Ex vivo

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Ex vivo brainstem: A. coronal view displaying the anterior portion of the tissue sample; B. sagittal view displaying the left-hand side of the tissue sample Ex vivo Brainstem sample.jpg
Ex vivo brainstem: A. coronal view displaying the anterior portion of the tissue sample; B. sagittal view displaying the left-hand side of the tissue sample

Ex vivo ( Latin for 'out of the living') refers to biological studies involving tissues, organs, or cells maintained outside their native organism under controlled laboratory conditions. These studies preserve the extracted materials' functional viability and structural integrity for limited periods by precisely managing the conditions. An intermediate approach between in vitro and in vivo studies, ex vivo models provide a balance between experimental control and physiological relevance, allowing for experiments that may be ethically or technically impractical in living subjects. While the foundational techniques of ex vivo experimentation were established in the 19th century, their formalization and refinement as a research methodology accelerated in the mid-20th century. Ex vivo procedures are widely applied in medical research, pharmacology, and biotechnology. [2] [3] [4]

Contents

Advantages and limitations

Ex vivo, meaning 'out of the living' in Latin, refers to biological studies involving tissues, organs, or cells maintained outside their native organism under controlled laboratory conditions. These studies preserve the extracted materials' functional viability and structural integrity for limited periods by precisely managing the conditions, which typically include oxygenation, temperature, nutrient delivery, and humidity. The conditions are often facilitated through cell culture media or specialized perfusion chambers. As an intermediate approach between in vitro studies, which traditionally use isolated cells in artificial environments, [a] and in vivo research, conducted within living organisms, ex vivo models preserve more of the native tissue architecture than traditional cell cultures, while offering greater experimental control than whole-organism studies. [6] [7] [8] By maintaining natural tissue organization, ex vivo models address some limitations of in vitro work, such as oversimplified cellular interactions, and mitigate the variability and systemic influences inherent to in vivo approaches. [9] [10] Ex vivo models may offer ethical benefits by reducing reliance on animal testing, allowing researchers to conduct physiologically relevant experiments without using entire living systems. [11] [12]

In cardiovascular research, a Langendorff-perfused heart preparation involves removing a heart from an organism and perfusing it with a nutrient-rich solution. This method maintains the structural integrity and electrical conduction pathways of the heart, making it suitable for studying phenomena like arrhythmias or drug effects in a more physiologically relevant setting without the complexities of an in vivo model. [13] [14] In dermatological research, human skin organ culture (hSOC) is a technique where excised human skin is maintained in an artificial medium while preserving its native structure. It is used to investigate wound healing, drug penetration, and toxicology. These models retain architectural features of the skin, enabling the study of conditions unique to humans, such as keloids, [b] which are not replicated in animal models. [7] [17] Skin explants from surgical procedures allow researchers to observe early-stage physiological responses to laser treatments in ways that closely resemble in vivo conditions, though processes like re-epithelialization occur more slowly than in living tissue. [18] In intervertebral disc research, ex vivo models that retain vertebral bone allow for testing potential drugs and investigating loading effects on disc degeneration and repair. [19] In biosensing and electroanalytical applications, ex vivo methods offer experimental flexibility unavailable in living systems. While many in vivo experiments favor micro- and nanoelectrodes to minimize invasiveness, larger electrodes are routinely used for specific purposes. Ex vivo approaches, by contrast, permit custom electrode geometries that interface precisely with biological tissues under controlled conditions, without the same constraints on size and invasiveness. This adaptability enables detailed examination of biological analytes and their physiological roles. Ex vivo electroanalytical methods are applied in neuroscience, pharmacology, and biomedical engineering to study neurotransmitter dynamics, metabolic activity, and disease-associated biomarkers. [20] : 161–164  [21] : 3–4 

However, ex vivo models have inherent limitations, including significant changes in biophysical properties of tissues after the organism's death, tissue degradation that increases with time, restricted viability duration, as well as absence of whole-body physiological responses, such as lack of blood circulation and nerve innervation. These constraints prevent the models from replicating long-term or systemic effects. [6] [22] The comparison between in vivo and ex vivo models can be complex due to these changes in tissue properties. For instance, studies have found large changes in electric fields between in vivo and ex vivo measurements that increased with postmortem time. [22]

