High-throughput multiparametric live-cell and immunofluorescence analyses demonstrate antifibrotic bioactivities of O. basilicum and A. membranaceus hairy roots on human hepatic stellate cells

IF 4.5 Q1 PLANT SCIENCES
Shannah Secret , Milena Mennecozzi , Tom Crabbe , Alessandra Devoto
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引用次数: 0

Abstract

Plants are a well-established source of compounds with biopharmaceutical activity, but the diversity and often low abundance of phytochemicals have hindered their medical application. Bridging this gap requires scalable growth conditions and sustainable extraction methodologies harnessed to robust and predictive assays of disease and cell toxicity. This study utilises plant hairy root biomass and constituent bioactives combined with high-throughput assays of fibrogenic progression using activated human hepatic stellate cells.
Fibrosis is a build-up of connective tissue components like collagen type I and fibronectin and is one of the underlying drivers of liver cirrhosis, a complex and often fatal condition arising from alcohol abuse, poor diet, or viral infection. New treatments are required to reverse the predicted steady increase in the global burden of this disease.
In this study, Astragalus membranaceus and Ocimum basilicum were transformed with Agrobacterium rhizogenes (Rhizobium rhizogenes) to provide reproducible and rapidly growing hairy root cultures. Titrations of methanol and pressed root extracts obtained from both species, altered selectively, the deposition of collagen type I and fibronectin, along variable effects on live readouts of stellate cells proliferation and death, distinguishing between Caspase-3/7 activation and membrane integrity maintenance. These results differ from the wider effects observed with the known anti-fibrotic compounds Nintedanib and Silymarin, which suggests that the extracts may prove to be a significant source of novel biopharmaceuticals useful in the treatment of liver cirrhosis. Remarkably, the concentrations showing activity were not toxic for primary human hepatocytes.
Moreover, the platform developed provides a novel integrated high-throughput sustainable extraction method and readout to elucidate the mechanism of action of unknown compounds with respect to previous studies.
高通量多参数活细胞和免疫荧光分析证实了basilicum和A. membranaceus毛状根对人肝星状细胞的抗纤维化生物活性
植物是具有生物制药活性的化合物的公认来源,但植物化学物质的多样性和往往低丰度阻碍了它们的医学应用。弥合这一差距需要可扩展的生长条件和可持续的提取方法,利用强大的疾病和细胞毒性预测分析。本研究利用植物毛状根生物量和成分生物活性,结合使用活化的人肝星状细胞进行高通量纤维化进展测定。纤维化是一种结缔组织成分的积累,如I型胶原蛋白和纤维连接蛋白,是肝硬化的潜在驱动因素之一,肝硬化是一种复杂且通常致命的疾病,由酗酒、不良饮食或病毒感染引起。需要新的治疗方法来扭转这一疾病全球负担的预期稳定增长。本研究利用根生农杆菌(Rhizobium rhizogenes)对黄芪和罗勒木进行转化,获得了可繁殖且生长迅速的毛状根培养物。从这两种植物中提取的甲醇和压根提取物的滴定,选择性地改变了I型胶原和纤维连接蛋白的沉积,以及对星状细胞增殖和死亡的实时读数的可变影响,区分了Caspase-3/7激活和膜完整性维持。这些结果不同于已知的抗纤维化化合物尼达尼布和水飞蓟素所观察到的更广泛的效果,这表明提取物可能被证明是用于治疗肝硬化的新型生物药物的重要来源。值得注意的是,显示活性的浓度对原代人肝细胞没有毒性。此外,开发的平台提供了一种新的集成高通量可持续提取方法和读数,以阐明未知化合物的作用机制。
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来源期刊
Current Plant Biology
Current Plant Biology Agricultural and Biological Sciences-Plant Science
CiteScore
10.90
自引率
1.90%
发文量
32
审稿时长
50 days
期刊介绍: Current Plant Biology aims to acknowledge and encourage interdisciplinary research in fundamental plant sciences with scope to address crop improvement, biodiversity, nutrition and human health. It publishes review articles, original research papers, method papers and short articles in plant research fields, such as systems biology, cell biology, genetics, epigenetics, mathematical modeling, signal transduction, plant-microbe interactions, synthetic biology, developmental biology, biochemistry, molecular biology, physiology, biotechnologies, bioinformatics and plant genomic resources.
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