RESUMO
BACKGROUND/AIM: We and others have previously shown that cell fusion plays an important role in cancer metastasis. Color coding of cancer and stromal cells with spectrally-distinct fluorescent proteins is a powerful tool, as pioneered by our laboratory to detect cell fusion. We have previously reported color-coded cell fusion between cancer cells and stromal cells in metastatic sites by using color-coded EL4 murine lymphoma cells and host mice expressing spectrally-distinct fluorescent proteins. Cell fusion occurred between cancer cells or, between cancer cells and normal cells, such as macrophages, fibroblasts, and mesenchymal stem cells. In the present study, the aim was to morphologically classify the fusion-hybrid cells observed in the primary tumor and multiple metastases EL4 formed from cells expressing red fluorescent protein (RFP) in transgenic mice expressing green fluorescent protein (GFP), in a syngeneic model. MATERIALS AND METHODS: RFP-expressing EL4 murine lymphoma cells were cultured in vitro. EL4-RFP cells were harvested and injected intraperitoneally into immunocompetent transgenic C57/BL6-GFP mice to establish a syngeneic model. Two weeks later, mice were sacrificed and each organ was harvested, cultured, and observed using confocal microscopy. RESULTS: EL4 intraperitoneal tumors (primary) and metastases in the lung, liver, blood, and bone marrow were formed. All tumors were harvested and cultured. In all specimens, RFP-EL4 cells, GFP-stromal cells, and fused yellow-fluorescent hybrid cells were observed. The fused hybrid cells showed various morphologies. Immune cell-like round-shaped yellow-fluorescent fused cells had a tendency to decrease with time in liver metastases and circulating blood. In contrast fibroblast-like spindle-shaped yellow-fluorescent fused cells increased in the intraperitoneal primary tumor, lung metastases, and bone marrow. CONCLUSION: Cell fusion between EL4-RFP cells and GFP stromal cells occurred in primary tumors and all metastatic sites. The morphology of the fused hybrid cells varied in the primary and metastatic sites. The present results suggest that fused cancer and stromal hybrid cells of varying morphology may play an important role in cancer progression.
Assuntos
Fusão Celular , Modelos Animais de Doenças , Proteínas Luminescentes , Linfoma , Camundongos Transgênicos , Proteína Vermelha Fluorescente , Células Estromais , Animais , Camundongos , Células Estromais/patologia , Células Estromais/metabolismo , Linhagem Celular Tumoral , Linfoma/patologia , Linfoma/genética , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Metástase Neoplásica , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Células Híbridas/patologiaRESUMO
Duchenne muscular dystrophy (DMD) is a severe X-linked disorder characterized by dystrophin gene mutations and mitochondrial dysfunction, leading to progressive muscle weakness and premature death of DMD patients. We developed human Dystrophin Expressing Chimeric (DEC) cells, created by the fusion of myoblasts from normal donors and DMD patients, as a foundation for DT-DEC01 therapy for DMD. Our preclinical studies on mdx mouse models of DMD revealed enhanced dystrophin expression and functional improvements in cardiac, respiratory, and skeletal muscles after systemic intraosseous DEC administration. The current study explored the feasibility of mitochondrial transfer and fusion within the created DEC cells, which is crucial for developing new therapeutic strategies for DMD. Following mitochondrial staining with MitoTracker Deep Red and MitoTracker Green dyes, mitochondrial fusion and transfer was assessed by Flow cytometry (FACS) and confocal microscopy. The PEG-mediated fusion of myoblasts from normal healthy donors (MBN/MBN) and normal and DMD-affected donors (MBN/MBDMD), confirmed the feasibility of myoblast and mitochondrial fusion and transfer. The colocalization of the mitochondrial dyes MitoTracker Deep Red and MitoTracker Green confirmed the mitochondrial chimeric state and the creation of chimeric mitochondria, as well as the transfer of healthy donor mitochondria within the created DEC cells. These findings are unique and significant, introducing the potential of DT-DEC01 therapy to restore mitochondrial function in DMD patients and in other diseases where mitochondrial dysfunction plays a critical role.
