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Book: Mitosis

Hinchcliffe, Edward H.

methods and protocols

(Methods in molecular biology ; 2415 ; Springer protocols)

2022

Author's details	edited by Edward H. Hinchcliffe
Series title	Methods in molecular biology ; 2415 Springer protocols
Collection
Keywords	Mitosis ; Mitosis/Research/Methodology
Subject code	571.844
Language	English
Size	xi, 256 Seiten, Illustrationen, Diagramme, 26 cm
Publisher	Humana Press
Publishing place	New York, NY
Publishing country	United States
Document type	Book
HBZ-ID	HT021186621
ISBN	978-1-0716-1903-2 ; 9781071619049 ; 1-0716-1903-9 ; 1071619047
Database	Catalogue ZB MED Medicine, Health

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Article ; Online: H3K27me3 in Diffuse Midline Glioma and Epithelial Ovarian Cancer: Opposing Epigenetic Changes Leading to the Same Poor Outcomes.

Day, Charles A / Hinchcliffe, Edward H / Robinson, James P

Cells

2022 Volume 11, Issue 21

Abstract: Histone post-translational modifications modulate gene expression through epigenetic gene regulation. The core histone H3 family members, H3.1, H3.2, and H3.3, play a central role in epigenetics. H3 histones can acquire many post-translational ... ...

Abstract	Histone post-translational modifications modulate gene expression through epigenetic gene regulation. The core histone H3 family members, H3.1, H3.2, and H3.3, play a central role in epigenetics. H3 histones can acquire many post-translational modifications, including the trimethylation of H3K27 (H3K27me3), which represses transcription. Triple methylation of H3K27 is performed by the histone methyltransferase Enhancer of Zeste Homologue 2 (EZH2), a component of the Polycomb Repressive Complex 2. Both global increases and decreases in H3K27me3 have been implicated in a wide range of cancer types. Here, we explore how opposing changes in H3K27me3 contribute to cancer by highlighting its role in two vastly different cancer types; (1) a form of glioma known as diffuse midline glioma H3K27-altered and (2) epithelial ovarian cancer. These two cancers vary widely in the age of onset, sex, associated mutations, and cell and organ type. However, both diffuse midline glioma and ovarian cancer have dysregulation of H3K27 methylation, triggering changes to the cancer cell transcriptome. In diffuse midline glioma, the loss of H3K27 methylation is a primary driving factor in tumorigenesis that promotes glial cell stemness and silences tumor suppressor genes. Conversely, hypermethylation of H3K27 occurs in late-stage epithelial ovarian cancer, which promotes tumor vascularization and tumor cell migration. By using each cancer type as a case study, this review emphasizes the importance of H3K27me3 in cancer while demonstrating that the mechanisms of histone H3 modification and subsequent gene expression changes are not a one-size-fits-all across cancer types.
MeSH term(s)	Humans ; Female ; Histones/metabolism ; Carcinoma, Ovarian Epithelial/genetics ; Glioma/genetics ; Glioma/pathology ; DNA Methylation ; Epigenesis, Genetic ; Ovarian Neoplasms/genetics
Chemical Substances	Histones
Language	English
Publishing date	2022-10-26
Publishing country	Switzerland
Document type	Journal Article ; Review ; Research Support, U.S. Gov't, Non-P.H.S. ; Research Support, Non-U.S. Gov't ; Research Support, N.I.H., Extramural
ZDB-ID	2661518-6
ISSN	2073-4409 ; 2073-4409
ISSN (online)	2073-4409
ISSN	2073-4409
DOI	10.3390/cells11213376
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article ; Online: Using Microinjection of Mammalian Cultured Cells to Study Cell Division.

Day, Charles / Langfald, Alyssa / Hinchcliffe, Edward H

Methods in molecular biology (Clifton, N.J.)

