Methods in cell biology最新文献_第10页

Organ-on-chip immunostaining method for three-dimensional identification and study of immune cells responding to drug-treated tumor cells. 器官芯片免疫染色法三维识别和研究免疫细胞对药物治疗肿瘤细胞的反应。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2025-02-05 DOI: 10.1016/bs.mcb.2025.01.005

Francesco Noto, Adele De Ninno, Mario Falchi, Luca Businaro, Giovanna Schiavoni, Fabrizio Mattei

{"title":"Organ-on-chip immunostaining method for three-dimensional identification and study of immune cells responding to drug-treated tumor cells.","authors":"Francesco Noto, Adele De Ninno, Mario Falchi, Luca Businaro, Giovanna Schiavoni, Fabrizio Mattei","doi":"10.1016/bs.mcb.2025.01.005","DOIUrl":"https://doi.org/10.1016/bs.mcb.2025.01.005","url":null,"abstract":"Epigenetic deregulation is implied in cancer initiation and resistance to antitumor drugs. In melanoma, aberrant DNA hypermethylation is frequently observed, resulting in the silencing of several genes involved in cell cycle regulation, apoptosis, tumor growth and drug resistance. DNA hypomethylating agents have been recently evaluated in both preclinical and clinical studies as a strategy to restore tumor suppressor genes and to increase immune recognition by tumors, highlighting their potential in pre-clinical models of melanoma. Advanced microfluidic system for the culture of complex three-dimensional cell, tissue and organ models have proven utility for oncoimmunology studies and drug testing. Here we present a protocol employing ad hoc fabricated microfluidic devices to reproduce a three-dimensional (3D) tumor microenvironment (TME) to study two aspects of the crosstalk between immune and cancerous cells under the effect of Decitabine (DAC), a DNA methyl transferase inhibitor (DNMTi). First, we evaluated the preferential migration of immune cells towards treated and non-treated melanoma cells inside the chip. Next, we identified a specific subpopulation of migrated immune cells, with an on-chip immunostaining protocol resulting in the acquisition and evaluation of 3D images on a Laser-Scanning Confocal Microscopy (LSCM) station for in-depth characterization of tumor-immune interactions. This protocol may find broad application for pre-clinical drug testing in cancer studies.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"196 ","pages":"209-223"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144667974","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Precise measurement of motor neuron dysfunction in Drosophila ALS model via climbing assay and leg imaging. 攀爬试验和腿部成像技术精确测量果蝇ALS模型运动神经元功能障碍。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2025-03-12 DOI: 10.1016/bs.mcb.2025.02.008

Yanan Wei, Hongyu Miao, Hadi Najafi, Woo Jae Kim

{"title":"Precise measurement of motor neuron dysfunction in Drosophila ALS model via climbing assay and leg imaging.","authors":"Yanan Wei, Hongyu Miao, Hadi Najafi, Woo Jae Kim","doi":"10.1016/bs.mcb.2025.02.008","DOIUrl":"https://doi.org/10.1016/bs.mcb.2025.02.008","url":null,"abstract":"Amyotrophic lateral sclerosis (ALS) is a severe neurodegenerative disorder characterized by progressive degeneration of motor neurons, leading to muscle weakness, paralysis, and death. While there is a plethora of studies focusing on many aspects of ALS, the pathogenesis of this disease is not well understood, and effective treatments are scarce. Drosophila melanogaster is a powerful model organism for studying ALS due to its genetic tractability and its evolutionarily conserved cellular and molecular processes which are also shared between the fly and human. Here, we introduce two simple and cost-effective methodologies for assessing motor neuron dysfunction in Drosophila: (1) Fast Inexpensive Climbing Test (FICT), and (2) Economical Leg Fluorescence Imaging (ELFI). These methods are established based on using basic equipment and straightforward procedures, making them accessible and applicable in various research and educational settings. FICT provides a non-invasive and high-throughput measure of motor dysfunction, while ELFI allows for direct visualization of fluorescently labeled cells in the Drosophila leg, facilitating the study of cell-cell communications in vivo. Our approach emphasizes the importance of both neuronal and glial contributions to ALS pathogenesis, offering valuable insights for the development of novel therapeutic strategies. These methods democratize access to ALS research tools, promoting global scientific collaboration and advancing our understanding of this devastating disease.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"197 ","pages":"127-148"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145033665","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Multidimensional profiling of cancer microenvironments in FFPE tissues by imaging mass cytometry. 肿瘤微环境在FFPE组织的多维谱成像细胞术。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-11-19 DOI: 10.1016/bs.mcb.2024.10.014

