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Trends and Prospects of Flow Cytometry in Cell and Molecular Biology

A special issue of International Journal of Molecular Sciences (ISSN 1422-0067). This special issue belongs to the section "Molecular Biology".

Deadline for manuscript submissions: 15 July 2024 | Viewed by 5263

Special Issue Editors

Prof. Dr. Claudio Ortolani

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Guest Editor
Department of Biomolecular Sciences, University of Urbino, Urbino, Italy
Interests: flow cytometry; hematology; immunology; clinical diagnostics
Special Issues, Collections and Topics in MDPI journals
Prof. Dr. José-Enrique O’Connor

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Guest Editor
Department of Biochemistry, School of Medicine, University of Valencia, 46010 Valencia, Spain
Interests: flow cytometry; cell biology; metabolism; biochemistry; immunology; stem cell biology; macrophage
Special Issues, Collections and Topics in MDPI journals
Dr. Monia Lenzi

E-Mail Website
Guest Editor
Department of Pharmacy and Biotechnology, Alma Mater Studiorum University of Bologna, 40126 Bologna, Italy
Interests: chemoprevention; genotoxicity; in vitro toxicology; bioactive molecules; natural compounds; flow cytometry
Special Issues, Collections and Topics in MDPI journals

Special Issue Information

Dear Colleagues,

Flow Cytometry is a methodology we consider mature but paradoxically keeps evolving rapidly, continuously offering new challenges even to those who think they know it deeply.

As proof of what has just been said, we could recall the new experimental fields for its application that have emerged in the last period.

Born as a technique for automatically evaluating cell features, such as physical parameters and the content of nucleic acids, it has become a multifaceted tool in a series of different sectors and an invaluable instrument in studying the biochemistry and molecular features of the cells.

On the one hand, the continuously growing availability of a panoply of fluorescent probes allows us to investigate a series of cellular activities in a new way; on the other hand, the appearance of new technologies, such as Raman cytometry and others, promises the arrival of an era in which it will be possible to acquire information on metabolism and cell functions through a label-free approach.

The ambition of this issue of the Journal is to take stock of what is happening in this field and help us understand the new directions toward which Flow Cytometry is moving.

I'm confident that, with the help of all of you, this special number of IJMS will be a success.

Prof. Dr. Claudio Ortolani
Prof. Dr. José-Enrique O’Connor
Dr. Monia Lenzi
Guest Editors

Manuscript Submission Information

Manuscripts should be submitted online at www.mdpi.com by registering and logging in to this website. Once you are registered, click here to go to the submission form. Manuscripts can be submitted until the deadline. All submissions that pass pre-check are peer-reviewed. Accepted papers will be published continuously in the journal (as soon as accepted) and will be listed together on the special issue website. Research articles, review articles as well as short communications are invited. For planned papers, a title and short abstract (about 100 words) can be sent to the Editorial Office for announcement on this website.

Submitted manuscripts should not have been published previously, nor be under consideration for publication elsewhere (except conference proceedings papers). All manuscripts are thoroughly refereed through a single-blind peer-review process. A guide for authors and other relevant information for submission of manuscripts is available on the Instructions for Authors page. International Journal of Molecular Sciences is an international peer-reviewed open access semimonthly journal published by MDPI.

Please visit the Instructions for Authors page before submitting a manuscript. There is an Article Processing Charge (APC) for publication in this open access journal. For details about the APC please see here. Submitted papers should be well formatted and use good English. Authors may use MDPI's English editing service prior to publication or during author revisions.