2021 Volume 2415, Page(s) 105–122

Abstract: The introduction of macromolecules directly into individual cells by microinjection is an important technique for manipulating mitotic cells. mRNA, purified proteins, or concentrated antibodies can all be injected directly into a single cell, and their ... ...

Abstract	The introduction of macromolecules directly into individual cells by microinjection is an important technique for manipulating mitotic cells. mRNA, purified proteins, or concentrated antibodies can all be injected directly into a single cell, and their effects monitored by live-cell imaging. The equipment necessary is relatively simple, and the technique can be easily mastered. Here we describe our microinjection setup, how to microinject cultured mammalian cells in mitosis, and how to analyze those cells by same-cell live and fixed imaging.
MeSH term(s)	Animals ; Cell Line ; Cells, Cultured ; Mammals ; Microinjections/methods ; Mitosis
Language	English
Publishing date	2021-12-22
Publishing country	United States
Document type	Journal Article
ISSN	1940-6029
ISSN (online)	1940-6029
DOI	10.1007/978-1-0716-1904-9_8
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Article: Centrosomes and the art of mitotic spindle maintenance.

Hinchcliffe, Edward H

International review of cell and molecular biology

2014 Volume 313, Page(s) 179–217

Abstract: The assembly of a bipolar spindle lies at the heart of mitotic chromosome segregation. In animal somatic cells, the process of spindle assembly involves multiple complex interactions between various cellular compartments, including an emerging ... ...

Abstract	The assembly of a bipolar spindle lies at the heart of mitotic chromosome segregation. In animal somatic cells, the process of spindle assembly involves multiple complex interactions between various cellular compartments, including an emerging antiparallel microtubule network, microtubule-associated motor proteins and spindle assembly factors, the cell's cortex, and the chromosomes themselves. The result is a dynamic structure capable of aligning pairs of sister chromatids, sensing chromosome misalignment, and generating force to segregate the replicated genome into two daughters. Because the centrosome lies at the center of the array of microtubule minus-ends, and the essential one-to-two duplication of the centrosome prior to mitosis is linked to cell cycle progression, this organelle has long been implicated as a device to generate spindle bipolarity. However, this classic model for spindle assembly is challenged by observations and experimental manipulations demonstrating that acentrosomal cells can and do form bipolar spindles, both mitotic and meiotic. Indeed, recent comprehensive proteomic analysis of centrosome-dependent versus independent mitotic spindle assembly mechanisms reveals a large, common set of genes required for both processes, with very few genes needed to differentiate between the two. While these studies cast doubt on an absolute role for the centrosome in establishing spindle polarity, it is clear that having too few or too many centrosomes results in abnormal chromosome segregation and aneuploidy. Here we review the case both for and against the role of the centrioles and centrosomes in ensuring proper assembly of a bipolar spindle, an essential element in the maintenance of genomic stability.
MeSH term(s)	Animals ; Cell Cycle Proteins/metabolism ; Centrosome/metabolism ; Centrosome/physiology ; Chromosomal Instability ; Chromosome Segregation ; Humans ; Microtubule Proteins/metabolism ; Mitosis ; Spindle Apparatus/metabolism ; Spindle Apparatus/physiology
Chemical Substances	Cell Cycle Proteins ; Microtubule Proteins
Language	English
Publishing date	2014
Publishing country	Netherlands
Document type	Journal Article ; Research Support, Non-U.S. Gov't ; Research Support, U.S. Gov't, Non-P.H.S. ; Review
ZDB-ID	2427220-6
ISSN	1937-6448 ; 0074-7696
ISSN	1937-6448 ; 0074-7696
DOI	10.1016/B978-0-12-800177-6.00006-2
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article: Manipulating cultured mammalian cells for mitosis research.

Day, Charles A / Langfald, Alyssa / Hinchcliffe, Edward H

Methods in cell biology

2020 Volume 158, Page(s) 43–61

Abstract: The study of mitosis has always relied on bulk-preparation biochemistry techniques (Mazia & Dan, 1952), but very early on lent itself to living, single cell microscopic techniques (Inoue, 1953; Taylor, 1959). Here we describe several of the methods used ... ...