Marieke E Ijsselsteijn, Noel F C C de Miranda

引用次数: 0

High-throughput sequencing technologies for cancer genomics. 癌症基因组学的高通量测序技术。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2025-03-05 DOI: 10.1016/bs.mcb.2025.02.018

Garima, Meenakshi Dhanawat, Kashish Wilson, Pramila Chaubey

{"title":"High-throughput sequencing technologies for cancer genomics.","authors":"Garima, Meenakshi Dhanawat, Kashish Wilson, Pramila Chaubey","doi":"10.1016/bs.mcb.2025.02.018","DOIUrl":"https://doi.org/10.1016/bs.mcb.2025.02.018","url":null,"abstract":"In investigations about transcriptomics, epigenomics, and genomics, high-throughput sequencing technologies have become indispensable. Several hundred million of DNA molecules may be sequenced at once thanks to high throughput sequencing (HTS) technologies, which can simultaneously analyze many DNA molecules. Traditionally, sequencing information has been clarified utilizing a low throughput technique known as Sanger sequencing. This added value makes it feasible to employ HTS to generate tremendous amounts of data, which enhances the comprehension of the transcriptome and genetic fingerprints of cells during various stages of evolution and pathology. By identifying somatic changes, morphological deviations, and repetitive changes across the human genome, techniques such as whole exome sequencing (WES) and whole genome sequencing (WGS) provide information about cancer formation as well as prospective therapies. Identifying tumor biology and discovering biomarkers rely on the examination of aberrant networks and variations in gene expression that RNA DNA sequencing, or RNA-Seq, offers. By identifying rare cell kinds and their function in carcinogenesis, the sequencing of one cell offers illumination on the wide range of cells observed across tumors. Metagenomics and chromatin immunoprecipitation sequencing (ChIP-Seq) delivers essential knowledge by discovering alterations that affect the epigenetic configuration and the microenvironment that accompanies tumors. Integrating these recent developments will allow the development of personalized treatments that use unique genetic traits to determine every cancer patient, offering more individualized treatments. The revolutionary implications of high-throughput genome sequencing for cancer research and treatment are addressed in this book chapter, particularly concerning cancer precision as well as effective treatment outcomes.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"198 ","pages":"103-133"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145206893","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Quantitative pre-clinical imaging of hypoxia and vascularity using MRI and PET. 定量临床前成像缺氧和血管的MRI和PET。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-11-19 DOI: 10.1016/bs.mcb.2024.10.016

Georgia Kanli, Selma Boudissa, Radovan Jirik, Tom Adamsen, Heidi Espedal, Hans Olav Rolfsnes, Frits Thorsen, Jesus Pacheco-Torres, Bassam Janji, Olivier Keunen

{"title":"Quantitative pre-clinical imaging of hypoxia and vascularity using MRI and PET.","authors":"Georgia Kanli, Selma Boudissa, Radovan Jirik, Tom Adamsen, Heidi Espedal, Hans Olav Rolfsnes, Frits Thorsen, Jesus Pacheco-Torres, Bassam Janji, Olivier Keunen","doi":"10.1016/bs.mcb.2024.10.016","DOIUrl":"10.1016/bs.mcb.2024.10.016","url":null,"abstract":"During hypoxia, tissues are subjected to an inadequate oxygen supply, disrupting the balance needed to maintain normal function. This deficiency can occur due to reduced oxygen delivery caused by impaired blood flow or a decline in the blood's ability to carry oxygen. In tumors, hypoxia and vascularization play crucial roles, shaping their microenvironments and influencing cancer progression, response to treatment and metastatic potential. This chapter provides guidance on the use of non-invasive imaging methods including Positron Emission Tomography and Magnetic Resonance Imaging to study tumor oxygenation in pre-clinical settings. These imaging techniques offer valuable insights into tumor vascularity and oxygen levels, aiding in understanding tumor behavior and treatment effects. For example, PET imaging uses tracers such as [18F]-fluoromisonidazole (FMISO) to visualize hypoxic areas within tumors, while MRI complements this with anatomical and functional images. Although directly assessing tumor hypoxia with MRI remains challenging, techniques like Blood Oxygen Level Dependent (BOLD) and Dynamic Contrast-Enhanced MRI (DCE-MRI) provide valuable information. BOLD can track changes in oxygen levels during oxygen challenges, while DCE-MRI offers real-time access to perfusion and vessel permeability data. Integrating data from these imaging modalities can help assess oxygen supply, refine treatment strategies, enhance therapeutic effectiveness, and ultimately improve patient outcomes.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"191 ","pages":"289-328"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143008160","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Single-cell RNA flow cytometry to assess intratumoral production of cytokines/chemokines. 单细胞RNA流式细胞术评估肿瘤内细胞因子/趋化因子的产生。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-11-20 DOI: 10.1016/bs.mcb.2024.10.013