History

Oswald Schmiedeberg2.jpg
Oswald Schmiedeberg
Oskar Langendorff 1913 (Paul Moennich).jpg
Oscar Langendorff

The foundations of ex vivo experimentation were established in the 19th century. In 1846, the German physiologist Carl Ludwig and one of his students, Carl Wild, conducted one of the earliest perfusion studies, in which the heart of a deceased animal was connected to the common carotid artery of a living donor animal. This arrangement allowed the donor's circulation to maintain perfusion of the recipient heart's coronary vessels, although the heart remained partially dependent on the living organism. [14] The earliest known studies on kidneys perfused outside the native organism were conducted by German physiologist Carl Eduard Loebell, who presented his findings in a doctoral dissertation [c] in 1849. [23] In 1866, Russian physiologist Elias von Cyon developed the isolated perfused frog heart preparation at the Carl Ludwig Institute of Physiology in Leipzig, Germany. This method was commonly used during the late 19th century and later served as the basis for the isolated perfused mammalian heart preparation. [24] In 1876, German physiologist Gustav von Bunge and German pharmacologist Oswald Schmiedeberg demonstrated the synthesis of hippuric acid in the isolated dog kidney. [23] In 1885, German physiologist Maximilian von Frey and Austrian biologist Max von Gruber, working at the Carl Ludwig Institute of Physiology, constructed an apparatus combining a mechanical pump with an early oxygenator that substituted the function of the heart and lungs in experiments on dogs. This device oxygenated blood outside the body and was a precursor to the heart-lung machine. [25]

In the 1880s, British physiologist Sydney Ringer developed a salt solution that sustained rhythmic contractions in the isolated frog heart. Later named Ringer's solution, it enabled extended observation of cardiac activity and supported controlled experimental studies on cardiac physiology in isolated preparations. [26] In 1895, German physiologist Oskar Langendorff introduced a method for isolated heart perfusion involving retrograde flow through the aorta to supply the coronary circulation. The Langendorff preparation allowed for direct measurement of cardiac function and precise control of perfusion parameters while minimizing systemic confounders inherent to in vivo models. It became a widely used technique in the study of cardiac physiology and remains a standard method in cardiovascular research. [14]

A heart-lung machine (upper right) in a coronary artery bypass surgery Coronary artery bypass surgery Image 657C-PH.jpg
A heart–lung machine (upper right) in a coronary artery bypass surgery

Throughout the 20th century, ex vivo techniques were adapted for a range of animal models. A significant refinement was the development of the working heart model, in which perfusate enters the left atrium and exits through the aorta, more closely replicating physiological flow conditions. Advances in instrumentation enabled detailed assessments of cardiac function, including pressure–volume relationships, oxygen consumption, and myocardial contractility. [27] [28] In 1953, American surgeon John Heysham Gibbon successfully employed a heart-lung machine during open-heart surgery on a human patient. [25]

Techniques

Demonstration of isolation of choroid from the mouse eye [29]
In situ lung function evaluation, and assessment of total lung capacity (TLC) and basal elastance after performing a recruitment maneuver [30]

Ex vivo research uses specialized techniques to sustain tissues or cells outside their natural environment while preserving their functional integrity. In ex vivo organ perfusion, whole organs are maintained in a functional state by circulating solutions that mimic physiological blood flow. This method enables researchers to investigate organ-level responses to drugs, disease states, or environmental changes under controlled conditions. By contrast, organ culture traditionally involves maintaining organ sections or small fragments in static or semi-static conditions without active perfusion. [31] [32]

Another widely used technique is organotypic slice cultures, where thin sections of organs are maintained on supportive matrices to preserve their three-dimensional structure and cellular interactions. This method enables the study of localized responses, tissue-specific dynamics, and physiological processes over extended periods. [33] [34] [35] An example is the use of hippocampal slices in neuroscience research to examine long-term cellular and synaptic activity. [36] In some cases, ex vivo electroporation, in which an electric field is applied to cells to facilitate the uptake of genetic material, is used to introduce DNA into cells within tissue slices, allowing researchers to study gene expression in a controlled environment. [37] : 241 