Assuntos
Distrofina , Mitocôndrias , Distrofia Muscular de Duchenne , Mioblastos , Distrofia Muscular de Duchenne/terapia , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/metabolismo , Distrofia Muscular de Duchenne/patologia , Humanos , Mitocôndrias/metabolismo , Animais , Distrofina/genética , Distrofina/metabolismo , Mioblastos/metabolismo , Mioblastos/citologia , Mioblastos/transplante , Camundongos , Terapia Baseada em Transplante de Células e Tecidos/métodos , Masculino , Camundongos Endogâmicos mdx , Células Híbridas , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Fusão CelularRESUMO
Allele-specific monoallelic gene expression is a unique phenomenon and a great resource for analyzing gene regulation. To study this phenomenon, we established new embryonic stem (ES) cell lines derived from F1 hybrid blastocysts from crosses between four mouse subspecies (Mus musculus domesticus, C57BL/6; M. musculus molossinus, MSM/Ms; M. musculus musculus, PWK; M. musculus castaneus, HMI/Ms) and analyzed the expression levels of undifferentiated pluripotent stem cell markers and karyotypes of each line. To demonstrate the utility of our cell lines, we analyzed the allele-specific expression pattern of the Inpp5d gene as an example. The allelic expression depended on the parental alleles; this dependence could be a consequence of differences in compatibility between cis- and trans-elements of the Inpp5d gene from different subspecies. The use of parental mice from four subspecies greatly enhanced genetic polymorphism. The F1 hybrid ES cells retained this polymorphism not only in the Inpp5d gene, but also at a genome-wide level. As we demonstrated for the Inpp5d gene, the established cell lines can contribute to the analysis of allelic expression imbalance based on the incompatibility between cis- and trans-elements and of phenotypes related to this incompatibility.
Assuntos
Desequilíbrio Alélico , Animais , Camundongos , Desequilíbrio Alélico/genética , Camundongos Endogâmicos C57BL , Alelos , Expressão Gênica/genética , Linhagem Celular , Fosfatidilinositol-3,4,5-Trifosfato 5-Fosfatases/genética , Polimorfismo Genético , Células-Tronco Embrionárias Murinas/citologia , Células-Tronco Embrionárias Murinas/metabolismo , Células Híbridas , Células-Tronco Embrionárias , Feminino , Especificidade da Espécie , MasculinoRESUMO
Persistently high, worldwide mortality from cancer highlights the unresolved challenges of disease surveillance and detection that impact survival. Development of a non-invasive, blood-based biomarker would transform survival from cancer. We demonstrate the functionality of ultra-high content analyses of a newly identified population of tumor cells that are hybrids between neoplastic and immune cells in patient matched tumor and peripheral blood specimens. Using oligonucleotide conjugated antibodies (Ab-oligo) permitting cyclic immunofluorescence (cyCIF), we present analyses of phenotypes among tumor and peripheral blood hybrid cells. Interestingly, the majority of circulating hybrid cell (CHC) subpopulations were not identified in tumor-associated hybrids. These results highlight the efficacy of ultra-high content phenotypic analyses using Ab-oligo based cyCIF applied to both tumor and peripheral blood specimens. The combination of a multiplex phenotypic profiling platform that is gentle enough to analyze blood to detect and evaluate disseminated tumor cells represents a novel approach to exploring novel tumor biology and potential utility for developing the population as a blood-based biomarker in cancer.
