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Nature Communications Jun 2022Precision oncology continues to challenge the "one-size-fits-all" dogma. Under the precision oncology banner, cancer patients are screened for molecular tumor... (Review)
Review
Precision oncology continues to challenge the "one-size-fits-all" dogma. Under the precision oncology banner, cancer patients are screened for molecular tumor alterations that predict treatment response, ideally leading to optimal treatments. Functional assays that directly evaluate treatment efficacy on the patient's cells offer an alternative and complementary tool to improve the accuracy of precision oncology. Unfortunately, traditional Petri dish-based assays overlook much tumor complexity, limiting their potential as predictive functional biomarkers. Here, we review past applications of microfluidic systems for precision medicine and discuss the present and potential future role of functional microfluidic assays as treatment predictors.
Topics: Biological Assay; Humans; Microfluidics; Neoplasms; Precision Medicine
PubMed: 35654785
DOI: 10.1038/s41467-022-30384-7 -
Micromachines Feb 2019In a forest of a hundred thousand trees, no two leaves are alike. Similarly, no two cells in a genetically identical group are the same. This heterogeneity at the... (Review)
Review
In a forest of a hundred thousand trees, no two leaves are alike. Similarly, no two cells in a genetically identical group are the same. This heterogeneity at the single-cell level has been recognized to be vital for the correct interpretation of diagnostic and therapeutic results of diseases, but has been masked for a long time by studying average responses from a population. To comprehensively understand cell heterogeneity, diverse manipulation and comprehensive analysis of cells at the single-cell level are demanded. However, using traditional biological tools, such as petri-dishes and well-plates, is technically challengeable for manipulating and analyzing single-cells with small size and low concentration of target biomolecules. With the development of microfluidics, which is a technology of manipulating and controlling fluids in the range of micro- to pico-liters in networks of channels with dimensions from tens to hundreds of microns, single-cell study has been blooming for almost two decades. Comparing to conventional petri-dish or well-plate experiments, microfluidic single-cell analysis offers advantages of higher throughput, smaller sample volume, automatic sample processing, and lower contamination risk, etc., which made microfluidics an ideal technology for conducting statically meaningful single-cell research. In this review, we will summarize the advances of microfluidics for single-cell manipulation and analysis from the aspects of methods and applications. First, various methods, such as hydrodynamic and electrical approaches, for microfluidic single-cell manipulation will be summarized. Second, single-cell analysis ranging from cellular to genetic level by using microfluidic technology is summarized. Last, we will also discuss the advantages and disadvantages of various microfluidic methods for single-cell manipulation, and then outlook the trend of microfluidic single-cell analysis.
PubMed: 30717128
DOI: 10.3390/mi10020104 -
Journal of Molecular Cell Biology Aug 2020Gastroesophageal cancers are leading causes of cancer death. Our attempts at adopting molecularly based treatment approaches have been slow and ineffective even though... (Review)
Review
Gastroesophageal cancers are leading causes of cancer death. Our attempts at adopting molecularly based treatment approaches have been slow and ineffective even though we begin to identify specific targetable gene mutations and pathways. It is clear that we should no longer treat all gastroesophageal cancers as a homogeneous disease, which is what we do when we use non-specific chemotherapy. However, we currently cannot monitor successful gene/pathway targeting, nor understand how/when tumors develop resistance, nor predict which patients will derive maximal benefit. To improve outcomes, we must precisely detail the heterogeneity of these tumors to then individualize cancer therapy as well as develop novel avenues to study and predict treatment effects in individual patients. To this end, patient-derived organoids, in which tumor cells from individual patients are grown in a Petri dish, are a new versatile system that allows for timely expandability, detailed molecular characterization, and genetic manipulation with the promise of enabling predictive assessment of treatment response. In this review, we will explore the development and basic techniques for organoid generation, and discuss the current and potential future applications of this exciting technology to study the basic science of carcinogenesis and to predict/guide cancer patient care in the clinics.
