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Drug Discovery Platform Targeting M. tuberculosis with Human Embryonic Stem Cell-Derived Macrophages.

ABSTRACT: A major limitation in anti-tuberculosis drug screening is the lack of reliable and scalable models for homogeneous human primary macrophage cells of non-cancer origin. Here we report a modified protocol for generating homogeneous populations of macrophage-like cells from human embryonic stem cells. The induced macrophages, referred to as iMACs, presented similar transcriptomic profiles and characteristic immunological features of classical macrophages and were permissive to viral and bacterial infection, in particular Mycobacterium tuberculosis (Mtb). More importantly, iMAC production was amenable to scale up. To evaluate iMAC efficiency in high-throughput anti-tuberculosis drug screening, we performed a phenotypic screening against intracellular Mtb, involving a library of 3,716 compounds that included FDA-approved drugs and other bioactive compounds. Our primary screen identified 120 hits, which were validated in a secondary screen by dose-intracellular and -extracellular Mtb assays. Our confirmatory studies identified a novel anti-Mtb compound, 10-DEBC, also showing activity against drug-resistant strains.

SUBMITTER: Han HW

PROVIDER: S-EPMC6915848 | biostudies-literature | 2019 Dec

REPOSITORIES: biostudies-literature

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Drug Discovery Platform Targeting M. tuberculosis with Human Embryonic Stem Cell-Derived Macrophages.

Han Hyo-Won HW Seo Hyang-Hee HH Jo Hye-Yeong HY Han Hyeong-Jun HJ Falcão Virgínia C A VCA Delorme Vincent V Heo Jinyeong J Shum David D Choi Jang-Hoon JH Lee Jin-Moo JM Lee Seung Hun SH Heo Hye-Ryeon HR Hong Seok-Ho SH Park Mi-Hyun MH Thimmulappa Rajesh K RK Kim Jung-Hyun JH

Stem cell reports 20191031 6

A major limitation in anti-tuberculosis drug screening is the lack of reliable and scalable models for homogeneous human primary macrophage cells of non-cancer origin. Here we report a modified protocol for generating homogeneous populations of macrophage-like cells from human embryonic stem cells. The induced macrophages, referred to as iMACs, presented similar transcriptomic profiles and characteristic immunological features of classical macrophages and were permissive to viral and bacterial i ...[more]

PMID: 31680058

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Project description:BackgroundInduced pluripotent stem cells (iPS) represent an innovative source for the standardized in vitro generation of macrophages (M?). M? show great promise in disease pathogenesis, particularly tuberculosis. However, there is no information about human iPS-derived (hiPS) macrophages (hiPS-M?) in response to tuberculosis infection.MethodsIn the present study, macrophages derived from hiPS were established via embryoid body (EB) formation by using feeder-free culture conditions, and the human monocyte cell line THP-1 (THP-1-M?) was used as control. iPS-M? were characterized by using morphology, Giemsa staining, nonspecific esterase staining (?-NAE), phagocytosis, and surface phenotype. Additionally, after treatment with Bacillus Calmette-Guérin (BCG) for 24 h, cell apoptosis was detected by using an Annexin V-FITC Apoptosis Detection assay. The production of nitric oxide (NO), expression of tumor necrosis factor alpha (TNF-?), activity of apoptosis-related protein cysteine-3 (Caspase-3) and expression of B-cell lymphoma-2 (Bcl-2) were analyzed.ResultsWith respect to morphology, surface phenotype, and function, the iPS-M? closely resembled their counterparts generated in vitro from a human monocyte cell line. iPS-M? exhibited the typically morphological characteristics of macrophages, such as round, oval, fusiform and irregular characteristics. The cells were Giemsa-stained-positive, ?-NAE-positive, and possessed phagocytic ability. iPS-M? express high levels of CD14, CD11b, CD40, CD68, and major histocompatibility complex II (MHC-II). Moreover, with regard to the apoptotic rate, the production of NO, expression of TNF-?, and activity of Caspase-3 and Bcl-2, iPS-M? closely resemble that of their counterparts generated in vitro from human monocyte cell line in response to BCG infection. The rate of apoptosis of BCG-treated iPS-M? was 37.77 ± 7.94% compared to that of the untreated group at 4.97 ± 1.60% (P < 0.01) by using Annexin V-FITC Apoptosis Detection. Additionally, the rate of apoptosis of BCG-treated THP-1-M? was 37.1 ± 2.84% compared to that of the untreated group at 6.19 ± 1.68% (P < 0.001). The expression of TNF-? and the production of NO were significantly increased (P < 0.001), and the activity of Caspase-3 was increased. However, the expression of Bcl-2 was inhibited (P < 0.001).ConclusionsOur results demonstrate that M? derived from hiPS perform the immunological function in response to Bacillus Calmette-Guérin infection by undergoing apoptosis, increasing the production of NO and expression of TNF-?. Thus, our study may help to overcome the limitations of research into certain rare diseases due to the lack of adequate supply of disease-specific primary cells.

Dataset Information

Drug Discovery Platform Targeting M. tuberculosis with Human Embryonic Stem Cell-Derived Macrophages.

Publications

Drug Discovery Platform Targeting M. tuberculosis with Human Embryonic Stem Cell-Derived Macrophages.

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