Primary cell culture

Short description: The culture of cells from fresh tissues

Primary cell culture

Primary interstinal organoid culture

Anatomical terminology

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Primary cell culture is the ex vivo culture of cells freshly obtained from a multicellular organism, as opposed to the culture of immortalized cell lines. In general, primary cell cultures are considered more representative of in vivo tissues than cell lines, and this is recognized legally in some countries such as the UK (Human Tissue Act 2004).[1] However, primary cells require adequate substrate and nutrient conditions to thrive and after a certain number of divisions they acquire a senescent phenotype, leading to irreversible cell cycle arrest.[2] The generation of cell lines stems from these two reasons. Primary cells can become immortalized either spontaneously (e.g. HeLa cells) or by genetic modification (e.g. HEK cells), at which point they become cell lines which can be subcultured indefinitely.[3]

Because of their requirements for viability, primary cell cultures did not become widespread until the 2000s. These cultures present several advantages over cell lines, including a better representation of the cellular heterogeneity of tissues, a more faithful transcriptomic and proteomic profile (especially when cultured in 3D) and more realistic functional responses, including drug responses.[4][5][6] In contrast, immortalized cell lines are known to become homogeneous through the natural selection of specific subpopulations, to undergo genetic drift and to acquire genetic aberrations. In many cases, cell lines have been misidentified, contaminated with other cells or infected with Mycoplasma, small intracellular bacteria that went undetected for decades.[4][7]

When whole or partial tissues are isolated and maintained ex vivo, the procedure is termed primary tissue culture. More specific terms include organotypic culture,[8] tissue slices[9] and explants.[10]

Neuronal primary cell cultures are cells collected from the brain of an organism. For example they can be used when examining substances effect on cell viability, which can further on be potential treatments for brain deficits.[11]

## Contents

* 1 Monolayer cultures
* 1.1 Monolayer cultures for the personalized therapy in aggressive thyroid cancer of follicular origin
* 2 Cell culture protocol
* 3 See also
* 4 References

## Monolayer cultures

"Monolayer cultures" refer to cell cultures where cells are grown in a single, flat layer on the surface of a culture dish or substrate. In a monolayer culture, cells adhere to the substrate and spread out in a two-dimensional arrangement. This type of cell culture is commonly used in laboratory settings for various purposes, including research, drug testing, and biotechnology.

Key features of monolayer cultures include:

Two-Dimensional Growth:Cells in monolayer cultures grow in a single plane, adhering to the surface of the culture vessel. This flat arrangement allows for easy observation and manipulation of individual cells.[12]

Adherence to Substrate:The cells attach to the surface of the culture dish or flask, and their growth and behavior can be influenced by the characteristics of the substrate.on the other words In cell culture, adherence to substrate describes a cell's capacity to adhere to a surface and proliferate.Many elements, including surface energy, substrate topography, and roughness, mediate the process of cell attachment.[13] The study of artificial polymer surfaces with varying chemical, topological, and mechanical cues that regulate cell activities has focused attention on the interaction between external surfaces and cells.[13] In a study that was published in the journal RSC Advances in 2021, the impact of roughness and surface energy on cell adhesion and growth was examined.The most advantageous circumstances for effective cell adhesion, development, and proliferation were discovered by the study to be moderate surface energy and intermediate roughness ratio.[13]

Cell Proliferation:Cells in monolayer cultures can undergo cell division and proliferation. This feature is crucial for experimental studies and the production of a larger number of cells for subsequent analyses.[12]

Observation and Imaging:The two-dimensional nature of monolayer cultures makes it convenient for microscopic observation and imaging. Researchers can easily visualize the cells, study their morphology, and monitor changes over time.[12]

Cell Differentiation:Depending on the cell type and culture conditions, monolayer cultures can be used to induce cell differentiation. This is particularly important in studying developmental processes and tissue-specific functions.[12]

### Monolayer cultures for the personalized therapy in aggressive thyroid cancer of follicular origin

Endocrine cancer with the highest incidence is thyroid cancer (TC). Differentiated thyroid cells (DTC) that originate from follicular thyroid cells account for over 90% of total thyroid cells (TC). Papillary TC (PTC), follicular TC (FTC), and Hürthle cell TC are examples of DTC. One percent of TC is anaplastic TC (ATC), which accounts for 15–40% of TC deaths.[14]

Mortality is one of the biggest obstacles to current treatment techniques against aggressive DTC or ATC. These strategies are not entirely effective against these conditions. Recent years have seen advancements in our knowledge of the molecular and genetic underpinnings of TC development as well as the introduction of novel medications, such as tyrosine kinase inhibitors (TKIs), which target the oncogenic or signaling kinases linked to cellular proliferation.[14]

Preclinical models have made use of thyroid cell lines that were isolated from tumoral cells and selected for their high rate of proliferation in vitro. As a result of their adaptation to in vitro growth circumstances, these cells actually lose the distinctive characteristics of the original tumor. Because of these factors, there are significant restrictions on the usage of these cell lines. More recently, monolayer cultures of human primary cells have been created, and their biological behavior has been studied. Furthermore, whereas human primary cell cultures may now be created from samples of fine-needle aspiration citology from aggressive dedifferentiated DTC or ATC, primary TC cells were previously only obtained by surgical biopsies. Without the use of useless medications, testing several TKIs in vitro on individual patients can aid in the development of novel, individualized treatments.[14]

Limitations of monolayer culture and motivations:

Scientists are investigating novel models that can more accurately mimic the structure and function of human organs because to the limitations of monolayer culture settings. Protocol improvements in recent times have led to the creation of three-dimensional (3D) organ-like architectures known as "organoids," which are able to exhibit properties of their corresponding real organs, such as morphological features, functional activities, and individual responses to particular pathogens.[15]