Topics: Animals; Carcinogenesis; Esophageal Neoplasms; Humans; Organoids; Stomach Neoplasms; Translational Research, Biomedical
PubMed: 32652008
DOI: 10.1093/jmcb/mjaa035 -
Histology and Histopathology Nov 2015Any tissue is made up of a heterogeneous mix of spatially distributed cell types. In response to any (patho) physiological cue, responses of each cell type in any given... (Review)
Review
Any tissue is made up of a heterogeneous mix of spatially distributed cell types. In response to any (patho) physiological cue, responses of each cell type in any given tissue may be unique and cannot be homogenized across cell-types and spatial co-ordinates. For example, in response to myocardial infarction, on one hand myocytes and fibroblasts of the heart tissue respond differently. On the other hand, myocytes in the infarct core respond differently compared to those in the peri-infarct zone. Therefore, isolation of pure targeted cells is an important and essential step for the molecular analysis of cells involved in the progression of disease. Laser capture microdissection (LCM) is powerful to obtain a pure targeted cell subgroup, or even a single cell, quickly and precisely under the microscope, successfully tackling the problem of tissue heterogeneity in molecular analysis. This review presents an overview of LCM technology, the principles, advantages and limitations and its down-stream applications in the fields of proteomics, genomics and transcriptomics. With powerful technologies and appropriate applications, this technique provides unprecedented insights into cell biology from cells grown in their natural tissue habitat as opposed to those cultured in artificial petri dish conditions.
Topics: Animals; Biomarkers; Cell Separation; Gene Expression Profiling; Genetic Markers; Genomics; Humans; Laser Capture Microdissection; Proteomics
PubMed: 25892148
DOI: 10.14670/HH-11-622 -
Biomedicines Apr 2024Stroke is a common neurological disorder, the second leading cause of death, and the third leading cause of disability. Unfortunately, the only approved drug for it is... (Review)
Review
Stroke is a common neurological disorder, the second leading cause of death, and the third leading cause of disability. Unfortunately, the only approved drug for it is tissue plasminogen, but the therapeutic window is limited. In this context, preclinical studies are relevant to better dissect the underlying mechanisms of stroke and for the drug screening of potential therapies. Brain organoids could be relevant in this setting. They are derived from pluripotent stem cells or isolated organ progenitors that differentiate to form an organ-like tissue, exhibiting multiple cell types that self-organize to form a structure not unlike the organ in vivo. Brain organoids mimic many key features of early human brain development at molecular, cellular, structural, and functional levels and have emerged as novel model systems that can be used to investigate human brain diseases including stroke. Brain organoids are a promising and powerful tool for ischemic stroke studies; however, there are a few concerns that need to be addressed, including the lack of vascularization and the many cell types that are typically present in the human brain. The aim of this review is to discuss the potential of brain organoids as a novel model system for studying ischemic stroke, highlighting both the advantages and disadvantages in the use of this technology.
PubMed: 38672231
DOI: 10.3390/biomedicines12040877 -
Cells May 2023Controversial reports have suggested that SARS-CoV E and 3a proteins are plasma membrane viroporins. Here, we aimed at better characterizing the cellular responses...
Controversial reports have suggested that SARS-CoV E and 3a proteins are plasma membrane viroporins. Here, we aimed at better characterizing the cellular responses induced by these proteins. First, we show that expression of SARS-CoV-2 E or 3a protein in CHO cells gives rise to cells with newly acquired round shapes that detach from the Petri dish. This suggests that cell death is induced upon expression of E or 3a protein. We confirmed this by using flow cytometry. In adhering cells expressing E or 3a protein, the whole-cell currents were not different from those of the control, suggesting that E and 3a proteins are not plasma membrane viroporins. In contrast, recording the currents on detached cells uncovered outwardly rectifying currents much larger than those observed in the control. We illustrate for the first time that carbenoxolone and probenecid block these outwardly rectifying currents; thus, these currents are most probably conducted by pannexin channels that are activated by cell morphology changes and also potentially by cell death. The truncation of C-terminal PDZ binding motifs reduces the proportion of dying cells but does not prevent these outwardly rectifying currents. This suggests distinct pathways for the induction of these cellular events by the two proteins. We conclude that SARS-CoV-2 E and 3a proteins are not viroporins expressed at the plasma membrane.