Khiem C Lam, Romina S Goldszmid

引用次数: 0

Flow cytometry-based monitoring of myeloid-derived suppressor cells in the peripheral blood of patients with solid tumors. 基于流式细胞术的实体瘤患者外周血髓源性抑制细胞监测。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-11-13 DOI: 10.1016/bs.mcb.2024.10.009

Anna-Jasmina Donaubauer, Ilka Scheer, Rainer Fietkau, Udo S Gaipl, Benjamin Frey

{"title":"Flow cytometry-based monitoring of myeloid-derived suppressor cells in the peripheral blood of patients with solid tumors.","authors":"Anna-Jasmina Donaubauer, Ilka Scheer, Rainer Fietkau, Udo S Gaipl, Benjamin Frey","doi":"10.1016/bs.mcb.2024.10.009","DOIUrl":"10.1016/bs.mcb.2024.10.009","url":null,"abstract":"Myeloid-derived suppressor cells (MDSCs) ameliorate inflammation by inhibiting T cell responses. In pathological conditions, such as autoimmunity, chronic infections or cancer they accumulate in the periphery. In cancer, MDSCs can also be part of the tumor microenvironment and are associated with a worse prognosis and limited response to immunotherapy. Nowadays attempts are made to specifically target MDSCs in cancer therapy. Still, the role of MDSCs in standard cancer treatment modalities, such as radiotherapy remains mostly elusive. Here, we describe a flow cytometry-based method to determine and monitor monocytic and granulocytic-derived MDSCs directly from whole blood in an easy, fast and reliable assay. As specific surface markers for MDSCs are lacking, the assay follows a gating strategy that excludes successively the main immune cells types and analyzes the remaining events for a set of molecules that are expressed on MDSCs. This assay is especially appropriate for longitudinal analyses and clinical trials and is suitable for being integrated into more complex immunophenotyping panels to generate a comprehensive immune status.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"191 ","pages":"135-150"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143008125","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Metallic nanoparticles biodistribution for the study of lymphoma in animal models. 金属纳米粒子在淋巴瘤动物模型研究中的生物分布。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-08-23 DOI: 10.1016/bs.mcb.2024.07.004

Barbara Chalhoub, Víctor Franco Puntes, Laura Mondragón

{"title":"Metallic nanoparticles biodistribution for the study of lymphoma in animal models.","authors":"Barbara Chalhoub, Víctor Franco Puntes, Laura Mondragón","doi":"10.1016/bs.mcb.2024.07.004","DOIUrl":"10.1016/bs.mcb.2024.07.004","url":null,"abstract":"T cell lymphoma constitutes a complex group of diseases, characterized by heterogeneous molecular features and clinical symptoms, and a dismal outcome no matter the therapeutic strategy chosen. In an attempt to improve patients' survival chances, treatment combinations (chemotherapy, radiotherapy, immunotherapy, gene therapy and thermotherapy) have been tested for their synergistic effects that may dramatically improve outcomes and reduce the side effects of each single modality treatment when therapeutic effects add up while side effects are distributed. In this context, nanoscale drug delivery agents have been developed and exploited to enhance the release of drugs in the treatment of several diseases, showing potential benefits in terms of pharmaceutical flexibility, selectivity, dose reduction and minimization of adverse effects. Inorganic materials (i.e., metal nanoparticles) can be used as imaging and radiotherapy agents demonstrating that nanoparticle-based therapies can combine and act as \"precision medicine\" for targeting tumors while leaving healthy tissue intact. Therefore, nanoparticles (NPs) appear as ideal platforms for multimodal therapy constituting a more than promising strategy in the search of effective combined treatments for T cell lymphoma. In our laboratory, we aim at validating these therapeutic strategies making use of metal NPs able to provide a diagnostic and therapeutic effect at the same time. Validation of the synthesized NPs will be possible thanks to the availability of an in vivo T cell lymphoma animal model also developed in the lab. Here, we describe basic protocols for the administration and biodistribution studies in solid tumors which could be of significant help for future therapies development and follow-up.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"192 ","pages":"159-180"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143039859","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Assessment of the primary cancer cell secretome using amino acid-analog labeling. 用氨基酸类似物标记评估原发性癌细胞分泌组。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-12-15 DOI: 10.1016/bs.mcb.2024.11.002

