journal
https://read.qxmd.com/read/38700834/a-coculture-system-for-modeling-intestinal-epithelial-fibroblast-crosstalk
#1
JOURNAL ARTICLE
Rebecca F Lee, Mei-Lan Li, Maria Figetakis, Kaelyn Sumigray
Epithelial organoid monoculture is a powerful tool to model stem cell dynamics in vitro. However, extensive efforts have recently revealed various niche players and their significant roles in regulating epithelial stem cells. Among these niche components, fibroblasts have been heavily recognized in the field as a critical niche signal secretor. Thus, understanding the roles of fibroblasts in epithelial dynamics has become increasingly relevant and crucial. This propels the development of approaches to coculture epithelial 3D organoids with fibroblasts to model epithelial-fibroblast crosstalk in vitro...
May 4, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38700833/microfluidic-chip-fabrication-for-tumor-cell-3d-culture-based-on-microwell-arrays
#2
JOURNAL ARTICLE
Lanjie Lei
Compared with traditional 2D cell culture, 3D cell culture more closely resembles the original state of cells in vivo and enables the establishment of in vivo-like microenvironments and cell-cell interactions, thereby providing valuable cellular materials for numerous studies. The direct establishment of in vitro patient tumor models can enhance drug testing, cancer research, and individualized precision therapy. In this study, we propose a microfluidic chip based on microwell arrays for 3D tumor cell culture...
May 4, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38700832/isolation-of-human-progenitor-epidermal-cells-on-collagen-type-iv-and-analysis-of-their-markers-with-flow-cytometry-and-pcr-methods
#3
JOURNAL ARTICLE
Milena Deptuła, Małgorzata Zawrzykraj, Paulina Słonimska, Michał Pikuła
Epidermal stem cells, located in the skin, together with keratinocytes are transplanted in regenerative therapies, e.g., for the treatment of burns or other wounds. Here, we describe the protocol of their enzymatic isolation from human skin. It includes separation of the epidermis form the dermis by incubation with dispase followed by cell isolation for epidermis by digestion with trypsin. Cell isolated with this method can be seeded on collagen IV-coated dishes. The methods of analysis of epidermal stem cells markers (e...
May 4, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38647865/production-of-neuroepithelial-organoids-from-human-induced-pluripotent-stem-cells-for-mimicking-early-neural-tube-development
#4
JOURNAL ARTICLE
Chunling Tang, Xinghui Wang, Eileen Gentleman, Nicholas A Kurniawan
Organoids have emerged as robust tools for unravelling the mechanisms that underly tissue development. They also serve as important in vitro systems for studying fundamentals of stem cell behavior and for building advanced disease models. During early development, a crucial step in the formation of the central nervous system is patterning of the neural tube dorsal-ventral (DV) axis. Here we describe a simple and rapid culture protocol to produce human neuroepithelial (NE) cysts and DV-patterned organoids from single human-induced pluripotent stem cells (hiPSCs)...
April 23, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38647864/assessment-of-stiffness-dependent-autophagosome-formation-and-apoptosis-in-embryonal-rhabdomyosarcoma-tumor-cells
#5
JOURNAL ARTICLE
Serap Sezen, Sevin Adiguzel, Atefeh Zarepour, Arezoo Khosravi, Joseph W Gordon, Saeid Ghavami, Ali Zarrabi
Remodeling of the extracellular matrix (ECM) eventually causes the stiffening of tumors and changes to the microenvironment. The stiffening alters the biological processes in cancer cells due to altered signaling through cell surface receptors. Autophagy, a key catabolic process in normal and cancer cells, is thought to be involved in mechano-transduction and the level of autophagy is probably stiffness-dependent. Here, we provide a methodology to study the effect of matrix stiffness on autophagy in embryonal rhabdomyosarcoma cells...
April 23, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38647863/measurement-of-myonuclear-accretion-in-vitro-and-in-vivo
#6
JOURNAL ARTICLE
Lola Lessard, Audrey Saugues, Julien Gondin, Rémi Mounier, Anita Kneppers
Adult skeletal muscle stem cells (MuSC) are the regenerative precursors of myofibers and also have an important role in myofiber growth, adaptation, and maintenance by fusing to the myofibers-a process referred to as "myonuclear accretion." Due to a focus on MuSC function during regeneration, myofibers remain a largely overlooked component of the MuSC niche influencing MuSC fate. Here, we describe a method to directly measure the rate of myonuclear accretion in vitro and in vivo using ethynyl-2'-deoxyuridine (EdU)-based tracing of MuSC progeny...
April 23, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38647862/organoid-culture-of-different-intestinal-segments-from-human-and-mouse
#7
JOURNAL ARTICLE
Yalong Wang, Ronghui Tan, Ye-Guang Chen
The intestine comprises distinct segments, each characterized by unique cell populations and functions. Intestinal organoids faithfully replicate the cellular composition and functions of the intestine. Over the past decade, the organoid model has garnered considerable attention for its application in investigation of organ development, renewal and functional performance. While the organoid culture systems for mouse small intestine and human large intestine have widely adopted, a comparison summary for different segments of the human or mouse intestine is lacking...