Assuntos
Células Neoplásicas Circulantes , Humanos , Células Neoplásicas Circulantes/patologia , Biomarcadores Tumorais , Células Híbridas/patologia , Anticorpos , FenótipoRESUMO
Although gene expression divergence has long been postulated to be the primary driver of human evolution, identifying the genes and genetic variants underlying uniquely human traits has proven to be quite challenging. Theory suggests that cell-type-specific cis-regulatory variants may fuel evolutionary adaptation due to the specificity of their effects. These variants can precisely tune the expression of a single gene in a single cell-type, avoiding the potentially deleterious consequences of trans-acting changes and non-cell type-specific changes that can impact many genes and cell types, respectively. It has recently become possible to quantify human-specific cis-acting regulatory divergence by measuring allele-specific expression in human-chimpanzee hybrid cells-the product of fusing induced pluripotent stem (iPS) cells of each species in vitro. However, these cis-regulatory changes have only been explored in a limited number of cell types. Here, we quantify human-chimpanzee cis-regulatory divergence in gene expression and chromatin accessibility across six cell types, enabling the identification of highly cell-type-specific cis-regulatory changes. We find that cell-type-specific genes and regulatory elements evolve faster than those shared across cell types, suggesting an important role for genes with cell-type-specific expression in human evolution. Furthermore, we identify several instances of lineage-specific natural selection that may have played key roles in specific cell types, such as coordinated changes in the cis-regulation of dozens of genes involved in neuronal firing in motor neurons. Finally, using novel metrics and a machine learning model, we identify genetic variants that likely alter chromatin accessibility and transcription factor binding, leading to neuron-specific changes in the expression of the neurodevelopmentally important genes FABP7 and GAD1. Overall, our results demonstrate that integrative analysis of cis-regulatory divergence in chromatin accessibility and gene expression across cell types is a promising approach to identify the specific genes and genetic variants that make us human.
Assuntos
Cromatina , Pan troglodytes , Humanos , Animais , Cromatina/genética , Células Híbridas , Neurônios Motores , Expressão GênicaRESUMO
Human cancer cell lines have an essential role in cancer research, but only authentic cell lines should be used as biological models. Authentication testing using short tandem repeat (STR) loci has shown that MGC-803 cells, which were reported to come from gastric adenocarcinoma, are similar to HeLa. In this study, we confirmed that the MGC-803 cell line contains genetic material from HeLa, including genetic sequence from human papilloma virus 18 (HPV18). Additional alleles were present on STR analysis that remained stable after extensive passaging and generation of mono-clones. This behavior is consistent with a hybrid cell line arising from cell-cell fusion. Further genetic analysis revealed that MGC-803 originated from donors with different genetic ancestries, one African (HeLa) and the other Asian. Transcriptomic analysis demonstrated that MGC-803 closely resembles HeLa and another nasopharyngeal-HeLa hybrid cell line CNE-2. Based on these findings, we conclude that MGC-803 is a hybrid cell line derived from HeLa and other cells, the latter derived from a different patient with Asian genetic ancestry.
Assuntos
Adenocarcinoma , Humanos , Células HeLa , Células Clonais , Alelos , Células Híbridas , Linhagem Celular TumoralRESUMO
This study uses personalized chronic lymphoblastic leukemia (CLL) cybrid cells to test various drugs/agents designed to improve mitochondrial function and cell longevity. Age-matched control (NL) and CLL cybrids were created. The NL and CLL cybrids were treated with ibrutinib (Ibr-10 µM), mitochondrial-targeted nutraceuticals such as alpha lipoic acid (ALA-1 mM), amla (Aml-300 µg), melatonin (Mel-1 mM), resveratrol (Res-100 µM) alone, or a combination of ibrutinib with nutraceuticals (Ibr + ALA, Ibr + Aml, Ibr + Mel, or Ibr + Res) for 48 h. MTT (3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazoliumbromide), H2DCFDA(2',7' Dichlorodihydrofluorescein diacetate), and JC1 assays were used to measure the cellular metabolism, intracellular ROS levels, and mitochondrial membrane potential (∆ψm), respectively. The expression levels of genes associated with antioxidant enzymes (SOD2, GPX3, and NOX4), apoptosis (BAX and CASP3), and inflammation (IL6, IL-1ß, TNFα, and TGFß) were measured using quantitative real-time PCR (qRT-PCR). CLL cybrids treated with Ibr + ALA, Ibr + Aml, Ibr + Mel, and Ibr + Res had (a) reduced cell survivability, (b) increased ROS production, (c) increased ∆ψm levels, (d) decreased antioxidant gene expression levels, and (e) increased apoptotic and inflammatory genes in CLL cybrids when compared with ibrutinib-alone-treated CLL cybrids. Our findings show that the addition of nutraceuticals makes the CLL cybrids more pro-apoptotic with decreased cell survival compared with CLL cybrids exposed to ibrutinib alone.
