Human Embryonic and Hepatic Stem Cell Differentiation Visualized in Two and Three Dimensions Based on Serial Sections

Abstract

Pluripotent human embryonic stem cells (hESCs) are characterized by two defining properties, self-renewal and differentiation. Self-renewing hESCs express transcription factors OCT4, SOX2, and NANOG, and surface markers SSEA-4 and TRA-1-60 and TRA-1-81 and their ability to differentiate into derivatives of the three germ layers show the differentiating potential. Studies suggest a certain microheterogeneity of the hESC colonies, in which not all cells in one colony of apparently undifferentiated cells express all the expected markers. We describe a technique to paraffin embed an entire hESC colony, and prepare 3-5 μm thick serial sections. Immunohistochemistry applied to individual sections produces a 2-dimensional survey of the developing hESC colony. Based on serial paraffin sections of the 2D-expression pattern, a new and useful 3D-visualization can be modeled. The actual 3D rendering of an entire colony is accomplished using 3D image processing software such as Mimics(®) or Amira(®). An extended version of this technique even allows for a high-magnification 3D-reconstruction of, e.g., hepatic stem cells in developing liver. These techniques combined allow for both a 2- and a 3-dimensional visualization of hESC colonies and stem cells in organs, which leads to new insights into and information about the interaction of stem cells with their surroundings.

Three‐dimensional digital reconstruction of skin epidermis and dermis

“…(The cell and dermis fibre can be segmented based on the threshold segmentation method.) The serial skin tissue sections were then imported and processed using the Mimics software (Vestentoft et al ., ). Using the threshold segmentation tool and manual segmentation tool, the grey value of skin tissue sections was adjusted until the required skin tissue structure was selected completely.…”

“…Conventional histological section can provide planar information and is helpful in distinguishing components of skin tissue. However, if two‐dimensional (2D) plane sections are used to assess 3D skin tissue then identification errors are easily made (Chang et al ., ; Dijkstra et al ., ; Weis et al ., ; Yamamoto et al ., ; Vestentoft et al ., ).…”

Understanding the three-dimensional world from two-dimensional immunofluorescent adjacent sections

“…Another drawback is that in most studies, serial sections are visualized using chromogen-based staining protocols, which limits the number of proteins that can be visualized at the same time. [ 2 3 4 ]…”