Keywords

  • biochemistry
  • molecular biology
  • cell function
  • cell metabolism
  • new cytometries
  • new applications
  • new methods
  • microbiology

Published Papers (6 papers)

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Research

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15 pages, 3884 KiB  
Article
Investigating Vα7.2+/CD161− T Cell and MAIT Cell Profiles Using Flow Cytometry in Healthy Subjects and Subjects with Atopic Dermatitis
by Parvind Singh, Krisztian Gaspar, Andrea Szegedi, Laszlo Sajtos, Sandor Barath and Zsuzsanna Hevessy
Int. J. Mol. Sci. 2024, 25(6), 3486; https://doi.org/10.3390/ijms25063486 - 20 Mar 2024
Viewed by 669
Abstract
This study investigates the roles of mucosal-associated invariant T (MAIT) cells and Vα7.2+/CD161− T cells in skin diseases, focusing on atopic dermatitis. MAIT cells, crucial for bridging innate and adaptive immunity, were analyzed alongside Vα7.2+/CD161− T cells [...] Read more.
This study investigates the roles of mucosal-associated invariant T (MAIT) cells and Vα7.2+/CD161− T cells in skin diseases, focusing on atopic dermatitis. MAIT cells, crucial for bridging innate and adaptive immunity, were analyzed alongside Vα7.2+/CD161− T cells in peripheral blood samples from 14 atopic dermatitis patients and 10 healthy controls. Flow cytometry and machine learning algorithms were employed for a comprehensive analysis. The results indicate a significant decrease in MAIT cells and CD69 subsets in atopic dermatitis, coupled with elevated CD38 and polyfunctional MAIT cells producing TNFα and Granzyme B (TNFα+/GzB+). Vα7.2+/CD161− T cells in atopic dermatitis exhibited a decrease in CD8 and IFNγ-producing subsets but an increase in CD38 activated and IL-22-producing subsets. These results highlight the distinctive features of MAIT cells and Vα7.2+/CD161− T cells and their different roles in the pathogenesis of atopic dermatitis and provide insights into their potential roles in immune-mediated skin diseases. Full article
(This article belongs to the Special Issue Trends and Prospects of Flow Cytometry in Cell and Molecular Biology)
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21 pages, 5694 KiB  
Article
Phenotypic Analysis of Hematopoietic Stem and Progenitor Cell Populations in Acute Myeloid Leukemia Based on Spectral Flow Cytometry, a 20-Color Panel, and Unsupervised Learning Algorithms
by Thomas Matthes
Int. J. Mol. Sci. 2024, 25(5), 2847; https://doi.org/10.3390/ijms25052847 - 29 Feb 2024
Viewed by 702
Abstract
The analysis of hematopoietic stem and progenitor cell populations (HSPCs) is fundamental in the understanding of normal hematopoiesis as well as in the management of malignant diseases, such as leukemias, and in their diagnosis and follow-up, particularly the measurement of treatment efficiency with [...] Read more.
The analysis of hematopoietic stem and progenitor cell populations (HSPCs) is fundamental in the understanding of normal hematopoiesis as well as in the management of malignant diseases, such as leukemias, and in their diagnosis and follow-up, particularly the measurement of treatment efficiency with the detection of measurable residual disease (MRD). In this study, I designed a 20-color flow cytometry panel tailored for the comprehensive analysis of HSPCs using a spectral cytometer. My investigation encompassed the examination of forty-six samples derived from both normal human bone marrows (BMs) and patients with acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS) along with those subjected to chemotherapy and BM transplantation. By comparing my findings to those obtained through conventional flow cytometric analyses utilizing multiple tubes, I demonstrate that my innovative 20-color approach enables a more in-depth exploration of HSPC subpopulations and the detection of MRD with at least comparable sensitivity. Furthermore, leveraging advanced analytical tools such as t-SNE and FlowSOM learning algorithms, I conduct extensive cross-sample comparisons with two-dimensional gating approaches. My results underscore the efficacy of these two methods as powerful unsupervised alternatives for manual HSPC subpopulation analysis. I expect that in the future, complex multi-dimensional flow cytometric data analyses, such as those employed in this study, will be increasingly used in hematologic diagnostics. Full article
(This article belongs to the Special Issue Trends and Prospects of Flow Cytometry in Cell and Molecular Biology)
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18 pages, 2099 KiB  
Article
Persistence of Chronic Lymphocytic Leukemia Stem-like Populations under Simultaneous In Vitro Treatment with Curcumin, Fludarabine, and Ibrutinib: Implications for Therapy Resistance
by Àngel Bistué-Rovira, Laura G. Rico, Jorge Bardina, Jordi Juncà, Isabel Granada, Jolene A. Bradford, Michael D. Ward, Roser Salvia, Francesc Solé and Jordi Petriz
Int. J. Mol. Sci. 2024, 25(4), 1994; https://doi.org/10.3390/ijms25041994 - 07 Feb 2024
Viewed by 968
Abstract
Leukemic stem cells (LSCs) possess similar characteristics to normal hematopoietic stem cells, including self-renewal capacity, quiescence, ability to initiate leukemia, and drug resistance. These cells play a significant role in leukemia relapse, persisting even after apparent remission. LSCs were first described in 1994 [...] Read more.
Leukemic stem cells (LSCs) possess similar characteristics to normal hematopoietic stem cells, including self-renewal capacity, quiescence, ability to initiate leukemia, and drug resistance. These cells play a significant role in leukemia relapse, persisting even after apparent remission. LSCs were first described in 1994 by Lapidot et al. Although they have been extensively studied in acute leukemia, more LSC research is still needed in chronic lymphocytic leukemia (CLL) to understand if reduced apoptosis in mature cells should still be considered as the major cause of this disease. Here, we provide new evidence suggesting the existence of stem-like cell populations in CLL, which may help to understand the disease as well as to develop effective treatments. In this study, we identified a potential leukemic stem cell subpopulation using the tetraploid CLL cell line I83. This subpopulation is characterized by diploid cells that were capable of generating the I83 tetraploid population. Furthermore, we adapted a novel flow cytometry analysis protocol to detect CLL subpopulations with stem cell properties in peripheral blood samples and primary cultures from CLL patients. These cells were identified by their co-expression of CD19 and CD5, characteristic markers of CLL cells. As previously described, increased alkaline phosphatase (ALP) activity is indicative of stemness and pluripotency. Moreover, we used this method to investigate the potential synergistic effect of curcumin in combination with fludarabine and ibrutinib to deplete this subpopulation. Our results confirmed the effectiveness of this ALP-based analysis protocol in detecting and monitoring leukemic stem-like cells in CLL. This analysis also identified limitations in eradicating these populations using in vitro testing. Furthermore, our findings demonstrated that curcumin significantly enhanced the effects of fludarabine and ibrutinib on the leukemic fraction, exhibiting synergistic effects (combination drug index, CDI 0.97 and 0.37, respectively). Our results lend support to the existence of potential stem-like populations in CLL cell lines, and to the idea that curcumin could serve as an effective adjuvant in therapies aimed at eliminating these populations and improving treatment efficacy. Full article
(This article belongs to the Special Issue Trends and Prospects of Flow Cytometry in Cell and Molecular Biology)
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Review