Assuntos
Leucemia Linfocítica Crônica de Células B , Leucemia Mieloide Aguda , Mitocôndrias , Humanos , Antioxidantes/metabolismo , Antioxidantes/farmacologia , Antioxidantes/uso terapêutico , Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Leucemia Linfocítica Crônica de Células B/metabolismo , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/metabolismo , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Espécies Reativas de Oxigênio/metabolismo , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Células Híbridas , Suplementos Nutricionais , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Expressão Gênica/efeitos dos fármacosRESUMO
Over the years, several methods have been developed for the segmentation of cell images. Most of the related techniques operate directly on the raw data (noisy cell samples) of the medical image which leads to adverse effects on the structure of leucocytes because the medical images are affected by multiple distortions (varying illumination, deficient background light intensity, and non-uniform staining). To overcome these problems, we came up with an improved solution that performs the qualitative enhancement of cell images for the smooth extraction of cell-nucleus. Although various segmentation methods have adopted an image improvement operation in practice. These methods also amplify the magnitude of image noise which leads to over-sampling and under-sampling of data points. This mis-labelling of data points is minimized by the developed approach which adopts a collaborative fusion strategy (CNN and Nuclear-norm approach) for the qualitative improvement of cell images. The enhanced cell samples were forwarded to the U-net (deep learning model) model for the semantic segmentation of cell images. The performance evaluation of the model was performed on three biomedical cell imaging datasets, which include the ALL-IDB (99.89% accuracy, 99.51% recall, and 99.01% precision), CellaVision (99.68% accuracy, 98.75% precision, and 97.94% specificity) and JTSC (98.45% accuracy, 97.42% precision, and 97.21% specificity) dataset. The results were compared with the state-of-art methods in which the adopted hybrid approach has overpowered the related techniques in the quantitative and qualitative domains.
Assuntos
Núcleo Celular , Processamento de Imagem Assistida por Computador , Processamento de Imagem Assistida por Computador/métodos , Células HíbridasRESUMO
BACKGROUND: Cancer's hallmark feature is its ability to evolve, leading to metastasis and recurrence. Although genetic mutations and epigenetic changes have been implicated, they don't fully explain the leukocytic traits that many cancers develop. Cell fusion between cancer and somatic cells, particularly macrophages, has been suggested as an alternative pathway for cancer cells to obtain new traits by acquiring exogenous genetic material. METHODS: This study aims to investigate the potential biological outcomes of tumor-myeloid cell fusion by generating tumor-macrophage hybrid cells. Two clones with markedly different tumorigenicity were selected, and RNA-seq was used to compare their RNA expressions with that of the control cells. Based on the results that the hybrid cells showed differential activation in several upstream regulator pathways that impact their biological behaviors, the hybrid cells' abilities to recruit stromal cells and establish angiogenesis as well as their cell cycle distributions were investigated through in vitro and in vivo studies. RESULTS: Although both hybrid clones demonstrated p53 activation and reduced growth rates, they exhibited distinct cell cycle distributions and ability to grow in vivo. Notably, while one clone was highly tumorigenic, the other showed little tumorigenicity. Despite these differences, both hybrid clones were potent environmental modifiers, exhibiting significant abilities to recruit stromal and immune cells and establish angiogenesis. CONCLUSIONS: The study revealed that tumor-somatic cell fusion is a potent environmental modifier that can modulate tumor survival and evolution, despite its relatively low occurrence. These findings suggest that tumor-somatic cell fusion could be a promising target for developing new cancer therapies. Furthermore, this study provides an experimental animal platform to investigate cancer-myeloid fusion and highlights the potential role of tumor-somatic cell fusion in modulating the tumor environment.
