine. However, the precise mechanisms of lineage specification of pluripotent stem cells are still largely to be explored. Identifying the role of various small synthetic peptides involved in cardiomyogenesis may provide new insights into pathways promoting cardiomyogenesis. Methods: In the present study, using a transgenic murine embryonic stem (ES) cell lineage expressing enhanced green fluorescent protein (EGFP) under the control of a-myosin heavy chain (alpha-MHC) promoter (p alpha MHC-EGFP), we investigated the cardiomyogenic effects of 7 synthetic peptides (Betrofin3, FGLs, FGL(L), hNgf_C2, EnkaminE, Plannexin and C3) on cardiac differentiation. The expression of several cardiac-specific markers was determined by RT-PCR whereas the structural and functional properties of derived cardiomyocytes were examined by immunofluorescence and electrophysiology, respectively. Results: The results revealed that Betrofin3, an agonist of brain derived neurotrophic factor (BDNF) peptide exerted the most striking pro-cardiomyogenic effect on ES cells. We found that BDNF receptor, TrkB expression was up-regulated during differentiation. Treatment of differentiating cells with Betrofin3 between days 3 and 5 enhanced the expression of cardiac-specific markers and improved cardiomyocyte differentiation and functionality as revealed by genes regulation, flow cytometry and patch clamp analysis. Thus Betrofin3 may exert its cardiomyogenic effects on ES cells via TrkB receptor. Conclusion: Taken together, the results suggest that Betrofin3 modulates BDNF signaling with positive cardiomyogenic effect in stage and dose-dependent manner providing an effective strategy to increase ES cell-based generation of cardiomyocytes and offer a novel therapeutic approach to cardiac pathologies where BDNF levels are impaired. Introduction Copyright (C) 2015 S. Karger AG, Base

Liver injury in malaria infected patients in Douala-Cameroon and its association with poor medical practice

Abstract Background Malaria is an endemic mosquito-borne disease in sub-Saharan regions, including Cameroon. Due to the obligatory hepatic stage of its pathogenic agents, malaria can induce liver damage if not properly treated. Hence, we assessed the impact of malaria infection on liver transaminases among febrile patients consulting at the Deido District Hospital, Douala-Cameroon, in regard to their attitude towards the practice of preventive measures, treatment, and management of malaria. Methods Over 10 weeks, 150 febrile patients and 28 healthy individuals serving as the control group were enrolled and their blood samples screened for Plasmodium species by Giemsa Staining and liver injury evaluated by measuring the serum level of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities. The socio-demographic characteristics of participants and their attitude towards the practice of preventive measures, treatment, and management of malaria were collected using a structured- questionnaire. Results Among tested febrile patients, 113 (75%) were malaria-positive. Females were more affected (65.5%) than males; the most affected age group were adults between 30-60 years (55.8%). A significant association (p˂0.05; relative risk [RR] = 1.424 or p˂0.05; RR = 1.947) was found between malaria infection and non-use of mosquito nets or insecticides, respectively. The serum level of ALT and AST activities in malaria-positive were significantly (p<0.05) increased, compared to healthy or malaria-negative individuals. Furthermore, transaminase activity was significantly (p<0.05) elevated in non-practitioners of preventive measures; and in patients who engaged in auto-medication or traditional medication, compared to those who sought treatment from health centers. Conclusion Our findings demonstrated that non-practice of preventive measures, improper treatment and management of malaria infection can lead to an abnormal increase in serum level of transaminases which may reflect liver injury