3D reconstruction of a tumorsphere created using SBF-SEM with ZEISS Sigma FE-SEM

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3D Ultrastructural Investigations of Tumorspheres with SBF-SEM

byZEISS Microscopy 22 March 2024 3 min read

User Story

3D Ultrastructural Investigations of Tumorspheres with SBF-SEM

Volume electron microscopy of 3D cancer cell spheroids reveals significant morphological differences compared to 2D cell cultures.

The tumorsphere, a 3D cell culture or spheroid derived from cancer stem cells, represents a valuable tool in cancer research and drug discovery due to its architecture closely resembling the physiological characteristics of in vivo tumors.

A recent article featured in the Journal of Structural Biology explored the ultrastructural morphological differences between tumorspheres and traditional monolayer cultured cells. This study utilized advanced sample preparation methodologies, including high-pressure freezing and freeze substitution, to preserve cellular structures in their native state. The images were captured using serial block-face scanning electron microscopy (SBF-SEM) with ZEISS Sigma FE-SEM.

The research was led by Dr. Mihnea Bostina, whose titles include Associate Professor at the University of Otago, Electron Microscopy Academic Lead at the Otago Micro and Nanoscale Imaging Facility, and President of Microscopy New Zealand. Collaborating on this endeavor were members of the Bostina Lab, including Nickhil Jadav, a PhD student specializing in structural oncology research within the Department of Microbiology and Immunology at the University of Otago, and Sailakshmi Velamoor, a former postdoctoral researcher at the University of Otago.

The application of SBF-SEM by this collaborative team revealed clear examples of critical morphological differences between tumorspheres and traditional 2D cell cultures. This not only reinforces the evidence supporting the superior in vivo modeling capabilities of 3D cultures but also underscores the profound insights that ultrastructural investigations with volume electron microscopy can uncover. These findings hold great promise for advancing our understanding of cancer biology and may have far-reaching implications for drug discovery and therapeutic development.

Our goal was to use SBF-SEM to explore tumorspheres' ultrastructure as they are known for being a superior cancer model compared to traditional 2D cell monolayers. The key question was how the 3D construction of cells in tumorspheres compared to those attached to a surface.

Dr. Mihnea Bostina Associate Professor at the University of Otago, Electron Microscopy Academic Lead at the Otago Micro and Nanoscale Imaging Facility, and President of Microscopy New Zealand

Volume Electron Microscopy of Tumorsphere

SBF-SEM with ZEISS Sigma FE-SEM

A tumorsphere - a 3D spheroid generated from cancer stem cells - is investigated using SBF-SEM with ZEISS Sigma FE-SEM to compare differences to traditional 2D cell cultures. This work is explored in detail in N. Jadav et al., Journal of Structural Biology.

Inside a Tumorsphere with Volume Electron Microscopy

The collaborative team reported very interesting results from their SBF-SEM studies of cancer cell spheroids.

In a 3D culture, cells displayed a distinct morphology: smaller, rounder, and more compact. Notably, central and peripheral cells within tumorspheres exhibited different characteristics. The team uncovered intricate cellular interconnectivities through tubular membranous structures facilitating cell-cell connections. Mitochondria showcased varied morphologies unique to tumorsphere cells. Another remarkable observation was the presence of long nuclear invaginations traversing the entire nucleus with individual mitochondria often present inside them in the vicinity of nucleoli.

We believe that volume electron microscopy with SBF-SEM will be highly valuable for comparing different types of cancer and examining tumorsphere responses to various cancer drugs or novel therapeutic agents.

Dr. Mihnea Bostina Associate Professor at the University of Otago, Electron Microscopy Academic Lead at the Otago Micro and Nanoscale Imaging Facility, and President of Microscopy New Zealand

Dr. Bostina's team with pages from their publication in the Journal of Structural Biology — First author Nickhil Jadav and SEM specialist Niki Hazelton with pages from the article in *The Journal of Structural Biology.*

The Importance of Tumorspheres in Cancer Research

In this study, Dr. Bostina and his team performed an ultrastructural comparative analysis between cancer cells cultured as monolayers and 3D cell cultured tumorspheres with SBF-SEM. The findings revealed numerous key differences in tumorspheres. Not only did they note differences in nuclear envelope invaginations, but they also discovered diversity in mitochondrial morphologies and intricate cellular interconnectivity.

These findings emphasize the importance of employing 3D cell culture models, such as tumorspheres, in research, drug discovery, and therapeutic studies as they more closely resemble in vivo conditions.

Read the full paper to see more of their beautiful ultrastructural work and in-depth analyses detailing the important differences between 3D cultured cells and traditional monolayers.