Assuntos
Neoplasias , Animais , Neoplasias/genética , Neoplasias/patologia , Células Híbridas/patologia , Fusão Celular , Comunicação Celular , Macrófagos/patologiaRESUMO
BACKGROUND: Bone metastasis is the leading cause of death in patients with prostate cancer (PCa) and currently has no effective treatment. Disseminated tumor cells in bone marrow often obtain new characteristics to cause therapy resistance and tumor recurrence. Thus, understanding the status of disseminated prostate cancer cells in bone marrow is crucial for developing a new treatment. METHODS: We analyzed the transcriptome of disseminated tumor cells from a single cell RNA-sequencing data of PCa bone metastases. We built a bone metastasis model through caudal artery injection of tumor cells, and sorted the tumor hybrid cells by flow cytometry. We performed multi-omics analysis, including transcriptomic, proteomic and phosphoproteomic analysis, to compare the difference between the tumor hybrid cells and parental cells. In vivo experiments were performed to analyze the tumor growth rate, metastatic and tumorigenic potential, drug and radiation sensitivity in hybrid cells. Single cell RNA-sequencing and CyTOF were performed to analyze the impact of hybrid cells on tumor microenvironment. RESULTS: Here, we identified a unique cluster of cancer cells in PCa bone metastases, which expressed myeloid cell markers and showed a significant change in pathways related to immune regulation and tumor progression. We found that cell fusion between disseminated tumor cells and bone marrow cells can be source of these myeloid-like tumor cells. Multi-omics showed the pathways related to cell adhesion and proliferation, such as focal adhesion, tight junction, DNA replication, and cell cycle, were most significantly changed in these hybrid cells. In vivo experiment showed hybrid cells had a significantly increased proliferative rate, and metastatic potential. Single cell RNA-sequencing and CyTOF showed tumor-associated neutrophils/monocytes/macrophages were highly enriched in hybrid cells-induced tumor microenvironment with a higher immunosuppressive capacity. Otherwise, the hybrid cells showed an enhanced EMT phenotype with higher tumorigenicity, and were resistant to docetaxel and ferroptosis, but sensitive to radiotherapy. CONCLUSION: Taken together, our data demonstrate that spontaneous cell fusion in bone marrow can generate myeloid-like tumor hybrid cells that promote the progression of bone metastasis, and these unique population of disseminated tumor cells can provide a potential therapeutic target for PCa bone metastasis.
Assuntos
Neoplasias Ósseas , Neoplasias da Próstata , Humanos , Masculino , Medula Óssea/patologia , Proteômica , Recidiva Local de Neoplasia/patologia , Neoplasias da Próstata/patologia , Neoplasias Ósseas/metabolismo , Células Híbridas/metabolismo , Células Híbridas/patologia , Células da Medula Óssea/patologia , RNA/metabolismo , Linhagem Celular Tumoral , Metástase Neoplásica/patologia , Microambiente TumoralRESUMO
The intrinsic features and functions of platelets and mesenchymal stem cells (MSCs) indicate their great potential in the treatment of intracerebral hemorrhage (ICH). However, neither of them can completely overcome ICH because of the stealth process and the complex pathology of ICH. Here, we fabricate hybrid cells for versatile and highly efficient ICH therapy by fusing MSCs with platelets and loading with lysophosphatidic acid-modified PbS quantum dots (LPA-QDs). The obtained LPA-QDs@FCs (FCs = fusion cells) not only inherit the capabilities of both platelets and MSCs but also exhibit clearly enhanced proliferation activated by LPA. After systemic administration, many proliferating LPA-QDs@FCs rapidly accumulate in ICH areas for responding to the vascular damage and inflammation and then efficiently prevent both the primary and secondary injuries of ICH but with no obvious side effects. Moreover, the treatment process can be tracked by near-infrared II fluorescence imaging with highly spatiotemporal resolution, providing a promising solution for ICH therapy.