Cell culture involves isolating individual cells from tissues and growing them in a medium enriched with nutrients and growth factors. While these cultures retain some functional characteristics of their tissue of origin, they often exhibit changes in phenotype and gene expression when removed from their native environment. Primary cell cultures, derived directly from tissues, more closely resemble physiological conditions than immortalized cell lines, making them essential for studying cellular behavior, disease mechanisms, and drug effects. [38] [39]

See also

Notes

  1. ↑ However, advanced in vitro models have evolved from simple 2D cultures to more complex 3D systems like organoids and organ-on-a-chip platforms, which aim to better replicate tissue architecture. This progression blurs the line between traditional in vitro and ex vivo models. [5]
  2. ↑ In animals, researchers can induce hypertrophic scars, which are somewhat similar to keloids, but keloid formation does not occur. Even in animal models designed to exhibit excessive fibrotic responses through genetic manipulation or specific treatments, the characteristic features of keloids—such as growth beyond the original wound margins and tendency to persist or recur without regression—are not observed. [15] [16]
  3. ↑ Titled De conditionibus, quibus secretiones in glandulis perficiuntur (transl. 'On the conditions under which secretions are produced in the glands'), the dissertation was presented at the University of Marburg, Germany. [23] This monograph, written in Latin, is available for browsing on Google Books and the Deutsche Digitale Bibliothek .

