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1.
Biomed Res Int ; 2019: 3126376, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-33204680

RESUMO

In the biomedical field, there is growing interest in using human stem cell-derived neurons as in vitro models for pharmacological and toxicological screening of bioactive compounds extracted from natural products. Lignosus rhinocerus (Tiger Milk Mushroom) is used by indigenous communities in Malaysia as a traditional medicine to treat various diseases. The sclerotium of L. rhinocerus has been reported to have medicinal properties, including various bioactivities such as neuritogenic, anti-inflammatory, and anticancer effects. This study aims to investigate the neuroprotective activities of L. rhinocerus sclerotial extracts. Human embryonic stem cell (hESC)-derived neural lineages exposed to the synthetic glucocorticoid, dexamethasone (DEX), were used as the in vitro models. Excess glucocorticoids have been shown to adversely affect fetal brain development and impair differentiation of neural progenitor cells. Screening of different L. rhinocerus sclerotial extracts and DEX on the hESC-derived neural lineages was conducted using cell viability and neurite outgrowth assays. The neuroprotective effects of L. rhinocerus sclerotial extracts against DEX were further evaluated using apoptosis assays and Western blot analysis. Hot aqueous and methanol extracts of L. rhinocerus sclerotium promoted neurite outgrowth of hESC-derived neural stem cells (NSCs) with negligible cytotoxicity. Treatment with DEX decreased viability of NSCs by inducing apoptosis. Coincubation of L. rhinocerus methanol extract with DEX attenuated the DEX-induced apoptosis and reduction in phospho-Akt (pAkt) level in NSCs. These results suggest the involvement of Akt signaling in the neuroprotection of L. rhinocerus methanol extract against DEX-induced apoptosis in NSCs. Methanol extract of L. rhinocerus sclerotium exhibited potential neuroprotective activities against DEX-induced toxicity in hESC-derived NSCs. This study thus validates the use of human stem cell-derived neural lineages as potential in vitro models for screening of natural products with neuroprotective properties.


Assuntos
Células-Tronco Embrionárias Humanas , Neuroproteção , Fármacos Neuroprotetores/farmacologia , Polyporaceae/metabolismo , Animais , Anexina A5 , Anexinas/análise , Apoptose/efeitos dos fármacos , Proteínas de Arabidopsis , Produtos Biológicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Dexametasona/efeitos adversos , Humanos , Malásia , Medicina Tradicional
2.
Virol J ; 13: 5, 2016 Jan 06.
Artigo em Inglês | MEDLINE | ID: mdl-26738773

RESUMO

BACKGROUND: The incidence of neurological complications and fatalities associated with Hand, Foot & Mouth disease has increased over recent years, due to emergence of newly-evolved strains of Enterovirus 71 (EV71). In the search for new antiviral therapeutics against EV71, accurate and sensitive in vitro cellular models for preliminary studies of EV71 pathogenesis is an essential prerequisite, before progressing to expensive and time-consuming live animal studies and clinical trials. METHODS: This study thus investigated whether neural lineages derived from pluripotent human embryonic stem cells (hESC) can fulfil this purpose. EV71 infection of hESC-derived neural stem cells (NSC) and mature neurons (MN) was carried out in vitro, in comparison with RD and SH-SY5Y cell lines. RESULTS: Upon assessment of post-infection survivability and EV71 production by the various types, it was observed that NSC were significantly more susceptible to EV71 infection compared to MN, RD (rhabdomyosarcoma) and SH-SY5Y cells, which was consistent with previous studies on mice. The SP81 peptide had significantly greater inhibitory effect on EV71 production by NSC and MN compared to the cancer-derived RD and SH-SY5Y cell lines. CONCLUSIONS: Hence, this study demonstrates that hESC-derived neural lineages can be utilized as in vitro models for studying EV71 pathogenesis and for screening of antiviral therapeutics.


Assuntos
Linhagem da Célula , Enterovirus Humano A/fisiologia , Células-Tronco Embrionárias Humanas/citologia , Células-Tronco Neurais/citologia , Células-Tronco Neurais/virologia , Neurônios/citologia , Neurônios/virologia , Animais , Biomarcadores , Diferenciação Celular , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Enterovirus Humano A/efeitos dos fármacos , Expressão Gênica , Humanos , Camundongos , Células-Tronco Neurais/efeitos dos fármacos , Células-Tronco Neurais/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Peptídeos/farmacologia , Replicação Viral/efeitos dos fármacos
3.
Stem Cells Int ; 2015: 105172, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26089911

RESUMO

Human pluripotent stem cells (hPSCs) derived from either blastocyst stage embryos (hESCs) or reprogrammed somatic cells (iPSCs) can provide an abundant source of human neuronal lineages that were previously sourced from human cadavers, abortuses, and discarded surgical waste. In addition to the well-known potential therapeutic application of these cells in regenerative medicine, these are also various promising nontherapeutic applications in toxicological and pharmacological screening of neuroactive compounds, as well as for in vitro modeling of neurodegenerative and neurodevelopmental disorders. Compared to alternative research models based on laboratory animals and immortalized cancer-derived human neural cell lines, neuronal cells differentiated from hPSCs possess the advantages of species specificity together with genetic and physiological normality, which could more closely recapitulate in vivo conditions within the human central nervous system. This review critically examines the various potential nontherapeutic applications of hPSC-derived neuronal lineages and gives a brief overview of differentiation protocols utilized to generate these cells from hESCs and iPSCs.

4.
Cytometry A ; 81(10): 888-95, 2012 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-22865628

RESUMO

Coral bleaching is of increasing concern to reef management and stakeholders. Thus far, quantification of coral bleaching tends to be heavily reliant on the enumeration of zooxanthellae, with less emphasis on assessment of photosynthetic or physiological condition, these being often assessed separately by techniques such as liquid chromatography. Traditional methods of enumeration using microscopy are time consuming, subjected to low precision and great observer error. In this study, we presented a method for the distinction of physoiological condition and rapid enumeration of zooxanthellae using flow cytometry (FCM). Microscopy verified that healthy looking/live versus damaged/dead zooxanthellae could be reliably and objectively distinguished and counted by FCM on the basis of red and green fluorescence and light scatter. Excellent correlations were also determined between FCM and microscopy estimates of cell concentrations of fresh zooxanthellae isolates from Pocillopora damicornis. The relative intensities of chlorophyll and ß-carotene fluorescences were shown to be important in understanding the results of increased cell counts in freshly isolated zooxanthellae experimentally exposed to high temperatures (34, 36, and 38°C) over 24 h, with ambient temperature (29°C) used as controls. The ability to simultaneously identify and enumerate subpopulations of different physiological states in the same sample provides an enormous advantage in not just determining bleaching responses, but elucidating adaptive response and mechanisms for tolerance. Therefore, this approach might provide a rapid, convenient, and reproducible methodology for climate change studies and reef management programs.


Assuntos
Antozoários/fisiologia , Contagem de Células/métodos , Dinoflagellida/citologia , Dinoflagellida/fisiologia , Animais , Clorofila/metabolismo , Mudança Climática , Citometria de Fluxo , Fluorescência , Lasers , Microscopia , Fotossíntese , Reprodutibilidade dos Testes , Simbiose , Temperatura , beta Caroteno/metabolismo
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