Assuntos
Hemorragia Cerebral , Células-Tronco Mesenquimais , Ratos , Animais , Humanos , Ratos Sprague-Dawley , Células Híbridas/patologia , Proliferação de CélulasRESUMO
This study aimed to demonstrate the feasibility of generating tumor cell vaccine models by single-cell surgery in a microfluidic device that integrates one-to-one electrofusion, shear flow reseparation, and on-device culture. The device was microfabricated from polydimethylsiloxane (PDMS) and consisted of microorifices (aperture size: â¼3 µm) for one-to-one fusion, and microcages for on-device culture. Using the device, we could achieve one-to-one electrofusion of leukemic plasmacytoid dendritic cells (DC-like cells) and Jurkat cells with a fusion efficiency of â¼ 80%. Fusion via the narrow microorifices allowed DC-like cells to acquire cytoplasmic contents of the Jurkat cells while preventing nuclei mixing. After fusion, the DC-like cells were selectively reseparated from the Jurkat cells by shear flow application to generate tumor nuclei-free antigen-recipient DC-like (tarDC-like) cells. When cultured as single cells on the device, these cells could survive under gentle medium perfusion with a median survival time of 11.5 h, although a few cells could survive longer than 36 h. Overall, this study demonstrates single-cell surgery in a microfluidic device for potential generation of dendritic cell vaccines which are uncontaminated with tumor nucleic materials. We believe that this study will inspire the generation of safer tumor cell vaccines for cancer immunotherapy.
Assuntos
Neoplasias , Humanos , Células Híbridas , Fusão Celular , Células Dendríticas/patologia , Antígenos de Neoplasias , Citoplasma , Dispositivos Lab-On-A-ChipRESUMO
Circulating tumor cells and hybrid cells formed by the fusion of tumor cells with normal cells are leading players in metastasis and have prognostic relevance. This study applies single-cell RNA sequencing to profile CD45-negative and CD45-positive circulating epithelial cells (CECs) in nonmetastatic breast cancer patients. CECs are represented by transcriptionally-distinct populations that include both aneuploid and diploid cells. CD45- CECs are predominantly aneuploid, but one population contained more diploid than aneuploid cells. CD45+ CECs mostly diploid: only two populations have aneuploid cells. Diploid CD45+ CECs annotated as different immune cells, surprisingly harbored many copy number aberrations, and positively correlated to tumor grade. It is noteworthy that cancer-associated signaling pathways areabundant only in one aneuploid CD45- CEC population, which may represent an aggressive subset of circulating tumor cells. Thus, CD45- and CD45+ CECs are highly heterogeneous in breast cancer patients and include aneuploid cells, which are most likely circulating tumor and hybrid cells, respectively, and diploid cells. DNA ploidy analysis can be an effective instrument for identifying tumor and hybrid cells among CECs. Further follow-up study is needed to determine which subsets of circulating tumor and hybrid cells contribute to breast cancer metastasis.
Assuntos
Neoplasias da Mama , Células Neoplásicas Circulantes , Humanos , Feminino , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Células Neoplásicas Circulantes/patologia , Células Epiteliais/patologia , Aneuploidia , Células Híbridas/patologiaRESUMO
Fusion among different cell populations represents a rare process that is mediated by both intrinsic and extracellular events. Cellular hybrid formation is relayed by orchestrating tightly regulated signaling pathways that can involve both normal and neoplastic cells. Certain important cell merger processes are often required during distinct organismal and tissue development, including placenta and skeletal muscle. In a neoplastic environment, however, cancer cell fusion can generate new cancer hybrid cells. Following survival during a subsequent post-hybrid selection process (PHSP), the new cancer hybrid cells express different tumorigenic properties. These can include elevated proliferative capacity, increased metastatic potential, resistance to certain therapeutic compounds, and formation of cancer stem-like cells, all of which characterize significantly enhanced tumor plasticity. However, many parts within this multi-step cascade are still poorly understood. Aside from intrinsic factors, cell fusion is particularly affected by extracellular conditions, including an inflammatory microenvironment, viruses, pH and ionic stress, hypoxia, and exosome signaling. Accordingly, the present review article will primarily highlight the influence of extracellular events that contribute to cell fusion in normal and tumorigenic tissues.