References

  1. ↑ Ford, Anastasia A.; Colon-Perez, Luis; Triplett, William T.; Gullett, Joseph M.; Mareci, Thomas H.; Fitzgerald, David B. (2013). "Imaging White Matter in Human Brainstem". Frontiers in Human Neuroscience. 7: 400. doi: 10.3389/fnhum.2013.00400 . PMC   3721683 . PMID   23898254.
  2. ↑ Makdisi, G; Makdisi, T; Jarmi, T; Caldeira, CC (2017). "Ex vivo lung perfusion review of a revolutionary technology". Annals of Translational Medicine. 5 (17): 343. doi: 10.21037/atm.2017.07.17 . PMC   5599284 . PMID   28936437.
  3. ↑ Griffiths, John R. (2022). "Magnetic resonance spectroscopy ex vivo: A short historical review". NMR in Biomedicine. 35 (4): e4740. doi:10.1002/nbm.4740. PMID   35415860.
  4. ↑ Maroli, Amith Sadananda; Powers, Robert (2023). "Closing the gap between in vivo and in vitro omics: using QA/QC to strengthen ex vivo NMR metabolomics". NMR in Biomedicine. 36 (4): e4594. doi:10.1002/nbm.4594. PMC   8821733 . PMID   34369014.
  5. ↑ Nairon, Kylie G.; Skardal, Aleksander (September 2021). "Biofabrication of advanced in vitro and ex vivo cancer models for disease modeling and drug screening". Future Drug Discovery. 3 (3). doi:10.4155/fdd-2020-0034.
  6. 1 2 Piglionico, Sofia Silvia; Pons, Coline; Romieu, Olivier; Cuisinier, Frédéric; Levallois, Bernard; Panayotov, Ivan Vladislavov (2023). "In vitro, ex vivo, and in vivo models for dental pulp regeneration". Journal of Materials Science: Materials in Medicine. 34 (15): 15. doi:10.1007/s10856-023-06718-2. PMC   10067643 . PMID   37004591.
  7. 1 2 Zhou, Lijuan; Zhang, Xianqi; Paus, Ralf; Lu, Zhongfa (2018). "The renaissance of human skin organ culture: A critical reappraisal". Differentiation. 104: 22–35. doi:10.1016/j.diff.2018.10.002. PMID   30391646.
  8. ↑ Mohizin, Abdul; Imran, Jakir Hossain; Lee, Kee Sung; Kim, Jung Kyung (February 2023). "Dynamic interaction of injected liquid jet with skin layer interfaces revealed by microsecond imaging of optically cleared ex vivo skin tissue model". Journal of Biological Engineering. 17 (1): 15. doi: 10.1186/s13036-023-00335-x . PMC   9969392 . PMID   36849998.
  9. ↑ Szczesny, Spencer E. (July 2020). "Ex vivo models of musculoskeletal tissues". Connective Tissue Research. 61 (3–4): 245–247. doi:10.1080/03008207.2020.1742418. PMID   32340565.
  10. ↑ Lilburn, D. M. L.; Tatler, A. L.; Six, J. S.; Lesbats, C.; Habgood, A.; Porte, J.; Hughes-Riley, T.; Shaw, D. E.; Jenkins, G.; Meersmann, T. (2016). "Investigating lung responses with functional hyperpolarized xenon-129 MRI in an ex vivo rat model of asthma". Magnetic Resonance in Medicine. 76 (4): 1224–1235. doi:10.1002/mrm.26003. PMC   5026173 . PMID   26507239.
  11. ↑ Ruppelt, Alicia; Pappers, Claire; de Mol, Bas; Spee, Bart; Rasponi, Marco; Stijnen, Marco (June 2022). "P1: Ex Vivo Liver Perfusion for Research Purposes. Can We Use Slaughterhouse Material to Reduce Animal Testing?". ASAIO Journal . 68 (Supplement 2): 91. doi:10.1097/01.mat.0000841220.50962.35.
  12. ↑ Tang, Shirley N.; Bonilla, Andres F.; Chahine, Nadeen O.; Colbath, Aimee C.; Easley, Jeremiah T.; Grad, Sibylle; Haglund, Lisbet; Le Maitre, Christine L.; Leung, Victor; McCoy, Annette M.; Purmessur, Devina; Tang, Simon Y.; Zeiter, Stephan; Smith, Lachlan J. (December 2022). "Controversies in spine research: Organ culture versus in vivo models for studies of the intervertebral disc". JOR Spine. 5 (4): e1235. doi:10.1002/jsp2.1235. PMC   9799089 . PMID   36601369.
  13. ↑ Nyman, Mathias; Stølen, Thomas O.; Johnsen, Andreas B.; Garten, Kristina; Burton, Francis L.; Smith, Godfrey L.; Loennechen, Jan P. (2024). "A comprehensive protocol combining in vivo and ex vivo electrophysiological experiments in an arrhythmogenic animal model". American Journal of Physiology – Heart and Circulatory Physiology. 326 (1): H203 –H215. doi:10.1152/ajpheart.00358.2023. PMC   11213483 . PMID   37975708.