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15 pages, 7680 KiB  
Review
SLAM Family Receptors in B Cell Chronic Lymphoproliferative Disorders
by Dominik Kľoc, Slavomír Kurhajec, Mykhailo Huniadi, Ján Sýkora, Tomáš Guman and Marek Šarišský
Int. J. Mol. Sci. 2024, 25(7), 4014; https://doi.org/10.3390/ijms25074014 - 04 Apr 2024
Viewed by 569
Abstract
The signaling lymphocytic activation molecule (SLAM) receptor family (SLAMF) consists of nine glycoproteins that belong to the CD2 superfamily of immunoglobulin (Ig) domain-containing molecules. SLAMF receptors modulate the differentiation and activation of a wide range of immune cells. Individual SLAMF receptors are expressed [...] Read more.
The signaling lymphocytic activation molecule (SLAM) receptor family (SLAMF) consists of nine glycoproteins that belong to the CD2 superfamily of immunoglobulin (Ig) domain-containing molecules. SLAMF receptors modulate the differentiation and activation of a wide range of immune cells. Individual SLAMF receptors are expressed on the surface of hematopoietic stem cells, hematopoietic progenitor cells, B cells, T cells, NK cells, NKT cells, monocytes, macrophages, dendritic cells, neutrophils, and platelets. The expression of SLAMF receptors was studied during normal B cell maturation. Several SLAMF receptors were also detected in cancer cell lines of B-lymphoid origin and in pathological B cells from patients with B cell chronic lymphoproliferative disorders (B-CLPD), the most frequent hematological malignancies in adults. This review summarizes current knowledge on the expression of SLAMF receptors and their adaptor proteins SAP and EAT-2 in B-CLPD. Several SLAMF receptors could be regarded as potential diagnostic and differential diagnostic markers, prognostic factors, and targets for the development of novel drugs for patients with B-CLPD. Full article
(This article belongs to the Special Issue Trends and Prospects of Flow Cytometry in Cell and Molecular Biology)
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19 pages, 1198 KiB  
Review
Applications of Flow Cytometry in Drug Discovery and Translational Research
by Sumana Ullas and Charles Sinclair
Int. J. Mol. Sci. 2024, 25(7), 3851; https://doi.org/10.3390/ijms25073851 - 29 Mar 2024
Viewed by 785
Abstract
Flow cytometry is a mainstay technique in cell biology research, where it is used for phenotypic analysis of mixed cell populations. Quantitative approaches have unlocked a deeper value of flow cytometry in drug discovery research. As the number of drug modalities and druggable [...] Read more.
Flow cytometry is a mainstay technique in cell biology research, where it is used for phenotypic analysis of mixed cell populations. Quantitative approaches have unlocked a deeper value of flow cytometry in drug discovery research. As the number of drug modalities and druggable mechanisms increases, there is an increasing drive to identify meaningful biomarkers, evaluate the relationship between pharmacokinetics and pharmacodynamics (PK/PD), and translate these insights into the evaluation of patients enrolled in early clinical trials. In this review, we discuss emerging roles for flow cytometry in the translational setting that supports the transition and evaluation of novel compounds in the clinic. Full article
(This article belongs to the Special Issue Trends and Prospects of Flow Cytometry in Cell and Molecular Biology)
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Other