Annemieke C Bouwman, Antoinette van Weverwijk, Onno B Bleijerveld, Liesbeth Hoekman, Bob J Ignacio, Kimberly M Bonger, Karin E de Visser

{"title":"Assessment of the primary cancer cell secretome using amino acid-analog labeling.","authors":"Annemieke C Bouwman, Antoinette van Weverwijk, Onno B Bleijerveld, Liesbeth Hoekman, Bob J Ignacio, Kimberly M Bonger, Karin E de Visser","doi":"10.1016/bs.mcb.2024.11.002","DOIUrl":"https://doi.org/10.1016/bs.mcb.2024.11.002","url":null,"abstract":"It is well established that reciprocal communication between cancer cells and other cells in the tumor microenvironment plays a crucial role in cancer progression and therapy response. There are multiple ways by which cells communicate, including direct cell-cell contact and the secretion of soluble mediators. The secretome of cancer cells contains valuable information to disentangle the complex conversation that is happening between cancer cells and neighboring or distant cells such as immune cells, fibroblasts and endothelial cells. Here, we provide a workflow of mapping the cancer cell secretome in an unbiased way using amino acid-analog labeling in combination with mass spectrometry. The generation of single cells from fresh tumors, isolation of primary cancer cells from a complex multi-cellular pool, and the detection of newly synthesized proteins that are secreted into the medium is described in detailed protocols. Using this experimental pipeline the secretome of cancer cells across different tumors can be determined, paving the way to unravel cell-cell communication networks in the tumor microenvironment, which may uncover novel therapeutic targets.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"196 ","pages":"43-65"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144667962","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Phenotypic assessment of dendritic cell maturation by cost-effective custom ELISA assays. 树突状细胞成熟的表型评估成本效益定制ELISA检测。

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2025-01-31 DOI: 10.1016/bs.mcb.2025.01.011

Peng Liu, Yuhong Pan, Misha Mao, Guido Kroemer, Oliver Kepp, Liwei Zhao

{"title":"Phenotypic assessment of dendritic cell maturation by cost-effective custom ELISA assays.","authors":"Peng Liu, Yuhong Pan, Misha Mao, Guido Kroemer, Oliver Kepp, Liwei Zhao","doi":"10.1016/bs.mcb.2025.01.011","DOIUrl":"https://doi.org/10.1016/bs.mcb.2025.01.011","url":null,"abstract":"Dendritic cells (DCs) are professional antigen-presenting cells that are pivotal in operating tumor immunosurveillance and orchestrating anticancer immune responses. Endowed with phagocytic and migratory capacities, DCs can capture and process tumor antigens, travel to lymphoid organs and prime naïve T cells, altogether leading to the clonal expansion of cytotoxic T lymphocytes (CTLs) that can specifically target and lyse cancer cells. Additionally, DCs contribute to the formation of immunological memory, ensuring durable therapeutic effects and long-term surveillance against tumor recurrence. Upon antigen engagement, DCs undergo a maturation process characterized by the production of specific cytokines as well as the increased expression of costimulatory molecules and chemokine receptors on their surface. Here we propose a panel of custom sandwich enzyme linked immunosorbent assays (ELISAs) for assessing DC maturation via the precise quantification of cytokines. This economic approach achieves high precision and reproducibility and can be readily applied in labs equipped with basic molecular cell biology facilities. With appropriate automatization, this protocol can be employed for high-throughput screening campaigns for the discovery of DC maturation modulators.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"196 ","pages":"271-290"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144667975","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0