April 23, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38647861/methods-for-generating-self-organizing-human-patterned-heart-organoids-using-pluripotent-stem-cells
#8
JOURNAL ARTICLE
Brett Volmert, Aitor Aguirre
Organoids derived from pluripotent stem cells exhibit notable similarities to organ development in vitro. Nonetheless, cardiac organoids generated to date possess immature phenotypes and are unable to model the full spectrum of heart development and disease. Here, we describe the developmental maturation of human heart organoids by controlled exposure to metabolic and hormonal factors over a 10-day period, mirroring key stages of human cardiac development and resulting in significant molecular, cellular, morphological, and functional changes...
April 23, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38634995/protocol-for-testing-the-effects-of-ssdna-aptamer-in-hela-and-mcf-7
#9
JOURNAL ARTICLE
Haregewoin Bezu Woldekidan, Zandile Nxumalo, Mutsa M Takundwa, Adugna Abdi Woldesemayat, Deepak B Thimiri Govinda Raj
Conventional approaches for treating tumors encompass chemotherapy, radiotherapy, and surgery. However, these methods come with their limitations when applied in clinical practice. Aptamers are often referred to as "chemical antibodies" and consist of short DNA or RNA molecules, designed to bind to a wide range of targets, including proteins or nucleic acid structures. They exhibit strong affinities and remarkable specificity for their target molecules, making them capable of functioning as therapeutic agents to directly impede tumor cell proliferation...
April 19, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38607594/evaluation-of-mitochondrial-phagy-mitophagy-in-human-non-small-adenocarcinoma-tumor-cells
#10
JOURNAL ARTICLE
Javad Alizadeh, Simone C da Silva Rosa, Marco Cordani, Saeid Ghavami
Non-small cell lung cancer (NSCLC) is a predominant form of lung cancer characterized by its aggressive nature and high mortality rate, primarily due to late-stage diagnosis and metastatic spread. Recent studies underscore the pivotal role of mitophagy, a selective form of autophagy targeting damaged or superfluous mitochondria, in cancer biology, including NSCLC. Mitophagy regulation may influence cancer cell survival, proliferation, and metastasis by modulating mitochondrial quality and cellular energy homeostasis...
April 13, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38578577/regeneration-of-volumetric-muscle-loss-using-mscs-encapsulated-in-prp-derived-fibrin-microbeads
#11
JOURNAL ARTICLE
Şükran Şeker, Özge Lalegül-Ülker, Ayşe Eser Elçin, Yaşar Murat Elçin
Volumetric muscle loss (VML) is one of the major types of soft tissue injury frequently encountered worldwide. In case of VML, the endogenous regenerative capacity of the skeletal muscle tissue is usually not sufficient for complete healing of the damaged area resulting in permanent functional musculoskeletal impairment. Therefore, the development of new tissue engineering approaches that will enable functional skeletal muscle regeneration by overcoming the limitations of current clinical treatments for VML injuries has become a critical goal...
April 6, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38578576/characterization-of-bitter-taste-receptor-dependent-autophagy-in-oral-epithelial-cells
#12
JOURNAL ARTICLE
Nisha Singh, Saeid Ghavami, Prashen Chelikani
Microbial dysbiosis is an important trigger in the development of oral diseases. Oral keratinocytes or gingival epithelial cells (GECs) offer protection against various microbial insults. Recent studies suggest that GECs expressed higher level of bitter taste receptor 14 (T2R14) compared to other taste receptors and toll-like receptors and act as innate immune sentinels. Macroautophagy or autophagy is a cellular conserved process involved in the regulation of host innate immune responses against microbial infection...
April 6, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38507214/standardizing-chondrocyte-isolation-and-articular-cartilage-decellularization-a-versatile-bioink-for-tissue-engineering-applications
#13
JOURNAL ARTICLE
Upasna Upadhyay, Kamma Srinivasulu, Lakshmi Kiran Chelluri
The extracellular matrix (ECM) is a noncellular component of tissues that provides structural and biochemical support to cells. The purpose of decellularization is to provide a tissue-specific niche to preserve the architecture, composition, and signaling molecules of the ECM. The current protocol discusses the standardization of chondrocyte isolation and the preparation of acellular ECM as a bioink additive from human native articular cartilage. Isolated chondrocytes with bioink additives provide a tissue-specific microenvironment...