Topics: Cricetinae; Animals; SARS-CoV-2; Cricetulus; COVID-19; Cell Membrane; CHO Cells
PubMed: 37296595
DOI: 10.3390/cells12111474 -
EJNMMI Physics Apr 2023Monte Carlo (MC) simulation in Nuclear Medicine is a powerful tool for modeling many physical phenomena which are difficult to track or measure directly. MC simulation...
PURPOSE
Monte Carlo (MC) simulation in Nuclear Medicine is a powerful tool for modeling many physical phenomena which are difficult to track or measure directly. MC simulation in SPECT/CT imaging is particularly suitable for optimizing the quantification of activity in a patient, and, consequently, the absorbed dose to each organ. To do so, validating MC results with real data acquired with gamma camera is mandatory. The aim of this study was the validation of the calibration factor (CF) and the recovery coefficient (RC) obtained with SIMIND Monte Carlo code for modeling a Siemens Symbia Intevo Excel SPECT-CT gamma camera to ensure optimal [Formula: see text]Tc and [Formula: see text]Lu SPECT quantification.
METHODS
Phantom experiments using [Formula: see text]Tc and [Formula: see text]Lu have been performed to measure spatial resolution and sensitivity, as well as to evaluate the CF and RC from acquired data. The geometries used for 2D planar imaging were (1) Petri dish and (2) capillary source while for 3D volumetric imaging were (3) a uniform filled cylinder phantom and (4) a Jaszczack phantom with spheres of different volumes. The experimental results have been compared with the results obtained from Monte Carlo simulations performed in the same geometries.
RESULTS
Comparison shows good accordance between simulated and experimental data. The measured planar spatial resolution was 8.3[Formula: see text] mm for [Formula: see text]Tc and 11.8±0.6 mm for [Formula: see text]Lu. The corresponding data obtained by SIMIND for [Formula: see text]Tc was 7.8±0.1 mm, while for [Formula: see text]Lu was 12.4±0.4 mm. The CF was 110.1±5.5 cps/MBq for Technetium and 18.3±1.0 cps/MBq for Lutetium. The corresponding CF obtained by SIMIND for [Formula: see text]Tc was 107.3±0.3 cps/MBq, while for [Formula: see text]Lu 20.4±0.7 cps/MBq. Moreover, a complete curve RCs vs Volume (ml) both for Technetium and Lutetium was determined to correct the PVE for all volumes of clinical interest. In none of the cases, a RC coefficient equal to 100 was found.
CONCLUSIONS
The validation of quantification parameters shows that SIMIND can be used for simulating both gamma camera planar and SPECT images of Siemens Symbia Intevo using [Formula: see text]Tc and [Formula: see text]Lu radionuclides for different medical purposes and treatments.
PubMed: 37029829
DOI: 10.1186/s40658-023-00547-6 -
Journal of Ophthalmology 2020To develop a method for the rapid isolation of rat RPE cells with high yield and maintain its epithelial state in modified culture system.
OBJECTIVE
To develop a method for the rapid isolation of rat RPE cells with high yield and maintain its epithelial state in modified culture system.
METHODS
The eyeballs were incubated with dispase. The retina was isolated with RPE attached and cut into several pieces. Following a brief incubation in growth medium, large RPE sheets can be harvested rapidly. RPE cells were divided into four groups and cultured for several weeks, that is, (1) in cell culture dishes with 10% FBS containing medium (CC dish-FBS), (2) in petri dishes with 10% FBS containing medium (Petri dish-FBS), (3) in cell culture dishes with N2 and B27 containing medium (CC dish-N2B27), and (4) in petri dishes with N2 and B27 containing medium (Petri dish-N2B27). Morphological and biological characteristics were investigated using light microscopy, Q-PCR, and western blot.
RESULTS
The retina would curl inwardly during the growth medium incubation period, releasing RPE sheets in the medium. Compared with low density group (5,000 cells/cm), RPE cells plated at high density (15,000 cells/cm) can maintain RPE morphology for a more extended period. Meanwhile, plating RPE cells at low density significantly reduced the expression of RPE cell type-specific genes (RPE65, CRALBP, and bestrophin) and increased the expression of EMT-related genes (N-cadherin, fibronectin, and -SMA), in comparison with the samples from the high density group. The petri dish culture condition reduced cell adhesion and thus inhibited RPE cell proliferation. As compared with other culture conditions, RPE cells in the petri dish-N2B27 condition could maintain RPE phenotype with increased expression of RPE-specific genes and decreased expression of EMT-related genes. The AKT/mTOR pathway was also decreased in petri dish-N2B27 condition.