Abstract	The study of mitosis has always relied on bulk-preparation biochemistry techniques (Mazia & Dan, 1952), but very early on lent itself to living, single cell microscopic techniques (Inoue, 1953; Taylor, 1959). Here we describe several of the methods used by our lab to study cell division in living cultured cells, including cold-induced mitotic arrest, cold-induced chromosome missegregation, same-cell live and fixed cell imaging, and microinjection of inactivating antibodies. We detail our imaging system based on an upright fluorescent microscope and spinning disk confocal, as well as the customized "HEKS" metal support slide imaging chambers.
MeSH term(s)	Animals ; Cell Culture Techniques/methods ; Cell Line ; Chlorocebus aethiops ; Humans ; Imaging, Three-Dimensional ; Mammals/physiology ; Microinjections ; Mitosis ; Rats ; Research ; Spindle Apparatus/metabolism
Language	English
Publishing date	2020-03-12
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
ISSN	0091-679X
ISSN	0091-679X
DOI	10.1016/bs.mcb.2020.02.001
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article ; Online: Probing cell shape regulation with patterned substratum: requirement of myosin II-mediated contractility.

Mader, Christopher C / Hinchcliffe, Edward H / Wang, Yu-Li

Soft matter

2020 Volume 3, Issue 3, Page(s) 357–363

Abstract: Adherent cells cultured on flat, homogeneous surfaces typically maintain an intact cell body with a polygonal or fan shape, despite active migration and strong mechanical interactions with the substratum. We hypothesized that, in addition to the ... ...

Abstract	Adherent cells cultured on flat, homogeneous surfaces typically maintain an intact cell body with a polygonal or fan shape, despite active migration and strong mechanical interactions with the substratum. We hypothesized that, in addition to the constraint of the surface membrane, an active mechanism may be involved in maintaining the shape and integrity of the cell body particularly where cells encounter complex topographic patterns of guidance cues. To detect if there is a mechanism that constrains cell shape, we plated NIH 3T3 fibroblasts on ring-patterned substrata 8-17.5 microns in width and 53-133 microns in outer diameter. Untreated cells have a limited angular span, encompassing an average of 108 degrees around the ring, even though these cells were able to cover a much larger surface when plated on flat surfaces of the same material. Treatment of 3T3 cells with a myosin II inhibitor, blebbistatin, induced a striking increase in the bending ability, causing cells to cover more than 60% of the ring. Inhibition of the Rho-dependent kinase with Y-27632 caused a similar but smaller increase in the angular span. Our results suggest that cell shape is controlled not only by the passive constraint of the surface membrane but also by an active mechanism driven by myosin II-mediated contractility under the regulation of Rho-dependent kinase. The inward surface tension-like forces allow the cell to maintain its integrity while navigating through complex physiological environments.
Language	English
Publishing date	2020-09-04
Publishing country	England
Document type	Journal Article
ZDB-ID	2191476-X
ISSN	1744-6848 ; 1744-683X
ISSN (online)	1744-6848
ISSN	1744-683X
DOI	10.1039/b606590b
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Article ; Online: H3K27me3 in Diffuse Midline Glioma and Epithelial Ovarian Cancer