## Cell culture protocol

For in vitro investigations to be conducted correctly, the cell culture protocol for a particular cell line must be optimized. The best culture conditions for different cell lines can differ significantly due to the heterogeneity of germ cell malignancies.[16]

## See also

* 3D cell culture
* Induced pluripotent stem cell
* Patient-derived xenograft

## References

1. ↑ "Guidelines for the use of cell lines in biomedical research". British Journal of Cancer 111 (6): 1021–1046. September 2014. doi:10.1038/bjc.2014.166. PMID 25117809.
2. ↑ "Cellular senescence: when bad things happen to good cells". Nature Reviews. Molecular Cell Biology 8 (9): 729–740. September 2007. doi:10.1038/nrm2233. PMID 17667954.
3. ↑ Culture of immortalized cells. New York: Wiley-Liss. 1996. ISBN 978-0-471-12134-3. https://books.google.com/books?id=BapqAAAAMAAJ.
4. ↑ 4.0 4.1 "The clinical relevance of cancer cell lines". Journal of the National Cancer Institute 105 (7): 452–458. April 2013. doi:10.1093/jnci/djt007. PMID 23434901.
5. ↑ "Efficacy of anti-cancer agents in cell lines versus human primary tumour tissue". Current Opinion in Pharmacology 10 (4): 375–379. August 2010. doi:10.1016/j.coph.2010.05.001. PMID 20570561.
6. ↑ "Organoid Profiling Identifies Common Responders to Chemotherapy in Pancreatic Cancer". Cancer Discovery 8 (9): 1112–1129. September 2018. doi:10.1158/2159-8290.CD-18-0349. PMID 29853643.
7. ↑ American Type Culture Collection Standards Development Organization Workgroup ASN-0002 (June 2010). "Cell line misidentification: the beginning of the end". Nature Reviews. Cancer 10 (6): 441–448. doi:10.1038/nrc2852. PMID 20448633.
8. ↑ "Preclinical model of organotypic culture for pharmacodynamic profiling of human tumors". Proceedings of the National Academy of Sciences of the United States of America 107 (18): 8352–8356. May 2010. doi:10.1073/pnas.0907676107. PMID 20404174. Bibcode: 2010PNAS..107.8352V.
9. ↑ "Ex vivo tumor culture systems for functional drug testing and therapy response prediction". Future Science OA 3 (2): FSO190. June 2017. doi:10.4155/fsoa-2017-0003. PMID 28670477.
10. ↑ "Organotypic culture of breast tumor explants as a multicellular system for the screening of natural compounds with antineoplastic potential". BioMed Research International 2015: 618021. 2015. doi:10.1155/2015/618021. PMID 26075250.
11. ↑ "Hydrogen Peroxide Induced Toxicity Is Reversed by the Macrocyclic IRAP-Inhibitor HA08 in Primary Hippocampal Cell Cultures". Current Issues in Molecular Biology 44 (10): 5000–5012. October 2022. doi:10.3390/cimb44100340. PMID 36286055.
12. ↑ 12.0 12.1 12.2 12.3 Harris, Andrew R.; Peter, Loic; Bellis, Julien; Baum, Buzz; Kabla, Alexandre J.; Charras, Guillaume T. (2012-10-09). "Characterizing the mechanics of cultured cell monolayers" (in en). Proceedings of the National Academy of Sciences 109 (41): 16449–16454. doi:10.1073/pnas.1213301109. ISSN 0027-8424. PMC 3478631. https://pnas.org/doi/full/10.1073/pnas.1213301109.
13. ↑ 13.0 13.1 13.2 Majhy, B.; Priyadarshini, P.; Sen, A. K. (2021). "Effect of surface energy and roughness on cell adhesion and growth – facile surface modification for enhanced cell culture" (in en). RSC Advances 11 (25): 15467–15476. doi:10.1039/D1RA02402G. ISSN 2046-2069. PMC 8698786. http://xlink.rsc.org/?DOI=D1RA02402G.
14. ↑ 14.0 14.1 14.2 Fallahi, Poupak; Ferrari, Silvia Martina; Elia, Giusy; Ragusa, Francesca; Patrizio, Armando; Paparo, Sabrina Rosaria; Marone, Gianni; Galdiero, Maria Rosaria et al. (February 2022). "Primary cell cultures for the personalized therapy in aggressive thyroid cancer of follicular origin" (in en). Seminars in Cancer Biology 79: 203–216. doi:10.1016/j.semcancer.2020.06.013. https://linkinghub.elsevier.com/retrieve/pii/S1044579X20301498.
15. ↑ Heydari, Zahra; Moeinvaziri, Farideh; Agarwal, Tarun; Pooyan, Paria; Shpichka, Anastasia; Maiti, Tapas K.; Timashev, Peter; Baharvand, Hossein et al. (December 2021). "Organoids: a novel modality in disease modeling" (in en). Bio-Design and Manufacturing 4 (4): 689–716. doi:10.1007/s42242-021-00150-7. ISSN 2096-5524. PMC 8349706. https://link.springer.com/10.1007/s42242-021-00150-7.
16. ↑ Lafin, John T.; Amatruda, James F.; Bagrodia, Aditya (2021), Bagrodia, Aditya; Amatruda, James F., eds., "Germ Cell Tumor Cell Culture Techniques" (in en), Testicular Germ Cell Tumors (New York, NY: Springer US) 2195: pp. 65–76, doi:10.1007/978-1-0716-0860-9_5, ISBN 978-1-0716-0859-3, PMID 32852757, https://link.springer.com/10.1007/978-1-0716-0860-9_5, retrieved 2024-01-03

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