  14. 1 2 3 Leivaditis, Vasileios; Mulita, Francesk; Dahm, Manfred; Grapatsas, Konstantinos; Papatriantafyllou, Athanasios; Bekou, Eleni; Verras, Georgios-Ioannis; Tasios, Konstantinos; Tchabashvili, Levan; Markakis, Konstantinos; Lozos, Vasileios; Koletsis, Efstratios (2024). "History of the development of isolated heart perfusion experimental model and its pioneering role in understanding heart physiology". Archives of Medical Science – Atherosclerotic Diseases. 9 (1): 109–121. doi:10.5114/amsad/188270. PMC   11289247 . PMID   39086622.
  15. ↑ Turobi, Naina Banun; Rahman, Fadel; Fernanda, Nathasya; Fawzy, Ahmad (March 2024). "Clinical and Molecular Insights into Hypertrophic Scars and Keloids: A Literature Review". International Journal of Medical Science and Clinical Research Studies. 4 (3): 387–399. doi:10.47191/ijmscrs/v4-i03-06.
  16. ↑ Hussein, Ramadan S. (March 2024). "Botulinum Neurotoxin BoNT-A in the Management of Hypertrophic Scars and Keloids: A Comprehensive Review". International Journal of Biomedicine. 14 (1): 15–19. doi:10.21103/Article14(1)_RA2.
  17. ↑ Sharma, J. R.; Lebeko, M.; Kidzeru, E. B.; Khumalo, N. P.; Bayat, A. (December 2019). "In vitro and ex vivo models for functional testing of therapeutic anti-scarring drug targets in keloids". Advances in Wound Care. 8 (12): 655–670. doi:10.1089/wound.2019.1040. PMC   6904937 . PMID   31827980.
  18. ↑ Cho, H.; Won, C.H.; Chang, S.E.; Lee, M.W.; Park, G. (2013). "Usefulness and Limitations of Skin Explants to Assess Laser Treatment". Medical Lasers. 2 (2): 58–63. doi:10.25289/ML.2013.2.2.58.
  19. ↑ Grant, Michael; Epure, Laura M.; Salem, Omar; AlGarni, Nadim; Ciobanu, Oana; Alaqeel, Maha; Antoniou, John; Mwale, Fackson (July 2016). "Development of a Large Animal Long-Term Intervertebral Disc Organ Culture Model That Includes the Bony Vertebrae for Ex Vivo Studies". Tissue Engineering Part C: Methods. 22 (7): 636–644. doi:10.1089/ten.tec.2016.0049. PMID   27216856.
  20. ↑ Patel, Bhavik A. (2021). "8. Measurement from ex vivo tissues". Electrochemistry for Bioanalysis. Elsevier. ISBN   9780128215357 – via Google Books.
  21. ↑ Michael, Adrian C.; Borland, Laura M. (2007). "1. An Introduction to Electrochemical Methods in Neuroscience". In Michael, Adrian C.; Borland, Laura M. (eds.). Electrochemical Methods for Neuroscience. CRC Press. ISBN   9781420005868 – via Google Books.
  22. 1 2 Opitz, Alexander; Falchier, Arnaud; Linn, Gary S.; Milham, Michael P.; Schroeder, Charles E. (May 2017). "Limitations of ex vivo measurements for in vivo neuroscience". Proceedings of the National Academy of Sciences of the United States of America . 114 (20): 5243–5246. Bibcode:2017PNAS..114.5243O. doi: 10.1073/pnas.1617024114 . PMC   5441777 . PMID   28461475.
  23. 1 2 3 Schurek, Hans-Joachim; Neumann, Klaus Hinrich; Schweda, Frank; Czogalla, Jan (2017). "1.1. Historical Aspects of the Perfusion of Isolated Kidneys". A Laboratory Manual of Kidney Perfusion Techniques. Münster, Germany: University of Münster. PMID   33369897.
  24. ↑ Skrzypiec-Spring, Monika; Grotthus, Bartosz; Szeląg, Adam; Schulz, Richard (2007). "Isolated heart perfusion according to Langendorff—Still viable in the new millennium". Journal of Pharmacological and Toxicological Methods. 55 (2): 113–126. doi:10.1016/j.vascn.2006.05.006. ISSN   1056-8719. PMID   16844390.
  25. 1 2 Zimmer, Heinz-Gerd (2003). "The heart-lung machine was invented twice--the first time by Max von Frey". Clinical Cardiology. 26 (9): 443–445. doi:10.1002/clc.4960260914. PMC   6654655 . PMID   14524605.
  26. ↑ Fye, W. B. (1984). "Sydney Ringer, calcium, and cardiac function". Circulation. 69 (4): 849–853. doi:10.1161/01.cir.69.4.849. PMID   6365353.