Charles A. Day / Edward H. Hinchcliffe / James P. Robinson

Cells, Vol 11, Iss 3376, p

Opposing Epigenetic Changes Leading to the Same Poor Outcomes

2022 Volume 3376

Abstract	Histone post-translational modifications modulate gene expression through epigenetic gene regulation. The core histone H3 family members, H3.1, H3.2, and H3.3, play a central role in epigenetics. H3 histones can acquire many post-translational modifications, including the trimethylation of H3K27 (H3K27me3), which represses transcription. Triple methylation of H3K27 is performed by the histone methyltransferase Enhancer of Zeste Homologue 2 (EZH2), a component of the Polycomb Repressive Complex 2. Both global increases and decreases in H3K27me3 have been implicated in a wide range of cancer types. Here, we explore how opposing changes in H3K27me3 contribute to cancer by highlighting its role in two vastly different cancer types; (1) a form of glioma known as diffuse midline glioma H3K27-altered and (2) epithelial ovarian cancer. These two cancers vary widely in the age of onset, sex, associated mutations, and cell and organ type. However, both diffuse midline glioma and ovarian cancer have dysregulation of H3K27 methylation, triggering changes to the cancer cell transcriptome. In diffuse midline glioma, the loss of H3K27 methylation is a primary driving factor in tumorigenesis that promotes glial cell stemness and silences tumor suppressor genes. Conversely, hypermethylation of H3K27 occurs in late-stage epithelial ovarian cancer, which promotes tumor vascularization and tumor cell migration. By using each cancer type as a case study, this review emphasizes the importance of H3K27me3 in cancer while demonstrating that the mechanisms of histone H3 modification and subsequent gene expression changes are not a one-size-fits-all across cancer types.
Keywords	diffuse midline glioma ; diffuse intrinsic pontine glioma ; epithelial ovarian cancer ; epigenetics ; histone H3 ; H3K27M ; Biology (General) ; QH301-705.5
Subject code	610
Language	English
Publishing date	2022-10-01T00:00:00Z
Publisher	MDPI AG
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: The centrosome and bipolar spindle assembly: does one have anything to do with the other?

Hinchcliffe, Edward H

Cell cycle (Georgetown, Tex.)

2011 Volume 10, Issue 22, Page(s) 3841–3848

Abstract: In vertebrate somatic cells the centrosome functions as the major microtubule-organizing center (MTOC), which splits and separates to form the poles of the mitotic spindle. However, the role of the centriole-containing centrosome in the formation of ... ...

Abstract	In vertebrate somatic cells the centrosome functions as the major microtubule-organizing center (MTOC), which splits and separates to form the poles of the mitotic spindle. However, the role of the centriole-containing centrosome in the formation of bipolar mitotic spindles continues to be controversial. Cells normally containing centrosomes are still able to build bipolar spindles after their centrioles have been removed or ablated. In naturally occurring cellular systems that lack centrioles - such as plant cells and many oocytes - bipolar spindles form in the complete absence of canonical centrosomes. These observations have led to the notion that centrosomes play no role during mitosis. However, recent work has re-examined spindle assembly in the absence of centrosomes, both in cells that naturally lack them, and those that have had them experimentally removed. The results of these studies suggest that an appreciation of microtubule network organization- both before and after nuclear envelope breakdown (NEB) - is the key to understanding the mechanisms that regulate spindle assembly and the generation of bipolarity.
MeSH term(s)	Animals ; Cell Line ; Cell Polarity ; Centrosome/physiology ; Centrosome/ultrastructure ; Chlorocebus aethiops ; Drosophila/cytology ; Drosophila/metabolism ; Drosophila/ultrastructure ; Female ; Mice ; Nuclear Envelope/metabolism ; Nuclear Envelope/ultrastructure ; Oocytes/cytology ; Oocytes/metabolism ; Oocytes/ultrastructure ; Sea Urchins/cytology ; Sea Urchins/metabolism ; Sea Urchins/ultrastructure ; Spindle Apparatus/metabolism ; Spindle Apparatus/physiology ; Spindle Apparatus/ultrastructure
Language	English
Publishing date	2011-11-15
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural
ZDB-ID	2146183-1
ISSN	1551-4005 ; 1538-4101 ; 1554-8627
ISSN (online)	1551-4005
ISSN	1538-4101 ; 1554-8627
DOI	10.4161/cc.10.22.18293
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Article ; Online: It's all about the pentiums: The use, manipulation, and storage of digital microscopy imaging data for the biological sciences.

