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10108 - Ki-67, Epidermal Hyperplasia

INTRODUCTION
The Ki-67 protein is expressed in the nucleus and is a marker for cell proliferation. Increased number of Ki-67 positive cells in epidermis indicates increased epidermal hyperplasia, a characteristic which is associated with various inflammatory skin diseases as atopic dermatitis and psoriasis [1].

This protocol can be used to assess the number of Ki-67 positive cells in epidermis. The protocol provides the number and area of Ki-67 positive cells within the epidermis region, as well as for the entire tissue section. Manual assessment of the outlined epidermis region can be necessary.

KEYWORDS
Ki-67, Ki67, skin, epidermis, quantification, digital pathology, image analysis

METHODS
The first image processing step is a rough outline of tissue. This is performed to limit the subsequent quantitative analysis and to facilitate numbering of tissue samples in cases with multiple samples per slide. The next processing step involves a segmentation of epidermis, followed by an exclusion of hair cells. The length of epidermis is calculated using the skeletonize feature. The rough outline and final region of interest (ROI) is seen in Figure 1 and 2, respectively. The final processing step includes a detection and quantification of positive nuclei in each ROI.

QUANTITATIVE OUTPUT VARIABLES
The output variables obtained from this protocol include:

  • Pos epi 1 - Pos epi 8: Area of positive nuclei in epidermis regions
  • Pos sec 1 - Pos sec 8: Area of positive nuclei in tissue sections
  • Pos epi numb 1 - Pos epi numb 8: Number of positive nuclei in epidermis regions
  • Pos sec numb 1 - Pos sec numb 8: Number of positive nuclei in tissue sections
  • Area epi 1 - Area epi 8: Area of epidermis regions
  • Area sec 1 - Area sec 8: Area of tissue sections
  • Length of epidermis 1 - Length of epidermis 8: Length of epidermis regions
     

WORKFLOW
The protocol exists in two versions: One for normal skin thickness and one for thickened skin. The user must manually choose which protocol to use. Each protocol is a sequence of five APPs.

Step 1: Load the APP for tissue detection “01 Outline Tissue”.
Step 2: Load the APP for epidermis detection “02 Detect Epidermis – rough”.
Step 3: Load the APP for removal of hair cells “03 Detect Epidermis – remove hair cells”. This APP outputs the variable “Length of epidermis 1 – Length of epidermis 8”.
Step 4: Load the cell detection protocol “04 Ki67 Cell Detection”.
Step 5: Load the quantification protocol “05 Cell Count”. Click the save button to transfer the results to the database. This APP outputs the first six variables noted in “Quantitative Output Variables”.

STAINING PROTOCOL
There is no staining protocol available.

ADDITIONAL INFORMATION
The APP utilizes the EngineTM and Viewer software modules, where EngineTM offers an execution platform to expand processing capability and speed of image analysis. Viewer gives a fast review together with several types of image adjustment properties ex. outlining of regions, annotations and direct measures of distance, curve length, radius, etc.

By adding the AuthorTM module the APP can be customized to fit other purposes. AuthorTM offers a comprehensive and dedicated set of tools for creating new fit-for-purpose analysis APPs, and no programming experience is required.

REFERENCES
1. Contrasting pathogenesis of atopic dermatitis and psoriasis—Part I: Clinical and pathologic concepts, E. Guttman-Yassky et al., The Journal of Allergy and Clinical Immunology (JACI) 127: 1110-8, 2011.

 

RUO
FIGURE 1
FIGURE 1
Initial rough outline of tissue and epidermis. The complete tissue area is referred to as a section.
FIGURE 2
FIGURE 2
Epidermis region after removal of hair cells.
FIGURE 3
FIGURE 3
Example of analyzing on thickened skin. Positive (red) and negative (blue) nuclei are labeled. The positive cells are afterwards quantified within the outlined epidermis region and for the entire tissue.
FIGURE 4
FIGURE 4
Example of analyzing on normal skin thickness. Positive (red) and negative (blue) nuclei are labeled. The positive cells are afterwards quantified within the outlined epidermis region and for the entire tissue.