CONCLUSION
The current study provided an alternative method for easy isolation of RPE cells with high yield and maintenance of its epithelial morphology in the petri dish-N2B27 condition.
PubMed: 32855817
DOI: 10.1155/2020/4892978 -
Brain Pathology (Zurich, Switzerland) Mar 2016Amyotrophic Lateral Sclerosis (ALS) is a complex multifactorial disorder, characterized by motor neuron loss with involvement of several other cell types, including... (Review)
Review
Amyotrophic Lateral Sclerosis (ALS) is a complex multifactorial disorder, characterized by motor neuron loss with involvement of several other cell types, including astrocytes, oligodendrocytes and microglia. Studies in vivo and in in vitro models have highlighted that the contribution of non-neuronal cells to the disease is a primary event and ALS pathogenesis is driven by both cell-autonomous and non-cell autonomous mechanisms. The advancements in genetics and in vitro modeling of the past 10 years have dramatically changed the way we investigate the pathogenic mechanisms involved in ALS. The identification of mutations in transactive response DNA-binding protein gene (TARDBP), fused in sarcoma (FUS) and, more recently, a GGGGCC-hexanucleotide repeat expansion in chromosome 9 open reading frame 72 (C9ORF72) and their link with familial ALS have provided new avenues of investigation and hypotheses on the pathophysiology of this devastating disease. In the same years, from 2007 to present, in vitro technologies to model neurological disorders have also undergone impressive developments. The advent of induced pluripotent stem cells (iPSCs) gave the field of ALS the opportunity to finally model in vitro not only familial, but also the larger part of ALS cases affected by sporadic disease. Since 2008, when the first human iPS-derived motor neurons from patients were cultured in a petri dish, several different techniques have been developed to produce iPSC lines through genetic reprogramming and multiple direct conversion methods have been optimised. In this review, we will give an overview of how human in vitro models have been used so far, what discoveries they have led to since 2007, and how the recent advances in technology combined with the genetic discoveries, have tremendously widened the horizon of ALS research.
Topics: Amyotrophic Lateral Sclerosis; Animals; Cellular Reprogramming Techniques; Drug Evaluation, Preclinical; Embryonic Stem Cells; Humans; Induced Pluripotent Stem Cells
PubMed: 26780562
DOI: 10.1111/bpa.12353 -
World Journal of Stem Cells Oct 2019Mesenchymal stem cells (MSCs) are stromal multipotent stem cells that can differentiate into multiple cell types, including fibroblasts, osteoblasts, chondrocytes,... (Review)
Review
Mesenchymal stem cells (MSCs) are stromal multipotent stem cells that can differentiate into multiple cell types, including fibroblasts, osteoblasts, chondrocytes, adipocytes, and myoblasts, thus allowing them to contribute to the regeneration of various tissues, especially bone tissue. MSCs are now considered one of the most promising cell types in the field of tissue engineering. Traditional petri dish-based culture of MSCs generate heterogeneity, which leads to inconsistent efficacy of MSC applications. Biodegradable and biocompatible polymers, poly(3-hydroxyalkanoates) (PHAs), are actively used for the manufacture of scaffolds that serve as carriers for MSC growth. The growth and differentiation of MSCs grown on PHA scaffolds depend on the physicochemical properties of the polymers, the 3D and surface microstructure of the scaffolds, and the biological activity of PHAs, which was discovered in a series of investigations. The mechanisms of the biological activity of PHAs in relation to MSCs remain insufficiently studied. We suggest that this effect on MSCs could be associated with the natural properties of bacteria-derived PHAs, especially the most widespread representative poly(3-hydroxybutyrate) (PHB). This biopolymer is present in the bacteria of mammalian microbiota, whereas endogenous poly(3-hydroxybutyrate) is found in mammalian tissues. The possible association of PHA effects on MSCs with various biological functions of poly(3-hydroxybutyrate) in bacteria and eukaryotes, including in humans, is discussed in this paper.
PubMed: 31692924
DOI: 10.4252/wjsc.v11.i10.764