Hornick, Jessica E / Hinchcliffe, Edward H

Molecular reproduction and development

2015 Volume 82, Issue 7-8, Page(s) 508–517

Abstract: Digital microscopy has revolutionized quantitative imaging, with binary-encoded computer files serving to capture and store imaging data sets for analysis. With the ever-present use of computers to generate and store imaging data, it becomes increasingly ...

Abstract	Digital microscopy has revolutionized quantitative imaging, with binary-encoded computer files serving to capture and store imaging data sets for analysis. With the ever-present use of computers to generate and store imaging data, it becomes increasingly important to understand how these files are created, and how they can be both used and mis-used. This is a particularly important task for the biologist who may have limited background in computer science. Here we discuss some of the basic aspects of digital data storage and use, including file types, storage media, and the choice between commercial and open-source software. Often, open-source software is written by a user or group of users, and then distributed to the scientific community at large. These can be important tools, but there are some hidden costs to this freeware that we will discuss. We will also compare open-source software to commercial imaging software, which is often written for use by non-computer scientists.
MeSH term(s)	Humans ; Image Processing, Computer-Assisted/methods ; Information Storage and Retrieval/methods ; Microscopy ; Software
Language	English
Publishing date	2015-07
Publishing country	United States
Document type	Journal Article ; Review
ZDB-ID	20321-x
ISSN	1098-2795 ; 1040-452X
ISSN (online)	1098-2795
ISSN	1040-452X
DOI	10.1002/mrd.22294
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Article ; Online: Overcoming translational barriers in H3K27-altered diffuse midline glioma: Increasing the drug-tumor residence time.

Power, Erica A / Rechberger, Julian S / Zhang, Liang / Oh, Ju-Hee / Anderson, Jacob B / Nesvick, Cody L / Ge, Jizhi / Hinchcliffe, Edward H / Elmquist, William F / Daniels, David J

Neuro-oncology advances

2023 Volume 5, Issue 1, Page(s) vdad033

Abstract: Background: H3K27-altered diffuse midline glioma (DMG) is the deadliest pediatric brain tumor; despite intensive research efforts, every clinical trial to date has failed. Is this because we are choosing the wrong drugs? Or are drug delivery and other ... ...

Abstract	Background: H3K27-altered diffuse midline glioma (DMG) is the deadliest pediatric brain tumor; despite intensive research efforts, every clinical trial to date has failed. Is this because we are choosing the wrong drugs? Or are drug delivery and other pharmacokinetic variables at play? We hypothesize that the answer is likely a combination, where optimization may result in a much needed novel therapeutic approach. Methods: We used in vitro drug screening, patient samples, and shRNA knockdown models to identify an upregulated target in DMG. A single small molecule protein kinase inhibitor with translational potential was selected for systemic and direct, loco-regional delivery to patient-derived xenografts (PDX) and genetically engineered mouse models (GEMM). Pharmacokinetic studies were conducted in non-tumor bearing rats. Results: Aurora kinase (AK) inhibitors demonstrated strong antitumor effects in DMG drug screens. Additional in vitro studies corroborated the importance of AK to DMG survival. Systemic delivery of alisertib showed promise in subcutaneous PDX but not intracranial GEMM and PDX models. Repeated loco-regional drug administration into the tumor through convection-enhanced delivery (CED) was equally inefficacious, and pharmacokinetic studies revealed rapid clearance of alisertib from the brain. In an effort to increase the drug to tumor residence time, continuous CED over 7 days improved drug retention in the rodent brainstem and significantly extended survival in both orthotopic PDXs and GEMMs. Conclusions: These studies provide evidence for increasing drug-tumor residence time of promising targeted therapies via extended CED as a valuable treatment strategy for DMG.
Language	English
Publishing date	2023-03-27
Publishing country	England
Document type	Journal Article
ZDB-ID	3009682-0
ISSN	2632-2498 ; 2632-2498
ISSN (online)	2632-2498
ISSN	2632-2498
DOI	10.1093/noajnl/vdad033
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