Assuntos
Carcinogênese , Células-Tronco Neoplásicas , Humanos , Fusão Celular , Linhagem Celular Tumoral , Células Híbridas , Carcinogênese/metabolismo , Células-Tronco Neoplásicas/metabolismo , Microambiente TumoralRESUMO
Triboelectric nanogenerators (TENGs) and dielectric elastomer generators (DEGs) are potentially promising energy conversion technologies, but they still have limitations due to their own intrinsic characteristics, including the low energy output of TENGs caused by the air breakdown effect, and external polarization voltage requirement for DEGs, which severely limit their practical applications. Herein, coupling TENG with DEG is proposed to build a mutual beneficial self-excitation hybrid generator (named TDHG) for harvesting distributed and low-quality mechanical energy (high entropy energy). Experimental results demonstrate that the output charges of this TDHG are enhanced by fivefold of that of the conventional charge-excitation TENG, and continuous operation of DEG is also realized by simple mechanical triggering. More importantly, owing to the high peak power contributed by TENG and the long output pulse duration guaranteed by DEG, the TDHG realizes a much higher energy conversion efficiency of 32% in comparison to either the TENG (3.6%) or DEG (13.2%). This work proposes a new design concept for hybridized energy harvester toward highly efficient mechanical energy harvesting.
Assuntos
Entropia , Frequência Cardíaca , Células Híbridas , Fenômenos FísicosRESUMO
Cardiac contractility assessment is of immense importance for the development of new therapeutics and their safe transition into clinical stages. While human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) hold promise to serve as a human-relevant model in preclinical phases of drug discovery and safety pharmacology, their maturity is still controversial in the scientific community and under constant development. We present a hybrid contractility and impedance/extracellular field potential (EFP) technology, adding significant pro-maturation features to an industry-standard 96-well platform. The impedance/EFP system monitors cellular functionality in real-time. Besides the beat rate of contractile cells, the electrical impedance spectroscopy readouts detect compound-induced morphological changes like cell density and integrity of the cellular monolayer. In the other component of the hybrid cell analysis system, the cells are cultured on bio-compliant membranes that mimic the mechanical environment of real heart tissue. This physiological environment supports the maturation of hiPSC-CMs in vitro, leading to more adult-like contractile responses including positive inotropic effects after treatment with isoproterenol, S-Bay K8644, or omecamtiv mecarbil. Parameters such as the amplitude of contraction force (mN/mm2) and beat duration also reveal downstream effects of compounds with influence on electrophysiological properties and calcium handling. The hybrid system provides the ideal tool for holistic cell analysis, allowing preclinical cardiac risk assessment beyond the current perspectives of human-relevant cell-based assays.
Assuntos
Células-Tronco Pluripotentes Induzidas , Adulto , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Miócitos Cardíacos/metabolismo , Contração Miocárdica , Fenômenos Eletrofisiológicos , Células Híbridas , Células CultivadasRESUMO
Selective differentiation of CD4+ T helper (Th) cells into specialized subsets such as Th1 and Th2 cells is a key element of the adaptive immune system driving appropriate immune responses. Besides those canonical Th-cell lineages, hybrid phenotypes such as Th1/2 cells arise in vivo, and their generation could be reproduced in vitro. While master-regulator transcription factors like T-bet for Th1 and GATA-3 for Th2 cells drive and maintain differentiation into the canonical lineages, the transcriptional architecture of hybrid phenotypes is less well understood. In particular, it has remained unclear whether a hybrid phenotype implies a mixture of the effects of several canonical lineages for each gene, or rather a bimodal behavior across genes. Th-cell differentiation is a dynamic process in which the regulatory factors are modulated over time, but longitudinal studies of Th-cell differentiation are sparse. Here, we present a dynamic transcriptome analysis following Th-cell differentiation into Th1, Th2, and Th1/2 hybrid cells at 3-h time intervals in the first hours after stimulation. We identified an early bifurcation point in gene expression programs, and we found that only a minority of ~20% of Th cell-specific genes showed mixed effects from both Th1 and Th2 cells on Th1/2 hybrid cells. While most genes followed either Th1- or Th2-cell gene expression, another fraction of ~20% of genes followed a Th1 and Th2 cell-independent transcriptional program associated with the transcription factors STAT1 and STAT4. Overall, our results emphasize the key role of high-resolution longitudinal data for the characterization of cellular phenotypes.