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17 pages, 11028 KiB  
Case Report
Flow Cytometry Analysis in Breast Implant-Associated Anaplastic Large Cell Lymphoma: Three Case Reports
by Veronica Davanzo, Alessandra Falda, Paola Fogar, Kathrin Ludwig, Jenny Zuin, Maria Cristina Toffanin, Marco Pizzi, Angelo Paolo Dei Tos and Daniela Basso
Int. J. Mol. Sci. 2024, 25(6), 3518; https://doi.org/10.3390/ijms25063518 - 20 Mar 2024
Viewed by 537
Abstract
Breast Implant-Associated-Anaplastic Large Cell Lymphoma (BIA-ALCL) is a rare T-cell non-Hodgkin lymphoma associated with breast prosthetic implants and represents a diagnostic challenge. The National Comprehensive Cancer Network (NCCN) guidelines, updated in 2024, recommend for diagnosis an integrated work-up that should include cell morphology, [...] Read more.
Breast Implant-Associated-Anaplastic Large Cell Lymphoma (BIA-ALCL) is a rare T-cell non-Hodgkin lymphoma associated with breast prosthetic implants and represents a diagnostic challenge. The National Comprehensive Cancer Network (NCCN) guidelines, updated in 2024, recommend for diagnosis an integrated work-up that should include cell morphology, CD30 immunohistochemistry (IHC), and flow cytometry (FCM). CD30 IHC, although the test of choice for BIA-ALCL diagnosis, is not pathognomonic, and this supports the recommendation to apply a multidisciplinary approach. A close collaboration between pathologists and laboratory professionals allowed the diagnosis of three BIA-ALCLs, presented as case reports, within a series of 35 patients subjected to periprosthetic effusions aspiration from 2018 to 2023. In one case, rare neoplastic cells were identified by FCM, and this result was essential in leading the anatomopathological picture as indicative of this neoplasm. In fact, the distinction between a lymphomatous infiltrate from reactive cells may be very complex in the cytopathology and IHC setting when neoplastic cells are rare. On the other hand, one limitation of FCM analysis is the need for fresh samples. In this study, we provide evidence that a dedicated fixative allows the maintenance of an unaltered CD30 expression on the cell surface for up to 72 h. Full article
(This article belongs to the Special Issue Trends and Prospects of Flow Cytometry in Cell and Molecular Biology)
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