  27. ↑ Neely, J. R.; Liebermeister, H.; Battersby, E. J.; Morgan, H. E. (1967). "Effect of pressure development on oxygen consumption by isolated rat heart". American Journal of Physiology. 212 (4): 804–814. doi:10.1152/ajplegacy.1967.212.4.804. PMID   6024443.
  28. ↑ Suga, H.; Sagawa, K. (1974). "Instantaneous pressure-volume relationships and their ratio in the excised, supported canine left ventricle". Circulation Research. 35 (1): 117–126. doi:10.1161/01.RES.35.1.117. PMID   4841253.
  29. ↑ Shao, Zhuo; Friedlander, Mollie; Hurst, Christian G.; Cui, Zhenghao; Pei, Dorothy T.; Evans, Lucy P.; Juan, Aimee M.; Tahir, Houda; Duhamel, François; Chen, Jing; Sapieha, Przemyslaw; Chemtob, Sylvain; Joyal, Jean-Sébastien; Smith, Lois E. H. (2013). "Choroid Sprouting Assay: An Ex Vivo Model of Microvascular Angiogenesis". PLOS ONE. 8 (7): e69552. Bibcode:2013PLoSO...869552S. doi: 10.1371/journal.pone.0069552 . PMC   3724908 . PMID   23922736. S2CID   466393.
  30. ↑ Bassani, Giulia Alessandra; Lonati, Caterina; Brambilla, Daniela; Rapido, Francesca; Valenza, Franco; Gatti, Stefano (2016). "Ex Vivo Lung Perfusion in the Rat: Detailed Procedure and Videos". PLOS ONE. 11 (12): e0167898. Bibcode:2016PLoSO..1167898B. doi: 10.1371/journal.pone.0167898 . PMC   5148015 . PMID   27936178.
  31. ↑ Menander, M.; Attawar, S.; Mahesh, BN.; Tisekar, O.; Mohandas, A. (2024). "Ex vivo lung perfusion and the Organ Care System: a review". Clinical Transplant Research. 38 (1): 23–36. doi:10.4285/ctr.23.0057. PMC   11075812 . PMID   38725180.
  32. ↑ Martins, Paulo N.; Del Turco, Serena; Gilbo, Nicholas (2022). "Organ Therapeutics During Ex-Situ Dynamic Preservation: A Look Into the Future". European Journal of Transplantation. 1 (1): 63–78. doi:10.57603/EJT-010.
  33. ↑ Peng, Michael; Margetts, Tyler J.; Sugali, Chenna Kesavulu; Rayana, Naga Pradeep; Dai, Jiannong; Sharma, Tasneem P.; Raghunathan, Vijay Krishna; Mao, Weiming (2022). "An ex vivo model of human corneal rim perfusion organ culture". Experimental Eye Research. 214: 108891. doi:10.1016/j.exer.2021.108891. PMC   8792355 . PMID   34896309.
  34. ↑ Humpel, Christian (2015). "Organotypic brain slice cultures: a review". Neuroscience. 305: 86–98. doi:10.1016/j.neuroscience.2015.07.086. PMC   4699268 . PMID   26254240.
  35. ↑ Siwczak, Fatina; Hiller, Charlotte; Pfannkuche, Helga; Schneider, Marlon R. (2023). "Culture of vibrating microtome tissue slices as a 3D model in biomedical research". Journal of Biological Engineering. 17 (1): 36. doi: 10.1186/s13036-023-00357-5 . PMC   10233560 . PMID   37264444.
  36. ↑ Jang, Sooah; Kim, Hyunjeong; Kim, Hye-Jin; Lee, Su Kyoung; Kim, Eun Woo; Namkoong, Kee; Kim, Eosu (2018). "Long-Term Culture of Organotypic Hippocampal Slice from Old 3xTg-AD Mouse: An ex vivo Model of Alzheimer's Disease". Psychiatry Investigation. 15 (2): 205–213. doi:10.30773/pi.2017.04.02. PMC   5900409 . PMID   29475217.
  37. ↑ Carter, Matt; Shieh, Jennifer C. (2015). "11. Gene Delivery Strategies". Guide to Research Techniques in Neuroscience (2nd ed.). Academic Press. ISBN   9780128005972 – via Google Books.
  38. ↑ Pan, Cuiping; Kumar, Chanchal; Bohl, Sebastian; Klingmueller, Ursula; Mann, Matthias (2009). "Comparative proteomic phenotyping of cell lines and primary cells to assess preservation of cell type-specific functions". Molecular & Cellular Proteomics. 8 (3): 443–450. doi: 10.1074/mcp.M800258-MCP200 . PMC   2649808 . PMID   18952599.
  39. ↑ Nilsson, Linnéa M.; Castresana-Aguirre, Miguel; Scott, Lena; Brismar, Hjalmar (2020). "RNA-seq reveals altered gene expression levels in proximal tubular cell cultures compared to renal cortex but not during early glucotoxicity". Scientific Reports. 10 (1): 108891. Bibcode:2020NatSR..1010390N. doi:10.1038/s41598-020-67361-3. PMC   7316724 . PMID   32587318.
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