Assuntos
Células Th1 , Células Th2 , Diferenciação Celular/genética , Células Híbridas , Ativação LinfocitáriaRESUMO
Mitochondrial cloning is a promising approach to achieve homoplasmic mitochondrial DNA (mtDNA) mutations. We previously developed a microfluidic device that performs single mitochondrion transfer from a mtDNA-intact cell to a mtDNA-less (ρ0) cell by promoting cytoplasmic connection through a microtunnel between them. In the present study, we described a method for generating transmitochondrial cybrids using the microfluidic device. After achieving mitochondrial transfer between HeLa cells and thymidine kinase-deficient ρ0143B cells using the microfluidic device, selective culture was carried out using a pyruvate and uridine (PU)-absent and 5-bromo-2'-deoxyuridine-supplemented culture medium. The resulting cells contained HeLa mtDNA and 143B nuclei, but both 143B mtDNA and HeLa nuclei were absent in these cells. Additionally, these cells showed lower lactate production than parent ρ0143B cells and disappearance of PU auxotrophy for cell growth. These results suggest successful generation of transmitochondrial cybrids using the microfluidic device. Furthermore, we succeeded in selective harvest of generated transmitochondrial cybrids under a PU-supplemented condition by removing unfused ρ0 cells with puromycin-based selection in the microfluidic device.
Assuntos
DNA Mitocondrial , Dispositivos Lab-On-A-Chip , Citoplasma/metabolismo , DNA Mitocondrial/genética , Células HeLa , Humanos , Células Híbridas , Mitocôndrias/genéticaRESUMO
Cancer is one of the most common diseases worldwide, and its treatment is associated with many challenges such as drug and radioresistance and formation of metastases. These difficulties are due to tumor heterogeneity, which has many causes. One may be the cell fusion, a process that is relevant to both physiological (e.g., wound healing) and pathophysiological (cancer and viral infection) processes. This literature review aimed to summarize the existing data on the hybrid/atypical forms of circulating cancer cells and their role in tumor progression. For that, the bioinformatics search in universal databases, such as PubMed, NCBI, and Google Scholar was conducted by using the keywords "hybrid cancer cells", "cancer cell fusion", etc. In this review the latest information related to the hybrid tumor cells, theories of their genesis, characteristics of different variants with data from our own researches are presented. Many aspects of the hybrid cell research are still in their infancy. However, with the level of knowledge already accumulated, circulating hybrids such as CAML and CHC could be considered as promising biomarkers of cancerous tumors, and even more as a new approach to cancer treatment.
Assuntos
Células Neoplásicas Circulantes , Contagem de Células , Fusão Celular , Humanos , Células Híbridas/patologia , Células Neoplásicas Circulantes/patologiaRESUMO
Mitochondria play important roles in the regulation of key cellular processes, including energy metabolism, oxidative stress response, and signaling towards cell death or survival, and are distinguished by carrying their own genome (mtDNA). Mitochondrial dysfunction has emerged as a prominent cellular mechanism involved in neurodegeneration, including Parkinson's disease (PD), a neurodegenerative movement disorder, characterized by progressive loss of dopaminergic neurons and the occurrence of proteinaceous Lewy body inclusions. The contribution of mtDNA variants to PD pathogenesis has long been debated and is still not clearly answered. Cytoplasmic hybrid (cybrid) cell models provided evidence for a contribution of mtDNA variants to the PD phenotype. However, conclusive evidence of mtDNA mutations as genetic cause of PD is still lacking. Several models have shown a role of somatic, rather than inherited mtDNA variants in the impairment of mitochondrial function and neurodegeneration. Accordingly, several nuclear genes driving inherited forms of PD are linked to mtDNA quality control mechanisms, and idiopathic as well as familial PD tissues present increased mtDNA damage. In this review, we highlight the use of cybrids in this PD research field and summarize various aspects of how and to what extent mtDNA variants may contribute to the etiology of PD.