BERKELEY, CA (UroToday.com) - Because they better mimic the physiological reality of a tissue, direct imaging of cellular 3D architectures, assemblage, and organization is of primary interest. [1] Beyond this, being able to directly monitor the dynamics of networking between 3D objects is highly relevant to closely follow the development of human organs such as the prostate.
Acini, the basic 3D structures constituting a secretory epithelium, are organized as tree-like structures in the glandular organ. The mechanisms regulating the development of acini and ductal structures are poorly understood. [2] Moreover, the disruption of apical polarity, 3D organization, and the progressive transformation of acini into spheroids are key events in tumor progression. Therefore, the analysis of cell polarity and branching morphogenesis is critical to assess epithelial development, integrity and homeostasis and to study the initiation and progression of epithelial tumors. Using a lensfree system along with holographic 3D image reconstruction, we showed that we could discriminate 3D objects according to their polarity (differentiated acini from tumor-like spheroids). [1] Beyond this result, we would like here to emphasize the remarkable potential of our lensfree imaging system, in particular to assess the dynamics of cell-cell and cell-ECM interactions in a 3D interaction network that would mimic the reality of tissues.
First, on the basis of recent unpublished data, we believe that the innovative combination between lensfree imaging and 3D cell culture provides a tool to examine the dynamics of cellular events occurring during epithelial development or carcinogenesis and to elucidate main events that control growth and differentiation of normal and cancerous epithelial cells. Cell movement is a key feature of many physiological and pathological processes including development, morphogenesis and tissue repair, and cancer invasion. [3, 4] Complex multicellular dynamics and tissue remodelling are supported by both single cell and collective cell migration processes. [5] If physiologically relevant 3D cell models have contributed to our understanding of cell migration, the signalling mechanisms and molecular actors driving migration in 3D remain poorly understood. [6] Moreover, cell migration and invasion of tissues are known to be controlled by external stimuli such as growth factors and cell–cell interactions. Distal effectors including extracellular matrix components are important in regulating sprouting and branching events. To analyse such dynamic cellular processes, confocal videomicroscopy is very effective at observing cellular and sub-cellular level, but because of small field of view, it misses the whole picture where the environment predominates. One major limitation would come, indeed, from a limited scope of imaging that under-estimates the intrinsic heterogeneity of cell migration strategies within a 3D context. [5] With lens-free imaging, we could very recently track in real-time, in a population of diverse 3D structures, the collective dynamics of cell-cell communication during epithelia tubulogenesis (unpublished data). The large field of view of our imaging system allowed us to observe the coexistence of various phenotypes and gave access to the representative cooperation between cells that collectively change their environment. We believe such a system that enables the rapid imaging and measurement of morphologic changes would greatly facilitate screening for environmental factors that affect epithelial homeostasis. Our system provides a tool to unravel biological processes at work in development and morphogenesis in a more physiological and dynamical context than traditional cell cultures.
Second, lens-free imaging can be extended to biomedical and drug screening applications, to follow cancer progression, and to evaluate anti-cancer drugs. To date, one key issue is the need for new predictive biomarkers in cancer. Especially in the area of prostate cancer, the issue of PSA (Prostatic Specific Antigen) testing remains controversial, both economically and socially. We currently took advantage of our recent technological innovations in parallelized lens-free imaging to perform RNAi-based high content screening in 3D cell culture of prostate cells. We aim to characterize signaling cascades and essential genes responsible for the formation of tumor-like spheroids and duct-like structures and on a longer term, to identify new biomarkers which could complement PSA testing and also to characterize new RNAi therapeutic agents in oncology. Interestingly, tumor spheroids are often considered good models that recapitulate features of cancer because they are characterized by hypoxic and necrotic regions. Therefore, spheroids are used in pharmaceutical labs in in vitro 3D spheroid-based assays for measuring tumor growth and evaluating anticancer drugs. Functional assays for the evaluation of tumor progression are mostly based on assays of spheroid migration and their ability to invade connective tissue. [7] However, those assays do not consider the status of spheroid polarity and organization that remains a critical readout to assess epithelial tissue integrity and homeostasis.
Hence, developing new diagnostic tests, not only based on 3D structures organization but also on the dynamics of duct-like formation and on the quantification of cell migration processes, would certainly constitute a novel approach to fight cancer. These technologies, and the expertise behind them, could be extended to study development and cancer biology of any secretory epithelium as well as to large scale screening for drug discovery or toxicology studies in organ-like models.
References:
- Dolega ME, Allier C, Vinjimore Kesavan S, Gerbaud S, Kermarrec F, Marcoux P, Dinten JM, Gidrol X, Picollet-D’hahan N (2013) Label-free analysis of prostate acini-like 3D structures by lensfree imaging. Biosensors & Bioelectronics 49: 176-183.
- Pohl M, Stuart RO, Sakurai H, Nigam SK (2000). Branching morphogenesis during kidney development. Annu Rev Physiol 62: 595–620.
- Nguyen-Ngoc KV, Cheung KJ, Brenot A, Shamir ER, Gray RS, Hines WC, Yaswen P, Werb Z, Ewald AJ (2012). ECM microenvironment regulates collective migration and local dissemination in normal and malignant mammary epithelium. PNAS 25, 109 (39) : E2595-604.
- Friedl P. & Gilmour D (2009). Collective cell migration in morphogenesis, regeneration and cancer. Nature Reviews Molecular Cell Biology 10, 445-457
- Friedl P, Sahai E, Weiss S, Yamada KM (2012). New dimensions in cell migration. Nat Rev Mol Cell Biol 13 (11) : 743-7.
- Doyle AD, Petrie RJ, Kutys ML, Yamada KM (2013). Dimensions in cell migration. Curr Opin Cell Biol 25 (5) : 642-9.
- Vinci M., Gowan S., Boxall F., Patterson L., Zimmermann M., Court W., Lomas C., Mendiola M., Hardisson D., Eccles S.A. (2012). Advances in establishment and analysis of three-dimensional tumor spheroid-based functional assays for target validation and drug evaluation. BMC Biol. 10, 29:1-20.
Written by:
Nathalie Picollet-D’hahan,a, b, c, * Monika E. Dolega,a, b, c Cédric Allier,d and Xavier Gidrola, b, c as part of Beyond the Abstract on UroToday.com. This initiative offers a method of publishing for the professional urology community. Authors are given an opportunity to expand on the circumstances, limitations etc... of their research by referencing the published abstract.
aCEA, iRTSV, Biologie à Grande Echelle, F-38054 Grenoble, France. bINSERM, U1038, F-38054 Grenoble, France cUniversité Grenoble-Alpes, F-38000 Grenoble, France dCEA, LETI, MINATEC, F-38054 Grenoble, France
* Corresponding Author:
Label-free analysis of prostate acini-like 3D structures by lensfree imaging - Abstract
More Information about Beyond the Abstract