March 21, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38507213/contribution-of-artificial-intelligence-to-the-identification-of-protein-protein-interactions-a-case-study-on-par-3-and-its-partner-adapter-molecule-crk
#14
JOURNAL ARTICLE
Fabio D'Amico
Protein-protein interactions (PPIs) are known to be involved in most cellular functions, and a detailed knowledge of such interactions is essential for studying their role in normal and pathological conditions. Significant progress is being made in the identification of PPIs through advances in computational methods. In particular, the AlphaFold2 machine learning-based model has been shown to accelerate drug discovery process by predicting the 3D structure of protein complexes. In this chapter, a straightforward protocol for predicting interprotein interactions between PAR-3 and its protein partner adapter molecule crk is provided...
March 21, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38507212/isolation-of-murine-neonatal-and-adult-osteomacs-to-examine-their-role-in-the-hematopoietic-niche
#15
JOURNAL ARTICLE
Safa F Mohamad, Melissa A Kacena
Maintenance of hematopoietic stem cell (HSC) function is an orchestrated event between multiple cell types, and crosstalk between these cell types is an essential part of HSC regulation. Among the cell groups of the niche involved in this process are a group of bone-resident macrophages known as osteomacs (OM). Previously, it was demonstrated that OM and osteoblasts contained within neonatal calvarial cells are critical to maintain hematopoietic function. Additionally, interactions between neonatal calvarial cells and megakaryocytes further enhance this hematopoietic activity...
March 21, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38507211/simplified-quantification-of-progenitor-zone-size-an-indicator-of-germ-stem-cell-niche-activity-in-the-nematode-caenorhabditis-elegans
#16
JOURNAL ARTICLE
Sarah R Fausett, Caroline A Laury, Rachel E Magallon, Christian Braendle
Germ stem cell (GSC) niches are fundamental for the maintenance of the immortal germ cell lineage across generations. In the nematode Caenorhabditis elegans, the simple GSC system has served as an important model for understanding stem cell biology and underlying genetic architecture. GSC niche activity in C. elegans is highly sensitive to subtle environmental and genetic variation. Quantifying variation in the C. elegans GSC niche is therefore essential; however, most methods to do so remain labor-intensive and time-consuming when screening large numbers of individuals...
March 21, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38502468/principles-of-indirect-co-culture-method-using-transwell
#17
JOURNAL ARTICLE
Mohammad Rasouli, Fatemeh Safari
The co-culture method is a simple type of cell culture method used to evaluate the effects of communication between various types of cells in an in vitro setting. In the co-culture method, two or more eukaryotic cell types, or eukaryotic and prokaryotic cells, are cultured together. The co-culture method reflects in vivo cell behaviors and thereby emerges as a pivotal technique with diverse applications in cancer research and cell biology. Two categories of co-culture methods (indirect methods and direct methods) are well known...
March 20, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38499918/evaluation-of-autophagy-in-conjunctival-fibroblasts
#18
JOURNAL ARTICLE
Parvaneh Mehrbod, Paola Brun, Umberto Rosani, Andrea Leonardi, Saeid Ghavami
Vernal keratoconjunctivitis (VKC) is a serious eye allergy characterized by poorly understood pathogenic mechanisms and a lack of effective treatments. Autophagy, a process involved in both triggering and suppressing immune and inflammatory responses, plays a role in VKC's pathophysiology. Understanding autophagy's involvement in VKC could lead to new treatment possibilities, such as utilizing specific topical substances to induce or inhibit autophagy and prevent severe complications of this eye condition. In our current protocol, we present a robust methodology established in our laboratory for studying autophagy in primary conjunctival fibroblasts...
March 19, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38499917/approaches-to-study-wound-induced-hair-neogenesis-wihn
#19
JOURNAL ARTICLE
Abhik Dutta, Dyuti Saha, Colin Jamora
Embryonic wound repair proceeds with complete regeneration of the tissue without any scar formation, whereas tissue repair in adults usually results in scars and the tissue does not completely regain its preinjured state. Wound-induced hair neogenesis (WIHN) in adult rodents results in de novo hair follicle formation in the center of large wounds, mimicking regeneration processes seen in fetal tissue. The investigation of WIHN therefore provides a unique quantitative framework for scrutinizing the mechanistic underpinnings of regenerative repair, which can have clinical implications in the context of scarless healing...
March 19, 2024: Methods in Molecular Biology
https://read.qxmd.com/read/38446407/measurement-of-autophagic-activity-in-cancer-cells-with-flow-cytometric-analysis-using-cyto-id-staining
#20
JOURNAL ARTICLE
Merve Şansaçar, Emel Başak Gencer Akçok
Autophagy is an evolutionarily conserved process providing the energy that cells need to survive, especially in stress situations, through catabolic processes. Considering the dual role of autophagy in cancer cells depending on the cellular context, it is crucial to comprehend the effect of drug candidates put forward to prevent cancer through the autophagy pathway. The CYTO-ID® Autophagy Detection Kit allows a rapid, specific and quantitative measurement of autophagic activity at the cellular level using a 488 nm-excitable green fluorescent detection reagent via flow cytometer...
March 7, 2024: Methods in Molecular Biology
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