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Measuring Fat and Vacuolar Space with Digital Pathology

Measuring Fat and Vacuolar Space with Digital Pathology. Frank Voelker, DVM, DACVP Principle Pathologist. Trevor Johnson, MS Director, Image Analysis. Originally An Algorithm for Measuring Fat. What was actually needed: An algorithm for measuring circumscribed clear spaces in tissue.

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Measuring Fat and Vacuolar Space with Digital Pathology

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  1. Measuring Fat and Vacuolar Space with Digital Pathology Frank Voelker, DVM, DACVP Principle Pathologist Trevor Johnson, MS Director, Image Analysis

  2. Originally An Algorithm for Measuring Fat What was actually needed: An algorithm for measuring circumscribed clear spaces in tissue • White Fat • Brown Fat • Lipid vacuoles in a variety of tissues (eg. Fatty liver or adrenal cortex) • Tissues composed of fibers or tubules (eg. skeletal muscle) • Tissues with hollow spaces or glandular lumena (eg. lung or thyroid gland) • Cerebral edema, demyelinization • Vacuoles in various cell types (eg. Lymphocytes)

  3. Algorithm Requirements • Data Output: • Percent area of vacuoles or circumscribed spaces. • Total number of spaces • Mean area/diameter/perimeter of spaces • Histograms of areas, diameters or perimeters • Shape and Area Threshold Controls • Hue Sensitivity Adjustment • Membrane Estimation or Completion

  4. Vacuole Algorithm Settings Input and Output Windows

  5. How the Image is Analyzed Define the tissue intensity Determine the maximum and minimum vacuole areas Determine how close each vacuole is allowed to resemble a circle and how tightly they are packed Determine if Vacuole Perimeter Estimation (VPM) is needed

  6. Measuring White Fat Cells

  7. Measuring White Fat

  8. Measuring White Fat – Removing Small and Elongated Vacuoles

  9. Measuring White Fat – Removing Small Vacuoles

  10. Morphometric Considerations

  11. Remove Tissue (Just View Vacuole Spaces in Green)

  12. Remove Tissue (Parameter Too Low Including Tissue)

  13. Include Small Vacuoles

  14. Exclude Large Vacuoles

  15. Vacuole Analysis Fatty Liver

  16. Fatty Liver

  17. Measuring Brown Fat Vacuoles

  18. Measuring Brown Fat Vacuoles

  19. Measuring Brown Fat with Nuclear Counting 173 Nuclei Area/No. Nuclei = Cell Size No. Nuclei/No. Fat Droplets = No. Droplets/Cell

  20. Measuring Pulmonary Alveoli

  21. Measuring Pulmonary Alveoli

  22. Adrenal Gland

  23. Vacuole Analysis Adrenal

  24. Measuring Adrenal Vacuoles with Nuclear Counts 187 Nuclei

  25. Measuring Adrenal Vacuoles

  26. Measuring Bone Marrow Vacuoles

  27. Measuring Bone Marrow Vacuoles

  28. Measuring Skeletal Muscle Fibers Type 1- Red Muscle. Slow twitch postural type muscle fiber. Aerobic oxidation. Type 2- White Muscle Fast twitch fibers. ATPase +. Anaerobic oxidation Muscle fiber alterations are important in: Denervation Statin-induced rhabdomyolysis Noninflammatory myopathies Congential myopathies Sports physiology

  29. Measuring Skeletal Muscle Fibers In H&E sections, skeletal muscle fibers are poorly demarcated

  30. Measuring Skeletal Muscle Fibers

  31. Muscle Fiber Histogram

  32. Summary An algorithm designed to measure vacuoles or circumscribed clear spaces in tissue was described in this presentation. The algorithm has proved useful in measuring clear spaces in a variety of tissues including white and brown fat, skeletal muscle fibers, pulmonary alveoli, fatty liver and adrenal cortex. Data outputs included percent as well as histograms and mean areas, diameters or perimeters of vacuoles or clear areas within tissue.

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